Spacer Oligonucleotide Typing Research Articles

Molecular epidemiology and drug resistance of Mycobacterium tuberculosis in a tertiary care hospital in northeastern Mexico.

Tuberculosis (TB) is a re-emerging disease considered a public health concern. In the present study, we analyzed the epidemiology and drug resistance of Mycobacterium tuberculosis strains isolated from patients with pulmonary TB. Mycobacterium tuberculosis isolates (n = 190) were obtained from patients with pulmonary TB admitted to Dr. José Eleuterio González University Hospital (UH). Each M. tuberculosis isolate was analyzed by spoligotyping (spacer oligonucleotide typing) and MIRU-VNTR (Mycobacterial Interspersed Repetitive Units-Variable Number Tandem Repeat). Drug resistance was evaluated using the Anyplex™ II MTB/MDR/XDR assay. The predominant spoligotypes observed were X1 (SIT 119, n = 46), T1 (SIT 53, n = 40), H3 (SIT 50, n = 13), Beijing (SIT 1, n = 11), and EAI2-Manila (SIT 19, n = 8). MIRU-VNTR analysis showed that the locus QUB-26 had the highest allelic variability. The observed drug resistance included monoresistance to rifampicin (2.6%; n = 5), isoniazid (3.2%; n = 6), and fluoroquinolones (1.6%; n = 3) as well as multidrug resistance (5.3%; n = 10). All of the Beijing strains were susceptible. Regarding comorbidities, 13.7% (26/190) of the patients were co-infected with TB and HIV (TB+HIV+), and 31.6% (55/190) had TB along with diabetes (TB + diabetes). The most prevalent lineages were X1 (SIT 119; 24.3%) and T1 (SIT 53; 21%). An alarming proportion (12.6%) of M. tuberculosis isolates presented drug resistance. To effectively manage TB, continuous surveillance of regional strain dissemination, drug resistance profiles, and TB-associated comorbidities is crucial.

Single-tube protocol for culture-independent spoligotyping of Mycobacterium tuberculosis based on MeltArray.

Spacer oligonucleotide typing (spoligotyping), the first-line genotyping assay for Mycobacterium tuberculosis (MTB), plays a fundamental role in the investigation of its epidemiology and evolution. In this study, we established a single-tube spoligotyping assay using MeltArray, a highly multiplex polymerase chain reaction (PCR) approach that runs on a real-time PCR thermocycler. The MeltArray protocol included an internal positive control, gyrB, to indicate the abundance of MTB via the quantification cycle and 43 spacers to identify the spoligotype via melting curve analysis. The entire protocol was completed in a single step within 2.5 hours. The lowest detectable copy number for the tested strains was 20 copies/reaction and thus sufficient for analyzing both culture and sputum samples. We conclude that MeltArray-based spoligotyping could be used immediately in low- and middle-income countries with a high tuberculosis burden, given its easy access, improved throughput, and potential applicability to clinical samples.

Genotyping methods and their contributions to the study of tuberculosis dynamic in Latin America.

Mycobacterium tuberculosis genotyping has impacted evolutionary studies worldwide. Nonetheless, its application and the knowledge generated depend on the genetic marker evaluated and the detection technologies that have evolved over the years. Here we describe the timeline of main genotypic methods related to M. tuberculosis in Latin America and the main findings obtained. Systematic searches through the PubMed database were performed from 1993 to May 2021. A total of 345 articles met the inclusion criteria and were selected. Spacer oligonucleotide typing (spoligotyping) was the most widely used method in Latin America, with decreasing use in parallel with increasing use of mycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-VNTR) and whole genome sequencing (WGS). Among the countries, Brazil, Mexico, and Argentina had the most publications, and a considerable part of the articles were in collaboration with Latin American or non-Latin American institutions; a small proportion of studies needed partnerships to perform the genotypic methods. The genotypic methods allowed the identification of M. tuberculosis genotypes with greater capacity for clonal expansion and revealed the predominance of the Euro-American lineage in Latin America. There was a notable presence of the Beijing family in Peru and Colombia. The data obtained demonstrated the importance of expanding collaborative networks of tuberculosis (TB) research groups to countries with low productivity in this area, the commitment of the few Latin American countries to advance TB research, as well as the inestimable value of building a Latin America database, considering ease of population mobility between countries.

