Molecular Characterization of HIV and Drug Resistance Mutations Among HIV Positive Children Attending Busia County Referral Hospital, Kenya

Abstract

Drug resistant Human Immunodeficiency Virus type 1 (HIV-1) variants are the main threat to current treatment programs. Overall, the emergence of HIV drug resistance is a complex and multifaceted problem, whose true extent has not been described in Kenya. Treatment failure is a major problem among those on antiretroviral therapy (ART) in Kenya. The resistance is associated with failure to adhere to therapy and low potency of some antiretroviral regimens. Monitoring HIV drug resistance is an important component of the World Health Organization’s (WHO) global HIV program. It is important to monitor for drug resistance in HIV positive children whose mothers have been on ART. The objectives of this study were to determine HIV drug resistant mutations and subtype diversity in HIV positive children born to HIV infected mothers attending Busia County Referral Hospital. To achieve this, archived plasma collected in 2011 from 65 HIV positive children aged between 6 weeks and 5 years were used. Nucleic acid (RNA) was extracted from plasma samples using the Qiagen RNA extraction kit according to the manufacturers’ instructions. One step reverse transcriptase-polymerase chain reaction (RT-PCR) and nested PCR were performed using primers targeting the protease and reverse transcriptase in the HIV-1 pol gene. Amplicons were further sequenced using the Big Dye chemistry, according to the manufacturer’s instructions. The Stanford University HIV drug resistance database and International AIDS Society (IAS) algorithm were used to determine the presence of drug resistant mutations. HIV subtype diversity was determined using Basic local alignment search tool (BLAST) as well as clustal W alignment and neighbor joining methods. The results showed that 16% (9/53) of the children had resistance mutations against Nucleoside reverse transcriptase inhibitor drugs and 24 % (13/53) had resistance mutations against Non-nucleoside reverse transcriptase inhibitor drugs. For protease inhibitor (PI) related mutations, minor mutations were found in 41.5 % (22/53) of the children. HIV-1 subtypes found included subtype A in 64 % (34/53) of the population, while 17 % (9/53) were subtype D and 5.7 % (3/53) were subtype C. Possible recombinants found included subtype A/C at 1.8 %(1/53), subtype A/D at 5.7 % (3/53), subtype K/A at 1.8 %(1/53) and subtype B/D 3.8 %(2/53). The observed drug resistant (DR) mutations call for continuous surveillance among children and establishment of HIV subtypes circulating among HIV infected mothers who are likely to transmit the virus to their infants.