Draft Genome Sequence of an Enterotoxigenic Escherichia coli Strain Carrying Genes for Colonization Surface Antigen 13 and a Heat-Labile Toxin.

Here, we report the draft genome of ESEI_597, an enterotoxigenic Escherichia coli (ETEC) strain harboring genes encoding colonization surface antigen 13 (CS13) and a heat-labile toxin. The ESEI_597 strain was isolated from an 8-month-old child living in Korogocho, Kenya, in 2013.

Differences in genetic flux in invasive Streptococcus pneumoniae associated with bacteraemia and meningitis.

Background: Genetic flux, a crucial process of pneumococcal evolution, is an essential aspect of bacterial physiology during human pathogenesis. However, the role of these genetic changes and the selective forces that drive them is not fully understood. Elucidating the underlying selective forces that determine the magnitude and direction (gene gain or loss) of gene transfer is important for better understanding the pathogenesis process, and may also highlight potential therapeutic and diagnostic targets. Methods: Here, we leveraged data from high throughput genome sequencing and robust probabilistic models to discover the magnitude and likely direction of genetic flux events, but not the source, in 209 multi-lineage invasive pneumococcal genomes generated from blood (n = 147) and CSF (n = 62) isolates, associated with bacteremia and meningitis respectively. The Gain and Loss Mapping Engine (GLOOME) was used to infer gene gain and loss more accurately by taking into account differences in rates of gene gain and loss among gene families, as well as independent evolution within and across lineages. Results: Our results show the likely extent and direction of gene fluctuations at different niche, during pneumococcal pathogenesis, highlighting that evolutionary dynamics are important for tissue-specific host invasion and survival. Conclusion: These findings improve insights on evolutionary dynamics during invasive pneumococcal disease, and highlight potential diagnostic and therapeutic targets.

COVID-19 risk factors: The curious case of Africa's governance and preparedness.

COVID-19 is now established in Africa, and requires appropriate prioritization of resources and customized control measures. Although there have been lower than predicted number of COVID-19 cases (6,839,159 (3.4% of global cases)) and fatalities (172,413 (4.1% of cases in Africa)) compared to global estimates from other regions, as of 3rd August 2021. The World Health Organization (WHO) has warned that poor mitigation strategies could worsen the current situation in African countries. Several aspects have been attributed to the lower COVID-19 magnitude observed in Africa that include: warmer climate, a youthful population, and previous experience in managing infectious diseases. However, the level of COVID-19 risk of exposure and vulnerability to develop complications varies greatly across the continent. At present, most COVID-19 disease trajectories have been predicted using mathematical models focused solely on demographic factors. We compared the global health security and governance indices in forty-one African countries, with a population of more than 2.5 million, and matched these to the magnitude of COVID-19 burden, to establish whether there is correlation. These findings suggests that more accurate comparisons of responses to COVID-19 can only be made within clusters of African countries that share similar governance and preparedness standards. We conclude that proper implementation of the continental framework on disaster preparedness and management is required for management of the COVID-19 pandemic.

Antimicrobial Resistance Profiles and Genes of Staphylococci Isolated from Mastitic Cow's Milk in Kenya.

Increasing numbers of potentially zoonotic multidrug-resistant (MDR) staphylococci strains, associated with mastitis in dairy cows, are being reported globally and threaten disease management in both animal and human health. However, the prevalence and antimicrobial resistance profiles of these strains, including methicillin-resistant staphylococci (MRS), in Kenya is not well known. This study investigated the drug resistance profiles and genes carried by 183 staphylococci isolates from 142 dairy cows representing 93 farms recovered from mastitis milk of dairy cows in two selected counties in Kenya. Staphylococci isolates were characterized by phenotypic characteristics, polymerase chain reaction (PCR) amplification, partial sequencing and susceptibility testing for 10 antimicrobial drugs. Detection of seven resistance genes to the various antimicrobial drugs was conducted using PCR. Overall, phenotypic resistance among the staphylococci ranged between 66.1% for ampicillin and 3.5% for fluoroquinolones. Twenty-five percent (25%) of S. aureus and 10.8% of the coagulase-negative staphylococci (CoNS) isolates, were methicillin-resistant staphylococci phenotypically (defined as resistance to cefoxitin disk diffusion). The most common genes found in S. aureus and CoNS were blaZ and strB at 44.3% and 26%, and 78% and 50%, respectively. MDR was observed in 29.67% and 16.3% of S. aureus and CoNS, respectively. These findings pose a threat to bovine mastitis treatment and management as well as human health.

