Open Science in Kenya: Where Are We?

According to the United Nations Educational, Scientific, and Cultural Organization (UNESCO), Open Science is the movement to make scientific research and data accessible to all. It has great potential for advancing science. At its core, it includes (but is not limited to) open access, open data, and open research. Some of the associated advantages are promoting collaboration, sharing and reproducibility in research, and preventing the reinvention of the wheel, thus saving resources. As research becomes more globalized and its output grows exponentially, especially in data, the need for open scientific research practices is more evident - the future of modern science. This has resulted in a concerted global interest in open science uptake. Even so, barriers still exist. The formal training curriculum in most, if not all, universities in Kenya does not equip students with the knowledge and tools to subsequently practice open science in their research. Therefore, to work openly and collaboratively, there is a need for awareness and training in the use of open science tools. These have been neglected, especially in most developing countries, and remain barriers to the cause. Moreover, there is scanty research on the state of affairs regarding the practice and/or adoption of open science. Thus, we developed, through the OpenScienceKE framework, a model to narrow the gap. A sensitize-train-hack-collaborate model was applied in Nairobi, the economic and administrative capital of Kenya. Using the model, we sensitized through seminars, trained on the use of tools through workshops, applied the skills learned in training through hackathons to collaboratively answer the question on the state of open science in Kenya. While the former parts of the model had 20-50 participants, the latter part mainly involved participants with a bioinformatics background, leveraging their advanced computational skills. This model resulted in an open resource that researchers can use to publish as open access cost-effectively. Moreover, we observed a growing interest in open science practices in Kenya through literature search and data mining and that lack of awareness and skills may still hinder the adoption and practice of open science. Furthermore, at the time of the analyses, we surprisingly found that out of the 20,069 papers downloaded from BioRXiv, only 18 had Kenyan authors, a majority of which are international (16) collaborations. This may suggest poor uptake of the use of preprints among Kenyan researchers. The findings in this study highlight the state of open science in Kenya and challenges facing its adoption and practice while bringing forth possible areas for primary consideration in the campaign toward open science. It also proposes a model (sensitize-train-hack-collaborate model) that may be adopted by researchers, funders and other proponents of open science to address some of the challenges faced in promoting its adoption in Kenya.

Ehrlichia spp. close to Ehrlichia ruminantium, Ehrlichia canis, and "Candidatus Ehrlichia regneryi" linked to heartwater-like disease in Kenyan camels (Camelus dromedarius).

We present findings from an outbreak of a heartwater-like disease in camels that killed at least 2000 adult animals in Kenya in 2016. Clinical signs included excitability, head pressing, aimless wandering, recumbency, and fast breathing followed by death after about 4 days. The observed morbidity in one herd was 40% with an average mortality of 7.5% in animals that received early antibiotic treatments. In untreated adults, the case fatality rate reached 100%. Gross pathology showed pulmonary edema, pleural exudate, hydrothorax, hydropericardium, ascites, enlarged "cooked" liver, nephrosis, and blood in the abomasum and intestine. Using established PCR-based protocols for tick-borne pathogens, a sequence close to Ehrlichia regneryi and Ehrlichia canis amplified in blood from two sick camels. We also amplified an Ehrlichia sp. sequence close to Ehrlichia ruminantium Welgevonden from a pool of Amblyomma spp. ticks collected from a sick camel and in a pool of Rhipicephalus spp. ticks from healthy camels.

Pathogens, endosymbionts, and blood-meal sources of host-seeking ticks in the fast-changing Maasai Mara wildlife ecosystem.

