Search Results (32)

This study investigated the genetic diversity of Mycobacterium avium subsp. paratuberculosis (Map)—the causative agent of paratuberculosis—isolated from different host species in Germany. A total of 500 isolates from 243 cattle herds and 9 other host species originating from 13 federal states were genotyped. A multi-target approach was applied, comprising IS900-RFLP with BstEII and PstI digestion; MIRU-VNTR; and SSR1, SSR8, and SSR9 analysis. In total, 93 combined genotypes were identified, 84 in cattle and 21 in non-cattle isolates. Ninety genotypes were assigned to the C-type group, and three genotypes (three from sheep and one from cattle) were assigned to the S-type/subtype III group. Cluster analysis divided genotypes into subgroups similar to those shown for WGS-SNP-based phylogenetic trees. New genotypes were revealed, including INMV262–267 and a specific sequence at locus VNTR7. Five genotypes that were predominant in cattle were also detected in sheep, goats, and deer. The majority of genotypes [61%] were identified only once. Polyclonal infections were observed in individual animals and herds, and various potential Map transmission linkages were uncovered. This high genotype richness of Map reflects the long history of paratuberculosis in Germany and intensive nationwide animal movement and international trading activity. Full article

Mycobacterium is one of the most dangerous pathogens of both animals and humans. Bovine tuberculosis (BTB) is a disease caused by mycobacteria belonging to the Mycobacterium tuberculosis complex (MTBC), which spreads mainly among domestic cattle but also to mammals other than cattle. The transmission of MTBC between different species requires research and epidemiological investigations to control its spread. When multiple species are a reservoir of infection, it poses a significant public health and veterinary concern. In this study, the diagnosis of alpaca, cattle, horses, dogs, a sheep and a cat from one farm suspected of bovine tuberculosis was performed. The animals (except for one horse, the dogs and the cat) were euthanised after the intradermal tuberculin tests. Mycobacterial isolation from animal tissue samples was performed. The obtained Mycobacterium strains were genotyped using spoligotyping and mycobacterial interspersed repetitive unit–variable number tandem repeat (MIRU-VNTR) methods. The isolates from a horse, two cows, a sheep and an alpaca were classified as Mycobacterium (M.) bovis. The single M. bovis spoligotype SB0666 pattern was isolated, and the MIRU-VNTR results presented the same 222632237401435 patterns. The molecular investigation uncovered information on the relationship of Mycobacterium bovis. Full article

Background Members of the Mycobacterium avium complex (MAC) have been documented to cause severe and disseminated infections in dogs, although such cases are sporadically reported. In this study, a comprehensive account of a rare case of generalised lymphadenomegaly caused by a mixed-strain infection with drug- and multidrug-resistant Mycobacterium avium subspecies hominissuis (Mah) in a Maremma sheepdog is presented. Methods Laboratory investigations, as well as the monitoring of the clinical signs displayed by the animal, were conducted throughout the course of a two-year drug therapy (based on rifampicin, azithromycin, and ciprofloxacin) and a two-year post-treatment follow-up period, until the death of the dog. Laboratory examinations included both solid and broth cultures from fine-needle aspiration samples of lymph nodes, molecular typing by 8-locus MIRUVNTR analysis and SNPs typing of five genetic regions (gyrB, rpsA, 3′hsp65, ITS and rpoB), and drug susceptibility testing towards seven antimycobacterial drugs. Results The results indicated the presence of two distinct genotypes of Mah, which exhibited different phenotypic characteristics, such as different drug susceptibility profiles and growth abilities in broth and solid media, suggesting a mixed-strain infection. Resistances to ethambutol alone, to ethambutol and clarithromycin, and to ethambutol, clarithromycin, rifampicin, and doxycycline were detected over the study. Conclusions Although the Mah strains isolated during the course of therapy showed sensitivity to the regiment, the complete eradication of the infection was never achieved. It has been hypothesised that the presence of drug-resistant and multidrug-resistant Mah strains in the animal may have been established at the onset of the infection or soon thereafter. The exposure to therapy has been suggested as a potential factor that could have favoured the growth of resistant strains, thereby rendering the therapy ineffective. The implications that the distinct phenotypic and genotypic profiles of Mah described here may have had for disease dynamics and control are discussed. Full article

