Search

Sustainable agriculture is crucial to ensuring food and nutritional security and improving livelihoods of millions across Africa. However, agricultural production is constrained by native and invasive pests. Indiscriminate insecticide use is often the first line of action to manage these pests, harming human and animal health, and impacting biodiversity. Thus, shifting to more ecologically sustainable alternatives is key to biodiversity conservation and sustainable agriculture. Biological control is a core component of integrated pest management (IPM) strategies, and natural enemies, like parasitoid wasps, are considered one of the most environmentally sustainable means of managing pests. However, the Anthropocene plays a significant role in the success and sustainability of this approach. Therefore, here, we provide a look at the influence of anthropological activities, the scope of the application of parasitoids as biocontrol agents of invasive pests, and the current trends in parasitoid utilization in Africa., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

The zoophytophagous mirid predator Nesidiocoris tenuis and the ectoparasitoid Stenomesius japonicus are important biological control agents for several agricultural pests including the invasive leafminer, Phthorimaea absoluta, a destructive pest of Solanaceous crops especially tomato in sub-Saharan Africa. However, little is known about how feeding by N. tenuis can influence the tritrophic interactions in the tomato plant. Here, we tested the hypothesis that N. tenuis phytophagy would influence the tritrophic olfactory interactions between the host plant tomato and pest, predator, and parasitoid. In olfactometer assays, P. absoluta females and N. tenuis adults were both attracted to constitutive volatiles released by the tomato plant. Whereas females of P. absoluta avoided volatiles released by N. tenuis-infested plants, S. japonicus females and N. tenuis adults were attracted to the induced volatiles. In coupled gas chromatography-electroantennographic detection (GC-EAD) recordings of intact and N. tenuis-infested plant volatiles, antennae of P. absoluta and S. japonicus females both detected eight components, whereas N. tenuis adults detected seven components which were identified by GC-mass spectrometry (GC-MS) as terpenes and green leaf volatiles (GLVs). Dose-response olfactometer bioassays revealed that the responses of P. absoluta, N. tenuis, and S. japonicus varied with the composition and concentration of blends and individual compounds tested from N tenuis-induced volatiles. Females of P. absoluta showed no preference for an eight-component blend formulated from the individual repellents including hexanal, (Z)-3-hexenyl butanoate, and δ-elemene identified in the volatiles. On the other hand, S. japonicus females were attracted to an eight-component blend including the attractants (E)-2-hexenal, (Z)-3-hexenol, methyl salicylate, β-phellandrene, and (E)-caryophyllene. Likewise, N. tenuis adults were attracted to a seven-component blend including the attractants β-phellandrene, δ-elemene, and (E)-caryophyllene identified in the volatiles. Our findings suggest that there is potential for the use of terpenes and GLVs to manage the insects in the tritrophic interaction., (© 2024. The Author(s).)

Plastic waste has recently become a major global environmental concern and one of the biggest challenges has been seeking for alternative management options. Several studies have revealed the potential of several coleopteran species to degrade plastics, and this is the first research paper on plastic-degradation potential by lesser mealworms from Africa. This study evaluated the whole mitogenomic profile of the lesser mealworm to further identify the insect. The ability of the mealworm to consume Polystyrene (PS) was also evaluated alongside its associated gut microbiota diversity. Our results showed a complete circular mitochondrial genome which clustered closely to the Alphitobius genus but also suggested that our insect might be a new subspecies which require further identification. During the PS feeding trials, overall survival rates of the larvae decreased when fed a sole PS diet while PS intake was observed to increase over a 30-day period. The predominant bacteria observed in larvae fed PS diets were Kluyvera, Lactococcus, Klebsiella, Enterobacter, and Enterococcus, while Stenotrophomonas dominated the control diet. These findings demonstrated that the newly identified lesser mealworm can survive on a PS diet and has a consortium of important bacteria strongly associated with PS degradation. This work provides a better understanding of bioremediation applications, paving the way for further research into the metabolic pathways of plastic-degrading microbes and bringing hope to solving plastic waste pollution while providing high-value insect protein towards a circular economy., (© 2024. The Author(s).)

