Decrypting the environmental sources of Mycobacterium canettii by high-throughput biochemical profiling.

Mycobacterium canettii is a smooth bacillus related to the Mycobacterium tuberculosis complex. It causes lymph nodes and pulmonary tuberculosis in patients living in countries of the Horn of Africa, including Djibouti. The environmental reservoirs of M. canettii are still unknown. We aimed to further decrypt these potential reservoirs by using an original approach of High-Throughput Carbon and Azote Substrate Profiling. The Biolog Phenotype profiling was performed on six clinical strains of M. canettii and one M. tuberculosis strain was used as a positive control. The experiments were duplicated and authenticated by negative controls. While M. tuberculosis metabolized 22/190 (11%) carbon substrates and 3/95 (3%) nitrogen substrates, 17/190 (8.9%) carbon substrates and three nitrogen substrates were metabolized by the six M. canettii strains forming the so-called corebiologome. A total at 16 carbon substrates and three nitrogen substrates were metabolized in common by M. tuberculosis and the six M. canettii strains. Moreover, at least one M. canettii strain metabolized 36/190 (19%) carbon substrates and 3/95 (3%) nitrogen substrates for a total of 39/285 (13%) substrates. Classifying these carbon and nitrogen substrates into ten potential environmental sources (plants, fruits and vegetables, bacteria, algae, fungi, nematodes, mollusks, mammals, insects and inanimate environment) significantly associated carbon and nitrogen substrates metabolized by at least one M. canettii strain with plants (p = 0.006). These results suggest that some plants endemic in the Horn of Africa may serve as ecological niches for M. canettii. Further ethnobotanical studies will indicate plant usages by local populations, then guiding field microbiological investigations in order to prove the definite environmental reservoirs of this opportunistic tuberculous pathogen.

Linezolid resistance in patients with drug-resistant TB and treatment failure in South Africa.

OBJECTIVES: Limited data exist on clinical associations and genotypic correlates of linezolid resistance in Mycobacterium tuberculosis. We aimed to describe mutations and clinical factors associated with phenotypic linezolid resistance from patients with drug-resistant TB at two public sector facilities in South Africa. METHODS: Adults and adolescents with treatment failure (culture positivity ≥4 months) on a linezolid-containing regimen were retrospectively identified. Phenotypic resistance, as defined by a linezolid MIC >1 mg/L, was assessed for retrieved isolates using broth microdilution. Targeted sequencing of rrl and rplC was performed, irrespective of growth on subculture. RESULTS: Thirty-nine patients with linezolid-based treatment failure were identified, 13 (33%) of whom had phenotypic or genotypic linezolid resistance after a median duration of 22 months (range = 7-32) of linezolid therapy. Paired MIC testing and genotyping was performed on 55 unique isolates. All isolates with phenotypic resistance (n = 16) were associated with known resistance mutations, most frequently due to the T460C substitution in rplC (n = 10); rrl mutations included G2814T, G2270C/T and A2810C. No mutations were detected in isolates with MICs at or below the critical concentration. CONCLUSIONS: Linezolid resistance occurred in a third of patients with drug-resistant TB and treatment failure. Resistance occurred late and was predicted by a limited number of mutations in rrl and rplC. Screening for genotypic resistance should be considered for patients with a positive culture after 4 months of linezolid therapy in order to optimize treatment and avoid the toxicity of ineffective linezolid therapy.

A new oligonucleotide array for the detection of multidrug and extensively drug-resistance tuberculosis.