A previous study in Pará, Northern Brazil, described a strain of Mycobacterium tuberculosis with a unique genotype (SIT2517/T1) associated with multidrug-resistant tuberculosis (MDR-TB). To improve our understanding of MDR-TB transmission dynamics of these strains within this region, we performed phenotypic and genotypic drug susceptibility testing (pDST/gDST), 24-loci mycobacterial interspersed repetitive units (MIRU-VNTR) genotyping, whole-genome sequencing (WGS) and geo-epidemiology analysis. Of the 28 SIT2517/T1 isolates, 19 (67.9%) could be genotyped by 24-loci MIRU-VNTR and 15 by WGS. All belonged to sublineage 4.1.1.3, distinct from other representative Lineage 4 isolates identified in Brazil. The MDR phenotype determined by pDST was confirmed by gDST, the latter also demonstrating the presence of additional mutations conferring pre-extensively drug-resistance (pre-XDR). Discrepancies between gDST and pDST were observed for pyrazinamide and fluoroquinolones. Thirteen out of 15 isolates analyzed by WGS were clustered when applying a 12 single nucleotide polymorphisms (SNPs) cutoff. The SIT2517/T1 isolates were distributed across the metropolitan regions of Belém and Collares municipalities, showing no geographic clustering. WGS-transmission network analysis revealed a high likelihood of direct transmission and the formation of two closely linked transmission chains. This study highlights the need to implement TB genomic surveillance in the Brazilian Amazon region. Full article

There has been very limited investigation regarding the genetic diversity of Mycobacterium tuberculosis (MTb) strains isolated from human immunodeficiency virus (HIV)-infected patients in Mexico. In this study, we isolated 93 MTb strains from pulmonary and extrapulmonary samples of HIV-infected patients treated in a public hospital in Mexico City to evaluate the genetic diversity using spoligotyping and mycobacterial interspersed repetitive unit-variable-number tandem-repeat (MIRU-VNTR) typing (based on 24 loci). The cohort comprised 80 male and 13 female individuals. There was a positive correlation between a high HIV viral load (>100,000 copies) and extrapulmonary tuberculosis (TB) (r = 0.306, p = 0.008). Lineage 4 was the most frequent lineage (79 strains). In this lineage, we found the H clade (n = 24), including the Haarlem, H3, and H1 families; the T clade (n = 22), including T1 and T2; the X clade (n = 15), including X1 and X3; the LAM clade (n = 14), including LAM1, LAM2, LAM3, LAM6, and LAM9; the S clade (n = 2); Uganda (n = 1); and Ghana (n = 1). We also found 12 strains in the EAI clade belonging to lineage 1, including the EAI2-Manila and EAI5 families. Interestingly, we identified one strain belonging to the Beijing family, which is part of lineage 2. One strain could not be identified. This study reports high genetic diversity among MTb strains, highlighting the need for a molecular epidemiological surveillance system that can help to monitor the spread of these strains, leading to more appropriate measures for TB control in HIV-infected patients. Full article