Endophytic fungal-based biopesticides are sustainable and ecologically-friendly biocontrol agents of several pests and diseases. However, their potential in managing tomato fusarium wilt disease (FWD) remains unexploited. This study therefore evaluated effectiveness of nine fungal isolates against tomato fusarium wilt pathogen, Fusarium oxysporum f. sp. lycopersici (FOL) in vitro using dual culture and co-culture assays. The efficacy of three potent endophytes that inhibited the pathogen in vitro was assessed against FWD incidence, severity, and ability to enhance growth and yield of tomatoes in planta. The ability of endophytically-colonized tomato (Solanum lycopersicum L.) plants to systemically defend themselves upon exposure to FOL were also assessed through defence genes expression using qPCR. In vitro assays showed that endophytes inhibited and suppressed FOL mycelial growth better than entomopathogenic fungi (EPF). Endophytes Trichoderma asperellum M2RT4, Hypocrea lixii F3ST1, Trichoderma harzianum KF2R41, and Trichoderma atroviride ICIPE 710 had the highest (68.84-99.61%) suppression and FOL radial growth inhibition rates compared to EPF which exhibited lowest (27.05-40.63%) inhibition rates. Endophytes T. asperellum M2RT4, H. lixii F3ST1 and T. harzianum KF2R41 colonized all tomato plant parts. During the in planta experiment, endophytically-colonized and FOL-infected tomato plants showed significant reduction of FWD incidence and severity compared to non-inoculated plants. In addition, these endophytes contributed to improved growth promotion parameters and yield. Moreover, there was significantly higher expression of tomato defence genes in T. asperellum M2RT4 colonized than in un-inoculated tomato plants. These findings demonstrated that H. lixii F3ST1 and T. asperellum M2RT4 are effective biocontrol agents against FWD and could sustainably mitigate tomato yield losses associated with fusarium wilt., (© 2024. The Author(s).)

The false codling moth (FCM), Thaumatotibia leucotreta, is a major quarantine pest native to Africa. Physical postharvest phytosanitary measures such as cold and heat treatments are championed to control its spread to new regions. However, the molecular changes that T. leucotreta undergoes as it attempts to adjust to its surroundings during the treatments and withstand the extreme temperatures remain largely unknown. The current study employs RNA-seq using the next-generation Illumina HiSeq platform to produce transcriptome profiles for differential gene expression analysis of T. leucotreta larvae under thermal stress. The transcriptome assembly analysis revealed 226,067 transcripts, clustering into 127,018 unigenes. In comparison to the 25 °C treated group, 874, 91, 159, and 754 individual differentially expressed genes (DEGs) co-regulated at -10, 0, 40, and 50 °C, respectively were discovered. Annotation of the DEGs by gene ontology (GO) revealed several genes, previously implicated in low and high-temperature stresses, including heat shock proteins, cytochrome P450, cuticle proteins, odorant binding proteins, and immune system genes. Kyoto Encyclopedia of Genes and Genomics (KEGG) classification analysis revealed that substantive DEGs were those involved in metabolic pathways such as thiamine, purine, folate, and glycerolipid metabolism pathways. The RT-qPCR validation of several significantly up- and down-regulated DEGs showed congruence between RNA-seq and qPCR data. This baseline study lays a foundation for future research into the molecular mechanisms underlying T. leucotreta's cold/heat tolerance by providing a thorough differential gene expression analysis that has identified multiple genes that may be associated with the insect's ability to withstand cold and heat., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Edible grasshopper, Ruspolia ruspolia , has nutritional and cherished cultural and economic importance to people from diverse cultures, particularly in over 20 African countries. It is consumed at home or commercially traded as sautéed, deep-fried, or boiled products. However, there is limited information on the hygiene practices of the vendors and the implications on the microbial safety of the final product. This research aimed at assessing the food safety knowledge, handling practices and shelf life of edible long-horned grasshopper products among vendors and the microbial safety of ready-to-eat products sold in 12 different markets in Uganda. Samples of raw, deep-fried and boiled grasshoppers were randomly collected from 74 vendors (62% street and 38% market vendors) and subjected to microbial analysis. Over 85% of the vendors surveyed had no public health food handler's certificate and >95% had limited post-harvest handling knowledge. Total aerobic bacteria (7.30-10.49 Log10 cfu/g), Enterobacteriaceae (5.53-8.56 Log10 cfu/g), yeasts and molds (4.96-6.01 Log10 cfu/g) total counts were significantly high and above the acceptable Codex Alimentarius Commission and Food Safety Authority of Ireland (FSAI) limits for ready-to-eat food products. Eight key pathogenic bacteria responsible for foodborne diseases were detected and these isolates were characterized as Bacillus cereus, Hafnia alvei, Serratia marcescens, Staphylococcus aureus, S. xylosus, S. scuiri, S. haemolyticus , and Pseudomonas aeruginosa . Findings from this study highlight the urgent need to create local and national food safety policies for the edible grasshopper " nsenene " subsector to regulate and guide street and market vending along the value chain, to prevent the transmission of foodborne diseases to consumers., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Mugo-Kamiri, Imungi, Njue, Diiro, Ombura, Akutse, Chrysantus, Khamis and Subramanian.)