Drug-resistant tuberculosis (TB) is a global crisis and a threat to health security. Since conventional drug susceptibility testing (DST) takes several weeks, we herein described a molecular assay to rapidly identify multidrug-resistant (MDR) and extensively drug-resistant (XDR) and reveal transmission associated-mutations of Mycobacterium tuberculosis complex (MTBC) isolates in 6 to 7 hours. An array was designed with 12 pairs of primers and 60 single nucleotide polymorphisms of 9 genes: rpoB, katG, inhA, ahpC, embB, rpsL, gyrA, rrs and eis. We assessed the performance of the array using 176 clinical MTBC isolates. The results of culture-based DST were used as the gold standard, the GenoType MTBDRplus and MTBDRsl tests were used for parallel comparison, and gene sequencing was performed to resolve the discordance. The sensitivities and specificities of the array are comparable to those of the MTBDRplus test for resistance to isoniazid (INH) (100.0%, 96.7%) and rifampicin (RIF) (99.4%, 96.7%) and of the MTBDRsl test for resistance to fluoroquinolones (FQs) (100%, 100%) and second-line injectable drugs (SLIDs) (98.3%, 100%). The sensitivities of the array for detecting resistance to ethambutol and streptomycin were 79.3% and 64.9%, respectively. The array has potential as a powerful tool for clinical diagnosis and epidemiological investigations.

Molecular Targets Related Drug Resistance Mechanisms in MDR-, XDR-, and TDR-Mycobacterium tuberculosis Strains.

Tuberculosis (TB) is a formidable infectious disease that remains a major cause of death worldwide today. Escalating application of genomic techniques has expedited the identification of increasing number of mutations associated with drug resistance in Mycobacterium tuberculosis. Unfortunately the prevalence of bacillary resistance becomes alarming in many parts of the world, with the daunting scenarios of multidrug-resistant tuberculosis (MDR-TB), extensively drug-resistant tuberculosis (XDR-TB) and total drug-resistant tuberculosis (TDR-TB), due to number of resistance pathways, alongside some apparently obscure ones. Recent advances in the understanding of the molecular/ genetic basis of drug targets and drug resistance mechanisms have been steadily made. Intriguing findings through whole genome sequencing and other molecular approaches facilitate the further understanding of biology and pathology of M. tuberculosis for the development of new therapeutics to meet the immense challenge of global health.

Extended spectrum of antibiotic susceptibility for tuberculosis, Djibouti.

In the Horn of Africa, there is a high prevalence of tuberculosis that is reported to be partly driven by multidrug-resistant (MDR) Mycobacterium tuberculosis strictu sensu strains. We conducted a prospective study to investigate M. tuberculosis complex species causing tuberculosis in Djibouti, and their in vitro susceptibility to standard anti-tuberculous antibiotics in addition to clofazimine, minocycline, chloramphenicol and sulfadiazine. Among the 118 mycobacteria isolates from 118 successive patients with suspected pulmonary tuberculosis, 111 strains of M. tuberculosis, five Mycobacterium canettii, one 'Mycobacterium simulans' and one Mycobacterium kansasii were identified. Drug-susceptibility tests performed on the first 78 isolates yielded nine MDR M. tuberculosis isolates. All isolates were fully susceptible to clofazimine, minocycline and chloramphenicol, and 75 of 78 isolates were susceptible to sulfadiazine. In the Horn of Africa, patients with confirmed pulmonary tuberculosis caused by an in vitro susceptible strain may benefit from anti-leprosy drugs, sulfamides and phenicol antibiotics.

Machine learning for classifying tuberculosis drug-resistance from DNA sequencing data.

Motivation: Correct and rapid determination of Mycobacterium tuberculosis (MTB) resistance against available tuberculosis (TB) drugs is essential for the control and management of TB. Conventional molecular diagnostic test assumes that the presence of any well-studied single nucleotide polymorphisms is sufficient to cause resistance, which yields low sensitivity for resistance classification. Summary: Given the availability of DNA sequencing data from MTB, we developed machine learning models for a cohort of 1839 UK bacterial isolates to classify MTB resistance against eight anti-TB drugs (isoniazid, rifampicin, ethambutol, pyrazinamide, ciprofloxacin, moxifloxacin, ofloxacin, streptomycin) and to classify multi-drug resistance. Results: Compared to previous rules-based approach, the sensitivities from the best-performing models increased by 2-4% for isoniazid, rifampicin and ethambutol to 97% (P < 0.01), respectively; for ciprofloxacin and multi-drug resistant TB, they increased to 96%. For moxifloxacin and ofloxacin, sensitivities increased by 12 and 15% from 83 and 81% based on existing known resistance alleles to 95% and 96% (P < 0.01), respectively. Particularly, our models improved sensitivities compared to the previous rules-based approach by 15 and 24% to 84 and 87% for pyrazinamide and streptomycin (P < 0.01), respectively. The best-performing models increase the area-under-the-ROC curve by 10% for pyrazinamide and streptomycin (P < 0.01), and 4-8% for other drugs (P < 0.01). Availability and implementation: The details of source code are provided at http://www.robots.ox.ac.uk/~davidc/code.php. Contact: david.clifton@eng.ox.ac.uk. Supplementary information: Supplementary data are available at Bioinformatics online.

