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Abstract Autophagy receptor NDP52 triggers bacterial autophagy against infection. However, the ability of NDP52 to protect against viral infection has not been established. We show that NDP52 binds to envelope proteins of hepatitis B virus (HBV) and triggers a degradation process that promotes HBV clearance. Inactivating NDP52 in hepatocytes results in decreased targeting of viral envelopes in the lysosome and increased levels of viral replication. NDP52 inhibits HBV at both viral entry and late replication stages. In contrast to NDP52-mediated bacterial autophagy, lysosomal degradation of HBV envelopes is independent of galectin 8 and ATG5. NDP52 forms complex with Rab9 and viral envelope proteins and links HBV to Rab9-dependent lysosomal degradation pathway. These findings reveal that NDP52 acts as a sensor for HBV infection, which mediates a unique antiviral response to eliminate the virus. This work also suggests direct roles for autophagy receptors in other lysosomal degradation pathways than canonical autophagy.

Abstract Background Few studies on neonatal severe bacterial infection are available in LMICs. Data are needed in these countries to prioritize interventions and decrease neonatal infections which are a primary cause of neonatal mortality. The BIRDY project (Bacterial Infections and Antimicrobial Drug Resistant among Young Children) was initially conducted in Madagascar, Senegal and Cambodia (BIRDY 1, 2012–2018), and continued in Madagascar only (BIRDY 2, 2018–2021). We present here the BIRDY 2 project whose objectives were (1) to estimate the incidence of neonatal severe bacterial infections and compare these findings with those obtained in BIRDY 1, (2) to identify determinants associated with severe bacterial infection and (3) to specify the antibiotic resistance pattern of bacteria in newborns. Methods The BIRDY 2 study was a prospective community-based mother and child cohort, both in urban and semi-rural areas. All pregnant women in the study areas were identified and enrolled. Their newborns were actively and passively followed-up from birth to 3 months. Data on clinical symptoms developed by the children and laboratory results of all clinical samples investigated were collected. A Cox proportional hazards model was performed to identify risk factors associated with possible severe bacterial infection. Findings A total of 53 possible severe bacterial infection and 6 confirmed severe bacterial infection episodes were identified among the 511 neonates followed-up, with more than half occurring in the first 3 days. For the first month period, the incidence of confirmed severe bacterial infection was 11.7 per 1,000 live births indicating a 1.3 -fold decrease compared to BIRDY 1 in Madagascar (p = 0.50) and the incidence of possible severe bacterial infection was 76.3, indicating a 2.6-fold decrease compared to BIRDY 1 in Madagascar (p

Abstract The human gut bacteriome is believed to have pivotal influences on human health and disease while the particular roles associated with the gut phageome have not been fully characterized yet with few exceptions. It is argued that gut microbiota can have a potential role in autism spectrum disorders (ASD). The public microbiota database of ASD and typically developing (TD) Chinese individuals were analyzed for phage protein-coding units (pPCU) to find any link between the phageome and ASD. The gut phageome of ASD individuals showed a wider diversity and higher abundance compared to TD individuals. The ASD phageome was associated with a significant expansion of Caudoviricetes bacteriophages. Phages infecting Bacteroidaceae and prophages encoded within Faecalibacterium were more frequent in ASD than in TD individuals. The expansion and diversification of ASD phageome can influence the bacterial homeostasis by imposing pressure on the bacterial communities. In conclusion, the differences of phages community in in ASD and TD can be used as potential diagnosis biomarkers of ASD. Further investigations are needed to verify the role of gut phage communities in the pathogenesis of ASD.

BackgroundAntibiotic resistance is a global public health issue, particularly in low- and middle-income countries (LMICs), where antibiotics required to treat resistant infections are not affordable. LMICs also bear a disproportionately high burden of bacterial diseases, particularly among children, and resistance jeopardizes progress made in these areas. Although outpatient antibiotic use is a major driver of antibiotic resistance, data on inappropriate antibiotic prescribing in LMICs are scarce at the community level, where the majority of prescribing occurs. Here, we aimed to characterize inappropriate antibiotic prescribing among young outpatient children and to identify its determinants in 3 LMICs.Methods and findingsWe used data from a prospective, community-based mother-and-child cohort (BIRDY, 2012 to 2018) conducted across urban and rural sites in Madagascar, Senegal, and Cambodia. Children were included at birth and followed-up for 3 to 24 months. Data from all outpatient consultations and antibiotics prescriptions were recorded. We defined inappropriate prescriptions as antibiotics prescribed for a health event determined not to require antibiotic therapy (antibiotic duration, dosage, and formulation were not considered). Antibiotic appropriateness was determined a posteriori using a classification algorithm developed according to international clinical guidelines. We used mixed logistic analyses to investigate risk factors for antibiotic prescription during consultations in which children were determined not to require antibiotics. Among the 2,719 children included in this analysis, there were 11,762 outpatient consultations over the follow-up period, of which 3,448 resulted in antibiotic prescription. Overall, 76.5% of consultations resulting in antibiotic prescription were determined not to require antibiotics, ranging from 71.5% in Madagascar to 83.3% in Cambodia. Among the 10,416 consultations (88.6%) determined not to require antibiotic therapy, 25.3% (n = 2,639) nonetheless resulted in antibiotic prescription. This proportion was much lower in Madagascar (15.6%) than in Cambodia (57.0%) or Senegal (57.2%) (p < 0.001). Among the consultations determined not to require antibiotics, in both Cambodia and Madagascar the diagnoses accounting for the greatest absolute share of inappropriate prescribing were rhinopharyngitis (59.0% of associated consultations in Cambodia, 7.9% in Madagascar) and gastroenteritis without evidence of blood in the stool (61.6% and 24.6%, respectively). In Senegal, uncomplicated bronchiolitis accounted for the greatest number of inappropriate prescriptions (84.4% of associated consultations). Across all inappropriate prescriptions, the most frequently prescribed antibiotic was amoxicillin in Cambodia and Madagascar (42.1% and 29.2%, respectively) and cefixime in Senegal (31.2%). Covariates associated with an increased risk of inappropriate prescription include patient age greater than 3 months (adjusted odds ratios (aOR) with 95% confidence interval (95% CI) ranged across countries from 1.91 [1.63, 2.25] to 5.25 [3.85, 7.15], p < 0.001) and living in rural as opposed to urban settings (aOR ranged across countries from 1.83 [1.57, 2.14] to 4.40 [2.34, 8.28], p < 0.001). Diagnosis with a higher severity score was also associated with an increased risk of inappropriate prescription (aOR = 2.00 [1.75, 2.30] for moderately severe, 3.10 [2.47, 3.91] for most severe, p < 0.001), as was consultation during the rainy season (aOR = 1.32 [1.19, 1.47], p < 0.001). The main limitation of our study is the lack of bacteriological documentation, which may have resulted in some diagnosis misclassification and possible overestimation of inappropriate antibiotic prescription.ConclusionIn this study, we observed extensive inappropriate antibiotic prescribing among pediatric outpatients in Madagascar, Senegal, and Cambodia. Despite great intercountry heterogeneity in prescribing practices, we identified common risk factors for inappropriate prescription. This underscores the importance of implementing local programs to optimize antibiotic prescribing at the community level in LMICs.