Frequency of Mycobacterium tuberculosis among Mycobacterium tuberculosis Complex Strains Isolated from Clinical Specimen

Background: Rapid and accurate detection of Mycobacterium tuberculosis (Mtb) is of primary importance for infection control and selection of anti-tuberculosis drugs. However, most clinical laboratories report Mtb complex (MTC) without reporting Mtb because MTC comprising Mtb, Mycobacterium bovis, Mycobacterium africanum, Mycobacterium microti, Mycobacterium caprae, and Mycobacterium pinnipedii have 99.9% similarity at the nucleotide level and identical 16S rRNA sequences. This study was conducted to analyze the species frequency of MTC isolates obtained from clinical specimens. Methods: Of 310 MTC isolates obtained from clinical samples in a tertiary care hospital from February 2017 to August 2018, MolecuTech Real TB-Taq (YD Diagnostics, Korea) real-time polymerase chain reaction (PCR) was performed,specifically to detect Mtb. For DNA showing Mtb negative results by Mtb-specific real-time PCR or pyrazinamide-resistant strains, PCR-based MTC typing, spacer oligonucleotide typing (spoligotyping), and exact tandem repeat D gene sequencing were performed. Results: All the 310 MTC isolates were identified to be Mtb. Two Mtb strains of the East-African-Indian 4-Vietnam genotype, which have not been reported in Korea, were also found. Conclusion: There was no zoonotic tuberculosis in this study. Since we investigated only 310 MTC isolates detected in only one medical institution, a multi-center study is needed to accurately know the prevalence of zoonotic tuberculosis in Korea.

Genetic surveillance and outcomes of pyrazinamide and fluoroquinolones-resistant tuberculosis in Taiwan

BackgroundPyrazinamide (PZA) and fluoroquinolone (FQ), particularly moxifloxacin (MXF), are essential drugs in the World Health Organization (WHO) recommended short-course regimen to treat drug-susceptible tuberculosis (TB). MethodsTo understand the extent of PZA and MXF susceptibility in general TB cases in Taiwan, we conducted retrospective analyses of 385 conservative Mycobacterium tuberculosis complex (MTBC) isolates identified from 4 TB laboratories in different regions of Taiwan. The case information was obtained from the TB registry. Genotypic drug susceptibility testing (DST) was performed by sequencing drug-resistance associated genes, PZA (pncA) and FQ (gyrA, and gyrB). Phenotypic DST was determined using the Bactec MGIT 960 system or the agar proportion method. Genotyping was carried out using spacer oligonucleotide typing. ResultsIn this study, 4.7% (18/385) cases' isolates harbored pncA mutations and 7.0% (27/385) cases’ isolates harbored gyrA or gyrB mutation. Notably, pncA mutation was associated with Beijing family genotypes (P = 0.028), East African-Indian (EAI) genotypes (P = 0.047) and MDR-TB (P < 0.001). Whereas, gyrA or gyrB mutation was associated with EAI genotypes (P = 0.020) and MDR-TB (P = 0.006). In addition, a statistically significant difference was found between the favorable outcomes using active and inactive PZA (P = 0.009) in 38 case isolates with any pncA, gyrA, or gyrB mutation. ConclusionWe concluded that routine PZA and FQ susceptibility tests are recommended for guiding the treatment of TB.

The potential risk of international spread of Mycobacterium bovis associated with movement of alpacas