Phthorimaea absoluta is a global constraint to tomato production and can cause up to 100% yield loss. Farmers heavily rely on synthetic pesticides to manage this pest. However, these pesticides are detrimental to human, animal, and environmental health. Therefore, exploring eco-friendly, sustainable Integrated Pest Management approaches, including biopesticides as potential alternatives, is of paramount importance. In this context, the present study (i) evaluated the efficacy of 10 Bacillus thuringiensis isolates, neem, garlic, and fenugreek; (ii) assessed the interactions between the most potent plant extracts and B. thuringiensis isolates, and (iii) evaluated the gut microbial diversity due to the treatments for the development of novel formulations against P. absoluta. Neem recorded the highest mortality of 93.79 ± 3.12% with an LT 50 value of 1.21 ± 0.24 days, Bt HD263 induced 91.3 ± 3.68% mortality with LT 50 of 2.63 ± 0.11 days, compared to both Bt 43 and fenugreek that caused < 50% mortality. Larval mortality was further enhanced to 99 ± 1.04% when Bt HD263 and neem were combined. Furthermore, the microbiome analyses showed that Klebsiella, Escherichia and Enterobacter had the highest abundance in all treatments with Klebsiella being the most abundant. In addition, a shift in the abundance of the bacterial genera due to the treatments was observed. Our findings showed that neem, garlic, and Bt HD263 could effectively control P. absoluta and be integrated into IPM programs after validation by field efficacy trials., (© 2024. The Author(s).)

This study sought to evaluate the genetic diversity of two invasive Phthorimaea species ( Phthorimaea operculella Zeller and Phthorimaea absoluta Meyrick), and identify potential niche overlap of both species. The complete mitogenomes of P. operculella and P. absoluta were sequenced and compared. Furthermore, the diversity within the family Gelechiidae was assessed. Subsequently, two species distribution models (MaxEnt and BIOCLIM) were used to identify niche overlaps of both species globally. The complete mitogenomes of both species were similar in size and structure, with a pairwise identity of 92.3%. The models showed a niche overlap of both species and revealed areas of marginal to high suitability for both pests in countries where they have not been reported. Therefore, these results present a compelling case for a deeper genetic and ecological investigation of the Gelechiidae family for taxonomic harmonization, an early warning for surveillance, stricter phytosanitary considerations and preventive management against the spread of the pests., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors.)

Phthorimaea absoluta is an invasive solanaceous plant pest with highly devastating effects on tomato plant. Heavy reliance on insecticide use to tackle the pest has been linked to insecticide resistance selection in P. absoluta populations. To underline insights on P. absoluta insecticide resistance mechanisms to diamides and avermectins, we evaluated the transcriptomic profile of parental (field-collected) and F8 (lab-reared) populations. Furthermore, to screen for the presence of organophosphate and pyrethroid resistance, we assessed the gene expression levels of acetylcholinesterase (ace1) and para-type voltage-gated sodium channel (VGSG) genes in the F1 to F8 lab-reared progeny of diamide and avermectin exposed P. absoluta field-collected populations. The VGSG gene showed up-regulation in 12.5% and down-regulation in 87.5% of the screened populations, while ace1 gene showed up-regulation in 37.5% and down-regulation in 62.5% of the screened populations. Gene ontology of the differentially expressed genes from both parental and eighth generations of diamide-sprayed P. absoluta populations revealed three genes involved in the metabolic detoxification of diamides in P. absoluta. Therefore, our study showed that the detoxification enzymes found could be responsible for P. absoluta diamide-based resistance, while behavioural resistance, which is stimulus-dependent, could be attributed to P. absoluta avermectin resistance., (© 2024. The Author(s).)