Abstract Background Anaemia occurs in children when the haemoglobin level in the blood is less than the normal (11 g/dL), the consequence is the decrease of oxygen quantity in the tissues. It is a prevalent public health problem in many low-income countries, including Madagascar, and data on risk factors are lacking. We used existing data collected within the pathophysiology of environmental enteric dysfunction (EED) in Madagascar and the Central African Republic project (AFRIBIOTA project) conducted in underprivileged neighbourhoods of Antananarivo to investigate the factors associated with anaemia in children 24 to 59 months of age. Methods Children included in the AFRIBIOTA project in Antananarivo for whom data on haemoglobin and ferritin concentrations were available were included in the study. Logistic regression modelling was performed to identify factors associated with anaemia. Results Of the 414 children included in this data analysis, 24.4% were found to suffer from anaemia. We found that older children (adjusted OR: 0.95; 95% CI: 0.93–0.98) were less likely to have anaemia. Those with iron deficiency (adjusted OR: 6.1; 95% CI: 3.4–11.1) and those with a high level of faecal calprotectin (adjusted OR: 2.5; 95% CI: 1.4–4.4) were more likely to have anaemia than controls. Conclusions To reduce anaemia in the children in this underprivileged area, more emphasis should be given to national strategies that improve children’s dietary quality and micronutrient intake. Furthermore, existing measures should be broadened to include measures to reduce infectious disease burden.

Abstract Klebsiella pneumoniae can lead to a wide range of diseases including pneumonia, bloodstream and urinary tract infections. During a short period of a pulmonary plague epidemic in October 2017 in Madagascar, 12 K. pneumoniae isolates were identified in ten sputum and two buboes aspirate samples. These isolates were from 12 patients suspected of plague, without epidemiological relationships, but were negative for Yersinia pestis in culture. Data were collected from the plague national surveillance system. The isolates were characterized by antimicrobial susceptibility testing and whole-genome sequencing. Real-time PCR was performed to confirm the presence of K. pneumoniae DNA in buboes. All isolates were identified as K. pneumoniae sensu stricto. Five isolates were extended-spectrum β-lactamases producers; eleven different sequence types were identified. Five isolates belonged to known hypervirulent sequence types. Our results demonstrate community-acquired pneumonia caused by K. pneumoniae isolates in patients suspected of plague stressing the importance of bed-side differential diagnosis.

Introduction Data regarding the acquisition of extended-spectrum beta-lactamase-producing Enterobacteriaceae (ESBL-PE) in neonates at the community level are scarce in low-income and middle-income countries (LMICs), where the burden of neonatal sepsis is high.Our study aims at identifying and quantifying the role of the different routes of ESBL-PE transmission for neonates, which are still undefined in the community in LMICs.Methods and analysis In a semirural community in Madagascar, 60 mothers and their neonates will be recruited at delivery, during which a maternal stool sample and meconium of the newborn will be collected. Home visits will be planned the day of the delivery and next at days 3, 7, 14, 21 and 28. Stool samples from the newborn, the mother and every other household member will be collected at each visit, as well as samples from the environment in contact with the newborn (food, surfaces and objects). Sociodemographic data and factors which might drive ESBL-PE acquisition will also be collected.We will analyse the isolated ESBL-PE using DNA sequencing methods to characterise clones, resistance genes and plasmids of ESBL-PE. To analyse these data globally, we will develop novel analytical approaches combining mathematical modelling and statistics. Finally, mathematical simulations will be performed to test different strategies of control of ESBL-PE transmission to neonates.In complement, we will conduct an anthropological investigation to understand local environments and practices that would contribute to neonatal ESBL-PE acquisition. In-depth interviews with members of 16 households will be conducted and 4 mother–newborn pairs will be followed by a participants’ observations methodology.Ethics and dissemination The study was approved by the ethical committee in Madagascar and by the institutional review board of Institut Pasteur, Paris, France.Findings will be reported to participating families, collaborators and local government; presented at national and international conferences and disseminated by peer-review publications.

Environmental Enteric Dysfunction (EED) is an associate driver of stunting in poor settings, and intestinal infections indirectly contribute to the pathophysiology underlying EED. Our work aimed at assessing whether enteric viral carriage is determinant to stunting. A total of 464 healthy and asymptomatic children, aged 2 to 5 years, were recruited, and classified as non-stunted, moderately stunted, or severely stunted. Among the recruited children, 329 stool samples were obtained and screened for enteric and non-enteric viruses by real-time polymerase chain reaction. We statistically tested for the associations between enteric viral and potential risk factors. Approximately 51.7% of the stool samples were positive for at least one virus and 40.7% were positive for non-enteric adenoviruses. No statistical difference was observed between virus prevalence and the growth status of the children. We did not find any statistically significant association between viral infection and most of the socio-demographic risk factors studied, except for having an inadequate food quality score or an over-nourished mother. In addition, being positive for Ascaris lumbricoides was identified as a protective factor against viral infection. In conclusion, we did not find evidence of a direct link between stunting and enteropathogenic viral carriage in our population.

Stunting and environmental enteric dysfunction (EED) may be responsible for altered gut and systemic immune responses. However, their impact on circulating immune cell populations remains poorly characterized during early life. A detailed flow cytometry analysis of major systemic immune cell populations in 53 stunted and 52 non-stunted (2 to 5 years old) children living in Antananarivo (Madagascar) was performed. Compared to age-matched non-stunted controls, stunted children aged 2-3 years old had a significantly lower relative proportion of classical monocytes. No significant associations were found between stunting and the percentages of effector T helper cell populations (Th1, Th2, Th17, Th1Th17, and cTfh). However, we found that HLA-DR expression (MFI) on all memory CD4+ or CD8+ T cell subsets was significantly lower in stunted children compared to non-stunted controls. Interestingly, in stunted children compared to the same age-matched non-stunted controls, we observed statistically significant age-specific differences in regulatory T cells (Treg) subsets. Indeed, in 2- to 3-year-old stunted children, a significantly higher percentage of memory Treg, whilst a significantly lower percentage of naive Treg, was found. Our results revealed that both innate and adaptive systemic cell percentages, as well as activation status, were impacted in an age-related manner during stunting. Our study provides valuable insights into the understanding of systemic immune system changes in stunted children.

