Spacer Oligonucleotide Typing Research Articles

Genotyping of Pulmonary Mycobacterium tuberculosis Isolates from Sudan Using Spoligotyping.

Tuberculosis (TB) remains a major public health problem worldwide due to its high risk of person-to-person transmission, morbidity and mortality [1]. Sudan has a high burden of tuberculosis. Spoligotyping (spacer oligonucleotide typing) a rapid method for genotyping of Mycobacterium tuberculosis using the principle of reverse hybridization. The ecology of the prevalent mycobacteria strain can vary depending on country and region. The aim of this study was to determine the genotyping of Mycobacterium tuberculosis isolated from Sudan using spoligotyping SPOLDB4. A total of 75 Mycobacterium tuberculosis sputum samples were collected from pulmonary Tuberculosis patients attending references Laboratories and diagnostic centers in Khartoum and Eastern Sudan in (2011-2013). The mycobacteria were genotyped using Spoligotyping technique and data obtained were analyzed and compared to the SPOLDB4 database. Among the 75 isolate analyzed, 57(76%) were identified by SPOLDB4 and 18 (24%) could not be matched to any lineages. The most prevalent genotype cluster was MANU2 38 (50.7%) followed by CASI Delhi 8 (10.7%). In the study SIT54 was the most common pattern 37 (49.3%) followed by SIT25 6(8%).

Genetic diversity of Mycobacterium tuberculosis isolated from tuberculosis patients in the Serengeti ecosystem in Tanzania

SummaryThis study was part of a larger cross-sectional survey that was evaluating tuberculosis (TB) infection in humans, livestock and wildlife in the Serengeti ecosystem in Tanzania. The study aimed at evaluating the genetic diversity of Mycobacterium tuberculosis isolates from TB patients attending health facilities in the Serengeti ecosystem. DNA was extracted from 214 sputum cultures obtained from consecutively enrolled newly diagnosed untreated TB patients aged ≥18 years. Spacer oligonucleotide typing (spoligotyping) and Mycobacterium Interspersed Repetitive Units and Variable Number Tandem Repeat (MIRU-VNTR) were used to genotype M. tuberculosis to establish the circulating lineages. Of the214 M. tuberculosis isolates genotyped, 55 (25.7%) belonged to the Central Asian (CAS) family, 52 (24.3%) were T family (an ill-defined family), 38 (17.8%) belonged to the Latin American Mediterranean (LAM) family, 25 (11.7%) to the East-African Indian (EAI) family, 25 (11.7%) comprised of different unassigned (‘Serengeti’) strain families, while 8 (3.7%) belonged to the Beijing family. A minority group that included Haarlem, X, U and S altogether accounted for 11 (5.2%) of all genotypes. MIRU-VNTR typing produced diverse patterns within and between families indicative of unlinked transmission chains. We conclude that, in the Serengeti ecosystem only a few successful families predominate namely CAS, T, LAM and EAI families. Other types found in lower prevalence are Beijing, Haarlem, X, S and MANU. The Haarlem, EAI_Somalia, LAM3 and S/convergent and X2 subfamilies found in this study were not reported in previous studies in Tanzania.

Study of Mycobacterium bovis genotypes in human and bovine isolates using spoligotyping, MIRUVNTR and RFLP-PCR

Aims and objectivesThe aim of this study is to investigate and detect the prevalence of Mycobacterium bovis subtypes (Mycobacterium bovis subtype bovis and Mycobacterium bovis subtype caprae) in humans and compare the genetic diversity of Mycobacterium bovis in humans and cattle with spoligotyping methods, as well as pyrazinamide susceptibility study of subtypes. MethodsExamining these subtypes with molecular epidemiology techniques is particularly important due to different treatment of M. bovis diverse subtypes in humans. Culture tests were performed on clinical samples that were isolated from Lowenstein-Jensen culture medium. Identification tests were performed to differentiate Mycobacterium bovis from Mycobacterium tuberculosis. DNA was extracted and spoligotyping (spacer oligonucleotide typing) was performed using the DRb and DRa primers. ResultsThe results were analyzed with the SPOLDB4 site. PCR-RFLP method of pncA gene was used to evaluate the resistance to pyrazinamide and pncA gene polymorphism. ConclusionsMycobacterium bovis subtype bovis in the Iranian population was reported with a frequency of 0.6% which was below the average of the previous reviews. In this study, all the strains were M. bovis subtype bovis resistant to pyrazinamide. No Mycobacterium bovis subtype caprae was detected. The only shared ST between humans and cattle was ST694. Mycobacterium bovis subtype bovis with ST 595 was reported as human bovis index.

