Temporal and spatial analysis of Plasmodium falciparum genomics reveals patterns of parasite connectivity in a low-transmission district in Southern Province, Zambia.

BACKGROUND: Understanding temporal and spatial dynamics of malaria transmission will help to inform effective interventions and strategies in regions approaching elimination. Parasite genomics are increasingly used to monitor epidemiologic trends, including assessing residual transmission across seasons and importation of malaria into these regions. METHODS: In a low and seasonal transmission setting of southern Zambia, a total of 441 Plasmodium falciparum samples collected from 8 neighbouring health centres between 2012 and 2018 were genotyped using molecular inversion probes (MIPs n = 1793) targeting a total of 1832 neutral and geographically informative SNPs distributed across the parasite genome. After filtering for quality and missingness, 302 samples and 1410 SNPs were retained and used for downstream population genomic analyses. RESULTS: The analyses revealed most (67%, n = 202) infections harboured one clone (monogenomic) with some variation at local level suggesting low, but heterogenous malaria transmission. Relatedness identity-by-descent (IBD) analysis revealed variable distribution of IBD segments across the genome and 6% of pairs were highly-related (IBD ≥ 0.25). Some of the highly-related parasite populations persisted across multiple seasons, suggesting that persistence of malaria in this low-transmission region is fueled by parasites "seeding" across the dry season. For recent years, clusters of clonal parasites were identified that were dissimilar to the general parasite population, suggesting parasite populations were increasingly fragmented at small spatial scales due to intensified control efforts. Clustering analysis using PCA and t-SNE showed a lack of substantial parasite population structure. CONCLUSION: Leveraging both genomic and epidemiological data provided comprehensive picture of fluctuations in parasite populations in this pre-elimination setting of southern Zambia over 7 years.

Cognitive and academic performance of rural Zambian youth exposed to HIV.

Studies focusing on children affected by HIV have shown that they have generally lower academic performance, however, few studies separate children who are HIV exposed and infected (CHEI) and those who are HIV exposed but uninfected (CHEU). Importantly, in rural sub-Saharan Africa, the majority of studies on CHEI and CHEU examine academic performance indirectly based on cognitive test scores. Therefore, studies assessing the effects of HIV on academic achievement directly for CHEI and CHEU are needed. This article evaluates the effects of HIV-infection on cognitive and academic performance by comparing CHEI (n = 82) and CHEU (n = 1045) aged 7-17 years old using cross-sectional data from an ongoing longitudinal study in a rural area of Zambia. Youth completed cognitive and academic assessments; their height and weight were assessed to generate Body Mass Index (BMI). Caregiver questionnaires provided information on youths' years in school and household socio-economic status (SES). Results indicated that while HIV infection status did explain some of the variance in performance between CHEI and CHEU, age, BMI, years of schooling and SES accounted for additional variance. The effect of years of schooling on both cognitive and academic performance demonstrated that CHEI's performance may be greatly improved by consistent school enrollment.

Pediatric Symptom Checklist-17: Factor Structure and Uniform Differential Item Functioning Across Gender and Age in HIV Orphans and Vulnerable Children in Zambia.

This study aimed to investigate the psychometric properties of the Pediatric Symptom Checklist-17 (PSC-17) in a sample of children orphaned or made vulnerable (OVC) by HIV in Zambia. Caregivers of 1,076 OVC (55.1% boys; Mage = 12.91 years) completed the PSC-17. Competing models, including confirmatory factor analysis (CFA), hierarchical CFA, bifactor CFA, exploratory structural equation modeling (ESEM), and bifactor ESEM, were tested to evaluate the optimal factor structure of the PSC-17. Results showed that the bifactor ESEM provided the best approximation of the PSC-17 data with a well-defined general psychosocial problems factor explaining 72% of the reliable variance in the total score and an internalizing factor containing 63% of reliable variance unique from the general factor. The observed overall psychosocial problems score was associated with lower academic achievement and working memory (with small effect sizes), supporting the discriminant validity of score interpretation. Results of multiple indicators multiple causes (MIMIC) analyses revealed that all items functioned equivalently across child gender and age.

Factors associated with antiretroviral therapy use among pregnant women in rural and urban settings in Southern Province, Zambia: 2016-2019.

