**7. Prolotherapy**

Prolotherapy derives its name from "proliferation therapy," or "proliferative injection ther‐ apy". It has also been called "regenerative injection therapy" ("RIT"), and some contem‐ porary authors name the therapy according to the injected solution. The precise mechanism of action is not known. It involves injecting a non-pharmacological and nonactive irritant solution into the body. This is hypothesized to reinitiate the inflammatory process which deposits additional new fibers, thereby repairing lax tendons or ligaments and also possibly promote the release of local growth factors. However, the precise mech‐ anism of action still remains unknown. It has been used for approximately 100 years, however, its modern application can be traced to the 1950s.This was when the prolothera‐ py injection protocols were formalized by George Hackett, a general surgeon in the U.S.,and were based on his clinical experience of over 30 years. The concept of creating irritation or injury to stimulate healing has been recorded as early as Roman times when hot needles were poked into the shoulders of injured gladiators [28].

There are no trials on the use of prolotherapy in a population with acute muscle tears. There is some evidence for its use in chronic pathologies such as LBP [29], groin pain [30], and knee OA [31]. However, existing evidence is inconclusive. Its actual use is based on "expert opinion" and some practitioners advocate a positive impact of the use of prolotherapy in athletes in term of return to play timeframes.


**6. Conclusions**

In this study, 50 Late Persian period and Macedonian period silver coins were studied by NDT metallurgical analysis. The results obtained from the SEM-EDS surface analysis of well-preserved areas sufficiently represent the concentrations of the coins' ground bulk. Correspondence was received between the SEM-EDS analysis results and the XRF analysis results. The results show that the coins are made of high purity silver with only small percentages of copper. All the coins were produced by similar techniques of casting flans and striking them by plastic deformation. Based on the average alloy composition values and SD, as well as the copper concentration distribution of the examined series, the five die combination issues of the Yehud series Type 5 O1/R1–O1/R5, Type 16 O2/R2, Type 24 O1/R2, and Type 31 O1/R1 groups, and the late addition Yehud coin type, each series was most likely produced by a different and controlled specific composition of silver–copper alloy. However, four Type 5 exceptional coins (IAA 153976 O1/R1, IAA 154383 O1/R2, IMJ 27387 O1/R2, and IAA 153981 O1/R4) revealed a different alloy composition with a much higher amount of copper and heterogeneity. This implies that at a certain stage of the minting process, a different batch of possibly recycled alloy was used instead of the standardized alloy that was recorded for all other coins of the same die connection. The current four-step methodology revealed novel information concerning the material culture of the southern Levant during the Late Persian period associated with minting production of silver coins. This four-step methodology can be used with additional bulk NDT methods by other researchers to study various ancient silver objects.

