Tuberculosis (TB), caused by Mycobacterium tuberculosis (MTB), is one of the top 10 causes of death worldwide. Drug-resistant tuberculosis (DR-TB) poses a major threat to the World Health ...Organization's “End TB” strategy which has defined its target as the year 2035. In 2019, there were close to 0.5 million cases of DRTB, of which 78% were resistant to multiple TB drugs. The traditional culture-based drug susceptibility test (DST - the current gold standard) often takes multiple weeks and the necessary laboratory facilities are not readily available in low-income countries. Whole genome sequencing (WGS) technology is rapidly becoming an important tool in clinical and research applications including transmission detection or prediction of DR-TB. For the latter, many tools have recently been developed using curated database(s) of known resistance conferring mutations. However, documenting all the mutations and their effect is a time-taking and a continuous process and therefore Machine Learning (ML) techniques can be useful for predicting the presence of DR-TB based on WGS data. This can pave the way to an earlier detection of drug resistance and consequently more efficient treatment when compared to the traditional DST.
Five mycobacterial isolates from sewage were classified as members of the genus
but presented inconclusive species assignments. Thus, the isolates (MYC017, MYC098, MYC101, MYC123 and MYC340) were ...analyzed by phenotypical, biochemical, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and genomic features to clarify their taxonomic position. Phenotypic analysis and biochemical tests did not distinguish these isolates from other non-pigmented mycobacteria. In contrast, MALDI-TOF MS analysis showed that isolates were not related to any previously described
species. Comparative genomic analysis showed values of ANI and dDDH between 81.59-85.56% and 24.4-28.8%, respectively, when compared to the genomes of species of this genus. In addition, two (MYC101 and MYC123) presented indistinguishable protein spectra from each other and values of ANI = 98.57% and dDDH = 97.3%, therefore being considered as belonging to the same species. Phylogenetic analysis grouped the five isolates within the
complex (MTC) but in a specific subclade and separated from the species already described and supported by 100% bootstrap value, confirming that they are part of this complex but different from earlier described species. According to these data, we propose the description of four new species belonging to the
genus: (i)
sp. nov. strain MYC017
(= ATCC TSD-296
= JCM 35364
), (ii)
sp. nov. strain MYC098
(= ATCC TSD-297
= JCM 35365T), (iii)
sp. nov. strain MYC101
(= ATCC TSD-298
= JCM 35366
) and MYC123 (= ATCC BAA-3216 = JCM 35367); and (iv)
sp. nov. strain MYC340
(= ATCC TSD-299
= JCM 35368
).
Brazil has the second largest number of leprosy cases worldwide, and the state of São Paulo has been considered non-endemic since 2006.
We analyzed 16 variable number tandem repeats loci and three ...single nucleotide polymorphisms loci of Mycobacterium leprae (M. leprae) in 125 clinical isolates from patients in different municipalities in the state.
The clustering pattern of M. leprae indicated that the transmission of leprosy persisted in the state and included scenarios of intra-extra-familial transmission in areas with low endemicity.
A significantly active circulation of M. leprae was observed. Therefore, surveillance and control measures must be implemented.
Rio de Janeiro is endemic for tuberculosis (TB) and presents the second largest prevalence of the disease in Brazil. Here, we present the bacterial population structure of 218 isolates of ...Mycobacterium tuberculosis, derived from 186 patients that were diagnosed between January 2008 and December 2009. Genotypes were generated by means of spoligotyping, 24 MIRU-VNTR typing and presence of fbpC103, RDRio and RD174. The results confirmed earlier data that predominant genotypes in Rio de Janeiro are those of the Euro American Lineages (99%). However, we observed differences between the classification by spoligotyping when comparing to that of 24 MIRU-VNTR typing, being respectively 43.6% vs. 62.4% of LAM, 34.9% vs. 9.6% of T and 18.3% vs. 21.5% of Haarlem. Among isolates classified as LAM by MIRU typing, 28.0% did not present the characteristic spoligotype profile with absence of spacers 21 to 24 and 32 to 36 and we designated these conveniently as "LAM-like", 79.3% of these presenting the LAM-specific SNP fbpC103. The frequency of RDRio and RD174 in the LAM strains, as defined both by spoligotyping and 24 MIRU-VNTR loci, were respectively 11% and 15.4%, demonstrating that RD174 is not always a marker for LAM/RDRio strains. We conclude that, although spoligotyping alone is a tool for classification of strains of the Euro-American lineage, when combined with MIRU-VNTRs, SNPs and RD typing, it leads to a much better understanding of the bacterial population structure and phylogenetic relationships among strains of M. tuberculosis in regions with high incidence of TB.
