Differential effects of frozen storage on the molecular detection of bacterial taxa that inhabit the nasopharynx

<p>Abstract</p> <p>Background</p> <p>Frozen storage often precedes metagenomic analysis of biological samples; however, the freezing process can have adverse effects on microbial composition. The effect of freezing on the detection of bacteria inhabiting the infant nasopharynx, a major reservoir of bacterial pathogens, was investigated.</p> <p>Methods</p> <p>16S ribosomal RNA (rRNA) gene-based terminal restriction fragment length polymorphism (T-RFLP) analysis of nasopharyngeal (NP) swabs from twelve Gambian infants was employed. NP swabs were analysed within hours of collection and then after 30 days of storage at -70°C.</p> <p>Results</p> <p>There was substantial heterogeneity among subjects with respect to the effect of freezing on the number of operational taxonomic units (OTUs) detected. Nevertheless, the mean number of OTUs decreased after frozen storage and the relative abundance for 72% of the OTUs changed by less than 0.5% after deep frozen storage. There were differences in the odds of detection and relative abundance of OTUs matched with <it>Moraxella sp</it>., <it>Haemophilus sp</it>./<it>Burkholderia sp.</it>, and <it>Pseudomonas sp</it>. A strong interaction between sex and the effect of freezing was found, whereby there was no significant change observed for males while the mean number of OTUs significantly declined among female infants following frozen storage.</p> <p>Conclusions</p> <p>Although frozen storage of biological samples is often necessary for archiving and logistic purposes, the potential effects on the number of taxa (composition) detected in microbial community studies are significant and should not be overlooked. Moreover, genetic factors such as sex may influence the integrity of nucleic acids during the freezing process.</p

Differential effects of frozen storage on the molecular detection of bacterial taxa that inhabit the nasopharynx

<p>Abstract</p> <p>Background</p> <p>Frozen storage often precedes metagenomic analysis of biological samples; however, the freezing process can have adverse effects on microbial composition. The effect of freezing on the detection of bacteria inhabiting the infant nasopharynx, a major reservoir of bacterial pathogens, was investigated.</p> <p>Methods</p> <p>16S ribosomal RNA (rRNA) gene-based terminal restriction fragment length polymorphism (T-RFLP) analysis of nasopharyngeal (NP) swabs from twelve Gambian infants was employed. NP swabs were analysed within hours of collection and then after 30 days of storage at -70°C.</p> <p>Results</p> <p>There was substantial heterogeneity among subjects with respect to the effect of freezing on the number of operational taxonomic units (OTUs) detected. Nevertheless, the mean number of OTUs decreased after frozen storage and the relative abundance for 72% of the OTUs changed by less than 0.5% after deep frozen storage. There were differences in the odds of detection and relative abundance of OTUs matched with <it>Moraxella sp</it>., <it>Haemophilus sp</it>./<it>Burkholderia sp.</it>, and <it>Pseudomonas sp</it>. A strong interaction between sex and the effect of freezing was found, whereby there was no significant change observed for males while the mean number of OTUs significantly declined among female infants following frozen storage.</p> <p>Conclusions</p> <p>Although frozen storage of biological samples is often necessary for archiving and logistic purposes, the potential effects on the number of taxa (composition) detected in microbial community studies are significant and should not be overlooked. Moreover, genetic factors such as sex may influence the integrity of nucleic acids during the freezing process.</p

Deploying Machine Learning Models Using Progressive Web Applications: Implementation Using a Neural Network Prediction Model for Pneumonia Related Child Mortality in The Gambia.