Assessment of Bactericidal Drug Activity and Treatment Outcome in a Mouse Tuberculosis Model Using a Clinical Beijing Strain.

Mycobacterium tuberculosis Beijing strains are associated with lower treatment success rates in tuberculosis (TB) patients. In contrast, laboratory strains such as H37Rv are often used in preclinical tuberculosis models. Therefore, we explored the impact of using a clinical Beijing strain on treatment outcome in our mouse tuberculosis model. Additionally, the predictive value of bactericidal activity on treatment outcome was assessed. BALB/c mice were infected with a Beijing strain and treated with one of 10 different combinations of conventional anti-TB drugs. Bactericidal activity was assessed by determining reductions in mycobacterial load after 7, 14, and 28 days and after 2, 3, and 6 months of treatment. Treatment outcome was evaluated after a 6-month treatment course and was based on lung culture status 3 months posttreatment. None of the anti-TB drug regimens tested could achieve 100% treatment success. Treatment outcome depended critically on rifampin. Four non-rifampin-containing regimens showed 0% treatment success compared to success rates between 81 and 95% for six rifampin-containing regimens. Bactericidal activity was predictive only for treatment outcome after 3 months of treatment. Our data advocate the use of multiple mycobacterial strains, including a Beijing strain, to increase the translational value of mouse TB models evaluating treatment outcome. Additionally, our findings support the notion that bactericidal activity in the first 2 months of treatment, as measured in clinical phase IIa/b trials, has limited predictive value for tuberculosis treatment outcome, thus emphasizing the need for better parameters to guide future phase IIII trials.

First Insight Into the Fluoroquinolone and Aminoglycoside Resistance of Multidrug-Resistant Mycobacterium tuberculosis in Saudi Arabia.

AbstractIn Saudi Arabia, there were no nationwide screening studies conducted so far to determine the aminoglycoside and fluoroquinolone resistance among multidrug-resistant tuberculosis (MDR-TB) isolates. Therefore, as the first attempt in the country, a retrospective analysis has been conducted on a nationwide collection of 2,956 M. tuberculosis clinical isolates screened with phenotypic drug susceptibility testing to define MDR-TB. Enrolled MDR-TB isolates were subjected to second-line drug susceptibility testing, detection of mutations conferring resistance to aminoglycosides and fluoroquinolone, followed by 24-loci mycobacterial interspersed repetitive unit-variable number of tandem repeat typing and spoligotyping. Overall, 83 isolates were identified as MDR-TB, and 13 (15.7%) isolates showed resistance to second-line drugs. Moxifloxacin (low level) showed higher resistant rates (10.8%) followed by ofloxacin (7.2%), capreomycin (3.6%), kanamycin (3.6%), and amikacin (2.4%). Overall fluoroquinolone resistance was 12%, whereas aminoglycoside resistance was 7.2%. Predominant mutations conferring resistance to fluoroquinolone were found in gyrA A90V and D94G, whereas aminoglycoside resistance was observed only with rrs gene A1401G mutation. The corresponding strain lineages predominated with Indo-Oceanic and East-African Indian origin. Interestingly, none of the isolates with second-line drug resistance was defined as extensively drug-resistant TB (XDR-TB). Surprisingly, many isolates (50.6%) were panresistant to first-line drugs. Saudi Arabia faces considerable burden of fluoroquinolone- and aminoglycoside-resistant MDR-TB. Higher incidence of panresistant MDR-TB reveals a threat for the emergence of XDR-TB strains in the near future.

DRUG RESISTANCE PATTERN AND MOLECULAR CHARACTERIZATION OF MYCOBACTERIUM TUBERCULOSIS STRAINS IN PUNJAB, PAKISTAN.