Background: The soil biota consists of a complex assembly of microbial communities and other organisms that vary significantly across farming systems, impacting soil health and plant productivity. Despite its importance, there has been limited exploration of how different cropping systems influence soil and plant root microbiomes. In this study, we investigated soil physicochemical properties, along with soil and maize-root microbiomes, in an agroecological cereal-legume companion cropping system known as push-pull technology (PPT). This system has been used in agriculture for over two decades for insect-pest management, soil health improvement, and weed control in sub-Saharan Africa. We compared the results with those obtained from maize-monoculture (Mono) cropping system., Results: The PPT cropping system changed the composition and diversity of soil and maize-root microbial communities, and led to notable improvements in soil physicochemical characteristics compared to that of the Mono cropping system. Distinct bacterial and fungal genera played a crucial role in influencing the variation in microbial diversity within these cropping systems. The relative abundance of fungal genera Trichoderma, Mortierella, and Bionectria and bacterial genera Streptomyces, RB41, and Nitrospira were more enriched in PPT. These microbial communities are associated with essential ecosystem services such as plant protection, decomposition, carbon utilization, bioinsecticides production, nitrogen fixation, nematode suppression, phytohormone production, and bioremediation. Conversely, pathogenic associated bacterial genus including Bryobacter were more enriched in Mono-root. Additionally, the Mono system exhibited a high relative abundance of fungal genera such as Gibberella, Neocosmospora, and Aspergillus, which are linked to plant diseases and food contamination. Significant differences were observed in the relative abundance of the inferred metabiome functional protein pathways including syringate degradation, L-methionine biosynthesis I, and inosine 5'-phosphate degradation., Conclusion: Push-pull cropping system positively influences soil and maize-root microbiomes and enhances soil physicochemical properties. This highlights its potential for agricultural and environmental sustainability. These findings contribute to our understanding of the diverse ecosystem services offered by this cropping system where it is practiced regarding the system's resilience and functional redundancy. Future research should focus on whether PPT affects the soil and maize-root microbial communities through the release of plant metabolites from the intercrop root exudates or through the alteration of the soil's nutritional status, which affects microbial enzymatic activities., (© 2024. The Author(s).)

Although edible rhinoceros beetle ( Oryctes sp. ) larvae are popularly consumed in many countries worldwide, they are prepared using different methods such as boiling, roasting, toasting, and deep-frying, whose effect on nutritional value and microbial safety is scarcely known. Here we investigated the effect of these methods on the nutritional value and microbial safety of Oryctes sp. larvae. Our hypothesis was that cooking the grubs using the four methods had no effect on their nutritional content and microbial loads and diversity. The grubs were analyzed for proximate composition, and fatty and amino acid profiles using standard chemical procedures; and microbial safety using standard culturing procedures. Deep-frying reduced protein and carbohydrate content, but elevated fat content. Boiling lowered ash content, but increased fibre and carbohydrate composition. Roasting and toasting increased protein and ash contents, respectively. Forty fatty acids were detected in the larvae, of which levels of only five were not significantly affected by cooking method, while the levels of the others were differentially affected by the different cooking methods. Amino acid profiles and levels were largely comparable across treatments, but lysine and arginine were higher in all cooked grubs than raw form. All the cooking methods eliminated Enterobacteriaceae, Shigella sp. and Campylobacter sp. from the grubs. Except boiling, all methods reduced total viable count to safe levels. Salmonella sp. were only eliminated by toasting and roasting; while boiling promoted growth of yeast and moulds. Staphylococcus aureus levels exceeded safety limits in all the cooking methods. These findings offer guidance on the type of method to use in preparing the grubs for desired nutritional and safety outcomes., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors.)

