Supporting evidence for a human reservoir of invasive non-Typhoidal Salmonella from household samples in Burkina Faso.

BACKGROUND: Salmonella Typhimurium and Enteritidis are major causes of bloodstream infection in children in sub-Saharan Africa. This study assessed evidence for their zoonotic versus human reservoir. METHODS: Index patients were children with blood culture confirmed Salmonella infection recruited during a microbiological surveillance study in Nanoro, rural Burkina between May 2013 and August 2014. After consent, their households were visited. Stool from household members and livestock (pooled samples per species) as well as drinking water were cultured for Salmonella. Isolates with identical serotype obtained from index patient and any household sample were defined as "paired isolates" and assessed for genetic relatedness by multilocus variable number tandem-repeat analysis (MLVA) and whole-genome sequencing (WGS). RESULTS: Twenty-nine households were visited for 32/42 (76.2%) eligible index patients: two households comprised two index patients each, and in a third household the index patient had a recurrent infection. Among the 32 index patients, serotypes were Salmonella Typhimurium (n = 26), Salmonella Enteritidis (n = 5) and Salmonella Freetown (n = 1). All Typhimurium isolates were sequence type (ST)313. Median delay between blood culture sampling and household visits was 13 days (range 6-26). Salmonella was obtained from 16/186 (8.6%) livestock samples (13 serotypes) and 18/290 (6.2%) household members (9 serotypes). None of the water samples yielded Salmonella. Paired Salmonella Typhimurium isolates were obtained from three households representing four index patients. MLVA types were identical in two pairs and similar in the third (consisting of two index patients and one household member). WGS showed a strong genetic relatedness with 0 to 2 core genome SNPs difference between pairs on a household level. Livestock samples did not yield any Salmonella Typhimurium or Salmonella Enteritidis, and the latter was exclusively obtained from blood culture. Other serotypes shared by human and/or livestock carriers in the same household were Salmonella Derby, Drac, Tennessee and Muenster. CONCLUSIONS/SIGNIFICANCE: The current study provides further evidence of a human reservoir for invasive non-Typhoidal Salmonella (iNTS) in sub-Saharan Africa.

Genomic and environmental risk factors for cardiometabolic diseases in Africa: methods used for Phase 1 of the AWI-Gen population cross-sectional study.

There is an alarming tide of cardiovascular and metabolic disease (CMD) sweeping across Africa. This may be a result of an increasingly urbanized lifestyle characterized by the growing consumption of processed and calorie-dense food, combined with physical inactivity and more sedentary behaviour. While the link between lifestyle and public health has been extensively studied in Caucasian and African American populations, few studies have been conducted in Africa. This paper describes the detailed methods for Phase 1 of the AWI-Gen study that were used to capture phenotype data and assess the associated risk factors and end points for CMD in persons over the age of 40 years in sub-Saharan Africa (SSA). We developed a population-based cross-sectional study of disease burden and phenotype in Africans, across six centres in SSA. These centres are in West Africa (Nanoro, Burkina Faso, and Navrongo, Ghana), in East Africa (Nairobi, Kenya) and in South Africa (Agincourt, Dikgale and Soweto). A total of 10,702 individuals between the ages of 40 and 60 years were recruited into the study across the six centres, plus an additional 1021 participants over the age of 60 years from the Agincourt centre. We collected socio-demographic, anthropometric, medical history, diet, physical activity, fat distribution and alcohol/tobacco consumption data from participants. Blood samples were collected for disease-related biomarker assays, and genomic DNA extraction for genome-wide association studies. Urine samples were collected to assess kidney function. The study provides base-line data for the development of a series of cohorts with a second wave of data collection in Phase 2 of the study. These data will provide valuable insights into the genetic and environmental influences on CMD on the African continent.

Population-based incidence, seasonality and serotype distribution of invasive salmonellosis among children in Nanoro, rural Burkina Faso.