BACKGROUND: Translating research outputs into practical tools for medical practitioners is a neglected area and could have a substantial impact. One of the barriers to implementing artificial intelligence (AI) and machine learning (ML) applications is their practical deployment in the field. Traditional web-based (i.e., server sided) applications are dependent on reliable internet connections, which may not be readily available in rural areas. Native mobile apps require device specific programming skills as well as contemporary hardware and software, with often rapid and unpredictable platform specific changes. This is a major challenge for using AI/ML tools in resource-limited settings. METHODS: An emerging technology, progressive web applications (PWAs), first introduced by Google in 2015, offers an opportunity to overcome the challenges of deploying bespoke AI/ML systems. The same PWA code can be implemented across all desktop platforms, iOS and Android phones and tablets. In addition to platform independence, a PWA can be designed to be primarily offline. RESULTS: We demonstrate how a neural network-based pneumonia mortality prediction triage tool was migrated from a typical academic framework (paper and web-based prototype) to a tool that can be used offline on any mobile phone-the most convenient deployment vehicle. After an initial online connection to download the software, the application runs entirely offline, reading data from cached memory, and running code via JavaScript. On mobile devices the application is installed as a native app, without the inconvenience of platform specific code through manufacturer code stores. DISCUSSION: We show that an ML application can be deployed as a platform independent offline PWA using a pneumonia-related child mortality prediction tool as an example. The aim of this tool was to assist clinical staff in triaging children for hospital admission, by predicting their risk of death. PWAs function seamlessly when their host devices lose internet connectivity, making them ideal for e-health apps that can help improve health and save lives in resource-limited settings in line with the UN Sustainable Development Goal 3 (SDG3)

A novel nano-iron supplement to safely combat iron deficiency and anaemia in young children: The IHAT-GUT double-blind, randomised, placebo-controlled trial protocol.

Background: Iron deficiency and its associated anaemia (IDA) are the leading forms of micronutrient malnutrition worldwide. Here we describe the rationale and design of the first clinical trial evaluating the efficacy and safety of an innovative nano iron supplement, iron hydroxide adipate tartrate (IHAT), for the treatment of IDA in young children (IHAT-GUT trial). Oral iron is often ineffective due to poor absorption and/or gastrointestinal adverse effects. IHAT is novel since it is effectively absorbed whilst remaining nanoparticulate in the gut, therefore should enable supplementation with fewer symptoms. Methods: IHAT-GUT is a three-arm, double-blind, randomised, placebo-controlled phase II trial conducted in Gambian children 6-35 months of age. The intervention consists of a 12-week supplementation with either IHAT, ferrous sulphate (both at doses bioequivalent to 12.5 mg Fe/day) or placebo. The trial aims to include 705 children with IDA who will be randomly assigned (1:1:1) to each arm. The primary objectives are to test non-inferiority of IHAT in relation to ferrous sulphate at treating IDA, and to test superiority of IHAT in relation to ferrous sulphate and non-inferiority in relation to placebo in terms of diarrhoea incidence and prevalence. Secondary objectives are mechanistic assessments, to test whether IHAT reduces the burden of enteric pathogens, morbidity, and intestinal inflammation, and that it does not cause detrimental changes to the gut microbiome, particularly in relation to Lactobacillaceae, Bifidobacteriaceae and Enterobacteriaceae. Discussion: This trial will test the hypothesis that supplementation with IHAT eliminates iron deficiency and improves haemoglobin levels without inducing gastrointestinal adverse effects. If shown to be the case, this would open the possibility for further testing and use of IHAT as a novel iron source for micronutrient intervention strategies in resource-poor countries, with the ultimate aim to help reduce the IDA global burden. Registration: This trial is registered at clinicaltrials.gov ( NCT02941081)

Prevalence of Panton-Valentine Leukocidin (PVL) and Antimicrobial Resistance in Community-Acquired Clinical Staphylococcus aureus in an Urban Gambian Hospital : A 11-Year Period Retrospective Pilot Study

This work is supported by MRCG. The source of funding has no role in the design and writing of the manuscript.Peer reviewedPublisher PD

Etiology of Pediatric Meningitis in West Africa Using Molecular Methods in the Era of Conjugate Vaccines against Pneumococcus, Meningococcus, and Haemophilus influenzae Type b.