ASSOCIATION OF MYCOBACTERIUM TUBERCULOSIS LINEAGES WITH IFN-Γ AND TNF-Α GENE POLYMORPHISMS AMONG PULMONARY TUBERCULOSIS PATIENT

The six major lineages of Mycobacterium tuberculosis [MTB] are found to be strongly associated with specific geographical outbreaks. But whether these bacterial lineages influence the host genetic polymorphism is uncertain. The present study was designed to evaluate the relevance of strain diversity and host genetic polymorphisms in susceptibility to pulmonary tuberculosis [PTB]. For this reason, single –nucleotide polymorphisms [SNPs] in interferon- γ [IFN-γ] receptor-1[G-611A], IFNG [G+ 2109A] and tumor necrosis factors [TNF-α] genes [at −238, 308,−857position] in patients [n=151] were analyzed and compared with controls [n=83]. The genetic diversity of M. tuberculosis isolates was performed using spacer oligonucleotide typing. Thereafter, the profile of IFN-γ and TNF-α allele frequency were investigated in each subtype of M.tuberculosis. The results showed C allele of TNF 857 and A allele of TNF 238 were more frequent in PTB cases [[TNF 857 C allele OR [CI95%] 0.6[0.4–0.9], p= 0.02] for TNF 238 A allele OR [CI95%] 5.5[3.4–9.0], p= 0.00]]. Similarly, G allele in IFNG+ 2109 A/G polymorphism were significantly more in patients than control subject[OR[CI95%] 0.3; p< 0.05]. The major identified clinical isolates of M. tuberculosis were EAI[42; 27.8% ], Haarlem[ 31; 20.5% ], CAS [ 23;15.2% ], Beijing[14; 9.2%], and T [11; 7.2% ] lineages. No correction was observed between strains diversity and frequency of SNPs in studied PTB cases. In conclusions, we exclude the possibility of genetic mutation in IFN-γ and TNF-α gene by different subtypes of M. tuberculosis. Although, our results supports a positive correlation between host SNPs and susceptibility to PTB.

Genotypes of Mycobacterium tuberculosis isolates in rural China: Using MIRU-VNTR and spoligotyping methods

The genotypes of Mycobacterium tuberculosis (MTB) have been found to be related to the risk of transmission and the development of drug resistance of this pathogen. Thus, exploring the molecular characteristics of MTB is helpful for understanding and controlling the spread of strains in areas with a high incidence of tuberculosis. We recruited 512 sputum smear-positive tuberculosis patients from 30 counties from 1 April to 30 June 2010; 503 MTB strains were isolated and 497 were successfully genotyped. We genotyped the strains based on a new 15-locus mycobacterial interspersed repetitive unit-variable number of tandem repeats (MIRU-VNTR) method in combination with spacer-oligonucleotide typing (spoligotyping) technology. Based on spoligotyping, 487 strains displayed known patterns, and 10 were absent from the current global spoligotyping database (SpolDB4). The predominant spoligotypes belonged to the Beijing or Beijing-like family (81.1%). When we used the new 15-locus (MIRU-15) set for the MIRU-VNTR analysis, 388 different patterns were identified, including 46 clusters and 342 unique patterns. The combination of spoligotyping and MIRU-15 demonstrated a high discriminatory power. The proportion of clusters varied significantly between the Beijing and non-Beijing family strains, but no significant association was observed between multidrug resistance and Beijing family strains. The present study demonstrated that the Beijing family strains are the most prevalent in rural China. Spoligotyping in combination with the new MIRU-15 technique is useful for the epidemiological analysis of MTB transmission and could be used as a first-line method for the large-scale genotyping of MTB.