The invasive tomato leaf miner, Phthorimaea absoluta, is conventionally controlled through chemical insecticides. However, the rise of insecticide resistance has necessitated sustainable and eco-friendly alternatives. Entomopathogenic fungi (EPF) have shown potential due to their ability to overcome resistance and have minimal impact on non-target organisms. Despite this potential, the precise physiological mechanisms by which EPF acts on insect pests remain poorly understood. To attain a comprehensive understanding of the complex physiological processes that drive the successful control of P. absoluta adults through EPF, we investigated the impacts of different Metarhizium anisopliae isolates (ICIPE 665, ICIPE 20, ICIPE 18) on the pest's survival , cellular immune responses, and gut microbiota under varying temperatures. The study unveiled that ICIPE 18 caused the highest mortality rate among P. absoluta moths, while ICIPE 20 exhibited the highest significant reduction in total hemocyte counts after 10 days at 25°C. Moreover, both isolates elicited notable shifts in P. absoluta's gut microbiota. Our findings revealed that ICIPE 18 and ICIPE 20 compromised the pest's defense and physiological functions, demonstrating their potential as biocontrol agents against P. absoluta in tomato production systems., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Maingi, Akutse, Ajene, Omolo and Khamis.)

Cabbage and Onion production in sub-Saharan Africa face numerous pest constraints that needs to be overcome to feed the rapidly growing population. This study aimed to establish the occurrence, incidence, and severity of soil-dwelling pests of cabbage and onions, and current management practices in five Counties of Kenya. Our findings revealed that most farmers grew hybrid vegetables on a small scale, which were highly dominated by various pest species (Delia platura, Maladera sp., and Agriotes sp. for cabbage and Atherigona orientalis and Urophorus humeralis for onion. The occurrence, incidence and severity of the various pest species on both crops varied considerably. Over 95% of the farmers relied on synthetic insecticides, which were applied weekly or bimonthly with limited success. Our findings demonstrate that invasive and polyphagous A. orientalis and D. platura were the most devastating pests of onion and cabbage without effective control options. Therefore, effective, sustainable, and affordable management strategies are required to control the spread of these pests to other crops in the region., (© 2023. The Author(s).)

Tuta absoluta feeds on solanaceous plants with preference on tomato. Management of the pest is mostly with chemical insecticides. This study identified insecticide resistant populations and predicted resistance to insecticides. Insecticide resistance development was modelled using system thinking, and system dynamics approaches. The model showed the pest resistance development is alarming with an exponential increase of the resistance strength mostly in recent years. Furthermore, we used seven insecticide-resistance gene markers to resolve the population structure and genetic differentiation of insecticide-resistant populations in Kenya. The genes for resistance (knockdown resistance ( kdr ) mutations, acetylcholinesterase (AChE) and voltage gated sodium channel (para)) were detected in all populations. Population structure analyses separated T. absoluta populations into three genetic clusters with resistant genes that are interconnected. A better insight on the population dynamics and the genetic structure T. absoluta resistant genes in Kenya will help estimate resistance strength and determine the most effective pest control strategies., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Authors. Published by Elsevier Ltd.)

The fall armyworm (FAW), Spodoptera frugiperda , an invasive pest originating from the Americas is a serious pest threatening cereal production and food security in Zambia. We studied the prevalence and abundance of natural enemies of FAW in three Agroecological regions (AERs I, II, and III) to identify those that could potentially serve as bio-control agents. Sampling of FAW parasitoids and predators was done along trunk roads at intervals of 10 km. Molecular sequence analysis and morphological characterization were used to identify natural enemies. Over 11 species of FAW natural enemies, including egg, egg-larval, and larval parasitoids, and predators, were identified in Zambia. The mean number of natural enemies and species richness was higher in AER I and IIa. Consequently, egg parasitism was highest in those two regions, at 24.5% and 12.2%, respectively. Larvae parasitism was highest in AER I (4.8%) and AER III (1.9), although no significant differences were observed. The most abundant and widely distributed parasitoid was Drino sp. (Diptera: Tachinidae), while Rhynocoris segmentarius (Germar) (Hemiptera: Reduviidae) and Belanogaster sp. (Hymenoptera: Vespidae) were the most prevalent predators. Our study reveals the presence of two natural enemies belonging to the genus Tiphia and Micromeriella , uncommon to FAW. Significant differences in the number of parasitoids were observed in polycropping, with the highest recovery of 12 ± 10% from maize + cowpeas + pumpkin and watermelon mixed cropping. The higher the rainfall, the lower the number of natural enemies recorded. Variations in rainfall patterns which affect FAW availability, cropping systems and the three AERs may explain natural enemies' species diversity in Zambia. The information provided in this study can aid the development of a national biological control programme for sustainable management of fall armyworm., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The authors SN, FK, PC, SS declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2023 Chipabika, Sohati, Khamis, Chikoti, Copeland, Ombura, Kachapulula, Tonga, Niassy and Sevgan.)

