Non-Tuberculous Mycobacterial isolates from Panama: A retrospective 5-year analysis (2017-2021).

BACKGROUND: The genus Mycobacterium includes well-known bacteria such as M. tuberculosis causing tuberculosis and M. leprae causing leprosy. Additionally, various species collectively termed non-tuberculous mycobacteria (NTM) can cause infections in humans and animals, affecting individuals across all age groups and health conditions. However, information on NTM infection prevalence in Panama is limited. METHODS: This study conducted a retrospective analysis of clinical records from 2017 to 2021, specifically focusing on patients with NTM isolates. Data were categorized by variables like sex, age, HIV status, and sample source. RESULTS: Among the 4430 clinical records analyzed, 698 were linked to patients with NTM isolates. Of these patients, 397 were male, and 301 were female. Most female patients with NTM isolates (n = 190) were aged >45 to 85 years, while most male patients (n = 334) fell in the >25 to 75 years age group. A noteworthy proportion of male patients (n = 65) were aged 25-35 years. A significant age difference between male (median [min-max] = 53 years [3-90]) and female (median [61 years [6-94]) patients was observed (p < 0.001). Regarding HIV status, 77 positive individuals were male, and 19 were female (p < 0.001). Most samples (n = 566) were sputum samples, with additional pulmonary-associated samples such as broncho-alveolar lavage, tracheal secretions, and pleural fluid samples. Among extrapulmonary isolates (n = 48), sources included catheter secretions, intracellular fluids, peritoneal fluid, blood cultures, cerebrospinal fluid, bone marrow samples, and capillary transplant lesions. Specifically, the analysis identified the pathogenic microorganisms responsible for mycobacteriosis in Panama during the specific period 2017-2021, as M. fortuitum (34.4%), M. intracellulare (20.06%), and M. abscessus (13.75%), respectively. CONCLUSIONS: This study highlights the growing public health concern of NTM infections in Panama. The research provides valuable insights into the prevalence and distribution of NTM species in the country, offering a foundation for the development and implementation of effective prevention and control strategies for NTM infections in Panama.

Simplified Model to Survey Tuberculosis Transmission in Countries Without Systematic Molecular Epidemiology Programs.

Systematic molecular/genomic epidemiology studies for tuberculosis surveillance cannot be implemented in many countries. We selected Panama as a model for an alternative strategy. Mycobacterial interspersed repetitive unit-variable-number tandem-repeat (MIRU-VNTR) analysis revealed a high proportion (50%) of Mycobacterium tuberculosis isolates included in 6 clusters (A-F) in 2 provinces (Panama and Colon). Cluster A corresponded to the Beijing sublineage. Whole-genome sequencing (WGS) differentiated clusters due to active recent transmission, with low single-nucleotide polymorphism-based diversity (cluster C), from clusters involving long-term prevalent strains with higher diversity (clusters A, B). Prospective application in Panama of 3 tailored strain-specific PCRs targeting marker single-nucleotide polymorphisms identified from WGS data revealed that 31.4% of incident cases involved strains A-C and that the Beijing strain was highly represented and restricted mainly to Colon. Rational integration of MIRU-VNTR, WGS, and tailored strain-specific PCRs could be a new model for tuberculosis surveillance in countries without molecular/genomic epidemiology programs.

Loss of zebrafish Ataxin-7, a SAGA subunit responsible for SCA7 retinopathy, causes ocular coloboma and malformation of photoreceptors.

Polyglutamine (polyQ) expansion in Ataxin-7 (ATXN7) results in spinocerebellar ataxia type 7 (SCA7) and causes visual impairment. SCA7 photoreceptors progressively lose their outer segments (OSs), a structure essential for their visual function. ATXN7 is a subunit of the transcriptional coactivator Spt-Ada-Gcn5 Acetyltransferase complex, implicated in the development of the visual system in flies. To determine the function of ATXN7 in the vertebrate eye, we have inactivated ATXN7 in zebrafish. While ATXN7 depletion in flies led to gross retinal degeneration, in zebrafish, it primarily results in ocular coloboma, a structural malformation responsible for pediatric visual impairment in humans. ATXN7 inactivation leads to elevated Hedgehog signaling in the forebrain, causing an alteration of proximo-distal patterning of the optic vesicle during early eye development and coloboma. At later developmental stages, malformations of photoreceptors due to incomplete formation of their OSs are observed and correlate with altered expression of crx, a key transcription factor involved in the formation of photoreceptor OS. Therefore, we propose that a primary toxic effect of polyQ expansion is the alteration of ATXN7 function in the daily renewal of OS in SCA7. Together, our data indicate that ATXN7 plays an essential role in vertebrate eye morphogenesis and photoreceptor differentiation, and its loss of function may contribute to the development of human coloboma.

Zebrafish P54 RNA helicases are cytoplasmic granule residents that are required for development and stress resilience.