Leprosy is an infectious disease caused by Mycobacterium leprae and frequently resulting in irreversible deformities and disabilities. Ticks play an important role in infectious disease transmission ...due to their low host specificity, worldwide distribution, and the biological ability to support transovarial transmission of a wide spectrum of pathogens, including viruses, bacteria and protozoa. To investigate a possible role for ticks as vectors of leprosy, we assessed transovarial transmission of M. leprae in artificially-fed adult female Amblyomma sculptum ticks, and infection and growth of M. leprae in tick cell lines. Our results revealed M. leprae RNA and antigens persisting in the midgut and present in the ovaries of adult female A. sculptum at least 2 days after oral infection, and present in their progeny (eggs and larvae), which demonstrates the occurrence of transovarial transmission of this pathogen. Infected tick larvae were able to inoculate viable bacilli during blood-feeding on a rabbit. Moreover, following inoculation with M. leprae, the Ixodes scapularis embryo-derived tick cell line IDE8 supported a detectable increase in the number of bacilli for at least 20 days, presenting a doubling time of approximately 12 days. As far as we know, this is the first in vitro cellular system able to promote growth of M. leprae. Finally, we successfully transformed a clinical M. leprae isolate by inserting the reporter plasmid pCHERRY3; transformed bacteria infected and grew in IDE8 cells over a 2-month period. Taken together, our data not only support the hypothesis that ticks may have the potential to act as a reservoir and/or vector of leprosy, but also suggest the feasibility of technological development of tick cell lines as a tool for large-scale production of M. leprae bacteria, as well as describing for the first time a method for their transformation.
Mycobacterium bovis is the causative agent of bovine tuberculosis (BTB), the pathogen responsible for serious economic impact on the livestock sector. In order to obtain data on isolated M. bovis ...strains and assist in the control and eradication program for BTB, a cross sectional descriptive molecular epidemiology study in the Brazilian Midwest was conducted. Through spoligotyping and 24-loci MIRU-VNTR methods, 37 clinical isolates of M. bovis circulating in the region were analyzed, 10 isolated from the state of Mato Grosso, 12 from the state of Mato Grosso do Sul and 15 from the state of Goiás. The spoligotyping analysis identified 10 distinct M. bovis profiles (SB0121 n = 14, SB0295 n = 6, SB0140 n = 6, SB0881 n = 3, SB1144 n = 2, SB1145 n = 2, SB0134 n = 1, SB1050 n = 1, SB1055 n = 1, SB1136 n = 1) grouped in six clusters and four orphan patterns. The MIRU-VNTR 24-loci grouped the same isolates in six clusters and 22 unique orphan patterns, showing higher discriminatory power than spoligotyping. When associating the results of both techniques, the isolates were grouped in five clusters and 24 unique M. bovis profiles. Among the 24-loci MIRU-VNTR evaluated, two, ETR-A and QUB 11b loci, showed high discriminatory ability (h = ≥ 0.50), while MIRU 16, MIRU 27, ETR-B, ETR-C, Mtub21 and QUB 26 loci showed moderate ability (h = 0.33 or h = 0.49) and were the most effective in evaluating the genotypic similarities among the clinical M. bovis isolate samples. Herein, the 29 patterns found amongst the 37 isolates of M. bovis circulating in the Brazilian Midwest can be due to the animal movement between regions, municipalities and farms, thus causing the spread of various M. bovis strains in herds from Midwest Brazil.
Isoniazid (INH), one of the most important drugs used in antituberculosis (anti-TB) treatment, is also the major drug involved in hepatotoxicity. Differences in INH-induced toxicity have been ...attributed to genetic variability at several loci, such as NAT2, CYP2E1, GSTM1 and GSTT1, that code for drug-metabolising enzymes. Our goal was to examine the polymorphisms in these enzymes as susceptibility factors to anti-TB drug-induced hepatitis in Brazilian individuals. In a case-control design, 167 unrelated active tuberculosis patients from the University Hospital of the Federal University of Rio de Janeiro, Brazil, were enrolled in this study. Patients with a history of anti-TB drug-induced acute hepatitis (cases with an increase to 3 times the upper limit of normal serum transaminases and symptoms of hepatitis) and patients with no evidence of anti-TB hepatic side effects (controls) were genotyped for NAT2, CYP2E1, GSTM1 and GSTT1 polymorphisms. Slow acetylators had a higher incidence of hepatitis than intermediate/rapid acetylators 22% (18/82) vs. 9.8% (6/61), odds ratio (OR), 2.86, 95% confidence interval (CI), 1.06-7.68, p = 0.04). Logistic regression showed that slow acetylation status was the only independent risk factor (OR 3.59, 95% CI, 2.53-4.64, p = 0.02) for the occurrence of anti-TB drug-induced hepatitis during anti-TB treatment with INH-containing schemes in Brazilian individuals.