Kenya's response to the COVID-19 pandemic: a balance between minimising morbidity and adverse economic impact.

Coronavirus disease 2019 (COVID-19) has ravaged the world's socioeconomic systems forcing many governments across the globe to implement unprecedented stringent mitigation measures to restrain its rapid spread and adverse effects. A disproportionate number of COVID-19 related morbidities and mortalities were predicted to occur in Africa. However, Africa still has a lower than predicted number of cases, 4% of the global pandemic burden. In this open letter, we highlight some of the early stringent countermeasures implemented in Kenya, a sub-Saharan African country, to avert the severe effects of the COVID-19 pandemic. These mitigation measures strike a balance between minimising COVID-19 associated morbidity and fatalities and its adverse economic impact, and taken together have significantly dampened the pandemic's impact on Kenya's populace.

Detecting bracoviral orthologs distribution in five tsetse fly species and the housefly genomes.

OBJECTIVE: Mutualism between endogenous viruses and eukaryotes is still poorly understood. Several endogenous double-stranded polydnaviruses, bracoviruses, homologous to those present in parasitic braconid wasp genomes were detected in the tsetse fly (Glossina morsitans morsitans). This is peculiar since tsetse flies do not share a reproductive lifestyle similar to wasps, but deliver fully developed larvae that pupate within minutes of exiting their mothers. The objective of this study is to investigate genomic distribution of bracoviral sequences in five tsetse fly species and the housefly, and examine its value as a potential vector control strategy target point. We use comparative genomics to determine the presence, distribution across Glossina species genomes, and evolutionary relationships of bracoviruses of five tsetse fly species and the housefly. RESULTS: We report on homologous bracoviruses in multiple Dipteran genomes. Phylogenetic reconstruction using within-species concatenated bracoviral orthologs shows great congruence with previously reconstructed insect species phylogenies. Our findings suggest that bracoviruses present in Diptera originate from a single integration event of the viral genome that occurred in an ancestor insect before the evolutionary radiation of different insect orders.

Prevalence of trypanosomes associated with drug resistance in Shimba Hills, Kwale County, Kenya.

OBJECTIVE: Animal African trypanosomiasis (AAT) is a life-threatening vector-borne disease, caused by trypanosome parasites, which are principally transmitted by tsetse flies. In Kenya, the prevalence of drug-resistant trypanosomes in endemic regions remains poorly understood. The objective of this study was to establish AAT point prevalence, drug susceptibility of associated trypanosomes, and measure infectivity by multiple AAT mammalian hosts to tsetse flies in Shimba hills, a resource-poor region with high bovine trypanosomiasis prevalence and morbidity rates at the coast of Kenya. We collected tsetse flies using traps (1 Ngu and 2 biconical), and then sorted them on sex and species. Trypanosomes present in tsetse flies were detected by first extracting all genomic DNA, and then performing PCR reactions with established primers of the internal transcribed spacer regions. Polymorphisms associated with trypanocide resistance in the TbAT1 gene were also detected by performing PCR reactions with established primers. RESULTS: Our findings suggest low trypanosome prevalence (3.7%), low trypanocide resistance, and low infectivity by multiple mammalian hosts to tsetse flies in Shimba hills. We conclude that enhanced surveillance is crucial for informing disease management practices that help prevent the spread of drug-resistant trypanosomiasis.