OBJECTIVES: To assess antiretroviral therapy (ART) coverage among pregnant women living with HIV and compare the characteristics of women who received and did not receive ART during pregnancy in Zambia. METHODS: A cross-sectional study was conducted at urban and rural health facilities in Southern Province, Zambia, from 2016 to 2019. Pregnant women living with HIV delivering at study sites were enrolled and administered a questionnaire, and the results of infant diagnostic testing for HIV at birth was documented. RESULTS: About 1184 mother/infant pairs were enrolled. ART coverage was 93.7%. Most women who did not receive ART during pregnancy reported HIV diagnosis at delivery (18.0%) or during pregnancy (57.7%). The primary reported reason for not receiving ART was not wanting to take the drugs. Women who did not receive ART during pregnancy were significantly younger, less likely to have disclosed their HIV-infection status to others, and less likely to have received antenatal care than women who received ART. ART use correlated with higher levels of education in urban but not rural sites. Overall, 1.0% of infants were infected with HIV at birth, including 0.8% of infants born to women who received ART and 4.1% of infants born to women who did not. CONCLUSIONS: Most women received ART according to guidelines, resulting in low perinatal transmission rates of HIV to infants. Efforts to increase ART coverage and prevent vertical transmission should focus on identifying incident HIV infections during pregnancy and strengthening counselling for newly diagnosed pregnant women.

Respiratory viruses in rural Zambia before and during the COVID-19 pandemic.

OBJECTIVES: With the emergence of the COVID-19 pandemic, restrictions were implemented globally to control the virus. Data on respiratory pathogens in sub-Saharan Africa during the COVID-19 pandemic are scarce. This analysis was conducted to evaluate patterns of respiratory pathogens in rural Zambia before and during the first year of the pandemic. METHODS: Surveillance was established in December 2018 at Macha Hospital in southern Zambia. Patients with respiratory symptoms in the outpatient and inpatient clinics were recruited. Nasopharyngeal samples were collected and tested for respiratory pathogens. The prevalence of respiratory symptoms and pathogens was evaluated and compared in the first (December 10, 2018-December 9, 2019) and second (December 10, 2019-November 30, 2020) years of surveillance. RESULTS: Outpatient visits and admissions for respiratory illness significantly decreased from the first to second year, especially among children. SARS-CoV-2 was not detected from any participants in Year 2. Among outpatients and inpatients with respiratory symptoms, the prevalence of respiratory syncytial virus and influenza viruses decreased from the first to second year. In contrast, the prevalence of rhinovirus/enterovirus, metapneumovirus and parainfluenza virus increased. CONCLUSIONS: The epidemiology of respiratory viruses in rural Zambia changed during the first year of the COVID-19 pandemic, suggesting that public health interventions may have had an impact on the introduction and circulation of respiratory pathogens in this area.

The NSEBA Demonstration Project: implementation of a point-of-care platform for early infant diagnosis of HIV in rural Zambia.

OBJECTIVES: To describe the experience and resource requirements of implementing point-of-care testing for early infant diagnosis of HIV in rural Zambia. METHODS: A demonstration project was conducted using a hub-and-spoke model in 2018-2019 at five clinics in rural Zambia. Two testing hubs were established, and all HIV-exposed infants were tested with the GeneXpert system. Data on costs, turnaround times and test results were collected. RESULTS: Seven hundred and eighty six tests were conducted. At the hubs, results were available a median of 2.4 (IQR: 2.1, 2.8) hours after sample collection and most mothers (84%) received same-day results. At the spoke facilities, results were available a median of 9 days (IQR: 7, 12) after sample collection and provided to the mother a median of 16 days (IQR: 10, 28) after sample collection. Eleven children tested positive, and 9 (82%) started treatment a median of 13 days (IQR: 7, 21) after sample collection and on the day mothers received results. In contrast, results from matching samples sent for routine testing were available a median of 38 days (IQR: 27, 61) after sample collection and provided to the mother a median of 91 days (IQR: 47, 135) after sample collection. CONCLUSIONS: Implementing point-of-care testing in a network of rural health centres in Zambia required significant initial and ongoing investment in infrastructure, training and supervision. However, point-of-care testing can rapidly diagnose HIV-infected infants, so they can benefit from early treatment.

Facility-based surveillance for influenza and respiratory syncytial virus in rural Zambia.