The fall armyworm (FAW) Spodoptera frugiperda , is a voracious pest of cereals native to the Americas and which invaded Africa in 2016. Chemical control is the main management option, which however remains ineffective and unsustainable. Fungal endophytes are increasingly used as alternative for the management of insect pests of economic importance. This study assessed the potential of eight endophytic fungal isolates to colonize maize plant and their ability to promote seedlings growth through seed and foliar inoculations, as well as their suppressive effects on FAW. Fungal colonization rates of different plant parts by the endophytes varied as per the inoculation methods. Beauveria bassiana ICIPE 279 colonized more than 60% of all the seedling parts while B . bassiana G1LU3 only colonized stem (25%) and leaf (5%) tissues through foliar inoculation. Trichoderma atroviride F2S21, T. asperellum M2RT4, T. harzianum F2R41, Trichoderma sp. F2L41, Hypocrea lixii F3ST1 and Fusarium proliferatum F2S51 successfully colonized all the plant parts and therefore were selected and further evaluated through seed inoculation for their endophytic persistence, effect on plant growth, and pathogenicity to Spodoptera frugiperda immature and adult stages. Weekly assessment showed varied effect of the endophytes on maize plant growth parameters compared to the control. During the first week, percentage colonization of the plant parts ranges between 90%-100%, 65%-100%, and 60%-100%, in the roots, stems, and leaves, respectively for all the five tested isolates. However, the colonization pattern/rates significantly decreased over time for H. lixii F3ST1 in the stems and leaves, and for T. harzianum F2R41 in the leaves and for T. asperellum M2RT4 in the roots. In addition, T. harzianum F2R41 outperformed all the other isolates in boosting the plant height, whereas H. lixii F3ST1 and T. asperellum M2RT4 outperformed all the other isolates in increasing the wet and dry shoots weight. Furthermore, the number of egg masses laid on endophytically-colonized maize plants varied among the treatments. Trichoderma asperellum M2RT4 and H. lixii F3ST1 endophytically-colonized maize plants significantly reduced the number of egg masses and the defoliation/feeding rates of the pest compared to the control. Additionally, T. harzianum F2R41 had the highest negative impact on the pupation and adult emergence of S. frugiperda with a female-biased sex ratio. Our findings indicate that T. asperellum M2RT4, T. harzianum F2R41, and H. lixii F3ST1 hold a potential to be developed as endophytic-fungal-based biopesticides for sustainable management of S. frugiperda and as plant growth promoters., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Kinyungu, Agbessenou, Subramanian, Khamis and Akutse.)

Integrated pest management (IPM) strategies are being promoted to suppress tephritid fruit fly infestation and reduce economic damage in mango production. However, research on their economic performance across different mango production scales (measured by the number of mango trees) is limited. This study estimated the economic benefits of IPM practices (parasitoids, orchard sanitation, food bait, biopesticides, male annihilation technique, and their combinations) in Kenya's small-, medium-, and large-scale mango production systems. We used the value-cost ratio (VCR) and net present value methods to estimate the heterogeneous economic performance of IPM practices using data from two unique farm surveys. On average, all IPM practices were profitable across various production scales. However, we found that these practices were more profitable for medium-scale farmers than for small- and large-scale farmers. The results show that farmers need a minimum of 9-17 trees, depending on the practice used, to break even and that there are little to no economic benefits to using IPM practices for farmers with more than 320 mango trees. The male annihilation technique was the most profitable practice, with a VCR of 36, and consequentially, the most adopted practice across all scales of production. Overall, we found significant heterogeneity in the profitability of IPM practices across different scales of production. The reason for the lack of profitability of IPM on large-scale farms remains unclear and warrants further investigation., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Mulungu, Muriithi, Kassie and Khamis.)

