Rosemary A. Audu

Management of HIV-1 infection with a combination of nevirapine, stavudine, and lamivudine: a preliminary report on the Nigerian antiretroviral program.

Objective:To evaluate treatment outcome in the first 12 months among HIV-positive patients managed with a combination of nevirapine + stavudine + lamivudine under the current national antiretroviral (ARV) program in Nigeria. Design:This was a prospective observational, cohort study on 50 ARV-naive patients who met the inclusion criteria for the program and had given informed consent. All patients were in stage 2 or stage 3 periods of infection based on World Health Organization clinical classification. The patients were treated with the generic brands of ARVs and treatment consisted of oral nevirapine (Nevimal, Cipla, Mumbai, India), 200 mg daily, lamivudine (Lamivir, Cipla), 150 mg twice daily, and stavudine (Stavir, Cipla), 40 mg twice daily. Prior to initiation of treatment, the clinical history and baseline data for each patient were documented. The levels of plasma HIV-1 RNA, CD4+ cell counts, frequency of opportunistic infections, and estimated body mass index were recorded at baseline and subsequently at intervals during treatment. Data obtained at the various sampling times for each parameter were compared against their baseline values. Results:Data on the plasma HIV-1 RNA levels indicated that between baseline and week 24, the median viral load of the patients decreased by 1.79 log10 copies/mL. Equally between baseline and week 48 the median CD4+ cell counts increased by 186 × 106 cells/L, the frequency of opportunistic infections decreased by 82%, the median body mass index increased by 4.8 kg/m2, and 36% experienced side effects, which were minor and transient. The most prevalent side effect recorded was skin rash associated with nevirapine. Good adherence to this triple regimen was recorded in >85% of the patients. Conclusions:The overall results within the 12-month treatment period indicated an effective suppression of viral replication, the reconstitution of the immune system, and improvement of the physical well-being of the study population. Though there may be differences in global distribution of the infecting HIV-1 subtypes, the clinical and biologic results of this study compared favorably to those documented in cohorts treated with branded and generic ARV drugs in some developed and developing countries. The cumulative data in this study further confirmed that the correct use of generic brands of ARVs is a feasible option in HIV care and support programs in resource-poor countries.

Genetic studies in the Nigerian population implicate an MSX1 mutation in complex oral facial clefting disorders.

Background Orofacial clefts are the most common malformations of the head and neck, with a worldwide prevalence of 1 in 700 births. They are commonly divided into CL(P) and CP based on anatomic, genetic, and embryologic findings. A Nigerian craniofacial anomalies study (NigeriaCRAN) was set up in 2006 to investigate the role of gene-environment interaction in the origin of orofacial clefts in Nigeria. Subjects and Methods DNA isolated from saliva from Nigerian probands was used for genotype association studies and direct sequencing of cleft candidate genes: MSX1, IRF6, FOXE1, FGFR1, FGFR2, BMP4, MAFB, ABCA4, PAX7, and VAX1, and the chromosome 8q region. Results A missense mutation A34G in MSX1 was observed in nine cases and four HapMap controls. No other apparent causative variations were identified. Deviation from Hardy Weinberg equilibrium (HWE) was observed in these cases (p = .00002). A significant difference was noted between the affected side for unilateral CL (p = .03) and bilateral clefts and between clefts on either side (p = .02). A significant gender difference was also observed for CP (p = .008). Conclusions Replication of a mutation previously implicated in other populations suggests a role for the MSX1 A34G variant in the development of CL(P).

Accumulation of Protease Mutations among Patients Failing Second-Line Antiretroviral Therapy and Response to Salvage Therapy in Nigeria

Background To date, antiretroviral therapy (ART) guidelines and programs in resource-limited settings (RLS) have focused on 1st- and 2nd-line (2 L) therapy. As programs approach a decade of implementation, policy regarding access to 3rd-line (3 L) ART is needed. We aimed to examine the impact of maintaining patients on failing 2 L ART on the accumulation of protease (PR) mutations. Methods and Findings From 2004–2011, the Harvard/APIN PEPFAR Program provided ART to >100,000 people in Nigeria. Genotypic resistance testing was performed on a subset of patients experiencing 2 L failure, defined as 2 consecutive viral loads (VL)>1000 copies/mL after ≥6 months on 2 L. Of 6714 patients who received protease inhibitor (PI)-based ART, 673 (10.0%) met virologic failure criteria. Genotypes were performed on 61 samples. Patients on non-suppressive 2 L therapy for <12 months prior to genotyping had a median of 2 (IQR: 0–5) International AIDS Society (IAS) PR mutations compared with 5 (IQR: 0–6) among patients failing for >24 months. Patients developed a median of 0.6 (IQR: 0–1.4) IAS PR mutations per 6 months on failing 2 L therapy. In 38% of failing patients no PR mutations were present. For patients failing >24 months, high- or intermediate-level resistance to lopinavir and atazanavir was present in 63%, with 5% to darunavir. Conclusions This is the first report assessing the impact of duration of non-suppressive 2 L therapy on the accumulation of PR resistance in a RLS. This information provides insight into the resistance cost of failing to switch non-suppressive 2 L regimens and highlights the issue of 3 L access.