BACKGROUND: While southern Africa experiences among the highest mortality rates from respiratory infections, the burden of influenza and respiratory syncytial virus (RSV) in rural areas is poorly understood. METHODS: We implemented facility-based surveillance in Macha, Zambia. Outpatients and inpatients presenting with influenza-like illness (ILI) underwent testing for influenza A, influenza B, and RSV and were prospectively followed for 3 to 5 weeks to assess clinical course. Log-binomial models assessed correlates of infection and clinical severity. RESULTS: Between December 2018 and December 2019, 17% of all outpatients presented with ILI and 16% of inpatients were admitted with an acute respiratory complaint. Influenza viruses and RSV were detected in 17% and 11% of outpatient participants with ILI, and 23% and 16% of inpatient participants with ILI, respectively. Influenza (July-September) and RSV (January-April) prevalence peaks were temporally distinct. RSV (relative risk [RR]: 1.78; 95% confidence interval [CI] 1.51-2.11), but not influenza, infection was associated with severe disease among patients with ILI. Underweight patients with ILI were more likely to be infected with influenza A (prevalence ratio [PR]: 1.72; 95% CI 1.04-2.87) and to have severe influenza A infections (RR: 2.49; 95% CI 1.57-3.93). CONCLUSIONS: Populations in rural Zambia bear a sizeable burden of viral respiratory infections and severe disease. The epidemiology of infections in this rural area differs from that reported from urban areas in Zambia.

Point-of-care p24 antigen detection for early infant diagnosis of HIV infection: cross-sectional and longitudinal studies in Zambia.

BACKGROUND: Early infant diagnosis of HIV infection is challenging in sub-Saharan Africa, particularly in rural areas, leading to delays in diagnosis and treatment. Use of a point-of-care test would overcome many challenges. This study evaluated the validity of a novel point-of-care p24 antigen detection test (LYNX) in rural and urban settings in southern Zambia. METHODS: Two studies were conducted: a cross-sectional study from 2014 to 2015 at Macha Hospital (LYNX Hospital study) and a longitudinal study from 2016 to 2018 at 12 health facilities in Southern Province, Zambia (NSEBA study). In both studies, children attending the facilities for early infant diagnosis were enrolled and a blood sample was collected for routine testing at the central lab and immediate on-site testing with the LYNX test. The performance of the LYNX test was measured in comparison to nucleic acid-based testing at the central lab. RESULTS: In the LYNX Hospital study, 210 tests were performed at a median age of 23.5 weeks (IQR: 8.9, 29.0). The sensitivity and specificity of the test were 70.0 and 100.0%, respectively. In the NSEBA study, 2608 tests were performed, including 1305 at birth and 1222 on children ≥4 weeks of age. For samples tested at birth, sensitivity was 13.6% (95% CI: 2.9, 34.9) and specificity was 99.6% (95% CI: 99.1, 99.9). While specificity was high for all ages, sensitivity increased with age and was higher for participants tested at ≥4 weeks of age (80.6%; 95% CI: 67.4, 93.7). Children with positive nucleic acid tests were more likely to be negative by the LYNX test if their mother received antiretroviral therapy during pregnancy (60.7% vs. 24.2%; p = 004). CONCLUSIONS: Considering the high specificity and moderate sensitivity that increased with age, the LYNX test could be of value for early infant diagnosis for infants ≥4 weeks of age, particularly in rural areas where centralized testing leads to long delays. Point-of-care tests with moderate sensitivity and high specificity that are affordable, easy-to-use, and easily implemented and maintained should be developed to expand access to testing and deliver same-day results to infants in areas where it is not feasible to implement nucleic acid-based point-of-care assays.

Treatment outcomes among children younger than five years living with HIV in rural Zambia, 2008-2018: a cohort study.

BACKGROUND: HIV testing and treatment guidelines for children in sub-Saharan Africa have evolved over time, such that children are now treated at younger ages. The objective of this study was to describe the treatment experience for immunologic, virologic, and growth outcomes among HIV-infected Zambian children younger than 5 years of age from 2008 to 2018. METHODS: Participants enrolled in a clinical cohort study in Macha, Zambia and initiating antiretroviral treatment before 5 years of age between 2008 and 2015 were included in the analysis and followed up to the end of 2018. Outcomes, including growth, CD4+ T-cell percentage, viral suppression, and mortality, were evaluated among all children using longitudinal and survival analyses. Comparisons by age at treatment initiation (< 1, 1 to < 2, and 2 to < 5 years) were also evaluated. RESULTS: Three hundred eighty-one children initiating treatment before 5 years of age between 2008 and 2015 were included in the analysis. Growth metrics and CD4+ T-cell percentage improved over time after treatment initiation. However, 20% of children remained underweight and 40% of children remained stunted after the first 36 months of treatment. 85% of children had a viral load < 400 copies/mL after 12 months of treatment. However, children < 1 year at treatment initiation were more likely to have a detectable viral load in the first 12 months of treatment and less likely to achieve viral suppression compared to older children. Mortality was highest in the first 12 months of treatment, among underweight children, and among children initiating treatment in 2008-2010 compared to 2011-2015. CONCLUSIONS: Most children initiating antiretroviral treatment from 2008 to 2015 in rural Zambia responded well to treatment. However, many children remained underweight and stunted, and experienced high mortality rates during the first few months of treatment. This supports continued efforts to improve early infant diagnosis, nutritional support, and pediatric drug formulations.