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., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Adams, Yusuf, Torto and Khamis.)

Bedbugs have experienced an extraordinary upsurge in the recent past across the world. This cross-sectional study aimed to explore the community perception of the pest outbreaks, the population dynamics, and dispersal patterns under different habitat systems. A survey was conducted within communities in nine counties in Kenya, where geographical coordinates of the sites of bedbug presence were recorded and maximum entropy distribution modelling (MaxEnt) was used to map and predict the potentially suitable habitat, while system thinking and system dynamics approach with Vensim PLE 8.0.9 software was applied to implement bedbug infestation dynamics. Our results indicated that majority of the respondents had ample knowledge on bedbugs and were concerned about the physico-psychologic and socio-economic health effects. Spatial distribution analysis showed regions in Kenya with optimal to suitable for bedbug occurrence in the whole country, and similar results were found at continental level across Africa. Furthermore, infestation dynamics results showed a rapid mobility of bedbug from one house to another. In terms of management strategies, the models showed that the combination of chemical with other control methods was considerably much more effective compared to the use of chemical approach only, appointing integrated pest management strategy as a better intervention approach in controlling the pest., (© 2022. The Author(s).)

Globally, most gut microbiota-related studies have focused on broilers due to their diverse microbial communities compared to that of layer chicken. However, in Africa few studies have been undertaken despite the increasing benefits to the poultry industry. The utilization of Insect-Based diets to improve the gastrointestinal function and gut health in poultry is increasingly gaining global attention. Here, we evaluated the potential roles of commercial black soldier fly larvae-based feeds (BSFLF) in reshaping the abundance, composition and diversity of the gut microbiota of layer chickens using high throughput Oxford nanopore Minion sequencing of the full length bacterial 16S rRNA gene. Two hundred and fifty ISA Brown layer chicks were reared in pens for a period of 20 weeks. The layer pullets were allotted 5 dietary treatments that were formulated as follows: control diet (T1): 100% FM + 0% BSFL, T2: 25% BSFL + 75% FM; T3: 50% BSFL + 50% FM; T4: 75% BSFL + 25% FM, and T5: 100% BSFL + 0% FM. Sampling was done from the eight major regions including oesophagus, crop, proventriculus, gizzard, duodenum, ileum, large intestines and ceca. Out of the 400,064 classified reads analyzed, the most dominant phyla identified across the feed treatments were Firmicutes, Bacteroidetes, Proteobacteria and Actinobacteria. The diet treatment with 100% inclusion levels of BSFL showed the highest intra-species alpha diversity and richness according to Chao1 and Shannon index. Intra-species beta diversity assessment revealed that the diet types significantly influenced the abundance of the microbiota, but differences between most abundant taxa were similar. There was increase in abundance of potentially beneficial bacteria (Lactobacillus, Bacteroides and Enterococcus) with increased inclusion levels of BSFLF in layer pullets diets. Across the different gut segments, Lactobacillus dominated all the eight regions and the ceca was the most diverse segment. Our findings unravel complex gut microbial shift in laying hen fed BSFLF and therefore underpins the potential roles of beneficial bacteria as promising prebiotics and probiotics in reshaping of the gut microbiota to maintain good gut health., (© 2022. The Author(s).)

