A 27-year-old male sought the dermatology care service for a skin lesion located on the right mandible, characterized by increased volume, erythema, and fistulous tracts. The dermatosis comprised an area of 3x4 cm and exuded a slightly seropurulent secretion (Figure 1A). He claimed it had been evolving for more than five months and mentioned having undergone previous treatment with different topical and oral antibiotics, without improvement. He claimed to be a permanent resident in Mexico City (19 25'10"N 99 08'44"O), but he had made frequent business trips to Veracruz state (19 26'05"N 96 22'59"O) in the last year for periods of one to two weeks. The suspected clinical diagnoses were actinomycosis, cutaneous tuberculosis, or sporotrichosis. Thus, a skin biopsy, histopathological analysis, and culture from a tissue sample were performed. Also, a blood count and blood chemistry were requested.
Regarding histopathology, the hematoxylin and eosin stain exhibited an epidermal hyperplasia and ulceration. Tissue slices (Figure 1B) exhibited an inflammatory infiltrate composed of histiocytes, plasma cells, lymphocytes, and polymorphonuclear located in the dermis and subcutaneous tissue (Figure 1C). There was also vascular damage. No fungal structure or mycobacteria was observed in special stains such as Gram, Ziehl-Neelsen, PAS [periodic acid-Schiff stain], and Giemsa. No fungal growth was observed in the culture media (Sabouraud Dextrose Agar with Cycloheximide and Chloramphenicol [Mycobiotic Agar], potato dextrose agar, and brain heart infusion). Surprisingly, parasites (amastigotes and trypomastigotes), as well as intracellular bacteria, were presented in the tissue samples (Figures 1D, 1E, and 1F). Furthermore, the bacterial culture analyses showed evidence for the growth of two different isolates. The former corresponded to a Gram-negative bacillus, Ziehl-Neelsen negative, and catalase-negative, identified by PCR and sequencing as Bacillus velenzensis (Figure 1G), and the second was identified as Corynebacterium sp. The antibiogram reported a Bacillus sp., sensitive to cephalothin, ciprofloxacin, fosfomycin, gentamicin, tetracycline and oxycline, resistant to nitrofurantoin and trimethoprim/sulfamethoxazole, and intermedium resistant to clindamycin, erythromycin, and penicillin. Moreover, culture media for fungi were evaluated weekly and did not present any growth during five weeks. Consequently, serological diagnoses were requested to confirm the diagnosis of Chagas disease or Leishmania sp. The initial tests, executed in 3 independent laboratories (blood bank laboratory [chemiluminescence architect Chagas kit (Abbott)], a research laboratory specialized in parasitic infections in our institution [ELISA home with the Ninoa strain as antigen]  , as well as an external private laboratory [chemiluminescence technique]), were reported as negative.
PCR analysis was done to identify Leishmania sp. and specific Leishmania species with DNA isolated from the tissue biopsies and blood; these tests showed negative results. However, a PCR established with the DNA satellite showed the presence of Trypanosoma cruzi DNA  . The sample sequence showed an identity of 99% with the T. cruzi ClBE07 clone (GenBank No AY520036). Furthermore, Bayesian phylogenetic analysis (Figure 2) supported the grouping of the sequence obtained inside T. cruzi DTUs and showed high posterior probability values (1.00) for DTU II. On the other hand, PCR-sequencing analysis to identify bacterial 16S revealed the presence of Bacillus amyloliquefaciens, also known as velezensis, in the tissue samples and culture (GenBank ascension No ON737906).
Laboratory tests were requested, including blood count and chemistry, reporting neutrophilia and eosinophilia. Other parameters were within the average reference values. Also, a complete cardiac assessment was performed, including thorax radiography, an electrocardiogram, and an echocardiogram. All tests were reported within the standard values. In addition, two independent laboratories (public and private) repeated ELISA and Western blot during the patient follow-up, and the results were positive four months after the original assessment.
The initial reports established a mixed cutaneous infection; thus, according to the antibiogram report, the patient received standard doses of cefalexin and ciprofloxacin with clinical improvement in three weeks. However, according to PCR and histology results that established Chagas disease (CD), the patient was referred to the Tropical Medicine Center, an authorized center for the diagnosis and management of Chagas disease in Mexico  , because the antiparasitic treatment for CD is not commercially available in Mexico. In addition, the CD is a mandatory report disease, because only special health authorities are able to provide the treatment (Secretaria de Salud). Sadly, current Mexican health care system lineages for Chagas treatment do not allow established treatment if the serology is negative, so for our patient, the treatment was delayed for several months, despite the parasite being visualized in tissue slices and the infection being confirmed with a PCR test  . Thus, the opportunity to treat the patient in the acute phase, when the benefit of treatment is more significant, was lost. Four months after the diagnosis, when the serology reported a positive result, treatment with nifurtimox was started, and doses of 240 mg at 8:00, and 180 mg at 15:00 and 22:00, were administered for 60 days. Afterwards, serology tests performed in August 2021 and January 2022 reported negative results. The last follow-up performed in July 2022 included a PCR and ELISA, which were also negative.