The impact of delayed treatment of uncomplicated P. falciparum malaria on progression to severe malaria: A systematic review and a pooled multicentre individual-patient meta-analysis.

BACKGROUND: Delay in receiving treatment for uncomplicated malaria (UM) is often reported to increase the risk of developing severe malaria (SM), but access to treatment remains low in most high-burden areas. Understanding the contribution of treatment delay on progression to severe disease is critical to determine how quickly patients need to receive treatment and to quantify the impact of widely implemented treatment interventions, such as 'test-and-treat' policies administered by community health workers (CHWs). We conducted a pooled individual-participant meta-analysis to estimate the association between treatment delay and presenting with SM. METHODS AND FINDINGS: A search using Ovid MEDLINE and Embase was initially conducted to identify studies on severe Plasmodium falciparum malaria that included information on treatment delay, such as fever duration (inception to 22nd September 2017). Studies identified included 5 case-control and 8 other observational clinical studies of SM and UM cases. Risk of bias was assessed using the Newcastle-Ottawa scale, and all studies were ranked as 'Good', scoring ≥7/10. Individual-patient data (IPD) were pooled from 13 studies of 3,989 (94.1% aged <15 years) SM patients and 5,780 (79.6% aged <15 years) UM cases in Benin, Malaysia, Mozambique, Tanzania, The Gambia, Uganda, Yemen, and Zambia. Definitions of SM were standardised across studies to compare treatment delay in patients with UM and different SM phenotypes using age-adjusted mixed-effects regression. The odds of any SM phenotype were significantly higher in children with longer delays between initial symptoms and arrival at the health facility (odds ratio [OR] = 1.33, 95% CI: 1.07-1.64 for a delay of >24 hours versus ≤24 hours; p = 0.009). Reported illness duration was a strong predictor of presenting with severe malarial anaemia (SMA) in children, with an OR of 2.79 (95% CI:1.92-4.06; p < 0.001) for a delay of 2-3 days and 5.46 (95% CI: 3.49-8.53; p < 0.001) for a delay of >7 days, compared with receiving treatment within 24 hours from symptom onset. We estimate that 42.8% of childhood SMA cases and 48.5% of adult SMA cases in the study areas would have been averted if all individuals were able to access treatment within the first day of symptom onset, if the association is fully causal. In studies specifically recording onset of nonsevere symptoms, long treatment delay was moderately associated with other SM phenotypes (OR [95% CI] >3 to ≤4 days versus ≤24 hours: cerebral malaria [CM] = 2.42 [1.24-4.72], p = 0.01; respiratory distress syndrome [RDS] = 4.09 [1.70-9.82], p = 0.002). In addition to unmeasured confounding, which is commonly present in observational studies, a key limitation is that many severe cases and deaths occur outside healthcare facilities in endemic countries, where the effect of delayed or no treatment is difficult to quantify. CONCLUSIONS: Our results quantify the relationship between rapid access to treatment and reduced risk of severe disease, which was particularly strong for SMA. There was some evidence to suggest that progression to other severe phenotypes may also be prevented by prompt treatment, though the association was not as strong, which may be explained by potential selection bias, sample size issues, or a difference in underlying pathology. These findings may help assess the impact of interventions that improve access to treatment.

Improving the efficiency of reactive case detection for malaria elimination in southern Zambia: a cross-sectional study.

