ABSTRACT
Background Although Leber hereditary optic neuropathy (LHON) and optic neuritis have different causes and managements,their clinical manifestations are difficult to be distinguished.Real-time fluorescence quantitative polymerase chain reaction(RTFQ-PCR) is a high flux,simple,rapid and specific detecting technology,so establishing a specific diagnosis method of LHON with RTFQ-PCR has a practical significance.Objective Purpose of the present study was to establish a real-time Taqman probe for mitochondrial DNA (mtDNA)11778G＞A mutation in LHON patients.Methods Primers and Taqman probe for mtDNA 11778G＞A mutation were designed based on mtDNA complete geneme.Eighty-four patients with LHON were selected from the LHON DNA bank of Molecular Biology Laboratory,Henan Eye Institute,and 40 normal physical examinees aged 18-20 years were from Henan People's Hospital.2 ml of periphery blood was collected from each individual.Based on the double-blindness principle,mtDNA 11778G＞A mutation was tested by both Taqman probe and sequencing to check the reliability of real-time Taqman probe.Results The mtDNA 11778G＞A mutation was found in 23 out of 84 patients,and 61 showed a negative result by the technique of real-time Taqman probe.The Ct values of 23 patients with mtDNA 11778G＞A mutation were 22.993 ±0.708,but those of 5 normal controls were 0.These findings showed a consistent rate of 100％ with the sequencing results.In addition,both the false positive rate and the false negative rate were zero.Conclusions Real-time Taqman probe technique is an accurate,convenient,sensitive,specific and intuitionistic method for the diagnosis of mtDNA 11778G＞A mutation in LHON patients.It is feasible and suitable to screen the LHON patients with mtDNA 11778G＞A mutation in a large scale.
ABSTRACT
To investigate the subtype distribution of human immunodeficiency virus-1(HIV-1) infection among men who have sex with men (MSM) in Zhengzhou, Henan Province, forty blood samples were collected from HIV-1 carriers, who acknowledged to have sex with men. The complete gag gene was amplified by RT-PCR and nested-PCR and sequenced. All sequences were edited by BioEdit and subtyped by genotyping software. Phylogenetic analysis of gag gene were then performed using the MEGA 3.1 software, the gene distances were calculated by Distance program. There were three different HIV-1 subtypes including B, CRF01-AE and CRF07-BC present among twenty four MSMs in Zhengzhou. Genotyping results showed that 33.33% (8/24) were B, 41.67% (10/24) were CRF01-AE and 25% (6/24) were CRF07-BC, and subtype CRF01-AE had become the most prevalent HIV-1 subtype in Zhengzhou, Henan province. In conclusion, recombinant HIV-1 strains are circulating in Henan province and the epidemiology is complicated.
Subject(s)
Adult , Humans , Male , Middle Aged , Young Adult , China , HIV-1 , Classification , Genetics , Homosexuality, Male , Phylogeny , Sequence Analysis, DNA , gag Gene Products, Human Immunodeficiency Virus , GeneticsABSTRACT
<p><b>OBJECTIVE</b>This study aimed to investigate the subtype distribution of gp41 gene of human immunodeficiency virus-1 (HIV-1) among men who have sex with men (MSM) in Zhengzhou.</p><p><b>METHODS</b>Thirty blood samples were collected from men who have sex with men infected by HIV. The complete gp41 gene was amplified by RT-PCR and nested-PCR and sequenced. All sequences were edited by Bioedit and subtyped with HIV sequence library US Los Alamos National Laboratory and online genotyping software provided by American National Center of Biotechnology Information. Phylogenetic analysis of gp41 gene was performed using the MEGA 3.1 software, and the genic dispersion rates among subtype of gp41 gene were analyzed.</p><p><b>RESULTS</b>A total of eighteen gene sequences of HIV-1 gp41 gene were obtained from thirty men who have sex with men infected by HIV, which belonged to subtype CRF15-01B (50% (9/18)), CRF01-AE (22% (4/18)), CRF07-B (22% (4/18)) and B (6% (1/18)), respectively. The intersubtype HIV-1 strains aggregate with according reference strains. The genetic distance inter-subtype of subtype CRF15-01B, CRF01-AE and CRF07-B were 0.050 ± 0.007, 0.052 ± 0.009 and 0.082 ± 0.012, respectively.</p><p><b>CONCLUSION</b>The prevalent subtypes of HIV-1 among among MSM in Zhengzhou was complicated and recombinant HIV-1 strains were the most prevalent strains.</p>