Tuberculosis (TB) is a cause of death from a single infectious agent Mycobacterium tuberculosis (MTB), leading to approximately 2.5 million deaths annually worldwide. Information regarding prevalence and pattern of drug resistance among TB patients in Pakistan remains inadequate due to the country's limited resources. This study compared conventional diagnostic techniques with a PCRbased assay targeting IS6110 sequence. In addition, MTB drug resistant profiles against four first-line drugs (ethambutol, isoniazid, rifampin, and streptomycin) from new and retreatment cases of TB. From 101 sputum samples microscopic examination of Ziehl-Neelsen-stained smears and culturing on Lowenstein Jensen medium resulted in 96% and 100% positives, compared to 98% by PCR. Prevalence of MDR-MTB was 41.5% and 58.5% among new (n = 51) and retreatment (n = 50) cases, but 10% of the former group were sensitive to all four first-line anti-TB drugs. Thus, MDR-MTB is highly prevalent among TB patients in Punjab Province, Pakistan (where the study was conducted) and, although PCR amplification of MTB-specific IS6110 sequence was rapid, it lacked the sensitivity of the culture assay.

Transmission of multi-drug resistant tuberculosis in Mongolia is driven by Beijing strains of Mycobacterium tuberculosis resistant to all first-line drugs.

BACKGROUND: Mongolia has high and rising rates of multi-drug resistant tuberculosis (MDR-TB). Spatio-temporal and programmatic evidence suggests a major contribution from MDR-TB transmission, but genotypic evidence has not been assessed. METHODS: All MDR-TB cases identified during 2012 were examined. Demographic and bacteriological data were obtained from the National Tuberculosis Reference Laboratory. Isolates of Mycobacterium tuberculosis from culture-confirmed category 1 treatment failures were genotyped using 24-loci mycobacterium interspersed repetitive unit (MIRU-24) analysis. RESULTS: Of the 210 MDR-TB cases identified, 115 (54.8%) were treatment failures (34.8% category 1; 20.0% category 2). Streptomycin resistance was present in 156 (74.3%) cases; including 55/73 (75.3%) category 1 treatment failures who had never been exposed to streptomycin. Among category 1 treatment failures, Beijing lineage strains predominated (88.0%; 59/67 of genotyped isolates). MIRU-24 clustering was documented in 62.7% (42/67) of strains; 55.2% (37/67) remained clustered when drug susceptibility test results were considered. In total 59.5% (25/42) of clustered strains were Beijing lineage and demonstrated in-vitro resistance to all first-line drugs tested. CONCLUSION: The MDR-TB epidemic in Mongolia appears to be driven by primary transmission of Beijing lineage strains resistant to all first-line drugs. Enhanced infection control strategies together with early MDR-TB case detection and appropriate treatment are necessary to limit escalation of the MDR-TB epidemic.

Turkish and Japanese Mycobacterium tuberculosis sublineages share a remote common ancestor.

Two geographically distant M. tuberculosis sublineages, Tur from Turkey and T3-Osaka from Japan, exhibit partially identical genotypic signatures (identical 12-loci MIRU-VNTR profiles, distinct spoligotyping patterns). We investigated T3-Osaka and Tur sublineages characteristics and potential genetic relatedness, first using MIRU-VNTR locus analysis on 21 and 25 samples of each sublineage respectively, and second comparing Whole Genome Sequences of 8 new samples to public data from 45 samples uncovering human tuberculosis diversity. We then tried to date their Most Recent Common Ancestor (MRCA) using three calibrations of SNP accumulation rate (long-term=0.03SNP/genome/year, derived from a tuberculosis ancestor of around 70,000years old; intermediate=0.2SNP/genome/year derived from a Peruvian mummy; short-term=0.5SNP/genome/year). To disentangle between these scenarios, we confronted the corresponding divergence times with major human history events and knowledge on human genetic divergence. We identified relatively high intrasublineage diversity for both T3-Osaka and Tur. We definitively proved their monophyly; the corresponding super-sublineage (referred to as "T3-Osa-Tur") shares a common ancestor with T3-Ethiopia and Ural sublineages but is only remotely related to other Euro-American sublineages such as X, LAM, Haarlem and S. The evolutionary scenario based on long-term evolution rate being valid until T3-Osa-Tur MRCA was not supported by Japanese fossil data. The evolutionary scenario relying on short-term evolution rate since T3-Osa-Tur MRCA was contradicted by human history and potential traces of past epidemics. T3-Osaka and Tur sublineages were found likely to have diverged between 800y and 2000years ago, potentially at the time of Mongol Empire. Altogether, this study definitively proves a strong genetic link between Turkish and Japanese tuberculosis. It provides a first hypothesis for calibrating TB Euro-American lineage molecular clock; additional studies are needed to reliably date events corresponding to intermediate depths in tuberculosis phylogeny.