Evaluation of the reliability of thiophen-2-carboxylic acid hydrazine result for the identification of Mycobacterium bovis

To evaluate the reliability of thiophen-2-carboxylic acid hydrazine (TCH) test for identification of Mycobacterium bovis (M. bovis). A total of 4069 clinically isolated strains were identified by P-Nitrobenzoic acid medium (500 mg/L) and TCH medium (5 mg/L). Mycobacterium tuberculosis (M. tuberculosis) complex strains susceptible to 5 mg/L TCH were further tested for susceptibility to 2 mg/L TCH. Spacer oligonucleotide typing (spoligotyping) and multi-loci PCR were also performed to identify TCH susceptible strains. Among the 4069 isolated strains there were 3929 strains belonging to M. tuberculosis complex (MTBC) of which 245 were susceptible to 5 mg/L TCH. Of these 245 strains, 20 were also susceptible to 2 mg/L TCH, while only 1 strain was identified to be M. bovis by both spoligotyping and multi-loci PCR. TCH susceptibility test (either 5 mg/L or 2 mg/L) is not a reliable method for identification of M. bovis.

Molecular Discrimination ofMycobacterium bovisin São Paulo, Brazil

Mycobacterium bovis, a member of the Mycobacterium tuberculosis complex, is the most common agent of cattle tuberculosis, a zoonosis that causes losses in meat and milk production in several countries. In order to support epidemiological studies aimed at controlling the disease, several methods for molecular discrimination of M. bovis isolates have recently been developed. The most frequently used are spacer oligonucleotide typing (spoligotyping), mycobacterial interspersed repetitive units (MIRU), and exact tandem repeat (ETR), but they all have different discriminatory power. In the present study, allelic diversity was calculated for each MIRU and ETR locus, and the Hunter-Gaston discriminatory index (HGI) was calculated for spoligotyping, 10 MIRUs, and 3 ETRs, in 116 isolates of M. bovis obtained from cattle. The analysis of allelic diversity indicated that MIRUs 16, 26, and 27, and ETRs A, B, and C, showed the greatest diversity between the assayed loci. The HGIs for each of the techniques were: spoligotyping=0.738381; MIRU=0.829835; and ETR=0.825337. The associations of the methods' improved discriminatory power were: spoligotyping+MIRU=0.930585; spoligotyping+ETR=0.931034; and MIRU+ETR=0.953373. The greatest discriminatory power was obtained when the three techniques were associated (HGI=0.98051). Considering the analyses of the present study, spoligotyping should be the first method to be used because it differentiates M. bovis from the other members of the Mycobacterium tuberculosis complex. As the associations of MIRU and ETR with spoligotyping resulted in nearly identical HGIs, ETR seems to be the best choice after spoligotyping, because it is faster and more economical than MIRU. Finally, MIRU should be the last method used. In spite of this finding, the choice of the method used should be based on the discriminatory power necessary for the objective at hand.

Molecular typing and drug susceptibility of Mycobacterium tuberculosis isolates from Chongqing Municipality, China

China’s tuberculosis (TB) burden is second only to that of India worldwide. In Chongqing, the largest municipality in southwestern China, although the prevalence of both TB and drug-resistant TB is higher than in other municipalities, the molecular characteristics and drug susceptibility phenotypes are poorly known. In this study, 297 Mycobacterium tuberculosis isolates from Chongqing were genotyped with spacer oligonucleotide typing (spoligotyping) and 28-locus MIRU-VNTR (24-locus MIRU-VNTR scheme and 4 other loci). Spoligotyping results were compared with drug-resistant profiles. Patients who showed clustering by both spoligotyping and 28-locus MIRU-VNTR were interviewed to investigate their detailed contact history. Our data demonstrated that the Beijing genotype was the most prevalent genotype, and ST1 was the most predominant lineage in Chongqing. The Beijing genotype was significantly associated with ethambutol resistance and multidrug-resistant phenotypes. A combination of the 10 most polymorphic loci permitted to achieve higher discriminatory power than 24-VNTR. In addition, a presumed transmission pathway was observed in a cluster of patients with the same MIRU-VNTR profile. The 10-VNTR locus set is suitable for genotyping of Mycobacterium tuberculosis in Chongqing.