Community-led data collection using Open Data Kit for surveillance of animal African trypanosomiasis in Shimba hills, Kenya.

OBJECTIVE: In Sub-Saharan Africa, there is an increase in trypanosome non-susceptibility to multiple trypanocides, but limited information on judicious trypanocide use is accessible to smallholder farmers and agricultural stakeholders in disease endemic regions, resulting in widespread multi-drug resistance. Huge economic expenses and the laborious nature of extensive field studies have hindered collection of the requisite large-scale prospective datasets required to inform disease management. We examined the efficacy of community-led data collection strategies using smartphones by smallholder farmers to acquire robust datasets from the trypanosomiasis endemic Shimba hills region in Kenya. We used Open Data Kit, an open-source smartphone application development software, to create a data collection App. RESULTS: Our study provides proof of concept for the viability of using smartphone Apps to remotely collect reliable large-scale information from smallholder farmers and veterinary health care givers in resource poor settings. We show that these datasets can be reliably collated remotely, analysed, and the findings can inform policies that improve farming practices and economic wellbeing while restricting widespread multi-drug resistance. Moreover, this strategy can be used to monitor and manage other infectious diseases in other rural, resource poor settings.

Modelling appropriate use of trypanocides to restrict wide-spread multi-drug resistance during chemotherapy of animal African trypanosomiasis.

Trypanocide resistance remains a huge challenge in the management of animal African trypanosomiasis. Paucity of data on the prevalence of multi-drug resistant trypanosomes has greatly hindered optimal veterinary management practices. We use mathematical model predictions to highlight appropriate drug regimens that impede trypanocide resistance development in cattle. We demonstrate that using drugs in decreasing resistance order results in a negligible increase in number of cattle with resistant infection, in contrast to a more pronounced increase from trypanocide use in increasing resistance order. We demonstrate that the lowest levels of trypanocide resistance are achieved with combination therapy. We also show that increasing the number of cattle treated leads to a progressive reduction in the number of cattle with drug resistant infections for treatments of up to 80% of the cattle population for the combination treatment strategy. Our findings provide an initial evidence-based framework on some essential practices that promote optimal use of the handful of trypanocides. We anticipate that our modest forecasts will improve therapeutic outcomes by appropriately informing on the best choice, and combination of drugs that minimize treatment failure rates.

Importance of human demographic history knowledge in genetic studies involving multi-ethnic cohorts.

Paucity of data from African populations due to under-representation in human genetic studies has impeded detailed understanding of the heritable human genome variation. This is despite the fact that Africa has sizeable genetic, cultural and linguistic diversity. There are renewed efforts to understand health problems relevant to African populations using more comprehensive datasets, and by improving expertise in health-related genomics among African scientists. We emphasise that careful consideration of the sampled populations from national and within-continental cohorts in large multi-ethnic genetic research efforts is required to maximise the prospects of identifying and fine-mapping novel risk variants in indigenous populations. We caution that human demographic history should be taken into consideration in such prospective genetic-association studies.

Peripheral blood RNA gene expression in children with pneumococcal meningitis: a prospective case-control study.