Environmental Enteric Dysfunction (EED) refers to an incompletely defined syndrome of inflammation, reduced absorptive capacity, and reduced barrier function in the small intestine. It is widespread among children and adults in low- and middle-income countries and is also associated with poor sanitation and certain gut infections possibly resulting in an abnormal gut microbiota, small intestinal bacterial overgrowth (SIBO) and stunting. We investigated bacterial pathogen exposure in stunted and non-stunted children in Antananarivo, Madagascar by collecting fecal samples from 464 children (96 severely stunted, 104 moderately stunted and 264 non-stunted) and the prevalence of SIBO in 109 duodenal aspirates from stunted children (61 from severely stunted and 48 from moderately stunted children). SIBO assessed by both aerobic and anaerobic plating techniques was very high: 85.3% when selecting a threshold of ≥105 CFU/ml of bacteria in the upper intestinal aspirates. Moreover, 58.7% of the children showed more than 106 bacteria/ml in these aspirates. The most prevalent cultivated genera recovered were Streptococcus, Neisseria, Staphylococcus, Rothia, Haemophilus, Pantoea and Branhamella. Feces screening by qPCR showed a high prevalence of bacterial enteropathogens, especially those categorized as being enteroinvasive or causing mucosal disruption, such as Shigella spp., enterotoxigenic Escherichia coli, enteropathogenic E. coli and enteroaggregative E. coli. These pathogens were detected at a similar rate in stunted children and controls, all showing no sign of severe diarrhea the day of inclusion but both living in a highly contaminated environment (slum-dwelling). Interestingly Shigella spp. was the most prevalent enteropathogen found in this study (83.3%) without overrepresentation in stunted children.

Background: Klebsiella pneumoniae (hereafter, Kp) is a major public health threat responsible for high levels of multidrug resistant (MDR) human infections. Besides, Kp also causes severe infections in the community, especially in Asia and Africa. Although most Kp infections are caused by endogenous intestinal carriage, little is known about the prevalence and microbiological characteristics of Kp in asymptomatic human carriage, and attached risk factors including environmental sources exposure. Methods: Here, 911 pregnant women from communities in Madagascar, Cambodia, and Senegal were screened for gut colonization by Kp. Characteristics of Kp strains (antimicrobial susceptibility, genomic diversity, virulence, and resistance genes) were defined, and associated risk factors were investigated. Results: Kp carriage rate was 55.9%, and Kp populations were highly heterogeneous (6 phylogroups, 325 sequence types, Simpson index 99.6%). One third of Kp isolates had acquired antimicrobial resistance genes. MDR-Kp (11.7% to 39.7%) and extended spectrum beta-lactamase (ESBL)-producing Kp (0.7% to 14.7%) varied among countries. Isolates with virulence genes were detected (14.5%). Environmental exposure factors including food, animal contacts, or hospitalization of household members were associated with carriage of Kp, antimicrobial resistance and hypervirulence. However, risk factors were country-specific and Kp subpopulation-specific. Conclusion: This large-scale multicenter study uncovers the huge diversity of Kp in human gut carriage, demonstrates that antimicrobial resistance is widespread in communities of three low-income countries, and underlines the challenges posed by Kp colonization to the control of antimicrobial resistance.

Abstract Background Child undernutrition is a global health issue that is associated with poor sanitation and an altered intestinal microbiota. Immunoglobulin (Ig) A mediates host-microbial homeostasis in the intestine, and acutely undernourished children have been shown to have altered IgA recognition of the fecal microbiota. We sought to determine whether chronic undernutrition (stunting) or intestinal inflammation were associated with antibody recognition of the microbiota using two geographically distinct populations from the Afribiota project. Fecal bacteria from 200 children between 2 and 5 years old in Antananarivo, Madagascar, and Bangui, Central African Republic (CAR), were sorted into IgA-positive (IgA+) and IgA-negative (IgA−) populations by flow cytometry and subsequently characterized by 16S rRNA gene sequencing to determine IgA-bacterial targeting. We additionally measured IgG+ fecal bacteria by flow cytometry in a subset of 75 children. Results Stunted children (height-for-age z-score ≤ −2) had a greater proportion of IgA+ bacteria in the fecal microbiota compared to non-stunted controls. This trend was consistent in both countries, despite the higher overall IgA-targeting of the microbiota in Madagascar, but lost significance in each country individually. Two of the most highly IgA-recognized bacteria regardless of nutritional status were Campylobacter (in CAR) and Haemophilus (in both countries), both of which were previously shown to be more abundant in stunted children; however, there was no association between IgA-targeting of these bacteria and either stunting or inflammatory markers. IgG-bound intestinal bacteria were rare in both stunted and non-stunted children, similar to levels observed in healthy populations. Conclusions Undernourished children carry a high load of intestinal pathogens and pathobionts. Our data suggest that stunted children have a greater proportion of IgA-recognized fecal bacteria. We moreover identify two putative pathobionts, Haemophilus and Campylobacter, that are broadly targeted by intestinal IgA. This study furthers our understanding of host-microbiota interactions in undernutrition and identifies immune-recognized microbes for future study.

Objectives: A molecular analysis was performed of two Providencia rettgeri (P. rettgeri) strains (Pr 297 and Pr 269) collected in 2007 and 2009 from wound swabs of patients admitted to the intensive care units at Joseph Ravoangy Andrianavalona hospital and the Military Hospital in Antananarivo, Madagascar. Methods: The two P. rettgeri isolates were subjected to susceptibility testing. Whole genome sequencing was performed to characterise the antibiotic resistance genes, genomic islands and mobilomes (integrons, plasmids and insertion sequences). Results: All isolates were found to be multidrug-resistant. Antibiotic-resistant genes described were amongst eight different classes of antimicrobial agents. Thirty insertion sequences and twelve genomic islands were predicted in each genome. Class 1 and class 2 integrons were found in both genomes, with gene cassette regions encompassing arr-2 - cmlA5 - blaOXA-10 - ant (3”)-Ia and dfrA1 - sat2 - ant (3”)-Ia – orfX, respectively. IncA/C2, ColM and ColE1-like plasmids were described harbouring blaCMY-30, qnrD and aac(6’)-Ib-cr4 genes, respectively. Phylogenetic analysis showed that Pr 297 and Pr 269 isolates were genetically identical and clustered with P. rettgeri strains described in the USA and Spain. Conclusions: It is believed that this is the first molecular characterisation of wound infection pathogens from Madagascan patients and the first description of P. rettgeri co-producing CMY-30, OXA-10 and AAC(6’)-Ib-cr4 enzymes. The diversity of the resistance determinants and mobile genetic elements was probably due to extensive horizontal gene transfer events, highlighting the need to conduct further molecular monitoring studies to understand the genomic plasticity of resistant bacteria in Madagascan hospitals.