**Supplementary Materials:** The following supporting information can be downloaded at: https: //www.mdpi.com/article/10.3390/ma16062200/s1, Figure S1. SEM-EDS analysis showing the spectra of a typical Yehud *gerah* silver coin (Type 5 O1/R3, IAA 153978, reverse); Figure S2. Images of the Yehud *gerah* Type 5 O1/R1 coins: (a) IAA 138139 reverse depicting an owl (SEM, BSE mode); (b) IAA 138139, reverse (multi-focal LM) showing the Paleo-Hebrew inscription YHD (Yeh[u]d); (c) IMJ 27398 obverse depicting a helmeted Athena (SEM, SE mode); (d) IMJ 34543 reverse depicting an owl (SEM, BSE mode); and (e) IMJ 34543 obverse depicting a helmeted Athena (SEM, BSE mode). The areas inside the squares were examined by EDS analysis; Table S1. SEM-EDS analysis results of the Yehud *gerah* Type 5 O1/R1 coins, where SA represents the scanned area. Only bright areas according to BSE mode of shiny metal were examined by EDS analysis; Figure S3. Images of Yehud *gerah* Type 5 O1/R2: (a) coin IAA 154383, reverse (multi-focal LM), showing the face of an owl; and (b) IAA 154383, reverse (SEM, BSE mode), where the bright areas represent shiny silver metal and the dark areas are covered with oxide and corrosion products. The areas inside the squares were examined by EDS analysis; Table S2. SEM-EDS analysis results of the Yehud *gerah* Type 5 O1/R2 coins, where SA represents the scanned area. Only bright areas according to BSE mode of shiny metal were examined by EDS analysis; Figure S4. Yehud *gerah* Type 5 O1/R3, IAA 153978 (obverse), SEM–EDS elemental mapping: (a) general view of the examined area; (b) the detected elements (Ag, Ca, O, Cl), where the green areas are rich in silver and red areas are rich in chlorine; (c) presence of Ag; (d) presence of Ca; (e) presence of O; and (f) presence of Cl; Table S3. SEM-EDS analysis results of the Yehud *gerah* Type 5 O1/R3 coins, where SA represents the scanned area. Only bright areas according to BSE mode of shiny metal were examined by EDS analysis; Table S4. SEM-EDS analysis results of the Yehud *gerah* Type 5 O1/R4 coins (each scanned area was 300 μm × 300 μm). Only bright areas according to BSE mode of shiny metal were examined by EDS analysis. RH represents Ramallah Hoard (nos. 2–6); Table S5. SEM-EDS analysis results of the Yehud *gerah* Type 5 O1/R5 coins. Only bright areas according to BSE mode of shiny metal were examined by EDS analysis; Figure S5. Yehud half *gerah* Type 16, O2/R2 coin, reverse depicting a falcon in flight: (a) Edom hoard no. 4 (SE mode); (b) Edom hoard no. 5 (SE mode); and (c) Edom hoard no. 6 (BSE mode), where the brighter areas according to the BSE mode are better preserved than the darker areas. The areas inside the squares were examined by EDS analysis; Table S6. SEM-EDS analysis results of the Yehud half *gerah* Type 16 O2/R2 coins. Only bright areas according to BSE mode of shiny metal were examined by EDS analysis; Figure S6. SEM images of the Trans-Jordan hoard Type 24 O1/R2, obverse depicting a facing head: (a) coin Trans-Jordan hoard no. 11 (SE mode); (b) coin Trans-Jordan hoard no. 13 (SE mode); (c,d) coin Trans-Jordan hoard no. 12 (SE mode and BSE mode, respectively). The areas inside the squares were examined by EDS analysis; Table S7. SEM-EDS analysis results of the Yehud quarter *obol* Type 24 O1/R2 coins, where SA represents the scanned area. Only bright areas according to BSE mode of shiny metal were examined by EDS analysis; Figure S7. SEM images of the Yehud *hemiobol* Type 31, O1/R1, reverse depicting a bird standing right, head reverted: (a) IMJ 34593 (BSE mode); and (b) IMJ 34591 (SE mode). The areas inside the squares were examined by EDS analysis; Table S8. SEM-EDS analysis results of the Yehud *hemiobol* Type 31 O1/R1 coins, where SA represents the scanned area. Only bright areas according to BSE mode of shiny metal were examined by EDS analysis; Table S9. SEM-EDS analysis results of the Yehud IAA 153976, IMJ 27424, IAA 101006, IAA 154383, IMJ 27383, Trans-Jordan hoard no. 11, and IMJ 34591 coins after roughly grinding the surface; Figure S8. The copper distribution of each silver coin group based on SEM-EDS chemical analysis results, where the Cu wt% concentration is presented vs. the relative no. of measurements (%): (a) Yehud *gerah* Type 5 O1/R1; (b) Yehud *gerah* Type 5 O1/R2; (c) Yehud *gerah* Type 5 O1/R3; (d) Yehud *gerah* Type 5 O1/R4; (e) Yehud *gerah* Type 5 O1/R5; (f) Yehud half *gerah* Type 16 O2/R2; (g) Yehud quarter *obol* Type 24 O1/R2; and (h) Yehud hemoibol Type 31 O1/R1; Figure S9. Yehud *gerah* coins Type 5 O1/R1 (ca. 350–333 BCE) (Athena/Owl): IAA 138139, IAA 153975, IMJ 27424, IMJ 27398, IMJ 34542, IMJ 34539, IMJ 34543, and IAA 153976; Figure S10. Yehud *gerah* coins Type 5 O1/R2 (ca. 350–333 BCE) (Athena/Owl): IAA 101006, IAA 177246, IAA 153977, IMJ 34538, IMJ 34553, IMJ 34537, IMJ 34554, IMJ 34620, IAA 154383, and IMJ 27387; Figure S11. Yehud *gerah* coins Type 5 O1/R3 (ca. 350–333 BCE) of the Yehud series (Athena/Owl): IAA 153979, IAA 153978, IMJ 34555, and IMJ 27425; Figure S12. Yehud *gerah* coins Type 5 O1/R4 of the Yehud series (Athena/Owl): RH2-RH6, IAA 153980, IMJ 34556, and IAA 153981; Figure S13. Yehud *gerah* coins Type 5 O1/R5 (ca. 350–333 BCE) of the Yehud series (Athena/Owl): IMJ 34558, and IMJ 27388; Figure S14. Yehud coins Type 16 O2/R2 (Persian king wearing a jagged crown/Falcon in flight) (ca. 350–333): Edom hoard nos. 1–6, IMJ 27383, IMJ 27414, and IMJ 34566; Figure S15. Yehud Attic standard quarter *obol*, Type 24 O1/R2 (Facing head/Owl) of the Macedonian period (ca. 320(?)–312 BCE): Trans-Jordan hoard nos. 11, 12, 13; Figure S16. Yehud coins Type 31 O1/R1 (Head of roaring lion/bird standing right, head reverted) (ca. 306–302/1 BCE): IMJ 34631, IMJ 34593, IMJ 34591, IMJ 34709, IMJ 34594, and IMJ 34715.