IntroductionThe study highlights the transboundary nature of tuberculosis (TB) in alpacas and the failure of current ante-mortem testing protocols (the tuberculin skin and Enferplex Camelid TB tests) to identify TB-free alpaca herds and individuals for export. Our research and the available literature indicate that the alpaca (Vicugna pacos) is extremely susceptible to Mycobacterium bovis infection, and that testing periodicity fails to take into account that animals do not manifest disease symptoms for a long time. The skin test failed to identify Mycobacterium bovis infection in two alpacas prior to their movement from the UK to Poland. The animals were purchased by a breeding centre in Poland, and were then shown at an international animal exhibition. The last owner of the alpacas before their deaths from TB bought the infected animals unwittingly in order to run rehabilitation activities with disabled children on his farm.Material and MethodsThoracic lymph node, lung and liver tissue samples obtained at necropsy were examined histopathologically after Ziehl–Neelsen staining. Tissue samples were homogenised and mycobacteria present there were cultured on Stonebrink’s medium during a 6-week incubation. A commercial test using polymorphism of the chromosomal direct repeat region provided species identification and additional identification was by spacer oligonucleotide typing and mycobacteria interspersed repetitive unit–variable number tandem repeat analysis with a gel electrophoresis protocol.ResultsThe microbiological examination confirmed multiorgan TB caused by the SB0666 spoligotype of Mycobacterium bovis.ConclusionDue to the suboptimal performance of current diagnostic tests for TB in alpacas, there is a risk that infected animals may be moved unwittingly. A risk of TB spread associated with the international movement of alpacas is implied by this study.

Drug resistant tuberculosis cases from the Copperbelt province and Northern regions of Zambia: Genetic diversity, demographic and clinical characteristics

Tuberculosis (TB) caused by Mycobacterium tuberculosis remains a major cause of death worldwide. Diverse genotypes have been demonstrated to drive the epidemiology of drug resistant (DR-) TB globally. Currently, there is limited knowledge on the genotypes and transmission dynamics of M. tuberculosis in Zambia. This study aimed to describe the genotypes of DR-TB from the Copperbelt and Northern regions of Zambia.Molecular typing tools of insertion sequence 6110-restriction fragment length polymorphism (IS6110-RFLP) and spacer oligonucleotide typing (spoligotyping) were applied.We demonstrate that diverse genotypes are associated with DR-TB in Zambia. The predominant genotype was lineage 4; other strains belonged to lineage 2 and 3. Genotypes previously identified as driving the epidemiology of drug susceptible TB have been identified as drivers of DR-TB. Genotyping analysis showed clustering of strains among patients from different regions of the country; suggesting that DR-TB is widespread.Molecular findings combined with phenotypic and epidemiologic findings play a critical role in identifying circulating genotypes and possible transmission chains. Clustering of drug resistant strains was demonstrated to be 48% and 86% according to IS6110-RFLP and spoligotyping, respectively. However, gaps in clinical and demographic data skew the interpretation, and call for data collection policy improvements.

One-step melting curve analysis-based McSpoligotyping reveals genotypes of Mycobacterium tuberculosis in a coastal city, China.

A one-step 3h melting curve analysis-based technology for spacer oligonucleotide typing (McSpoligotyping) analyzed with the SITVIT2 database was applied to detect epidemiological events of Mycobacterium tuberculosis (MTB) in a coastal city of China from 2016 to 2018. 306 MTB isolates were identified by Fuzhou tuberculosis designated hospitals between 2016 and 2018. The results showed that the MTB isolates were divided into Beijing family and non-Beijing families, accounting for 45.42% (139/306) and 54.58% (167/306), respectively. H and T families were the prevalent genotypes in non-Beijing families. Herein, the spoligotyping technology has practical application for the classification and tracing of tuberculosis in the public.

Genetic characterization of Mycobacterium tuberculosis complex isolates circulating in Abuja, Nigeria.

ObjectiveNigeria ranks fourth among the high tuberculosis (TB) burden countries. This study describes the prevalence of drug resistance and the genetic diversity of Mycobacterium tuberculosis in Abuja’s Federal Capital Territory.Materials and methodsTwo hundred and seventy-eight consecutive sputum samples were collected from adults with presumptive TB during 2013–2014. DNA was extracted from Löwenstein–Jensen cultures and analyzed for the identification of nontuberculous mycobacteria species, detection of drug resistance with line probe assays, and high-throughput spacer oligonucleotide typing (spoligotyping) using microbead-based hybridization.ResultsTwo hundred and two cultures were positive for M. tuberculosis complex, 24 negative, 38 contaminated, and 15 positive for nontuberculous mycobacteria. Five (2.5%) M. tuberculosis complex isolates were resistant to rifampicin (RIF) and isoniazid (multidrug resistant), nine (4.5%) to RIF alone, and 15 (7.4%) to isoniazid alone; two RIF-resistant isolates were also resistant to fluoroquinolones and ethambutol, and one multidrug resistant isolate was also resistant to ethambutol. Among the 180 isolates with spoligotyping results, 164 (91.1%) were classified as lineage 4 (Euro-American), 13 (7.2%) as lineage 5 (West African 1), two (1.1%) as lineage 2 (East Asia), and one (0.6%) as lineage 6 (West African 2). One hundred and fifty-six (86.7%) isolates were grouped in 17 clusters (2–108 isolates/cluster), of which 108 (60.0%) were grouped as L4.6.2/Cameroon (spoligotype international type 61).ConclusionThe description of drug resistance prevalence and genetic diversity of M. tuberculosis in this study may be useful for improving TB control in Nigeria.

