**Contents**

Reprinted from: *Diagnostics* **2020**, *10*, 960, doi:10.3390/diagnostics10110960 ............ **129**

## **Andreas Hahn, Hagen Frickmann and Ulrike Loderst¨adt**

Reprinted from: *Diagnostics* **2022**, *12*, 716, doi:10.3390/diagnostics12030716 ............. **185**

## **About the Editor**

## **Tivani P. Mashamba-Thompson**

Tivani P. Mashamba-Thompson is a Medical Scientist (registered by the Health Professions Council of South Africa) and a Professor of Diagnostics Research. She was appointed as a Full Professor and a Deputy Dean of Research and Postgraduate Studies for the Faculty of Health Sciences, University of Pretoria, with effect from 1 January 2021. She also serves as a nonexecutive board member of the National Health Laboratory Services (NHLS) as a Deputy Chairperson of the NHLS Research and Innovation Committee. She is also a member of the South African Department of Health Office of Health Standards Compliance technical task team. She has previously served as a diagnostics expert advisor for Abbott International.

## *Editorial* **Implementation Science: Bridging the Gap between Point-of-Care Diagnostics Research and Practice**

**Tivani P. Mashamba-Thompson**

Faculty of Health Sciences, University of Pretoria, Pretoria 0002, South Africa<EMAIL_ADDRESS>
The advent of the novel Coronavirus 2019 (COVID-19) pandemic has fuelled technological innovation and led to the increased research on development and deployment of new diagnostics for use at point-of-care (POC). The rapid uptake of the newly developed diagnostics requires a systematic approach to bridge the research-to-practice gap. Implementation science (IS) involves the use of evidence-based practices (EBPs) that are characterised by both quality improvement and dissemination methods aiming to promote the scaling up of health interventions such as POC diagnostics to enhance quality and outcomes [1]. This research approach employs transdisciplinary quantitative and qualitative designs with solid grounding in theory. Implementation science studies are designed to enable identification of factors that impact uptake of health interventions across multiple levels, including the patient, provider, clinic, facility, organisation, and often the broader community and policy environment. In this Special Issue, we present a summary of twelve studies that employed implementation science approaches demonstrating research aimed at optimising implementation various kinds of point-of-care (POC) diagnostics among different population groups and different healthcare settings globally.

Nucleic acid amplification tests (NAATs), such as the reverse-transcriptase polymerase chain reaction (RT-PCR) tests, were the first to be developed and widely deployed at the beginning of the COVID-19 pandemic. These tests were designed to detect viral RNA. A RT-PCR-positive result is highly specific for the presence of viral nucleic acid. A study on portable, easy-to-use SARS-CoV-2 RT-PCR POC diagnostic device showed that the POC test's performance was comparable to that of the conventional RT-qPCR tests [2]. It also showed that the POC RT-PCT displayed the characteristics of a POC test, with a short turnaround time for quick patient isolation. The implementation of the first official approved self-administered rapid antigen tests (RATs), CoviSelf, was assessed as part of a community-level COVID-19 pandemic response in rural India [3]. Results of the study show that self-administered RATs have potential in rural settings as they are cheap, quick and reasonably reliable. However, the tests kits were found to not be user-friendly and required equitable distribution to minimise the spread of COVID-19. One study conducted at a hospital setting in Germany evaluated the lung ultrasound (LUS) in 101 symptomatic patients with suspected COVID-19 infection at hospital admission [4]. Results of this evaluation demonstrate that early LUS examination as part of in-patient admission provides a diagnostic gain and is valuable for the clarification of SARS-CoV-2-suspected patients at hospital admission. An assessment of the correlation between hospital-based lung ultrasound (LUS) and chest X-ray (CXR) findings in 247 COVID-19 patients in Spain showed positive results [5]. A significant correlation between LUS findings and CXR in patients with suspected or confirmed COVID-19 infection and supported use of the LUS exam as the initial POC diagnostic imaging test was demonstrated.

Interventions such as mobile Health (mHealth) have been shown to help enhance health service delivery and access to disease diagnosis. Increased availability and use of mHealth to help improve access to diagnostics is recommended for resource-limited settings. Osei et al., 2021 conducted a study to examine the availability and use of mHealth

**Citation:** Mashamba-Thompson, T.P. Implementation Science: Bridging the Gap between Point-of-Care Diagnostics Research and Practice. *Diagnostics* **2022**, *12*, 1648. https://doi.org/10.3390/ diagnostics12071648

Received: 5 July 2022 Accepted: 5 July 2022 Published: 6 July 2022

**Publisher's Note:** MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

**Copyright:** © 2022 by the author. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

for disease diagnosis and treatment support by healthcare professionals in the Ashanti Region of Ghana [6]. The findings in this study show minimal use of mHealth for disease diagnosis and treatment support by healthcare professionals at rural clinics, which in turn affects the accessibility of medical care in such resource constrained settings. One of the contributing factors for equitable accessibility, distribution and availability of POC tests is the agility of supply chain management systems. Maluleke et al., 2021, conducted a scoping review to systematically map evidence on supply chain management systems for POC diagnostics services with a focus on optimising the SARS-CoV-2 testing capacity in resourcelimited settings [7]. The results of the review showed has showed that there is limited research evidence on POC diagnostics supply chain management systems globally. In addition to ensuring accessibility of new POC diagnostics to those who need it, acceptance by stakeholders is key to enabling uptake and appropriate usage of available diagnostics. Thirty-one stakeholders involved in adoption of POC ultrasound (POCUS) implementation in a US academic medical centre were interviewed to determine their perspective on this diagnostic intervention [8]. The Practical Robust Implementation and Sustainability Model (PRISM) was employed to guide framework analysis for the data collected from interviews with stakeholders. Stakeholders considered the following overarching themes to be important for the adoption and fidelity of POCUS by clinicians and health systems: clinical impact; efficiency; cost; development of credentialing policies; and robust quality assurance processes.

The need to optimise and monitor POC diagnostics quality management systems has been emphasised in other studies [9–11]. A systematic review and meta-analysis of global evidence showed moderate accuracy of mobile-linked POC diagnostics in detecting infections and recommended the development and deployment of more highly accurate mHealth-linked POC diagnostics [11]. Evaluation of the prognostic capacity of ΔLA (delta lactate) (correlation between prehospital lactate (pLA) and hospital lactate (hLA)) with respect to in-hospital two-day mortality among emergency department patients was also performed. Results of the evaluation demonstrate that lactate clearance in the initial moments of ED care appears to be a more reliable prognostic index than a baseline lactate value taken alone [9]. Hahn et al., 2021, employed a theorical model to demonstrate sensitivity-optimised screening as a "diagnostics as prevention" strategy for managing infectious diseases using HIV infections as a prototype [10]. The model was designed to increase case definitions for diagnostic test sensitivity of by compensating for the limited sensitivity of a test in the early stage of a disease. The model also enabled inclusion of known symptoms of the respective disease stage and RDT-based exposition prevention in a pandemic. This concept was widely used for the management of COVID-19 through applying rapid diagnostic tests with imperfect diagnostic accuracy.

Despite the technological advancements presented by the COVID-19 pandemic, disruption to services directed at management of existing infectious and deadly pandemics, such as TB was reported during the early stages of the pandemic. Dlangalala et al., 2021 systematically mapped available evidence on TB services at the primary healthcare (PHC) level during the COVID-19 period using as scoping review study [12]. The study revealed that pandemic mitigation strategies, as well as the fear and stigma experienced at the beginning of the pandemic, may have limited uptake of TB diagnostic services at PHC level. The presented poor TB service up-take may also be a result of poor health literacy, which has been shown to be generally low among vulnerable populations. In this context, health literacy would be defined in line with access to technology enabling disease screening, diagnosis and linkage to care, i.e., diagnostics literacy. The COVID-19 pandemic has revealed diagnostics literacy as one of the unmet needs among vulnerable populations that continue to experience short- and longer-term socio-economic consequences. To address this unmet need a multi-level diagnostics literacy advocacy model was proposed to help improve diagnostic uptake among vulnerable populations [13]. Sustainable implementation of the proposed model will require involvement of all key stakeholders.

**Funding:** This research received no external funding.

**Conflicts of Interest:** The author declares no conflict of interest.

## **References**

## *Brief Report* **Analytical Evaluation of Visby Medical RT-PCR Portable Device for Rapid Detection of SARS-CoV-2**

**Adriana Renzoni 1,\*, Francisco Perez 2, Marie Thérèse Ngo Nsoga 2,3, Sabine Yerly 1, Erik Boehm 1, Angèle Gayet-Ageron 4, Laurent Kaiser 1,2,3 and Manuel Schibler 1,3**

**Abstract:** Extended community testing constitutes one of the main strategic pillars in controlling the COVID-19 pandemic. Reverse transcription PCR (RT-PCR) targeting the SARS-CoV-2 genome on nasopharyngeal swab samples is currently the reference test. While displaying excellent analytical sensitivity and specificity, this test is costly, often requires a substantial turnaround time, and, more importantly, is subject to reagent and other material shortages. To complement this technology, rapid antigen tests have been developed and made available worldwide, allowing cheap, quick, and decentralized SARS-CoV-2 testing. The main drawback of these tests is the reduced sensitivity when RT-PCR is the gold standard. In this study, we evaluate Visby an innovative, portable, easy-to-use RT-PCR point-of-care (POC) diagnostic device. Our retrospective analysis shows that overall, compared to the Cobas 6800 RT-qPCR assay (Roche), this RT-PCR POC technology detects SARS-CoV-2 RNA with 95% sensitivity (95%CI = 86.3–99%) and 100% specificity (95% CI = 80.5–100%). For samples with cycle-threshold values below 31, we observed 100% sensitivity (95% CI = 66.4–100%). While showing an analytical sensitivity slightly below that of a standard RT-qPCR system, the evaluated Visby RT-PCR POC device may prove to be an interesting diagnostic alternative in the COVID-19 pandemic, potentially combining the practical advantages of rapid antigen tests and the robust analytical performances of nucleic acid detection systems.

**Keywords:** SARS-CoV-2; rapid diagnostic techniques; POCT techniques

## **1. Introduction**

Complementary to prevention measures, such as social distancing, mask wearing, and hand hygiene, extensive population screening with isolation of positive cases is one of the key means of controlling the COVID-19 pandemic caused by severe SARS-CoV-2 [1]. SARS-CoV-2 RNA detection by RT-qPCR) in nasopharyngeal respiratory samples is currently considered to represent the gold standard for COVID-19 diagnosis. This technology, known for its high analytical sensitivity and specificity, has been applied to clinical virology diagnostics since the 1990s. On the other hand, it is costly, requires complex laboratory infrastructure, often has a high turnaround time, and even more importantly, RT-qPCR reagents and plastic materials are subject to shortages in the current crisis [2].

Rapid antigen tests are increasingly widely used, allowing for user-friendly, low-cost, quick, and decentralized testing. The main limitation of these tests is the reduced sensitivity in comparison to RT-qPCR, around 90% in the best-case scenario. Innovative nucleic acid

**Citation:** Renzoni, A.; Perez, F.; Ngo Nsoga, M.T.; Yerly, S.; Boehm, E.; Gayet-Ageron, A.; Kaiser, L.; Schibler, M. Analytical Evaluation of Visby Medical RT-PCR Portable Device for Rapid Detection of SARS-CoV-2. *Diagnostics* **2021**, *11*, 813. https:// doi.org/10.3390/diagnostics11050813

Academic Editor: Tivani P. Mashamba-Thompson

Received: 8 March 2021 Accepted: 28 April 2021 Published: 29 April 2021

**Publisher's Note:** MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

**Copyright:** © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

detection point-of-care (POC) diagnostic technologies are now being developed, potentially combining the excellent analytical performance of RT-qPCR and the practical advantages of rapid antigen tests [3,4].

An alternative to the detection of antigen is the amplification of nucleic acid by reverse transcription in combination with isothermal methods such as: loop-mediated isothermal amplification (RT-LAMP) [5–7], recombinase polymerase amplification (RT-RPA) [8–10], and nicking-endonuclease amplification reactions (RT-NEAR) [11–14]; which do not need a thermal cycler, potentially enabling their use at POC facilities. The reference standard of RT-PCR also detects viral nucleic acid, and some devices capable of RT-PCR have been developed that are suitable for POC use, such as the DNAnudge Covid-19 test.