Paleomicrobiology of Human Tuberculosis.

Tuberculosis is a significant global disease today, so understanding its origins and history is important. It is primarily a lung infection and is transmitted by infectious aerosols from person to person, so a high population density encourages its spread. The causative organism is Mycobacterium tuberculosis, an obligate pathogen in the M. tuberculosis complex that also contains closely related species, such as Mycobacterium bovis, that primarily infect animals. Typical bone lesions occur in about 5% of untreated infections. These can be recognized in historical and archaeological material, along with nonspecific paleopathology such as new bone formation (periostitis), especially on ribs. Based on such lesions, tuberculosis has been found in ancient Egypt, pre-Columbian America, and Neolithic Europe. The detection of M. tuberculosis ancient DNA (aDNA) by using PCR led to the development of the new field of paleomicrobiology. As a result, a large number of tuberculosis cases were recognized in mummified tissue and bones with nonspecific or no lesions. In parallel with these developments, M. tuberculosis cell wall lipid biomarkers have detected tuberculosis suggested by paleopathology and confirmed aDNA findings. In well-preserved cases, molecular typing has identified M. tuberculosis lineages and genotypes. The current interest in targeted enrichment, shotgun sequencing, and metagenomic analysis reveals ancient mixed infections with different M. tuberculosis strains and other pathogens. Identification of M. tuberculosis lineages from samples of known age enables the date of the emergence of strains and lineages to be calculated directly rather than by making assumptions on the rate of evolutionary change.

Personalizing therapy for multidrug resistant TB: the potential of Rapid Whole Genome Sequencing.

Multidrug resistant tuberculosis is an increasing problem globally. The current gold standard in drug sensitivity testing is slow and cumbersome. To tackle drug resistance effectively, a more rapid method of testing is required. Current molecular tests are fast, but only offer information on a limited number of genetic loci. Whole genome sequencing presents an attractive alternative that can provide comprehensive, clinically relevant information on all described loci. Although the standard approach to whole genome sequencing of Mycobacterium tuberculosis is slow due to the requirement of culture, this article will describe recent advances that mean it has the potential to provide results within days.

Molecular typing of drug-susceptible and -resistant Mycobacterium tuberculosis in Johannesburg, South Africa.

SETTING: Knowledge about spoligotyping families of drug-susceptible and drug-resistant Mycobacterium tuberculosis circulating in the Johannesburg area, South Africa, is lacking. OBJECTIVE: To determine the genetic diversity of M. tuberculosis isolates circulating in the Johannesburg area and to compare the results with both national and international databanks. DESIGN: Five hundred cultured M. tuberculosis isolates from within the greater Johannesburg metropolitan area collected from January 2009 to December 2010 were obtained from the National Health Laboratory Services (NHLS) Mycobacteriology Referral Laboratory, Johannesburg, in MGIT vials. The isolates were specimens from individuals with tuberculosis (TB) symptoms and known TB patients submitted to the NHLS for routine mycobacterial culture and drug susceptibility testing. The isolates were genotyped using spoligotyping. RESULTS: Spoligotyping generated 62 shared types, with 92% (458/500) of the sample size matching pre-existing shared types. Of the 62 shared types, eight families were predominant (clustering from 16 to 132), representing 64% (340/500) of the sample. The Beijing family (135/500) predominated (P < 0.001). CONCLUSION: TB incidence in Johannesburg does not appear to be due to clonality, but is rather due to diverse circulating strains, namely the Beijing family, followed by the S, Latin American-Mediterranean and T families.