The Beijing genotype is the most distributed M. tuberculosis family in Kazakhstan. In this study, we identified dominant Beijing clusters in Kazakhstan and assessed their drug susceptibility profiles and association with the most widely spread mutation Ser531Leu of the rpoB gene and the mutation Ser315Thr of the katG gene associated with resistance to rifampicin and isoniazid, respectively. M. tuberculosis isolates (n = 540) from new TB cases were included in the study. MIRU-VNTR genotyping was performed for 540 clinical isolates to determine M. tuberculosis families using 24 loci. RD analysis was additionally performed for the Beijing isolates. The identification of mutations in the drug-resistance genes of M. tuberculosis was performed with allele-specific real-time PCR and Sanger sequencing. The Beijing genotype was identified in 60% (324/540) of the clinical isolates. Central Asian/Russian cluster 94-32 was the most distributed cluster among the Beijing isolates (50.3%; 163/324). Three other dominant Beijing clusters were identified as 94-33 (3.4%; 11/324), 100-32 (3.1%; 10/324) and 99-32 (3.1%; 10/324). The Beijing genotype was associated with drug-resistant TB (p < 0.0001), including multidrug-resistant TB (p < 0.0001), in our study. An association of the mutation Ser531Leu of the rpoB gene with the Beijing genotype was found (p < 0.0001; OR = 16.0000; 95%CI: 4.9161–52.0740). Among the Beijing isolates, cluster 94-32 showed an association with MDR-TB (p = 0.021). This is why the evaluation of the Beijing genotype and its clusters is needed to control MDR-TB in Kazakhstan. Full article

This study investigated the intra- and inter-herd diversity of Mycobacterium avium subsp. paratuberculosis (MAP) isolates from four goat herds in Thuringia (Germany) that were affected by paratuberculosis for several years. The main focus was on the characterization and distribution of genotypes among animals and the environment of goat herd 1. This study included 196 isolates from the feces of 121 infected goats, various tissues from 13 clinically diseased goats, 29 environmental samples from herd 1, and additionally, 22 isolates of different origin from herds 2 to 4. The isolates, sampled between 2018 and 2022, were genotyped using short-sequence-repeat (SSR) analysis, mycobacterial-interspersed repetitive units–variable-number tandem repeat (MIRU–VNTR) analysis, and a single nucleotide polymorphism (SNP)-based assay for phylogenetic grouping. All the isolates belonged to the MAP-C group. In herd 1, one predominant genotype was determined, while two other genotypes were identified very rarely and only in fecal and environmental samples. One of three further genotypes was found in each of herds 2 to 4. The assignment of genotypes to different phylogenetic clades suggested six different infection strains. The results indicated no epidemiological links between the examined herds. Based on the current MAP genotyping data from Germany, possible sources of infection are MAP-contaminated barns previously used by infected cattle and the purchase of sub-clinically infected goats. Full article

Non-tuberculous mycobacteria (NTM) are ubiquitous organisms, of which some, especially those of the Mycobacterium avium complex (MAC), may be opportunistic animal and human pathogens. Infection with NTM can interfere with tuberculosis (TB) diagnosis and induce zoonoses, especially in immunocompromised individuals. Diseases caused by NTM have become more readily recognized; however, they are likely still underestimated. In this study, we identified and genotyped Mycobacterium avium strains that were isolated during TB monitoring among free-living carnivorous animals from southeastern Poland. In 2011–2020, lymph node samples from 192 such animals were tested for mycobacteria. A total of 41 isolates of M. avium strains were detected with the use of IS901, IS900, IS1245, and mycobacterial interspersed repeat unit–variable number tandem repeat (MIRU-VNTR) identification. Thirty-three were identified as M. avium subsp. avium. These strains were derived from 1 beech marten (Martes foina), 1 common buzzard (Buteo buteo), 2 European badgers (Meles meles), 3 wolves (Canis lupus), and 26 red foxes (Vulpes vulpes). One strain isolated from a wolf was identified as M. avium subsp. hominissuis. The results show the widespread occurrence of MAC bacilli in the studied environment and additionally comprise new data on the molecular characteristics of M. avium subspecies carried by free-living southeastern Polish carnivores. Full article