Recently, Visby Medical Inc. (Visby) has developed and begun manufacturing small palm-sized RT-PCR devices capable of detecting SARS-CoV-2. This device contains all necessary reagents, and is operated simply by loading a sample and sequentially pushing buttons; thus representing a self-contained easy-to-use RT-PCR POC diagnostic device. The device makes use of a continuous flow serpentine PCR channel combined with an enzyme linked detection chemistry to produce a colorimetric signal that is easily observable by the user. The device, originally developed for detection of bacterial targets in a sexually transmitted infection panel [15] has been adapted for rapid RT-PCR identification of the SARS-CoV-2 N gene and has received an FDA EUA for use in high and moderate complexity settings. More recently, on 8 February 2021, the Visby test received EUA for use in CLIAwaived settings (https://www.fda.gov/media/145917/download, accessed on 28 April 2021). Of note, the device may only be used once before requiring disposal or refurbishment by the manufacturer. Visby is intended for testing individuals suspected of COVID-19 by their health care provider using nasopharygeal, anterior nasal or mid-turbinate swabs collected into transport media,

In this study, we tested the new RT-PCR POC Device developed by Visby to detect the presence of SARS-CoV-2 RNA in a retrospective study using frozen SARS-CoV-2 positive nasopharyngeal samples using Cobas 6800 RT-qPCR assay (Roche) as the gold standard.

## **2. Material and Methods**

## *2.1. Patient Samples and Assay Controls*

According to the Swiss Ethics Committee on research involving leftover clinical samples, this study occurred in the context of a method quality validation in an emergency setting and therefore did not require any authorization from our ethics committee. The vast majority of positive samples come from outpatient symptomatic patients enrolled in a COVID-19 screening center at the Geneva University Hospitals. Sixty-one confirmed positive SARS-CoV-2 nasopharyngeal swab (NPS) samples were tested. Sample positivity, that is samples with a cycle-threshold (Ct) value below 35, was previously determined by using the Cobas 6800 SARS-CoV-2 RT-PCR assay (Roche, Switzerland) and stored in Copan UTMTM or an in-house DMEM-based media that has been validated in our institution [16]. Samples were collected around 6 weeks or less prior to device testing and frozen at −80 ◦C. Patient specific data include demographics, symptom severity, and date of symptoms onset. Seventeen SARS-CoV-2 PCR negative NPS samples were tested as negative controls. Two external positive and negative controls (NATSARS (COV2)-ERC and NATSARS (COV2)- NEG, respectively) were obtained from ZeptoMetrix (Distributed by Helvetica Health Care, Geneva, Switzerland). External controls were performed when different operators were running the experiments. In total, 17 samples were considered as controls in this study.

## *2.2. Visby Medical POC Device Testing*

Analysis of samples and external controls was performed following the manufacturer protocol (https://www.fda.gov/media/142228/download, accessed on 28 April 2021) with some modifications, as we performed a retrospective study using frozen SARS-CoV-2 samples. Briefly, specimens were mixed by inversion 5 times, and 650 μL was inserted into provided dilution buffer using the special Pasteur pipette provided with the device

(Visby Medical) (Figure 1A). The diluted sample was mixed five times by inversion, 650 μL was taken with a second Pasteur pipette and dispensed immediately into the device (Button No. 1). After loading the sample; sample extraction, reverse transcription, PCR amplification, and result visualization were started by sequentially pushing down buttons 1, 2 and 3 and plugging the device in. A successful device start is indicated by a white light. After 30 min, a green or red check mark appears confirming a valid or invalid device performance, respectively (Figure 1B). Samples run in invalid devices were retested using a new device.

**Figure 1.** (**A**) Schematic steps for Visby Medical kit procedure. (1) Kit contents. (2) Patient sample preparation, with dilution buffer and cassette. (3–5) The sample is put into the dilution buffer and mixed by inversion. (6–7) 650 uL of the diluted sample is inserted into the device using the pastette. (8) Buttons 1, 2, and 3 are pushed, and the device is connected to electrical power. (9) Results displayed. A white checkmark appears, indicating that the test is in progress (see description below). (**B**) The left panel shows the white or green or red check marks denoting correct power and processing of the device, a valid test result ready to be read, or an invalid result due to an electrical error, respectively. The right panel shows an example of a valid (green light) and positive (visible upper and lower purple spots) or negative (only the upper purple spot is visible).

Results of valid devices were displayed in the test result window as visible purple marks (Figure 1B). A purple mark in the upper position (indicated by a "√" symbol) indicates a valid sample extraction and amplification reaction, which was determined by detecting the human 18S ribosomal structural RNA present in nasopharyngeal samples. A purple mark in the lower position (indicated by a "+" symbol) indicates a positive amplification of SARS-CoV-2 nucleic acid. Visualization of both upper and lower position purple marks denotes a positive SARS-CoV-2 sample, while visualization of a single upper position purple mark denotes a negative SARS-CoV-2 sample (Figure 1B). The external positive and negative controls were used to ensure that test reagents are working correctly. Each control was run when a new operator was running the tests. Results interpretation was performed in a blinded manner by two laboratory staff members.

## *2.3. Comparison RT-qPCR Assay*

The validated Roche Cobas 6800 SARS-CoV-2 RNA extraction and RT-qPCR assay were used as a reference assay (Cobas SARS-CoV-2 Ref<PHONE_NUMBER>0; Cobas SARS-CoV-2 Control kit Ref:<PHONE_NUMBER>0; Cobas 6800/8800 Buffer Negative Control kit Ref<PHONE_NUMBER>0; Roche, Switzerland). In case of discrepant results between the Cobas and Visby assays, viral RNA genome detection was also performed using GeneXpert (Ref: XPRSARS-CoV2-10 from Cepheid). Cobas targets ORF1a/b and a pan-Sarbecovirus conserved region of the E gene, while GeneXpert targets N and E genes.

## *2.4. Evaluation of Analytical Performances*

The sensitivity and specificity of the device were determined by comparison to the Cobas assay, according to sensitivity = TP/(TP + FN); Specificity = TN/(TN + FP); where TP = true-positives, FN = false-negatives, TN = true-negatives, and FP = falsepositives. Confidence intervals of 95% (95% CI) were calculated using the Clopper–Pearson method [17]. We reported overall sensitivity and specificity, as well as sensitivity by delay from symptoms onset and by categories of Ct values.

## *2.5. Limit of Detection (LoD)*

As recommended by the manufacturer, a SARS-CoV-2 negative nasopharyngeal cell matrix was prepared to perform LoD analyses. First, nasopharyngeal samples, which were previously verified to be SARS-CoV-2 negative, were pooled to obtain a total of 12 mL. The pooled nasopharyngeal sample was retested by Cobas RT-qPCR to ensure that the sample pool was negative and immediately frozen at −80 ◦C. SARS-CoV-2 viral stock was produced as follows: SARS-CoV-2 isolates were collected from ex vivo infections of airway epithelia cultured in an air-liquid interface as previously described [18]. Briefly, after 3 h of apical virus inoculation, in vitro differentiated respiratory tissues (MucilAirTM, Epithelix SARL, Geneva, Switzerland) were washed three times with PBS (Phosphate Buffered Saline, Sigma, St. Louis, MO, USA) and incubated in MucilAirTM medium at 33 ◦C and 5% CO2. Four days post infection, respiratory tissues culture supernatant is removed and 200μL of MucilAirTM medium was added apically and viral culture supernatant was collected after 20 min of incubation at 33 ◦C at 5% CO2. Viral load in the culture supernatant was determined by RNA quantification. Briefly, RNA was extracted with NucliSens easyMAG (bioMérieux, Marcy-l'Étoile, France), and quantified by RT-qPCR using SuperScript™ III Platinum™ One-Step qRT-PCR Kit (Invitrogen, Carlsbad, CA, USA) in a CFX96 Thermal Cycler (Bio-Rad, Hercules, CA, USA). RT-qPCR was performed using a specific set of primers and probes targeting the SARS-CoV-2 E gene (forward primer: 5 -ACAGGTACGTTAATAGTTAATAGCGT-3 , reverse primer: 5 -ATATTGCAGCAGTACG CACACA-3 , and the probe: 5 -6-FAM- ACACTAGCCATCCTTACTGCGCTTCG-BBQ-3 ) [19]. In vitro transcribed SARS-CoV-2 E gene RNA (EVAg, Essen, Germany) was used as a reference standard to convert Ct values into RNA copies/mL. Data were analyzed using Bio-Rad CFX maestro software (Bio-Rad, Essen, Germany). Quantified culture supernatant was finally diluted in PBS to a concentration of 2.1 × <sup>10</sup><sup>6</sup> copies/mL.

## **3. Results**

## *3.1. Visby POC Device and Cobas SARS-CoV-2 Nucleic Acid Detection Concordance*

Frozen samples were thawed, dispensed immediately into dilution buffer provided with the device, and further processed to avoid nucleic acid damage as much as possible. After 30 min of sample processing, purple mark recording was performed immediately in devices displaying the green light. An invalid run occurred for a single sample, but after retesting, yielded a valid result.

The clinical accuracy of the device compared to Cobas RT-qPCR was calculated. From the 61 Cobas-positive samples (Ct values from 15.5 to 34), the device detected 58 positive samples with 3 false-negative samples. All 17 Cobas-negative samples were also found to be negative by the device (Table 1). We observed a sensitivity of 95.1% (CI 95% = 86.3–99%) and a specificity of 100% (CI 95% = 80.5–100%). An analysis by Ct values showed that the device achieved 100% sensitivity in samples with Ct values below 35 (Table 2). As shown in Table 3, the device achieves > 95% sensitivity with tests performed from 2 days after onset of symptoms, and 100% (73.5–100%) from 4 days post symptoms onset. We further evaluated Visby results in relation to viral loads and days post onset of symptoms, and observed that the few false-negative results were observed with three samples with low viral loads (Figure 2).

Since discrepant results might be explained by the effects of sample freezing on RNA stability, we retested the discordant samples with the Cobas assay. Nearly identical positive Cobas Ct values were found, showing that RNA degradation upon thawing samples could not explain the negative device results (Table S1). Discrepant sample results might be explained by detection of different genes used by the device (N-gene) and the Cobas (E and Orf1 genes) assay. We therefore analyzed discrepant samples with the SARS-CoV-2 GeneXpert assay, targeting N and E genes, however, both genes were detected (Table S1).

**Table 1.** Sensitivity and specificity of the Visby device.

Sensitivity: 95% (95% CI 86–99)/specificity: 100% (95% CI 80.5–100).

**Table 2.** Sensitivity of Visby device depending on RT-qPCR CT values.

**Table 3.** Sensitivity of Visby device by the delay since symptom onset.

**Figure 2.** Detection of SARS-CoV-2 by Visby device based on viral loads and the time of symptom onset. SARS-CoV-2 viral loads by the time of symptom onset for the symptomatic and RT-qPCR positive individuals. Black and red dots represent positive or negative Visby results, respectively. Dotted line: 6 log10 SARS-CoV-2 RNA copies/mL shows the VL threshold limit determining the number of culturable viruses. Note: Samples classified at day 0 include 4 samples with unknown dates of symptoms.

## *3.2. Analytical Limit of Detection (LoD)*

A clinical SARS-CoV-2 isolate was cultured in an air-liquid interface respiratory epithelium system (mucilAir). Viral supernatant concentration was quantified using Egene standards from the European virus archive and diluted to a final concentration of 2.1 × <sup>10</sup><sup>6</sup> copies/mL. Live virus from cell culture supernatant were serially diluted (105, 104, 103, 102, 101 copies/mL) in SARS-CoV-2 negative nasopharyngeal cell matrix to evaluate the LoD. We tested a single replicate of each dilution and found them all positive, showing a Visby positive detection with low viral loads (102 copies/mL). Our data are in agreement with the manufacturer LoD determination, which showed a 95% detection rate for 1112 genomic copies/mL and a detection rate of 45% for 125 genomic copies/mL (Visby medical package insert).

## **4. Discussion and Conclusions**

The aim of this study was to evaluate the Visby POC device to detect the presence of SARS-CoV-2 RNA in previously collected frozen clinical nasopharyngeal samples, in comparison to the Cobas 6800 RT-qPCR assay (Roche). The devices detected SARS-CoV-2 RNA with 95% sensitivity and 100% specificity.

Few discrepant results were observed that might be explained by potential N-gene mutations that are undetectable by the device, similarly to previously published observations showing that failure to detect SARS-CoV-2 by the Cobas 6800 assay was linked to E-gene mutations [20]. This is a possible scenario, but an unlikely one, as extensive in silico analysis of variants were detected by Visby primers and probes (https: //www.fda.gov/media/142228/download, accessed on 28 April 2021). Having excluded potential issues related to different viral genes targeted with both assays, the reduced sensitivity observed with the devices probably results from the different technologies applied, which differ in technical details such as buffers and sample processing methods. Visby's test is not directly compatible with most viral transport media (manufacturer package insert). While dilution of samples collected in transport media mitigates the inhibitory effects of the transport media, it reduces the test sensitivity, which could account for the 3 Cobas-positive samples that the device missed. However, this dilution step did not prevent detection of other samples with similar viral loads. We cannot exclude the possibility of PCR inhibiting substances within these nasal samples that prevent gene amplification exclusively with the device methodology [21].

Dilution of the sample collected into transport media to mitigate the inhibitory effects of the transport media may reduce the sensitivity of the test. While a remnant sample evaluation was performed in this study, it is reasonable to assume that direct swab elution with no dilution step would yield an improvement in sensitivity. A prospective clinical study would be important to assess the clinical sensitivity of the direct swab method.