INTRODUCTION: Invasive pneumococcal disease (IPD), caused by Streptococcus pneumoniae, is a leading cause of pneumonia, meningitis and septicaemia worldwide, with increased morbidity and mortality in HIV-infected children. OBJECTIVES: We aimed to compare peripheral blood expression profiles between HIV-infected and uninfected children with pneumococcal meningitis and controls, and between survivors and non-survivors, in order to provide insight into the host inflammatory response leading to poorer outcomes. DESIGN AND SETTING: Prospective case-control observational study in a tertiary hospital in Malawi. PARTICIPANTS: Children aged 2 months to 16 years with pneumococcal meningitis or pneumonia. METHODS: We used the human genome HGU133A Affymetrix array to explore differences in gene expression between cases with pneumococcal meningitis (n=12) and controls, and between HIV-infected and uninfected cases, and validated gene expression profiles for 34 genes using real-time quantitative PCR (RT-qPCR) in an independent set of cases with IPD (n=229) and controls (n=13). Pathway analysis was used to explore genes differentially expressed. RESULTS: Irrespective of underlying HIV infection, cases showed significant upregulation compared with controls of the following: S100 calcium-binding protein A12 (S100A12); vanin-1 (VNN1); arginase, liver (ARG1); matrix metallopeptidase 9 (MMP9); annexin A3 (ANXA3); interleukin 1 receptor, type II (IL1R2); CD177 molecule (CD177); endocytic adaptor protein (NUMB) and S100 calcium-binding protein A9 (S100A9), cytoskeleton-associated protein 4 (CKAP4); and glycogenin 1 (GYG1). RT-qPCR confirmed differential expression in keeping with microarray results. There was no differential gene expression in HIV-infected compared with HIV-uninfected cases, but there was significant upregulation of folate receptor 3 (FOLR3), S100A12 in survivors compared with non-survivors. CONCLUSION: Children with IPD demonstrated increased expression in genes regulating immune activation, oxidative stress, leucocyte adhesion and migration, arginine metabolism, and glucocorticoid receptor signalling.

Comparative genomics identifies male accessory gland proteins in five Glossina species.

Accessory gland proteins (ACPs) are important reproductive proteins produced by the male accessory glands (MAGs) of most insect species. These proteins are essential for male insect fertility, and are transferred alongside semen to females during copulation. ACPs are poorly characterized in Glossina species (tsetse fly), the principal vector of the parasite that causes life-threatening Human African Trypanosomiasis and Animal trypanosomiasis in endemic regions in Africa. The tsetse fly has a peculiar reproductive cycle because of the absence of oviposition. Females mate once and store sperm in a spermathecal, and produce a single fully developed larva at a time that pupates within minutes of exiting their uterus. This slow reproductive cycle, compared to other insects, significantly restricts reproduction to only 3 to 6 larvae per female lifespan. This unique reproductive cycle is an attractive vector control strategy entry point. We exploit comparative genomics approaches to explore the diversity of ACPs in the recently available whole genome sequence data from five tsetse fly species ( Glossina morsitans, G. austeni, G. brevipalpis, G. pallidipes and G. fuscipes). We used previously described ACPs in Drosophila melanogaster and Anopheles gambiae as reference sequences. We identified 36, 27, 31, 29 and 33 diverse ACP orthologous genes in G. austeni, G. brevipalpis, G. fuscipes, G. pallidipes and G. morsitans genomes respectively, which we classified into 21 functional classes. Our findings provide genetic evidence of MAG proteins in five recently sequenced Glossina genomes. It highlights new avenues for molecular studies that evaluate potential field control strategies of these important vectors of human and animal disease.

Comparative Genomic Analysis of Meningitis- and Bacteremia-Causing Pneumococci Identifies a Common Core Genome.

Streptococcus pneumoniae is a nasopharyngeal commensal that occasionally invades normally sterile sites to cause bloodstream infection and meningitis. Although the pneumococcal population structure and evolutionary genetics are well defined, it is not clear whether pneumococci that cause meningitis are genetically distinct from those that do not. Here, we used whole-genome sequencing of 140 isolates of S. pneumoniae recovered from bloodstream infection (n = 70) and meningitis (n = 70) to compare their genetic contents. By fitting a double-exponential decaying-function model, we show that these isolates share a core of 1,427 genes (95% confidence interval [CI], 1,425 to 1,435 genes) and that there is no difference in the core genome or accessory gene content from these disease manifestations. Gene presence/absence alone therefore does not explain the virulence behavior of pneumococci that reach the meninges. Our analysis, however, supports the requirement of a range of previously described virulence factors and vaccine candidates for both meningitis- and bacteremia-causing pneumococci. This high-resolution view suggests that, despite considerable competency for genetic exchange, all pneumococci are under considerable pressure to retain key components advantageous for colonization and transmission and that these components are essential for access to and survival in sterile sites.

High multiple carriage and emergence of Streptococcus pneumoniae vaccine serotype variants in Malawian children.

BACKGROUND: Carriage of either single or multiple pneumococcal serotypes (multiple carriage) is a prerequisite for developing invasive pneumococcal disease. However, despite the reported high rates of pneumococcal carriage in Malawi, no data on carriage of multiple serotypes has been reported previously. Our study provides the first description of the prevalence of multiple pneumococcal carriage in Malawi. METHODS: The study was conducted in Blantyre and Karonga districts in Malawi, from 2008 to 2012. We recruited 116 children aged 0-13 years. These children were either HIV-infected (N = 44) or uninfected (N = 72). Nasopharyngeal samples were collected using sterile swabs. Pneumococcal serotypes in the samples were identified by microarray. Strains that could not be typed by microarray were sequenced to characterise possible genetic alterations within the capsular polysaccharide (CPS) locus. RESULTS: The microarray identified 179 pneumococcal strains (from 116 subjects), encompassing 43 distinct serotypes and non-typeable (NT) strains. Forty per cent (46/116) of children carried multiple serotypes. Carriage of vaccine type (VT) strains was higher (p = 0.028) in younger (0-2 years) children (71 %, 40/56) compared to older (3-13 years) children (50 %, 30/60). Genetic variations within the CPS locus of known serotypes were observed in 19 % (34/179) of the strains identified. The variants included 13-valent pneumococcal conjugate vaccine (PCV13) serotypes 6B and 19A, and the polysaccharide vaccine serotype 20. Serotype 6B variants were the most frequently isolated (47 %, 16/34). Unlike the wild type, the CPS locus of the 6B variants contained an insertion of the licD-family phosphotransferase gene. The CPS locus of 19A- and 20-variants contained an inversion in the sugar-biosynthesis (rmlD) gene and a 717 bp deletion within the transferase (whaF) gene, respectively. CONCLUSIONS: The high multiple carriage in Malawian children provides opportunities for genetic exchange through horizontal gene transfer. This may potentially lead to CPS locus variants and vaccine escape. Variants reported here occurred naturally, however, PCV13 introduction could exacerbate the CPS genetic variations. Further studies are therefore recommended to assess the invasive potential of these variants and establish whether PCV13 would offer cross-protection. We have shown that younger children (0-2 years) are a reservoir of VT serotypes, which makes them an ideal target for vaccination.

Genetic diversity of the Pneumococcal CbpA: Implications for next-generation vaccine development.

Pneumococci are capable of vaccine escape by genetic recombination at the targeted capsular locus, significantly reducing long-term vaccine effectiveness. Recently, efforts have been redirected to understanding pneumococcal biology related to potential next-generation vaccine candidates. A variety of serotype-independent protein antigens capable of inducing protective immune responses in tissue culture and animal models of infection have been identified. However, ideal vaccine candidates that are conserved across all genotypes, provide broad population coverage, and induce T-cell dependent immune responses are still under investigation. We examined whether immune responses due to the highly polymorphic CbpA antigen are due to a conserved domain capable of evoking specific immune "memory" across all genotypes of pneumococci. We defined the genotypes in a global dataset of 213 pneumococcal isolates. This isolate collection was genotypically diverse and ideal for establishing the presence of conserved CbpA epitopes as potential protein vaccine candidates. Examination of the CbpA locus sequence was highly polymorphic at both the nucleic acid and amino acid level. Despite this high polymorphism some domains are broadly conserved and consist of different amino acid residues with the same physicochemical properties, and therefore have similar tertiary structures. The two most common domains identified in the CbpA gene are modular teichoic acid phosphorylcholine esterase Pce (2bib:A), and R2 domain (1w9r:A). These conserved domains are immunogenic, therefore capable of inducing long-term host immune responses; moreover they are extracellularly located and thus accessible. We proposed their evaluation as suitable next-generation CbpA-fusion protein vaccine candidates.