The role of questing ticks in the epidemiology of tick-borne diseases in Kenya's Maasai Mara National Reserve (MMNR), an ecosystem with intensified human-wildlife-livestock interactions, remains poorly understood. We surveyed the diversity of questing ticks, their blood-meal hosts, and tick-borne pathogens to understand potential effects on human and livestock health. By flagging and hand-picking from vegetation in 25 localities, we collected 1,465 host-seeking ticks, mostly Rhipicephalus and Amblyomma species identified by morphology and molecular analysis. We used PCR with high-resolution melting (HRM) analysis and sequencing to identify Anaplasma, Babesia, Coxiella, Ehrlichia, Rickettsia, and Theileria pathogens and blood-meal remnants in 231 tick pools. We detected blood-meals from humans, wildebeest, and African buffalo in Rh. appendiculatus, goat in Rh. evertsi, sheep in Am. gemma, and cattle in Am. variegatum. Rickettsia africae was detected in Am. gemma (MIR = 3.10) that had fed on sheep and in Am. variegatum (MIR = 250) that had fed on cattle. We found Rickettsia spp. in Am. gemma (MIR = 9.29) and Rh. evertsi (MIR = 200), Anaplasma ovis in Rh. appendiculatus (MIR = 0.89) and Rh. evertsi (MIR = 200), Anaplasma bovis in Rh. appendiculatus (MIR = 0.89), and Theileria parva in Rh. appendiculatus (MIR = 24). No Babesia, Ehrlichia, or Coxiella pathogens were detected. Unexpectedly, species-specific Coxiella sp. endosymbionts were detected in all tick genera (174/231 pools), which may affect tick physiology and vector competence. These findings show that ticks from the MMNR are infected with zoonotic R. africae and unclassified Rickettsia spp., demonstrating risk of African tick-bite fever and other spotted-fever group rickettsioses to locals and visitors. The protozoan pathogens identified may also pose risk to livestock production. The diverse vertebrate blood-meals of questing ticks in this ecosystem including humans, wildlife, and domestic animals, may amplify transmission of tick-borne zoonoses and livestock diseases.

Arboviruses and Blood Meal Sources in Zoophilic Mosquitoes at Human-Wildlife Interfaces in Kenya.

Background: Zoophilic mosquitoes play an important role in the transmission of arboviruses of medical importance at human-wildlife interfaces, yet arbovirus surveillance efforts have been focused mostly on anthropophilic mosquitoes. Understanding the diversity of zoophilic mosquitoes and their associated feeding patterns and arboviruses can inform better vector control strategies. Materials and Methods: We morphologically identified mosquitoes collected from two game reserves in Kenya, the Maasai Mara National Reserve (MMNR) and locations near the Shimba Hills National Reserve (SHNR). Representative mosquitoes were also identified by cytochrome c oxidase subunit 1 (COI) barcode sequencing. In addition, we identified the vertebrate hosts of mosquito blood meals from the contents of each mosquito's abdomen by high-resolution melting (HRM) analysis and sequencing of COI, 16S ribosomal RNA, and cytochrome b gene PCR products. Similarly, mosquito arbovirus infections were identified by HRM analysis and sequencing of Alphavirus- and Flavivirus-specific RT-PCR products. Results: Of 2858 mosquitoes collected, 51 were engorged with blood meals from seven different vertebrate hosts, including humans, birds, domestic, and peridomestic animals and wildlife. Culex was the most abundant mosquito genus, with Culex pipiens being the most abundant species in both study regions. Among MMNR samples, we detected dengue serotype-2 virus (DENV-2) for the first time in Aedes tarsalis and Aedes tricholabis, as well as Sindbis virus in male Cx. pipiens. We also detected DENV-2 in Aedes aegypti sampled from locations near the SHNR. Human and diverse wildlife blood meals were identified, including bushbuck blood in the dengue-infected Ae. tarsalis and both human and hippopotamus blood in a single Eretmapodites chrysogaster mosquito. Conclusions: Our findings highlight the potential risk of sylvatic dengue and Sindbis transmission to humans by zoophilic mosquitoes at human-wildlife interfaces in Africa. Of specific importance, we provide evidence of sylvatic DENV-2 in Ae. tarsalis and Ae. tricholabis, representing potential new dengue vectors.