Stress granules are cytoplasmic foci that directly respond to the protein synthesis status of the cell. Various environmental insults, such as oxidative stress or extreme heat, block protein synthesis; consequently, mRNA will stall in translation, and stress granules will immediately form and become enriched with mRNAs. P54 DEAD box RNA helicases are components of RNA granules such as P-bodies and stress granules. We studied the expression, in cytoplasmic foci, of both zebrafish P54 RNA helicases (P54a and P54b) during development and found that they are expressed in cytoplasmic granules under both normal conditions and stress conditions. In zebrafish embryos exposed to heat shock, some proportion of P54a and P54b helicases move to larger granules that exhibit the properties of genuine stress granules. Knockdown of P54a and/or P54b in zebrafish embryos produces developmental abnormalities restricted to the posterior trunk; further, these embryos do not form stress granules, and their survival upon exposure to heat-shock conditions is compromised. Our observations fit the model that cells lacking stress granules have no resilience or ability to recover once the stress has ended, indicating that stress granules play an essential role in the way organisms adapt to a changing environment.

A stem cell proliferation burst forms new layers of P63 expressing suprabasal cells during zebrafish postembryonic epidermal development.

Organ growth during development is a highly regulated process with both temporal and spatial constraints. Epidermal stratification is essential for skin growth and development. Although the zebrafish has been well studied, it is not known when and how epidermal stratification occurs. This is because beyond the first five days of development our knowledge is currently limited. We found that epidermal stratification in zebrafish begins when the larvae reach a standard length (SL) of 6 mm at approximately 25 days of age. Over the next four days (from a SL of 6 to 9 mm), epidermis thickness increases almost four-fold. This represents a sudden increase in organ size, since for the previous 20 days of development, the epidermis has been only two layers thick. This pattern is different from that observed in mammals that undergo continuous stratification from E14.5-E18.5. To study how stem cell proliferation gives rise to the new epidermal layers, we used a combination of markers: one for cell proliferation (proliferating cell nuclear-antigen PCNA) and one for epidermal stem cells (P63 transcription factor). We identified, throughout the stratification process, two different waves of cell division. Initially, the most basal epidermal cells divided and generated a subset of suprabasal cells (possibly transient-amplifying cells); within the next several days, the basal cells stopped dividing, and the suprabasal cells began proliferation, giving rise to most of the cell types in the new layers. This part of the process is similar to what has been recently found during epidermal stratification in mammals.

High clustering rates of multidrug-resistant Mycobacterium tuberculosis genotypes in Panama.

BACKGROUND: Tuberculosis continues to be one of the leading causes of death worldwide and in the American region. Although multidrug-resistant tuberculosis (MDR-TB) remains a threat to TB control in Panama, few studies have focused in typing MDR-TB strains. The aim of our study was to characterize MDR Mycobacterium tuberculosis clinical isolates using PCR-based genetic markers. METHODS: From 2002 to 2004, a total of 231 Mycobacterium tuberculosis isolates from TB cases country-wide were screened for antibiotic resistance, and MDR-TB isolates were further genotyped by double repetitive element PCR (DRE-PCR), (GTG)5-PCR and spoligotyping. RESULTS: A total of 37 isolates (0.85%) were resistant to both isoniazid (INH) and rifampicin (RIF). Among these 37 isolates, only two (5.4%) were resistant to all five drugs tested. Dual genotyping using DRE-PCR and (GTG)5-PCR of MDR Mycobacterium tuberculosis isolates revealed eight clusters comprising 82.9% of the MDR-TB strain collection, and six isolates (17.1%) showed unique fingerprints. The spoligotyping of MDR-TB clinical isolates identified 68% as members of the 42 (LAM9) family genotype. CONCLUSION: Our findings suggest that MDR Mycobacterium tuberculosis is highly clustered in Panama's metropolitan area corresponding to Panama City and Colon City, and our study reveals the genotype distribution across the country.

The zebrafish mutants for the V-ATPase subunits d, ac45, E, H and c and their variable pigment dilution phenotype.

BACKGROUND: The V-ATPase is a proton pump that creates an acidic medium, necessary for lysosome function and vesicular traffic. It is also essential for several developmental processes. Many enzymes, like the V-ATPase, are assemblies of multiple subunits, in which each one performs a specific function required to achieve full activity. In the zebrafish V-ATPase 15 different subunits form this multimeric complex and mutations in any of these subunits induce hypopigmentation or pigment dilution phenotype. We have previously found variability in the pigment dilution phenotype among five of the V-ATPase zebrafish mutants. This work presents additional information about such differences and is an update from a previous report. FINDINGS: We describe the variable phenotype severity observed among zebrafish V-ATPase pigment dilution mutants studying mRNA expression levels from their corresponding genes. At the same time we carried out phylogenetic analysis for this genes. CONCLUSIONS: Based in the similarities between different pigment dilution mutants we suggest that there is an essential role for V-ATPases in melanosome biogenesis and melanocyte survival. Neither variable expression levels for the different V-ATPase subunits studied here or the presence of duplicated genes seems to account for the variable phenotype severity from this group of mutants. We believe there are some similarities between the pigment dilution phenotype from zebrafish V-ATPase insertional mutants and pigment mutants obtained in a chemical screening ("Tubingen pigmentation mutants"). As for some of these "Tubingen mutants" the mutated gene has not been found we suggest that mutations in V-ATPase genes may be inducing their defects.