In the present analytical study, the POC Device performs less well at Ct values ≥35; however, the reduction in sensitivity is relatively unimportant since high Ct values probably indicate a low transmission risk [22]. Furthermore, the sensitivity displayed by the device is still substantially higher than that of the 85% to 90% of the best antigen tests [23–27].

This device operates by detecting viral nucleic acids such as the reference standard (RT-PCR) and isothermal amplification tests. RT-LAMP amplification unfortunately has issues such as: non-specific amplification that can be a problem in later steps as a result of "carryover contamination", "product cross contamination" [28–30], or primer hybridization [31].

In contrast, the device evaluated here uses the same robust and well established principle used by the gold-standard tests, and thus is expected to be reliable and accurate. Other RT-PCR devices suitable for POC diagnostics are on the market (i.e., NudgeBox or LIAT) but are less portable than the Visby POC device evaluated here, and may be unavailable or have a sample processing time of up to 90 min that may be prohibitive for some envisioned POC uses [4].

It is important to note the limitations of our study. The selection of samples tested is not representative of an epidemiological trend because only frozen samples were used, and no prospective recruitment of patients was performed with both methods concomitantly. No estimation of positive or negative predictive values can be performed as samples were not consecutively collected and prevalence is not interpretable. Future clinical evaluations, including head-to-head comparisons with antigen tests and isothermal tests, testing of SARS-CoV-2 asymptomatic-contacts, and screening healthcare workers will help to better position SARS-CoV-2 nucleic acid detection among the POC diagnostics.

In summary, the Visby device appears to be an attractive alternative POC test, performing nearly as well as conventional RT-qPCR tests, while displaying the appealing characteristics of a POC test. It is available in a portable format with minimal requirement of technical skills. After sample acquisition, less than 1 min is needed for treatment and loading samples onto de VISBY device. Viral extraction, amplification and easy-readout are obtained after exactly 30 min. This short turnaround time allows for rapid patient isolation or orientation decisions, e.g., in nursing homes or even hospital settings lacking more complex laboratory equipment. A potential disadvantage includes the generation of waste that remains an issue to be addressed. The company is exploring recycling components to reduce generated waste.

**Supplementary Materials:** The following are available online at https://www.mdpi.com/article/ 10.3390/diagnostics11050813/s1. Table S1: Positive and negative RT-qPCR Cobas samples tested on RT-PCR Visby diagnostic device. C = corncordant result; NC = Non-concordant results. INV = invalid result.

**Author Contributions:** A.R., conception, experimental manipulation and analysis, writing the manuscript. F.P., experimental manipulation and critical reading of the manuscript. M.T.N.N. experimental manipulation and critical reading of the manuscript. S.Y., conception and critical reading of the manuscript. E.B., conception and critical reading of the manuscript. A.G.-A., statistical analysis and critical reading of the manuscript. L.K., conception and critical reading of the manuscript. M.S., conception, experimental analysis, writing and critical reading of the manuscript. All authors have read and agreed to the published version of the manuscript.

**Funding:** Marie Thérèse Ngo Nsoga is a beneficiary of the excellence grant from the Swiss Confederation and the grant from the humanitarian commission of the University Hospital of Geneva.

**Institutional Review Board Statement:** According to the Swiss Ethics Committee on research involving leftover clinical samples, this study occurred in the context of a method quality validation in an emergency setting and therefore did not require any authorization from our ethics committee.

**Acknowledgments:** We thanks Pascal Cherpillod for the pictures.

**Conflicts of Interest:** The authors declare that there are no conflicts of interest.

## **References**

## *Article* **Can Self-Administered Rapid Antigen Tests (RATs) Help Rural India? An Evaluation of the CoviSelf Kit as a Response to the 2019–2022 COVID-19 Pandemic**

**Marika Vicziany 1,\* and Jaideep Hardikar <sup>2</sup>**

**Abstract:** This paper evaluates India's first officially approved self-administered rapid antigen test kit against COVID-19, a device called CoviSelf. The context is rural India. Rapid antigen tests (RATs) are currently popular in situations where vaccination rates are low, where sections of the community remain unvaccinated, where the COVID-19 pandemic continues to grow and where easy or timely access to RTPCR (reverse transcription-polymerase chain reaction) testing is not an option. Given that rural residents make up 66% of the Indian population, our evaluation focuses on the question of whether this self-administered RAT could help protect villagers and contain the Indian pandemic. CoviSelf has two components: the test and IT (information technology) parts. Using discourse analysis, a qualitative methodology, we evaluate the practicality of the kit on the basis of data in its instructional leaflet, reports about India's 'digital divide' and our published research on the constraints of daily life in Indian villages. This paper does not provide a scientific assessment of the effectiveness of CoviSelf in detecting infection. As social scientists, our contribution sits within the field of qualitative studies of medical and health problems. Self-administered RATs are cheap, quick and reasonably reliable. Hence, point-of-care testing at the doorsteps of villagers has much potential, but realising the benefits of innovative, diagnostic medical technologies requires a realistic understanding of the conditions in Indian villages and designing devices that work in rural situations. This paper forms part of a larger project regarding the COVID-19 pandemic in rural India. A follow-up study based on fieldwork is planned for 2022–2023.

**Keywords:** COVID-19; Indian pandemic; rapid antigen tests (RATs); CoviSelf; CoWIN; digital divide; rural India; Indian villagers; poverty; discourse analysis; qualitative medical/health research strategies

## **1. Introduction**

As the Indian COVID-19 pandemic enters a new phase in 2022 with the rising dominance of the highly infectious Omicron variant, the use of self-administered RATs is increasing in India and other countries as a way of relieving the pressure on stressed clinical and hospital settings that administer and analyse reverse transcription polymerase chain reaction (hereafter RTPCR) tests [1]. As a third pandemic wave emerged in India, on 10 February 2022, the Indian Council of Medical Research (hereafter ICMR) announced that a positive result from a self-administered RAT was the equivalent of a positive result from an RTPCR procedure and that repeat testing using the latter was no longer necessary [2]. In this paper, we evaluate the first self-administered RAT to receive official approval by ICMR some nine months earlier, on 10 May 2021 [3]—CoviSelf. ICMR is the 'apex body in India for the formulation, coordination and promotion of biomedical research' [4] and sits within the Department of Health Research (Ministry of Health and Family Welfare), Government of India. As the first Indian made, self-administered RAT to receive official recognition,

**Citation:** Vicziany, M.; Hardikar, J. Can Self-Administered Rapid Antigen Tests (RATs) Help Rural India? An Evaluation of the CoviSelf Kit as a Response to the 2019–2022 COVID-19 Pandemic. *Diagnostics* **2022**, *12*, 644. https://doi.org/ 10.3390/diagnostics12030644

Academic Editor: Tivani P. Mashamba-Thompson

Received: 22 November 2021 Accepted: 21 February 2022 Published: 6 March 2022

**Publisher's Note:** MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

**Copyright:** © 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

CoviSelf deserves our scrutiny. This diagnostic device for home use was developed by the private Indian company Mylab Discovery Solutions Pvt Ltd, based in Pune in the state of Maharashtra [5]. The tone of the company's press release of 10 May 2021 reflects the excitement surrounding the development of this innovative testing kit during increasingly desperate times [3]:

With CoviSelfTM, Mylab aims to make testing reach *the doorstep of every Indian* to help them fight the second and any subsequent waves of [*sic*] pandemic. *Now, any citizen can test for Covid-19 themselves*, isolate and seek treatment quickly. Early detection can help save thousands of lives and significantly reduce [*sic*] burden on hospitals. The test can be purchased without a prescription from local pharmacies and online channel partners [italics added by authors of this article]

The claims about delivering a diagnostic tool for containing COVID-19 to 'every Indian ... any citizen' are ambitious and admirable but need to be tested. Our analysis is not based on a scientific or laboratory assessment of the accuracy of this testing device. Few reliable studies of RAT evaluations have been published; perhaps the best-known exception is the UK's Cochrane Report, published in March 2021 [6]. Until detailed and systematic quantitative data are published regarding Indian RAT devices, the official list of ICMR-approved RATs is our only source of information about the four criteria used to validate and approve the RATs distributed in India, including those used in home testing [7]. Moreover, official criteria are not an exact guide to how RATs function in medical, health and home settings. In mid-January 2022, ICMR published a list of eight self-test kits that it had approved, in addition to another list of five that it did not approve [7]. At the top of the approved list appears CoviSelf by Mylab.

As the first government sanctioned, self-administered RAT for COVID-19, our analysis of CoviSelf also provides a benchmark for evaluating the others that have become available in India since 10 May 2021. However, a scientific appraisal of the sensitivity and specificity of the RATs in India is not our aim in this paper [8]. Rather, our evaluation is based on the extent to which the CoviSelf kit is compatible with the practical needs and concerns of Indian villagers. We do this by considering the characteristics of CoviSelf in the context of the socioeconomic circumstances in Indian villages. Some 66% of Indians live in rural areas [9], so the applicability of CoviSelf has widespread implications for addressing the country's pandemic while India waits for additional medical technologies to become available—for example, anti-COVID medication [10]. Yet, there is nothing inevitable about the arrival of such drugs or the trajectory that the pandemic could take. Given this, lessons can be learnt about the longer-term contribution that self-administered RATs could make to villagers' wellbeing by analysing the practicality of the CoviSelf kit. Mylab's aim for CoviSelf to reach 'every Indian and any citizen' is best tested against the socioeconomic parameters that define villages, where the majority of Indians live.

Our paper provides a qualitative evaluation of CoviSelf within a broad socioeconomic context. In 2008, Sir Michael Rawlins stressed the importance of recognising the value of qualitative research for health interventions. He questioned the hierarchy of evidence that gave greater importance to randomised clinical trials as the basis for health decisions. 'Observational evidence' or qualitative data, including 'expert opinion', formed an important component of therapeutic recommendations, he said [11]. Rawlins' advice has been taken seriously and new health-medical literature has emerged in response [12,13], including an appreciation of the value of analysing documents of both a textual [14] and non-textual kind. This article belongs to this growing school of thought regarding the significance of qualitative research and its role in supporting better health and medical initiatives. Our analysis sits within the framework of discourse analysis, using textual sources to analyse CoviSelf. The words and language of these texts constitute our data. Our interest in the socioeconomic context of new diagnostic medical devices was prompted by one of our engineering colleagues, who asked: 'what is the point of us inventing diagnostic tools unless we know how these fit into the daily life and needs of ordinary people?' [15].

## **2. RATs in Clinical Settings in India**

We are not aware of any published research assessing the self-administered CoviSelf kit, so we turn to what is known about RATs in clinical settings during India's COVID-19 pandemic. 'Clinical settings' are those in which trained professionals administer the test in either an institutional or private environment, including clinics, hospitals, schools, offices, prisons, military facilities etc. This experience speaks to the value of making self-administered RATs accessible to all Indians.

By April 2021, 49% of all the tests for COVID-19 in India used RATs, 'principally the SD Biosensor test (SD Biosensor Standard Q COVID 19 Antigen test)' [16]. This was in response to the second wave of COVID-19 in the early months of 2021, a wave defined by the Delta strain and the shift of the Indian pandemic from towns to villages. By contrast, during the first wave of COVID-19 between December 2019 and October 2020 [17], only about a third of the infections occurred in rural India, but that increased in 2021 to over 50 percent according to the State Bank of India [18]. By mid-May 2021, 69% of India's 748 districts (the lowest administrative level in the country) reported rates of infection above 10% [19]. The spread of COVID-19 generated interest in RATs as a way of saving the time taken for testing by RTPCR [20] and keeping track of the pandemic [21]. Although many extra RTPCR laboratories were set up throughout India during the first wave, the establishment and running costs of molecular biological testing facilities can be a limitation during times of a raging pandemic. In Bihar and Uttar Pradesh, states where rural healthcare facilities are especially limited, RATs played an even larger role by accounting for 87% and 59% of all the tests, respectively [16]. The cost of a RAT test is almost half or less than the cost of an RTPCR, an added incentive in the poorer states of northern India.

One review of RATs in India stated that they greatly 'helped us ... in detecting and diagnosing COVID-19 at its early stage and also by large scale screening of communities residing in hot-spot areas with high incidence of disease' [22]. The Cochrane Report confirmed the value of all the above factors when using RATs for widespread diagnostic purposes [23]. Another study spoke of the benefits of RATs involving 'triage and emergencies' needing priority diagnoses [24]. RATs have long been used for diagnosing other illnesses, such as pharyngitis caused by Streptococcus bacteria [25] and Bancroftian filariasis [26]. Despite this, before the pandemic, there was a tendency amongst some Indian medical professionals to see RATs as an unreliable diagnostic tool [27]. That view has changed.