BACKGROUND: Bloodstream infections (BSI) caused by Salmonella Typhi and invasive non-Typhoidal Salmonella (iNTS) frequently affect children living in rural sub-Saharan Africa but data about incidence and serotype distribution are rare. OBJECTIVE: The present study assessed the population-based incidence of Salmonella BSI and severe malaria in a Health and Demographic Surveillance System in a rural area with seasonal malaria transmission in Nanoro, Burkina Faso. METHODS: Children between 2 months-15 years old with severe febrile illness were enrolled during a one-year surveillance period (May 2013-May 2014). Thick blood films and blood cultures were sampled and processed upon admission. Population-based incidences were corrected for non-referral, health seeking behavior, non-inclusion and blood culture sensitivity. Adjusted incidence rates were expressed per 100,000 person-years of observations (PYO). RESULTS: Among children < 5 years old, incidence rates for iNTS, Salmonella Typhi and severe malaria per 100,000 PYO were 4,138 (95% Confidence Interval (CI): 3,740-4,572), 224 (95% CI: 138-340) and 2,866 (95% CI: 2,538-3,233) respectively. Among those aged 5-15 years, corresponding incidence rates were 25 (95% CI: 8-60), 273 (95% CI: 203-355) and 135 (95% CI: 87-195) respectively. Most iNTS occurred during the peak of the rainy season and in parallel with the increase of Plasmodium falciparum malaria; for Salmonella Typhi no clear seasonal pattern was observed. Salmonella Typhi and iNTS accounted for 13.3% and 55.8% of all 118 BSI episodes; 71.6% of iNTS (48/67) isolates were Salmonella enterica serovar Typhimurium and 25.4% (17/67) Salmonella enterica serovar Enteritidis; there was no apparent geographical clustering. CONCLUSION: The present findings from rural West-Africa confirm high incidences of Salmonella Typhi and iNTS, the latter with a seasonal and Plasmodium falciparum-related pattern. It urges prioritization of the development and implementation of Salmonella Typhi as well as iNTS vaccines in this setting.

Severe anaemia associated with Plasmodium falciparum infection in children: consequences for additional blood sampling for research.

BACKGROUND: Plasmodium falciparum infection may cause severe anaemia, particularly in children. When planning a diagnostic study on children suspected of severe malaria in sub-Saharan Africa, it was questioned how much blood could be safely sampled; intended blood volumes (blood cultures and EDTA blood) were 6 mL (children aged <6 years) and 10 mL (6-12 years). A previous review [Bull World Health Organ. 89: 46-53. 2011] recommended not to exceed 3.8 % of total blood volume (TBV). In a simulation exercise using data of children previously enrolled in a study about severe malaria and bacteraemia in Burkina Faso, the impact of this 3.8 % safety guideline was evaluated. METHODS: For a total of 666 children aged >2 months to <12 years, data of age, weight and haemoglobin value (Hb) were available. For each child, the estimated TBV (TBVe) (mL) was calculated by multiplying the body weight (kg) by the factor 80 (ml/kg). Next, TBVe was corrected for the degree of anaemia to obtain the functional TBV (TBVf). The correction factor consisted of the rate 'Hb of the child divided by the reference Hb'; both the lowest ('best case') and highest ('worst case') reference Hb values were used. Next, the exact volume that a 3.8 % proportion of this TBVf would present was calculated and this volume was compared to the blood volumes that were intended to be sampled. RESULTS: When applied to the Burkina Faso cohort, the simulation exercise pointed out that in 5.3 % (best case) and 11.4 % (worst case) of children the blood volume intended to be sampled would exceed the volume as defined by the 3.8 % safety guideline. Highest proportions would be in the age groups 2-6 months (19.0 %; worst scenario) and 6 months-2 years (15.7 %; worst case scenario). A positive rapid diagnostic test for P. falciparum was associated with an increased risk of violating the safety guideline in the worst case scenario (p = 0.016). CONCLUSIONS: Blood sampling in children for research in P. falciparum endemic settings may easily violate the proposed safety guideline when applied to TBVf. Ethical committees and researchers should be wary of this and take appropriate precautions.

The impact of clinical research activities on communities in rural Africa: the development of the Clinical Research Unit of Nanoro (CRUN) in Burkina Faso.