Effect of HIV‐1 infection and increasing immunosuppression on menstrual function

Aim:  The aim of this study was to determine the prevalence, pattern and determinants of menstrual abnormalities in HIV‐positive Nigerian women.

Improving quality in national reference laboratories: The role of SLMTA and mentorship

Background The Nigerian Institute of Medical Research houses two reference laboratories: the virology and tuberculosis laboratories. Both were enrolled in the Strengthening Laboratory Management Toward Accreditation (SLMTA) programme. Objective To describe the impact of SLMTA and discuss factors affecting the results, with an emphasis on mentorship. Methods The SLMTA programme was implemented from April 2010 through November 2012. Participants attended three workshops and executed quality improvement projects; laboratory auditors evaluated performance using a standard checklist. The virology laboratory did not receive mentorship; however, the tuberculosis laboratory had an international mentor who visited the laboratory four times during the programme, spending two to four weeks embedded within the laboratory during each visit. Results There was an overall improvement in the performance of both laboratories, with the virology laboratory increasing 13% (from 80% at baseline to 93% at exit audit) and the tuberculosis laboratory increasing 29% (from 66% to 95%). These scores were maintained nine months later at the surveillance audit. Conclusion The SLMTA programme resulted in improved and sustained quality management performance for both laboratories. Mentoring was a possible factor in the substantial improvement made by the tuberculosis laboratory and should be considered in order to augment the training received from the SLMTA workshops.

Prevalence and risk factors of asymptomatic bacteriuria among pregnant Nigerians infected with HIV

Objective: There are conflicting report on the association of HIV infection and asymptomatic bacteriuria (ASB). Most of these studies were from areas with low HIV burden. This study determined the prevalence and risk factors of ASB in HIV positive pregnant women. Methods: A cross sectional study among HIV positive pregnant women seen at a large PMTCT clinic in Lagos Nigeria. The women were evaluated for ASB at first clinic attendance. Blood samples were also collected for viral load, CD4 count and hemoglobin levels assessment. Data were managed with SPSS for windows version 19. Results: 102 (18.1%) women out of 563 studied were found positive for asymptomatic bacteriuria. Ninety-seven (95.1%) of the positive samples yielded single bacterial isolates. Escherichia coli (44.3%) and Proteus mirabilis (21.6%) were the most common bacterial isolates. Previous urinary tract infection (OR: 4.3), HIV-1 RNA greater than 10,000 copies/ml (OR: 3.9), CD4 count <200 cells/mm3 (OR: 1.4) and maternal hemoglobin <11 g/dl (OR: 1.4) were factors significantly associated with ASB after controlling for possible confounders. Conclusion: ASB is common in HIV positive pregnant women in our environment and is associated with previous UTI, high viral load, low CD4 count and maternal hemoglobin <11 g/dl.

Experience of quality management system in a clinical laboratory in Nigeria

Issues Quality-management systems (QMS) are uncommon in clinical laboratories in Nigeria, and until recently, none of the nation’s 5 349 clinical laboratories have been able to attain the certifications necessary to begin the process of attaining international accreditation. Nigeria’s Human Virology Laboratory (HVL), however, began implementation of a QMS in 2006, and in 2008 it was determined that the laboratory conformed to the requirements of ISO 9001:2000 (now 2008), making it the first diagnostic laboratory to be certified in Nigeria. The HVL has now applied for the World Health Organization (WHO) accreditation preparedness scheme. The experience of the QMS implementation process and the lessons learned therein are shared here. Description In 2005, two personnel from the HVL spent time studying quality systems in a certified clinical laboratory in Dakar, Senegal. Following this peer-to-peer technical assistance, several training sessions were undertaken by HVL staff, a baseline assessment was conducted, and processes were established. The HVL has monitored its quality indicators and conducted internal and external audits; these analyses (from 2007 to 2009) are presented herein. Lessons learned Although there was improvement in the pre-analytical and analytical indicators analysed and although data-entry errors decreased in the post-analytical process, the delay in returning laboratory test results increased significantly. There were several factors identified as causes for this delay and all of these have now been addressed except for an identified need for automation of some high-volume assays (currently being negotiated). Internal and external audits showed a trend of increasing non-conformities which could be the result of personnel simply becoming lax over time. Application for laboratory accreditation, however, could provide the renewed vigour needed to correct these non-conformities. Recommendation This experience shows that sustainability of the QMS at present is a cause for concern. However, the tiered system of accreditation being developed by WHO–Afro may act as a driving force to preserve the spirit of continual improvement.