Genomic Investigation of a Mycobacterium tuberculosis Outbreak Involving Prison and Community Cases in Florida, United States.

.We used whole-genome sequencing to investigate a tuberculosis outbreak involving U.S.-born persons in the prison system and both U.S.- and foreign-born persons in the community in Florida over a 7-year period (2009–2015). Genotyping by spacer oligonucleotide typing and 24-locus mycobacterial interspersed repetitive unit-variable number tandem repeat suggested that the outbreak might be clonal in origin. However, contact tracing could not link the two populations. Through a multidisciplinary approach, we showed that the cluster involved distinct bacterial transmission networks segregated by country of birth. The source strain is of foreign origin and circulated in the local Florida community for more than 20 years before introduction into the prison system. We also identified novel transmission links involving foreign and U.S.-born cases not discovered during contact investigation. Our data highlight the potential for spread of strains originating from outside the United States into U.S. “high-risk” populations, such as prisoners, with subsequent movement back to the general community.

Spoligotyping and whole-genome sequencing analysis of lineage 1 strains of Mycobacterium tuberculosis in Da Nang, Vietnam.

BackgroundSpacer oligonucleotide typing (spoligotyping), a widely used, classical genotyping method for Mycobacterium tuberculosis complex (MTBC), is a PCR-based dot-blot hybridization technique to detect the genetic diversity of the direct repeat (DR) region. Of the seven major MTBC lineages in the world, lineage 1 (Indo-Oceanic) mostly corresponds to the East African–Indian (EAI) spoligotype family in East Africa and Southeast Asia.ObjectivesWe investigated the genomic features of Vietnamese lineage 1 strains, comparing spoligotype patterns using whole-genome sequencing (WGS) data.MethodsM. tuberculosis strains isolated in Da Nang, Vietnam were subjected to conventional spoligotyping, followed by WGS analysis using a high-throughput sequencer. Vietnamese lineage 1 strains were further analyzed with other lineage 1 strains obtained from a public database.ResultsIndicating a major spoligotype in Da Nang, 86 (46.2%) of the 186 isolates belonged to the EAI family or lineage 1. Although typical EAI4-VNM strains are characterized by the deletion of spacers 26 and 27, 65 (75.6%) showed ambiguous signals on spacer 26. De novo assembly of the entire DR region and in silico spoligotyping analysis suggested the absence of spacer 26, and direct sequencing revealed that the 17th spacer sequence not used for conventional typing, was cross-hybridized to the spacer 26 probe. Vietnamese EAI4-VNM, other EAI-like strains, and those showing a non-EAI pattern lacking many spacers formed a monophyletic group separate from other EAI families in the world.ConclusionInformation about the alignment of spacers in the entire DR region obtained from WGS data provides a clue for the determination of experimentally ambiguous spoligo patterns. WGS data also helped to analyze the hidden relationships between apparently distinct spoligo patterns.

Association between Mycobacterium tuberculosis lineage and site of disease in Florida, 2009–2015