BackgroundSevere bacterial infections (SBIs) are a leading cause of neonatal deaths in low- and middle-income countries (LMICs). However, most data came from hospitals, which do not include neonates who did not seek care or were treated outside the hospital. Studies from the community are scarce, and few among those available were conducted with high-quality microbiological techniques. The burden of SBI at the community level is therefore largely unknown. We aimed here to describe the incidence, etiology, risk factors, and antibiotic resistance profiles of community-acquired neonatal SBI in 3 LMICs.Methods and findingsThe BIRDY study is a prospective multicentric community-based mother and child cohort study and was conducted in both urban and rural areas in Madagascar (2012 to 2018), Cambodia (2014 to 2018), and Senegal (2014 to 2018). All pregnant women within a geographically defined population were identified and enrolled. Their neonates were actively followed from birth to 28 days to document all episodes of SBI. A total of 3,858 pregnant women (2,273 (58.9%) in Madagascar, 814 (21.1%) in Cambodia, and 771 (20.0%) in Senegal) were enrolled in the study, and, of these, 31.2% were primigravidae. Women enrolled in the urban sites represented 39.6% (900/2,273), 45.5% (370/814), and 61.9% (477/771), and those enrolled in the rural sites represented 60.4% (1,373/2,273), 54.5% (444/814), and 38.1% (294/771) of the total in Madagascar, Cambodia, and Senegal, respectively. Among the 3,688 recruited newborns, 49.6% were male and 8.7% were low birth weight (LBW). The incidence of possible severe bacterial infection (pSBI; clinical diagnosis based on WHO guidelines of the Integrated Management of Childhood Illness) was 196.3 [95% confidence interval (CI) 176.5 to 218.2], 110.1 [88.3 to 137.3], and 78.3 [59.5 to 103] per 1,000 live births in Madagascar, Cambodia, and Senegal, respectively. The incidence of pSBI differed between urban and rural sites in all study countries. In Madagascar, we estimated an incidence of 161.0 pSBI per 1,000 live births [133.5 to 194] in the urban site and 219.0 [192.6 to 249.1] pSBI per 1,000 live births in the rural site (p = 0.008). In Cambodia, estimated incidences were 141.1 [105.4 to 189.0] and 85.3 [61.0 to 119.4] pSBI per 1,000 live births in urban and rural sites, respectively (p = 0.025), while in Senegal, we estimated 103.6 [76.0 to 141.2] pSBI and 41.5 [23.0 to 75.0] pSBI per 1,000 live births in urban and rural sites, respectively (p = 0.006). The incidences of culture-confirmed SBI were 15.2 [10.6 to 21.8], 6.5 [2.7 to 15.6], and 10.2 [4.8 to 21.3] per 1,000 live births in Madagascar, Cambodia, and Senegal, respectively, with no difference between urban and rural sites in each country. The great majority of early-onset infections occurred during the first 3 days of life (72.7%). The 3 main pathogens isolated were Klebsiella spp. (11/45, 24.4%), Escherichia coli (10/45, 22.2%), and Staphylococcus spp. (11/45, 24.4%). Among the 13 gram-positive isolates, 5 were resistant to gentamicin, and, among the 29 gram-negative isolates, 13 were resistant to gentamicin, with only 1 E. coli out of 10 sensitive to ampicillin. Almost one-third of the isolates were resistant to both first-line drugs recommended for the management of neonatal sepsis (ampicillin and gentamicin). Overall, 38 deaths occurred among neonates with SBI (possible and culture-confirmed SBI together). LBW and foul-smelling amniotic fluid at delivery were common risk factors for early pSBI in all 3 countries. A main limitation of the study was the lack of samples from a significant proportion of infants with pBSI including 35 neonatal deaths. Without these samples, bacterial infection and resistance profiles could not be confirmed.ConclusionsIn this study, we observed a high incidence of neonatal SBI, particularly in the first 3 days of life, in the community of 3 LMICs. The current treatment for the management of neonatal infection is hindered by antimicrobial resistance. Our findings suggest that microbiological diagnosis of SBI remains a challenge in these settings and support more research on causes of neonatal death and the implementation of early interventions (e.g., follow-up of at-risk newborns during the first days of life) to decrease the burden of neonatal SBI and associated mortality and help achieve Sustainable Development Goal 3.

Abstract Background The present study aimed to perform a deep phenotypic and genotypic analysis of 15 clinical carbapenem-resistant Acinetobacter baumannii (CRAb) strains isolated in Madagascar between 2008 and 2016 from diverse sources. Methods CRAb isolates collected from the Clinical Biology Centre of the Institut Pasteur of Madagascar, from the neonatal unit of Antananarivo military hospital, and from intensive care units of Mahajanga Androva and Antananarivo Joseph Ravoahangy Andrianavalona (HJRA) hospitals were subjected to susceptibility testing. Whole-genome sequencing allowed us to assess the presence of antibiotic-resistance determinants, insertion sequences, integrons, genomic islands and potential virulence factors in all strains. The structure of the carO porin gene and deduced protein (CarO) were also assessed in CRAb isolates. Results All isolates were found to be multidrug-resistant strains. Antibiotic-resistance genes against six classes of antimicrobial agents were described. The four carbapenem-resistance genes: bla OXA-51 like , bla OXA-23 , bla OXA-24 , and bla OXA-58 genes were detected in 100, 53.3, 13.3, and 6.6% of the isolates, respectively. Additionally, an ISAba1 located upstream of bla OXA-23 and bla ADC-like genes was observed in 53.3 and 66.7% of isolates, respectively. Further, Tn2006 and Tn2008 were found associated to the ISAba1-bla OXA-23 structure. An 8051-bp mobilizable plasmid harbouring the bla OXA-24 gene was isolated in two strains. In addition, 46.7% of isolates were positive for class 1 integrons. Overall, five sequences types (STs), with predominantly ST2, were detected. Several virulence genes were found in the CRAb isolates, among which two genes, epsA and ptk, responsible for the capsule-positive phenotype, were involved in A. baumannii pathogenesis. Conclusions This study revealed the presence of high-level carbapenem resistance in A. baumannii with the first description of OXA-24 and OXA-58 carbapenemases in Madagascar. This highlights the importance of better monitoring and controlling CRAb in Madagascan hospitals to avoid their spread.