BACKGROUND: Reactive case detection (RCD) seeks to enhance malaria surveillance and control by identifying and treating parasitaemic individuals residing near index cases. In Zambia, this strategy starts with passive detection of symptomatic incident malaria cases at local health facilities or by community health workers, with subsequent home visits to screen-and-treat residents in the index case and neighbouring (secondary) households within a 140-m radius using rapid diagnostic tests (RDTs). However, a small circular radius may not be the most efficient strategy to identify parasitaemic individuals in low-endemic areas with hotspots of malaria transmission. To evaluate if RCD efficiency could be improved by increasing the probability of identifying parasitaemic residents, environmental risk factors and a larger screening radius (250 m) were assessed in a region of low malaria endemicity. METHODS: Between January 12, 2015 and July 26, 2017, 4170 individuals residing in 158 index and 531 secondary households were enrolled and completed a baseline questionnaire in the catchment area of Macha Hospital in Choma District, Southern Province, Zambia. Plasmodium falciparum prevalence was measured using PfHRP2 RDTs and quantitative PCR (qPCR). A Quickbird™ high-resolution satellite image of the catchment area was used to create environmental risk factors in ArcGIS, and generalized estimating equations were used to evaluate associations between risk factors and secondary households with parasitaemic individuals. RESULTS: The parasite prevalence in secondary (non-index case) households was 0.7% by RDT and 1.8% by qPCR. Overall, 8.5% (n = 45) of secondary households had at least one resident with parasitaemia by qPCR or RDT. The risk of a secondary household having a parasitaemic resident was significantly increased in proximity to higher order streams and marginally with increasing distance from index households. The adjusted OR for proximity to third- and fifth-order streams were 2.97 (95% CI 1.04-8.42) and 2.30 (95% CI 1.04-5.09), respectively, and that for distance to index households for each 50 m was 1.24 (95% CI 0.98-1.58). CONCLUSION: Applying proximity to streams as a screening tool, 16% (n = 3) more malaria-positive secondary households were identified compared to using a 140-m circular screening radius. This analysis highlights the potential use of environmental risk factors as a screening strategy to increase RCD efficiency.

The CYB5R3c .350C>G and G6PD A alleles modify severity of anemia in malaria and sickle cell disease.

Genetic modifiers of anemia in Plasmodium falciparum infection and sickle cell disease (SCD) are not fully known. Both conditions are associated with oxidative stress, hemolysis and anemia. The CYB5R3 gene encodes cytochrome b5 reductase 3, which converts methemoglobin to hemoglobin through oxidation of NADH. CYB5R3c.350C > G encoding CYB5R3T117S , the most frequent recognized African-specific polymorphism, does not have known functional significance, but its high allele frequency (23% in African Americans) suggests a selection advantage. Glucose-6-phosphate dehydrogenase (G6PD) is essential for protection from oxidants; its African-polymorphic X-linked A+ and A- alleles, and other variants with reduced activity, coincide with endemic malaria distribution, suggesting protection from lethal infection. We examined the association of CYB5R3c.350C > G with severe anemia (hemoglobin <5 g/dL) in the context of G6PD A+ and A- status among 165 Zambian children with malaria. CYB5R3c.350C > G offered protection against severe malarial anemia in children without G6PD deficiency (G6PD wild type or A+/A- heterozygotes) (odds ratio 0.29, P = .022) but not in G6PD A+ or A- hemizygotes/homozygotes. We also examined the relationship of CYB5R3c.350C > G with hemoglobin concentration among 267 children and 321 adults and adolescents with SCD in the US and UK and found higher hemoglobin in SCD patients without G6PD deficiency (ß = 0.29, P = .022 children; ß = 0.33, P = .004 adults). Functional studies in SCD erythrocytes revealed mildly lower activity of native CYB5R3T117S compared to wildtype CYB5R3 and higher NADH/NAD+ ratios. In conclusion, CYB5R3c.350C > G appears to ameliorate anemia severity in malaria and SCD patients without G6PD deficiency, possibly accounting for CYB5R3c.350C > G selection and its high prevalence.

Acceptability and feasibility of testing for HIV infection at birth and linkage to care in rural and urban Zambia: a cross-sectional study.

