Best of AIDS 2003: Epidemiology, Social, Cultural, and Political
Best of AIDS 2003: Epidemiology, Social, Cultural, and Political
Objective: To further define the genetic diversity of HIV-1 in Kenya using approaches that clearly distinguish subtypes from inter-subtype recombinants.
Design: Near full genome sequencing and analysis were used, including sensitive new tools for detection and mapping of recombinants.
Methods: Purified peripheral blood mononuclear cell DNA from 41 HIV-1 positive blood donations collected from six hospitals across southern Kenya was used to amplify near full-length genomes by nested PCR. These were sequenced on an ABI 3100 automated sequencer and analyzed phylogenetically.
Results: Among 41 near full-length genomes, 25 were non-recombinant (61%) and 16 were recombinant (39%). Of the 25 pure subtypes, 23 were subtype A, one was subtype C and one was subtype D. Most recombinants consisted of subtype A and either subtype C or subtype D; a few contained A2, a recently identified sub-subtype. Two A2/D recombinants had identical breakpoints and may represent a circulating recombinant form. A third A2/ D recombinant had the same structure as a previously described Korean isolate, and these may constitute a second A2-containing circulating recombinant form.
Conclusions: In Kenya, 93% of HIV-1 genomes were subtype A or A-containing recombinant strains. Almost 40% of all strains were recombinant. Vaccine candidates tested in Kenya should be based on subtype A strains, but the methods used for evaluation of breakthrough infections during future vaccine trials should be capable of identifying non-A subtypes, the A2 sub-subtype, and recombinants.
Next year 10,000 Kenyans will volunteer to take part in the final trial of an AIDS vaccine developed jointly in Kenya and the UK. The vaccine is based upon data gathered from years of follow-up on how the immune systems of prostitutes at Nairobi's Majengo slum were responding to the virus. The study will take 3 years to complete and if the results prove encouraging it will take another 2 years to license and bring the vaccine to mass production.
Determining the genetic make-up of circulating strains will be critical for this and other future vaccine work. Previously research based on partial sequences of the HIV genome showed that at least 70% of circulating strains in Kenya were subtype A, the rest were mostly subtype D with small percentages being subtypes C, G, or recombinant. In contrast, a recent study published in AIDS by the Walter Reed Army Institute of Research demonstrated that over 90% of HIV-1 fully sequenced genomes were actually subtype A or A-containing recombinant strains and almost half of all these strains were recombinant. "The great amount of genetic diversity that this paper documents makes it clear that this type of screening assay with the capacity to detect recombinants is critical", states Dr. Jean Carr one of the study's investigators.
The Walter Reed study was based upon 41 HIV-1 positive blood samples obtained from six hospitals across southern Kenya: Kericho, Kisii, and Kisumu District Hospitals, Coast General Hospital (Mombasa), Rift Valley Provincial Hospital (Nukuru) and Kenyatta National Hospital (Nairobi). Purified peripheral blood mononuclear cell DNA from these blood donations was then used to amplify near full-length genomes by nested PCR. These were then sequenced on a ABI 3100 automated sequencer and analysed phylogenetically.
The future aim of this team is to expand their work by fully sequencing peripheral blood mononuclear cells from 500 anonymous individuals from all over the country. They believe that vaccine candidates tested in Kenya should be based on subtype A strains, but the methods used for evaluation of breakthrough infections during vaccine trials, like the one occurring next year, should be capable of identifying non-A subtypes, the A2 sub-subtype, and recombinants. The hope is that their work can be used to quickly detect any future skewing of viral populations in Kenya that has occurred because of vaccine pressure.
(Enlarge Image)
Phylogenetic analysis of near full-length HIV-1 genomes of Kenyan blood bank samples. (a) Forty-one near full-length genome sequences were phylogenetically analyzed by neighbor-joining with reference strains from subtypes A-J, CRF01_AE and CRF02_AG. Recombinant or hypermutated strains are indicated. (b) Twenty-one near full-length genome sequences were phylogenetically analyzed by neighbor-joining followed by bootstrap analysis with reference strains from subtypes A-J, CRF01_AE and CRF02_AG. Only non-recombinant, non-hypermutated sequences were included on this tree. Bootstrap values . 70% were considered significant and included on the tree. The scale bars at the bottom of each figure show the branch length of 10% genetic distance.
(Enlarge Image)
Phylogenetic analysis of near full-length HIV-1 genomes of Kenyan blood bank samples. (a) Forty-one near full-length genome sequences were phylogenetically analyzed by neighbor-joining with reference strains from subtypes A-J, CRF01_AE and CRF02_AG. Recombinant or hypermutated strains are indicated. (b) Twenty-one near full-length genome sequences were phylogenetically analyzed by neighbor-joining followed by bootstrap analysis with reference strains from subtypes A-J, CRF01_AE and CRF02_AG. Only non-recombinant, non-hypermutated sequences were included on this tree. Bootstrap values . 70% were considered significant and included on the tree. The scale bars at the bottom of each figure show the branch length of 10% genetic distance.
