Fredrick A. Okoth

Identification of env CRF-10 among HIV Variants Circulating in Rural Western Kenya

As part of a program to determine the genetic diversity of human immunodeficiency virus in rural Kenya, we carried out a molecular analysis of the C2-V3 region of HIV-infected blood samples obtained from 30 antenatal clinic attendees of seven health centers in western Kenya. Direct sequencing was carried out on the envelope C2-V3 region of proviral DNA. On phylogenetic analysis with reference strains, 20 were subtype Al, 2 were subtype D, 1 was subtype C, 1 was subtype G, 1 was CRF-10, 2A/D, 2A/C, and 2 were unclassified. The presence of CRF-10 and the great variety of subtypes and recombinants in such a limited sample size suggest that western Kenya may be a potential hotspot for HIV recombination in the country.

Active Generation and Selection for HIV Intersubtype A/D Recombinant Forms in a Coinfected Patient in Kenya

To investigate the in vivo evolution of recombinant HIV, we followed up on a mother who was initially coinfected with subtypes A and D in Kenya. Blood samples were obtained in 1996 and 2002, and HIV pol and env genes were amplified by PCR, cloned, sequenced, and phylogenetically analyzed. As for the 1996 sample most of the clones generated from the pol and env genes clustered either with subtypes A and D reference strains. However, two clones from the pol gene were found to be independent recombinants between subtypes A and D by RIP analysis, suggesting active generation of recombinant forms. As for the 2002 sample, all the clones from the pol gene clustered only with the subtype A reference strain, while all the env clones clustered only with subtype D, denoting a dominance of an A/D recombinant form. These results indicate that in patients dually infected with subtypes A and D there is an ongoing generation and selection for A/D recombinant forms.

HIV Type 1 Subtype Surveillance in Central Kenya

Human immunodeficiency virus 1 (HIV-1) infection is characterized by genetic diversity such that specific viral subtypes are predominant in specific geographic areas. To determine circulating subtypes of HIV-1 in different parts of central Kenya, a cross-sectional study was carried out on HIV-1-positive blood samples collected from consenting individuals in eight hospitals of Kenyas central province. Proviral DNA was extracted from peripheral blood mononuclear cells. Polymerase chain reaction and direct sequencing using primers generated from a highly conserved region of HIV-1 env gp41 were carried out. Ninety-six samples were successfully amplified and sequenced. Analysis of the sequences showed that a majority of them belonged to subtype A1 (67/96, 69.8%), followed by subtypes D (18, 18.7%) and C (11/96, 11.5%). Consistent with findings in other parts of Kenya, HIV-1 subtype A1 was the most dominant virus in circulation. Continued surveillance of circulating subtypes of HIV-1 in Kenya is important in determining the evolution of the HIV/AIDS epidemic in Kenya.

HIV Type 1 Subtype Diversity and Drug Resistance among HIV Type 1-Infected Kenyan Patients Initiating Antiretroviral Therapy

The treatment of HIV-1 infection with antiretroviral drugs has greatly improved the survival of those who are infected. However, HIV-1 diversity and drug resistance are major challenges in patient management, especially in resource-poor countries. To evaluate HIV-1 genetic diversity and drug resistance-associated mutations among drug-naive patients in Kenya prior to antiretroviral therapy (ART), a genetic analysis of HIV-1 pol-RT and env-gp41 was performed on samples collected from 53 (18 males and 35 females) consenting patients between April and June 2005. The average age, baseline CD4(+) T cell counts, and viral loads were 38 (range, 24-62) years, 475 (range, 203-799) cells/mm(3), and 4.7 (range, 3.4-5.9) log(10) copies/ml, respectively. Phylogenetic analysis revealed that 40 samples (75.5%) were concordant subtypes for the two genes and 13 (24.5%) were discordant, suggesting possible recombination and/or dual infections. Prevalent subtypes included A1/A1(pol-RT/env-gp41), 31 (58.5%); D/D, 9 (16.9%); A1/C, 2 (3.8%); A1/D, 4 (7.5%); G/A1, 2 (3.8%); A1/A2, 1 (1.9%); C/A1, 2 (3.8%); D/A1, 1(1.9%); and D/A2, 1 (1.9%). Major reverse transcriptase inhibitor (RTI) resistance-associated mutations were found in four patients (7.5%). Of these patients, three had nucleoside RTI resistance mutations, such as M184V, K65R, D67N, K70R, and K219Q. Nonnucleoside RTI resistance-associated mutations K103N and Y181C were detected in three patients and one patient, respectively. Multiple drug resistance mutations were observed in this drug-naive population. With increasing numbers of patients that require treatment and the rapid upscaling of ART in Kenya, HIV-1 drug resistance testing is recommended before starting treatment in order to achieve better clinical outcomes.