BackgroundMycobacterium tuberculosis is characterized into four global lineages with strong geographical restriction. To date one study in the United States has investigated M. tuberculosis lineage association with tuberculosis (TB) disease presentation (extra-pulmonary versus pulmonary). We update this analysis using recent (2009–2015) data from the State of Florida to measure lineage association with pulmonary TB, the infectious form of the disease. MethodsM. tuberculosis lineage was assigned based on the spacer oligonucleotide typing (spoligotyping) patterns. TB disease site was defined as exclusively pulmonary or extra-pulmonary. We used ORs to measure the association between M. tuberculosis lineages and pulmonary compared to extra-pulmonary TB. The final multivariable model was adjusted for patient socio-demographics, HIV and diabetes status. ResultsWe analyzed 3061 cases, 83.4% were infected with a Euro-American lineage, 8.4% Indo-Oceanic and 8.2% East-Asian lineage. The majority of the cases (86.0%) were exclusively pulmonary. Compared to the Indo-Oceanic lineage, infection with a Euro-American (AOR=1.87, 95% CI: 1.21, 2.91) or an East-Asian (AOR=2.11, 95% CI: 1.27, 3.50) lineage favored pulmonary disease compared to extra-pulmonary. In a sub-analysis among pulmonary cases, strain lineage was not associated with sputum smear positive status, indicating that the observed association with pulmonary disease is independent of host contagiousness. ConclusionAs an obligate pathogen, M. tuberculosis' fitness is directly correlated to its transmission potential. In this analysis, we show that M. tuberculosis lineage is associated with pulmonary disease presentation. This association may explain the predominance in a region of certain lineages compared to others.

Evaluation of 24-locus MIRU-VNTR genotyping in Mycobacterium tuberculosis cluster investigations in four jurisdictions in the United States, 2006–2010

The U.S. Centers for Disease Control and Prevention (CDC) uses a combination of spacer oligonucleotide typing (spoligotyping) and mycobacterial interspersed repetitive units-variable number of tandem repeats (MIRU-VNTR) analyses as part of the National TB Genotyping Service (NTGS). The NTGS expansion from 12-locus MIRU-VNTR (MIRU12) to 24-locus MIRU-VNTR (MIRU24) in 2009 enhanced the ability to discriminate Mycobacterium tuberculosis strains. In the current study, we investigated the MIRU24 concordance among epidemiologic-linked tuberculosis (TB) patients in four U.S. health jurisdictions. We also evaluated the programmatic benefits of combining MIRU24 and spoligotyping with epidemiologic evidence in identifying potential recent TB transmission. We examined 342 TB patients in 42 spoligotype/MIRU12 (PCRType) clusters (equivalent to 46 spoligotype/MIRU24 [GENType] clusters) to identify epidemiologic links among cases. GENType clusters, when compared to PCRType clusters, had 12 times higher odds of epidemiologic links being identified if patients were younger than 25 years and 3 times higher odds if patients resided in the same zip code, or had HIV infection. Sixty (18%) fewer PCRType-clustered patients would need investigations if clusters are defined using GENType instead of PCRType. An important advantage of defining clusters by MIRU24 is resource savings related to the reduced number of clustered cases needing investigation.

Genotyping and rifampicin and isoniazid resistance in Mycobacterium bovis strains isolated from the lymph nodes of slaughtered cattle

In developing nations, 10–20% of the human cases of tuberculosis are caused by Mycobacterium bovis. However, this percentage may be underestimated because most laboratories in developing countries do not routinely perform mycobacterial cultures, and only a few have the systems in place to identify M. bovis. There are few studies investigating genotypic diversity and drug resistance in M. bovis from animal and/or human infections.The genotypic diversity of M. bovis strains obtained from bovine lymph nodes were investigated by spacer oligonucleotide typing (spoligotyping) and mycobacterial interspersed repetitive unit–variable-number tandem repeat typing (MIRU-VNTR). The phenotypic resistance to isoniazid and rifampicin and MIC values of the isolates were determined using the resazurin microtiter assay plate method (REMA). The evaluation of the possible genetic basis for such resistance was performed with GenoType MTBDRplus. Sixty-seven isolates were obtained, of which 11 (16%) were MDR-TB, 8 (12%) were isoniazid-resistant, and 2 (3%) were rifampicin-resistant. Mutations associated with drug resistance were not found. Genotyping techniques enabled the grouping of the strains into 12 clusters and 21 isolates with unique profiles.The high frequency of M. bovis reinforces the impact of the pathogen as a major causal agent of bovine tuberculosis in the study area. The resistance of the strains to drugs used for first-line treatment of human tuberculosis raises public health concerns. Further studies are required to elucidate the basis of drug resistance and genotypic diversity in M. bovis.

