Angela A. Garibaldi Week 3

Prior knowledge of the HIV virus:

• Currently I don't know very much about the biological aspects of the HIV virus. I only know that it makes the immune system very weak and is a RNA retro virus.
• The following are questions about HIV I would like answered:
  • 1. What is the overall molecular mechanism of the disease?
    2. What type of virus is it specifically and what does this imply?
    3. What are the mechanisms of some of the methods of treatment currently used?

"Becoming an Expert in Pubmed"

Note: Version of Pubmed used is The following are the searching processes used to find review articles relevant to answering the above questions:

1. My first search query was: HIV [TI] evolution Limits:links to free full text, abstracts review, english.

• [TI] specifies that only papers with "HIV" in the title of any papers.

• I've highlighted the PubMed ID number of the article that I chose. This is what is used to complete the bibliography code and used to easily find the paper again.

• The following review seemed most relevant and clear as an introductory paper

1. Cohen MS, Hellmann N, Levy JA, DeCock K, and Lange J. The spread, treatment, and prevention of HIV-1: evolution of a global pandemic. J Clin Invest. 2008 Apr;118(4):1244-54. DOI:10.1172/JCI34706 |

Further Articles for Investigation:

1. Williams KC and Burdo TH. HIV and SIV infection: the role of cellular restriction and immune responses in viral replication and pathogenesis. APMIS. 2009 May;117(5-6):400-12. DOI:10.1111/j.1600-0463.2009.02450.x |

Markham Article

10 Biological Terms

1. seroconverting- serotype- The genotype of a unicellular organism as defined by antisera against antigenic determinants expressed on the surface. Therefore; the conversion of the serotype.Dictionary of Cell and Molecular Biology
2. non-synonymous mutation- a.k.a. a missense mutation -A form of point mutation resulting in a codon that codes for a different amino acid, and thus, causes the synthesis of a protein with an altered amino acid sequence during translation. biology-online.org dictionary
3. CD4 T-Cells- A form of T lymphocyte with CD4 receptor on the cell surface that recognizes antigens of a virus-infected cell. This type of T lymphocyte releases lymphokines upon stimulation of its recognition of antigens of a virus-infected cell. Upon its antigen recognition it is activated, stimulating, in turn, B lymphocytes and killer T lymphocytes during the antibody formation.It is the target cell of AIDS virus, infecting and eventually killing the lymphocyte by the virus. biology-online.org dictionary
4. CD4- A 55-kD glycoprotein that serves as differentiation antigen found on the surface of T lymphocytes and macrophages. The CD in CD4 is an abbreviation for cluster of differentiation. It belongs to the immunoglobulin supergene family. The presence of CD4 characterizes the helper/inducer cell. It also serves as HIV receptors where the virus binds directly with its envelope protein gp120. biology-online.org dictionary
5. Nested PCR -Variety of polymerase chain reaction, in which specificity is improved by using two sets of primers sequentially. An initial PCR is performed with the ‘outer’ primer pairs, then a small aliquot is used as a template for a second round of PCR with the ‘inner’ primer pair. Dictionary of Cell and Molecular Biology
6. PBMC (peripheral blood mononuclear cells) - A mixture of monocytes and lymphocytes; blood leucocytes from which granulocytes have been separated and removed.Dictionary of Cell and Molecular Biology

Outline of the Article:

INTRODUCTION:

1. stable host environment -breeds best adapted viruses to dominate and all mutations after are not represented in the gene pool as much.
2. Different viruses will be similar in genetic makeup with few variants because mutations will be more neutral
3. Unstable host environment - caused by the host's immune response or specifically in HIV differential display of co-receptors.
4. If destabilizing force selected strongly against the variants, the gene pool of the viruses that were most dominant to begin with will survive best.
5. (Frequency dependent selection)If destabilizing force selected strongly against the virus variant that is most abundant, the viral load will decrease overall. Genetic diversity will maintain due to the gene pool of the viral variants in fewer numbers due to their different variants.
6. These remaining viral variants will propagate and lead to variants that can surpass the selective power (such as the specific immune system response )
7. This experiment analyzes the sequence patters of the viral variants using a wider scope of time points than previous studies.

• • ( Analysis has shown divergent patterns between nonprogressor patients and rapid progression patients. Higher genetic diversity patterns in viral variants point to faster CD4 T Cell decline.)

METHODS

1. 15 patients checked in 6 month intervals: Rapid progressors (<200 CD4T-cell count) Moderate (200-650) NonProgressors (>650)
2. Sequence HIV-1 env genes by amplifying a region of the env gene with Nested PCR from the PBMCs, cloned into pUC19, sequenced with Sanger chain termination method. Used only samples that could be detected at lowest dilution.
3. Significant because viral DNA is mostly acquired from these unactivated host (due to recent infection) PBMCs and can only last a few days in unactivated Tcells. (In other words they won't have time to change significantly) Therefore, the DNA in these cells should be very similar to the RNA of the viruses circulating in the plasma at the time.
4. Used the single round PCR samples that could be detected at the lowest dilution in the second round PCRs in order to specify clones that had unique viral genome template.
5. used reverse transcription-PCR to measure plasma viral loads
6. 'Correlation analysis was used, comparing individuals with a different status (non, moderate, rapid) at similar time points over the course of a year in order to determine the correlation between genetic diversity or mutational divergence via CD4 T-cell count and sequencing results.
7. Phylogenetic trees have taxa color coded based on time point. red=V1,orange=V2, green=V3, light blue=V4, dark blue= V5, purple=V6, brown=V7, gray=V8
8. dS/dN Ratios- determined a baseline sequence for each individual and then compared it with observed strains over time. Differences between the base and the subsequent strain was labeled either synonymous or nonsynonymous.
9. dS,dN and dS/dN ratios were averaged over all strains across the board for each visit.