Three-gene PCR and high-resolution melting analysis for differentiating vertebrate species mitochondrial DNA for biodiversity research and complementing forensic surveillance.

Reliable molecular identification of vertebrate species from morphologically unidentifiable tissue is critical for the prosecution of illegally-traded wildlife products, conservation-based biodiversity research, and identification of blood-meal hosts of hematophagous invertebrates. However, forensic identification of vertebrate tissue relies on sequencing of the mitochondrial cytochrome oxidase I (COI) 'barcode' gene, which remains costly for purposes of screening large numbers of unknown samples during routine surveillance. Here, we adapted a rapid, low-cost approach to differentiate 10 domestic and 24 wildlife species that are common in the East African illegal wildlife products trade based on their unique high-resolution melting profiles from COI, cytochrome b, and 16S ribosomal RNA gene PCR products. Using the approach, we identified (i) giraffe among covertly sampled meat from Kenyan butcheries, and (ii) forest elephant mitochondrial sequences among savannah elephant reference samples. This approach is being adopted for high-throughput pre-screening of potential bushmeat samples in East African forensic science pipelines.

Vertical transmission of naturally occurring Bunyamwera and insect-specific flavivirus infections in mosquitoes from islands and mainland shores of Lakes Victoria and Baringo in Kenya.

BACKGROUND: Many arboviruses transmitted by mosquitoes have been implicated as causative agents of both human and animal illnesses in East Africa. Although epidemics of arboviral emerging infectious diseases have risen in frequency in recent years, the extent to which mosquitoes maintain pathogens in circulation during inter-epidemic periods is still poorly understood. This study aimed to investigate whether arboviruses may be maintained by vertical transmission via immature life stages of different mosquito vector species. METHODOLOGY: We collected immature mosquitoes (egg, larva, pupa) on the shores and islands of Lake Baringo and Lake Victoria in western Kenya and reared them to adults. Mosquito pools (≤25 specimens/pool) of each species were screened for mosquito-borne viruses by high-resolution melting analysis and sequencing of multiplex PCR products of genus-specific primers (alphaviruses, flaviviruses, phleboviruses and Bunyamwera-group orthobunyaviruses). We further confirmed positive samples by culturing in baby hamster kidney and Aedes mosquito cell lines and re-sequencing. PRINCIPAL FINDINGS: Culex univittatus (2/31pools) and Anopheles gambiae (1/77 pools) from the Lake Victoria region were positive for Bunyamwera virus, a pathogenic virus that is of public health concern. In addition, Aedes aegypti (3/50), Aedes luteocephalus (3/13), Aedes spp. (2/15), and Culex pipiens (1/140) pools were positive for Aedes flaviviruses at Lake Victoria, whereas at Lake Baringo, three pools of An. gambiae mosquitoes were positive for Anopheles flavivirus. These insect-specific flaviviruses (ISFVs), which are presumably non-pathogenic to vertebrates, were found in known medically important arbovirus and malaria vectors. CONCLUSIONS: Our results suggest that not only ISFVs, but also a pathogenic arbovirus, are naturally maintained within mosquito populations by vertical transmission, even in the absence of vertebrate hosts. Therefore, virus and vector surveillance, even during inter-epidemics, and the study of vector-arbovirus-ISFV interactions, may aid in identifying arbovirus transmission risks, with the potential to inform control strategies that lead to disease prevention.

Molecular Detection of Tick-Borne Pathogen Diversities in Ticks from Livestock and Reptiles along the Shores and Adjacent Islands of Lake Victoria and Lake Baringo, Kenya.