Compared with the 'gold standard' of RTPCRs, RATs are less accurate, although they are highly valuable during a rapidly escalating health crisis. One recent study based on 2168 outpatient samples reported a false negative rate from RATs of 4.3% in the first wave of the pandemic and 8.1% in the second wave [17]. It is for this reason that negative test results from RATs still need to be confirmed by RTCPR tests. However, in desperate situations where the status of a person with a possible COVID-19 infection needs to be quickly established, the small margin of inaccuracy in RATs is not a prohibitive factor to their use. Detailed reports about ICMR-approved RATs are only available to state governments, but we found one published study of the first RAT named in the ICMR list of 6 January 2022 [28]. Gupta et al. evaluated the Standard Q rapid antigen detection test (produced by SD Biosensor, Inc., Gurugram) and showed that the test accuracy was 95.4% and the 'overall sensitivity and specificity of the test were 81.8 and 99.6 per cent' [23]. The authors concluded that RATs with these kinds of properties could help alleviate the pressure on emergency departments in hospitals by allowing 'patients showing positive results [to be]... immediately triaged', thereby reducing the risks of COVID-19 infection spreading to other patients and staff in waiting rooms. ICMR then issued an advisory to Indian hospitals saying that the test should be 'adopted in [their] diagnostic algorithms'. Unfortunately, the use of RATs in clinical settings is seriously constrained in rural India, where medical and health professionals are in short supply [29]. In that scenario, the question is whether a self-administered COVID-19 RAT might make the benefits of new diagnostic technologies more accessible to villagers. The need for rapid diagnostic assessments of

COVID-19 infections in rural areas is evident from the situation that emerged from the start of pandemic in India.

## *The Urgent Need for Self-Administered RATs in Villages*

The negative effect of the pandemic has arrived on top of a long-term agrarian crisis in India. Agriculture has not been profitable for decades, and many farmers have been driven to suicide [30]. Poor, small, semi-literate farming families (including those who lease or share crop land, marginal farmers and labourers) have chosen to migrate to the cities for casual employment. When the Indian government declared an immediate national lockdown on 24 March 2020 due to the pandemic, millions of migrant workers found themselves walking home over hundreds of kilometres along India's railway lines, or catching buses, trains or any other possible conveyance. Of the estimated 139 million migrant workers, perhaps a fifth formed a wave of reverse migration as they returned to their natal village homes in search of sustenance and emotional security. Plans to assist them during the pandemic were constrained because the central government had no idea of the scale of the problem due to a lack of statistics. What we do know is that about 6.3 million migrant workers were taken to state transport hubs closest to their villages by special trains between May and August 2020 [31]. There, they were left to walk to their homes, which were often hundreds of kilometres distant. Perhaps three to four times that number travelled home by other means. Back in their villages, they often returned to subsistence farming.

The official data regarding the mortality and morbidity rates of COVID-19 in India are no better than the data on migrant labour, and government estimates have been criticised for their lack of realism. Alternative estimates suggest that the total cumulative deaths in India have been between 2.5 and 7 times higher than the reported number of less than half a million [32–36]. Estimates of the total number of rural deaths *from* COVID-19 are significantly lower than the real figures because India's percentage of medically certified causes of death (MCCD)—based on the signing of the 'Medical Certificate of Cause of Death' by a medical professional—is about 20% [37] of all the deaths at the national level. This low level reflects the fact that the 'Medical Certificate of Cause of Death' is typically signed in urban hospitals and clinics where the majority of Indian doctors are based. What people die of in rural areas is largely unknown unless special surveys are conducted. Thus, we can assume that the rural deaths from COVID-19 accounted for much more than the reported 55% of the total deaths between 2020 and 2022.

The impact of COVID-19 s devastation was revealed on 5 May 2021, showing that, in the preceding week, COVID-19 had become India's leading cause of death, outstripping ischemic heart disease (the previous leading cause of mortality) by more than 2.5 times [38]. In summary, Indian villages remain in desperate need of medical attention during the ongoing COVID-19 pandemic. Given the inadequacies of rural health provision (see Section 4.3.2), self-administered RATs hold out the promise of giving villagers timely and useful information about how to protect their health. Our earlier study of the positive attitudes of villagers to point-of-care blood testing at their doorsteps suggests that what holds back the health of rural dwellers is not their opposition to innovative allopathic technologies but their poverty and lack of access to adequate healthcare (see Sections 4.3.2 and 4.3.3).

## **3. Materials and Methods**

This article employs qualitative data and methods to evaluate the self-administered Indian rapid antigen test CoviSelf.

## *3.1. Qualitative Framework*

Content analysis and discourse analysis are two of many qualitative research methods that can be applied. The former assesses the frequency with which certain words, phrases or metaphors appear in a particular documentary text. This approach would not suit our purposes because we wish to understand the structure of the main document rather than focus on the frequency of particular words. Hence, we employ discourse analysis, where discourse is defined as 'contextually sensitive written and spoken language produced as part of the interaction between speakers and hearers and writers and readers' [39]. In our analysis, the speaker or writer is the producer of CoviSelf and the hearers or readers are the users of it.

## *3.2. Our Main Data Source*

The main document we have analysed is the text of the printed 'patient information leaflet' that is part of the six components of the CoviSelf kit (Figure 1). Throughout this article, we refer to this document as the 'instruction leaflet' or the 'leaflet' [40]. Mylab calls this leaflet the 'user manual'. We downloaded the English language version of the leaflet from the Mylab website [41]. We treat the leaflet as an agent reflecting the urgent diagnostic needs thrown up by the COVID-19 pandemic in India. As the first officially approved, self-administered, Indian made rapid antigen test for COVID-19, the text of this document has its own 'potency as well as capacity' [42]. We argue that the information embedded in the leaflet does not constitute 'inert data' but rather that the data are directed to particular social actors [42] who, in turn, interact with the data when they use the kit. That interaction involves the user of the kit reading the data (in the form of instructions) in the leaflet, interpreting it, interpreting the results of the COVID-19 test and interacting with any external agents who may be involved in the testing process—for example, any person helping the user to administer the test or the Government of India database that seeks to capture the user's test results.

**Figure 1.** The components of the self-administered CoviSelf RAT kit. (Showing in clockwise order from the bottom left-hand side: the plastic bag for disposing of used items, the CoviSelf package, the instruction leaflet and, in the centre, the sterile nasal swab, the test card and the pre-filled extraction tube.) Source: Ms Anusha Kesarkar-Gavankar, photograph of 18 November 2021.

Our discourse analysis involved a close reading of the language of the instruction leaflet by checking for the three things required to effectively communicate the instructions to the consumer: first, we interrogated the text to see whether the assumptions it made about the user's handling of the testing device were reasonable; second, we asked whether the language of the leaflet was clear, consistent and non-technical; and, third, we evaluated those sections of the leaflet that instruct the user about the IT components of the kit. From the start of our analysis, we were struck by the two main characteristics of the leaflet: it addresses both the test component and a special IT feature.

The leaflet was defined by us as the main document because it ends up in the hands of those who purchase the CoviSelf RAT kit. For this reason, we read the text multiple times to cross-check our findings, test out alternative interpretations of the data and resolve any inconsistencies that emerged. In reporting the 'Results' of our analysis below, we arranged our findings according to the sequence involved in using the self-administered CoviSelf RAT in order to place ourselves in the situation faced by a user. In evaluating the leaflet, we engaged in the 'core activity' of discourse analysis by thinking and categorising the '...actions, intentions, characters, events....' [40] as revealed by this document.

## *3.3. Contextualising Our Main Data Source*

An important part of our research method was to acknowledge that the meaning of the leaflet is not self-evident. The reader of the leaflet is not a passive, predictable sponge that merely absorbs the prefigured information provided in the printed text. Rather, particular readers construct the meaning of the instruction leaflet within specific cultural and socioeconomic contexts that vary from user to user. In other words, there is a process of negotiating taking place between the text and context, and it is this that enables the reader to construct meaning. Early in our evaluation, discourse analysis compelled us to ask where the CoviSelf diagnostic device sat within the broader framework of India's IT revolution. For instance, which Indian consumers had smart phones capable of downloading the necessary app referred to in the leaflet? Mylab's press release of 10 May 2021 stressed the benefits to Indian citizens of the kit's features because they could now buy the diagnostic device 'without a prescription from local pharmacies and online channel partners', and, in using the kit, the consumer would promote the traceability of the pandemic through the company's 'AI-powered mobile App' [3]. Our 'Results' below question the assumption that lies behind Mylab's statement. We ask, does 'every Indian and any citizen' have access to the information technology that Mylab has taken for granted?

## *3.4. Our Second and Third Data Sources*

These questions led us to the second data source—a group of documents produced by the Indian media, professional associations and the Supreme Court of India about India's digital divide. The digital divide stressed the disadvantages suffered by rural residents in accessing electronic information.

Following the logic of discourse analysis, we then widened the context for understanding the CoviSelf leaflet even further by using a third data source, namely our own 'expert opinion', as documented in peer-reviewed, published research about living conditions in Indian villages. Fieldwork in central India before the pandemic showed us that villagers were receptive to point-of-care blood testing at their doorsteps [43]. We assumed, therefore, that there might be positive responses to the idea of using the self-administered CoviSelf RAT kit because the demand for modern medical interventions is driven by the desire to prolong longevity. This may well be a universal motivation, but our findings remind us that the specific cultural context, in Prior's words, renders the 'unremarkable, remarkable' [42]. With this third data source, we explored the interplay between the digital divide in rural India and the multiple constraints faced by villagers in accessing timely health and medical care.

## *3.5. The Limitations of This Study*

With the ongoing COVID-19 pandemic, it has not been possible to undertake fieldwork because the desperate circumstances facing rural residents prevents them from participating in time-consuming, in-depth interviews. This is the main limitation of our present study. On the other hand, a discourse analysis of the three data sources (specified above) allowed us to refine our research questions and hypotheses for future research on village responses to the pandemic. In-depth interviews in rural areas during the next two years will constitute

the fourth and final data source for an expanded study of the current paper. The locus of this extended study will build on our existing work in Wardha District (Maharashtra), the site of our research on point-of-care blood testing technologies. That earlier period of fieldwork was informed by our prior analysis of classical and modern Hindu texts that suggested that there were unlikely to be any cultural objections to modern diagnostic blood technologies. Our hypothesis for further fieldwork (in 2022–2023) on self-administered RATs in rural India questions the value of CoviSelf in villages. As a result of the present paper, we have come to the view that self-administered RATs for detecting COVID-19 infection would be more suitable to villagers if they did not include an IT component and if they were free. This hypothesis is not based on any questions about cultural resistance to the innovative CoviSelf device but rather doubts about its current practicality and cost. The urgent need for self-administered RATs in rural India suggests that investing in the redesign of this diagnostic device is well worthwhile.

## **4. Results**

Our results are discussed under three main headings. Firstly, we report the findings from our discourse analysis of the main document, namely the kit's printed instruction leaflet. In the second part, we explain the findings from our analysis of the data regarding India's digital divide and its impact on rural India. In Section 3, we use our own data from previous studies of Indian villages. All three parts locate the CoviSelf kit within the wider socioeconomic context of Indian villages in conformity with discourse analysis, which seeks to understand how particular readers construct the meaning of different kinds of documents.

## *4.1. The CoviSelf Instruction Leaflet*

Our analysis of the language of the leaflet focuses on four issues affecting the practicality of this self-administered RAT, but we begin our discourse analysis by considering the structure of that document. After that, the next two findings deal with the processes by which a test result is obtained, and the last two findings report on different aspects of the IT components of the kit. The data take the form of the leaflet's instructions to the reader. Where appropriate, short comments are made about the practicality of the text in terms of its limitations and benefits.

## 4.1.1. The Structure of the Instruction Leaflet

The layout of the leaflet is an important consideration in our discourse analysis because it tells us how the information provided to the user has been structured by Mylab, i.e., the order of priority given to various points. The leaflet takes the form of a folded, double-sided sheet of printed paper measuring 21 cm × 57 cm. One side is in English and the reverse side in Hindi. The leaflet is also available in eight other Indian languages representing different linguistic regions. The online version of each leaflet is identical in information and size to the copy included in the CoviSelf kit. The text is accompanied by diagrams illustrating all the instructions and how to read the positive, negative and invalid results. The only exceptions are the text in column one (both top and bottom panels, see below) and the bottom part of columns seven and eight, stating the FAQs (frequently asked questions), the limitations of the procedures, Mylab's contact details and a QR code that can be scanned to access a video demonstration about how to use the kit.

Both sides of the leaflet are divided into eight columns, each with two panels (top and bottom). When folded, the leaflet measures about 10 cm × 7 cm. Column one, top panel, is taken up by the front cover of the leaflet, and the panel beneath it explains the kit's intended use, kit storage and stability and the principles of the test; this last section explains some of the science behind the test device. Column two, top panel, is headed 'Kit Contents' and names the six components that make up the kit. The panel beneath that explains the 'test preparation', which includes washing and thoroughly drying the hands. The explanatory video that can be downloaded by some users says that a table top should

be sanitised so that the contents of the kit can be laid out [44]. No such instruction appears in the leaflet.