HIV-1 subtype and viral tropism determination for evaluating antiretroviral therapy options: an analysis of archived Kenyan blood samples.

BackgroundInfection with HIV-1 is characterized by genetic diversity such that specific viral subtypes are predominant in specific geographical areas. The genetic variation in HIV-1 pol and env genes is responsible for rapid development of resistance to current drugs. This variation has influenced disease progression among the infected and necessitated the search for alternative drugs with novel targets. Though successfully used in developed countries, these novel drugs are still limited in resource-poor countries. The aim of this study was to determine HIV-1 subtypes, recombination, dual infections and viral tropism of HIV-1 among Kenyan patients prior to widespread use of antiretroviral drugs.MethodsRemnant blood samples from consenting sexually transmitted infection (STI) patients in Nairobi were collected between February and May 2001 and stored. Polymerase chain reaction and cloning of portions of HIV-1 gag, pol and env genes was carried out followed by automated DNA sequencing.ResultsTwenty HIV-1 positive samples (from 11 females and 9 males) were analyzed. The average age of males (32.5 years) and females (26.5 years) was significantly different (p value < 0.0001). Phylogenetic analysis revealed that 90% (18/20) were concordant HIV-1 subtypes: 12 were subtype A1; 2, A2; 3, D and 1, C. Two samples (10%) were discordant showing different subtypes in the three regions. Of 19 samples checked for co-receptor usage, 14 (73.7%) were chemokine co-receptor 5 (CCR5) variants while three (15.8%) were CXCR4 variants. Two had dual/mixed co-receptor use with X4 variants being minor population.ConclusionHIV-1 subtype A accounted for majority of the infections. Though perceived to be a high risk population, the prevalence of recombination in this sample was low with no dual infections detected. Genotypic co-receptor analysis showed that most patients harbored viruses that are predicted to use CCR5.

Genetic diversity of HIV type 1 along the coastal strip of Kenya.

A study on the genetic diversity of HIV-1 subtypes present along the coastal strip of Kenya, i.e., Kilifi, Mombasa, Msambweni, and Malindi districts, was carried out. DNA sequences for regions encoding a portion of the env-gp41 region of the virus were generated by PCR and sequenced directly. Eighty six samples that were successfully sequenced were analyzed. From the analysis, 86% (74) were subtype A1, 5% (4) were subtype C, 8% (7) were subtype D, and 1% (1) was subtype G. This study shows that HIV-1 subtype A1 is the most dominant subtype in circulation in this region.

The changing trend of HIV type 1 subtypes in Nairobi.

Monitoring the distribution of HIV-1 subtypes and recombinants among infected individuals has become a priority in HIV therapy. A laboratory analysis of samples collected from HIV-positive patients attending an STI clinic in Nairobi was done between March and May 2004. PCR was carried out on pol (intergrase) and env (C2V3) regions and resulting data on the 54 samples successfully analyzed revealed the following as circulating subtypes: 35/54(65%) were A1/A1, 5/54(9%) were A/C, 4/54 (7%) were A1/D, 1/54 (2%) was C/D, 1/54 (2%) was D/D, 1/54 (2%) was A1/A2, 1/54 (2%)was G/G, 1/54 (2%) was A2/D, 1/54 (2%) was C/C, and 4/54 (7%) were CRF02_ AG. The results show an increase in HIV-1 recombinants with the emergence of A1/A2 and an increase in CRF02_AG recombinants. Subtype diversity in the advent of ARV use will impact negatively on treatment outcomes. As such, increased viral evolution and recombination will call for continuous evaluation of available anti-HIV regimens for better management of those infected with HIV-1.

Prevalence of Nevirapine-Associated Resistance Mutations after Single Dose Prophylactic Treatment among Antenatal Clinic Attendees in North Rift Kenya

The use of single dose nevirapine to prevent mother-to-child transmission of HIV has been reported to induce drug-resistant mutations and reduce options for antiretroviral treatment for HIV-infected mothers and their children. To explore the status of nevirapine-resistant HIV genotypes in rural hospitals in the North Rift Valley Province of Kenya, samples collected 3 months after single dose nevirapine from 36 mothers and their children were analyzed. Resistance mutations were genotypically evaluated through proviral DNA amplification, cloning, and sequencing. Ten mothers (27.8%) had antiretroviral-associated resistance mutations of whom four (11.1%) had specific nevirapine (NNRTI) resistance-associated mutations. Three mothers (8.3%) transmitted the infection to their infants. This presence of nevirapine mutations in rural antenatal clinic attendees confirms the importance of integrating antiretroviral resistance monitoring as a key component in programs geared to prevention of HIV mother-to-child transmission.

HIV type 1 genetic diversity in Moyale, Mandera, and Turkana based on env-C2-V3 sequences.

The genetic diversity of HIV-1 subtypes circulating in three districts of northern Kenya, i.e., Turkana, Mandera, and Moyale, was studied. DNA sequences encoding a portion of the env-C2-V3 region of the virus were amplified by PCR and sequenced directly. One hundred and fifty-nine samples were successfully sequenced in the env-C2-V3 region and analyzed. From the analysis, 57% were subtype A1, 27% were subtype C, 9% were subtype D, and the remaining 7% were unclassified. This study showed that HIV-1 subtype A1 was the dominant subtype in circulation in this region, though there was a significant percentage of HIV-1 subtype C in circulation there.

Survey on prevalence and risk factors on HIV-1 among pregnant women in North-Rift, Kenya: a hospital based cross-sectional study conducted between 2005 and 2006.

BackgroundThe HIV/AIDS epidemic in Kenya is a major public-health problem. Estimating the prevalence of HIV in pregnant women provides essential information for an effective implementation of HIV/AIDS control measures and monitoring of HIV spread within a country. The objective of this study was to determine the prevalence of HIV infection, risk factors for HIV/AIDS and immunologic (lymphocyte profile) characteristics among pregnant women attending antenatal clinics in three district hospitals in North-Rift, Kenya.MethodsBlood samples were collected from pregnant women attending antenatal clinics in three district hospitals (Kitale, Kapsabet and Nandi Hills) after informed consent and pre-test counseling. The samples were tested for HIV antibodies as per the guidelines laid down by Ministry of Health, Kenya. A structured pretested questionnaire was used to obtain demographic data. Lymphocyte subset counts were quantified by standard flow cytometry.ResultsOf the 4638 pregnant women tested, 309 (6.7%) were HIV seropositive. The majority (85.1%) of the antenatal attendees did not know their HIV status prior to visiting the clinic for antenatal care. The highest proportion of HIV infected women was in the age group 21–25 years (35.5%). The 31–35 age group had the highest (8.5%) HIV prevalence, while women aged more than 35 years had the lowest (2.5%).Women in a polygamous relationship were significantly more likely to be HIV infected as compared to those in a monogamous relationship (p = 0.000). The highest HIV prevalence (6.3%) was recorded among antenatal attendees who had attended secondary schools followed by those with primary and tertiary level of education (6% and 5% respectively). However, there was no significant relationship between HIV seropositivity and the level of education (p = 0.653 and p = 0.469 for secondary and tertiary respectively). The mean CD4 count was 466 cells/mm3 (9–2000 cells/mm3). Those that had less than 200 cells/mm3 accounted for 14% and only nine were on antiretroviral therapy.ConclusionSeroprevalence of HIV was found to be consistent with the reports from the national HIV sentinel surveys. Enumeration of T-lymphocyte (CD4/8) should be carried out routinely in the antenatal clinics for proper timing of initiation of antiretroviral therapy among HIV infected pregnant women.