Drug Resistance Mutation Profiles Research Articles

Evaluation of the GenoType® MTBDRplus assay and identification of a rare mutation for improving MDR-TB detection

To assess the new GenoType® MTBDRplus assay for the rapid detection of multidrug-resistant tuberculosis (MDR-TB) in comparison with DNA sequencing to identify drug resistance mutation profiles in China. Using MTBDRplus, drug susceptibility testing (DST) and DNA sequencing, 237 Mycobacterium tuberculosis strains were tested. The sensitivity of MTBDRplus was 75.0% (126/168) for isoniazid (INH) resistant strains, and 93.5% (157/168) for rifampicin (RMP) resistant strains. It correlated well with sequencing, with 94.9% and 99.6% agreement for each strain category and 100% specificity for all categories. The two most common rpoB mutations were S531L (53.6%, 90/168) and D516G (17.3%, 29/168) in RMP-resistant strains. INH resistance was dominated by the katG 315 locus (S to T, N, R, I) mutation (73.7%, 124/168), and a rare katG mutation, S315N (6.5%, 11/168), not covered by MTBDRplus was identified. The mutation combination inhA-15/inhA-8 and katG315 (34 strains) was characteristically displayed in MDR-TB strains (23.5%), but not in INH-monoresistant strains. Although Genotype MTBDRplus is a rapid and reliable molecular test for detecting MDR-TB, a significant proportion of strains in China contain a rare katG S315N mutation that would be missed by the assay. Further improvements may be achieved by incorporating this mutation into the assay to increase sensitivity in detecting INH resistance in China.

The human immunodeficiency virus‐1 genotype diversity and drug resistance mutations profile of volunteer blood donors from Chinese blood centers

The global human immunodeficiency virus (HIV)-1 epidemic is becoming increasingly diverse and complex. Molecular epidemiologic characteristics were studied for HIV-1-infected blood donors from five Chinese regions to determine genotype diversity and drug resistance mutations (DRMs) profile. HIV-1 confirmed-reactive serum samples were collected from 172 blood donors from five blood centers during 2007 to 2010. HIV-1 Pol including whole protease and partial reverse transcriptase genes was amplified, sequenced, and analyzed for the subtype determination and drug resistance profile description. A total of 113 amplified sequences including 82 from Kunming blood center and 31 from four other blood centers had the following genotype characteristics: G (0.9%), B (2.7%), circulating recombinant form (CRF) 01_AE (32.7%), CRF07_BC (22.1%), and CRF08_BC (41.6%). Female donors represent 45.1% of all cases and 63.9% cases with DRMs. The prevalence of samples with potential low or higher resistance among Chinese blood donors is 4.4%. HIV-1 infection in Chinese blood donors is genetically diverse and the subtype distribution reflects that from the high-risk populations. Our results support continuous molecular epidemiologic surveillance for HIV-1 in blood donors as a part of a comprehensive HIV control program.

Affordable in-house antiretroviral drug resistance assay with good performance in non-subtype B HIV-1

The introduction of antiretroviral (ARV) therapy in resource-poor settings is effective in suppressing HIV-1 replication and prolonging life of infected individuals. This has led to a demand for affordable HIV-1 drug resistance assays, since treatment failure due to development of drug resistance is common. This study developed and evaluated an affordable “in–house” genotyping assay to monitor HIV-1 drug resistance in Africa, particularly South Africa. An “in-house” assay using automated RNA extraction, and subtype C specific PCR and sequencing primers was developed and successfully evaluated 396 patient samples (viral load ranges 1000–1.6 million RNA copies/ml). The “in-house” assay was validated by comparing sequence data and drug resistance profiles from 90 patient and 10 external quality control samples to data from the ViroSeq™ HIV-1 Genotyping kit. The “in-house” assay was more efficient, amplifying all 100 samples, compared to 91 samples using Viroseq. The “in house” sequences were 99.2% homologous to the ViroSeq sequences, and identical drug resistance mutation profiles were observed in 96 samples. Furthermore, the “in-house” assay genotyped 260 of 295 samples from seven African sites, where 47% were non-subtype C. Overall, the newly validated “in-house” drug resistance assay is suited for use in Africa as it overcomes the obstacle of subtype diversity.