Between the top and bottom panels is a central box that states: 'Download the Mylab CoviSelf App from Play Store/App Store'. Column three, top panel, instructs the user to open the CoviSelf App and activate it; the bottom panel has instructions for preparing the pre-filled extraction tube. Column four is named 'Step 1' and instructs the user how to collect the nasal sample. Column five, designated 'Step 2', has instructions for placing the swab into the pre-filled extraction tube. 'Step 3' appears at the top of column six and tells the user how and where to place the sample on the test card. 'Step 4', beneath 'Step 3', asks the user to take a photo of the test card and '*wait for the App to analyse and display your Covid-19 test results*' (authors' italics). Under 'Step 4' appear instructions for disposing of the waste once the test has been completed.

Columns seven and eight, the leaflet's last two columns, have two sections: the top explains the appearance of the test card when positive, negative or invalid test results appear. The bottom section provides additional information to the user: three FAQs are listed, five limitations to the procedure are stated and there is a QR code that can be scanned if the user wants to watch a video demonstration of how to use the kit. The bottom strip across both columns gives the name of the manufacturer and contact details.

Our analysis of the structure of the leaflet suggests that the order in which the instructions are arranged encourages the reader to use the IT component and to depend on it for 'analysing' the test results. Instructions about using the IT features are given at the start and end of the leaflet. The first instruction appears prominently in the middle of column two and asks the user to 'Download the Mylab CoviSelf App'. This is followed by instructions in column three saying '*Open the Mylab CoviSelf App and fill in the credentials*'. After that, the user is told to scan the QR code on the test card. More significantly, 'Step 4' is presented to the user as the last step in the sequence of actions needed to complete the testing procedure. It asks the user to take a photograph of the test card and '*wait for the App to analyse and display your Covid-19 test results*'. However, the test card has already completed both of these functions: it has analysed the nasal sample thanks to the bioactive ingredients used in the test process and it has displayed the test result. In the instruction leaflet, there is a section that explains how the test card tells users whether they have a positive, negative or invalid test result. However, these explanations appear at the end of the leaflet, just before the FAQs section in columns seven and eight. A more logical order would have been for information about the test results to be labelled 'Step 4'. The function of uploading a photograph of the test card to the app could then have been labelled as an optional 'Step 5'. This new 'Step 5' would also need to give the user an explanation for why the photograph of the test result should be uploaded to the app. In other words, the user should be given the choice of taking or not taking a photograph of the test result that has appeared on the test card. Instead, the instructions and current layout of the leaflet create the misleading impression that the IT functions of the kit are an integral part of the testing process. The true purposes of the IT functions are analysed below (Section 4.1.5). We also address the risks that these pose.

4.1.2. The Ease of Using the Kit (the Relevant Headings in the Instruction Leaflet Are Steps 1, 2 and 3)

After scanning the QR code on the test card, the user is told to remove the pre-filled extraction tube from its packaging, hold it erect and tap the tube so that the 'extraction buffer settles at the bottom of the tube'. The user is warned to be careful not to spill the contents of the pre-filled extraction tube—but no reason is given for this, so the user has no way of knowing the critical nature of this instruction. The fluid in the tube contains the reagent needed for testing the nasal sample. 'Step 1' then follows: it involves removing the swab from its packet by holding the tail end opposite the swab head. The swab head goes into the nostrils and so should not be touched because that could risk contamination. It is rolled five times in each nostril to collect cell samples.

Removing the swab from its package with one hand may be difficult if the user is already holding the pre-filled extraction tube in the other hand. Thus, there is a risk of spillage and contamination if the tube or swab are put down on a non-sterile surface. A second person could help by holding the pre-filled extraction tube, but, if that person has not washed and dried their hands thoroughly, there is another risk of contamination. In any case, instructions about this alternative approach are not given in the leaflet; it is assumed that the user is nimble.

'Step 2' comes next. On removing the swab from the nostrils, the swab head is inserted into the pre-filled extraction tube. The bottom of the tube needs to be pinched while the swab stick is swirled ten times to ensure that the nasal sample mixes properly with the pre-filled liquid. After that, the swab stick needs to be broken 'at its breakpoint', but this point is hard to see. The lower half of the swab, with the swab head, remains inside the tube. The top broken off part is placed into the disposable bag to be thrown away at the end of the testing process.

Juggling these different tasks might not be so hard for an experienced user, but firsttime users risk spilling the tube's contents, contaminating the swab head or not collecting sufficient material from the nostrils. Nevertheless, supposing all has gone smoothly, the swab head is now inside the pre-filled extraction tube, which is then sealed with the attached cap. The test can now move to 'Step 3'. The user is instructed to press the bottom part of the tube and place two full drops of the liquid into the sample well at the far end of the test card near the letter T (in Figure 2, the well is located on the far right of the image). Within 10 to 15 min, a test result should appear in the test window.

**Figure 2.** The CoviSelf test card. (From the far left: the QR code, the control point C, the test point T and the sample well for receiving the nasal mixture). Source: Ms Anusha Kesarkar-Gavankar, photograph of 18 November 2021.

While waiting for the test results, the user can place all the discarded items into the disposable bag and 'disinfect all surfaces that the specimen may have touched' in addition to washing their hands after throwing the bag into household waste (bottom of column six). Soap and water are the best disinfectants against the coronavirus, and soap is readily available in Indian villages. However, what villagers might not know is that effective cleaning requires the user to scrub their hands 'for at least 20 s' [45].

4.1.3. Reading the Test Result and the Clarity of the Leaflet's Language (the Relevant Headings in the Instruction Leaflet Are Positive Result, Negative Result and Invalid)

The user can read the results on the test card, which has two letters in the centre beneath the test window: C and T. C is the 'control' point and T is the 'detection' point. If the test result is *positive*, two lines appear in the test window located in the centre of the card—one line next to the letter C and the second line next to T. The T line can be any shade of pink or purple, but all shades, no matter how light, indicate a positive test result. If only one line appears next to the letter C and there is no line next to the letter T, it means that the test result is *negative* for the virus. If no line appears in the test window next to the letter C, the result is invalid. The text should, however, clearly state that the test has failed.

The language in which the above information is expressed in the leaflet is complex and technical. For example, in the case of a positive test result, the leaflet reads: 'If both the quality control line 'C' and the detection line appear, novel coronavirus antigen has been detected and the result is positive for antigen'. Antigen is not a well understood term in most countries, and neither is the notion of 'detection line' clear. However, the labels next to the diagrams illustrating the appearance of a positive and negative result are better: the 'detection line' is described as the 'T test line'.

If a negative test result appears, the 'symptomatic' user is advised to immediately proceed with an RTPCR test because 'RAT[s] are likely to miss [a] few positive cases presenting with a low viral load'. We have inserted the word '[a]' into this quotation because, without it, the actual sentence says the opposite of what is intended. This problem in the English language version of the leaflet does not, however, occur in the Hindi version.

The leaflet also uses inconsistent terminology that can confuse users. For example, the test card is also called the test device or cassette, the pre-filled extraction tube is also called the pre-filled buffer tube and the liquid in the tube is called an extraction buffer and the antigen buffer.

4.1.4. How to Respond to the Test Results (the Relevant Headings in the Instruction Leaflet Are Positive Result and Negative Result)

From the above account, it is obvious that the CoviSelf kit is a stand-alone device that has the capacity to tell the user whether they are or are not infected with the COVID-19 virus. However, the kit contains another component—an IT factor, which is problematic beyond our analysis of the misleading importance given to the app in the instruction leaflet (see Section 4.1.1).

The leaflet states that all positive results are 'true positives' and the user is advised to follow 'home isolation and care as per the ICMR and Ministry of Health and Family Welfare protocol'. A website address with further instructions is provided for the user. All users lacking access to the internet will be excluded from further information—and we are speaking of millions of Indian citizens.

The advice is clearer in the case of a negative result. Here, the leaflet tells a user who has symptoms but receives a negative result to isolate at home and get an RTPCR test as soon as possible. Despite the misleading English language text explaining why more testing is required (see Section 4.1.3), users are familiar with the acronym RTPCR because, during the pandemic, such tests have become a routine procedure for hospital admissions. Nevertheless, users may not see the point of having an RTPCR test if their CoviSelf test results are negative. There is no compelling explanation in the leaflet to justify the expense and inconvenience of visiting a clinic or hospital to get an RTPCR test. The leaflet only states that a low viral load could mean that some positive cases are missed, but the words 'low viral load' are not automatically meaningful to most readers, who will not know about the science of virus testing.

More importantly, the user is not encouraged to take *another self-administered test* if the first result is negative. If the viral load is low at the time of the first test, a second test could pick up a true infection if the viral load has increased. There has now been considerable research and experience to show that, in the words of Richard Hatchett, 'the antigen tests are less sensitive if you give just one. But if you can do it in a sequential way, they become cumulatively as sensitive as a PCR' [46].

If the user is a villager, the advice to have a follow-up RTPCR test involves a timeconsuming and expensive trip to an urban medical institution (see Section 4.3.2 below). 'Isolating at home' in the case of symptomatic individuals is also advice that is virtually impossible to follow as, in most villages in India, a whole family lives in a single room. In such cases, isolation at home actually increases the risk of all family members becoming infected.  In India, recognizing the symptoms for COVID-19 is also difficult because the virus mimics the signs for many other illnesses—for example, other respiratory conditions accounted for a total of about 15% of Indian deaths between 2010 and 2013 [47] and much higher levels of morbidity.

4.1.5. The Test Results and the Objectives of the ICMR (the Relevant Section in the Instruction Leaflet Is Step 4)

Instructions about the IT component of the kit are stated at the beginning and end of the leaflet. Prior to conducting the test, the user is asked to download the Mylab CoviSelf app from either Google Play or the App Store; a mobile phone message is then received asking the user to enter their personal details and scan the unique QR code printed on one end of the test card (on the left hand side of the image in Figure 2). The personal details are: name, age, gender, address, pin code, mobile and Indian ID number (or *Aadhar* card number). Except for the Indian ID number, which is an 'optional' detail, all the information is stated as being 'mandatory as per ICMR guidelines'. Once the user has completed these tasks, they are ready to conduct the test. The result should appear within 15 min. The user is then asked to take a photograph of the test result and '*Wait for the App to analyse and display your COVID-19 test results*'.

There are two problems with these instructions. First, making the personal details 'mandatory' sounds ominous even if ICMR has no way of enforcing this. In Indian villages, such statements are taken seriously (see Section 5 'Discussion' about '*sarkar*'). Secondly, it is misleading to state that the test result for COVID-19 infection is being analysed by the Mylab app. As already explained above, the result of the test appears within 15 min on the test card. What the leaflet should be saying is that, once the test result has been photographed, it is uploaded to Mylab records and then forwarded to the ICMR database, which forms part of the records of the Government of India. Equally important, there is no provision in the leaflet for the user to give their consent to sharing their personal details or test results with the government.

The role of ICMR is only explained on the Mylab CoviSelf website, where there is a section headed FAQs. From this, we can learn about the destination of the uploaded test result in response to the following question: 'What are the benefits of reporting the result to ICMR?' The following answer is given [48]:

ICMR is the regulatory body in India which is responsible for the curb of COVID-19 pandemic along with other important regulatory bodies. Reporting the results helps the body and authorities curb the spread of the disease. It thus becomes our moral obligation to help ICMR by reporting our test result data.

It is significant that the above FAQ and answer do not appear in the user leaflet, although the FAQ section in that leaflet takes up a total of 20 lines, suggesting that there was enough space to include the five lines about the role of the ICMR database. Helping the government to collect better records of COVID-19 infection is a worthy objective, but it could end up being counterproductive to do so without the prior agreement of the user (see Section 5 'Discussion'). Moreover, it is just as important to protect the right of Indian citizens to keep their personal details private and their medical records confidential [49].

From the user's perspective, is there any benefit to activating the IT component of CoviSelf? The analytical methods of discourse analysis tell us that the answer to this question depends on who the user is.

After the user sends a photograph of the test result to the Mylab CoviSelf app (and ultimately to the ICMR) via their mobile phone, they receive a notification of their COVID-19 status. This can be beneficial for several reasons. First, it might reassure those users who are uncertain about their reading of the results on their test card. The response from the Mylab app gives them a statement about their test result, and this validation enables them to act accordingly. Second, many middle-class Indians have used this statement as evidence of their COVID-19 negative status when boarding domestic flights and entering hotels. Such certification has enabled some professionals and holiday seekers to be mobile inside India [50]. One hopes that such travellers have been cautious and only travelled when they were asymptomatic. However, as already noted, everyone's tolerance for, and interpretation of, 'symptoms' differs.

These benefits are counterbalanced by the multiple problems that arise from trying to use the IT components of CoviSelf. Many urban users have complained about the difficulties (see Section 6 'Conclusions'). It is regrettable that the instruction leaflet conflates the two components of CoviSelf, i.e., the test and the IT functions, because this gives users the impression that the kit will not work without activating the IT features. In rural India, the inadequate provision of healthcare services suggests that the potential value of a self-administered COVID-19 RAT is high, but the IT component compromises the device's usefulness for the reasons we explain in the next two sections.