English

There is scarcity of data on molecular epidemiology of tuberculosis (TB) in many of the high burden countries of sub-Saharan Africa including Nigeria. This may be due to inadequate diagnostic and research capacities. There is a need to build capacity on molecular epidemiologic techniques in order to enhance TB diagnosis, treatment and prevention strategies. Some of the more popular molecular typing methods being used globally include IS6110-based restriction fragment length polymorphism (RFLP),polymerase chain reaction (PCR) -based methods such as spacer oligonucleotide typing or spoligotyping, mycobacterial interspersed repetitive units-multilocus variable number tandem repeats (MIRU-VNTR), single –nucleotide polymorphisms and large –sequence polymorphism analysis. Among other things, molecular technique(s) to be adopted for use in a particular setting depend on the state of the infrastructure including laboratory facilities, presence of skilled personnel and cost. In Nigeria however, availability of PCR-based diagnostic assay –MTBDR plus assay (HAIN- Lifesciences, Germany) in some TB reference laboratories may serve as a springboard for introduction of advanced TB molecular epidemiologic tools in the country. Key words: Molecular epidemiologic tools, tuberculosis, Nigeria.

Spoligotyping and drug resistance analysis of Mycobacterium tuberculosis strains from national survey in China.

BackgroundTuberculosis (TB), caused by Mycobacterium tuberculosis complex (MTBC), is one of the major causes of death in the world today. Although China has the second largest global case rate of tuberculosis, a systematic study of TB prevalence in China has not been completed. From 2006 to 2007, the base line surveillance of tuberculosis was carried out by Ministry of Health, and more than 4000 representative strains were selected from 31 provinces in China.Methodology/Principal FindingsThe aim of the present research was to survey the genotypes of representative Mycobacterium tuberculosis (M. tuberculosis) strains from China using spacer oligonucleotide typing (spoligotyping), and to analyze the relationship between genotype and drug resistance for the first time. A total of 4017 clinical isolates were collected from 2007 to 2008 throughout China. Among those M. tuberculosis isolates, 2500 (62.2%) isolates were Beijing genotypes. The percentage of Beijing genotypes in northern China was higher than in southern China (76.5% vs. 53.2%). Additionally, the frequencies of rifampin-resistant, ofloxacin-resistant and multidrug-resistant isolates were significantly higher in Beijing genotype strains than non-Beijing strains. Furthermore, a novel genotype named “China Southern genotype (CS)” was only isolated from Fujian and Guangdong provinces. Hence, it is very practical to uncover the reason for prevalence of the CS type in southern China.Conclusions/SignificanceIn conclusion, Beijing family genotypes were still the predominant genotype throughout China, which exhibited a greater correlation with rifampin-resistance, ofloxacin-resistance and MDR phenotypes than other TB spoligotypes, and some regions of China showed several unique characters in the distribution of M. tuberculosis genotypes. Our research represents an important contribution for the TB control and research community, which completes broad pictures on drug resistance levels and distribution of M. tuberculosis strain types over China.

Molecular genetic analysis of Mycobacterium tuberculosis strains spread in different patient groups in St. Petersburg, Russia

Molecular epidemiological features of Mycobacterium tuberculosis strains among different patient groups in Russia have not been studied well. The aim of our study was to compare the genotypes of M. tuberculosis strains circulating among tuberculosis (TB) patients from different groups: homeless, human immunodeficiency virus (HIV)-infected, prisoners, and the general population of St. Petersburg citizens. One hundred and forty-two M. tuberculosis complex isolates from different TB patient groups were studied using the spacer oligonucleotide typing (spoligotyping) method. The majority of the studied M. tuberculosis isolates in all groups belonged to the Beijing family (55% among homeless; 77% among HIV-infected; 60% among the general population; 81% among prisoners). There were no significant differences in the Beijing family prevalence among homeless patients, HIV/TB co-infected patients, and the general population of TB patients. The lowest genetic diversity of the pathogen was detected among imprisoned patients. The results of our study demonstrate that M. tuberculosis strains circulating among patients from high-risk groups are also spread among the general population of St. Petersburg citizens.

