Background & objectives:Mycobacterium tuberculosis (M. tuberculosis) has four homologous mammalian cell entry (mce) operons (mce1-4) that encode exported proteins and have a possible role in the virulence mechanism of this pathogen. The expression of mce operon is considered to be complex and not completely understood. Although expression of mce operon at different in vitro growth phases has been studied earlier, its expression in different M. tuberculosis isolates under different growth phases is not yet studied. The present preliminary study was conducted on a limited number of isolates to know the trend of expression pattern of mce operon genes in different M. tuberculosis isolates under different growth stages.Methods:In this study, we monitored the transcriptional profile of selected mce operon genes (mce1A, mce1D, mce2A, mce2D, mce3A, mce3C) in different M. tuberculosis isolates (MDR1, MDR2, and sensitive isolate) at early exponential and stationary phases using real-time quantitative PCR.Results:The expression ratio of all selected mce operon genes in all M. tuberculosis isolates was reduced at the initial phase and increased substantially at a later phase of growth. Higher expression of mce1 operon genes was found in all M. tuberculosis isolates as compared to other mce operon genes (mce2 and mce3 operons) at stationary growth phase.Interpretation & conclusions:The higher expression of mce operon genes at stationary phase (as compared to early exponential phase) suggested growth phase dependent expression of mce operon genes. This indicated that the mce operon genes might have a role in M. tuberculosis survival and adaptation on the onset of adverse condition like stationary phase. Identification of differentially expressed genes will add to our understanding of the bacilli involved in adaptation to different growth conditions.
Background Characterization of drug resistance mutations and signature pattern in the archived DNA of HIV-1 infected North Indian patients.Methods Blood samples were collected from 9 patients enrolled in ART Centre, S.N. Medical College, Agra, North India from 1 year to = < 7 years. DNA was isolated and amplified for protease and reverse transcriptase genes using nested primers from WHO dried blood spot protocol 2010. The polymerase chain reaction products were sequenced and drug resistance mutation patterns were analysed using the HIV drug resistance database, Stanford University, USA. Various computational tools and websites like Stanford HIV drug resistance database, Viral epidemiological signature pattern analysis (VESPA), Hypermutation, SNAP version 2.1.1, Entropy were utilized for analysis of the sequence data.Results This study enables an analysis of archived DNA samples in patients having lower viral load and where the situation of HIV RNA isolation was negligible. Lamivudine associated drug-resistant mutations such as M184V/M184I, nevirapine associated mutations Y181C and H221Y and efavirenz associated mutations M230I were observed in two patients. No mutations were observed in the remaining seven patients. The signature pattern analysis of 9 protease and reverse transcriptase genes identified the conservation of amino acid sequences as compared to the reference sequence. The signature pattern of nucleotide substitution (synonymous to non-synonymous ratio) of the protease gene was 5.02 and the reverse transcriptase gene was 7.01. No hyper mutation was observed in the protease and reverse transcriptase gene of the archived DNA.Conclusions The analysis of the drug resistance mutation in the protease and reverse transcriptase gene of the archived DNA of HIV-1 subtype C infected patients over 1 to = < 7 years of first-line ART may be helpful in the treatment guideline in North Indian patients. A few signature amino acids were persisted in the reverse transcriptase and protease gene of similar HIV-1 subtype C infected patients over 1 to = < 7 years of first line ART in comparison to the reference sequence. This archived DNA drug resistance mutation testing could be an important tool when RNA testing becomes unsuccessful.
The aim of this study was to analyze the sociodemographic, clinical, immunologic, and risk factors of drug-resistant (>1000 copies/ml) and virologically suppressed (<1000 to <40 copies /ml to target not detected levels) patients at the ART center, Sarojini Naidu Medical College, Agra, North India. A total of 193 patients on first-line antiretroviral therapy were included those were recruited from December 2009 to November 2016. The patients included in this study had a CD4 cell count of ≤ 350 cubic/mm. The details of Demographic, clinical, and social factors were collected in a patient information leaflet and statistically analyzed. After viral load, two groups of patients were observed. The drug-resistant group (N=58) had a viral load ≥1000 copies/ml and virologically suppressed group (N=135) had a viral load <1000 copies/ml to target not detected level. A comparison statement result was presented in both groups of drug-resistant and virologically suppressed patients. Males were observed in the highest frequency (42, 72.41%). The heterosexual mode of transmission was predominant (40, 68.96%, and 98,72.59%). The highest number of married couples and illiterates in the two groups. Tuberculosis was observed in the highest numbers in the two groups. The analysis of socioeconomic factors of North Indian patients will be a concern issue in the demographic profiles of HIV-1 patients. The clinical features, analysis would help clinicians in further therapy implementation, monitoring CD4 and Viral load counts of North Indian patients.
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