BACKGROUND: Early infant diagnosis is important for timely identification of HIV-infected infants and linkage to care. Testing at birth has been implemented to facilitate earlier diagnosis of HIV infection but may present new challenges. This study was conducted to understand the acceptability and feasibility of birth testing in urban and rural settings in southern Zambia. METHODS: This cross-sectional study was conducted at 11 hospitals and clinics in Livingstone, Choma, and Macha in Southern Province, Zambia from 2016 to 2018. Infants born to pregnant women living with HIV at the sites were eligible for enrollment. After enrollment, a questionnaire was administered to the mother and a dried blood spot card was collected from infants for testing at a central laboratory. When results were available, mothers were notified to return to the clinic. Acceptability of birth testing was evaluated based on the proportion of women who agreed to participate and the reasons for non-participation among women who declined. Feasibility of testing at birth was evaluated using turnaround times for returning results, the proportion of women receiving results, and linkage to care for infants testing positive. RESULTS: One thousand four hundred three women were approached for the study. A small proportion declined due to refusal of birth testing (0 to 8.2% across sites). One thousand two hundred ninety women agreed to have their infants tested. The proportion of mothers receiving results ranged from 51.6 to 92.1%, and was significantly lower at the hospital than clinics in Livingstone (51.6% vs. 69.8%; p < 0.0001) and Macha (69.5% vs. 85.7%; p < 0.0001) but not Choma (85.7% vs. 92.1%; p = 0.34). For mothers who received test results, the median turnaround time from sample collection was 67 days in Livingstone and 53 days in Macha and Choma. Overall, 23 (1.8%) infants tested positive for HIV but only 8 (34.8%) were linked to care a median of 68 days (range: 29, 784) after sample collection. CONCLUSIONS: While testing at birth was acceptable, this study highlights the operational challenges under a centralized laboratory testing system. Point-of-care platforms are needed for rapid testing and return of results so HIV-infected children can be identified, linked to care, and treated as early as possible.

Tracking Progress Toward Elimination of Mother to Child Transmission of HIV in Zambia: Findings from the Early Infant Diagnosis of HIV Program (2009-2017).

BACKGROUND: We carried out analyses of early infant testing results at Livingstone Central Hospital in Zambia to assess time of testing, linkages to care and availability of test results for clinical decision making. METHODS: We abstracted data from registers of HIV-exposed infants who had dried blood spots cards (DBS) collected for DNA-PCR from January 2009 to December 2017. Only those tested from 2014 to 2017 had additional data which were used to estimate risk factors for mother-to-child HIV transmission using logistic regression models. RESULTS: DBS were collected from 2630 children. The proportion of HIV-positive tests decreased from 21% in 2009 to 2% in 2016 and 2017. Median turnaround time for results was 9 weeks (IQR: 5, 15) for HIV-negative, 7 weeks (IQR: 5, 13) for HIV-positive children. Only 2% of infants whose mothers took antiretroviral therapy (ART) were HIV positive, while 18% of infants whose mothers took short course antiretroviral drugs (ARVs) were infected. Infants of mothers who did not take ARVs had 9 times the odds of an HIV positive test (OR = 8.9, 95% CI: 3.6, 22.6). Infants of mothers who received short course ARVs were 40% less likely to get an HIV test within the first 2 months of life (OR = 0.6, 95% CI: 0.4, 0.9) compared to infants of mothers who received ART. Only 52% had a third test at median age 52 weeks (IQR: 50, 54). CONCLUSIONS: Long turnaround time for test results and low retention in care after the initial HIV test were critical challenges to clinical decision making.

Who is Advocating for Children and Adolescents With HIV Infection?

Despite significant gains in the past 10 years in the treatment and care of children and adolescents with HIV infection, there is still need for advocacy for this group. The author discusses how far we have come and the need for on-going efforts, not only for appropriate child-friendly drug formulations, but also for cooperation between quantitative and qualitative scientists to further the existing knowledge base of how best to help these children with a chronic disease.

Genetic Evidence of Focal Plasmodium falciparum Transmission in a Pre-elimination Setting in Southern Province, Zambia.

BACKGROUND: Southern Province, Zambia has experienced a dramatic decline in Plasmodium falciparum malaria transmission in the past decade and is targeted for elimination. Zambia's National Malaria Elimination Program recommends reactive case detection (RCD) within 140 m of index households to enhance surveillance and eliminate remaining transmission foci. METHODS: To evaluate whether RCD captures local transmission, we genotyped 26 microsatellites from 106 samples collected from index (n = 27) and secondary (n = 79) cases detected through RCD in the Macha Hospital catchment area between January 2015 and April 2016. RESULTS: Participants from the same RCD event harbored more genetically related parasites than those from different RCD events, suggesting that RCD captures, at least in part, infections related through local transmission. Related parasites clustered in space and time, up to at least 250 m from index households. Spatial analysis identified a putative focal transmission hotspot. CONCLUSIONS: The current RCD strategy detects focal transmission events, although programmatic guidelines to screen within 140 m of index households may fail to capture all secondary cases. This study highlights the utility of parasite genetic data in assessing programmatic interventions, and similar approaches may be useful to malaria elimination programs seeking to tailor intervention strategies to the underlying transmission epidemiology.