Rapid and accurate detection of tuberculosis (TB) drug resistance is critical for the successful treatment and control of TB. Here, we investigated resistance to anti-TB drugs and genetic variations in 215 drug-resistant Mycobacterium tuberculosis isolates in Korea. Genetic variations were observed in rpoB Ser531Leu, katG Ser315Thr, and gyrA Asp94Gly; however, the minimum inhibitory concentrations varied, which can be attributed to other resistance mechanisms. Examination of genetic relatedness among drug-resistant isolates revealed that the cluster size of resistant bacteria was less than six strains, suggesting no evidence of a large-scale epidemic caused by a specific strain. However, rpoC mutants of the rifampicin-resistant isolates were composed of five types of clusters, suggesting that these compensatory mutations advance propagation. In the present study, more than 90% of the resistance mechanisms to major anti-TB drugs were identified, and the effect of each mutation on drug resistance was estimated. With the clinical application of recent next-generation sequencing-based susceptibility testing, the present study is expected to improve the clinical utilization of genotype-based drug susceptibility testing for the diagnosis and treatment of patients with drug-resistant TB. Full article

Bovine tuberculosis is endemic in Nigeria with control measures as provided by the laws of the country being minimally enforced mostly at the abattoirs only. This study focused on bovine tuberculosis in Adamawa and Gombe States. Tuberculosis lesions were observed in 183 of 13,688 slaughtered cattle in the regions between June and December 2020. Analysis of tissue samples resulted in 17 Mycobacterium bovis isolates, predominantly from Gombe State. Spoligotyping identified four spoligotypes, including SB0944, SB1025, SB1104, and one novel pattern. MIRU-VNTR analysis further differentiated these spoligotypes into eight profiles. All isolates belonged to the Af1 clonal complex. The study emphasises the need for broader coverage and more isolates to comprehensively understand the molecular epidemiology of bovine tuberculosis in Nigeria. To enhance research and surveillance, a cost-effective approach is proposed, utilising a discriminatory VNTR panel comprising five or nine loci. The five-locus panel consists of ETR-C, QUB26, QUB11b, MIRU04, and QUB323. Alternatively, the nine-locus panel includes ETR-A, ETR-B, QUB11a, and MIRU26. Implementing this approach would provide valuable insights into the genetic diversity of M. bovis strains in Nigeria. These findings are crucial for developing effective control measures and minimising the impact of bovine tuberculosis on both animal and human health. Full article

Mycobacterium bovis infects cattle and wildlife, and also causes a small proportion of tuberculosis cases in humans. In most European countries, M. bovis infections in cattle have been drastically reduced, but not eradicated. Here, to determine the M. bovis circulation within and between the human, cattle, and wildlife compartments, we characterized by spoligotyping and mycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-VNTR) typing the genetic diversity of M. bovis isolates collected from humans, cattle, and wildlife in France from 2000 to 2010. We also assessed their genetic structure within and among the different host groups, and across time and space. The M. bovis genetic structure and its spatiotemporal variations showed different dynamics in the human and animal compartments. Most genotypes detected in human isolates were absent in cattle and wildlife isolates, possibly because in patients, M. bovis infection was contracted abroad or was the reactivation of an old lesion. Therefore, they did not match the genetic pool present in France during the study period. However, some human-cattle exchanges occurred because some genotypes were common to both compartments. This study provides new elements for understanding M. bovis epidemiology in France, and calls for increased efforts to control this pathogen worldwide. Full article

The aim of this study was to assess the diversity of minisatellite VNTR loci in Mycobacterium bovis/M. caprae isolates in Bulgaria and view their position within global M. bovis diversity. Forty-three M. bovis/M. caprae isolates from cattle in different farms in Bulgaria were collected in 2015–2021 and typed in 13 VNTR loci. The M. bovis and M. caprae branches were clearly separated on the VNTR phylogenetic tree. The larger and more geographically dispersed M. caprae group was more diverse than M. bovis group was (HGI 0.67 vs. 0.60). Overall, six clusters were identified (from 2 to 19 isolates) and nine orphans (all loci-based HGI 0.79). Locus QUB3232 was the most discriminatory one (HGI 0.64). MIRU4 and MIRU40 were monomorphic, and MIRU26 was almost monomorphic. Four loci (ETRA, ETRB, Mtub21, and MIRU16) discriminated only between M. bovis and M. caprae. The comparison with published VNTR datasets from 11 countries showed both overall heterogeneity between the settings and predominantly local evolution of the clonal complexes. To conclude, six loci may be recommended for primary genotyping of M. bovis/M. caprae isolates in Bulgaria: ETRC, QUB11b, QUB11a, QUB26, QUB3232, and MIRU10 (HGI 0.77). VNTR typing based on a limited number of loci appears to be useful for primary bTB surveillance. Full article