Quality assurance for molecular epidemiology of tuberculosis methods in the mycobacterium reference laboratory

In the National Mycobacterium Reference Laboratory of the Israeli National Public Health Laboratory (hereafter referred to as “the laboratory”), three methods are employed for the molecular epidemiology of Mycobacterium tuberculosis: IS6110 restriction fragment length polymorphism typing (RFLP typing), 43 spacer oligonucleotide typing (spoligotyping), and 24 loci Mycobacterial Interspersed Repeating Unit—Variable Number of Tandem Repeats typing (MIRU-VNTR typing). In this article, we describe the main practical aspects concerning quality assurance of these methods that are based on standardized, internationally agreed upon conditions, including consensus reference strains and markers. All three methods were validated by classical epidemiology (confirmed transmission) and clinical information. The laboratory has introduced a new 5 colors, 4 primer sets multiplex modification of the optimal 24-miru typing system that includes an easily produced in-house internal standard for the high-throughput capillary electrophoresis system. Quantitative measurement of the internal standards yielded statistics for measurement uncertainty that include the frequency distribution, mean, standard deviation, 95% confidence interval and coefficient of variation. Use of the new internal standard developed in our laboratory allowed us to introduce the first quantitative evaluation of the system performance of the AB3130xl capillary electrophoresis genetic analyzer for MIRU-VNTR typing. The results are discussed in terms of expected accuracy and precision of MIRU-VNTR results, and possible implications for forensic microsatellite typing which may be much more sensitive to the observed intra- and inter-plate variation.

Detection of Mycobacterium bovis and Mycobacterium tuberculosis from Cattle: Possible Public Health Relevance.

Mycobacterium bovis and Mycobacterium tuberculosis infect both animals and humans. The disease epidemiology by these agents differs in developed and developing countries due to the differences in the implementation of the prevention and control strategies. The present study describes the detection of M. bovis and M. tuberculosis from specimens of lungs and pulmonary lymph nodes of four cattle died in an organized herd of 183 cattle in the state of Himachal Pradesh, India, with inconclusive skin test results. Identification and distinction of these closely related mycobacterial species was done by PCR-RFLP targeting hsp65 gene followed by spacer oligonucleotide typing. Mixed infection of M. bovis and M. tuberculosis was detected in one cattle.

Genotyping of 210 Mycobacterium tuberculosis strains with Spoligotyping and MIRU-VNTR among pediatric tuberculosis patients in Chongqing

To evaluate the application of spacer oligonucleotide typing (Spoligotyping) and mycobacterial interspersed repetitive unit-variable-number tandem repeat (MIRU-VNTR) analysis in the molecular-epidemiological study of tuberculosis and to discuss the characteristics of pediatric Mycobacterium (M.) tuberculosis strains in Chongqing. M. tuberculosis strains isolated and typed by Spoligotyping and MIRU-VNTR respectively, from the children patients in Chongqing and to compare the results from both methods, epidemiologically. By means of Spoligotyping, 210 clinical isolates were divided into 2 gene groups, displaying 44 genotypes. Among them, the biggest group was M. tuberculosis Beijing family, including 130 strains (61.90%), using the Spoligotyping. From the results of MIRU-VNTR, 24 loci showed different polymorphism and the HGI of different loci set (12 old loci, 15 basic loci and 24-loci set) increased accordingly. The subtle difference in HGI was originated from one locus ETR-B, which was included in the 24-locus system. The diversity of each loci and MIRU-VNTR set for non-Beijing genotype strains was higher than that of the Beijing genotype strains. In this study, it was preliminarily confirmed the existence of high polymorphism of M. tuberculosis while the Beijing Family was the main genotype and main prevalent strain in children of Chongqing area. Spoligotyping prior to 15-locus with ETR-B combination seemed more suitable for the massive epidemiological investigation of pediatric tuberculosis patients.