Drug resistance and population structure of M.tuberculosis isolates from prisons and communities in Ethiopia

BackgroundThe population structure and drug resistance pattern of Mycobacterium tuberculosis complex (MTBC) isolates in Ethiopian prisons and some communities is still unknown.MethodsA comparative cross sectional study was conducted on 126 MTBC strains isolated from prisons and communities in southwestern, southern and eastern Ethiopia. Phenotypic drug susceptibility testing was performed with the MGIT960 system. Combined 24-loci Mycobacterium interspersed repetitive unit-variable number tandem repeat and spacer oligonucleotide typing methods were used to study the MTBC population structure. The obtained data from prisons and communities were compared using statistical tests and regression analysis.ResultsA diverse population structure with 11 different lineages and sub-lineages was identified. The predominant strains were the recently described Ethiopia_H37Rv like (27.52%) and Ethiopia_3 (16.51%) with equal lineage distribution between prisons and communities. 28.57% of prison strains and 31.82% of community strains shared the identical genotype with at least one other strain. The multidrug-resistance (MDR) prevalence of the community was 2.27% whereas that of prisons was 9.52%. The highest mono resistance was seen against streptomycin (15.89%).ConclusionTuberculosis in communities and prisons is caused by a variety of MTBC lineages with predominance of local Ethiopian lineages. The increasing prevalence of MDR MTBC strains is alarming. These findings suggest the need for new approaches for control of MDR tuberculosis in Ethiopia.Electronic supplementary materialThe online version of this article (doi:10.1186/s12879-016-2041-x) contains supplementary material, which is available to authorized users.

Molecular Strain Typing of Mycobacterium tuberculosis: a Review of Frequently Used Methods.

Tuberculosis, caused by the bacterium Mycobacterium tuberculosis, remains one of the most serious global health problems. Molecular typing of M. tuberculosis has been used for various epidemiologic purposes as well as for clinical management. Currently, many techniques are available to type M. tuberculosis. Choosing the most appropriate technique in accordance with the existing laboratory conditions and the specific features of the geographic region is important. Insertion sequence IS6110-based restriction fragment length polymorphism (RFLP) analysis is considered the gold standard for the molecular epidemiologic investigations of tuberculosis. However, other polymerase chain reaction-based methods such as spacer oligonucleotide typing (spoligotyping), which detects 43 spacer sequence-interspersing direct repeats (DRs) in the genomic DR region; mycobacterial interspersed repetitive units–variable number tandem repeats, (MIRU-VNTR), which determines the number and size of tandem repetitive DNA sequences; repetitive-sequence-based PCR (rep-PCR), which provides high-throughput genotypic fingerprinting of multiple Mycobacterium species; and the recently developed genome-based whole genome sequencing methods demonstrate similar discriminatory power and greater convenience. This review focuses on techniques frequently used for the molecular typing of M. tuberculosis and discusses their general aspects and applications.

Genetic diversity of the Mycobacterium tuberculosis East African–Indian family in three tropical Asian countries

The Beijing lineage of Mycobacterium tuberculosis (MTB) is the most predominant MTB strain in Asian countries and is spreading worldwide, however, the East African-Indian (EAI) lineage is also particularly prevalent in many tropical Asian countries. The evolutionary relationships among MTB EAI isolates from Taiwan and those of tropical Asian countries remain unknown. The EAI strains collected from patients in Taiwan were analyzed using spacer oligonucleotide typing and mycobacterial interspersed repetitive unit-variable number of tandem repeats (MIRU-VNTR) typing, and compared with published profiles from Cambodia and Singapore to investigate potential epidemiological linkages. Among the three countries, the EAI lineage was most prevalent in Cambodia (60%; Singapore, 25.62%; and Taiwan, 21.85%), having also the highest rates of multidrug resistance and lowest rates of clustering of MTB isolates. We describe a convenient method using seven selected MIRU-VNTR loci for first-line typing to discriminate Beijing and EAI lineages. A potential epidemiological linkage in these tropical Asian countries is also discussed based on a minimum-spanning tree constructed using 24 MIRU-VNTR loci of MTB EAI strains. This study identified evolutionary relationships among MTB EAI isolates from Taiwan and those of two other tropical Asian countries, Cambodia and Singapore.

