Search

Abstract Background In Africa, food legumes such as cowpeas and beans constitute important sources of proteins for millions of rural and urban populations. However, attacks by multiple pest species can reduce yields by up to 80%. Small-holder farmers protect their crops against pests using conventional insecticides, thereby contributing to insecticide resistance and jeopardizing food safety. To date, no alternative sustainable practices are available to reduce insecticide use in the management of legume pests. This study aimed to provide a sustainable alternative to insecticide use based on semiochemicals to manage multiple legume pest species and enhance food productivity. Results Using coupled gas chromatography-mass spectrometry (GC–MS) and coupled GC-electroantennographic detection (EAD) analyses, we identified 2-methylbutyl 2-methylbutanoate as the male-produced aggregation pheromone of the legume pest, Clavigralla elongata, a coreid bug species specific to East Africa. In multi-site field trials conducted in West Africa and East Africa, 2-methylbutyl 2-methylbutanoate and its analogue, isopentyl butanoate, previously identified from the pan-tropical coreid bug, C. tomentosicollis, both lured multiple legume insect pest species including from the Clavigralla genus, the legume pod-borer Maruca vitrata, flower thrips Megalurothrips sjostedti, and whitefly Bemisia tabaci into traps. Additionally, both pheromones lured the Clavigralla natural enemy parasitoid Gryon fulviventris into traps. The whitefly was only captured in pheromone-baited traps in East Africa. Deployment of an optimized pheromone trapping system significantly reduced legume pests and increased natural enemy density by up to sixfold compared to farmer practice (insecticide-treatment) and control. Legume yields for pheromone and insecticide treatments were comparable, ~ 320–590% higher than untreated controls. Conclusion These findings establish coreid bug pheromones as effective novel multifunctional semiochemical-based tools for sustainable legume pest management and production without insecticide use. Graphical abstract

Abstract Background Surveillance of the host–anopheline mosquitoes’ interaction is important for assessing malaria transmission risk and guiding vector control. We assume that changes in malaria vector species’ feeding habits, as well as the surrounding environment, have a substantial impact on varied malaria transmission. In this study, we determined the vertebrate host feeding patterns of anopheline mosquitoes to characterize entomologic risk factors for malaria in Jabi Tehnan, Northwestern Ethiopia. Methods Blood-fed anophelines surveyed during malaria surveillance in Jabi Tehnan district of northwestern Ethiopia were utilized in this study. They were collected using Centers for Disease Control and Prevention (CDC) light traps deployed in selected households per village, placed indoors and outdoors, spanning three agroecological settings (dry mountain, plateau, and semiarid highlands) between June 2020 and May 2021. The engorged mosquitoes were analyzed for host blood meal sources and Plasmodium infection via polymerase chain reaction (PCR) and/or sequencing. Infection rates and bovine and human blood indices were calculated and compared for abundant species; between indoors and outdoors and between agroecology using a chi-squared test for equality of proportion in R package at a significant level of p ≤ 0.05. Results A total of 246 mosquitoes were successfully typed (indoor, 121; outdoor, 125), with greater relative abundance indoors in mountain and plateau highlands, and outdoors in semiarid areas. Despite ecological differences in blood-fed capture rates, cattle served as the most utilized blood meal source by 11 anopheline species with an overall bovine blood index (BBI) of 74.4%. This trend was dictated by Anopheles gambiae s.l. (198/246; BBI = 73.7%), which exhibited the most plastic feeding habits that included humans (human blood index = 15.7%) and other livestock and rodents. A total of five anopheline species (An. gambiae s.l., An. funestus s.l., An. coustani s.l., An. pretoriensis, and An. pharoensis) fed on humans, of which the first three were found infected with Plasmodium parasites. Most of the infected specimens were An. arabiensis (5.6%, 11/198) and had recently fed mainly on cattle (72.7%, 8/11); one each of infected An. funestus s.l. and An. coustani s.l. had fed on humans and cattle, respectively. Conclusions The results demonstrate communal feeding on cattle by anophelines including primary and secondary malaria vectors. This study also indicates the importance of cattle-targeted interventions for sustainable control of malaria vectors in the study areas. Graphical abstract

Abstract Improved understanding of mosquito–plant feeding interactions can reveal insights into the ecological dynamics of pathogen transmission. In wild malaria vectors Anopheles gambiae s.l. and An. funestus group surveyed in selected dryland ecosystems of Kenya, we found a low level of plant feeding (2.8%) using biochemical cold anthrone test but uncovered 14-fold (41%) higher rate via DNA barcoding targeting the chloroplast rbcL gene. Plasmodium falciparum positivity was associated with either reduced or increased total sugar levels and varied by mosquito species. Gut analysis revealed the mosquitoes to frequently feed on acacia plants (~ 89%) (mainly Vachellia tortilis) in the family Fabaceae. Chemical analysis revealed 1-octen-3-ol (29.9%) as the dominant mosquito attractant, and the sugars glucose, sucrose, fructose, talose and inositol enriched in the vegetative parts, of acacia plants. Nutritional analysis of An. longipalpis C with high plant feeding rates detected fewer sugars (glucose, talose, fructose) compared to acacia plants. These results demonstrate (i) the sensitivity of DNA barcoding to detect plant feeding in malaria vectors, (ii) Plasmodium infection status affects energetic reserves of wild anopheline vectors and (iii) nutrient content and olfactory cues likely represent potent correlates of acacia preferred as a host plant by diverse malaria vectors. The results have relevance in the development of odor-bait control strategies including attractive targeted sugar-baits.

Abstract Among the factors affecting the effectiveness of malaria control is poor knowledge of the entomologic drivers of the disease. We investigated anopheline populations as part of a baseline study to implement house screening of windows and doors as a supplementary malaria control tool towards elimination in Jabi Tehnan district, Amhara Regional State of Ethiopia. The samples were surveyed monthly using CDC light traps between June 2020 and May 2021. Mosquito trap density ( 80% of total anopheline captures and contributed ~ 42% of Plasmodium-infected mosquitoes. On the other hand, morphologically scored Anopheles funestus s.l., constituting about 6% of anopheline collections, accounted for 50% of sporozoite-infected mosquitoes. Most of the infected An. funestus s.l. specimens (86.7%) were grouped with previously unknown or undescribed Anopheles species previously implicated as a cryptic malaria vector in the western Kenyan highlands, confirming its wider geographic distribution in eastern Africa. Other species with Plasmodium infection included An. longipalpis C, An. theileri, An. demillioni, and An. nili. Cumulatively, 77.8% of the infected mosquitoes occurred outdoors. These results suggest efficient malaria parasite transmission despite the low vector densities, which has implications for effective endpoint indicators to monitor malaria control progress. Additionally, the largely outdoor infection and discovery of previously unknown and cryptic vectors suggest an increased risk of residual malaria transmission and, thus, a constraint on effective malaria prevention and control.

Abstract In the Americas, the fall armyworm (Spodoptera frugiperda) exists in two genetically distinct strains, the corn (C) and rice (R) strains. Despite their names, these strains are not associated with host plant preferences but have been shown to vary in pheromone composition and male responses. Recently, S. frugiperda was detected in Africa as an invasive species, but knowledge about variation in strain types, pheromone composition and inter-strain mating of populations of the pest in the continent has not been fully examined. Therefore, this study aimed to investigate variations, if any in the pheromone composition of female moths, male moth responses, and mating between C and R mitotypes of S. frugiperda populations in Kenya, as well as their geographic distribution. Strains (mitotypes) of S. frugiperda were identified using mitochondrial DNA (mtDNA) markers, and their pheromonal composition determined by coupled gas chromatography–mass spectrometric (GC–MS) analysis. Male moth responses to these compounds were evaluated using GC-electroantennographic detection (EAD), electroantennogram (EAG), and wind tunnel assays. Oviposition assays were used to determine whether R and C mitotype moths could mate and produce eggs. The results showed that both the R and C mitotypes were present, and there were no statistically significant differences in their distribution across all sampled locations. Five pheromone compounds including (Z)-7-dodecenyl acetate (Z7-12:OAc), (Z)-7-tetradecenyl acetate (Z7-14:OAc), (Z)-9-tetradecenyl acetate (Z9-14:OAc), (Z)-11-tetradecenyl acetate (Z11-14:OAc) and (Z)-11-hexadecenyl acetate (Z11-16:OAc), were detected in the pheromone glands of female moths of both mitotypes, with Z9-14:OAc being the most abundant. The relative percentage composition of Z9-14:OAc was similar in both mitotypes. However, the R mitotype had a 2.7 times higher relative percentage composition of Z7-12:OAc compared to the C mitotype moth, while the C mitotype moth had a 2.4 times higher relative percentage composition of Z11-16:OAc than the R mitotype moth. Male moths of both mitotypes exhibited similar responses to the pheromone compounds, showing the strongest responses to Z9-14:OAc and Z7-12:OAc in electrophysiological and behavioural assays. There was mating between R and C mitotypes with egg production comparable to mating within the same mitotype. Our results revealed that differences between the two S. frugiperda mitotypes are characterized by female moth pheromone composition rather than male moth responses to the pheromones, and that this does not prevent hybridisation between the mitotypes, which may have implications for their management.

IntroductionPreviously, we documented that the malaria vector Anopheles gambiae responds to volatile emissions from the root exudate water of the invasive plant, Parthenium hysterophorus. However, the origin of the volatiles remains to be investigated. Here, we isolated bacteria from the root exudate water of the plant, test the influence of their volatiles in gravid An. gambiae oviposition, and examined relationships between volatile profiles and oviposition.MethodsBacteria from root exudate water of P. hysterophorus were isolated using culture on Luria Bertani medium and identified by sequencing the 16S rRNA gene. Cultures of individual isolates were evaluated for egg laying response by gravid An. gambiae and number of eggs laid compared using generalized linear models relative to those in crude bacteria-mixture. Headspace volatile emissions of the bacterial isolates were analyzed by gas chromatography coupled to mass spectrometry (GC-MS) and relationships between volatile organic compound (VOC) profiles and gravid mosquito oviposition examined using Random Forest Analysis. Proximate analysis was performed to assess the difference in volatile chemistry among the different isolates.ResultsThree isolates were identified as Gram-negative bacteria belonging to two families: Enterobacteriaceae (Enterobacter sp. and Enterobacter mori) and Alcaligenaceae (Alcaligens aquatilis). An. gambiae laid 3-fold more eggs in cultures of A. aquatilis than in those of Enterobacter sp. In turn, approx. 4-fold more eggs were laid in cultures of E. mori than A. aquatilis. Overall, 16 VOCs were identified in the headspace of the isolates belonging to the chemical classes benzenoids, pyrazines, aldehydes, terpenes, alcohols, alkanes, and indoles. Random Forest Analysis identified 10 compounds contributing the most to the attraction of odors of the bacteria isolates to oviposition. Specifically, dodecane and indole were emitted in higher amounts in odors of Enterobacter sp than the other two species. Proximate analysis revealed differential attraction of the isolates on the gravid mosquito to be associated with their volatile profiles.ConclusionOur results provide first report of E. mori or A. aquatilis mediating attractive oviposition responses in An. gambiae in support of the important role microbes play in insect oviposition. The potential use of the microbes and associated volatiles in malaria vector management needs further investigation.

Abstract Background The fall armyworm (FAW), Spodoptera frugiperda, is a devastating invasive pest and a threat to food security in Africa, with yield losses of 20–50%. Recent studies highlighted the importance of cereal crops such as maize and sorghum as the most preferred host plants for FAW oviposition. In the current work, we investigated the olfactory responses of FAW and its key larval endoparasitoid Cotesia icipe to odours from the preferred host (maize) in the presence of six potential companion crops including beans, groundnut, sweet potato, greenleaf- and silverleaf desmodium, and cassava. We hypothesized that odours released by companion crops in maize-based intercropping systems would alter host preferences of FAW for oviposition and its parasitoid responses. Results In dual choice oviposition bioassays, FAW laid significantly more eggs on maize than on the other plants. However, in the multiple-choice bioassays, significantly fewer eggs were laid on maize when companion plants were present except cassava. While wind tunnel bioassays confirmed the differential behavioural responses of FAW, we found that its larval endoparasitoid C. icipe was attracted to volatiles from the companion plants tested individually and/or when they were combined with maize. Coupled gas chromatography–mass spectrometry (GC–MS) analysis detected several potential behaviour-modifying compounds including (Z)-3-hexenyl acetate, (E)-β-ocimene, (E)-4,8-dimethyl-1,3,7-nonatriene, (E)-β-caryophyllene, camphor, methyl salicylate and (E, E)-4,8,12-trimethyl-1,3,7,11-tridecatetraene. Conclusions Our findings provide evidence supporting diversified maize cropping system could reduce FAW damage by repelling the pest while simultaneously recruiting its natural enemies. Hence, diversifying cereal cropping system with companion crops could serve as an ecologically sustainable FAW management strategy. Graphical Abstract

Phlebotomine sand flies are medically important as vectors of the protozoan parasites that cause leishmaniasis and other bacterial and viral pathogens. Previous work demonstrated that both sexes of certain species of sandflies are attracted to 1-octen-3-ol (octenol). Since 1-octen-3-ol exists as two enantiomeric isomers ─ ((R)-(-)- (R-form) and (S)-(+)- (S-form), we tested the hypothesis that the two enantiomeric forms and racemic mixture (R/S) attracted different sand fly species. We carried out field trials in a leishmaniasis endemic foci in Baringo County, Kenya. In a randomized design, trap captures of sandflies in CDC light traps baited with the R-, S- and racemic (R/S) forms of 1-octen-3-ol in hexane varied with the form and dose of the compound. Interestingly, of the captured species, only Phlebotomus martini, the vector of the parasite causing visceral leishmaniasis, exhibited a dose-dependent response to octenol; captures of both sexes of the species being generally 1.7-fold higher with the R- than S-form. There was no significant effect of treatment on captures of Sergentomyia species (S. schwetzi, S. antennata, S, clydei). Our findings have implications for surveillance of sandfly populations as part of leishmaniasis epidemiologic investigation.

IntroductionCulicoides biting midges (Diptera: Ceratopogonidae) are vectors of arboviral pathogens that primarily affect livestock represented by Schmallenberg virus (SBV), epizootic hemorrhagic disease virus (EHDV) and bluetongue virus (BTV). In Kenya, studies examining the bionomic features of Culicoides including species diversity, blood-feeding habits, and association with viruses are limited.MethodsAdult Culicoides were surveyed using CDC light traps in two semi-arid ecologies, Baringo and Kajiado counties, in Kenya. Blood-fed specimens were analysed through polymerase chain reaction (PCR) and sequencing of cytochrome oxidase subunit 1 (cox1) barcoding region. Culicoides pools were screened for virus infection by generic RT-PCR and next-generation sequencing (NGS).ResultsAnalysis of blood-fed specimens confirmed that midges had fed on cattle, goats, sheep, zebra, and birds. Cox1 barcoding of the sampled specimens revealed the presence of known vectors of BTV and epizootic hemorrhagic disease virus (EHDV) including species in the Imicola group (Culicoides imicola) and Schultzei group (C. enderleni, C. kingi, and C. chultzei). Culicoides leucostictus and a cryptic species distantly related to the Imicola group were also identified. Screening of generated pools (11,006 individuals assigned to 333 pools) by generic RT-PCR revealed presence of seven phylogenetically distinct viruses grouping in the genera Goukovirus, Pacuvirus and Orthobunyavirus. The viruses showed an overall minimum infection rate (MIR) of 7.0% (66/333, 95% confidence interval (CI) 5.5-8.9). In addition, full coding sequences of two new iflaviruses, tentatively named Oloisinyai_1 and Oloisinyai_2, were generated by next-generation sequencing (NGS) from individual homogenate of Culicoides pool.ConclusionThe results indicate a high genetic diversity of viruses in Kenyan biting midges. Further insights into host-vector-virus interactions as well as investigations on the potential clinical significance of the detected viruses are warranted.

Intraguild interactions are known to trigger competition among phytophagous insects that depend on the same food source, which may have implications on natural enemy performance. Here, we tested the hypothesis that intraguild interactions between the greenhouse whitefly, Trialeurodes vaporariorum, and the South American tomato pinworm, Phthorimaea absoluta on the host plant tomato would affect performance of the pinworm ectoparasitoid Bracon nigricans. In oviposition bioassays, P. absoluta females significantly laid more eggs and were attracted to tomato plants infested with T. vaporariorum than uninfested plants. Additionally, T. vaporariorum infestation decreased B. nigricans parasitism on P. absoluta larvae and attraction to tomato plants. In coupled gas chromatography-electroantennographic detection (GC-EAD) recordings, antennae of P. absoluta and B. nigricans females detected 18 and 15 components, respectively from the herbivore-induced volatiles, which were identified by GC-mass spectrometry (GC-MS) as green leaf volatiles (GLVs) and terpenes. In dose-response behavioral assays, P. absoluta and B. nigricans responses varied with the composition and concentration of blends and individual compounds tested. Whereas P. absoluta was attracted to GLVs and terpenes, B. nigricans avoided most of these chemicals, especially the GLV (E)-2-hexenal and terpenes δ-3-carene, and (E)-caryophyllene. The results suggest that herbivore intraguild interactions can influence the behavioral ecology and performance of natural enemies which may have evolutionary significance. Also, these specific GLVs and terpenes can be exploited in the management of P. absoluta.

Abstract Outdoor biting by anopheline mosquitoes is one of the contributors to residual malaria transmission, but the profile of vectors driving this phenomenon is not well understood. Here, we studied the bionomics and genetically characterized populations of An. gambiae and An. funestus complexes trapped outdoors in three selected dryland areas including Kerio Valley, Nguruman and Rabai in Kenya. We observed a higher abundance of Anopheles funestus group members (n = 639, 90.6%) compared to those of the An. gambiae complex (n = 66, 9.4%) with An. longipalpis C as the dominant vector species with a Plasmodium falciparum sporozoite rate (Pfsp) of 5.2% (19/362). The known malaria vectors including An. funestus s.s. (8.7%, 2/23), An. gambiae (14.3%, 2/14), An. rivulorum (14.1%, 9/64), An. arabiensis (1.9%, 1/52) occurred in low densities and displayed high Pfsp rates, which varied with the site. Additionally, six cryptic species found associated with the An. funestus group harbored Pf sporozoites (cumulative Pfsp rate = 7.2%, 13/181). We detected low frequency of resistant 119F-GSTe2 alleles in An. funestus s.s. (15.6%) and An. longipalpis C (3.1%) in Kerio Valley only. Evidence of outdoor activity, emergence of novel and divergent vectors and detection of mutations conferring metabolic resistance to pyrethroid/DDT could contribute to residual malaria transmission posing a threat to effective malaria control.

ABSTRACT Arboviruses are among emerging pathogens of public and veterinary health significance. However, in most of sub-Saharan Africa, their role in the aetiologies of diseases in farm animals is poorly described due to paucity of active surveillance and appropriate diagnosis. Here, we report the discovery of a previously unknown orbivirus in cattle collected in the Kenyan Rift Valley in 2020 and 2021. We isolated the virus in cell culture from the serum of a clinically sick cow aged 2 to 3 years, presenting signs of lethargy. High-throughput sequencing revealed an orbivirus genome architecture with 10 double-stranded RNA segments and a total size of 18,731 bp. The VP1 (Pol) and VP3 (T2) nucleotide sequences of the detected virus, tentatively named Kaptombes virus (KPTV), shared maximum similarities of 77.5% and 80.7% to the mosquito-borne Sathuvachari virus (SVIV) found in some Asian countries, respectively. Screening of 2,039 sera from cattle, goats, and sheep by specific RT-PCR identified KPTV in three additional samples originating from different herds collected in 2020 and 2021. Neutralizing antibodies against KPTV were found in 6% of sera from ruminants (12/200) collected in the region. In vivo experiments with new-born and adult mice induced body tremors, hind limb paralysis, weakness, lethargy, and mortality. Taken together, the data suggest the detection of a potentially disease-causing orbivirus in cattle in Kenya. Its impact on livestock, as well as its potential economic damage, needs to be addressed in future studies using targeted surveillance and diagnostics. IMPORTANCE The genus Orbivirus contains several viruses that cause large outbreaks in wild and domestic animals. However, there is little knowledge on the contribution of orbiviruses to diseases in livestock in Africa. Here, we report the identification of a novel presumably disease-causing orbivirus in cattle, Kenya. The virus, designated Kaptombes virus (KPTV), was initially isolated from a clinically sick cow aged 2 to 3 years, presenting signs of lethargy. The virus was subsequently detected in three additional cows sampled in neighboring locations in the subsequent year. Neutralizing antibodies against KPTV were found in 10% of cattle sera. Infection of new-born and adult mice with KPTV caused severe symptoms and lead to death. Together, these findings indicate the presence of a previously unknown orbivirus in ruminants in Kenya. These data are of relevance as cattle represents an important livestock species in farming industry and often is the main source of livelihoods in rural areas of Africa.

The tomato leafminer, Tuta absoluta is a destructive invasive pest of cultivated tomato and other Solanaceae plants, with yield losses of 80-100%. Mirid predators are key natural enemies of T. absoluta, but they also feed on host plants in the absence of their prey. Management of T. absoluta is a challenge due to its high biotic potential, resistance to many insecticides and the absence of sufficiently adapted auxiliary fauna in its new dispersion zones. Olfaction plays an important role in the tritrophic interaction between tomato, its herbivore pest T. absoluta and its mirid predators, which can be influenced by non-host plant odors. However, how non-host odours shape this interaction is poorly understood. Previously, we had demonstrated belowground crop protection properties of certain Asteraceae plants against the root-knot nematode Meloidogyne incognita, pest of tomato and other Solanaceae plants. Additionally, Asteraceae plants impact negatively on feeding behavior of above-ground pests of Solanaceae plants, including the greenhouse whitefly (Trialeurodes vaporariorum) and green peach aphid (Myzus persicae). Here, we tested the hypothesis that foliar volatiles from some of these non-host Asteraceae plants can influence the tomato-T. absoluta-mirid predator tritrophic interaction. In olfactometer assays, T. absoluta females were attracted to volatiles of the Solanaceae host plants tomato and giant nightshade but avoided volatiles of the Asteraceae plants, blackjack and marigold, and the positive control, wild tomato, when tested alone or in combination with the host plants. Coupled gas chromatography-mass spectrometry analysis showed that host and non-host plants varied in their emission of volatiles, mainly monoterpenes and sesquiterpenes. Random forest analysis combined with behavioral assays identified monoterpenes as the host plant attractive blend to T. absoluta and its mirid predator, with sesquiterpenes identified as the non-host plant repellent blend against T. absoluta. Contrastingly, the mirid predator was indifferent to the non-host plant repellent sesquiterpenes. Our findings indicate that terpenes influence the tomato-T. absoluta-mirid predator tritrophic interaction. Further, our results emphasize the importance of studying crop protection from a holistic approach to identify companion crops that serve multi-functional roles.

Phleboviruses are emerging pathogens of public health importance. However, their association with ticks is poorly described, particularly in Africa. Here, adult ticks infesting cattle, goats and sheep were collected in two dryland pastoralist ecosystems of Kenya (Baringo and Kajiado counties) and were screened for infection with phleboviruses. Ticks mainly belonged to the species Rhipicephalus appendiculatus, Hyalomma impeltatum, and Hyalomma rufipes. A fragment of the RNA-dependent RNA polymerase (RdRp) gene was identified in thirty of 671 tick pools, of which twenty-nine were from livestock sampled in Baringo county. Phylogenetic analyses revealed that twenty-five sequences were falling in three clades within the group of tick-associated phleboviruses. The sequences of the three clades showed nucleotide distances 8%, 19% and 22%, respectively, to previously known viruses suggesting that these sequence fragments may belong to three distinct viruses. Viruses of the group of tick-associated phleboviruses have been found in several countries and continents but so far have not been associated with disease in humans or animals. In addition, five sequences were found to group with the sandfly-associated phleboviruses Bogoria virus, Perkerra virus and Ntepes virus recently detected in the same region. Further studies are needed to investigate the transmission and maintenance cycles of these viruses, as well as to assess their potential to infect vertebrates.

ABSTRACT Ngari virus (NRIV) is a mosquito-borne reassortant orthobunyavirus that causes severe febrile illness and hemorrhagic fever in humans and small ruminants. Due to limited diagnostics and surveillance, NRIV has only been detected sporadically during Rift Valley fever virus outbreaks. Little is known on its interepidemic maintenance and geographic distribution. In this study, sera from cattle, goats, and sheep were collected through a cross-sectional survey after the rainy seasons between 2020 and 2021 in two pastoralist-dominated semiarid ecosystems, Baringo and Kajiado counties in Kenya. NRIV was detected in 11 apparently healthy animals (11/2,039, 0.54%) by RT-PCR and isolated in cell culture from seven individuals. Growth analyses displayed efficient replication in cells from sheep and humans in contrast to weak replication in goat cells. NRIV infection of a wide variety of different vector cells showed only rapid replication in Aedes albopictus cells but not in cells derived from other mosquito species or sandflies. Phylogenetic analyses of complete-coding sequences of L, M, and S segments of four viruses showed that the Kenyan sequences established a monophyletic clade most closely related to a NRIV sequence from a small ruminant from Mauritania. NRIV neutralizing reactivity in cattle, goats, and sheep were 41.6% (95% CI = 30 to 54.3), 52.4% (95% CI = 37.7 to 66.6), and 19% (95% CI = 9.7 to 33.6), respectively. This is the first detection of NRIV in livestock in Kenya. Our results demonstrate active and undetected circulation of NRIV in the three most common livestock species highlighting the need for an active one-health surveillance of host networks, including humans, livestock, and vectors. IMPORTANCE Surveillance of vectors and hosts for infection with zoonotic arthropod-borne viruses is important for early detection and intervention measures to prevent outbreaks. Here, we report the undetected circulation of Ngari virus (NRIV) in apparently healthy cattle, sheep, and goats in Kenya. NRIV is associated with outbreaks of hemorrhagic fever in humans and small ruminants. We demonstrate the isolation of infectious virus from several animals as well as presence of neutralizing antibodies in 38% of the tested animals. Our data indicate active virus circulation and endemicity likely having important implications for human and animal health.

Crimean–Congo haemorrhagic fever virus (CCHFV) is the causative agent of CCHF, a fatal viral haemorrhagic fever disease in humans. The maintenance of CCHFV in the ecosystem remains poorly understood. Certain tick species are considered as vectors and reservoirs of the virus. Diverse animals are suspected as amplifiers, with only scarce knowledge regarding rodents in virus epidemiology. In this study, serum samples from febrile patients, asymptomatic livestock (cattle, donkeys, sheep, and goats), and peridomestic rodents from Baringo (Marigat) and Kajiado (Nguruman) counties within the Kenyan Rift Valley were screened for acute CCHFV infection by RT-PCR and for CCHFV exposure by ELISA. RT-PCR was performed on all livestock samples in pools (5–7/pool by species and site) and in humans and rodents individually. CCHFV seropositivity was significantly higher in livestock (11.9%, 113/951) compared to rodents (6.5%, 6/93) and humans (5.9%, 29/493) (p = 0.001). Among the livestock, seropositivity was the highest in donkeys (31.4%, 16/51), followed by cattle (14.1%, 44/310), sheep (9.8%, 29/295) and goats (8.1%, 24/295). The presence of IgM antibodies against CCHFV was found in febrile patients suggesting acute or recent infection. CCHFV RNA was detected in four pooled sera samples from sheep (1.4%, 4/280) and four rodent tissues (0.83%, 4/480) showing up to 99% pairwise nucleotide identities among each other. Phylogenetic analyses of partial S segment sequences generated from these samples revealed a close relationship of 96–98% nucleotide identity to strains in the CCHFV Africa 3 lineage. The findings of this study suggest active unnoticed circulation of CCHFV in the study area and the involvement of livestock, rodents, and humans in the circulation of CCHFV in Kenya. The detection of CCHF viral RNA and antibodies against CCHFV in rodents suggests that they may participate in the viral transmission cycle.

Background: Thymol as a natural biological template can be modified chemically since the hydroxyl group makes it a candidate for structural modification. Thus, this study incorporated the triazole moiety on thymol and the chlorination of thymol moiety to help improve its biological potency. Materials and methods: A series of ten 1,2,3-triazole-thymol derivatives 1–10 were synthesized from thymol, by a click reaction between O-propargyl terminal alkyne of thymol and its chlorothymol with benzyl azide and substituted benzyl azides. Their structures were confirmed by spectroscopic methods (1H-NMR, 13C-NMR, IR, GC-MS-EI/CI and LC-ESI-QTOF-MS). The Well diffusion method using Müeller-Hinton agar plates was used to demonstrate the antimicrobial activities of the synthesized triazole-thymol derivatives on selected bacterial strains; Escherichia coli ATCC 25922, Staphylococcus aureus ATCC25923, Methicillin resistant S. aureus (MRSA), Pseudomonas aeruginosa ATCC 29853, E. coli ESBL, Klebsiella pneumoniae NCTC 13438 and Meropenem Resistant E. coli. Results: All the synthesized triazole-thymol derivatives showed significant but variable antibacterial activity against the seven medically important bacterial strains tested. The compound 4-((4-chloro-2-isopropyl-5-methylphenoxy)methyl)-1-(2-nitrobenzyl)-1H-1,2,3triazole (9) demonstrated a higher antibacterial activity with a mean zone of inhibition (38.7 mm) compared with ampicillin as the positive control which gave a zone size of 30.0 mm. In addition, the compound showed a three-fold potency than the parent compound, thymol (11.0 mm) against MRSA at a concentration of 100 μg/ml. Conclusion: These results provide additional evidence of the exploitation of natural products like thymol as leads for drug development against medically important bacterial pathogens.

Abstract Gravid female Anopheles gambiae mosquitoes identify suitable oviposition sites through a repertoire of cues, but the influence of allelochemicals, especially root phytochemicals in modulating this behavior and impacting subsequent progeny bionomics remains unexplored. We addressed these questions in the malaria vector Anopheles gambiae and its invasive host plant Parthenium hysterophorus. Using chemical analysis combined with laboratory behavioral assays, we demonstrate that a blend of terpenes, namely α-pinene, α-phellandrene, β-phellandrene, 3-carene and (E)-caryophyllene emitted from P. hysterophorus root exudate treated-water attracted gravid females. However, fewer eggs (55%) hatched in this treatment than in control water (66%). The sesquiterpene lactone parthenin, identified in both the natural aquatic habitat harboring P. hysterophorus and root exudate-treated water was found to be responsible for the ovicidal effect. Moreover, larvae exposed to parthenin developed 2 to 3 days earlier but survived 4 to 5 days longer as adults (median larval survival time = 9 days (all replicates);11 to 12 days as adults) than the non-exposed control (median larval survival time = 11 days (reps 1 & 2), 12 days (rep 3); 6 to 7 days as adults). These results improve our understanding of the risk and benefits of oviposition site selection by gravid An. gambiae females and the role root exudate allelochemicals could play on anopheline bionomics, with potential implications in malaria transmission.

Insect-specific flaviviruses (ISFs), although not known to be pathogenic to humans and animals, can modulate the transmission of arboviruses by mosquitoes. In this study, we screened 6665 host-seeking, gravid and blood-fed mosquitoes for infection with flaviviruses and assessed the vertebrate hosts of the blood-fed mosquitoes sampled in Baringo and Kajiado counties; both dryland ecosystem counties in the Kenyan Rift Valley. Sequence fragments of two ISFs were detected. Cuacua virus (CuCuV) was found in three blood-fed Mansonia (Ma.) africana. The genome was sequenced by next-generation sequencing (NGS), confirming 95.8% nucleotide sequence identity to CuCuV detected in Mansonia sp. in Mozambique. Sequence fragments of a potential novel ISF showing nucleotide identity of 72% to Aedes flavivirus virus were detected in individual blood-fed Aedes aegypti, Anopheles gambiae s.l., Ma. africana and Culex (Cx.) univittatus, all having fed on human blood. Blood-meal analysis revealed that the collected mosquitoes fed on diverse hosts, primarily humans and livestock, with a minor representation of wild mammals, amphibians and birds. The potential impact of the detected ISFs on arbovirus transmission requires further research.

Trypanosomes are important global livestock and human pathogens of public health importance. Elucidating the chemical mechanisms of trypanosome-relevant host interactions can enhance the design and development of a novel, next-generation trypanosomosis diagnostics. However, it is unknown how trypanosome infection affects livestock volatile odors. Here, we show that Trypanosoma congolense and Trypanosoma vivax infections induced dihydro-β- ionone and junenol, while abundance of dihydro-α-ionone, phenolics, p-cresol, and 3-propylphenol significantly elevated in cow urine. These biomarkers of trypanosome infection are conserved in cow breath and the urine metabolites of naturally infected cows, regardless of population, diet, or environment differences. Furthermore, treating trypanosome-infected cows reduced the levels of these indicators back to the pre-infection levels. Finally, we demonstrated that the potential of some specific biomarkers of phenolic origin may be used to detect active trypanosome infections, including low-level infections that are not detectable by microscopy. The sensitivity and specificity of biomarkers detection are suited for rapid, robust, and non-invasive trypanosomosis diagnosis under field conditions.

Abstract Background In a recent study using DNA barcoding, we identified the plants fed upon by four Afro-tropical mosquito species that vector dengue, malaria, and Rift Valley fever. Herein, we have expanded on this study by investigating the role of three of the plants, Pithecellobium dulce (Fabaceae), Leonotis nepetifolia (Lamiaceae), and Opuntia ficus-indica (Cactaceae), on the survival, fecundity, and egg viability of the dengue vector Aedes aegypti. Methods We tested these effects using females that received (i) an initial three rations of blood meals and (ii) no blood meal at all. Two controls were included: age-matched females fed on glucose solution with or without an initial blood meal and those fed exclusively on blood meals. Data were collected daily over a 30-day period. The amino acid contents of Ae. aegypti guts and their respective diets were detected by coupled liquid chromatography-mass spectrometry. Results Females fed on P. dulce and an exclusively blood meal diet had a shorter survival than those fed on glucose. On the other hand, females fed on L. nepetifolia survived longer than those fed exclusively on blood meals, whereas those fed on O. ficus-indica had the shortest survival time. With an initial blood meal, females fed on L. nepetifolia laid 1.6-fold more eggs while those fed on the other diets laid fewer eggs compared to those fed exclusively on blood meals. Hatching rates of the eggs laid varied with the diet. Mass spectroscopic analysis of gut contents of mosquitoes exposed to the different diets showed qualitative and quantitative differences in their amino acid levels. Conclusion Our findings highlight the central role of plant nutrients in the reproductive fitness of dengue vectors, which may impact their disease transmission potential.

Abstract The sources of animal odours are highly diverse, yet their ecological importance, in host–vector communication, remains unexplored. Here, using the camel (host)–Stomoxys calcitrans (vector) interaction, we collected and analyzed the Volatile Organic Compounds (VOCs) of camels from four of its different odour sources: breath, body (skin), urine, and dung. On non-metric model multivariate analyses of VOCs we show that substantial chemo-diversity exists between metabolic products associated with an individual camel. VOCs from the four metabolic products were distinct and widely segregated. Next, we show electrophysiologically, that VOCs shared between metabolic products activated more Olfactory Sensory Neurons (OSNs) and elicited strong behavioural attractive responses from S. calcitrans under field conditions independent of geography. In our extended studies on house flies, the behavioural response to these VOCs appears to be conserved. Overall, our results establish that VOCs from a range of metabolic products determine host–vector ecological interactions and may provide a more rigorous approach for discovery of unique and more potent attractants.

Hantaviruses are zoonotic rodent-borne viruses that are known to infect humans and cause various symptoms of disease, including hemorrhagic fever with renal and cardiopulmonary syndromes. They have a segmented single-stranded, enveloped, negative-sense RNA genome and are widely distributed. This study aimed to investigate the circulation of rodent-borne hantaviruses in peridomestic rodents and shrews in two semi-arid ecologies within the Kenyan Rift Valley. The small mammals were trapped using baited folding Sherman traps set within and around houses, then they were sedated and euthanatized through cervical dislocation before collecting blood and tissue samples (liver, kidney, spleen, and lungs). Tissue samples were screened with pan-hantavirus PCR primers, targeting the large genome segment (L) encoding the RNA-dependent RNA polymerase (RdRp). Eleven of the small mammals captured were shrews (11/489, 2.5%) and 478 (97.5%) were rodents. A cytochrome b gene-based genetic assay for shrew identification confirmed the eleven shrews sampled to be Crocidura somalica. Hantavirus RNA was detected in three (3/11, 27%) shrews from Baringo County. The sequences showed 93–97% nucleotide and 96–99% amino acid identities among each other, as well as 74–76% nucleotide and 79–83% amino acid identities to other shrew-borne hantaviruses, such as Tanganya virus (TNGV). The detected viruses formed a monophyletic clade with shrew-borne hantaviruses from other parts of Africa. To our knowledge, this constitutes the first report published on the circulation of hantaviruses in shrews in Kenya.

Aedes simpsoni complex has a wide distribution in Africa and comprises at least three described sub-species including the yellow fever virus (YFV) vector Ae. bromeliae. To date, the distribution and relative contributions of the sub-species and/or subpopulations including bionomic characteristics in relation to YF transmission dynamics remain poorly studied. In this study conducted in two areas with divergent ecosystems: peri-urban (coastal Rabai) and rural (Rift Valley Kerio Valley) in Kenya, survival rate was estimated by parity in Ae. simpsoni s.l. mosquitoes sampled using CO2-baited BG Sentinel traps. We then applied PCR targeting the nuclear internal transcribed spacer 2 (ITS2), region followed by sequencing and phylogenetic analytics to identify the sibling species in the Ae. simpsoni complex among parous and blood fed cohorts. Our results show that Ae. bromeliae was the most dominant sub-species in both areas, exhibiting high survival rates, human blood-feeding, and potentially, high vectorial capacity for pathogen transmission. We document for the first time the presence of Ae. lilii in Kenya and potentially yet-to-be described species in the complex displaying human feeding tendencies. We also infer a wide host feeding range on rodents, reptile, and domestic livestock besides humans especially for Ae. bromeliae. This feeding trend could likely expose humans to various zoonotic pathogens. Taken together, we highlight the utility of genotype-based analyses to generate precision surveillance data of vector populations for enhanced disease risk prediction and to guide cost-effective interventions (e.g. YF vaccinations).

Abstract Background In insects, oviposition decisions may lead to egg deposition in substrates with different larval density and nutritional levels. Individuals developing in such substrates may present plasticity in their phenotype. Here, we investigated the effect of two factors related to oviposition decisions, namely larval density and substrate quality, on the wing size and wing shape of the stable fly, Stomoxys calcitrans L. (Diptera: Muscidae). Methods We reared S. calcitrans larvae at different densities (5, 15 and 25) and on different substrates (camel, cow, donkey and sheep dung). For each fly that emerged, we recorded body weight, and detached, slide-mounted and photographed the right wing. Next, we collected 15 landmarks on each photographed wing, and applied geometric morphometric analysis to assess variation in wing size and wing shape of S. calcitrans across the different larval densities and substrate types. Results We observed that wing size and wing shape of S. calcitrans were affected by larval density and the nature of the developmental substrate. Flies reared in a group of 5 had larger wing centroid size, wing length, wing width, wing area and wing loading compared with those reared in a group of 25. Also, flies developed in donkey and sheep dung had larger wing centroid size, wing length, wing width, wing area and wing loading in comparison with those grown in camel and cow dung. Canonical variate analysis followed by discriminant analysis revealed significant wing shape variation in S. calcitrans across the different densities and substrates. Wing size had a significant but weak positive effect on wing shape. Conclusions This study demonstrates the high sensitivity of S. calcitrans wings to variation in larval density and developmental substrate, and that use of landmark-based geometric morphometric analysis could improve our understanding of how flies of veterinary importance respond to environmental variability.

We describe a novel virus, designated Ntepes virus (NPV), isolated from sand flies in Kenya. NPV has the characteristic phlebovirus trisegmented genome architecture and is related to, but distinct from, Gabek Forest phlebovirus. Diverse cell cultures derived from wildlife, livestock, and humans were susceptible to NPV, with pronounced permissiveness in swine and rodent cells. NPV infection of newborn mice caused rapid and fatal illness. Permissiveness for NPV replication in sand fly cells, but not mosquito cells, suggests a vector-specific adaptation. Specific neutralizing antibodies were found in 13.9% (26/187) of human serum samples taken at the site of isolation of NPV as well as a disparate site in northeastern Kenya, suggesting a wide distribution. We identify a novel human-infecting arbovirus and highlight the importance of rural areas in tropical Africa for arbovirus surveillance as well as extending arbovirus surveillance to include hematophagous arthropods other than mosquitoes.

The bioecology of phlebotomine sand flies is intimately linked to the utilization of environmental resources including plant feeding. However, plant feeding behavior of sand flies remains largely understudied for Afrotropical species. Here, using a combination of biochemical, molecular, and chemical approaches, we decipher specific plant-feeding associations in field-collected sand flies from a dry ecology endemic for leishmaniasis in Kenya. Cold-anthrone test indicative of recent plant feeding showed that fructose positivity rates were similar in both sand fly sexes and between those sampled indoors and outdoors. Analysis of derived sequences of the ribulose-1,5-bisphosphate carboxylase large subunit gene (rbcL) from fructose-positive specimens implicated mainly Acacia plants in the family Fabaceae (73%) as those readily foraged on by both sexes of Phlebotomus and Sergentomyia. Chemical analysis by high performance liquid chromatography detected fructose as the most common sugar in sand flies and leaves of selected plant species in the Fabaceae family. Analysis of similarities (ANOSIM) of the headspace volatile profiles of selected Fabaceae plants identified benzyl alcohol, (Z)-linalool oxide, (E)-β-ocimene, p-cymene, p-cresol, and m-cresol, as discriminating compounds between the plant volatiles. These results indicate selective sand fly plant feeding and suggest that the discriminating volatile organic compounds could be exploited in attractive toxic sugar- and odor- bait technologies control strategies.

Phlebotomine sand flies transmit many viral protozoan and bacterial pathogens of public health importance. Knowledge of the ecologic factors influencing their distribution at local scale can provide insights into disease epidemiology and avenues for targeted control. Animal sheds, termite mounds and houses are important peri-domestic and domestic habitats utilized by different sand flies as resting or breeding habitats. However, our knowledge for selection of these habitats by sand flies remains poor. Here, we tested the hypothesis that these habitat types harbor different composition of sand fly species and differ in their volatile chemistry that could influence sand fly selection. To achieve this, we employed CDC light traps following a cross-sectional survey to investigate the distribution of sand flies in the three habitats in an endemic site for leishmaniasis in Kenya. The study was carried out during the dry season, when sand flies are optimally abundant in 2018 and 2020. Sand fly abundance did not vary between the habitats, but species-specific differences in abundance was evident. Measures of sand fly community structure (Shannon diversity and richness) were highest in animal shed, followed by termite mound and lowest inside human dwelling (house). This finding indicates broader attraction of both sexes of sand flies and females of varying physiological states to animal sheds potentially used as breeding or resting sites, but also as a signal for host presence for a blood meal. Furthermore, gas chromatography-mass spectrometric analysis of volatiles collected from represented substrates associated with these habitats viz: human foot odor on worn socks (houses indoors), cow dung (animal sheds) and termite mounds (enclosed vent), revealed a total of 47 volatile organic compounds. Of these, 26, 35 and 16 were detected in human socks, cow dung and enclosed termite vent, respectively. Of these volatiles, 1-octen-3-ol, 6-methyl-5-hepten-2-one, α-pinene, benzyl alcohol, m-cresol, p-cresol and decanal, previously known as attractants for sandflies and other blood-feeding insects, were common to the habitats. Our results suggest that habitat volatiles may contribute to the composition of sand flies and highlight their potential for use in monitoring sand fly populations.

ABSTRACT Sandfly-transmitted phleboviruses (family Phenuiviridae, order Bunyavirales) are associated with febrile illness and infections of the nervous system in humans. These viruses are almost exclusively found in tropical areas of the New World and restricted to semiarid and temperate zones in the Old World. Here, we discovered seven strains of four previously unknown phleboviruses, named Bogoria virus (BOGV), Embossos virus (EMRV), Kiborgoch virus (KBGV), and Perkerra virus (PERV), as well as the recently discovered Ntepes virus, in sandflies collected in the Kenyan Rift Valley. The genomes have a tripartite organization with conserved termini typical of phleboviruses. LOBV, PERV, and EMBV showed low similarity to known phleboviruses, with less than 55% pairwise amino acid identities in the RNA-directed RNA polymerase (RdRp) proteins, and defined a highly diversified monophyletic clade in sister relationship to the sandfly fever Sicilian serocomplex. All three viruses failed to react with sandfly fever Sicilian virus antisera in recombinant immunofluorescence assays (rIFA), suggesting that the viruses belong to a yet-unknown serogroup. In contrast, KBGV was closely related to Toscana virus (84% identity of RdRp proteins) and shared a most recent common ancestor with the clade comprising sandfly fever Naples and Toscana viruses. KBGV reacted with sandfly fever Naples and Toscana virus antisera in rIFA. The genetic diversity of the detected viruses and their phylogenetic positions implies that the Old World sandfly-borne phleboviruses originated from sub-Saharan Africa. Importantly, our findings suggest that diseases associated with sandfly-borne phlebovirus infections may also affect the Kenyan population. IMPORTANCE Studies on the genetic diversity of arthropod-borne viruses circulating in rural regions can provide critical early indications on new emerging viruses essential for global epidemic preparedness. In this study, we describe the discovery of four phleboviruses in sandflies from the Kenyan Rift Valley. The novel viruses are related to the two medically important serocomplexes, sandfly fever Naples and sandfly fever Sicilian, that are associated with febrile illness and neuroinvasive infections and which were previously not known to occur in sub-Saharan Africa. Knowledge on the occurrence of sandfly-borne phleboviruses in Kenya and elsewhere in Africa can help to decipher their contributions in the etiologies of fevers of unknown origin in patients. Our findings on five genetically diverse phleboviruses detected in Kenya suggest that the common ancestor of Old World phleboviruses existed in sub-Saharan Africa, a hot spot for emerging arboviruses.

Insects that transmit many of the world’s deadliest animal diseases, for instance trypanosomosis, find their suitable hosts and avoid non-preferred hosts mostly through olfactory cues. The waterbuck repellent blend (WRB) comprising geranylacetone, guaiacol, pentanoic acid, and δ-octalactone derived from waterbuck skin odor is a repellent to some savannah-adapted tsetse flies and reduces trap catches of riverine species. However, the cellular and molecular mechanisms associated with detection and coding of the repellent odors remain to be elucidated. Here, we demonstrated that WRB inhibited blood feeding in both Glossina pallidipes Austen, 1903 and Glossina fuscipes fuscipes Newstead, 1910. Using the DREAM (Deorphanization of Receptors based on Expression Alterations in odorant receptor mRNA levels) technique, combined with ortholog comparison and molecular docking, we predicted the putative odorant receptors (ORs) for the WRB in G. f. fuscipes, a non-model insect. We show that exposure of G. f. fuscipes in vivo to WRB odorant resulted in up- and downregulation of mRNA transcript of several ORs. The WRB component with strong feeding inhibition altered mRNA transcript differently as compared to an attractant odor, showing these two odors of opposing valence already segregate at the cellular and molecular levels. Furthermore, molecular dynamics simulations demonstrated that the predicted ligand–OR binding pockets consisted mostly of hydrophobic residues with a few hydrogen bonds but a stable interaction. Finally, our electrophysiological response showed the olfactory sensory neurons of G. f. fuscipes tuned to the tsetse repellent components in different sensitivity and selectivity.

Potato (Solanum tuberosum) is a widely consumed staple food crop worldwide whose production is threatened by potato cyst nematodes (PCN). To infect a host, PCN eggs first need to be stimulated to hatch by chemical components in the host root exudates, yet it remains unknown how most root exudate components influence PCN behavior. Here, we evaluated the influence of eight compounds identified by LC-QqQ-MS in the root exudate of potato on the hatching response of the PCN, Globodera rostochiensis at varying doses. The eight compounds included the amino acids tyrosine, tryptophan and phenylalanine; phytohormones zeatin and methyl dihydrojasmonate; steroidal glycoalkaloids α-solanine and α-chaconine and the steroidal alkaloid solanidine. We additionally tested two other Solanaceae steroidal alkaloids, solasodine and tomatidine, previously identified in the root exudates of tomato, an alternative host for PCN. In dose-response assays with the individual compounds, the known PCN hatching factors α-chaconine and α-solanine stimulated the highest number of eggs to hatch, ∼47 and ∼42%, respectively, whereas the steroidal alkaloids (aglycones), solanidine and solasodine and potato root exudate (PRE) were intermediate, 28% each and 21%, respectively, with tomatidine eliciting the lowest hatching response 13%. However, ∼60% of the hatched juveniles failed to emerge from the cyst, which was compound- and concentration-dependent. The amino acids, phytohormones and the negative control (1% DMSO in water), however, were generally non-stimulatory. The use of steroidal glycoalkaloids and their aglycones in the suicidal hatching of PCN offers promise as an environmentally sustainable approach to manage this pest.

Jingmen tick virus (JMTV) is an arbovirus with a multisegmented genome related to those of unsegmented flaviviruses. The virus first described in Rhipicephalus microplus ticks collected in Jingmen city (Hubei Province, China) in 2010 is associated with febrile illness in humans. Since then, the geographic range has expanded to include Trinidad and Tobago, Brazil, and Uganda. However, the ecology of JMTV remains poorly described in Africa. We screened adult ticks (n = 4550, 718 pools) for JMTV infection by reverse transcription polymerase chain reaction (RT-PCR). Ticks were collected from cattle (n = 859, 18.88%), goats (n = 2070, 45.49%), sheep (n = 1574, 34.59%), and free-ranging tortoises (Leopard tortoise, Stigmochelys pardalis) (n = 47, 1.03%) in two Kenyan pastoralist-dominated areas (Baringo and Kajiado counties) with a history of undiagnosed febrile human illness. Surprisingly, ticks collected from goats (0.3%, 95% confidence interval (CI) 0.1–0.5), sheep (1.8%, 95% CI 1.2–2.5), and tortoise (74.5%, 95% CI 60.9–85.4, were found infected with JMTV, but ticks collected from cattle were all negative. JMTV ribonucleic acid (RNA) was also detected in blood from tortoises (66.7%, 95% CI 16.1–97.7). Intragenetic distance of JMTV sequences originating from tortoise-associated ticks was greater than that of sheep-associated ticks. Phylogenetic analyses of seven complete-coding genome sequences generated from tortoise-associated ticks formed a monophyletic clade within JMTV strains from other countries. In summary, our findings confirm the circulation of JMTV in ticks in Kenya. Further epidemiological surveys are needed to assess the potential public health impact of JMTV in Kenya.

Abstract Background Most malaria vectors belong to species complexes. Sibling species often exhibit divergent behaviors dictating the measures that can be deployed effectively in their control. Despite the importance of the Anopheles funestus complex in malaria transmission in sub-Saharan Africa, sibling species have rarely been identified in the past and their vectoring potential remains understudied. Methods We analyzed 1149 wild-caught An. funestus (senso lato) specimens from 21 sites in Kenya, covering the major malaria endemic areas including western, central and coastal areas. Indoor and outdoor collection tools were used targeting host-seeking and resting mosquitoes. The identity of sibling species, infection with malaria Plasmodium parasites, and the host blood meal sources of engorged specimens were analyzed using PCR-based and sequencing methods. Results The most abundant sibling species collected in all study sites were Anopheles funestus (59.8%) and Anopheles rivulorum (32.4%) among the 1062 successfully amplified specimens of the An. funestus complex. Proportionally, An. funestus dominated in indoor collections whilst An. rivulorum dominated in outdoor collections. Other species identified were Anopheles leesoni (4.6%), Anopheles parensis (2.4%), Anopheles vaneedeni (0.1%) and for the first time in Kenya, Anopheles longipalpis C (0.7%). Anopheles funestus had an overall Plasmodium infection rate of 9.7% (62/636), predominantly Plasmodium falciparum (59), with two infected with Plasmodium ovale and one with Plasmodium malariae. There was no difference in the infection rate between indoor and outdoor collections. Out of 344 An. rivulorum, only one carried P. falciparum. We also detected P. falciparum infection in two non-blood-fed An. longipalpis C (2/7) which is the first record for this species in Kenya. The mean human blood indices for An. funestus and An. rivulorum were 68% (93/136) and 64% (45/70), respectively, with feeding tendencies on a broad host range including humans and domestic animals such as cow, goat, sheep, chicken and pig. Conclusions Our findings underscore the importance of active surveillance through application of molecular approaches to unravel novel parasite-vector associations possibly contributed by cryptic species with important implications for effective malaria control and elimination.

Abstract Background Small targets comprising panels of blue and insecticide-treated black netting material each 0.25 × 0.25 m have been shown to attract and kill Glossina fuscipes fuscipes Newstead, 1910 (Diptera: Glossinidae) thereby reducing its population density by over 90% in field trials. However, their attractive ability has not been fully exploited for sampling purposes. Therefore, in this study we assessed the effectiveness of using sticky small targets as sampling tools for G. f. fuscipes in western Kenya. We also determined the influence of colour on the landing response of female and male flies on sticky small targets. Methods Using a series of randomised block experiments, the numbers of tsetse flies caught with sticky small targets were compared with those caught with biconical traps. A negative binomial regression was used to model fly catches. Odds ratios as measures of association between the landing response on the blue or black panel of the sticky small target and the sex of flies were obtained from a multiple logistic regression. Results The results showed that sticky small targets caught 13.5 and 3.6 times more female and male tsetse flies than biconical traps (Z = 9.551, P < 0.0001 and Z = 5.978, P < 0.0001, respectively). Females had a 1.7 times likelihood of landing on the black panel than males (Z = 2.25, P = 0.025). Conclusion This study suggests that sticky small targets are an effective sampling tool for G. f. fuscipes. Therefore, we recommend the use of sticky small targets as an alternative to biconical traps for observational and experimental investigations of G. f. fuscipes.

Abstract Background Anopheles arabiensis is a key vector for the transmission of human malaria in sub-Saharan Africa. Over the past 10,000 years, humans have successfully cultivated grasses and altered the landscape, creating An. arabiensis favourable environments that contain excellent habitats for both larvae and adults. Sugarcane is the most expanding agricultural system in sub-Saharan Africa, and is linked to the increased threat of malaria in rural communities. The prolific production and wind dispersal of sugarcane pollen, together with standing pools of water, often provide, as a result of irrigation, a nutrient-rich environment for the offspring of gravid malaria mosquitoes. Results In the present study, sugarcane pollen-associated volatiles from two cultivars are shown to attract gravid An. arabiensis in a still air two-port olfactometer and stimulate egg laying in an oviposition bioassay. Through combined gas chromatography and electroantennographic detection, as well as combined gas chromatography and mass spectrometric analyses, we identified the bioactive volatiles and generated a synthetic blend that reproduced the full behavioural repertoire of gravid mosquitoes in the Y-tube assay. Two subtractive odour blends, when compared with the full blend, were significantly more attractive. These three and four-component subtractive blends share the compounds (1R)-(+)-α-pinene, nonanal and benzaldehyde, of which, (1R)-(+)-α-pinene and nonanal are found in the attractive odour blends from rice plants and maize pollen. In pairwise comparisons, the rice synthetic odour blend was more attractive to gravid mosquitoes than either of the pollen blends, whereas the pollen blends did not differ in attraction. Conclusions The attraction of gravid females to sugarcane pollen volatiles demonstrated in this study, together with the previously found grass-associated volatiles, raise the potential of developing a bioactive chimeric blend to attract gravid malaria mosquitoes. This is discussed in relation to the development of novel and cost-effective vector control measures.

Abstract Background The edible and medicinal leguminous plant Cassia tora L. (Fabaceae) is known to possess insecticidal properties against a wide range of plant-feeding insects. However, the bioactivity of extracts of this plant and their constituents against vectors of medical importance has been largely unexplored. We investigated the mosquito larvicidal activity of the seed extract and its major anthraquinones against larvae of the African malaria vector Anopheles gambiae (s.s.). Methods Third-fourth instar larval mortality was observed after 24, 48, 72 and 96 h of exposure to varying doses of the extracts, and two anthraquinones isolates identified using liquid chromatography- quadrupole time of flight mass spectrometry (LC-QtoF-MS). The mosquito larval mortality was evaluated relative to the natural insecticide azadirachtin. Results Fractionation of the crude extract decreased mosquito larvicidal activity, however, larvicidal activity increased with increasing dose of the treatment and exposure time. The known anthraquinones aurantio-obtusin and obtusin were identified as key larvicidal compounds. Aurantio-obtusin and obtusin, exhibited similar toxicity to larvae of A. gambiae (s.s.) with LD50 values of 10 and 10.2 ppm, respectively. However, the two anthraquinones were four- and ~ six-fold less potent than that of the crude seed extract and azadirachtin, which had comparable LD50 values of 2.5 and 1.7 ppm, respectively. Conclusion Both aurantio-obtusin and obtusin showed mosquito larvicidal activity which were comparable to their respective fractions although they were less potent relative to the crude extract and azadirachtin. Further studies need to be conducted on C. tora for its exploitation as a potential eco-friendly tool in mosquito larval source reduction.