Genotypes and Characteristics of Clustering and Drug Susceptibility of Mycobacterium tuberculosis Isolates Collected in Heilongjiang Province, China

For the last decade China has occupied second place, after India, among the top five countries with high burdens of tuberculosis (TB). Heilongjiang Province is located in northeastern China. The prevalence of drug-resistant TB in Heilongjiang Province is higher than the average level in China. To determine the transmission characteristics of Mycobacterium tuberculosis strains isolated in this area and their genetic relationships, especially among the Beijing family strains, we investigated their genotypes. From May 2007 to October 2008, 200 M. tuberculosis isolates from patients presenting pulmonary TB were analyzed by molecular typing using PCR-based methods: spacer-oligonucleotide typing (spoligotyping), Beijing family-specific PCR (detection of the deletion of region of difference 105 [RD105]), and mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) analysis. Different combinations of MIRU-VNTR loci were evaluated to define the genotypes and clustering characteristics of the local strains. We found that Beijing family strains represented 89.5% of the isolates studied. However, the rates of multidrug-resistant (MDR) M. tuberculosis among Beijing and non-Beijing family strains were not statistically different. The 15-locus set is considered the optimal MIRU-VNTR locus combination for analyzing the M. tuberculosis strains epidemic in this area, while the 10-locus set is an ideal set for first-line molecular typing. We found that the clustering rate of all the M. tuberculosis isolates analyzed was 10.0% using the 15-locus set typing. We conclude that the Beijing family genotype is predominant and that highly epidemic TB and MDR TB are less likely associated with the active transmission of M. tuberculosis in the study area.

Characteristics of multidrug-resistant Mycobacterium tuberculosis in Taiwan: A population-based study

A population-based study was performed to characterize the genotype and drug-resistant patterns of multidrug-resistant tuberculosis (MDR-TB) in Taiwan. From 2007 to 2008, we analyzed 494 MDR Mycobacterium tuberculosis complex isolates using spacer oligonucleotide typing and drug susceptibility testing. The majority of cases occurred in the age groups of 45–54 (24.3%) and ≥65 (23.1%). Of the 494 MDR isolates, 25.1% were resistant to ethambutol, 15.6% were resistant to streptomycin, 27.1% were resistant to all four first-line anti-tuberculosis drugs, 28.9% were resistant to ofloxacin, and 8.7% were extensively drug-resistant (XDR). Compared with the SpolDB4, 86 spoligotypes were identified in 492 isolates. We observed 427 (86.8%) isolates belonging to 49 known spoligotypes and 65 isolates (13.2%) in 37 undesignated spoligotypes. Beijing lineages (50.0%) were the predominant genotype, followed by Haarlem (18.2%) and East-African-Indian (EAI) (5.7%). Geographically, Beijing lineages were predominant in all regions, whereas Haarlem lineages were predominant only in the east (28.1%) and EAI (11.3%) only in the south. Beijing lineages are statistically associated with MDR in younger age groups and eastern Taiwan. Furthermore, we found that Beijing ST1 (46.1%), Haarlem3 ST50 (7.1%) and ST742 (4.7%), and EAI2_MANILA ST19 (3.9%) were the prevalent groups. Thus, continuous surveillance with more thorough genotyping and epidemiological investigation is crucial for the prevention of further dissemination, the determination of the temporal and spatial trends of multi-drug resistance, and the emergence of XDR-TB in Taiwan.