Mycobacterium tuberculosis (Mtb) Central Asian Strain (CAS) Lineage 3 (L3) genotype is predominantly found in East-Africa, Central-Asia, Western-Asia, and South-Asia; however, a new spoligotyping CAS/SIT2545 was found in northern regions of Brazil. We aimed to characterize and describe the genetic diversity and perform a phylogenetic assessment of this novel genotype. We performed 24-MIRU-VNTR loci and Whole-genome sequencing (WGS) of six Brazilian isolates previously spoligotyped. The libraries were prepared using a Nextera-XT kit and sequenced in a NextSeq 550 Illumina instrument. We performed lineage assignment and genomic characterization. From publicly available genomes of Mtb L3 and other lineages, we created a robust dataset to run the MTBSeq pipeline and perform a phylogenetic analysis. MIRU-VNTR and WGS confirmed CAS/SIT2545 belongs to L3. Out of 1691 genomes, 1350 (79.83%) passed in quality control (genomic coverage > 95%). Strain 431 differed in 52 single nucleotide variants (SNV), confirming it does not belong to the same transmission chain. The eight genomes from a global dataset clustered closer to Brazilian strains differed in >52 SNVs. We hypothesized L3 and L1 were introduced in Brazilian Northern in the same historical event; however, there is a need for additional studies exploring the genetic diversity of Mtb Brazilian Northern. Full article

Introduction: Zoonoses have recently become an increasing public health problem. Zoonoses are estimated to account for 60% of all emerging infectious diseases. One particularly important zoonosis is human tuberculosis, especially tuberculosis due to Mycobacterium bovis (M. bovis), which is naturally resistant to pyrazinamide (PZA). Material and Methods: The patient had a pulmonary form of tuberculosis accompanied by a cough and fever. At the same time, the disease was also confirmed in 20 out of 25 cattle on the farm. The clinical specimen (sputum) was examined in accordance with the European Union (EU) laboratories’ methodology. Tissue materials from cattle were verified in the National Veterinary Research Institute (NVRI), in the Bovine tuberculosis (BTB) Reference Laboratory, Pulawy, Poland and tested in accordance with the guidelines for the laboratory diagnosis of BTB. Results: All M. bovis isolates represented one spoligotype, SB0120. The results of mycobacterial interspersed repetitive unit variable number tandem repeat (MIRU-VNTR) evaluation showed the same genetic pattern. Conclusions: Findings from this study suggest the first confirmed interspecific transmission of Mycobacterium bovis, between a farmer and his cattle, in Poland. Present findings support the increasing concern regarding zoonotic TB that has been highlighted elsewhere. Full article

Mycobacterium tuberculosis complex (MTBC) Lineage 3 (L3) strains are abundant in world regions with the highest tuberculosis burden. To investigate the population structure and the global diversity of this major lineage, we analyzed a dataset comprising 2682 L3 strains from 38 countries over 5 continents, by employing 24-loci mycobacterial interspersed repetitive unit-variable number of tandem repeats genotyping (MIRU-VNTR) and drug susceptibility testing. We further combined whole-genome sequencing (WGS) and phylogeographic analysis for 373 strains representing the global L3 genetic diversity. Ancestral state reconstruction confirmed that the origin of L3 strains is located in Southern Asia and further revealed multiple independent introduction events into North-East and East Africa. This study provides a systematic understanding of the global diversity of L3 strains and reports phylogenetic variations that could inform clinical trials which evaluate the effectivity of new drugs/regimens or vaccine candidates. Full article