**Author Contributions:** Conceptualization, M.C., D.A., H.G. and O.T.; study objectives, M.C., D.A., H.G. and O.T.; research methodology, M.C., D.A., H.G. and O.T.; formal analysis, M.C. and D.A.; discussion of results, M.C., D.A., H.G. and O.T.; conclusions, M.C., D.A., H.G. and O.T.; writing introduction, H.G. and O.T.; writing—technological background to research, M.C. and D.A.; writing original draft preparation, D.A.; project administration, H.G. and O.T.; funding acquisition, H.G. and O.T.; supervision, O.T. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research forms part of an Israel Science Foundation (ISF) project (No. 2883/20); Corpus of Samarian Coinage (ca. Fourth Century BCE); PI: Oren Tal.

**Institutional Review Board Statement:** Not applicable.

**Informed Consent Statement:** Not applicable.

**Data Availability Statement:** The data supporting the reported results can be found in the Supplementary Materials file.

**Acknowledgments:** The authors are indebted to the ISF and would also like to thank Tomer Reuveni from the Wolfson Applied Materials Research Center at the Tel Aviv University for his SEM technical assistance. We are also grateful to Robert Kool, head of the coin department at the IAA, for providing the controlled archaeological excavation coins for our project.

**Conflicts of Interest:** The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.


**5. Conclusions**

It is often di fficult to guarantee the absence of *Legionella* from water distribution systems, even if a disinfection system is in place. Our data revealed that di fferences in three buildings belonging to the same structure were linked to building size, water consumption rates, the number of outlets, and their intended use. WTP 828 performed well in terms of reducing *Legionella* contamination, but only a change in the study approach (adequate risk assessment plan, increase in monitoring samples sites, and alteration of the WTP 828 dosage in relation to the *Legionella* levels) facilitated the discovery of di fferences in *Legionella* colonization and an understanding of disinfectant activity dynamics.

Further investigations are needed to elucidate how the dose of disinfectant a ffects the presence of specific strains in each building and to generate a risk map highlighting the phylogenetic correlations between strains. The assessment of changes in colonization dynamics will be useful for controlling the concentration and type of disinfectant that can be used in a water system (i.e., shock or continuous treatment, bacterial resistance development) in relation to accommodation works and technical operations in the water network that could support *Legionella* proliferation.

The low cost of WTP 828, the dosage by a pump, the easy maintenance procedures and simple and safe check of disinfectant residue at distal outlets, suggest that the approach used in this study could be a valid alternative to traditional disinfection methods.

**Supplementary Materials:** The following are available online at http://www.mdpi.com/2076-0817/8/4/209/s1. Figure S1. Site map of MCH, Cotignola (RA), Italy. A representative view of MCH and a map of its three buildings: MCH picture (A) and MCH planimetry (B); Table S1. MCH structure and water outlet characteristics.

**Author Contributions:** S.C. and L.G. conceived, designed the experiments, wrote the paper, and performed the experiments; A.G. and G.F.S. performed the sequence analysis; A.D. performed the data analysis; D.M., A.C., F.S., and A.M. provided disinfectants and performed the water sample collection; T.P. and P.S. provided technical support to study the MCH hot water network.

**Funding:** This work was supported by a research grant (RFO 2017) from University of Bologna, Italy.

**Acknowledgments:** The authors would like to thank the MCH sta ff for helping with the study, the technical sta ff at Water Team S.r.l. for assistance and provision of the WTP 828 disinfectant, Eta Beta S.r.l. for technical assistance, and Graziella Ciani for help with the microbiological analyses.