Mycobacterium bovis BCG-Russia Clinical Isolate with Noncanonical Spoligotyping Profile

<i>Mycobacterium bovis</i> BCG-Russia Clinical Isolate with Noncanonical Spoligotyping Profile

The Most Predominant Spoligopatterns of Mycobacterium tuberculosis Isolates among Iranian, Afghan-Immigrant, Pakistani and Turkish Tuberculosis Patients: A Comparative Analysis

Background: The aim of this study was to identify prevalent spoligopatterns of Mycobacterium tuberculosis (MTB) strains within Iran and its neighboring countries. Methods: The spacer oligonucleotide typing (spoligotyping) was performed on 1,742 MTB strains isolated from Iranian and Afghan patients. Subsequently, these results were compared with published results of Pakistan and Turkey. Results: Based on the international spoligotyping database T, Central Asian Strain (CAS), East African-Indian and Latin American and Mediterranean families were prevalent in the region. T was dominant amongIranian (32.3%) and Turkish (36.5%), whereas CAS was more prevalent among Pakistani (61.3%) and Afghan-immigrant (27.4%) TB patients. Conclusion: The majority of strains isolated from Pakistan and Afghanistan belonged to ancient MTB genotypes (principal genetic group, PGG, I). In contrast, in Turkey there was prevalence of modern TB (PGG II and III), whereas in Iran, the prevalence of ancient and modern TB was almost equal. Therefore, Iran may be considered as the connecting geographical location between ancient and modern TB.

Tracing the origins of Mycobacterium bovis tuberculosis in humans in the USA to cattle in Mexico using spoligotyping

ObjectivesTo compare genotypes of Mycobacterium bovis strains from humans in Southern California with genotypes of M. bovis strains in cattle in Mexico and the USA to explore the possible origins of human infections. MethodsWe conducted a descriptive analysis of M. bovis genotypes from a binational population of humans and cattle using spacer oligonucleotide typing (spoligotyping). ResultsOne hundred six human M. bovis spoligotypes were compared to spoligotypes from 496 Mexican cattle and 219 US cattle. Twelve spoligotype patterns were identified among human cases and 126 spoligotype patterns were detected in cattle. Over 91% (97/106) of the human M. bovis isolates had spoligotypes that were identical to those found in Mexican cattle. Four human cases had spoligotypes that matched both cattle born in Mexico and in the USA. Nine human cases had spoligotypes that did not match cattle born in Mexico or the USA. ConclusionsOur data indicate that the population of M. bovis strains causing human TB disease in Southern California is closely related to the M. bovis strain population found in Mexican cattle and supports existing epidemiological evidence that human M. bovis disease in San Diego likely originated from Mexican cattle.

A conformal Bayesian network for classification of Mycobacterium tuberculosis complex lineages

BackgroundWe present a novel conformal Bayesian network (CBN) to classify strains of Mycobacterium tuberculosis Complex (MTBC) into six major genetic lineages based on two high-throuput biomarkers: mycobacterial interspersed repetitive units (MIRU) and spacer oligonucleotide typing (spoligotyping). MTBC is the causative agent of tuberculosis (TB), which remains one of the leading causes of disease and morbidity world-wide. DNA fingerprinting methods such as MIRU and spoligotyping are key components in the control and tracking of modern TB. ResultsCBN is designed to exploit background knowledge about MTBC biomarkers. It can be trained on large historical TB databases of various subsets of MTBC biomarkers. During TB control efforts not all biomarkers may be available. So, CBN is designed to predict the major lineage of isolates genotyped by any combination of the PCR-based typing methods: spoligotyping and MIRU typing. CBN achieves high accuracy on three large MTBC collections consisting of over 34,737 isolates genotyped by different combinations of spoligotypes, 12 loci of MIRU, and 24 loci of MIRU. CBN captures distinct MIRU and spoligotype signatures associated with each lineage, explaining its excellent performance. Visualization of MIRU and spoligotype signatures yields insight into both how the model works and the genetic diversity of MTBC.ConclusionsCBN conforms to the available PCR-based biological markers and achieves high performance in identifying major lineages of MTBC. The method can be readily extended as new biomarkers are introduced for TB tracking and control. An online tool (http://www.cs.rpi.edu/~bennek/tbinsight/tblineage) makes the CBN model available for TB control and research efforts.