BackgroundThis study aimed to compare the prevalence of intestinal parasite infestations (IPIs) in stunted children, compared to control children, in Ankasina and Andranomanalina Isotry (two disadvantaged neighborhoods of Antananarivo, Madagascar), to characterize associated risk factors and to compare IPI detection by real-time PCR and standard microscopy techniques.Methodology/principal findingsFecal samples were collected from a total of 410 children (171 stunted and 239 control) aged 2-5 years. A single stool sample per subject was examined by simple merthiolate-iodine-formaldehyde (MIF), Kato-Katz smear and real-time PCR techniques. A total of 96.3% of the children were infested with at least one intestinal parasite. The most prevalent parasites were Giardia intestinalis (79.5%), Ascaris lumbricoides (68.3%) and Trichuris trichiura (68.0%). For all parasites studied, real-time PCR showed higher detection rates compared to microscopy (G. intestinalis [77.6% (n = 318) versus 20.9% (n = 86)], Entamoeba histolytica [15.8% (n = 65) versus 1.9% (n = 8)] and A. lumbricoides [64.1% (n = 263) versus 50.7% (n = 208)]). Among the different variables assessed in the study, age of 4 to 5 years (AOR = 4.61; 95% CI, (1.35-15.77)) and primary and secondary educational level of the mother (AOR = 12.59; 95% CI, (2.76-57.47); AOR = 9.17; 95% CI, (2.12-39.71), respectively) were significantly associated with IPIs. Children drinking untreated water was associated with infestation with G. intestinalis (AOR = 1.85; 95% CI, (1.1-3.09)) and E. histolytica (AOR = 1.9; 95% CI, (1.07-3.38)). E. histolytica was also associated with moderately stunted children (AOR = 0.37; 95% CI, 0.2-0.71). Similarly, children aged between 4 and 5 years (AOR = 3.2; 95% CI (2.04-5.01)) and living on noncemented soil types (AOR = 1.85; 95% CI, (1.18-2.09)) were associated with T. trichiura infestation.Conclusions/significanceThe prevalence of IPIs is substantial in the studied areas in both stunted and control children, despite the large-scale drug administration of antiparasitic drugs in the country. This high prevalence of IPIs warrants further investigation. Improved health education, environmental sanitation and quality of water sources should be provided.

Abstract Background Globally one out of four children under 5 years is affected by linear growth delay (stunting). This syndrome has severe long-term sequelae including increased risk of illness and mortality and delayed psychomotor development. Stunting is a syndrome that is linked to poor nutrition and repeated infections. To date, the treatment of stunted children is challenging as the underlying etiology and pathophysiological mechanisms remain elusive. We hypothesize that pediatric environmental enteropathy (PEE), a chronic inflammation of the small intestine, plays a major role in the pathophysiology of stunting, failure of nutritional interventions and diminished response to oral vaccines, potentially via changes in the composition of the pro- and eukaryotic intestinal communities. The main objective of AFRIBIOTA is to describe the intestinal dysbiosis observed in the context of stunting and to link it to PEE. Secondary objectives include the identification of the broader socio-economic environment and biological and environmental risk factors for stunting and PEE as well as the testing of a set of easy-to-use candidate biomarkers for PEE. We also assess host outcomes including mucosal and systemic immunity and psychomotor development. This article describes the rationale and study protocol of the AFRIBIOTA project. Methods AFRIBIOTA is a case-control study for stunting recruiting children in Bangui, Central African Republic and in Antananarivo, Madagascar. In each country, 460 children aged 2–5 years with no overt signs of gastrointestinal disease are recruited (260 with no growth delay, 100 moderately stunted and 100 severely stunted). We compare the intestinal microbiota composition (gastric and small intestinal aspirates; feces), the mucosal and systemic immune status and the psychomotor development of children with stunting and/or PEE compared to non-stunted controls. We also perform anthropological and epidemiological investigations of the children’s broader living conditions and assess risk factors using a standardized questionnaire. Discussion To date, the pathophysiology and risk factors of stunting and PEE have been insufficiently investigated. AFRIBIOTA will add new insights into the pathophysiology underlying stunting and PEE and in doing so will enable implementation of new biomarkers and design of evidence-based treatment strategies for these two syndromes.

Severe bacterial infections are a leading cause of death among neonates in low-income countries, which harbor several factors leading to emergence and spread of multidrug-resistant bacteria. Low-income countries should prioritize interventions to decrease neonatal infections; however, data are scarce, specifically from the community. To assess incidence, etiologies, and antimicrobial drug–resistance patterns of neonatal infections, during 2012–2014, we conducted a community-based prospective investigation of 981 newborns in rural and urban areas of Madagascar. The incidence of culture-confirmed severe neonatal infections was high: 17.7 cases/1,000 live births. Most (75%) occurred during the first week of life. The most common (81%) bacteria isolated were gram-negative. The incidence rate for multidrug-resistant neonatal infection was 7.7 cases/1,000 live births. In Madagascar, interventions to improve prevention, early diagnosis, and management of bacterial infections in neonates should be prioritized.

Abstract Background Pneumococcus kills over one million children annually and over 90 % of these deaths occur in low-income countries especially in Sub-Saharan Africa (SSA) where HIV exacerbates the disease burden. In SSA, serotype 1 pneumococci particularly the endemic ST217 clone, causes majority of the pneumococcal disease burden. To understand the evolution of the virulent ST217 clone, we analysed ST217 whole genomes from isolates sampled from African and Asian countries. Methods We analysed 226 whole genome sequences from the ST217 lineage sampled from 9 African and 4 Asian countries. We constructed a whole genome alignment and used it for phylogenetic and coalescent analyses. We also screened the genomes to determine presence of antibiotic resistance conferring genes. Results Population structure analysis grouped the ST217 isolates into five sequence clusters (SCs), which were highly associated with different geographical regions and showed limited intracontinental and intercontinental spread. The SCs showed lower than expected genomic sequence, which suggested strong purifying selection and small population sizes caused by bottlenecks. Recombination rates varied between the SCs but were lower than in other successful clones such as PMEN1. African isolates showed higher prevalence of antibiotic resistance genes than Asian isolates. Interestingly, certain West African isolates harbored a defective chloramphenicol and tetracycline resistance-conferring element (Tn5253) with a deletion in the loci encoding the chloramphenicol resistance gene (catpC194), which caused lower chloramphenicol than tetracycline resistance. Furthermore, certain genes that promote colonisation were absent in the isolates, which may contribute to serotype 1’s rarity in carriage and consequently its lower recombination rates. Conclusions The high phylogeographic diversity of the ST217 clone shows that this clone has been in circulation globally for a long time, which allowed its diversification and adaptation in different geographical regions. Such geographic adaptation reflects local variations in selection pressures in different locales. Further studies will be required to fully understand the biological mechanisms which makes the ST217 clone highly invasive but unable to successfully colonise the human nasopharynx for long durations which results in lower recombination rates.

Improved methods for the detection and characterization of carried Neisseria meningitidis isolates are needed. We evaluated a multiplex PCR algorithm for the detection of a variety of carriage strains in the meningitis belt. To further improve the sensitivity and specificity of the existing PCR assays, primers for gel-based PCR assays (sodC, H, Z) and primers/probe for real-time quantitative PCR (qPCR) assays (porA, cnl, sodC, H, E, Z) were modified or created using Primer Express software. Optimized multiplex PCR assays were tested on 247 well-characterised carriage isolates from six countries of the African meningitis belt. The PCR algorithm developed enabled the detection of N. meningitidis species using gel-based and real-time multiplex PCR targeting porA, sodC, cnl and characterization of capsule genes through sequential multiplex PCR assays for genogroups (A, W, X, then B, C, Y and finally H, E and Z). Targeting both porA and sodC genes together allowed the detection of meningococci with a sensitivity of 96% and 89% and a specificity of 78% and 67%, for qPCR and gel-based PCR respectively. The sensitivity and specificity ranges for capsular genogrouping of N. meningitidis are 67% - 100% and 98%-100% respectively for gel-based PCR and 90%-100% and 99%-100% for qPCR. We developed a PCR algorithm that allows simple, rapid and systematic detection and characterisation of most major and minor N. meningitidis capsular groups, including uncommon capsular groups (H, E, Z).

