Diversity and Divergence in HIV-1 Viral Variants between patients with high CD4+ T Cell Variability and Patients with Rapid CD4+ T Cell Decline Kevin Paiz-Ramirez Janelle N. Ruiz Biology Professor Kam D. Dahlquist, Ph.D. Department of Biology Loyola Marymount University March 2nd, 2009

Outline I. HIV-1 Background II. Markham et al. Previous Studies III. Research Questions IV. Results A. Diversity Between Groups B. Divergence Between Groups V. Interpretations between synonymous & non- synonymous Groups VI. Limitations To Results VII. Interpretations In Light of Recent Studies VIII. References

Background HIV-1 Human Immunodeficiency Virus, is categorized into 3 stages Seroconversion ~7-8 weeks Latency ~variable period of years Acquired Immune Deficency Syndrome Center for Disease Control’s Public Health Image Library #10000

Markham, et al. Markham et al. explored analyses of synonymous and non-synonymous differences and rates of divergence to reveal selective heterogeneity as a function of disease progression category We chose to examine if there are differences in HIV-1 diversity or divergence between participants with high CD4 T-Cell variability within the study, between visits as compared to participants with linear progression

Research Question Are their differences in HIV-1 diversity or divergence between participants with high CD4 T cell variability between visits as compared to participants with rapid CD4+ T Cell decline (slope ~ -1)? Linear ProgressorsHigh Variability

How Were Subjects Chosen? Linear Progressors: Patients with rapid CD4+ T Cell Decline

High Variability: Patients with high CD4 T cell variability between visits How Were Subjects Chosen?

High Variability: Patients with high CD4 T cell variability between visits How Were Subjects Chosen?

Low Variability: Patients with low CD4 T cell variability between visits How Were Subjects Chosen?

Diversity and Divergence Between Groups SubjectClassification Number of visits # of Clones (all visits)SThetaMin Difference Max Difference Median dS/dN 4Linear Progressor Linear Progressor High Variability High Variability High Variability Low Variability

Diversity and Divergence Between Groups SubjectClassification Number of visits # of Clones (all visits)SThetaMin Difference Max Difference Median dS/dN 4Linear Progressor Linear Progressor High Variability High Variability High Variability Low Variability

Low Variability DIVERSITY: Comparing Number of Clones for all Visits Across Groups

Diversity and Divergence Between Groups SubjectClassification Number of visits # of Clones (all visits)SThetaMin Difference Max Difference Median dS/dN 4Linear Progressor Linear Progressor High Variability High Variability High Variability Low Variability

Diversity and Divergence Between Groups SubjectClassification Number of visits # of Clones (all visits)SThetaMin Difference Max Difference Median dS/dN 4Linear Progressor Linear Progressor High Variability High Variability High Variability Low Variability

DIVERSITY: Comparing the S Value Across Groups Subject 4 Subject 10 Subject 12 Subject 8 Subject 6 Subject 5 Linear ProgressorsHigh Variability Low Variability Group # S Value

DIVERSITY: Comparing the S Value Across Groups Subject 4 Subject 10 Subject 12 Subject 8 Subject 5 Linear ProgressorsHigh Variability Low Variability Group # S Value P Value: Subject 6

DIVERSITY: Comparing the S Value Across Groups Subject 4 Subject 10 Subject 12 Subject 8 Subject 6 Subject 5 Linear ProgressorsHigh Variability Low Variability Group # S Value P Value:

Diversity and Divergence Between Groups SubjectClassification Number of visits # of Clones (all visits)SThetaMin Difference Max Difference Median dS/dN 4Linear Progressor Linear Progressor High Variability High Variability High Variability Low Variability

DIVERGENCE: Comparing Theta Across Groups Subject 4 Subject 10 Subject 12 Subject 8 Subject 6 Subject 5 Group # Theta Linear ProgressorsHigh Variability Low Variability

DIVERGENCE: Comparing Theta Across Groups Subject 4 Subject 10 Subject 12 Subject 8 Subject 6 Subject 5 Group # Theta Linear ProgressorsHigh Variability Low Variability P Value:

DIVERGENCE: Comparing Theta Across Groups Subject 4 Subject 10 Subject 12 Subject 8 Subject 6 Subject 5 Group # Theta Linear ProgressorsHigh Variability Low Variability P Value:

Diversity and Divergence Between Groups SubjectClassification Number of visits # of Clones (all visits)SThetaMin Difference Max Difference Median dS/dN 4Linear Progressor Linear Progressor High Variability High Variability High Variability Low Variability

DIVERGENCE: Comparing Min and Max Differences Across Groups SubjectClassification # of Clones (all visits)Min DifferenceMax Difference 4Linear Progressor Linear Progressor High Variability High Variability High Variability Low Variability P Value: >.05

DIVERGENCE: Comparing Min and Max Differences Across Groups SubjectClassification # of Clones (all visits)Min DifferenceMax Difference 4Linear Progressor Linear Progressor High Variability High Variability High Variability Low Variability P Value: >.05

Research Question Are their differences in HIV-1 diversity or divergence between participants with high CD4 T cell variability between visits as compared to participants with rapid CD4+ T Cell decline (slope ~ -1)? Linear ProgressorsHigh Variability

Differences exist in diversity and divergence of HIV-1 variants b/w Groups Linear Progressors show higher diversity HIV-1 variants as shown by higher number of clones and higher S value compared to subjects with High Variability in CD4 count. Linear Progressors show higher divergence in HIV-1 variants as shown by higher value of theta compared to subjects with High Variability in CD4 count. Hypothesis supported?

Limitations to Results Only looked at first 5 visits to maintain consistency and due to limitations of analytical software Issues with Subject 8 versus Subject 6 in high variability group Missing data for subject 4 (visits 5-7) Only one subjects in low variability group; therefore could not run T Tests with this group

Synonymous versus Non-Synonymous mutations Between Groups SubjectClassification Number of visits # of Clones (all visits)SThetaMin Difference Max Difference Median dS/dN 4Linear Progressor Linear Progressor High Variability High Variability High Variability Low Variability P Value: >.05

Synonymous versus Non-Synonymous mutations Between Groups SubjectClassification Number of visits # of Clones (all visits)SThetaMin Difference Max Difference Median dS/dN 4Linear Progressor Linear Progressor High Variability High Variability High Variability Low Variability P Value: >.05

Synonymous versus Non-Synonymous mutations Between Groups Appears to be no significant differences in synonymous versus non-synonymous mutations in HIV-1 variants between linear progressors and high variability group Participants with low variability in CD4 count appear to show selection against Non- synonymous mutations in HIV-1 variants

Templeton, et al. Templeton’s article examined selection of 15 subjects similar to the Markham study, but further investigated a broader range of potential selective contexts. Overall disease progression categories: CD4 and CD8 T- Cell counts Mutations leading to amino acid substitutions were subject to positive selection over a broad range of clinical conditions in nonsyncytium inducing form (NSI) It was determined that the study must be focused in the V3 region

Templeton, et al. The results indicated that the V3 region is subject to selection, but cannot be summarized in a marginal fashion to a single variable such as CD4 counts. Diverse patterns of V3 region evolution depending on rapid and non-progressors. The V3 region is context dependent. Further studies of this region could be explored in the HIV-1 Genome.

References Markham RB, Wang WC, Weisstein AE, Wang Z, Munoz A, Templeton A, Margolick J, Vlahov D, Quinn T, Farzadegan H, and Yu XF. Patterns of HIV-1 evolution in individuals with differing rates of CD4 T cell decline. Proc Natl Acad Sci U S A 1998 Oct 13; 95(21) Templeton Alan R, Reichert R, Weisstein A, Xia-Fang Y, Markham RB. Selection in Context: Patterns of Natural Selection in the Glycoprotein 120 Region of Human Immunodeficiency Virus 1 Within Infected Individuals. Genetics Society of America USA 2004, Dec 20: