Isolation and phylogenetic characterization of arboviruses circulating among phlebotomine sandflies in parts of North Rift, Kenya

BackgroundUntil recently, arbovirus surveillance is mainly focused on mosquito and tick vectors, resulting in the discovery of several mosquito- and tick-borne arboviruses. However, the role of sandflies in arbovirus transmission and disease has remained largely unexplored. This study sought to isolate and characterize arboviruses from phlebotomine sandflies from selected pastoral ecozones in the North Rift region of Kenya.MethodsSandflies were collected from selected sites in North Rift Kenya between 2015 and 2018. They were sorted and pooled by sex, site, and collection date. The pools were homogenized and inoculated onto Vero cells for virus isolation. The positive pools were analyzed by polymerase chain reaction targeting different arboviruses. The isolates were further characterized by high-throughput sequencing using Illumina Miseq platform.ResultsApproximately 28,226 sandflies translating to 824 pools were sampled from the selected regions. A total of 11 showed reproducible cytopathic effects on Vero cells. We identified five arboviruses: sindbis (n = 4) from Kacheliba and Baringo, Chandipura (n = 4) from Turkana and Baringo, Koutango (n = 1) and Ntepes (n = 1) from Baringo, and Bogoria (n = 1) from Kacheliba. The percent identities of the identified viruses were approximately 80% to 98% compared to known viruses in GenBank, suggesting that some of them could be novel viruses.ConclusionThis study successfully isolated and characterized five arboviruses from sandflies. The findings suggest that sandflies are potential hosts of a wide range of arboviruses and are therefore important vectors to consider in arbovirus surveillance and evaluated for their ability to transmit them. Further studies are needed to determine the public health importance and extent of exposure of these viruses to humans and livestock populations

The evolving SARS-CoV-2 epidemic in Africa: Insights from rapidly expanding genomic surveillance

INTRODUCTION Investment in Africa over the past year with regard to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequencing has led to a massive increase in the number of sequences, which, to date, exceeds 100,000 sequences generated to track the pandemic on the continent. These sequences have profoundly affected how public health officials in Africa have navigated the COVID-19 pandemic. RATIONALE We demonstrate how the first 100,000 SARS-CoV-2 sequences from Africa have helped monitor the epidemic on the continent, how genomic surveillance expanded over the course of the pandemic, and how we adapted our sequencing methods to deal with an evolving virus. Finally, we also examine how viral lineages have spread across the continent in a phylogeographic framework to gain insights into the underlying temporal and spatial transmission dynamics for several variants of concern (VOCs). RESULTS Our results indicate that the number of countries in Africa that can sequence the virus within their own borders is growing and that this is coupled with a shorter turnaround time from the time of sampling to sequence submission. Ongoing evolution necessitated the continual updating of primer sets, and, as a result, eight primer sets were designed in tandem with viral evolution and used to ensure effective sequencing of the virus. The pandemic unfolded through multiple waves of infection that were each driven by distinct genetic lineages, with B.1-like ancestral strains associated with the first pandemic wave of infections in 2020. Successive waves on the continent were fueled by different VOCs, with Alpha and Beta cocirculating in distinct spatial patterns during the second wave and Delta and Omicron affecting the whole continent during the third and fourth waves, respectively. Phylogeographic reconstruction points toward distinct differences in viral importation and exportation patterns associated with the Alpha, Beta, Delta, and Omicron variants and subvariants, when considering both Africa versus the rest of the world and viral dissemination within the continent. Our epidemiological and phylogenetic inferences therefore underscore the heterogeneous nature of the pandemic on the continent and highlight key insights and challenges, for instance, recognizing the limitations of low testing proportions. We also highlight the early warning capacity that genomic surveillance in Africa has had for the rest of the world with the detection of new lineages and variants, the most recent being the characterization of various Omicron subvariants. CONCLUSION Sustained investment for diagnostics and genomic surveillance in Africa is needed as the virus continues to evolve. This is important not only to help combat SARS-CoV-2 on the continent but also because it can be used as a platform to help address the many emerging and reemerging infectious disease threats in Africa. In particular, capacity building for local sequencing within countries or within the continent should be prioritized because this is generally associated with shorter turnaround times, providing the most benefit to local public health authorities tasked with pandemic response and mitigation and allowing for the fastest reaction to localized outbreaks. These investments are crucial for pandemic preparedness and response and will serve the health of the continent well into the 21st century