In low and middle income countries (LMICs), where the burden of neonatal sepsis is the highest, the spread of extended spectrum beta-lactamase-producing enterobacteriaceae (ESBL-PE) in the community, potentially contributing to the neonatal mortality, is a public health concern. Data regarding the acquisition of ESBL-PE during the neonatal period are scarce. The routes of transmission are not well defined and particularly the possible key role played by pregnant women. This study aimed to understand the neonatal acquisition of ESBL-PE in the community in Madagascar. The study was conducted in urban and semi-rural areas. Newborns were included at birth and followed-up during their first month of life. Maternal stool samples at delivery and six stool samples in each infant were collected to screen for ESBL-PE. A Cox proportional hazards model was performed to identify factors associated with the first ESBL-PE acquisition. The incidence rate of ESBL-PE acquisition was 10.4 cases/1000 newborn-days [95% CI: 8.0-13.4 cases per 1000 newborn-days]. Of the 83 ESBL-PE isolates identified, Escherichia coli was the most frequent species (n = 28, 34.1%), followed by Klebsiella pneumoniae (n = 20, 24.4%). Cox multivariate analysis showed that independent risk factors for ESBL-PE acquisition were low birth weight (adjusted Hazard-ratio (aHR) = 2.7, 95% CI [1.2; 5.9]), cesarean-section, (aHR = 3.4, 95% CI [1.7; 7.1]) and maternal use of antibiotics at delivery (aHR = 2.2, 95% CI [1.1; 4.5]). Our results confirm that mothers play a significant role in the neonatal acquisition of ESBL-PE. In LMICs, public health interventions during pregnancy should be reinforced to avoid unnecessary caesarean section, unnecessary antibiotic use at delivery and low birth weight newborns.

We identified 3 lineages among multidrug-resistant (MDR) Salmonella enterica serotype Typhi isolates in the Gulf of Guinea region in Africa during the 2000s. However, the MDR H58 haplotype, which predominates in southern Asia and Kenya, was not identified. MDR quinolone-susceptible isolates contained a 190-kb incHI1 pST2 plasmid or a 50-kb incN pST3 plasmid.

Antimicrobial resistance (AMR) is a major threat to human, animal health, and environment worldwide. For human, transmission occurred through a variety of routes both in health-care settings and community. In animals, AMR was reported in livestock, pets, and wildlife; transmission of AMR can be zoonotic with the probably most important route being foodborne transmission. The Indian Ocean Commission (IOC), composed of Comoros, Madagascar, Mauritius, Reunion (France), and Seychelles recognized the surveillance of AMR in both animal and human as a main public health priority for the region. Mayotte, French overseas territory, located in Comoros archipelago, was also included in this review. This review summarized our best epidemiological knowledge regarding AMR in Indian Ocean. We documented the prevalence, and phenotypic and genotypic profiles of prone to resistance Gram-positive and Gram-negative bacteria both in animals and humans. Our review clearly pointed out extended-spectrum β-lactamase and carbapenemase-producing Enterobacteriaceae as main human and animal health issue in IOC. However, publications on AMR are scarce, particularly in Comoros, Mayotte, and Seychelles. Thus, research and surveillance priorities were recommended (i) estimating the volume of antimicrobial drugs used in livestock and human medicine in the different territories [mainly third generation cephalosporin (3GC)]; (ii) developing a “One Health” surveillance approach with epidemiological indicators as zoonotic foodborne pathogen (i.e., couple Escherichia coli resistance to 3GC/carbapenems); (iii) screening travelers with a history of hospitalization and consumption of antibiotic drug returning from at risk areas (e.g., mcr-1 transmission with China or hajj pilgrims) allowing an early warning detection of the emergence for quick control measures implementation in IOC.

Serum bactericidal antibody titres that correlate with protection against invasive meningococcal disease have been characterised. However, titres that are associated with protection against acquisition of pharyngeal carriage of Neisseria meningitidis are not known.Sera were obtained from the members of a household in seven countries of the African meningitis belt in which a pharyngeal carrier of N. meningitidis had been identified during a cross-sectional survey. Serum bactericidal antibody titres at baseline were compared between individuals in the household of the carrier who became a carrier of a meningococcus of the same genogroup during six months of subsequent follow-up and household members who did not become a carrier of a meningococcus of this genogroup during this period.Serum bacterial antibody titres were significantly higher in carriers of a serogroup W or Y meningococcus at the time of recruitment than in those who were not a carrier of N. meningitidis of the same genogroup. Serum bactericidal antibody titres to a strain of N. meningitis of the same genogroup as the index cases were no different in individuals who acquired carriage with a meningococcus of the same genogroup as the index case than in those who did not become a carrier during six months of follow-up.Serum bacterial antibody titres to N. meningitidis of genogroup W or Y in the range of those acquired by natural exposure to meningococci of these genogroups, or with cross-reactive bacteria, are not associated with protection against acquisition of carriage with meningococci of either of these genogroups.

BACKGROUND:To mitigate the burden of pneumococcal infections in Niger, a 13-valent pneumococcal vaccine, PCV13, was introduced for routine child vaccination in July 2014. In order to provide pre-vaccine baseline data and allow appreciation of changes on carriage due to vaccination, we analyzed retrospectively pneumococcal isolates obtained from healthy, 0 to 2 year old children prior to the vaccine introduction. METHODS:From June 5, 2007, to May 26, 2008, 1200 nasopharyngeal swabs were collected from healthy 0 to 2 year old children and analyzed by standard microbiological methods. Serotyping was done by SM-PCR and the data were analyzed with R version 2.15.0 (2012-03-30). RESULTS:Streptococcus pneumoniae was detected in 654/1200 children (54.5%) among whom 339 (51.8%) were males. The ages of the study subjects varied from few days to 26 months (mean = 7.1, median = 6, 95% CI [6.8-7.4]). Out of 654 frozen isolates, 377 (54.8%) were able to be re-grown and analyzed. In total, 32 different serogroups/serotypes were detected of which, the most prevalent were 6/(6A/6B/6C/6D) (15.6%), 23F (10.6%), 19F (9.3%), 14 (9%), 19A (5.6%), 23B (4.0%), 25F/38 (3.7%), 18/(18A/18B/18C/18F) (2.9%) and PCR non-typeable (16.4%). Eleven serogroups/serotypes accounting for 57.3% (216/377) were of PCV13 types. Of the 211/377 (56%) isolates tested for drug sensitivity, 23/211 (10.9%), 24/211 (11.4%), 9/211(4.3%) and 148/210 (70.5%) were respectively resistance to oxacillin, chloramphenicol, erythromycin and tetracycline. Thirteen of the oxacillin resistant isolates were additionally multidrug-resistant. No resistance was however detected to gentamycin500μg and to fluoroquinolones (ø Norfloxacin5μg 3 months and presence in family of more than one sibling aged < 6 years were significant risk factors for carriage. CONCLUSION:A global rate of 54.5% pneumococcal carriage was detected in this study. The introduced PCV13 vaccine should cover 57.3% (216/377) of circulating serogroups/serotypes, among which were those resistant to antibiotics. Age > 3 months and presence in family of children aged < 6 years were significant factors for pneumococcal carriage. The present data should help understanding post vaccine introduction changes in pneumococcal carriage and infections for better action.

Knowledge of rotavirus epidemiology is necessary to make informed decisions about vaccine introduction and to evaluate vaccine impact. During April 2010–March 2012, rotavirus surveillance was conducted among 9,745 children 80% of isolates during the second year. Hospitalization and death rates and severe dehydration among rotavirus case-patients did not differ during the 2 years. The emergence of G12P[8] warrants close attention to the characteristics of associated epidemics and possible prevention measures.

Neisseria meningitidis (Nm), Haemophilus influenzae (Hi), and Streptococcus pneumoniae (Sp) are the lead causes of bacterial meningitis. Detection of these pathogens from clinical specimens using traditional real-time PCR (rt-PCR) requires DNA extraction to remove the PCR inhibitors prior to testing, which is time consuming and labor intensive. In this study, five species-specific (Nm-sodC and -ctrA, Hi-hpd#1 and -hpd#3 and Sp-lytA) and six serogroup-specific rt-PCR tests (A, B, C, W, X, Y) targeting Nm capsular genes were evaluated in the two direct rt-PCR methods using PerfeCTa and 5x Omni that do not require DNA extraction. The sensitivity and specify of the two direct rt-PCR methods were compared to TaqMan traditional rt-PCR, the current standard rt-PCR method for the detection of meningitis pathogens. The LLD for all 11 rt-PCR tests ranged from 6,227 to 272,229 CFU/ml for TaqMan, 1,824-135,982 for 5x Omni, and 168-6,836 CFU/ml for PerfeCTa. The diagnostic sensitivity using TaqMan ranged from 89.2%-99.6%, except for NmB-csb, which was 69.7%. For 5x Omni, the sensitivity varied from 67.1% to 99.8%, with three tests below 90%. The sensitivity of these tests using PerfeCTa varied from 89.4% to 99.8%. The specificity ranges of the 11 tests were 98.0-99.9%, 97.5-99.9%, and 92.9-99.9% for TaqMan, 5x Omni, and PerfeCTa, respectively. PerfeCTa direct rt-PCR demonstrated similar or better sensitivity compared to 5x Omni direct rt-PCR or TaqMan traditional rt-PCR. Since the direct rt-PCR method does not require DNA extraction, it reduces the time and cost for processing CSF specimens, increases testing throughput, decreases the risk of cross-contamination, and conserves precious CSF. The direct rt-PCR method will be beneficial to laboratories with high testing volume.

[This corrects the article DOI: 10.1371/journal.pone.0147928.].

The pattern of epidemic meningococcal disease in the African meningitis belt may be influenced by the background level of population immunity but this has been measured infrequently. A standardised enzyme-linked immunosorbent assay (ELISA) for measuring meningococcal serogroup A IgG antibodies was established at five centres within the meningitis belt. Antibody concentrations were then measured in 3930 individuals stratified by age and residence from six countries. Seroprevalence by age was used in a catalytic model to determine the force of infection. Meningococcal serogroup A IgG antibody concentrations were high in each country but showed heterogeneity across the meningitis belt. The geometric mean concentration (GMC) was highest in Ghana (9.09 μg/mL [95% CI 8.29, 9.97]) and lowest in Ethiopia (1.43 μg/mL [95% CI 1.31, 1.57]) on the margins of the belt. The force of infection was lowest in Ethiopia (λ = 0.028). Variables associated with a concentration above the putative protective level of 2 μg/mL were age, urban residence and a history of recent vaccination with a meningococcal vaccine. Prior to vaccination with the serogroup A meningococcal conjugate vaccine, meningococcal serogroup A IgG antibody concentrations were high across the African meningitis belt and yet the region remained susceptible to epidemics.

Melioidosis, caused by the bacterium Burkholderia pseudomallei, is an infectious disease of humans or animals, and the specific environmental conditions that are present in western Indian Ocean islands are particularly suitable for the establishment/survival of B. pseudomallei. Indeed, an increasing number of new cases have been reported in this region (Madagascar, Mauritius, Réunion (France), and Seychelles, except Comoros and Mayotte (France)), and are described in this review. Our review clearly points out that further studies are needed in order to investigate the real incidence and burden of melioidosis in the western Indian Ocean and especially Madagascar, since it is likely to be higher than currently reported. Thus, research and surveillance priorities were recommended (i) to improve awareness of melioidosis in the population and among clinicians; (ii) to improve diagnostics, in order to provide rapid and effective treatment; (iii) to implement a surveillance and reporting system in the western Indian Ocean; and (iv) to investigate the presence of B. pseudomallei in environmental samples, since we have demonstrated its presence in soil samples originating from the yard of a Madagascan case.

Invasive meningococcal disease (IMD) is a major cause of bacterial meningitides and septicaemia. This study shows the results of the laboratory-based surveillance of IMD in Belgium over the period 1997-2012.The results are based on microbiological and molecular laboratory surveillance of 2997 clinical isolates of N. meningitides received by the Belgian Meningococcal Reference Centre (BMRC) over the period 1997-2012.Serogroup B has always been a major cause of meningococcal disease in Belgium, with P3.4 as most frequent serotype till 2008, while an increase in non-serotypable strains has been observed in the last few years. Clonal complexes cc-41/44 and cc-269 are most frequently observed in serogroup B strains. In the late nineties, the incidence of serogroup C disease increased considerably and peaked in 2001, mainly associated with phenotypes C:2a:P1.5,2, C:2a:P1.5 and C:2a:P1.2 (ST-11/ET-37 clonal complex). The introduction of the meningococcal C conjugate vaccine has been followed by an 88% significant decrease in serogroup C disease from 2001 to 2004 nationally, yet sharper in Flanders (92%) compared to Wallonia (77%). Since 2008 a difference in incidence of serogroup C was observed in Flanders (0-0.1/100,000) versus Wallonia (0.1-0.3/100,000).This study showed the change in epidemiology and strain population over a 16 years period spanning an exhaustive vaccination campaign and highlights the influence of regional vaccination policies with different cohorts sizes on short and long-term IMD incidences.

