The prevalence of hepatitis b virus infection in HIV-Positive and HIV-Negative infants: KwaZulu-Natal, South Africa

Background: The prevalence of hepatitis B virus (HBV) amongst South African infants and children has been reported in the pre-HIV era. Despite the reported high prevalence of HIV in the general population of South Africa; the rate of HIV/HBV co-infection amongst infants and children remains poorly reported.Objectives: We describe the prevalence of HBV infection amongst HIV-positive and HIV-negative infants by molecular methods of diagnosis using dried blood spot samples.Methods: This retrospective cross-sectional study was conducted between July 2011 and December 2011 in an academic referral laboratory offering viral diagnostic services to the entire KwaZulu-Natal province of South Africa. A total of 322 study samples were collected from discarded residual dried blood spot samples following routine infant diagnosis of HIV. Equal proportions of HIV-positive and HIV-negative infant specimens were studied. Statistical differences in the prevalence of HBV between the HIV-positive and HIV-negative samples were calculated using the Pearson chi-square test; and a p-value 0.05 was considered statistically significant. Further testing for HBV DNA using a nested polymerase chain reaction method was performed.Results: The overall prevalence of HBV was 10%. In the HIV-positive group; 21 of 161 infants tested positive for HBV compared with 12 of 161 HIV-negative infants who tested positive for HBV. The proportion of infants infected with HBV was marginally higher amongst HIV positiveinfants (13.0%; 95% CI 6.8-19.9) compared with HIV-negative infants (7.5%; 95% C I2.5-13.7; P

Real-time polymerase chain reaction optimised for hepatitis c virus detection in dried blood spots from HIV-Exposed infants, KwaZulu-Natal, South Africa

Background: There is a paucity of data on the prevalence of hepatitis C virus (HCV) in children, particularly in sub-Saharan Africa. A major obstacle in resource-limited settings for polymerase chain reaction (PCR) testing is the necessity for specimen transportation and storage at low temperatures. There are numerous recent studies of using real-time HCV PCR for diagnosis and screening of plasma and serum, but few have looked at using dried blood spot (DBS)specimens. Objectives: The aim of this study was to optimise a real-time HCV PCR method to detect HCV RNA from infant DBS specimens for use as a tool for HCV surveillance in KwaZulu-Natal, South Africa. Method: The LightCycler® 2.0 instrument was used for the HCV PCR using the LightCycler® RNA Master SYBR Green I kit. Template volume, primer concentration and primer annealing temperatures were optimised and the method was used on 179 DBS specimens from HIV-exposed infants in KwaZulu-Natal. Results: Primer concentrations adjusted to 0.25 µM and a template volume of 10 µL improved the PCR amplification. Primer annealing temperatures lowered from 65°C to 58°C resulted in higher quantities of amplified PCR product. The limit of detection of the optimised HCV PCR assay was between 1200 IU/mL and 3580 IU/mL of HCV RNA. HCV was not detected in any of the 179 DBS specimens.Conclusion: The optimised real-time HCV PCR on infant DBS specimens performed well, but HCV was not found in this surveillance study. HIV infection may have little impact on the vertical transmission of HCV in this region

Dried blood spot specimen quality and validation of a new pre-analytical processing method for qualitative HIV-1 PCR, KwaZulu-Natal, South Africa

Background: Poor quality dried blood spot (DBS) specimens are usually rejected by virology laboratories; affecting early infant diagnosis of HIV. The practice of combining two incompletely-filled DBS in one specimen preparation tube during pre-analytical specimen processing (i.e.; the two-spot method) has been implemented to reduce the number of specimens being rejected for insufficient volume.Objectives: This study analysed laboratory data to describe the quality of DBS specimens and the use of the two-spot method over a one-year period; then validated the two-spot method against the standard (one-spot) method.Methods: Data on HIV-1 PCR test requests submitted in 2014 to the Department of Virology at Inkosi Albert Luthuli Central Hospital in KwaZulu-Natal province; South Africa were analysed to describe reasons for specimen rejection; as well as results of the two-spot method. The accuracy; lower limit of detection and precision of the two-spot method were assessed.Results: Of the 88 481 specimens received; 3.7% were rejected for pre-analytical problems. Of those; 48.9% were rejected as a result of insufficient specimen volume. Two health facilities had significantly more specimen rejections than other facilities. The two-spot method prevented 10 504 specimen rejections. The Pearson correlation coefficient comparing the standard to the two-spot method was 0.997.Conclusions: The two-spot method was comparable with the standard method of pre-analytical specimen processing. Two health facilities were identified for targeted retraining on specimen quality. The two-spot method of DBS specimen processing can be used as an adjunct to retraining; to reduce the number of specimens rejected and improve linkage to care