Starch branching enzymes and their genes in Sorghum

Starch is an important raw material both for food and non-food purposes. It is synthesized and stored in source and sink tissues in plants. The starch deposited in amyloplasts of storage tissues possesses several physico-chemical properties, which makes it desirable for diverse applications. For industrial applications, starches high in either amylose or amylopectin are preferred in order to minimize chemical modification. The main purpose in our research is to understand starch synthesis in the sorghum endosperm, and use this information to generate transgenic sorghum with novel starches for both human diet and industrial application. In starch biosynthesis, the activity and expression profile of starch branching enzymes is important in determining both the yield and proportion of amylose and amylopectin. This thesis describes some of the characteristics of starch branching enzymes and their genes in sorghum. The genes encoding two SBE isoforms, SBEIIa and SBEIIb, were cloned and characterised. Comparison of the SBEIIb amino acid sequence with sequences from related species revealed a conserved core stretch of amino acids believed to harbor the catalytic site of the enzyme. Spatial expression patterns showed that the activity of sorghum sbeI and sbeIIb are seed-specific, while sorghum sbeIIa is expressed in endosperm, embryonic and vegetative tissues. We demonstrated that endosperm-specific expression of barley sbeIIb is governed by the second intron of the barley sbeIIb gene, and that the same second intron can confer endosperm specific expression to both sorghum sbeIIb and barley sbeIIa. This implies that a similar regulation factor in barley, which interacts with sequences within the intron, is also present in sorghum. Deletion mutagenesis of the sorghum sbeIIb promoter showed that a, 1-kb fragment 5´ of the transcriptional start site constitutes the minimal promoter capable of high level expression. Diurnal expression profiles suggested that an endogenous oscillator in the endosperm controls expression of sbeI, sbIIa and sbeIIb

Dengue Outbreak in Mombasa City, Kenya, 2013-2014: Entomologic Investigations.

Dengue outbreaks were first reported in East Africa in the late 1970s to early 1980s including the 1982 outbreak on the Kenyan coast. In 2011, dengue outbreaks occurred in Mandera in northern Kenya and subsequently in Mombasa city along the Kenyan coast in 2013-2014. Following laboratory confirmation of dengue fever cases, an entomologic investigation was conducted to establish the mosquito species, and densities, causing the outbreak. Affected parts of the city were identified with the help of public health officials. Adult Ae. aegypti mosquitoes were collected using various tools, processed and screened for dengue virus (DENV) by cell culture and RT-PCR. All containers in every accessible house and compound within affected suburbs were inspected for immatures. A total of 2,065 Ae. aegypti adults were collected and 192 houses and 1,676 containers inspected. An overall house index of 22%, container index, 31.0% (indoor = 19; outdoor = 43) and Breteau index, 270.1, were observed, suggesting that the risk of dengue transmission was high. Overall, jerry cans were the most productive containers (18%), followed by drums (17%), buckets (16%), tires (14%) and tanks (10%). However, each site had specific most-productive container-types such as tanks (17%) in Kizingo; Drums in Nyali (30%) and Changamwe (33%), plastic basins (35%) in Nyali-B and plastic buckets (81%) in Ganjoni. We recommend that for effective control of the dengue vector in Mombasa city, all container types would be targeted. Measures would include proper covering of water storage containers and eliminating discarded containers outdoors through a public participatory environmental clean-up exercise. Providing reliable piped water to all households would minimize the need for water storage and reduce aquatic habitats. Isolation of DENV from male Ae. aegypti mosquitoes is a first observation in Kenya and provides further evidence that transovarial transmission may have a role in DENV circulation and/or maintenance in the environment

Crimean-Congo Hemorrhagic Fever Virus in Hyalommid Ticks, Northeastern Kenya

As part of ongoing arbovirus surveillance, we screened ticks obtained from livestock in northeastern Kenya in 2008 to assess the risk for human exposure to tick-borne viruses. Of 1,144 pools of 8,600 Hyalomma spp. ticks screened for Congo-Crimean hemorrhagic fever virus by reverse transcription PCR, 23 pools were infected, demonstrating a potential for human exposure

Sugar-mediated semidian oscillation of gene expression in the cassava storage root regulates starch synthesis

Starch branching enzyme (SBE) activity in the cassava storage root exhibited a diurnal fluctuation, dictated by a transcriptional oscillation of the corresponding SBE genes. The peak of SBE activity coincided with the onset of sucrose accumulation in the storage, and we conclude that the oscillatory mechanism keeps the starch synthetic apparatus in the storage root sink in tune with the flux of sucrose from the photosynthetic source. When storage roots were uncoupled from the source, SBE expression could be effectively induced by exogenous sucrose. Turanose, a sucrose isomer that cannot be metabolized by plants, mimicked the effect of sucrose, demonstrating that downstream metabolism of sucrose was not necessary for signal transmission. Also glucose and glucose-1-P induced SBE expression. Interestingly, induction by sucrose, turanose and glucose but not glucose-1-P sustained an overt semidian (12-h) oscillation in SBE expression and was sensitive to the hexokinase (HXK) inhibitor glucosamine. These results suggest a pivotal regulatory role for HXK during starch synthesis. Abscisic acid (ABA) was another potent inducer of SBE expression. Induction by ABA was similar to that of glucose-1-P in that it bypassed the semidian oscillator. Both the sugar and ABA signaling cascades were disrupted by okadaic acid, a protein phosphatase inhibitor. Based on these findings, we propose a model for sugar signaling in regulation of starch synthesis in the cassava storage root