## *4.2. India's Digital Divide*

India's digital divide is part of a global problem and influences the value of the CoviSelf kit because it contains an IT feature. In 2002, the Secretary General of the UN spoke of the digital divide as the widening gap between the 'haves' and 'have nots' in accessing the new information and communications technologies and the dangers this posed by excluding the world's poor from the 'emerging knowledge-based global economy' [51]. The key technologies for such access were the internet and mobile phones. Two years earlier, UN members agreed that one of the Millennial Goals was to 'ensure that the benefits of new technologies, especially information and communication technologies, are available to all'. The data we have used to provide the general context for understanding the implications of this digital divide for CoviSelf come from the following sources: the Indian press, a 2021 report by the Internet and Mobile Indian Association (hereafter IAMAI) and a court case by the Software Freedom Law Centre before the Supreme Court of India in 2021.

## 4.2.1. The Indian Press

The phrase 'Indian press reports about India's digital divide' revealed over 46 million hits on the Google search engine. A report in 2019 from the *Economic Times* stated that India had an estimated 450 million smart phone users and about 550 million users of 'feature phones' [52]. A feature phone is defined by Kantar IMRB/MMA as 'A mobile phone that incorporates a fixed set of functions beyond voice calling and text messaging such as limited web browsing and e-mail, ability to play music *but generally cannot download apps from an online market place*' [authors' emphasis] [53]. Whilst most mobile feature phones allow users to access the internet for entertainment, they typically lack the capacity to support complex internet apps, such as the one used by CoviSelf. This places the innovative CoviSelf device beyond the majority of Indians despite the hopes that Mylab has of reaching all citizens.

## 4.2.2. The Internet and Mobile Association of India (IAMAI) Report of 2021

A more fundamental problem is the unreliability of telecommunication signals in many parts of India, especially rural areas [54]. The 2021 IAMAI report on internet usage in urban and rural India shows unsurprisingly that states with relatively good telecommunications infrastructure have a higher percentage of active internet users (hereafter AIUs) amongst their population (e.g., 61% in Maharashtra) than states lacking such facilities (e.g., 24% in Bihar) (Table 1). This reflects the level of urbanisation: 45% of Maharashtra's population live in towns and cities compared with 11% in Bihar [55]. For the whole of India, AIUs represent about 31% of the rural population compared with 67% percent of urban residents (Table 1). The percentage of people who have what is often called 'connectivity' in India may be less than this as many AIUs might have more than one phone and more than one internet account.

**Table 1.** Internet usage in urban and rural India in 2020 (based on active internet users (AIUs)).

Source: Collated from Kantar, *Internet Adoption in India: ICUBE 2020*, Internet and Mobile Association of India/Kantar: Delhi, India, June 2021, (https://images.assettype.com/afaqs/2021-06/b9a3220f-ae2f-43db-a0b4-3 6a372b243c4/KANTAR\_ICUBE\_2020\_Report\_C1.pdf, accessed on 20 February 2022).

## 4.2.3. Civil Society and the Supreme Court of India

The exclusion of the rural poor from internet access is well known within Indian elite circles, yet, despite this, the Government of India created the CoWIN app in early 2021 to make it easier for Indian citizens to get appointments for vaccinations against COVID-19. The idea behind this innovative app was to stop citizens standing in long queues for vaccinations—thus saving time, reducing the risks of cross-infection and taking the pressure off clinics, hospitals and vaccination centres. Unfortunately, when the new system of appointments was announced on 18 April 2021, it also became mandatory for all Indians aged from 18 to 44 to make online appointments via the CoWIN app. The Software Freedom Law Centre (hereafter SFLC) in New Delhi wrote to the Indian Ministry of Health objecting to the mandate and, after being ignored, raised a Sou Moto case in the Supreme Court of India against the ministry [56]. The SFLC argued that the CoWIN app [57] failed to address the 'digital exclusion' of Indians who lacked appropriate mobile phones and internet access and it also failed to protect the privacy of citizens [58]. Justice Chandrachud (Supreme Court) described the government's assumptions behind the CoWIN portal as 'far-fetched' and 'exclusionary of the rural areas' [59].

The objections against the CoWIN app (before the mandatory requirements were removed) apply equally to the IT component of CoviSelf. However, there has been no public outcry against CoviSelf because buying the device in India is a matter of personal choice rather than something mandated by the state. Nevertheless, this should not blind us to the fact that the digital divide deprives millions of citizens from the benefits of this self-administered RAT. The government's failure to address the needs of villagers in the provision of self-administered RATs is inconsistent with its strong campaign to improve rural hygiene since 2014 through the Swachh Bharat Mission to end, for example, open defecation and the risk of faecal infection [47]. This contradiction in the government's rural development programs cannot be addressed in this paper because the topic is too large and complex, but, in the following section, we consider the practicality of CoviSelf for Indian villagers.

## *4.3. The Conditions of Life in Rural India*

In extending the context in which to understand the benefits and limitations of CoviSelf, we have drawn on our own research published in peer-reviewed international journals and books. Vicziany has published on the poverty and marginalisation of *Dalits* (former 'untouchables') [60], the family planning program [61,62], food security [63–65], Koli villages in Mumbai [66] and rural health [29,43,47], including work on the potential of point-of-care blood testing in villages [29,43]. Hardikar has reported extensively on questions of rural poverty, co-authored work with Vicziany on poor farmers, agriculture and health [29,43] and has published two books on India's farming crisis [30,67].

## 4.3.1. Village Poverty and Employment

The bulk of our research has dealt with poor, semi-literate people in marginalised and socio-economically disadvantaged households. Our most recent research in Wardha District (Maharashtra) involved 36 in-depth interviews with farming groups defined by various criteria: a high proportion of families living below the poverty line, households with very little irrigated land, poor farmers, landless villagers and tribal or *Dalit* families [43]. Our findings showed that poor rural households are preoccupied by three major concerns. Their first priority is to secure a source of income through any kind of employment- even if they farm; supplementary sources of income are important, in particular through shortor long-term migration to urban centres. Those who remain in villages seek to diversify their income locally—for example, by serving as government agents for the public food distribution system, working as linesmen for electricity companies or providing local taxi services.

Our understanding of the drivers of poverty in rural India and the constraints it imposes on the capacity of villagers to avail themselves of the benefits of modern technologies was confirmed by the Supreme Court's judgement in the CoWIN case (see Section 4.2.3). The judges noted that the minimum internet tariff plan would equal '4–5% of the month's income' of urban and rural people living below the poverty line [68]. Accessing internet data is, therefore, something that they cannot afford given that the poor are also heavily indebted. The burden of poverty extends into all aspects of life, including the lack of consumer power to buy the CoviSelf kit that retails for about Rs. 250. Given that the daily wage for a farm labourer at the top end of the range of agricultural jobs (i.e., ploughing and tilling) is about Rs. 365 [69], the cost of CoviSelf is equal to 68% of the daily wage that sustains a whole family. For the unemployed and those living below the poverty line of Rs 1316 and Rs 896 *a month* in urban and rural India, respectively [68], the expense of the CoviSelf kit is unthinkable. Moreover, the benefits of all self-administered RATs are greater if used more than once over a number of days to check the onset and end of infection. Such costs are beyond the means of India's poor whether they are marginal farmers, labourers, the under-employed or the unemployed.

Our research on rural poverty is supported by other quantitative data showing that 27.9% of India's population is suffering from multidimensional poverty [70], defined as multiple and overlapping conditions of deprivation. The MPI (multidimensional poverty index) goes beyond previous attempts to measure poverty using simple headcounts and monetary estimates of the poverty line. As a result, the MPI replaced the Human Poverty Index (HPI) of the United Nations Development Programme in 2010. The data show that interstate variations in poverty in India are stark and support our previous reference to the large regional disparities between, for example, Maharashtra and Bihar. The proportion of the population in these two states that is multidimensionally poor is 15% and 52%, respectively [71]. Poverty weighs especially heavily on disadvantaged social groups throughout India (such as the *Dalits* and tribal people), with five out of six Indians belonging to these communities suffering from multidimensional poverty [72]. Rural economic insecurity is the overall context in which we need to evaluate the practicality of CoviSelf, the affordability of mobile phones and internet access and the future of self-administered RATs for villagers.

## 4.3.2. Village Health and Diagnostic Medical Interventions

The second priority of Indian villagers is their health. India's medical system is a plural one, ranging from trained allopaths (i.e., doctors trained in the 'western' medical system) and Ayurdevic-Unani practitioners to herbalists, soothsayers, spirit men, midwives, gurus, spiritual mediums and 'quacks'. The Maharashtra Medical Association, for example, has been engaged in a campaign to marginalise all but allopaths, yet the plurality remains because there are insufficient allopaths willing to serve in rural areas. Even in states that have seemingly adequate rural health infrastructure (such in Maharashtra), the human resource constraints are serious. The result is that villagers bypass government rural medical institutions and travel considerable distances to clinics and hospitals in or near towns [43]. While this is costly, rural residents have the satisfaction of getting a more timely and reliable diagnosis of their condition, a prescribed treatment regime and admission to hospital in urgent cases.

The healthcare sector is now dominated by private services, with many hospitals being part of larger systems of education involving private medical colleges. Providing medical training and degrees has become a lucrative business in India and attracted many entrepreneurs. The appearance of the private health sector has resulted in villagers experiencing a medical revolution, with entrepreneurs working to make their hospitals more accessible and affordable to the poor by offering 'membership cards' and using their political links to help residents in need of attention. In return, these entrepreneurs and politicians are rewarded when their beneficiaries vote for them in elections [43].

Yet, despite this, the shortage of medical and health professionals in rural areas has not been alleviated. The private entrepreneurs that have developed western based medical institutions are urban based. In the government's rural health infrastructure there are large human resource shortfalls and vacancies relative to requirements [73]. At the village level in India, health care is left to what can best be described as a pre-modern private sector. One rare study reported that the village level private sector accounted for 68% of local healthcare and that 75% of villages had at least one health care provider. However, of the 3373 providers surveyed, only 8% were allopaths with an MBBS qualification and 24% were AYUSH providers with degrees in alternative medicine, namely in Ayurveda, Yoga, Unani, Siddhi and Homeopathy. The remaining 68% lacked any formal training [74]. In other words, the size of the private sector reflects the poor relationship that exists between villagers and government funded institutions. It does not mean that the private sector provides better health and medical services, even if these are more accessible.

For all the above reasons, villagers with life-threatening health problems go straight to the nearest hospitals, even though these are expensive. 'Free' treatment in government institutions, subsidised health care in the private sector and the support of an emerging insurance system have failed to reduce the out-of-pocket (hereafter OoP) expenses for inpatients and outpatients. Our study of the potential for point-of-care blood testing at the village level showed that there are many potential rural innovators because OoP costs are too high. Of the 36 villagers we interviewed between 2017 and 2018, 61% were receptive to the idea of introducing simple blood tests into their villages [43]—using bioactive paper, such tests would require only a few drops of blood and allow villagers to avoid the many costs incurred by travelling to town-based institutions. More generally, we discovered that medical innovations do not come as surprises to Indian villagers; their receptivity to blood tests at their doorsteps reflected their experience with modern medical practices, ranging from complex procedures, such as liver transplants and transfusions for accidents through to regular blood testing and treatment for genetic blood diseases [43].

Rural residents place a high value on their health, and, because of this, they are willing to spend large amounts of money on medical diagnoses and treatments to address their illnesses and increase their longevity. Many rural households go into debt to cover medical expenses. There is no evidence in the available literature or our fieldwork to suggest hesitancy arising from their distrust of modern medicine; rather, they may be distrustful of the government institutions charged with the delivery of rural healthcare.

## 4.3.3. Village Religion and Modern Medicine

The third preoccupation of villagers is to ensure that their family and local religious traditions are respected, including the worship of goddesses. The COVID-19 pandemic has seen the proliferation of Corona goddesses. This could easily be misunderstood by non-Indian observers thinking that here is a contemporary example of Indian fatalism and attitudes opposed to allopathic medicine. However, our research showed that goddess worship, including blood sacrifices to appease village and household deities, sat comfortably alongside the demand for modern, scientific health interventions. One of the villagers that we met explained that her husband had died of alcoholism despite the fact that a goat had been sacrificed to the family deity on his birth. We asked her why the mother goddess, Firasti-Aai, had not provided her husband with lifelong protection against ill fortune. She exonerated the deity and blamed her husband:

He had good health but he brought it upon himself by drinking too much. How is our deity responsible for that? [43].

Powerful religious beliefs, in other words, do not cancel out personal responsibility. We found no evidence of any clash between local religious traditions and innovative medical practices.