**Conflicts of Interest:** S.C., L.G., A.G., G.F.S., and A.D., declare no competing interest; T.P. and P.S. are employed by GVM Care & Research; D.M. and A.C. by Eta-Beta S.r.l. and F.S.; and A.M. by Water Team S.r.l. Water Team S.r.l. provided assistance and provision of the WTP 828 disinfectant and Eta Beta S.r.l. provided technical assistance for disinfection treatment.

**Ethics Approval and Consent to Participate:** Not applicable.

#### **Consent for Publication:** Not applicable.

**Availability of Data Materials:** The datasets generated and analyzed during the study are not publicly available due to ongoing further analysis on the data and protected by hospital privacy policies. The corresponding authors can response to any further questions about them.


**1. Introduction**

The livelihood and economies of most Sub Saharan African (SSA) countries are driven by rainfed agriculture [1–3]. About 96% of agricultural lands in SSA are rainfed [1,4]. Agriculture employs over 65% of the active labour force of the region, the majority of whom are practicing subsistence rainfed farming [5]. The agricultural sector is also responsible for 75% of SSA domestic trade [6,7]. It adds significantly to the economy of the region by contributing up to 15–20% to the Gross Domestic Product (GDP) [1,4,8,9]. In 2000, about 80% of the cereals consumed in SSA were domestically produced locally [4]. However, West Africa has been identified as one of the most vulnerable regions of the world owing to its low adaptive capacity and a fast-growing population, with many citizens whom are faced with malnourishment [1,10–12]. An adverse change in the climate over West Africa, both spatially and temporally, coupled with inadequate institutional and economic capacity to cope or adapt to its impact could become a determinant threat to agricultural production. Thus, food security and socio-economic activities across the region may be a ffected [13–17].

Climate strongly a ffects rainfed agriculture with direct consequences on food security [18–20]. This has resulted in di fferent studies focusing on the response of crops and agriculture to the impact of increased greenhouse gas emissions across di fferent regions of SSA owing to malnutrition and the need to improve food security [1,11,20–22]. Extreme changes in climate are projected to increase [23], translating into increased occurrence of both droughts and floods, which already account for 70% of economic losses through soil erosion and drought in West Africa [24,25]. The fifth Intergovernmental Panel on Climate Change (IPCC) assessment reported a projected warming across the di fferent seasons over SSA to be larger than the global annual mean temperature increase [23]. The projected warming (1.5–4 ◦C by 2100) is likely to a ffect the agricultural sector by a reduction of up to 50% in crop yield and 90% in revenue across the region by the end of the century [7,23]. However, this may be further aggravated in regions like West Africa where the climate is warming faster and may lead to a radical departure from the regions' historical variability [26].

The definition of departure varies across disciplines. Broadly, a departure refers to a deviation or variation from a norm, standard rule, or behaviour. It can also mean starting out on a new course of action. In climate science, climate departure can be defined as a shift in the climate pattern of a region outside the range of historical variability and may be described in terms of mean local temperature exceeding historical highs [27–29]. Mora et al. [29] described climate departure as the year in which the average temperature of the coldest year after 2005 was warmer than the historic hottest year at a given location. Here we have defined climate departure as a deviation from the historical mean and/or variance of the local climate of an area or region induced by global warming [30].

The projected global warming level and the timing over the continent may intensify the impact of climate change on crop suitability. Severe temperature fluctuations and other extreme weather conditions such as droughts and floods may also threaten crop suitability thresholds. They vary spatially, resulting in potential yield declines where crop growth conditions are currently suitable and possible yield increases in other areas [31,32]. Challinor et al. [33], for instance, projected a future decline in crop yield of up to 5% for every degree of warming above the historical level in Africa.

Given the current state of climate departure research and the direct impact of climate on crop production systems (particularly rainfed), we are interested in the climate change induced crop realizations when climate departs from historical variability, which we term crop–climate departure. This study explores and proposes the information value of a comprehensive definition of crop–climate departure as "a departure from historical crop suitability threshold, whether in terms of variability, mean or both, due to warming of the climate over a location both in space and time resulting from climate change whether of radical climatic nature or not". This is in the context of recent climate historical variability and future climate projections using three West African weather stations, within three Food and Agriculture Organizations (FAO) Agro-Ecological Zones (AEZs). Mora et al. [29] suggests that West Africa will experience a climate departure with a mean temperature about two decades (2029) earlier than the global mean temperature (2047). Thus, we use the region as our proof of concept and to examine any likely large-scale crop suitability consequent changes the region may already be experiencing. Section 2 describes the data and methods used. Results from the study are outlined in Section 3. The discussion of the results and concluding remarks and recommendations for future are in Sections 4 and 5, respectively.

## **2. Data and Methods**