Molecular, epidemiological and infectivity characterisation of a Mycobacterium tuberculosis strain prevalent in Madrid (original) (raw)
AIDS, 1999
Background: As prisons have a high prevalence of tuberculosis and HIV infection, we studied the possibility of multiple tuberculosis infections in a selected population of HIV-infected inmates. Methods: Two groups of patients with special characteristics were selected from 226 HIV-infected inmates diagnosed with tuberculosis in the prisons of Madrid (Spain) between 1993 and 1994. The first group contained nine patients who remained culture positive 4 months after the initiation of therapy and the second group contained 28 patients with Mycobacterium tuberculosis isolated from different anatomical sites. DNA typing with IS6110 was performed on all isolates from these patients. In some patients a secondary DNA typing with the plasmid pTBN12 was performed. Results: Two patients from group A had a second M. tuberculosis strain obtained 4 and 18 months after the initial isolate, with different IS6110 and pTBN12 patterns. In one patient the second strain was multidrug resistant and in the other patient both strains had the same drug-susceptible pattern. The clinical and microbiologic evidence in both patients was consistent with the presence of active tuberculosis caused by a new strain of M. tuberculosis. In group B, the isolates from 27 patients shared similar fingerprint pattern; however, in one patient isolates from sputum and urine showed different IS6110 and pTBN12 patterns, although both strains showed the same drug-susceptible phenotype. Conclusion: This study provides evidence that HIV-infected inmates living under conditions of high environmental infectivity can be infected with two different strains of M. tuberculosis. This finding has implications for the tuberculosis-control programs in prison.
Journal of Clinical Microbiology, 2011
We ran a comparative analysis of all patients for whom a positive culture of Mycobacterium tuberculosis complex was available between April 2004 and October 2005 and whose HIV serology results were known, with spoligotyping results (n ؍ 163) split into 49 HIV-positive patients and 114 HIV-negative patients. Spoligotype international type 373 (SIT373) (T1 lineage), which was highly prevalent among the HIV ؉ patients, was totally absent from the HIV ؊ population, suggesting that we had a specific clone affecting nearly 1/3 of all HIVtuberculosis (TB)-coinfected patients. Among the LAM10-CAM sublineage strains, we had only a single strain of SIT403 among HIV ؊ patients (0.88%), as opposed to 12.25% of the HIV ؉ population ( 2 ؍ 10.77; P < 0.01), indicating a strong association between the strain and the HIV ؉ population. The LAM10-CAM lineage spoligotype SIT61 was prevalent among the 2 subsets (37.72% in HIV ؊ versus 12.24% in HIV ؉ populations), though, with a significant difference between the 2 groups ( 2 ؍ 10.53; P < 0.01). However, there was no significant difference for SIT53 (T1 lineage) in the 2 subsets: 6.14 versus 8.2% ( 2 ؍ 0.22; P > 0.05). A total of 7/49, or 14.3%, other SITs among HIV ؉ patients were not found among the HIV ؊ patients. When added to the most prevalent SIT among HIV ؉ patients (SIT373; n ؍ 16), 23/49, or 47%, isolates among HIV-TBcoinfected patients were unique. We conclude that further studies should be carried out to investigate the evolution of these genotypes and others in the emergence of multidrug resistance and control of tuberculosis in Nigeria.
Clinical Infectious Diseases, 1996
Management of mycobacterial infection is species specific; however, treatment is prompted by positive smears or cultures, often several weeks before species identification. The objective of this study was to determine the species distribution of mycobacterial isolates from various body sites in patients infected with human immunodeficiency virus (HIV). All mycobacterial isolates recovered at St. Paul's Hospital (Vancouver, British Columbia, Canada) from April 1989 to March 1993 were reviewed. Among 357 HIV-positive patients with mycobacterial infections, 64% (96) of the sputum isolates were Mycobacterium avium complex (MAC), 18% were Mycobacterium tuberculosis, and 17% were Mycobacterium kansasii. Lymph node involvement (25 patients) was due to either MAC (72%) or M. tuberculosis (24%). Two hundred ninety-eight episodes of mycobacteremia were due to MAC (98%), M. tuberculosis (1%), and M. kansasii (1%). Similarly, cultures of 84 bone marrow biopsy specimens (99%), 19 intestinal biopsy specimens (100%), and 30 stool specimens (97%) yielded predominantly MAC. These results have implications for initial therapy, particularly in areas where rapid methods for species identification are not readily available. Because of considerable geographic variation, development of guidelines for selection of initial therapy depends on regional determination of species distribution in HIV-related mycobacterial infections. Mycobacterial infections are common in individuals infected with HIV. Three species, Mycobacterium avium complex (MAC), Mycobacterium tuberculosis, and Mycobacterium kansasii, account for almost all mycobacterial disease in patients with AIDS . Knowledge of the likelihood of various species being responsible for disease is important for initial drug selection [1], infection control practices [2], and prognosis . When conventional techniques are used, 4 to 8 weeks may lapse from the time of preliminary demonstration of acid-fast bacilli by smear examination or culture to species identification. Nucleic acid probes for hybridization have been successfully employed for rapid identification of species [5]; however, in many countries, they may not be widely available. Because of the lesser importance and frequency of nontuberculous mycobacteria infections before the AIDS epidemic, the practice of many clinicians has been to initiate therapy for M tuberculosis infection on the basis of positive smears or cultures of specimens from any site, pending species identification. We studied the relative frequency of different mycobacterial species infecting HIV-positive patients at St. Paul's Hospital (Vancouver,
PLoS ONE, 2014
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 fbpC 103 , RD Rio 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 fbpC 103 . The frequency of RD Rio 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/RD Rio 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.