Outbreak of extrapulmonary tuberculosis infection associated with acupuncture point injection.

Mycobacterium tuberculosis infection is rarely reported to be associated with acupuncture practices. We performed a retrospective outbreak investigation of a unique outbreak of 33 extrapulmonary M. tuberculosis infections related to acupuncture point injection therapy (AIT) among clients who visited a private traditional Chinese medicine clinical centre in China. The lumps, abscesses and ulcers occurred mostly on the neck, shoulders, waist, knees and hips, localized at acupuncture point meridian sites. These symptoms appeared from January to November 2011, with a peak cluster of infections in September 2011 (nine cases). M. tuberculosis Beijing strain was isolated and confirmed by DNA sequencing. All diagnosed patients were treated empirically with appropriate antibiotic treatment, and their condition improved. Our study indicated that this outbreak was most likely resulted from contaminated AIT. Drafting standard guidelines for AIT is urgently needed, and routine medical supervision should be provided, including obligating health providers to perform routine physical examinations that include testing for infectious diseases.

Avaliação da atividade antimicobacteriana da lignana diidrocubebina extraída da Piper cubeba e de seus derivados semissintéticos / Evaluation of antimycobacterial activity of dihydrocubebin lignan extracted from Piper cubeba and its semisynthetic derivatives

RESUMO: A atividade antimicobacteriana de diidrocubebina (1), uma lignana dibenzilbutanodioica obtida a partir de extrato etanólico de sementes da Piper cubeba, e seus derivados foram avaliados in vitro contra três diferentes cepas de Mycobacterium utilizando o método de microdiluição. Dentre as lignanas avaliadas 3 e 4 foram as mais ativas, exibindo valores de CIM de 62,5 µg/mL contra M. avium e M. tuberculosis, respectivamente. Os derivados 2-6 obtidos por síntese parcial possuem diferentes substituintes nos carbonos 9 e 9 ', que alteram polaridade, solubilidade e limitam as rotações livres entre C8-C8' em relação de material (1) de partida. As diferenças estruturais entre estes compostos podem fornecer informações importantes sobre a relação estrutura-atividade antimicobacteriana do esqueleto dibenzilbutanodioico, obtido a partir de fonte natural, como um possível alvo para o desenvolvimento de drogas mais potentes contra a tuberculose

ABSTRACT: Evaluation of antimycobacterial activity of dihydrocubebin lignan extracted from Piper cubeba and its semisynthetic derivatives. The antimycobacterial activity of the dihydrocubebin (1), a dibenzylbutanedioiclignan obtained from ethanolic extract of Piper cubeba seeds, and its derivatives were examined in vitro against three different strains of Mycobacterium using amicrodilution method. Among the lignans evaluated, the 3 and 4 samples were the most active ones, displaying MIC values of 62.5 µg/mL against M. avium and M. tuberculosis, respectively. The derivatives 2-6, obtained for partial synthesis, had different substituents in the carbons 9 and 9', fact thatalters the polarity, solubility and restricts the free rotations between the bonds C8-C8' in relation to the starting material (1). The structural differences among these compounds provide important information about the antimycobacterial structure-activity relationship of the dibenzylbutanodioic skeleton, obtained from natural source, such as a possible target for the development of more powerful drugs against tuberculosis

Treatment outcomes of multidrug-resistant tuberculosis in Switzerland.