To determine whether cats were a risk for transmission of Salmonella to humans, we evaluated the excretion of Salmonella by pet cats. Rectal-swab specimens were taken from 278 healthy house cats, from 58 cats that died of disease, and from 35 group-housed cats. Group-housed cats were kept in one room with three cat trays and a common water and feed tray. Eighteen (51.4%) of 35 group-housed cats, 5 (8.6%) of 58 diseased cats (5/58), and 1 (0.36%) of 278 healthy house cats excreted Salmonella. Salmonella isolates were of serotypes Typhimurium, Enteritidis, Bovismorbificans and 4:i:-. Acquired antimicrobial resistance was found in serotype Typhimurium (resistance to ampicillin, chloramphenicol, and tetracycline; to ampicillin; and to chloramphenicol) and 4:i:- strains (resistance to ampicillin, chloramphenicol, sulfonamides, trimethoprim, and sulfamethoxazole/trimethoprim). Cats that excrete Salmonella can pose a public health hazard to people who are highly susceptible to Salmonella, such as children, the elderly, and immunocompromised persons.

BACKGROUND:Epidemics of meningococcal meningitis (MM) recurrently strike the African Meningitis Belt. This study aimed at investigating factors, still poorly understood, that influence annual incidence of MM serogroup A, the main etiologic agent over 2004-2010, at a fine spatial scale in Niger. METHODOLOGY/PRINCIPAL FINDINGS:To take into account data dependencies over space and time and control for unobserved confounding factors, we developed an explanatory Bayesian hierarchical model over 2004-2010 at the health centre catchment area (HCCA) level. The multivariate model revealed that both climatic and non-climatic factors were important for explaining spatio-temporal variations in incidence: mean relative humidity during November-June over the study region (posterior mean Incidence Rate Ratio (IRR) = 0.656, 95% Credible Interval (CI) 0.405-0.949) and occurrence of early rains in March in a HCCA (IRR = 0.353, 95% CI 0.239-0.502) were protective factors; a higher risk was associated with the percentage of neighbouring HCCAs having at least one MM A case during the same year (IRR = 2.365, 95% CI 2.078-2.695), the presence of a road crossing the HCCA (IRR = 1.743, 95% CI 1.173-2.474) and the occurrence of cases before 31 December in a HCCA (IRR = 6.801, 95% CI 4.004-10.910). At the study region level, higher annual incidence correlated with greater geographic spread and, to a lesser extent, with higher intensity of localized outbreaks. CONCLUSIONS:Based on these findings, we hypothesize that spatio-temporal variability of MM A incidence between years and HCCAs result from variations in the intensity or duration of the dry season climatic effects on disease risk, and is further impacted by factors of spatial contacts, representing facilitated pathogen transmission. Additional unexplained factors may contribute to the observed incidence patterns and should be further investigated.

OBJECTIVE:Detection of meningococcal carriers is key to understanding the epidemiology of Neisseria meningitidis, yet no gold standard has been established. Here, we directly compare two methods for collecting pharyngeal swabs to identify meningococcal carriers. METHODS:We conducted cross-sectional surveys of schoolchildren at multiple sites in Africa to compare swabbing the posterior pharynx behind the uvula (U) to swabbing the posterior pharynx behind the uvula plus one tonsil (T). Swabs were cultured immediately and analyzed using molecular methods. RESULTS:One thousand and six paired swab samples collected from schoolchildren in four countries were analyzed. Prevalence of meningococcal carriage was 6.9% (95% CI: 5.4-8.6%) based on the results from both swabs, but the observed prevalence was lower based on one swab type alone. Prevalence based on the T swab or the U swab alone was similar (5.2% (95% CI: 3.8-6.7%) versus 4.9% (95% CI: 3.6-6.4%) respectively (p=0.6)). The concordance between the two methods was 96.3% and the kappa was 0.61 (95% CI: 0.50-0.73), indicating good agreement. CONCLUSIONS:These two commonly used methods for collecting pharyngeal swabs provide consistent estimates of the prevalence of carriage, but both methods misclassified carriers to some degree, leading to underestimates of the prevalence.

BACKGROUND: The epidemiology of pneumococcal meningitis in the African 'meningitis belt' is poorly studied. In order to ensure an effective vaccination strategy and post-vaccination surveillance, we examined the serotype distribution patterns of pneumococcal meningitis in Niger over the period 2003-2011. METHODS: Cerebrospinal fluid (CSF) samples were collected from different health facilities throughout Niger in the frame of the national microbiological surveillance of meningitis. Determination of the serotype of CSF positive for pneumococci was performed using a sequential multiplex PCR method (SM-PCR) adapted with a national algorithm in which 32 different serotypes were covered and grouped into eight consecutive PCR. RESULTS: The SM-PCR assay could predict the Sp serotype for 779 CSF (88.7%), 98 CSF (11.3%) were not-typeable in our national-adapted algorithm. In total, 26 different serotypes were identified. Serotype 1 (n = 393) was the most prevalent and accounted for 45.3% of infections, followed by serogroups/serotypes 12F/(12A)/(44)/(46) (7.3%), 6/(6A/6B/6C/6D) (5.4%), 14 (5.2%), 5 (4.6%), 23F (4.2%), 45 (3.6%), 2 (3.1%), 18/(18A/18B/18C/18F) (2.9%) and 17 others serotypes with a prevalence of less than 2%. The proportion of serotype 1 in infants(

BackgroundMeningococcal meningitis is a major health problem in the "African Meningitis Belt" where recurrent epidemics occur during the hot, dry season. In Niger, a central country belonging to the Meningitis Belt, reported meningitis cases varied between 1,000 and 13,000 from 2003 to 2009, with a case-fatality rate of 5-15%.Methodology/principal findingsIn order to gain insight in the epidemiology of meningococcal meningitis in Niger and to improve control strategies, the emergence of the epidemics and their diffusion patterns at a fine spatial scale have been investigated. A statistical analysis of the spatio-temporal distribution of confirmed meningococcal meningitis cases was performed between 2002 and 2009, based on health centre catchment areas (HCCAs) as spatial units. Anselin's local Moran's I test for spatial autocorrelation and Kulldorff's spatial scan statistic were used to identify spatial and spatio-temporal clusters of cases. Spatial clusters were detected every year and most frequently occurred within nine southern districts. Clusters most often encompassed few HCCAs within a district, without expanding to the entire district. Besides, strong intra-district heterogeneity and inter-annual variability in the spatio-temporal epidemic patterns were observed. To further investigate the benefit of using a finer spatial scale for surveillance and disease control, we compared timeliness of epidemic detection at the HCCA level versus district level and showed that a decision based on threshold estimated at the HCCA level may lead to earlier detection of outbreaks.Conclusions/significanceOur findings provide an evidence-based approach to improve control of meningitis in sub-Saharan Africa. First, they can assist public health authorities in Niger to better adjust allocation of resources (antibiotics, rapid diagnostic tests and medical staff). Then, this spatio-temporal analysis showed that surveillance at a finer spatial scale (HCCA) would be more efficient for public health response: outbreaks would be detected earlier and reactive vaccination would be better targeted.