The prudent course of action for believers wishing to prolong their lives is to propitiate the goddesses and simultaneously address their ill-health by resorting to appropriate medical solutions. Since independence in 1947, Indian governments have also promoted widespread immunisation for infectious diseases, and this has contributed to a decline in death rates. However, despite great demand in rural India, the present pandemic has been characterised by shortages of vaccines, oxygen, hospital beds, masks and drugs, such as remdesivir. There has not been any cultural resistance to these measures. For most of 2020 and 2021, vaccinations against COVID-19 in rural India lagged seriously behind those in urban areas, demonstrating the 'urban bias' that Indian development policies suffer from. By mid-May 2021, only about 15% of rural residents had received at least one dose of vaccine [75], partly because urban–rural disparities cause India's cold chain to be unevenly distributed [76]. By early 2022, however, the vaccination gap had closed thanks to a new focus on villages, although urban–rural differences remain, with 79% of eligible urban residents fully vaccinated compared with 69% of rural dwellers in late January 2022 [77].

## **5. Discussion**

The unique status of the India-made, self-administered CoviSelf RAT kit is based on official approval by ICMR. In the context of a pandemic, that places a special responsibility on the government to ensure that this diagnostic device can benefit all Indian citizens. The IT component of the kit fails this basic test because millions of people in India do not have access to the internet and thus the app embedded in the CoviSelf kit is out of reach for them. India's digital divide has prevented universal internet access because there are too many poor households that lack the financial resources to buy sufficiently sophisticated mobile phones or pay for online information. There is also the matter of the privacy of the user for, as explained above, personal details are being entered into a Government of India database without prior consent from the individual users of CoviSelf. These realities fly in the face of Mylab's objective of bringing CoviSelf to the 'doorstep of every Indian ... any citizen', as stated in the company's press release of 10 May 2021 [3].

Might the official approval of CoviSelf be defended by the argument that the privacy of the poorest Indian citizens cannot be compromised if they are unable to download the Mylab app or upload their test results?

Such a defence makes no sense for a number of reasons. First, villagers come and go between villages and towns, and they might well purchase a CoviSelf kit thinking that it will help to protect their families from infection. The poor are attracted to innovations that hold out the promise of a better life, and unsuitable decisions can be made as a result. Second, even if they cannot read the CoviSelf leaflet, villagers can go to a local 'fixer', who, for a small fee, will read the leaflet and try to handle the IT component of CoviSelf on their smart phone. This 'solution' has been reported for rural residents accessing other government apps [54]. Should this happen, the villager becomes caught up in all the limitations of the kit. When the fixer follows the leaflet, the first thing they will read is the instructions for downloading the Mylab app and the last thing they will read will be about sending the image on the test card to Mylab. The fixer might not realise that the test result is before them on the test card and that nothing further needs to be done to find out whether the user has tested positive or negative for the virus.

The self-administered CoviSelf RAT kit, therefore, fails two tests of importance in the world's largest democracy: the right of all Indians to access new technologies approved by the Indian government and the right to privacy. The IT component of CoviSelf carries an additional risk, namely that it could create suspicion about the real purpose of this self-test diagnostic device and also about other government-approved medical technologies. If the government hopes to contain the spread of COVID-19 and harness self-administered RATs to that end, it should consider using local hospitals with good reputations in rural areas. For example, in Wardha District, we discovered that the Kasturba and Datta Meghe Hospitals are highly regarded by villagers for their excellent services, including rural outreach work [43]. It would not be difficult to involve the professionals from such institutions in the promotion and use of CoviSelf. Such professional backing could also help to disseminate more accurate information about the nature of COVID-19 and how to avoid infection. Trusted health professionals could explain to villagers the health risks posed by *symptomless* individuals who are nevertheless carrying COVID-19. Normally, villagers do not seek diagnosis or treatment for symptomless family members, as we know from the failures of the government's anti-TB program [78].

The prerequisites for such outreach are twofold: make the kit free to all citizens and remove the IT component. Given the multidimensional nature and extent of Indian poverty, the first point is significant. The second point also matters if CoviSelf is to be trusted by villagers. The present design of CoviSelf is likely to raise the suspicions of villagers: if medical professionals are seen entering a villager's personal details into a smart phone, it would soon become widely known that the test result and private information were being sent by Mylab to the Government of India. The Hindi word for government is '*sarkar*', and, amongst India's poor, it inspires fear rather than confidence. For complex historical reasons, '*sarkar*' has never been a trusted institution in India. The fear would be not only about the possible uses of such data but also anxiety regarding some kind of government or police action in the event that they test positive.

## **6. Conclusions**

Our evaluation of India's first officially approved self-administered COVID-19 RAT kit uses the framework of discourse analysis to ask whether CoviSelf is of any practical value in rural India. Drawing on data from a wide range of qualitative, textual sources, we have identified a number of significant problems with the language of the instruction leaflet and the assumptions it makes about the living conditions in rural India; specifically, its promotion of the IT features of this diagnostic device betrays a lack of insight into why India's 'have-nots' have been excluded from the information technology revolution.

The key lesson emerging from our evaluation is that, if CoviSelf is to be made more practical and accessible to Indian villagers, the IT functions should be removed and the kit made cost-free to users. Such changes would also automatically protect the privacy of the user and their test result. The ambitions of Mylab could well be realised by such adjustments, namely the production of a self-administered, Indian-made COVID-19 diagnostic device for every Indian. In removing the embedded IT features of CoviSelf, the instruction leaflet would need to be rewritten, and that would provide an opportunity to address the other language problems that we found in the text.

Such changes would also help frustrated urban middle-class users whose struggles with the kit are recorded on the Google Store website. Many users wrote that the kit was a waste of money, some suggesting that it was a scam, others that the instructions were wrong and that the test card was not working properly. One frustrated customer (Ramana G) wrote on 1 November 2021 [79]:

I had to take three tests and the result is still inconclusive, what is the problem. The 1st test ... I couldn't squiz [*sic*] the liquid onto sample level and I broke the well bottom and the lines so I exited the App. But then I tried the second time and did everything right and found that results were inconclusive as I scanned later than 20 min... I tried third time and exactly waited for 15mins and it's the same problem.

Some three months later, another customer (Arjita Mukherjee) on the same website commented:

I followed all instructions yet I didn't receive my results. Just said Invalid Casette, low server problem. Second, I understand that there might be server issue, then what's the point of keeping another 15 min time interval when the whole kit gets invalid after 20 min. Just keep the test result appearance that's it. Really disappointed. The manual rtpcrs are far safe and better I suppose.

This kind of feedback allows us to predict a villager's likely experience with CoviSelf in its present form. The online customer reviews confirm our evaluation of the instructional leaflet—namely that there are serious communication problems with the language in the leaflet, the instructions to users and the IT features.

The potential for self-administered RATs in Indian villages, especially in the face of the highly infectious Omicron strain, remains undiminished. The majority of Indians live in rural areas where access to timely medical diagnosis, assistance and drugs is seriously constrained. Modifying the CoviSelf kit to remove the limitations that we have identified in this paper could assist with its equitable distribution as an effective, risk-minimising device against galloping infections in villages. Richard Hatchett has given a prescient warning at the height of the Omicron wave: 'unpredictably, the virus appears to have the capacity to become, essentially, a pandemic at any time ... access to diagnostics—and updated diagnostics, are absolutely critical to managing an infectious disease crisis' [46]. Given this, the distribution of self-administered RATs such as CoviSelf should accompany vaccines and anti-viral drugs as an essential part of India's pandemic planning.

**Author Contributions:** M.V.: lead author; conceptualisation; methodology; formal analysis; resources and data curation; original draft preparation; writing, review and editing; supervision; project administration; 40 years of fieldwork experience in India. J.H.: validation; data curation; review and edition; 20 years of fieldwork experience in India. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research received no external funding.

**Informed Consent Statement:** Not applicable.

**Acknowledgments:** We are very grateful to Leon Piterman (General Practice, Monash Medical Faculty, Melbourne), Naiyana Tikky Wattanapenpaiboon (Monash Asia Initiative), Ashok Vivek Patel (International Association of Rural Health and Medicine, Pune, Maharashtra) and the three reviewers for their invaluable comments on an earlier draft of this article. We also thank Vivien Seyler (*South Asia: J. South Asian Stud.*) for her editorial support and Anusha Kesarkar-Gavankar (IITB-Monash Research Academy, Mumbai, India) for her work as our research assistant. Our special appreciation also goes to Ashok Gulati (Agriculture, ICRIER, New Delhi, India) for his advice on some data issues.

**Conflicts of Interest:** Neither author has any connections with the Indian government, Covi-Self or any other stakeholder discussed in this paper. The paper did not involve any human or animal experiments.

## **References**

## *Article* **Significance of Lung Ultrasound in Patients with Suspected COVID-19 Infection at Hospital Admission**

**Holger Gutsche 1, Thomas G. Lesser 1, Frank Wolfram 1,\* and Torsten Doenst <sup>2</sup>**

**Abstract:** With a lung ultrasound (LUS) the typical findings are interstitial pneumonia. COVID-19 pneumonia is often manifested in sub-pleural areas, which is preferably detected by sonography. An RT-PCR test cannot always ensure a safe differentiation of COVID-19- and non-diseased cases. Clinically challenging is that a reliable and time efficient decision regarding COVID-19 suspects requiring isolation. Therefore, this study was aimed at evaluating the significance of LUS in symptomatic patients with COVID-19 suspicion at hospital admission. A total of 101 patients admitted to a suspect ward with COVID-19-typical symptoms were assessed. All patients received prospectively a standardized LUS at admission. Patients were classified as LUS-positive and -negative cases based on a specific LUS score. The RT-PCR test in combination with the clinical findings served as a reference. Correctly classified were 14/15 COVID-19 diseased suspects as LUS-positive (sensitivity: 93.3%). Twenty-seven out of 61 non-positive cases were classified as false positive with LUS (specificity: 55.7%). In 34/35 patients who were assessed as LUS negative, no COVID-19 disease was detected during the hospitalization. The PPV and NPV of the LUS were 34.1% and 97.1%. LUS is a valuable tool in symptomatic patients for the assessment of COVID-19-disease. The high negative predictive value of LUS is helpful to rule out the disease.

**Keywords:** lung ultrasound; COVID-19; POCUS

## **1. Introduction**

The COVID-19 pandemic poses a major challenge to the health care system. Among other things, the timely and safe differentiation between symptomatic patients with COVID-19 disease and those not infected by Sars-COV-2, is a challenge particularly for hospitalized patients. Due to the very non-specific symptoms such as fever, cough, cold, or unclear infection parameters, a high number of patients without COVID-19 disease are admitted to isolated COVID-wards. Especially elderly, often multi-morbid, patients are thereby exposed to an additional risk of infection, and treatment of their underlying disease and comorbidities might be delayed. To this end, medical resources are tied up in such cases, and this increases the strain upon the health care system in a pandemic situation.

With currently available testing methods, notably the RT-PCR test, the absence of Sars-Cov2 infection can only identified correctly and expeditiously to a limited extent. The false negative rate of the RT-PCR test is being reported up to 38% [1], which can arise as a result of poor sampling quality, improper collection, or unfavorable transport conditions [2]. Furthermore, due to limited test capacities, results can be delayed for several days, or positive test results might be present only after the second or third sample [3]. Therefore, a single negative RT-PCR test result on respiratory specimens is insufficient to rule out COVID-19 disease [4–6].

Symptomatic COVID-19 disease is dominantly associated with pneumonia. For imaging and monitoring of those, chest X-Ray (CXR) is of minor importance compared

**Citation:** Gutsche, H.; Lesser, T.G.; Wolfram, F.; Doenst, T. Significance of Lung Ultrasound in Patients with Suspected COVID-19 Infection at Hospital Admission. *Diagnostics* **2021**, *11*, 921. https://doi.org/10.3390/ diagnostics11060921

Academic Editor: Tivani P. Mashamba-Thompson

Received: 29 March 2021 Accepted: 17 May 2021 Published: 21 May 2021

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with computed tomography (CT) because of missing specificity [7]. Lung CT of COVID-19 pneumonia shows typical findings of interstitial pneumonia with a sensitivity of 94% [8]. It is recommended in case of urgent COVID-19 suspicion and a negative RT-PCR. However, the examination is not practicable as an incoming screening examination due to high patient volume in a pandemic situation, transport problems, hygiene guidelines, and associated radiation exposure.

The lung ultrasound is becoming a more and more important point-of-care imaging modality in the differential diagnostics of pulmonary and pleural diseases [9]. In the case of interstitial pneumonia, typical ultrasound findings such as an irregular or fragmented pleural line, B-lines, white lung syndrome (WLS), or consolidations can be collected [10]. Since COVID-19 pneumonia manifests mainly peripheral/subpleural, an ideal prerequisite for the use of LUS is available [11]. Early stages of pneumonia can be detected with ultrasound, and the findings correlate well with those by computed tomography [12–14]. Due to its portability, LUS allows bedside imaging in absence of radiation and could be a valuable diagnostic screening tool for the early detection or exclusion of patients with Sars-COV-2 infections.

Therefore, this study is aimed to investigate the significance of LUS in symptomatic patients with COVID-19 suspicion at the time of hospitalization. For this, the diagnostic value of a lung ultrasound with reference to the RT-PCR test with the clinical findings was determined.