Loss-of-Function GRHL3 Variants Detected in African Patients with Isolated Cleft Palate

In contrast to the progress that has been made toward understanding the genetic etiology of cleft lip with or without cleft palate, relatively little is known about the genetic etiology for cleft palate only (CPO). A common coding variant of grainyhead like transcription factor 3 (GRHL3) was recently shown to be associated with risk for CPO in Europeans. Mutations in this gene were also reported in families with Van der Woude syndrome. To identify rare mutations in GRHL3 that might explain the missing heritability for CPO, we sequenced GRHL3 in cases of CPO from Africa. We recruited participants from Ghana, Ethiopia, and Nigeria. This cohort included case-parent trios, cases and other family members, as well as controls. We sequenced exons of this gene in DNA from a total of 134 nonsyndromic cases. When possible, we sequenced them in parents to identify de novo mutations. Five novel mutations were identified: 2 missense (c.497C>A; p.Pro166His and c.1229A>G; p.Asp410Gly), 1 splice site (c.1282A>C p.Ser428Arg), 1 frameshift (c.470delC; p.Gly158Alafster55), and 1 nonsense (c.1677C>A; p.Tyr559Ter). These mutations were absent from 270 sequenced controls and from all public exome and whole genome databases, including the 1000 Genomes database (which includes data from Africa). However, 4 of the 5 mutations were present in unaffected mothers, indicating that their penetrance is incomplete. Interestingly, 1 mutation damaged a predicted sumoylation site, and another disrupted a predicted CK1 phosphorylation site. Overexpression assays in zebrafish and reporter assays in vitro indicated that 4 variants were functionally null or hypomorphic, while 1 was dominant negative. This study provides evidence that, as in Caucasian populations, mutations in GRHL3 contribute to the risk of nonsyndromic CPO in the African population.

Development and Implementation Challenges of a Quality Assured HIV Infant Diagnosis Program in Nigeria Using Dried Blood Spots and DNA Polymerase Chain Reaction

Nigeria has one of the highest HIV burdens as well as mother-to-infant transmission rates in the world. A pilot program using polymerase chain reaction (PCR)-based testing of dried blood spot (DBS) specimens was implemented to enable early identification of HIV-infected infants and timely referral and linkage to care. From February 2007 to October 2008, whole blood was collected by finger prick to prepare DBS from infants <18 months presenting in six public mother-and-child health facilities in Lagos, Nigeria. The DBS were tested using the Roche Amplicor HIV-1 DNA Test, v1.5. To monitor laboratory testing quality, all of the PCR-positive and 10% of the PCR-negative DBS were retested by the same method at another reference laboratory. Three hundred and sixty-five randomly selected infants were screened using HIV rapid tests (RT) according to the national algorithm and RT-negative and PCR-positive specimens were also tested using Genscreen enzyme-linked immunosorbent assay (EIA) (Bio-Rad, France). The turnaround time (TAT) from sample collection, testing, and dispatching of results from each health facility was monitored. A total of 1,273 infants with a median age of 12.6 weeks (1 day to 71.6 weeks) participated in the program and 280 (22.0%) were PCR positive. HIV transmission levels varied greatly in the different health facilities ranging from 7.1% to 38.4%. Infants aged 48 to 72 weeks had the highest level of PCR positivity (41.1%). All PCR-positive specimens were confirmed by retesting. The mean turnaround time from DBS collection to returning of the laboratory result to the health facilities was 25 days. Three infants were found to be HIV antibody negative by rapid tests but were positive by both PCR and the fourth generation EIA. The DBS-based PCR program accurately identified all of the HIV-infected infants. However, many programmatic challenges related to the laboratory and TAT were identified.

Proficiency testing for HIV, tuberculosis and malaria diagnosis in clinical laboratories in Nigeria

Background Proficiency testing (PT) is a means of verifying the reliability of laboratory results, but such programmes are not readily available to laboratories in developing countries. This project provided PT to laboratories in Nigeria. Objectives To assess the proficiency of laboratories in the diagnosis of HIV, tuberculosis and malaria. Methods This was a prospective study carried out between 2009 and 2011. A structured questionnaire was administered to 106 randomly-selected laboratories. Forty-four indicated their interest in participation and were enrolled. Four rounds of pre-characterised plasma panels for HIV, sputum films for tuberculosis and blood films for malaria were distributed quarterly by courier over the course of one year. The results were returned within two weeks and scores of ≥ 80% were reported as satisfactory. Mentoring was offered after the first and second PT rounds. Results Average HIV PT scores increased from 74% to 95% from the first round to the third round, but decreased in the fourth round. For diagnosis of tuberculosis, average scores increased from 42% in the first round to 78% in the second round; but a decrease to 34% was observed in the fourth round. Malaria PT performance was 2% at first, but average scores increased between the second and fourth rounds, culminating in a fourth-round score of 39%. Many participants requested training and mentoring. Conclusions There were gross deficiencies in the quality of laboratory services rendered across Nigeria. In-country PT programmes, implemented in conjunction with mentoring, will improve coverage and diagnosis of HIV, tuberculosis and malaria.