Isolation, identification and DNA fingerprinting of Mycobacterial Isolates from AIDS patients
International Journal of Molecular and Clinical Microbiology, 2011
Tuberculosis (TB) is one of the most important AIDS associated infectious diseases worldwide. It is a leading cause of illness and death among people with HIV/AIDS in resource-poor areas of the world. The annual incidence of TB among indigenous Iranians stands at 14 cases per 100,000 inhabitants. This study aimed to identify Mycobacterium infection among Iranian HIV positive patients. Two sputum specimens were collected from smear positive AIDS patients. Samples were cultured on Lowenstein-Jensen media for three weeks. DNA was extracted from two samples based on van Embden protocol. To identify Mycobacterium tuberculosis complex, a PCR was conducted to amplify a 245 bp fragment of IS6110 element, followed by RD12 method to confirm PCR result. Whole DNA-RFLP with PvuII restriction enzyme was employed to genotype the cultured isolates. The results obtained by colonial morphology, PCR, and RD12 methods showed that both isolates were belonging to M. tuberculosis, and Genotyping of the isolates by RFLP technique displayed that two isolates were belonging to different strains of M. tuberculosis.
Infection Genetics and Evolution
One of the high tuberculosis (TB) incidence countries in the world, Brazil is characterized by considerable differences in TB incidence on regional and state level. In the present study, we describe Brazilian spoligotypes of 1991 Mycobacterium tuberculosis complex (MTC) clinical isolates from patients residents of 11 states from different regions of the country, diagnosed between 1996 and 2005. By performing spoligotyping on a large number of M. tuberculosis clinical isolates, one of the main objectives of this study was to determine the major genotype families causing TB in Brazil and to verify the region-associated genotype distribution. We observed a total of 577 distinct spoligopatterns, 12.6% of these corresponded to orphan patterns while 87.4% belonged to 326 shared-types (SITs). Among the latter, 86 SITs (isolated from 178 patients) had been observed for the first time in this study, the most frequent being SIT2517 which belonged to the T3-ETH lineage and was exclusively found among patients residents of Belém, the capital of the state of Pará (n = 8 isolates). Irrespective of shared-type labeling, a total of 19.5% strains were unique (unclustered) in our study as opposed to 80.5% clustered isolates (189 clusters, size range from 2 to 205 isolates). The three largest clusters were SIT42 of the Latin-America & Mediterranean (LAM) 9 clade (10.3%), SIT53 of the T clade (7.6%), and SIT50 of the Haarlem clade (5.4%). The predominant MTC lineages in Brazil in decreasing order belonged to the LAM (46%); the ill-defined T (18.6%); the Haarlem (12.2%), the X (4.7%), the S (1.9%), and the East African Indian (EAI) (0.85%) families. The rest of clades grouped together as Mycobacterium africanum, Mycobacterium bovis, Beijing, Central Asian (CAS), and the Manu types, represented less than 1% of the strains. Finally, about 15% of the isolates showed spoligotype signatures that were not yet classified among well-defined lineages. In conclusion, we provide hereby a first insight into the population structure of MTC isolates in Brazil, showing the predominance of both LAM and T family and the existence of region-associated genotypes.