Abstract Plant volatile signatures are often used as cues by herbivores to locate their preferred hosts. Here, we report on the volatile organic compounds used by the subterranean root-knot nematode (RKN) Meloidogyne incognita for host location. We compared responses of infective second stage juveniles (J2s) to root volatiles of three cultivars and one accession of the solanaceous plant, Capsicum annum against moist sand in dual choice assays. J2s were more attracted to the three cultivars than to the accession, relative to controls. GC/MS analysis of the volatiles identified common constituents in each plant, five of which were identified as α-pinene, limonene, 2-methoxy-3-(1-methylpropyl)-pyrazine, methyl salicylate and tridecane. We additionally identified thymol as being specific to the accession. In dose-response assays, a blend of the five components elicited positive chemotaxis (71–88%), whereas individual components elicited varying responses; Methyl salicylate (MeSA) elicited the highest positive chemotaxis (70–80%), α-pinene, limonene and tridecane were intermediate (54–60%), and 2-methoxy-3-(1-methylpropyl)-pyrazine the lowest (49–55%). In contrast, thymol alone or thymol combined with either the preferred natural plant root volatiles or the five-component synthetic blend induced negative chemotaxis. Our results provide insights into RKN-host plant interactions, creating new opportunities for plant breeding programmes towards management of RKNs.

Oviposition site selection by gravid female insects is an important determinant in species distribution, abundance, and population dynamics. Females may assess the suitability of a potential oviposition substrate by using cues from conspecific or heterospecific individuals already present. Here, we assessed whether the presence of conspecific or heterospecific larvae and parasites influenced oviposition decisions by the stable fly, Stomoxys calcitrans (Linneaus). Using dual and multiple-choice oviposition bioassays, we found that gravid female S. calcitrans avoided substrates with conspecific larvae, the larvae of house flies, Musca domestica (Linneaus), and the mite Macrocheles muscaedomesticae (Scopoli). Avoidance of conspecific and heterospecific larvae persisted in the dark, suggesting that this behavior is mediated by chemical rather than visual cues. When we reared S. calcitrans in the presence of conspecific larvae and the larvae of house flies at different densities we found that this negatively affected emergence time, larval weight, larval survival, pupal weight, pupal survival, and adult weight. We also demonstrated that individuals of S. calcitrans developed in the presence of mites exhibited low egg hatchability, and poor larval and adult survival. Our study provides additional support for the “preference-performance” hypothesis in S. calcitrans, with gravid females preferring to lay eggs on a substrate that will enhance offspring fitness. We recommend that the chemical cues involved in avoidance by gravid female S. calcitrans of substrates with conspecific and heterospecific larvae should be elucidated. This could lead to the discovery of repellent chemicals important for S. calcitrans management.

BackgroundA blend of compounds (pentanoic acid, guaiacol, δ-octalactone and geranylacetone) identified in waterbuck (Kobus defassa) body odour referred to as waterbuck repellent compounds (WRC) and a synthetic repellent 4-methylguaiacol have previously been shown to repel tsetse flies from the morsitans group. However, these repellents have not been evaluated on palpalis group tsetse flies. In this study, we evaluated the effect of these repellents on catches of Glossina fuscipes fuscipes (major vector of human sleeping sickness) in biconical traps and on sticky small targets which are visually attractive to palpalis group flies. The attractive devices were baited with different doses and blends of the repellent compounds. We also assessed the effect of removal of individual constituents in the synthetic blend of WRC on catches of G. f. fuscipes.Methodology/principal findingsThe study was conducted in western Kenya on four islands of Lake Victoria namely Big Chamaunga, Small Chamaunga, Manga and Rusinga. The tsetse fly catches from the treatments were modeled using a negative binomial regression to determine their effect on catches. In the presence of WRC and 4-methylguaiacol (released at ≈2 mg/h and ≈1.4 mg/h respectively), catches of G. f. fuscipes were significantly reduced by 33% (PConclusions/significanceWe conclude that WRC and 4-methylguaiacol reduce catches of G. f. fuscipes at stationary visually attractive traps and suggest that they may serve as broad spectrum repellents for Glossina species. We recommend further studies to investigate the effects of these compounds on reduction of G. f. fuscipes attracted to human hosts as this may lead to development of new strategies of reducing the prevalence and incidence of sleeping sickness.

The composition and structure of microbial communities associated with mosquitoes remain poorly understood despite their important role in host biology and potential to be harnessed as novel strategies for mosquito-borne disease control. We employed MiSeq sequencing of the 16S rRNA gene amplicons to characterize the bacterial flora of field-collected populations of Aedes mcintoshi and Aedes ochraceus, the primary vectors of Rift Valley fever (RVF) virus in Kenya. Proteobacteria (53.5%), Firmicutes (22.0%) and Actinobacteria (10.0%) were the most abundant bacterial phyla accounting for 85.5% of the total sequences. Non-metric multi-dimensional scaling plots based on Bray-Curtis dissimilarities revealed a clear grouping of the samples by mosquito species, indicating that the two mosquito species harbored distinct microbial communities. Microbial diversity, richness and composition was strongly influenced by the site of mosquito collection and overall, Ae. ochraceus had significantly higher microbial diversity and richness than Ae. mcintoshi. Our findings suggest that host species and site of collection are important determinants of bacterial community composition and diversity in RVF virus vectors and these differences likely contribute to the spatio-temporal transmission dynamics of RVF virus.

BackgroundAfrican trypanosomosis, primarily transmitted by tsetse flies, remains a serious public health and economic challenge in sub-Saharan Africa. Interventions employing natural repellents from non-preferred hosts of tsetse flies represent a promising management approach. Although zebras have been identified as non-preferred hosts of tsetse flies, the basis for this repellency is poorly understood. We hypothesized that zebra skin odors contribute to their avoidance by tsetse flies.Methodology/principal findingsWe evaluated the effect of crude zebra skin odors on catches of wild savannah tsetse flies (Glossina pallidipes Austen, 1903) using unbaited Ngu traps compared to the traps baited with two known tsetse fly management chemicals; a repellent blend derived from waterbuck odor, WRC (comprising geranylacetone, guaiacol, pentanoic acid and δ-octalactone), and an attractant comprising cow urine and acetone, in a series of Latin square-designed experiments. Coupled gas chromatography-electroantennographic detection (GC/EAD) and GC-mass spectrometry (GC/MS) analyses of zebra skin odors identified seven electrophysiologically-active components; 6-methyl-5-hepten-2-one, acetophenone, geranylacetone, heptanal, octanal, nonanal and decanal, which were tested in blends and singly for repellency to tsetse flies when combined with Ngu traps baited with cow urine and acetone in field trials. The crude zebra skin odors and a seven-component blend of the EAD-active components, formulated in their natural ratio of occurrence in zebra skin odor, significantly reduced catches of G. pallidipesby 66.7% and 48.9% respectively, and compared favorably with the repellency of WRC (58.1%- 59.2%). Repellency of the seven-component blend was attributed to the presence of the three ketones 6-methyl-5-hepten-2-one, acetophenone and geranylacetone, which when in a blend caused a 62.7% reduction in trap catch of G. pallidipes.Conclusions/significanceOur findings reveal fundamental insights into tsetse fly ecology and the allomonal effect of zebra skin odor, and potential integration of the three-component ketone blend into the management toolkit for tsetse and African trypanosomosis control.

The need for easily biodegradable and less toxic chemicals in drug development and pest control continues to fuel the exploration and discovery of new natural molecules. Like certain plants, some insects can also respond rapidly to microbial infections by producing a plethora of immune-induced molecules that include antibacterial and antifungal peptides/polypeptides (AMPs), among other structurally diverse small molecules. The recent recognition that new natural product-derived scaffolds are urgently needed to tackle life-threatening pathogenic infections has been prompted by the health threats posed by multidrug resistance. Although many researchers have concentrated on the discovery of AMPs, surprisingly, edible insect-produced AMPs/small molecules have received little attention. This review will discuss the recent advances in the identification and bioactivity analysis of insect AMPs, with a focus on small molecules associated with the microbiota of selected African edible insects. These molecules could be used as templates for developing next-generation drugs to combat multidrug-resistant pathogens.

The African weaver ant, Oecophylla longinoda, is used as a biological control agent for the management of pests. The ant has several exocrine glands in the abdomen, including Dufour’s, poison, rectal, and sternal glands, which are associated with pheromone secretions for intra-specific communication. Previous studies have analyzed the gland secretions of Dufour’s and poison glands. The chemistry of the rectal and sternal glands is unknown. We re-analyzed the secretions from Dufour’s and poison glands plus the rectal and sternal glands to compare their chemistries and identify additional components. We used the solid-phase microextraction (SPME) technique to collect gland headspace volatiles and solvent extraction for the secretions. Coupled gas chromatography–mass spectrometry (GC-MS) analysis detected a total of 78 components, of which 62 were being reported for the first time. These additional components included 32 hydrocarbons, 12 carboxylic acids, 5 aldehydes, 3 alcohols, 2 ketones, 4 terpenes, 3 sterols, and 1 benzenoid. The chemistry of Dufour’s and poison glands showed a strong overlap and was distinct from that of the rectal and sternal glands. The different gland mixtures may contribute to the different physiological and behavioral functions in this ant species.