Objective: To further define the genetic diversity of HIV-1 in Kenya using approaches that clearly distinguish subtypes from inter-subtype recombinants.
Design: Near full genome sequencing and analysis were used, including sensitive new tools for detection and mapping of recombinants.
Methods: Purified peripheral blood mononuclear cell DNA from 41 HIV-1 positive blood donations collected from six hospitals across southern Kenya was used to amplify near full-length genomes by nested PCR. These were sequenced on an ABI 3100 automated sequencer and analyzed phylogenetically.
Results: Among 41 near full-length genomes, 25 were non-recombinant (61%) and 16 were recombinant (39%). Of the 25 pure subtypes, 23 were subtype A, one was subtype C and one was subtype D. Most recombinants consisted of subtype A and either subtype C or subtype D; a few contained A2, a recently identified sub-subtype. Two A2/D recombinants had identical breakpoints and may represent a circulating recombinant form. A third A2/ D recombinant had the same structure as a previously described Korean isolate, and these may constitute a second A2-containing circulating recombinant form.
Conclusions: In Kenya, 93% of HIV-1 genomes were subtype A or A-containing recombinant strains. Almost 40% of all strains were recombinant. Vaccine candidates tested in Kenya should be based on subtype A strains, but the methods used for evaluation of breakthrough infections during future vaccine trials should be capable of identifying non-A subtypes, the A2 sub-subtype, and recombinants.
Next year 10,000 Kenyans will volunteer to take part in the final trial of an AIDS vaccine developed jointly in Kenya and the UK. The vaccine is based upon data gathered from years of follow-up on how the immune systems of prostitutes at Nairobi's Majengo slum were responding to the virus. The study will take 3 years to complete and if the results prove encouraging it will take another 2 years to license and bring the vaccine to mass production.
Determining the genetic make-up of circulating strains will be critical for this and other future vaccine work. Previously research based on partial sequences of the HIV genome showed that at least 70% of circulating strains in Kenya were subtype A, the rest were mostly subtype D with small percentages being subtypes C, G, or recombinant. In contrast, a recent study published in AIDS by the Walter Reed Army Institute of Research demonstrated that over 90% of HIV-1 fully sequenced genomes were actually subtype A or A-containing recombinant strains and almost half of all these strains were recombinant. "The great amount of genetic diversity that this paper documents makes it clear that this type of screening assay with the capacity to detect recombinants is critical", states Dr. Jean Carr one of the study's investigators.
The Walter Reed study was based upon 41 HIV-1 positive blood samples obtained from six hospitals across southern Kenya: Kericho, Kisii, and Kisumu District Hospitals, Coast General Hospital (Mombasa), Rift Valley Provincial Hospital (Nukuru) and Kenyatta National Hospital (Nairobi). Purified peripheral blood mononuclear cell DNA from these blood donations was then used to amplify near full-length genomes by nested PCR. These were then sequenced on a ABI 3100 automated sequencer and analysed phylogenetically.
The future aim of this team is to expand their work by fully sequencing peripheral blood mononuclear cells from 500 anonymous individuals from all over the country. They believe that vaccine candidates tested in Kenya should be based on subtype A strains, but the methods used for evaluation of breakthrough infections during vaccine trials, like the one occurring next year, should be capable of identifying non-A subtypes, the A2 sub-subtype, and recombinants. The hope is that their work can be used to quickly detect any future skewing of viral populations in Kenya that has occurred because of vaccine pressure.
(Enlarge Image)
Phylogenetic analysis of near full-length HIV-1 genomes of Kenyan blood bank samples. (a) Forty-one near full-length genome sequences were phylogenetically analyzed by neighbor-joining with reference strains from subtypes A-J, CRF01_AE and CRF02_AG. Recombinant or hypermutated strains are indicated. (b) Twenty-one near full-length genome sequences were phylogenetically analyzed by neighbor-joining followed by bootstrap analysis with reference strains from subtypes A-J, CRF01_AE and CRF02_AG. Only non-recombinant, non-hypermutated sequences were included on this tree. Bootstrap values . 70% were considered significant and included on the tree. The scale bars at the bottom of each figure show the branch length of 10% genetic distance.
(Enlarge Image)
Phylogenetic analysis of near full-length HIV-1 genomes of Kenyan blood bank samples. (a) Forty-one near full-length genome sequences were phylogenetically analyzed by neighbor-joining with reference strains from subtypes A-J, CRF01_AE and CRF02_AG. Recombinant or hypermutated strains are indicated. (b) Twenty-one near full-length genome sequences were phylogenetically analyzed by neighbor-joining followed by bootstrap analysis with reference strains from subtypes A-J, CRF01_AE and CRF02_AG. Only non-recombinant, non-hypermutated sequences were included on this tree. Bootstrap values . 70% were considered significant and included on the tree. The scale bars at the bottom of each figure show the branch length of 10% genetic distance.
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