BACKGROUND: The opportunities for developing new drugs and vaccines for malaria control look brighter now than ten years ago. However, there are few places in sub-Saharan Africa with the necessary infrastructure and expertise to support such research in compliance to international standards of clinical research (ICH-GCP). The Clinical Research Unit of Nanoro (CRUN) was founded in 2008 to provide a much-needed GCP-compliant clinical trial platform for an imminent large-scale Phase 3 malaria vaccine trial. A dynamic approach was used that entailed developing the required infrastructure and human resources, while engaging local communities in the process as key stakeholders. This provided a better understanding and ownership of the research activities by the local population. CASE DESCRIPTION: Within five years (2008-2013), the CRUN set up a fully and well-equipped GCP-compliant clinical trial research facility, which enabled to attract 25 grants. The research team grew from ten health workers prior to 2008 to 254 in 2013. A Health and Demographic Surveillance System (HDSS), which covers a total population of about 60,000 people in 24 villages was set up in the district. The local community contributed to the development of the facility through the leadership of the king and the mayor of Nanoro. As a result of their active advocacy, the government extended the national electrical grid to the new research center, and later to the entire village. This produced a positive impact on the community's quality of life. The quality of health care improved substantially, due to the creation of more elaborate clinical laboratory services and the acquisition of state-of-the-art equipment. CONCLUSION: Involving the community in the key steps of establishing the centre provided the foundation for what was to become the CRUN success story. This experience demonstrates that when clinical trials research sites are carefully developed and implemented, they can have a positive and powerful impact on local communities in resource-poor settings, well beyond the task of generating expected study data.

Frequency of severe malaria and invasive bacterial infections among children admitted to a rural hospital in Burkina Faso.

BACKGROUND: Although severe malaria is an important cause of mortality among children in Burkina Faso, data on community-acquired invasive bacterial infections (IBI, bacteremia and meningitis) are lacking, as well as data on the involved pathogens and their antibiotic resistance rates. METHODS: The present study was conducted in a rural hospital and health center in Burkina Faso, in a seasonal malaria transmission area. Hospitalized children (<15 years) presenting with T≥38.0°C and/or signs of severe illness were enrolled upon admission. Malaria diagnosis and blood culture were performed for all participants, lumbar puncture when clinically indicated. We assessed the frequency of severe malaria (microscopically confirmed, according to World Health Organization definitions) and IBI, and the species distribution and antibiotic resistance of the bacterial pathogens causing IBI. RESULTS: From July 2012 to July 2013, a total of 711 patients were included. Severe malaria was diagnosed in 292 (41.1%) children, including 8 (2.7%) with IBI co-infection. IBI was demonstrated in 67 (9.7%) children (bacteremia, n = 63; meningitis, n = 6), 8 (11.8%) were co-infected with malaria. Non-Typhoid Salmonella spp. (NTS) was the predominant isolate from blood culture (32.8%), followed by Salmonella Typhi (18.8%), Streptococcus pneumoniae (18.8%) and Escherichia coli (12.5%). High antibiotic resistance rates to first line antibiotics were observed, particularly among Gram-negative pathogens. In addition, decreased ciprofloxacin susceptibility and extended-spectrum beta lactamase (ESBL) production was reported for one NTS isolate each. ESBL production was observed in 3/8 E. coli isolates. In-hospital mortality was 8.2% and case-fatality rates for IBI (23.4%) were significantly higher compared to severe malaria (6.8%, p<0.001). CONCLUSIONS: Although severe malaria was the main cause of illness, IBI were not uncommon and had higher case-fatality rates. The high frequency, antibiotic resistance rates and mortality rates of community acquired IBI require improvement in hygiene, better diagnostic methods and revision of current treatment guidelines.

Effectiveness of artesunate-amodiaquine vs. artemether-lumefantrine for the treatment of uncomplicated falciparum malaria in Nanoro, Burkina Faso: a non-inferiority randomised trial.

OBJECTIVES: Artemisinin-based combination therapies (ACTs) are essential for the effective control of falciparum malaria in endemic countries. However, in most countries, such choice has been carried out without knowing their effectiveness when deployed in real-life conditions, that is, when treatment is not directly observed. We report here the results of a study assessing the effectiveness of the two ACTs currently recommended in Burkina Faso for the treatment of uncomplicated malaria, that is, artemether-lumefantrine (AL) and artesunate-amodiaquine (ASAQ). METHODS: Between September 2008 and January 2010, 340 children were randomised to one of the two study arms and followed up for 42 days. Treatment was administered according to routine practices, that is, the first dose was given by study nurses who explained to the parent/guardian how to administer the other doses at home during the following 2 days. RESULTS: The results showed a significantly higher unadjusted adequate clinical and parasitological response in the ASAQ (58.4%) than in the AL arm (46.1%) at day 28 but these trends were similar after correction with PCR data (ASAQ (89.7%) and AL (89.8%)). New infections started to appear after day 14, first in the AL and then in the ASAQ arm but at day 42 day of follow-up we observed no difference in the occurrence of recrudescent infection. CONCLUSION: Despite a lower cure rate than those reported in efficacy studies in which the treatment administration was directly observed, both AL and ASAQ can still be used for the treatment of uncomplicated malaria in Burkina Faso.

