Mycobacterium tuberculosis lineage 7 strains are associated with prolonged patient delay in seeking treatment for pulmonary tuberculosis in Amhara Region, Ethiopia.

Recent genotyping studies of Mycobacterium tuberculosis in Ethiopia have reported the identification of a new phylogenetically distinct M. tuberculosis lineage, lineage 7. We therefore investigated the genetic diversity and association of specific M. tuberculosis lineages with sociodemographic and clinical parameters among pulmonary TB patients in the Amhara Region, Ethiopia. DNA was isolated from M. tuberculosis-positive sputum specimens (n=240) and analyzed by PCR and 24-locus mycobacterial interspersed repetitive unit-variable-number tandem-repeat (MIRU-VNTR) analysis and spoligotyping. Bioinformatic analysis assigned the M. tuberculosis genotypes to global lineages, and associations between patient characteristics and genotype were evaluated using logistic regression analysis. The study revealed a high diversity of modern and premodern M. tuberculosis lineages, among which approximately 25% were not previously reported. Among the M. tuberculosis strains (n=138) assigned to seven subgroups, the largest cluster belonged to the lineage Central Asian (CAS) (n=60; 26.0%), the second largest to lineage 7 (n=36; 15.6%), and the third largest to the lineage Haarlem (n=35; 15.2%). Four sublineages were new in the MIRU-VNTRplus database, designated NW-ETH3, NW-ETH1, NW-ETH2, and NW-ETH4, which included 24 (10.4%), 18 (7.8%), 8 (3.5%), and 5 (2.2%) isolates, respectively. Notably, patient delay in seeking treatment was significantly longer among patients infected with lineage 7 strains (Mann-Whitney test, P<0.008) than in patients infected with CAS strains (adjusted odds ratio [AOR], 4.7; 95% confidence interval [CI], 1.6 to 13.5). Lineage 7 strains also grew more slowly than other M. tuberculosis strains. Cases of Haarlem (OR, 2.8; 95% CI, 1.2 to 6.6) and NW-ETH3 (OR, 2.8; 95% CI, 1.0 to 7.3) infection appeared in defined clusters. Intensified active case finding and contact tracing activities in the study region are needed to expedite diagnosis and treatment of TB.

Evolution of extensively drug-resistant Mycobacterium tuberculosis from a susceptible ancestor in a single patient.

BACKGROUND: Mycobacterium tuberculosis is characterized by a low mutation rate and a lack of genetic recombination. Yet, the rise of extensively resistant strains paints a picture of a microbe with an impressive adaptive potential. Here we describe the first documented case of extensively drug-resistant tuberculosis evolved from a susceptible ancestor within a single patient. RESULTS: Genome sequences of nine serial M. tuberculosis isolates from the same patient uncovered a dramatic turnover of competing lineages driven by the emergence, and subsequent fixation or loss of single nucleotide polymorphisms. For most drugs, resistance arose through independent emergence of mutations in more than one clone, of which only one ultimately prevailed as the clone carrying it expanded, displacing the other clones in the process. The vast majority of mutations identified over 3.5 years were either involved in drug resistance or hitchhiking in the genetic background of these. Additionally, RNA-sequencing of isolates grown in the absence of drug challenge revealed that the efflux-associated iniBAC operon was up-regulated over time, whereas down-regulated genes include those involved in mycolic acid synthesis. CONCLUSIONS: We observed both rapid acquisitions of resistance to antimicrobial compounds mediated by individual mutations as well as a gradual increase in fitness in the presence of antibiotics, likely driven by stable gene expression reprogramming. The rapid turnover of resistance mutations and hitchhiking neutral mutations has major implications for inferring tuberculosis transmission events in situations where drug resistance evolves within transmission chains.

Dominant Mycobacterium tuberculosis lineages in elderly patients born in Norway.

BACKGROUND: During the previous century Norway had a high incidence of tuberculosis, but no molecular epidemiological studies could be performed and these previously epidemic strains have been disappearing during the last decades. Currently, tuberculosis among native Norwegians is in the elimination phase, and it is still not known what type of M. tuberculosis was so efficiently controlled during the second half of the 20th century. However, many elderly Norwegian-born people still develop TB that cannot be clustered to imported or recently transmitted strains of M. tuberculosis. Thus, the majority of these cases are results of reactivation of disease that was transmitted many decades ago. METHODOLOGY/PRINCIPAL FINDINGS: A total of 213 strains of M. tuberculosis isolated during 1998-2005, from patients born in Norway before 1950 were genotyped in the current study. The findings demonstrated a highly homogenous M. tuberculosis population among the patients. A total of 40% belonged to the T-family, were 35% were assigned to T1 sub- family (T2 = 0, 93%, T3 = 1, 4% and T4 = 2, 3%). As many as 35% of the isolates belonged to the Haarlem family, were 15% were assigned to Haarlem1 and 19% to Haarlem3. The remaining 25% belonged to 15 different other families. The RFLP-patterns indicated that the isolates were not a result of recent transmission, but rather represented well established strains that apparently dominated in Norway many decades ago. CONCLUSIONS/SIGNIFICANCE: The T 1, Haarlem 1, and Haarlem 3 families of M. tuberculosis were abundant among patients born in Norway before 1950. The M. tuberculosis cases represented reactivated disease that had been acquired before 1994 and were likely to have been latent for several decades. Thus, the current study indicated that the T 1, Haarlem 1, and Haarlem 3 families may have been common in Norway, when tuberculosis represented a serious public health threat during the first half of the 20th century.