Despite the implementation of effective conjugate vaccines against the three main bacterial pathogens that cause meningitis, Streptococcus pneumoniae, Haemophilus influenzae type b (Hib), and Neisseria meningitidis serogroup A, the burden of meningitis in West Africa remains high. The relative importance of other bacterial, viral, and parasitic pathogens in central nervous system infections is poorly characterized. Cerebrospinal fluid (CSF) specimens were collected from children younger than 5 years with suspected meningitis, presenting at pediatric teaching hospitals across West Africa in five countries including Senegal, Ghana, Togo, Nigeria, and Niger. Cerebrospinal fluid specimens were initially tested using bacteriologic culture and a triplex real-time polymerase chain reaction (PCR) assay for N. meningitidis, S. pneumoniae, and H. influenzae used in routine meningitis surveillance. A custom TaqMan Array Card (TAC) assay was later used to detect 35 pathogens including 15 bacteria, 17 viruses, one fungus, and two protozoans. Among 711 CSF specimens tested, the pathogen positivity rates were 2% and 20% by the triplex real-time PCR (three pathogens) and TAC (35 pathogens), respectively. TAC detected 10 bacterial pathogens, eight viral pathogens, and Plasmodium. Overall, Escherichia coli was the most prevalent (4.8%), followed by S. pneumoniae (3.5%) and Plasmodium (3.5%). Multiple pathogens were detected in 4.4% of the specimens. Children with human immunodeficiency virus (HIV) and Plasmodium detected in CSF had high mortality. Among 220 neonates, 17% had at least one pathogen detected, dominated by gram-negative bacteria. The meningitis TAC enhanced the detection of pathogens in children with meningitis and may be useful for case-based meningitis surveillance

Geospatial analysis and impact of targeted development of breast cancer care in The Gambia: a cross-sectional study

Background The Gambia has one of the lowest survival rates for breast cancer in Africa. Contributing factors are late presentation, delays within the healthcare system, and decreased availability of resources. We aimed to characterize the capacity and geographic location of healthcare facilities in the country and calculate the proportion of the population with access to breast cancer care. Methods A facility-based assessment tool was administered to secondary and tertiary healthcare facilities and private medical centers and clinics in The Gambia. GPS coordinates were obtained, and proximity of service availability and population analysis were performed. Distance thresholds of 10, 20, and 45 km were chosen to determine access to screening, pathologic diagnosis, and surgical management. An additional population analysis was performed to observe the potential impact of targeted development of resources for breast cancer care. Results All 102 secondary and tertiary healthcare facilities and private medical centers and clinics in The Gambia were included. Breast cancer screening is mainly performed through clinical breast examination and is available in 52 facilities. Seven facilities provide pathologic diagnosis and surgical management of breast cancer. The proportion of the Gambian population with access to screening, pathologic diagnosis, and surgical management is 72, 53, and 62%, respectively. A hypothetical targeted expansion of resources would increase the covered population to 95, 62, and 84%. Conclusions Almost half of the Gambian population does not have access to pathologic diagnosis and surgical management of breast cancer within the distance threshold utilized in the study. Mapping and population analysis can identify areas for targeted development of resources to increase access to breast cancer care

SARS-CoV-2 seroprevalence in pregnant women during the first three COVID-19 waves in The Gambia

OBJECTIVES: SARS-CoV-2 transmission in Sub-Saharan Africa has probably been underestimated. Population-based seroprevalence studies are needed to determine the extent of transmission in the continent. METHODS: Blood samples from a cohort of Gambian pregnant women were tested for SARS-CoV-2 total receptor binding domain (RBD) IgM/IgG before (Pre-pandemic: October-December 2019), and during the pandemic (Pre-wave1: February-June 2020; Post-wave1: October-December 2020, Post-wave2: May-June 2021; and Post-wave3: October-December 2021). Samples reactive for SARS-CoV-2 total RBD IgM/IgG were tested in specific S1- and nucleocapsid (NCP) IgG assays. RESULTS: SARS-CoV-2 total RBD IgM/IgG seroprevalence was 0.9% 95%CI (0.2, 4.9) in Pre-pandemic; 4.1% (1.4, 11.4) in Pre-wave1; 31.1% (25.2, 37.7) in Post-wave1; 62.5% (55.8, 68.8) in Post-wave2 and 90.0% (85.1, 93.5) in Post-wave3. S-protein IgG and NCP-protein IgG seroprevalence also increased at each Post-wave period. Although S-protein IgG and NCP-protein IgG seroprevalence was similar at Post-wave1, S-protein IgG seroprevalence was higher at Post-wave2 and Post-wave3, [prevalence difference (PD) 13.5 (0.1, 26.8) and prevalence ratio (PR) 1.5 (1.0, 2.3) in Post-wave2; and 22.9 (9.2, 36.6) and 1.4 (1.1, 1.8) in Post-wave3 respectively, p<0.001]. CONCLUSION: SARS-CoV-2 transmission in The Gambia during the first three COVID-19 waves was high, differing significantly from official numbers of COVID-19 cases reported. Our findings are important for policy makers in managing the near-endemic COVID-19