## **2. Materials and Methods**

## *2.1. Patients*

In the period from April to June 2020, patients with clinical suspicion of COVID-19 disease underwent prospective and standardized LUS after admission to the COVID-19 suspect ward of the SRH Wald-Klinikum Gera. The symptoms for inclusion were cough, dyspnea, unclear fever (>38 ◦C), as well as an increase in laboratory infection parameters (CrP > 10 mg/L; PCT > 0.5 mg/L). Patient characteristics were separately documented from the digitally stored patient data after anonymization. These included ages, gender, symptoms, computed tomography of the chest (if done), and laboratory findings, as well as secondary diseases.

Inclusion criteria were an age over 18 years, no pregnancy, and LUS examination within 48 h after hospital admission. Exclusion was performed in need of immediate ICU transfer and an uncooperative condition.

## *2.2. Lung Ultrasound Examination*

All examinations were carried out by one experienced LUS investigator with a background of six years in thoracic and vascular surgery with daily practice of general and lung sonography. The examiner received prior to the study a general LUS training according to EFSUMB and AIUM recommendations and COVID-19 specific online training.

The systematic LUS examination was performed bedside on six thoracic areas of the right (R) and left (L) hemi-thorax in an insulating room at the COVID-suspect ward.

The investigation was conducted on the basis of international recommendations for COVID-19 lung ultrasound examination [15,16]. Herein the probe was placed onto the intercostal place to ensure imaging without rib shadowing. The investigator scanned for suspicious or pathological LUS feature within each areal.

The following chest areas were investigated:

The examinations were performed with a clinical sonography scanner (LOGIQ/VENUE, GE HealthCare, Solingen, Germany) using a linear probe (9L) or in obese patients with a convex probe (C1-5). The scanner specific lung ultrasound pre-set was used with a median MI of 1.1 (0.8–1.2) and TIS of 0.1 (0.0–0.2). The operator adjusted the placement of the single focus onto the pleural line. LUS frames of five seconds were initially recorded from each areal to ensure the monitoring of at least one breath cycle. The video sequences were evaluated blindly by two LUS experienced investigators (FW, HG) independent of each other, and without knowledge of patients' condition including the RT-PCR status. In the case of different assessments, a third clinician (TL) was consulted for consensus formation. The definitive classification of all 12 lung areas was carried out by means of a lung score of 0–3 based on the international COVID-19 LUS scoring system according to Soldati et al. [16]:

Patients were classified as "LUS-COVID-positive" if a score of 2 or 3 was present in one area out of 12. A classification as "LUS-COVID-negative" was made when a score of 0 or 1 was found throughout all areas. The patient was excluded from the study if more than one area was missing or could not be assessed.

## *2.3. RT-PCR Reference*

The patients suspected for COVID-19 disease already received a RT-PCR test in the emergency department (ED) at admission. In the case of negative test results and continued symptomatic course, a repeated RT-PCR test was carried out by sampling an irritant sputum or throat rinse water. As soon as a positive single test was present during the course of the disease, the patient was classified as "COVID-19-diseased". All RT-PCR tests were carried out by SYNLAB Medical Care Center Gera (SYNLAB MVZ, Weiden, Germany). A "first RT-PCR" test was defined if taken and successfully analyzed at the ED in the moment of admission. Out of 76 included cases a first RT-PCR test was successfully performed on 68 cases. Four RT-PCR samples could not be evaluated due to material defects and four patients were admitted to the COVID-19 ward due to a positive ambulatory COVID-19 test without detailed documentation.

## *2.4. Statistical Analysis*

The statistical analysis was performed using MedCalc (Vers. 19.1.7 Medcalc LTD, Ostend, Belgium). Quantitative variables are presented as median and Inter Quartile Range (25th percentile, 75th percentile), and categorical as counts and proportion. Statistical evaluation was performed on the categorical variables with the Fisher's Exact–Boschloo test, and on continuous data with Mann–Whitney U tests. The tests' power was estimated based on the resulting counts [17]. In addition, sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and diagnostic accuracy were evaluated for the LUS score, first RT-PCR Test, and in a combination of both. The positive count for combination of LUS and RT-PCR test was made when an LUS-COVID-positive or positive first RT-PCR test result was present. The estimates are reported with 95% confidence interval (CI). A two-sided *p*-value < 0.05 was defined as significant.

## **3. Results**

## *3.1. Patient Characteristics*

A total of 101 patients with clinical suspicion of COVID-19 disease were examined using LUS. After excluding 25 cases, 76 patients were included for the study. A summary of studies patient selection is shown in Figure 1.

**Figure 1.** Flow chart of the patient selection and classification according to the lung ultrasound and reference standard.

Baseline characteristics, secondary diseases and clinical symptoms of the study patients are presented in Table 1. The median age was 75.5 years. Both genders were almost equally distributed (male 48.7% and female 51.3%). The median length of stay on the COVID-19 suspect ward was 4.7 days for all patients, 2.4 days for non-COVID-19-diseased and 13.9 days for COVID-19-diseased patients. In 15 (19.7%) of the patients, COVID-19 disease was finally diagnosed, in 61 patients (80.3%) COVID-19 disease had been excluded after the end of the diagnostics. With regard to the incidence of fever, cough, or dyspnea as well as blood gases pO2 and pCO2, no differences were found between COVID-19 and non-COVID-19-diseased patients (*p* > 0.05). The loss of taste and/or sense of smell were found significantly more commonly found among patients with confirmed COVID-19 disease (*p* = 0.047). No significant difference with regard to the secondary diseases was present between non- and COVID-19-diseased cases.

**Table 1.** Baseline characteristics of the study patients with secondary diseases and clinical symptoms.

A *p*-value < 0.05 was defined as significant.

## *3.2. Diagnostic Value of Lung Ultrasound and in Combination with RT-PCR Test*

The LUS examination was performed at the COVID-suspect ward within a median time delay of 18 h 5 min (9 h 48 min–23 h 30 min) after admission to the hospital's ED. Fourteen out of the fifteen COVID-19-diseased patients were classified LUS-COVIDpositive (sensitivity: 93.3%; CI: 68.1% to 99.8%). Of 61 non-COVID-19-diseased patients, 27 LUS-COVID-positive cases were false-positive classified with LUS (specificity: 55.7%; CI: 42.5% to 68.5%). In 34 patients who were assessed as negative by LUS (LUS-COVIDnegative), no COVID-19 disease was detected in the further course of the hospitalization. One patient was classified LUS-COVID-negative in the presence of COVID-19 disease and therefore judged to be false-negative. This patient was admitted with symptomatic cough and an increased infection parameter (CrP = 47 mg/L) due to a fracture from an external, not COVID-19 treating, hospital. No fever or dyspnea and lung infiltrations on CXR were present.

The positive and negative predictive value (PPV, NPV) of the LUS was 34.2% (CI: 27.5% to 41.5%) and 97.1% (CI: 83.5% to 99.6%), respectively. The first RT-PCR test, carried out at hospital admission such as in the ED (68 of 72 RT-PCR tests), showed a sensitivity of 90.9% (CI: 58.7% to 99.8%), a specificity of 100% (CI: 93.7% to 100%), resulting in a PPV of 100% and an NPV of 98.3% (CI: 89.8% to 99.7%). Until a reliable result was available, approximately 2.1 (CI: 1.9 to 2.3) RT-PCR tests were performed, therefore non-COVID-19 diseased patients were discharged from the COVID-19 ward after 2.4 days (CI: 2.0 to 2.9). The result of the first RT-PCR test combined with the LUS-COVID classification, revealed a sensitivity of 100% (CI: 71.5% to 100%) and thus a negative predictive value of 100% (Table 2).

**Table 2.** Diagnostic significance of LUS score classification, first RT-PCR test, and in combination of both in COVID-19 suspected patients.

LUS, lung ultrasound; PPV, positive predictive value; NPV, negative predictive value.

## *3.3. Pathological Lung Ultrasound Findings*

More than two B-lines, white lung syndrome, pleural fragmentation, and consolidations were found significantly more frequently in COVID-19-diseased than in non-COVID-19-diseased patients. When COVID-19 was present, more than two B-lines appear in all patients, WLS in 66.7%, subpleural consolidation in 53.3%, and pleural fragmentation in 93.3%. COVID-19 patients were significantly more often classified as LUS-COVID-positive than non-COVID-19-diseased cases (93.3% vs. 44.3%, *p* < 0.01). For COVID-19-diseased patients an LUS score of 3 was found in 53.3%, a score of 2 in 40%, whereas only one patient (1.6%) showed an LUS score 1, characterized by a thickened pleural line in three areas and increased B-lines in two areas. Pleural effusion was found more frequently in non-COVID-19-diseased (23%) than in COVID-19-diseased (6.7%) patients which revealed no statistical significance. A summary of LUS score and pathological findings are presented in Table 3.

**Table 3.** Classification results and lung ultrasound characteristics of study patients.

A *p*-value < 0.05 was defined as significant, \* indicate Fischer's exact test with zero counts, see limitation.

The most common LUS findings in non-COVID-19-diseased patients were thickened pleura (98.4%), increased B-lines (75.4%), and a pleural fragmentation (42.6%). Subpleural consolidation as a typical finding in interstitial pneumonia was shown in 26.2%. LUS scores of 3 and 2 were present in 26.2% and 18.0%, respectively. An LUS score 1 was significantly more common in non-COVID-19-diseased than in COVID-19-diseased patients (54.1% vs. 6.7%).

Non-pathological, aerated lung in all areas (score 0) was not present in all COVID-19 diseased, and in only one non-COVID-19-diseased (1.6%) patient, which was not significant.

## **4. Discussion**

We were able to show that the early lung ultrasound examination of inpatient admission provides a diagnostic gain and is valuable in the clarification of Sars-COV-2 suspected patients at hospital admission. With LUS, 93.3% of the COVID-19-positive cases could be classified correctly. In 97.1% of the LUS-negative cases, no COVID-19 disease was present. The NPV increases to 100% when the LUS findings are combined with the first RT-PCR test, carried out at hospital admission.

In contrast, symptomatic screening (fever, cough, dyspnea, blood gas) is insufficient, as our baseline characteristic shows. COVID-19 specific symptoms are slightly more frequently found on diseased patients, which was not significant. Although the loss of taste and smell is a significant symptom it is not applicable for screening due to the low incidence (6.8%).

The duration on a COVID suspect ward (2.4 days) for non-COVID-19-diseased cases is signally inappropriate, causing a delay of therapy, risk of infection, and an unnecessary consumption of resources. The RT-PCR test shows a valuable diagnostic accuracy for the study specific cohort of symptomatic suspects at hospital admission. However, the LUS provides a higher sensitivity than the first RT-PCR test, but with a much lower specificity. Using LUS in combination with an RT-PCR test could help reducing the isolation time on a suspect ward, and therefore reduce the strain on the health care system in a pandemic situation.

The specificity of the LUS examination is low (55.7%), as the sonographic findings are not only COVID-19 specific. Such pathological LUS signs can occur in other diseasecausing interstitial pneumonia, or are present due to underlying chronical lung disease, as frequently found in patients requiring hospitalization. Therefore, LUS cannot provide an etiological diagnosis. Comparable results were found in a recent study by Sorlini et al. [18], reporting a sensitivity and specificity of 92.0% and 64.9%, respectively. The higher specificity can be explained by the higher prevalence of the COVID-19 disease (74.7%) in the cohort studied.

In addition, Volpicelli et al. [19] reported an LUS sensitivity of 90.2% and specificity of 50.5% investigating early LUS under various inclusion criteria. Narinx et al. [20] found an LUS sensitivity of 93.3% and specificity of 21.3% for patients at ED admission. Highly interesting, is the work of Pivetta et al. [21], with a high LUS sensitivity (94.4%) and specificity (95%). In contrast to our work, all the above studies classified the appearance of increased B Lines and WLS as a COVID-19 related LUS sign, which would represent a score of 1 in our study. This choice, different inclusion criteria, and the clinical situation of the LUS examiner being aware of the patients' condition (not blinded), might explain the high specificity found by Pivetta et al. However, these studies in addition to our results, demonstrate the potential of LUS, but also the need for a deeper definition of COVID-19 specific LUS features and for an appropriate definition of inclusion criteria.

The LUS score classification has a decisive influence on the test results of the LUS. A lower cut off, that includes score 1 to be LUS-COVID-positive, leads to a high sensitivity and high false positive rate. LUS as an early diagnostic method of non-COVID-19 diseased cases should be focused on the exclusion of the disease to avoid hospital admission. Our chosen cut off provides a justifiable NPV, which is to our knowledge the highest one reported so far. Therefore, we recommend expressing suspicion of COVID-19 disease only with an LUS score of ≥2 (Figure 2) for patients in early hospital admission. In our study, a high prevalence of cardiac disease (82%) and hypertension (80%) were present in the non-COVID-19-disease group, which are often associated with increased lung water. These underlying pathologies frequently manifest pathological LUS artefacts such as B-lines and WLS (score 1) [9].