Determinants of the Sympatric Host-Pathogen Relationship in Tuberculosis.

Major contributions from pathogen genome analysis and host genetics have equated the possibility of Mycobacterium tuberculosis co-evolution with its human host leading to more stable sympatric host–pathogen relationships. However, the attribution to either sympatric or allopatric categories depends on the resolution or grain of genotypic characterization. We explored the influence on the sympatric host-pathogen relationship of clinical (HIV infection and multidrug-resistant tuberculosis [MDRTB]) and demographic (gender and age) factors in regards to the genotypic grain by using spacer oligonucleotide typing (spoligotyping) for classification of M. tuberculosis strains within the Euro-American lineage. We analyzed a total of 547 tuberculosis (TB) cases, from six year consecutive sampling in a setting with high TB-HIV coinfection (32.0%). Of these, 62.0% were caused by major circulating pathogen genotypes. The sympatric relationship was defined according to spoligotype in comparison to the international spoligotype database SpolDB4. While no significant association with Euro-American lineage was observed with any of the factors analyzed, increasing the resolution with spoligotyping evidenced a significant association of MDRTB with sympatric strains, regardless of the HIV status. Furthermore, distribution curves of the prevalence of sympatric and allopatric TB in relation to patients’ age showed an accentuation of the relevance of the age of onset in the allopatric relationship, as reflected in the trimodal distribution. On the contrary, sympatric TB was characterized by the tendency towards a typical (standard) distribution curve. Our results suggest that within the Euro-American lineage a greater degree of genotyping fine-tuning is necessary in modeling the biological processes behind the host-pathogen interplay. Furthermore, prevalence distribution of sympatric TB to age was suggestive of host genetic determinisms driven by more common variants.

Spillover of Mycobacterium bovis from wildlife to livestock, South Africa.

During August 2012–February 2013, bovine tuberculosis was detected in communal livestock bordering the Greater Kruger National Park Complex (GKNPC) in South Africa. Using spacer oligonucleotide and variable number tandem repeat typing, we identified the Mycobacterium bovis strain endemic in GKNPC wildlife. Our findings indicate bovine tuberculosis spillover from GKNPC wildlife to neighboring livestock.

Genetic Diversity and Drug Resistance of 133 Mycobacterium tuberculosis Isolates from Jiangxi Province, China

The genotypes of Mycobacterium tuberculosis (M. tuberculosis) have been found to differ in their resistance to different drugs. Although there is a high incidence of tuberculosis (TB) in Jiangxi, knowledge of the genotypes of M. tuberculosis in this province is limited in recent years. In this study, we investigated the relationship between genetic diversity and drug resistance in M. tuberculosis isolates collected from Jiangxi during January to October, 2014. A total of 133 M. tuberculosis isolates collected from the Jiangxi Chest Hospital were genotyped using both spacer oligonucleotide typing (spoligotyping) and 24-locus mycobacterial interspersed repetitive units-variable number of tandem repeats (MIRU-VNTR). The resistance of these isolates to four first-line, and four second-line, anti-TB drugs, namely, isoniazid, rifampicin, ethambutol, streptomycin, capreomycin, amikacin, levofloxacin and protionamide, was then tested. The results indicate that the Beijing family was the most prevalent genotype (75.94%), followed by the T1 family (13.53%), the MANU2 family (1.50%), the T2 family (0.75%) and the Beijing-like genotype (0.75%). We also found nine new genotypes that did not match those in the Spoldb4.0 database. The 24-locus MIRU-VNTR method had a low clustering rate (10.53%) and a high HGDI (0.9723), and proved a high resolution method for genotyping M. tuberculosis isolates. More than half of the M. tuberculosis isolates collected in the present study were resistant to anti-TB drugs (56.39%, 75/133), and the majority of these were resistant to more than one drug (81.33%, 61/75). The Beijing family is the most prevalent TB strain in Jiangxi province, China. More than half the M. tuberculosis isolates collected were resistant to anti-TB drugs, and the majority was resistant to more than one drug. There was, however, no relationship between genetic diversity and drug resistance. Moreover, our results suggest that treatment history can lead to the development of drug resistance (P < 0.05), which supports the more moderate use of medication in the treatment of TB.