The effect of live attenuated influenza vaccine on pneumococcal colonisation densities among children aged 24-59 months in The Gambia: a phase 4, open label, randomised, controlled trial.

BACKGROUND: Influenza and other respiratory viruses promote Streptococcus pneumoniae proliferation in the upper respiratory tract. We sought to investigate for what we believe is the first time, the effect of intranasal live attenuated influenza vaccine (LAIV) on nasopharyngeal S pneumoniae density in a low-income to middle-income country population with high pneumococcal carriage rates. METHODS: In an open-label, randomised, controlled trial in The Gambia, 330 healthy children aged 24-59 months were randomly assigned 2:1 to receive one trivalent LAIV dose at enrolment (day 0, intervention) or at the end of active follow-up (day 21, control). The investigator team were initially masked to block size and randomisation sequence to avoid allocation bias. Group allocation was later revealed to the investigator team. The primary outcome was PCR-quantified day 7 and 21 pneumococcal density. Asymptomatic respiratory viral infection at baseline and LAIV strain shedding were included as covariates in generalised mixed-effects models, to assess the effect of LAIV and other variables on pneumococcal densities. The study is registered at ClinicalTrials.gov, NCT02972957, and is closed to recruitment. FINDINGS: Between Feb 8 and April 12, 2017, and Jan 15 and March 28, 2018, of 343 children assessed for eligibility, 213 in the intervention group and 108 in the control group completed the study and were included in the final analysis. Although no significant differences were seen in pneumococcal carriage or density at each timepoint when comparing groups, changes from baseline were observed in the LAIV group. The baseline S pneumoniae carriage prevalence was high in both LAIV and control groups (75%) and increased by day 21 in the LAIV group (85%, p=0·0037), but not in the control group (79%, p=0·44). An increase in pneumococcal density from day 0 amounts was seen in the LAIV group at day 7 (+0·207 log10 copies per μL, SE 0·105, p=0·050) and day 21 (+0·280 log10 copies per μL, SE 0·105, p=0·0082), but not in the control group. Older age was associated with lower pneumococcal density (-0·015 log10 copies per μL, SE 0·005, p=0·0030), with the presence of asymptomatic respiratory viruses at baseline (+0·259 log10 copies per μL, SE 0·097, p=0·017), and greater LAIV shedding at day 7 (+0·380 log10 copies per μL, SE 0·167, p=0·024) associated with higher pneumococcal density. A significant increase in rhinorrhoea was reported in the LAIV group compared with the control group children during the first 7 days of the study (103 [48%] of 213, compared with 25 [23%] of 108, p<0·0001), and between day 7 and 21 (108 [51%] of 213, compared with 28 [26%] of 108, p<0·0001). INTERPRETATION: LAIV was associated with a modest increase in nasopharyngeal pneumococcal carriage and density in the 21 days following vaccination, with the increase in density lower in magnitude than previously described in the UK. This increase was accelerated when LAIV was administered in the presence of pre-existing asymptomatic respiratory viruses, suggesting that nasopharyngeal S pneumoniae proliferation is driven by cumulative mixed-viral co-infections. The effect of LAIV on pneumococcal density is probably similar to other respiratory viral infections in children. Our findings provide reassurance for the use of LAIV to expand influenza vaccine programmes in low-income to middle-income country populations with high pneumococcal carriage. FUNDING: Wellcome Trust