Predictors of loss to follow-up among children on long-term antiretroviral therapy in Zambia (2003-2015).

BACKGROUND: Retention in care is critical for children living with HIV taking antiretroviral therapy (ART). Loss to follow-up (LTFU) is high in HIV treatment programs in resource limited settings. We estimated the cumulative incidence of LTFU and identified associated risk factors among children on ART at Livingstone Central Hospital (LCH), Zambia. METHODS: Using a retrospective cohort study design, we abstracted data from medical records of children who received ART between 2003 and 2015. Loss to follow-up was defined as no clinical and pharmacy contact for at least 90 days after the child missed their last scheduled clinical visit. Non-parametric competing risks models were used to estimate the cumulative incidence of death, LTFU and transfer. Cause-specific Cox regression was used to estimate the hazard ratios of the risk factors of LTFU. RESULTS: A total of 1039 children aged 0-15 years commenced ART at LCH between 2003 and 2015. Median duration of follow-up was 3.8 years (95% CI: 1.2-6.5), median age at ART initiation was 3.6 years (IQR: 1.3-8.6), 179 (17%) started treatment during their first year of life. At least 167 (16%) were LTFU and we traced 151 (90%). Of those we traced, 39 (26%) had died, 71 (47%) defaulted, 20 (13%) continued ART at other clinics and 21 (14%) continued treatment with gaps. The cumulative incidence of LTFU for the entire cohort was 2.7% (95% CI: 1.9-3.9) at 3 months, 4.1% (95% CI: 2.9-5.4) at 6 months and 14.1% (95% CI: 12.4-16.9) after 5 years on ART. Associated risk factors were: 1) non-disclosure of HIV status at baseline, aHR = 1.9 (1.2-2.9), 2) No phone ownership, aHR = 2.1 (1.6-2.9), 3) starting treatment between 2013 to 2015, aHR = 5.6 (2.2-14.1). CONCLUSION: Among the children LTFU mortality and default were substantially high. Children who started treatment in recent years (2013-2015) had the highest hazard of LTFU. Lack of access to a phone and non-disclosure of HIV-status to the index child was associated with higher hazards of LTFU. We recommend re-enforcement of client counselling and focused follow-up strategies using modern technology such as mobile phones as adjunct to current approaches.

Long-term survival outcomes of HIV infected children receiving antiretroviral therapy: an observational study from Zambia (2003-2015).

BACKGROUND: In 2017, 64% of children living with HIV in Zambia accessed Antiretroviral Therapy (ART). Despite expanded ART coverage, there is paucity of information on effectiveness of pediatric ART in reducing mortality. The aim of this research is to describe treatment outcomes, measure mortality rates and assess predictors of mortality among children receiving ART. METHODS: Using a retrospective cohort study design, we abstracted routinely collected clinical data from medical records of children from birth to 15 years old, who had received ART for at least 6 months at Livingstone Central Hospital in Southern Province Zambia, between January 2003 and June 2015. The primary outcome was death. Cause of death was ascertained from medical records and death certificates. Distribution of survival times according to baseline covariates were estimated using Kaplan Meier and Cox Proportional Hazards methods. RESULTS: Overall, 1039 children were commenced on ART during the study period. The median age at treatment initiation was 3.6 years (IQR: 1.3-8.6) and 520 (50%) children were female. Of these, 71 (7%) died, 164 (16%) were lost to follow-up, 210 (20%) transferred and 594 (56%) were actively on treatment. After 4450 person years, mortality rate was 1.6/100 (95% CI: 1.4-1.8). Mortality was highest during the first 3 months of treatment (11.7/100 (95% CI: 7.6-16.3). In multivariable proportional hazards regression, the adjusted hazards of death were highest among children aged < 1 year (aHR = 3.1 (95% CI: 1.3-6.4), compared to those aged 6-15 years, WHO stage 4 (aHR =4.8 (95% CI: 2.3-10), compared to WHO stage 1 and 2. In the sensitivity analysis to address bias due to loss to follow-up, mortality increased 5 times when we assumed that all the children who were lost to follow up died within 90 days of their last visit. CONCLUSION: We observed low attrition due to mortality among children on ART. Loss to follow-up was high (16%). Mortality was highest during the first 3 months of treatment. Children aged less than one year and those with advanced WHO disease stage had higher mortality. We recommend effective interventions to improve retention in care and early diagnosis of HIV in children.