Drug Resistance Mutation Profile and Accumulation Kinetics in Human Immunodeficiency Virus-Positive Individuals Infected with Subtypes B and F Failing Highly Active Antiretroviral Therapy Are Influenced by Different Viral Codon Usage Patterns

The major human immunodeficiency virus type 1 subtype circulating in Brazil is B, followed by F and C. We have genotyped 882 samples from Brazilian patients for whom highly active antiretroviral therapy failed, and we found subtype B and the unique recombinant B/F1 forms circulating. Due to codon usage variation, there is a significantly lower incidence of the substitutions L210W, Q151M, and F116Y in subtype F1 isolates than in the subtype B counterparts.

Profile of drug resistance mutations among HIV-1-infected Tunisian subjects failing antiretroviral therapy

Three years after the introduction of antiretroviral therapy (ART) in Tunisia (North Africa), we aimed to determine the prevalence of drug resistance mutations in Tunisian HIV-1-infected patients failing ART. Plasma samples of 80 patients were tested for genotypic resistance using two distinct line probe assays, LiPA HIV-1 reverse transcriptase RT and LiPA HIV-1 protease assay. Of the 80 patients, 82.5% showed resistance to at least one antiretroviral molecule. In the RT gene, resistance to nucleoside RT inhibitors (NRTIs) and non-nucleoside RT inhibitors (NNRTIs) were recognized in 66.25 and 37.5%, respectively, with M184V, T215Y and K103N being the codons most frequently involved. Resistance to protease inhibitors (PIs) was found in 46.25% of cases. Despite the presence of different mutations, the viral variants were still susceptible to other RTIs and PIs that are currently not available in Tunisia. Thus, alternative therapeutic options exist but are not yet accessible.

Human immunodeficiency virus type 1 (HIV-1) genotyping in Rio de Janeiro, Brazil: assessing subtype and drug-resistance associated mutations in HIV-1 infected individuals failing highly active antiretroviral therapy

In order to assess the human immunodeficiency virus type 1 (HIV-1) drug resistance mutation profiles and evaluate the distribution of the genetic subtypes in the state of Rio de Janeiro, Brazil, blood samples from 547 HIV-1 infected patients failing antiretroviral (ARV) therapy, were collected during the years 2002 and 2003 to perform the viral resistance genotyping at the Renageno Laboratory from Rio de Janeiro (Oswaldo Cruz Foundation). Viral resistance genotyping was performed using ViroSeq Genotyping System (Celera Diagnostic-Abbott, US). The HIV-1 subtyping based on polymerase (pol) gene sequences (protease and reverse transcriptase-RT regions) was as follows: subtype B (91.2%), subtype F (4.9%), and B/F viral recombinant forms (3.3%). The subtype C was identified in two patients (0.4%) and the recombinant CRF_02/AG virus was found infecting one patient (0.2%). The HIV-1 genotyping profile associated to the reverse transcriptase inhibitors has shown a high frequency of the M184V mutation followed by the timidine-associated mutations. The K103N mutation was the most prevalent to the non-nucleoside RT inhibitor and the resistance associated to protease inhibitor showed the minor mutations L63P, L10F/R, and A71V as the more prevalent. A large proportion of subtype B was observed in HIV-1 treated patients from Rio de Janeiro. In addition, we have identified the circulation of drug-resistant HIV-1 subtype C and are presenting the first report of the occurrence of an African recombinant CRF_02/AG virus in Rio de Janeiro, Brazil. A clear association between HIV-1 subtypes and protease resistance mutations was observed in this study. The maintenance of resistance genotyping programs for HIV-1 failing patients is important to the management of ARV therapies and to attempt and monitor the HIV-1 subtype prevalence in Brazil.