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Edit Comment Close Premium member Presentation Transcript Slide1: HIV STUDIES Laboratory diagnosis Serology Viral load Genotyping CD4/CD8 Research Epidemiology Immune response Co-receptors Virus subtypes Slide2: Epidemiology Immune response Genetic factors Virus subtypes Slide3: Research in epidemiology Central America collaborative project -PASCA, M. of Health and the University of Washington -Study population: Sex workers and men that have sex with men from CA -Preliminary results HIV prevalence: SW: 2% MHSM: 11% Other STDs C. trachomatis: 15% NG: 4% NG+CT: 2% Prevalence of HIV, Sifilis y Trichomonas and any other STD in CSW : Prevalence of HIV, Sifilis y Trichomonas and any other STD in CSW Prevalence of HIV, Sifilis y Trichomonas and any other STD in CSW by place of work : Prevalence of HIV, Sifilis y Trichomonas and any other STD in CSW by place of work Prevalence of HIV, Sifilis y Trichomonas and any other STD in CSW by type of activity : Prevalence of HIV, Sifilis y Trichomonas and any other STD in CSW by type of activity Prevalence of N. gonorrhoeae and C. trachomatis in CSW in Panama, N=434: Prevalence of N. gonorrhoeae and C. trachomatis in CSW in Panama, N=434Slide8: Epidemiology Immune response Genetic factors Virus subtypes Immune markers of progression: Immune markers of progression Define less expensive markers of progression in HIV infected patients Study population: 75 HIV+ patients, 75 HIV- subjects Markers: CD4, CD8, total lymphocytes Beta 2-microglobulin P-24 Immunoglobulins Pascale, J. M. et al. Immunological markers of disease progression in patients infected with the Human Immunodeficiency Virus. Clin Diag Lab Immunol Vol.4, No. 4, 1997.Slide10: T lymphocytes in HIV+ and HIV- subjects CELLS/MICROLITER HIV- SUBJECTS 766 1120 2596 1112 784 2393 708 238 1451 CD4/CD8 1.46 0.70 0.34 HIV+ PATIENTS ASYMPTOMATIC AIDSSlide11: B-2 MICROGLOBULIN CONCENTRATION (mg/l) 2.2 + 0.7 3.4 + 2.1 5.7 + 3.6 HIV- SUBJECTS ASYMPTOMATIC AIDS HIV + PATIENTS B-2 Microglobulin Beta 2 Microglobulin in plasma of HIV+ and HIV- subjects Slide12: p-24 in sera CD4 cells/ul p-24 (pg/ml) 52 40 18 <200 200-500 >500 HIV- p24 levelsSlide13: IL-2 and TNF CD4 cells/ul IL-2 and TNF alpha (pg/ml) <200 200-500 >500 HIV- TNF IL-2 Slide14: IgG, IgA and IgM levels in HIV+ and HIV- subjects CD4 cells/ul Ig concentration mg/dl <200 200-500 >500 HIV - Total IgG Total IgA Total IgM Percentages of HIV+ patients showing specific levels of immune markers of progression: Percentages of HIV+ patients showing specific levels of immune markers of progression Immune marker Asymptomatic AIDS (%) (%) CD4 <200 cells/ml 4 96 TL <1750 cells/ ml 15 89 B2-M >4 mg/l 25 81 IgA >450 mg/dl 34 72 TL+B2M 11 91Slide16: Epidemiology Immune response Genetic factors Virus subtypes Co-receptors used by the HIV to invade cells: Co-receptors used by the HIV to invade cells CCR5 Macrophages APC CXCR4 T cells CCR5 mutations: CCR5 mutations CCR5 is coded by the CMKBR5 gen. At least 20 mutations of the CCR5 receptors have been identified. The 32 deletion protects homozygous subjects from HIV infection, and delays progression of the disease in heterozygous patients.Receptor del CCR5-wt: Receptor del CCR5-wtReceptor de CCR5-32: Receptor de CCR5-32Methodology: Methodology DNA extraction: - Buffy coat Amplification of DNA: - One tube PCR Electrophoresis: - 2% Agarose-TBEResults: Results 188 bp Wt/Wt 156 bp Wt/32 32 Deletion : 32 Deletion Results: Results 1- From the 83 patients 8 were (wt/32). 2- 75 patients were wt/wt. 3- All (wt/32) survived more than 11 years without treatment (slow progressors). 4-None of the (wt/32) children were infected.Slide26: Epidemiology Immune response Genetic factors Virus subtypes Slide27: HIV-1 SUBTYPESSlide28: Determine HIV-1 subtypes circulating in Panama. Evaluate the genetic diversity of the HIV subtypes identified. Correlate HIV subtypes variability with transmission patterns, immune response, and progression. OBJECTIVESSlide29: Why gp41? Small fragment Highly conserved region between M,N and O (Agwale et al (2001) J. Clin. Microbiol. 39:2110-2114). Can be used for phylogenetic analyses Study Samples: Study Samples Subjects: 25 HIV positive patients from Saint Thomas Hospital that came for viral load evaluation. Method: Gp41 Sequencing Slide31: Methodology DNA extraction gp41 Nested PCR Chunfu Y. et al. J. Clin. Microbiol, Vol. 38, 1999. Sequencing 1058 pb 1516 pb 460bpSlide32: SEQUENCING 1-Genotyping: drug resistance 2-HIV subtypesPanama vs other Subtypes: Panama vs other Subtypes Subtype B Other SubtypesFuture Research: Future Research 1-To determine the importance of other mutations in progression of the infection. 2-Immune mediators of resistance and/or progression. 3-Circulation of different HIV subtypes and recompbinant forms in Panama.Where we are??: Where we are?? 1-What do we have? More than 300 samples (plasma and buffy coat) of HIV infected patients. 2-What facilities do we have? Flow cytometer, ABI-310 sequencer, viral load and genotying equipments. 3-What do we need? Improved softwares for subtyping and genetic analysis of recombinant forms.Slide36: INSTITUTO CONMEMORATIVO GORGAS You do not have the permission to view this presentation. In order to view it, please contact the author of the presentation.