Introduction:
Several polymorphisms altering the
NAT2
activity have already been identified. The geographical distribution of
NAT2
variants has been extensively studied and has been demonstrated to ...vary significantly among different ethnic population. Here, we describe the genetic variability of human N-acetyltransferase 2 (
NAT2
) gene and the predominant genotype-deduced acetylation profiles of Brazilians.
Methods:
A total of 964 individuals, from five geographical different regions, were genotyped for
NAT2
by sequencing the entire coding exon.
Results:
Twenty-three previously described NAT2 single nucleotide polymorphisms (SNPs) were identified, including the seven most common ones globally (c.191G>A, c.282C>T, c.341T>C, c.481C>T, c.590G>A, c.803A>G and c.857G>A). The main allelic groups were NAT2*5 (36%) and NAT2*6 (18.2%), followed to the reference allele
NAT2*4
(20.4%). Combined into genotypes, the most prevalent allelic groups were
NAT2*5/*5
(14.6%),
NAT2*5/*6
(11.9%) and
NAT2*6/*6
(6.2%). The genotype deduced
NAT2
slow acetylation phenotype was predominant but showed significant variability between geographical regions. The prevalence of slow acetylation phenotype was higher in the Northeast, North and Midwest (51.3%, 45.5% and 41.5%, respectively) of the country. In the Southeast, the intermediate acetylation phenotype was the most prevalent (40.3%) and, in the South, the prevalence of rapid acetylation phenotype was significantly higher (36.7%), when compared to other Brazilian states (
p
< 0.0001). Comparison of the predicted acetylation profile among regions showed homogeneity among the North and Northeast but was significantly different when compared to the Southeast (
p
= 0.0396). The Southern region was significantly different from all other regions (
p
< 0.0001).
Discussion:
This study contributes not only to current knowledge of the
NAT2
population genetic diversity in different geographical regions of Brazil, but also to the reconstruction of a more accurate phenotypic picture of
NAT2
acetylator profiles in those regions.
The World Health Organization (WHO) has classified human zoonotic tuberculosis (TB) due to Mycobacterium bovis as a neglected issue in the developing world. In a recent cross-sectional study in ...Brazil, three of 189 TB patients presented with a coinfection of M. bovis and M. tuberculosis and were selected as cases for this study.
The aim was to evaluate risk factors (RF) for zoonotic TB in an urban area of Brazil in order to guide preventive programmes.
A matched case-control study was carried out nested within a cross-sectional study. For each of the three cases, 14 age- and sex-matched controls (TB due to M. tuberculosis) were selected.
Zoonotic potential exposures (ZE) and extrapulmonary TB (EPTB) were independently associated with zoonotic TB in multivariate analyses.
ZE by occupation and consumption of raw milk and derivative products that place individuals in direct and indirect contact with animals and their excretions/secretions increase the risk for zoonotic TB in Brazil, especially among those with EPTB. Therefore, measures such as efficient control of bovine TB, distribution of pasteurised milk and its derivative products, and the diagnosis and monitoring of zoonotic TB in humans are essential steps, especially in developing countries where bovine TB is enzootic, and further studies are necessary.
The Latin American Mediterranean (LAM) spoligotype family is one of the most successful genotype of Mycobacterium tuberculosis worldwide and particularly prevalent in South-America. Within this ...family, a sublineage named Region of Difference Rio (RD.sup.Rio) was reported initially in Brazil and is characterized by a genomic deletion of about 26.3 kb. This lineage seems to show a specific adaptation to the Euro-Latin American population. In this context, we sought to evaluate the LAM family and the presence of the RD.sup.Rio genotype in samples from three Latin American countries including Paraguay, Venezuela and Argentina. To detect LAM strains reliably we applied a typing scheme using spoligotyping, 12 loci MIRU-VNTR, the Ag85C.sup.103 SNP and the regions of difference RD.sup.Rio and RD174. IS6110-RFLP results were also used when available. Genotyping of 413 M. tuberculosis isolates from three Latin-American countries detected LAM (46%) and the ill-defined T clade (16%) as the most frequent families. The highest clustering rate was detected in the sample population from the city of Caracas in Venezuela. We observed considerable differences in the presence of the RD.sup.Rio lineage, with high frequency in Caracas-Venezuela (55%) and low frequency in Buenos Aires-Argentina (11%) and Paraguay (10%). The molecular markers (RD174, Ag85C.sup.103, MIRU02-MIRU40 signature) of the RD.sup.Rio lineage were essentially confirmed. For the LAM family, the most polymorphic loci were MIRU40, MIRU31, MIRU10, MIRU26, MIRU16 and the least polymorphic MIRU24, MIRU20, MIRU04, MIRU23. Our results suggest a differential adaptation of LAM-sublineages in neighboring populations and that RD.sup.Rio strains spread regionally with different rates of distribution. The Ag85C SNP and RDs (RD174, RD.sup.Rio) tested in this study can in fact facilitate molecular epidemiological studies of LAM strains in endemic settings and low-income countries.
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