Although diverse tick-borne pathogens (TBPs) are endemic to East Africa, with recognized impact on human and livestock health, their diversity and specific interactions with tick and vertebrate host species remain poorly understood in the region. In particular, the role of reptiles in TBP epidemiology remains unknown, despite having been implicated with TBPs of livestock among exported tortoises and lizards. Understanding TBP ecologies, and the potential role of common reptiles, is critical for the development of targeted transmission control strategies for these neglected tropical disease agents. During the wet months (April-May; October-December) of 2012-2013, we surveyed TBP diversity among 4,126 ticks parasitizing livestock and reptiles at homesteads along the shores and islands of Lake Baringo and Lake Victoria in Kenya, regions endemic to diverse neglected tick-borne diseases. After morphological identification of 13 distinct Rhipicephalus, Amblyomma, and Hyalomma tick species, ticks were pooled (≤8 individuals) by species, host, sampling site, and collection date into 585 tick pools. By supplementing previously established molecular assays for TBP detection with high-resolution melting analysis of PCR products before sequencing, we identified high frequencies of potential disease agents of ehrlichiosis (12.48% Ehrlichia ruminantium, 9.06% Ehrlichia canis), anaplasmosis (6.32% Anaplasma ovis, 14.36% Anaplasma platys, and 3.08% Anaplasma bovis,), and rickettsiosis (6.15% Rickettsia africae, 2.22% Rickettsia aeschlimannii, 4.27% Rickettsia rhipicephali, and 4.95% Rickettsia spp.), as well as Paracoccus sp. and apicomplexan hemoparasites (0.51% Theileria sp., 2.56% Hepatozoon fitzsimonsi, and 1.37% Babesia caballi) among tick pools. Notably, we identified E. ruminantium in both Amblyomma and Rhipicephalus pools of ticks sampled from livestock in both study areas as well as in Amblyomma falsomarmoreum (66.7%) and Amblyomma nuttalli (100%) sampled from tortoises and Amblyomma sparsum (63.6%) sampled in both cattle and tortoises at Lake Baringo. Similarly, we identified E. canis in rhipicephaline ticks sampled from livestock and dogs in both regions and Amblyomma latum (75%) sampled from monitor lizards at Lake Victoria. These novel tick-host-pathogen interactions have implications on the risk of disease transmission to humans and domestic animals and highlight the complexity of TBP ecologies, which may include reptiles as reservoir species, in sub-Saharan Africa.

Arbovirus and insect-specific virus discovery in Kenya by novel six genera multiplex high-resolution melting analysis.

A broad diversity of arthropod-borne viruses (arboviruses) of global health concern are endemic to East Africa, yet most surveillance efforts are limited to just a few key viral pathogens. Additionally, estimates of arbovirus diversity in the tropics are likely to be underestimated as their discovery has lagged significantly over past decades due to limitations in fast and sensitive arbovirus identification methods. Here, we developed a nearly pan-arbovirus detection assay that uses high-resolution melting (HRM) analysis of RT-PCR products from highly multiplexed assays to differentiate broad diversities of arboviruses. We differentiated 15 viral culture controls and seven additional synthetic viral DNA sequence controls, within Flavivirus, Alphavirus, Nairovirus, Phlebovirus, Orthobunyavirus and Thogotovirus genera. Among Bunyamwera, sindbis, dengue and Thogoto virus serial dilutions, detection by multiplex RT-PCR-HRM was comparable to the gold standard Vero cell plaque assays. We applied our low-cost method for enhanced broad-range pathogen surveillance from mosquito samples collected in Kenya and identified diverse insect-specific viruses, including a new clade in anopheline mosquitoes, and Wesselsbron virus, an arbovirus that can cause viral haemorrhagic fever in humans and has not previously been isolated in Kenya, in Culex spp. and Anopheles coustani mosquitoes. Our findings demonstrate how multiplex RT-PCR-HRM can identify novel viral diversities and potential disease threats that may not be included in pathogen detection panels of routine surveillance efforts. This approach can be adapted to other pathogens to enhance disease surveillance and pathogen discovery efforts, as well as the study of pathogen diversity and viral evolutionary ecology.