OBJECTIVE: To assess outcomes 24 months after treatment start for multidrug-resistant tuberculosis (MDR-TB). METHODS: Cohort study of all culture-positive MDR-TB cases notified in Switzerland from 01/2003 to 07/2010. RESULTS: Fifty-one cases were observed, with a median age of 26 years (range 2-56). Twenty-seven were male, five of Swiss origin, 46 of foreign origin (Asia 18, Africa 13, former Soviet Union 8), including 21 asylum seekers and refugees. Twelve had received a previous treatment for TB and 24 had not (15 unknown). Forty-four cases were pulmonary of which 25 were known to be sputum smear positive. All but two strains showed additional resistances: 29 to ethambutol, 27 to pyrazinamide, 6 to a fluoroquinolone, 5 to amikacin. None was resistant to both of the latter two classes. Molecular analyses showed three pairs of identical strains. Fluoroquinolones were used in 48 patients and second-line injectable drugs in 37. The median duration of MDR treatment was 18 months (range 1-26). The outcome after 24 months was successful in 39 (76%) and unsatisfactory in 12 (24%) patients: two deaths from TB; two treatments terminated owing to side effects of drugs and one owing to pregnancy; four defaults from treatment at months 0, 4, 8, and 21; two transfers abroad with unknown outcome; one outcome unknown. There was no significant association of unfavourable outcomes with age, sex, origin, previous treatment, treatment delay, resistance pattern, and classes of drugs used. CONCLUSIONS: MDR-TB in Switzerland occurs mostly in persons of foreign origin. Results of decentralised treatments were satisfactory.

Mycobacterium tuberculosis complex strains and drug susceptibility in a cattle-rearing region of Cameroon.

SETTING: Seven district hospitals in the Adamaoua Region of Cameroon, June 2009 to May 2010. OBJECTIVES: To identify species among Mycobacterium tuberculosis complex (MTC) strains responsible for pulmonary tuberculosis (PTB) and determine their susceptibility to anti-tuberculosis drugs. DESIGN: Sputum specimens were collected from 509 consecutively admitted adults and cultured. Identification of cultured strains was mainly based on culture growth characteristics and standard biochemical tests with spoligotyping for confirmation on a subset of strains. Drug susceptibility testing was performed using the indirect proportion method. RESULTS: Growth of MTC strains occurred in specimens of 445/509 (87.4%) patients: 433 (97.3%) were identified as M. tuberculosis, 10 (2.3%) as M. africanum and 2 (0.4%) as M. bovis. The overall resistance rate was 7.9%, with 7.3% resistance in new cases and 21.1% in previously treated cases. Isoniazid resistance in new cases was most common (4.2%), followed by streptomycin (3.3%), rifampicin (1.9%) and ethambutol (0.9%). Multidrug-resistant tuberculosis was more frequent in previously treated than in new cases (10.5% vs. 1.4%, P < 0.05). CONCLUSION: Although the Adamaoua Region is a stock-farming area, M. tuberculosis is the predominant MTC species responsible for PTB. Anti-tuberculosis drug resistance in new and previously treated cases is well established, underscoring the need to reinforce the DOTS strategy.

Association between Mycobacterium tuberculosis complex phylogenetic lineage and acquired drug resistance.

BACKGROUND: Development of resistance to antituberculosis drugs during treatment (i.e., acquired resistance) can lead to emergence of resistant strains and consequent poor clinical outcomes. However, it is unknown whether Mycobacterium tuberculosis complex species and lineage affects the likelihood of acquired resistance. METHODS: We analyzed data from the U.S. National Tuberculosis Surveillance System and National Tuberculosis Genotyping Service for tuberculosis cases during 2004-2011 with assigned species and lineage and both initial and final drug susceptibility test results. We determined univariate associations between species and lineage of Mycobacterium tuberculosis complex bacteria and acquired resistance to isoniazid, rifamycins, fluoroquinolones, and second-line injectables. We used Poisson regression with backward elimination to generate multivariable models for acquired resistance to isoniazid and rifamycins. RESULTS: M. bovis was independently associated with acquired resistance to isoniazid (adjusted prevalence ratio = 8.46, 95% CI 2.96-24.14) adjusting for HIV status, and with acquired resistance to rifamycins (adjusted prevalence ratio = 4.53, 95% CI 1.29-15.90) adjusting for homelessness, HIV status, initial resistance to isoniazid, site of disease, and administration of therapy. East Asian lineage was associated with acquired resistance to fluoroquinolones (prevalence ratio = 6.10, 95% CI 1.56-23.83). CONCLUSIONS: We found an association between mycobacterial species and lineage and acquired drug resistance using U.S. surveillance data. Prospective clinical studies are needed to determine the clinical significance of these findings, including whether rapid genotyping of isolates at the outset of treatment may benefit patient management.