Accuracy of PfHRP2 versus Pf-pLDH antigen detection by malaria rapid diagnostic tests in hospitalized children in a seasonal hyperendemic malaria transmission area in Burkina Faso.

BACKGROUND: In most sub-Saharan African countries malaria rapid diagnostic tests (RDTs) are now used for the diagnosis of malaria. Most RDTs used detect Plasmodium falciparum histidine-rich protein-2 (PfHRP2), though P. falciparum-specific parasite lactate dehydrogenase (Pf-pLDH)-detecting RDTs may have advantages over PfHRP2-detecting RDTs. Only few data are available on the use of RDTs in severe illness and the present study compared Pf-pLDH to PfHRP2-detection. METHODS: Hospitalized children aged one month to 14 years presenting with fever or severe illness were included over one year. Venous blood samples were drawn for malaria diagnosis (microscopy and RDT), culture and complete blood count. Leftovers were stored at -80 °C and used for additional RDT analysis and PCR. An RDT targeting both PfHRP2 and Pf-pLDH was performed on all samples for direct comparison of diagnostic accuracy with microscopy as reference method. PCR was performed to explore false-positive RDT results. RESULTS: In 376 of 694 (54.2%) included children, malaria was microscopically confirmed. Sensitivity, specificity, positive predictive value (PPV) and negative predictive value were 100.0, 70.9, 69.4 and 100.0%, respectively for PfHRP2-detection and 98.7, 94.0, 91.6 and 99.1%, respectively for Pf-pLDH-detection. Specificity and PPV were significantly lower for PfHRP2-detection (p <0.001). For both detection antigens, specificity was lowest for children one to five years and in the rainy season. PPV for both antigens was highest in the rainy season, because of higher malaria prevalence. False positive PfHRP2 results were associated with prior anti-malarial treatment and positive PCR results (98/114 (86.0%) samples tested). CONCLUSION: Among children presenting with severe febrile illness in a seasonal hyperendemic malaria transmission area, the present study observed similar sensitivity but lower specificity and PPV of PfHRP2 compared to Pf-pLDH-detection. Further studies should assess the diagnostic accuracy and safety of an appropriate Pf-pLDH-detecting RDT in field settings and if satisfying, replacement of PfHRP2 by Pf-pLDH-detecting RDTs should be considered.

A phase 3 trial of RTS,S/AS01 malaria vaccine in African infants.

BACKGROUND: The candidate malaria vaccine RTS,S/AS01 reduced episodes of both clinical and severe malaria in children 5 to 17 months of age by approximately 50% in an ongoing phase 3 trial. We studied infants 6 to 12 weeks of age recruited for the same trial. METHODS: We administered RTS,S/AS01 or a comparator vaccine to 6537 infants who were 6 to 12 weeks of age at the time of the first vaccination in conjunction with Expanded Program on Immunization (EPI) vaccines in a three-dose monthly schedule. Vaccine efficacy against the first or only episode of clinical malaria during the 12 months after vaccination, a coprimary end point, was analyzed with the use of Cox regression. Vaccine efficacy against all malaria episodes, vaccine efficacy against severe malaria, safety, and immunogenicity were also assessed. RESULTS: The incidence of the first or only episode of clinical malaria in the intention-to-treat population during the 14 months after the first dose of vaccine was 0.31 per person-year in the RTS,S/AS01 group and 0.40 per person-year in the control group, for a vaccine efficacy of 30.1% (95% confidence interval [CI], 23.6 to 36.1). Vaccine efficacy in the per-protocol population was 31.3% (97.5% CI, 23.6 to 38.3). Vaccine efficacy against severe malaria was 26.0% (95% CI, -7.4 to 48.6) in the intention-to-treat population and 36.6% (95% CI, 4.6 to 57.7) in the per-protocol population. Serious adverse events occurred with a similar frequency in the two study groups. One month after administration of the third dose of RTS,S/AS01, 99.7% of children were positive for anti-circumsporozoite antibodies, with a geometric mean titer of 209 EU per milliliter (95% CI, 197 to 222). CONCLUSIONS: The RTS,S/AS01 vaccine coadministered with EPI vaccines provided modest protection against both clinical and severe malaria in young infants. (Funded by GlaxoSmithKline Biologicals and the PATH Malaria Vaccine Initiative; RTS,S ClinicalTrials.gov number, NCT00866619.).