Evidence for histidine-rich protein 2 immune complex formation in symptomatic patients in Southern Zambia.

BACKGROUND: Rapid diagnostic tests based on histidine-rich protein 2 (HRP2) detection are the primary tools used to detect Plasmodium falciparum malaria infections. Recent conflicting reports call into question whether α-HRP2 antibodies are present in human host circulation and if resulting immune complexes could interfere with HRP2 detection on malaria RDTs. This study sought to determine the prevalence of immune-complexed HRP2 in a low-transmission region of Southern Zambia. METHODS: An ELISA was used to quantify HRP2 in patient sample DBS extracts before and after heat-based immune complex dissociation. A pull-down assay reliant on proteins A, G, and L was developed and applied for IgG and IgM capture and subsequent immunoprecipitation of any HRP2 present in immune complexed form. A total of 104 patient samples were evaluated using both methods. RESULTS: Immune-complexed HRP2 was detectable in 17% (18/104) of all samples evaluated and 70% (16/23) of HRP2-positive samples. A majority of the patients with samples containing immune-complexed HRP2 had P. falciparum infections (11/18) and were also positive for free HRP2 (16/18). For 72% (13/18) of patients with immune-complexed HRP2, less than 10% of the total HRP2 present was in immune-complexed form. For the remaining samples, a large proportion (≥ 20%) of total HRP2 was complexed with α-HRP2 antibodies. CONCLUSIONS: Endogenous α-HRP2 antibodies form immune complexes with HRP2 in the symptomatic patient population of a low-transmission area in rural Southern Zambia. For the majority of patients, the percentage of HRP2 in immune complexes is low and does not affect HRP2-based malaria diagnosis. However, for some patients, a significant portion of the total HRP2 was in immune-complexed form. Future studies investigating the prevalence and proportion of immune-complexed HRP2 in asymptomatic individuals with low HRP2 levels will be required to assess whether α-HRP2 antibodies affect HRP2 detection for this portion of the transmission reservoir.

Controlled release spatial repellent devices (CRDs) as novel tools against malaria transmission: a semi-field study in Macha, Zambia.

BACKGROUND: The emergence of mosquitoes that can avoid indoor-deployed interventions, such as treated bed nets and indoor residual spraying, threatens the mainstay of malaria control in Zambia. Furthermore, the requirement for high coverage of these tools poses operational challenges. Spatial repellents are being assessed to supplement these vector control tools, but limitations exist in the residual effect of the repellent and the need for external power or heat for diffusion of the volatiles. METHODS: A semi-field evaluation of a novel controlled release spatial repellent device (CRD) was conducted in Macha, Zambia. These devices emanate metofluthrin with no need for external power. Devices were deployed in huts within the semi-field system (SFS). Female Anopheles gambiae sensu stricto released within the SFS were trapped overnight by light traps and collected by aspiration the next morning inside and outside of huts to determine the extent of mosquito repellency and the impact on host-seeking and survival. Experiments studied the impact of number of devices as well as the presence of hut occupants. The study was complemented with numerical methods based on computational fluid dynamics to simulate spatial distribution of metofluthrin. RESULTS: Presence of CRDs was associated with significant reductions in indoor counts of mosquitoes, regardless of whether huts were occupied or not. Repellency ranged from 15 to 60% compared to huts with no devices. Reducing the number of devices from 16 to 4 had little impact on repellency. When huts were occupied, indoor mosquito host-seeking was higher in the presence of CRDs, whilst survival was significantly reduced. CONCLUSIONS: This study demonstrated that deployment of as few as four CRDs within a hut was associated with reduced indoor mosquito densities. As would be expected, presence of occupants within huts, resulted in greater indoor catches (both with and without devices). The increased indoor mosquito host-seeking and mortality in huts when devices were present may be explained by the excito-repellency activity of metofluthrin. These semi-field experiments provide preliminary data on the utility of CRD spatial repellents to reduce indoor densities of An. gambiae mosquitoes. Studies will further investigate the impact of CRDs on mosquito behaviour as well as epidemiological protective efficacy.