Globally, there is growing interest to integrate insect-derived ingredients into food products. Knowledge of consumer perception to these food products is growing rapidly in the literature, but similar knowledge on the use of oils from African edible insects remains to be established. In this study, we (1) compared the chemistry of the oils from two commonly consumed grasshoppers, the desert locust Schistocerca gregaria and the African bush-cricket Ruspolia differens with those of olive and sesame oils; (2) compared the proximate composition of a baked product (cookie) prepared from the oils; (3) identified the potential volatiles and fatty acids contributing to the aroma and taste; and (4) examined acceptance and willingness to pay (WTP) for the baked product among consumers with no previous experience of entomophagy. Our results showed that the insect oils were compositionally richer in omega-3 fatty acids, flavonoids, and vitamin E than the plant oils. Proximate analysis and volatile chemistry revealed that differences in aroma and taste of the cookies were associated with their sources of oils. Consumers’ acceptance was high for cookies prepared with R. differens (95%) and sesame (89%) oils compared to those with olive and S. gregaria oils. Notably, cookies prepared with insect oils had more than 50% dislike in aroma and taste. Consumers’ willingness to pay for cookies prepared with insect oils was 6–8 times higher than for cookies containing olive oil, but 3–4 times lower than cookies containing sesame oil. Our findings show that integrating edible insect oils into cookies, entices people to ‘‘take the first step” in entomophagy by decreasing insect-based food products neophobia, thereby, contributing to consumers’ acceptance of the baked products. However, future research should explore the use of refined or flavored insect oils for bakery products to reduce off-flavors that might have been perceived in the formulated food products

Abstract Background Lack of effective vaccines and therapeutics for important arboviral diseases such as Rift Valley fever (RVF) and dengue, necessitates continuous monitoring of vector populations for infections in them. Plant-based lures as surveillance tools has the potential of targeting mosquitoes of both sexes and females of varied physiological states; yet such lures are lacking for vectors of these diseases. Here, we present evidence of the effectiveness of linalool oxide (LO), a single plant-based lure previously developed for malaria vectors in trapping RVF vectors, Aedes mcintoshi and Aedes ochraceus, and dengue vector, Aedes aegypti. Methods For RVF vectors, we used CDC traps to evaluate the performance of LO against three vertebrate-based lures: CO2 (dry ice), BioGent (BG) lure, and HONAD (a blend of aldehydes) in 2 experiments with Completely Randomized design: 1) using unlit CDC traps baited separately with LO, HONAD and BG-lure, and unlit CDC trap + CO2 and lit CDC trap as controls, 2) similar treatments but with inclusion of CO2 to all the traps. For dengue vectors, LO was evaluated against BG lure using BG sentinel traps, in a 3 × 6 Latin Square design, first as single lures and then combined with CO2 and traps baited with CO2 included as controls. Trap captures were compared between the treatments using Chi square and GLM. Results Low captures of RVF vectors were recorded for all lures in the absence of CO2 with no significant difference between them. When combined with CO2, LO performance in trapping these vectors was comparable to BG-lure and HONAD but it was less effective than the lit CDC trap. In the absence of CO2, LO performed comparably with the BG-lure in trapping female Ae. aegypti, but with significantly higher males recorded in traps baited with the plant-based lure. When CO2 was added, LO was significantly better than the BG-lure with a 2.8- fold increase in captures of male Ae. aegypti. Conclusions These results highlight the potential of LO as a generalist plant-based lure for mosquito disease vectors, pending further assessment of possible specificity in their response profile to the different stereoisomers of this compound.

Abstract Background Cues that guide gravid Anopheles gambiae sensu lato to oviposition sites can be manipulated to create new strategies for monitoring and controlling malaria vectors. However, progress towards identifying such cues is slow in part due to the lack of appropriate tools for investigating long-range attraction to putative oviposition substrates. This study aimed to develop a relatively easy-to-use bioassay system that can effectively analyse chemical attraction of gravid Anopheles gambiae sensu stricto. Methods BG-Sentinel™ mosquito traps that use fans to dispense odourants were modified to contain aqueous substrates. Choice tests with two identical traps set in an 80 m2 screened semi-field system were used to analyse the catch efficacy of the traps and the effectiveness of the bioassay. A different batch of 200 gravid An. gambiae s.s. was released on every experimental night. Choices tested were (1) distilled versus distilled water (baseline) and (2) distilled water versus soil infusion. Further, comparisons were made of distilled water and soil infusions both containing 150 g/l of Sodium Chloride (NaCl). Sodium Chloride is known to affect the release rate of volatiles from organic substrates. Results When both traps contained distilled water, 45 % (95 confidence interval (CI) 33–57 %) of all released mosquitoes were trapped. The proportion increased to 84 % (95 CI 73–91 %) when traps contained soil infusions. In choice tests, a gravid female was twice as likely to be trapped in the test trap with soil infusion as in the trap with distilled water (odds ratio (OR) 1.8, 95 % CI 1.3–2.6). Furthermore, the attraction of gravid females towards the test trap with infusion more than tripled (OR 3.4, 95 % CI 2.4–4.8) when salt was added to the substrates. Conclusion Minor modifications of the BG-Sentinel™ mosquito trap turned it into a powerful bioassay tool for evaluating the orientation of gravid mosquitoes to putative oviposition substrates using olfaction. This study describes a useful tool for investigating olfactory attraction of gravid An. gambiae s.s. and provides additional evidence that gravid mosquitoes of this species are attracted to and can be baited with attractive substrates such as organic infusions over a distance of several metres.

Abstract Background Effective surveillance and estimation of the biting fraction of Aedes aegypti is critical for accurate determination of the extent of virus transmission during outbreaks and inter-epidemic periods of dengue and chikungunya fever. Here, we describe the development and use of synthetic human odor baits for improved sampling of adult Ae. aegypti, in two dengue and chikungunya fevers endemic areas in Kenya; Kilifi and Busia counties. Methods We collected volatiles from the feet and trunks of two female and two male volunteers aged between 25 and 45 years. We used coupled gas chromatography- electroantennographic detection (GC/EAD) analysis to screen for antennally-active components from the volatiles and coupled GC-mass spectrometry (GC/MS) to identify the EAD-active components. Using randomized replicated designs, we compared the efficacies of Biogents (BG) sentinel traps baited with carbon dioxide plus either single or blends of the identified compounds against the BG sentinel trap baited with carbon dioxide plus the BG commercial lure in trapping Ae. aegypti. The daily mosquito counts in the different traps were subjected to negative binomial regression following the generalized linear models procedures. Results A total of ten major EAD-active components identified by GC/MS as mainly aldehydes and carboxylic acids, were consistently isolated from the human feet and trunk volatiles from at least two volunteers. Field assays with synthetic chemicals of the shared EAD-active components identified from the feet and trunk gave varying results. Ae. aegypti were more attracted to carbon dioxide baited BG sentinel traps combined with blends of aldehydes than to similar traps combined with blends of carboxylic acids. When we assessed the efficacy of hexanoic acid detected in odors of the BG commercial lure and volunteers plus carbon dioxide, trap captures of Ae. aegypti doubled over the trap baited with the commercial BG lure. However, dispensing aldehydes and carboxylic acids together in blends, reduced trap captures of Ae. aegypti by ~45%-50%. Conclusions Our results provide evidence for roles of carboxylic acids and aldehydes in Ae. aegypti host attraction and also show that of the carboxylic acids, hexanoic acid is a more effective lure for the vector than the BG commercial lure.

The global spread of vector-borne diseases remains a worrying public health threat, raising the need for development of new combat strategies for vector control. Knowledge of vector ecology can be exploited in this regard, including plant feeding; a critical resource that mosquitoes of both sexes rely on for survival and other metabolic processes. However, the identity of plant species mosquitoes feed on in nature remains largely unknown. By testing the hypothesis about selectivity in plant feeding, we employed a DNA-based approach targeting trnH-psbA and matK genes and identified host plants of field-collected Afro-tropical mosquito vectors of dengue, Rift Valley fever and malaria being among the most important mosquito-borne diseases in East Africa. These included three plant species for Aedes aegypti (dengue), two for both Aedes mcintoshi and Aedes ochraceus (Rift Valley fever) and five for Anopheles gambiae (malaria). Since plant feeding is mediated by olfactory cues, we further sought to identify specific odor signatures that may modulate host plant location. Using coupled gas chromatography (GC)-electroantennographic detection, GC/mass spectrometry and electroantennogram analyses, we identified a total of 21 antennally-active components variably detected by Ae. aegypti, Ae. mcintoshi and An. gambiae from their respective host plants. Whereas Ae. aegypti predominantly detected benzenoids, Ae. mcintoshi detected mainly aldehydes while An. gambiae detected sesquiterpenes and alkenes. Interestingly, the monoterpenes β-myrcene and (E)-β-ocimene were consistently detected by all the mosquito species and present in all the identified host plants, suggesting that they may serve as signature cues in plant location. This study highlights the utility of molecular approaches in identifying specific vector-plant associations, which can be exploited in maximizing control strategies such as such as attractive toxic sugar bait and odor-bait technology.

Abstract Background Methods currently used in sampling adult Aedes aegypti, the main vector of dengue and chikungunya viruses are limited for effective surveillance of the vector and accurate determination of the extent of virus transmission during outbreaks and inter - epidemic periods. Here, we document the use of natural human skin odours in baited traps to improve sampling of adult Ae. aegypti in two different endemic areas of chikungunya and dengue in Kenya – Kilifi and Busia Counties. The chemistry of the volatiles released from human odours and the Biogent (BG)-commercial lure were also compared. Methods Cotton socks and T-shirts were used to obtain natural human skin volatiles from the feet and trunk of three volunteers (volunteers 1 and 2 in Kilifi and volunteers 2 and 3 in Busia). Using Latin square design, we compared the efficacies of BG sentinel traps baited with carbon dioxide plus (a) no bait, (b) human feet volatiles, (c) human trunk volatiles each against (c) a control (Biogent commercial lure) at the two sites. Coupled gas chromatography-mass spectrometry (GC-MS) was used to identify and compare candidate attractants released by the commercial lure and human odours. Results Ae. aegypti captured in the trap baited with feet odours from volunteer 2 and trunk odours from the same volunteer were significantly higher than in the control trap in Busia and Kilifi respectively, [IRR = 5.63, 95% CI: 1.15 - 28.30, p = 0.030] and [IRR = 3.99, 95% CI: 0.95-16.69, p = 0.049]. At both sites, Ae. aegypti captures in traps baited with either the feet or trunk odours from volunteers 1 and 3 were not significantly different from the control. Major qualitative differences were observed between the chemical profiles of human odours and the commercial BG-lure. Aldehydes, fatty acids and ketones dominated human odour profiles, whereas the BG-lure released mainly hexanoic acid. Conclusions Our results suggest that additional candidate attractants are present in human skin volatiles which can help to improve the efficacy of lures for trapping and surveillance of Ae. aegypti.