BMX: a tool for computing bacterial phyletic composition from orthologous maps.

BACKGROUND: New sequencing technologies have made it possible to explore genetic diversity at higher resolution in microbial populations. However, our understanding evolutionary relationships, and comparison of closely and distantly related bacterial genomes from these massive datasets remains a formidable challenge. Numerous clustering algorithms that group genomic data based on homology have been developed, but new tools are still required to analyse the resultant orthologous maps to understand functional genetic similarities and their phyletic patterns (patterns of presence of absence of genes). FINDINGS: Bacterial Makeup eXplorer (BMX) implements an algorithm that swiftly and efficiently facilitates the determination of the number of orthologs in prokaryotic genomes employing a reference free approach, which may be further exploited to transfer of gene annotations. BMX is able to integrate orthologous maps of highly diverse prokaryotic genomes therefore making it possible to perform robust and scalable, multi-platform, high quality annotation transfer and gene-by-gene composition assessment method. In addition results are presented in the form of publication quality figures. CONCLUSIONS: BMX allows extensive data analysis of orthologous map databases to understand underlying biological relationships. Furthermore, BMX is portable across different platforms and can be installed easily. In summary, BMX allows higher resolution analysis of genomes from diverse bacterial populations.

Population genetic structure of Streptococcus pneumoniae in Kilifi, Kenya, prior to the introduction of pneumococcal conjugate vaccine.

BACKGROUND: The 10-valent pneumococcal conjugate vaccine (PCV10) was introduced in Kenya in 2011. Introduction of any PCV will perturb the existing pneumococcal population structure, thus the aim was to genotype pneumococci collected in Kilifi before PCV10. METHODS AND FINDINGS: Using multilocus sequence typing (MLST), we genotyped >1100 invasive and carriage pneumococci from children, the largest collection genotyped from a single resource-poor country and reported to date. Serotype 1 was the most common serotype causing invasive disease and was rarely detected in carriage; all serotype 1 isolates were members of clonal complex (CC) 217. There were temporal fluctuations in the major circulating sequence types (STs); and although 1-3 major serotype 1, 14 or 23F STs co-circulated annually, the two major serotype 5 STs mainly circulated independently. Major STs/CCs also included isolates of serotypes 3, 12F, 18C and 19A and each shared ≤ 2 MLST alleles with STs that circulate widely elsewhere. Major CCs associated with non-PCV10 serotypes were predominantly represented by carriage isolates, although serotype 19A and 12F CCs were largely invasive and a serotype 10A CC was equally represented by invasive and carriage isolates. CONCLUSIONS: Understanding the pre-PCV10 population genetic structure in Kilifi will allow for the detection of changes in prevalence of the circulating genotypes and evidence for capsular switching post-vaccine implementation.

Piliation of invasive Streptococcus pneumoniae isolates in the era before pneumococcal conjugate vaccine introduction in Malawi.

The pneumococcal pilus has been shown to be an important determinant of adhesion and virulence in mouse models of colonization, pneumonia, and bacteremia. A pilus is capable of inducing protective immunity, supporting its inclusion in next-generation pneumococcal protein vaccine formulations. Whether this vaccine target is common among pneumococci in sub-Saharan Africa is uncertain. To define the prevalence and genetic diversity of type I and II pili among invasive pneumococci in Malawi prior to the introduction of the 13-valent pneumococcal conjugate vaccine (PCV13) into routine childhood immunization, we examined 188 Streptococcus pneumoniae isolates collected between 2002 and 2008 (17% serotype 1). In this region of high disease burden, we found a low frequency of invasive piliated pneumococci (14%) and pilus gene sequence diversity similar to that seen previously in multiple global pneumococcal lineages. All common serotypes with pilus were covered by PCV13 and so we predict that pilus prevalence will be reduced in the Malawian pneumococcal population after PCV13 introduction.