2021
Tuberculosis (TB) remains an important public health problem in developing countries like the Philippines. The success of the National TB Control Program depends on a clear understanding of the dynamics of transmission and spread of TB in high-risk populations in the community. We conducted a molecular epidemiologic analysis of M. tuberculosis isolates collected from inmates with pulmonary TB in selected prisons in the Philippines. A total of 25 isolates were characterized and genotyped using Spoligotyping and 15-loci MIRU-VNTR (mycobacterial interspersed repetitive units-variable number of tandem repeats) typing. The majority of the patients were male (84%) and aged 30-49 yr old (68%). Eighteen (72%) of the culture-positive patients had severe pulmonary TB, 13 (52%) were smear-positive, and seven (28%) were classified as having a high bacillary load. Twenty isolates (80%) were susceptible to all the first-line drugs. Two (8%) were multidrugresistant (MDR) and isolated from patients in the same prison, one of which was resistant to all first-line drugs. Three isolates (12%) were streptomycin-monoresistant. There were nine identified Spoligo-International Types (SITs), with SIT19 as the predominant (40%). One isolate (4%) did not match any SIT in the SpoIDB4 database, while three were not assessed due to inadequate DNA for analysis. The distribution of strains according to major M. tuberculosis clades were as follows: EAI2_Manilla (48%) > LAM2 (20%) > LAM6 (8%) = LAM9 (8%). Spoligotyping identified two clusters and 13 genotypes (four unique strains) with a Hunter-Gaston discriminatory index (HGDI) of 0.83. MIRU-VNTR typing identified two clusters and 23 genotypes (HGDI = 0.993). Combined Spoligotyping and MIRU-VNTR typing also identified two clusters and 23 genotypes (HGDI = 0.993). There were no significant associations shown among host demographic factors, severity of the disease, drug resistance, and M. tuberculosis strain. We conclude that our patient population was infected predominantly by M. tuberculosis belonging to the EAI2_Manila clade.
Molecular Strain Typing ofMycobacterium tuberculosis: a Review of Frequently Used Methods
Journal of Korean Medical Science, 2016
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 reactionbased 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-sequencebased 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.
Journal of Clinical Tuberculosis and Other Mycobacterial Diseases, 2018
Background: Tuberculosis is a serious infection that is common in people living with HIV and increases the mortality and morbidity from the diseases. The study of genetic diversity among strains of M. tuberculosis has a great impact in studying pathogenicity and transmissibility, design for vaccines production, identification of nominee genes for drug targets, and improving molecular diagnostic techniques. The aim of this study was to characterize Mycobacterium tuberculosis (Mtb) isolated from suspected pulmonary tuberculosis among people living with HIV. Method: A total of 143 sputum samples was collected and transported to Akililu Lemma TB laboratory. The collected samples were processed for culture using Lowenstein-Jensen medium. For 45 culture positive isolates, genotyping of mycobacterial DNA was performed by spoligotyping and isolates were assigned to families using the SpolDB4 and the model-based program 'SPOTCLUST'. Categorical data were analyzed by Chi-square test. Result: A high level of diversity was found among the 45 isolates. Twenty six different Spoligo patterns were obtained. The T (46.7%), Family33 (44.4%) and Central Asian (CAS): (4.4%) families were the dominant isolates comprising 91.5% of the total strains. Of 44% of the Euro-American, 6/20(30%) and 9/20(45%), identified were lineage belonged to Spoligo-International-Type (SIT 336 ) and SIT 149 . Of the total strains, 12 (22%) were unique and have not been described in SpolDB4 to date. Conclusion: We found the high diversity of Mtb in pulmonary tuberculosis patients in this setting. T 3_ ETH family identified as the numerous M.tuberculosis strains circulating in the community.
Scientific Reports
structure but included the maximum likelihood-inferred ancestral nucleotide positions from a virtual ancestor 11 , by using the Burrows-Wheeler Aligner. SNP calls were made with SAMtools and VarScan (coverage of at least 20×). We kept only the homozygous calls (present in at least 90% of the reads in a specific position) and filtered out potential calling errors by omitting variants detected in repetitive regions, phages, PE/PPE regions, and SNPs close to indels (10 bp window) or in areas with an anomalous accumulation of variants (three or more SNPs in 10 bp). Alignments and SNP variants were visualized and checked with the IGV program. Multiple comparisons between the SNPs from the clustered isolates were made using an in-house script written in R software (R Foundation for Statistical Computing, Vienna, Austria 2011, www.R-project.org). The median-joining networks were constructed from the SNP matrix generated using NETWORK 5.0.0.1. Median vectors (mv) were defined when the distribution of SNPs indicated the existence of a non-sequenced node corresponding to non-sampled genotype in the cluster. The chronology of acquisition of the SNPs is represented from left to right in the networks.