Background: Drug resistance in Mycobacterium tuberculosis is primarily attributed to mutations in target genes. These mutations incur a fitness cost and result in bacterial generations that are less fit, which subsequently acquire compensatory mutations to restore fitness. We hypothesize that mutations in specific drug target genes influence bacterial metabolism and cellular function, which affects its ability to develop subsequent resistance to additional agents. We aim to determine whether the sequential acquisition of drug resistance and specific mutations in a well-defined clinical M. tuberculosis strain promotes or limits the development of additional resistance.Methods & Materials: In vitro mutants resistant to pretomanid, linezolid, moxifloxacin, rifampicin and kanamycin were generated from a pan-susceptible clinical strain from the Beijing lineage. The resistant phenotypes to the anti-TB agents were confirmed by the broth microdilution assay and genetic mutations were identified by targeted gene sequencing. Growth of mono-resistant mutants was done in enriched medium for 14 days to assess in vitro fitness. Double resistant mutants were generated against anti-TB drug combinations at concentrations 5x and 10x the MIC. Subsequently, mutation frequencies for these anti-TB drugs in the different monoresistant backgrounds were determined.Results: The initial level of resistance and the mutation frequencies observed for the mono-resistant mutants were comparable to those previously reported. Targeted gene sequencing revealed the presence of known and clinically relevant mutations in the mutants resistant to linezolid, rifampicin, kanamycin and moxifloxacin. Significant growth defects were observed for mutants grown under in vitro conditions compared to the sensitive progenitor. Mutation frequencies determination in the mono-resistant mutants revealed a significant increase in mutation frequency against rifampicin and kanamycin, but a significant decrease in mutation frequency against linezolid and sutezolid. This suggests that these mono-resistant mutants are more prone to develop resistance to rifampicin and kanamycin, but less prone to develop resistance against linezolid and sutezolid. Even though kanamycin and linezolid both inhibit protein synthesis, these compounds target different subunits of the ribosome, thereby leading to different outcomes in terms of fitness in the mutants with impaired cellular function.Conclusion: These observations showed that oxazolidinone treatment is instrumental in limiting the development of multidrug resistance in M. tuberculosis in vitro.

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BackgroundIncreased antibiotic resistance is a significant problem associated with higher morbidity, mortality, and costs in managing urinary tract infections (UTI). This study aims to analyze the antimicrobial resistance profile of uropathogens isolated from children with the first attack of community-acquired UTI.Materials and MethodsThis retrospective cohort study was conducted between January 2010-December 2016, with the patients aged one month to 18 years diagnosed with the first attack of community-acquired UTI at the pediatric outpatient clinic of a tertiary care hospital. Patients' age, sex, UTI history, clinical findings at admission, urine analysis, urine culture, and antibiogram results were recorded.ResultsA total of 1086 patients and 1086 cultures were included in the study. The male/female ratio was 1/5.3. The mean age of the children was 73.7±47.1 (0.4-215.9) months. 16.0% were aged 1-24 months. E. coli was the most common causative agent found to be 85.1% in frequency. The overall antimicrobial resistance was found to be highest for ampicillin, followed by piperacillin and trimethoprim-sulfamethoxazole (63.5, 41.6 and 38.1%, respectively). Imipenem, amikacin, and tobramycin had the least resistance (0.5, 0.5, and 7.6%, respectively). The overall antimicrobial resistance against amoxicillin-clavulanate, ampicillin-sulbactam, cefuroxime, ceftriaxone was 19.4, 24, 25.9 and 21.1%; respectively.Extended-spectrum beta-lactamase positivity was detected in 5.6% (n=61) of samples.ConclusionOur study showed significant increase in antimicrobial resistance to the most common antibiotics which are prescribed before urine culture results and antibiotic sensitivities are available. The resistance rates for amoxicillin-clavulanate, ampicillin-sulbactam, cefuroxime, ceftriaxone were almost 20% or above in children with the first UTI attack in an outpatient setting.