The efficiency of an autodissemination technique in controlling adult whiteflies, Trialeurodes vaporariorum Westwood (Hemiptera: Aleyrodidae) on tomato, Solunum lycopersicum was investigated with previously identified potent fungal isolates of Metarhizium anisopliae ICIPE 18, ICIPE 62 and ICIPE 69 under screenhouse or semi-field conditions. The autodissemination device was inoculated with dry conidia of the M. anisopliae isolates, while control insects were exposed to a fungus-free device. Sampling for conidia uptake, conidial viability and persistence, and insect mortality was done at 1, 2, 3, 5 and 8 days post-exposure, and collected insects were monitored for mortality over ten days. Overall, mortality was higher in insects exposed to ICIPE 18 (62.8%) and ICIPE 69 (61.8%) than in those exposed to ICIPE 62 (42.6%), with median lethal times, (LT 50 ) ranging between 6.73-8.54 days. The control group recorded the lowest mortality rates (18.9%). A general linear reduction in conidial viability with exposure time was observed, although this was more pronounced with M. anisopliae ICIPE 62. Insects exposed to M. anisopliae ICIPE 69 also recorded the highest conidia uptake, hence selected for further evaluation with a T. vaporariorum attractant volatile organic compound, ( E )-2-hexenal. The volatile inhibited fungal germination in laboratory compatibility tests, therefore, spatial separation of M. anisopliae ICIPE 69 and ( E )-2-hexenal in the autodissemination device was conducted. The inhibitory effects of the volatile were significantly reduced by spatial separation at a distance of 5 cm between the fungus and the volatile, which was found to be more suitable and chosen for the subsequent experiments. Results showed that ( E )-2-hexenal did not influence conidia uptake by the insects, while fungal viability and the subsequent mortality variations were more related to duration of exposure. The fungus-volatile compatibility demonstrated with spatial separation provides a basis for the optimisation of the volatile formulation to achieve better T. vaporariorum suppression with an excellent autodissemination efficiency when used in the management of whiteflies under screenhouse conditions., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Paradza, Khamis, Yusuf, Subramanian and Akutse.)

The oriental fruit fly, Bactrocera dorsalis (Hendel), and marula fruit fly, Ceratitis cosyra (Walker), are major fruit-infesting tephritids across sub-Saharan Africa. Biological control of these pests using parasitic wasps has been widely adopted but with varying levels of success. Most studies investigating host-parasitoid models have focused on functional and evolutionary aspects leaving a knowledge gap about the physiological mechanisms underpinning the efficacy of parasitoids as biocontrol agents of tephritids. To better understand these physiological mechanisms, we investigated changes in the cellular immune responses of C. cosyra and B. dorsalis when exposed to the parasitic wasps, Diachasmimorpha longicaudata (Ashmaed) and Psyttalia cosyrae (Wilkinson). We found that B. dorsalis was more resistant to parasitisation, had a higher hemocyte count, and encapsulated more parasitoid eggs compared to C. cosyra , achieving up to 100% encapsulation when exposed to P. cosyra e. Exposing B. dorsalis to either parasitoid species induced the formation of a rare cell type, the giant multinucleated hemocyte, which was not observed in C. cosyra . Furthermore, compared to P. cosyra e-parasitized larvae, those of both host species parasitized by D. longicaudata had lower encapsulation rates, hemocyte counts and spreading abilities and yielded a higher number of parasitoid progeny with the highest parasitoid emergence (72.13%) recorded in C. cosyra . These results demonstrate that cellular immune responses are central to host-parasitoid interaction in tephritid fruit flies and further suggest that D. longicaudata presents greater potential as a biocontrol agent of B. dorsalis and C. cosyra in horticultural cropping systems., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Gwokyalya, Herren, Weldon, Khamis, Ndlela and Mohamed.)

Long-horned grasshopper (Ruspolia differens Serville) is a tasty delicacy in over 20 African countries. This study evaluated the impact of diverse post-harvest thermal treatment (blanching, boiling, toasting, and deep-frying) on the nutrients, total flavonoid content and sterols preservation of R. differens products. Crude protein, ash, and fibre of R. differens was drastically reduced by deep-frying technique. There was increase in Omega-3 (α-linolenic acid), Omega-6 fatty acid (linoleic and arachidonic acids) and sterols [(22Z)-27-Norergosta-5,22-dien-3β-ol, cholesterol, campesterol, cholest-4-ene-3-one and β-sitosterol] and flavonoids (2-3 folds) during blanching compared to other techniques. The iron and zinc content increased significantly in blanched and boiled products of R. differens. Thus, losses of nutrients, total flavonoid content and sterols during processing of R. differens for food can be mitigated by employing blanching technique, which is cheaper and least time-consuming. The implications of these dietary and therapeutic compounds on human nutrition and health are discussed., (Copyright © 2022 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

The Asian citrus psyllid ( Diaphorina citri Kuwayama) is a key pest of Citrus sp. worldwide, as it acts as a vector for Candidatus Liberibacter asiaticus, the bacterial pathogen that causes citrus Huanglongbing. Diaphorina citri has been reported in Kenya, Tanzania, and more recently in Ethiopia. This study assessed the genetic diversity and phylogeographic structure of the pest to gain insights into the potential sources of its introduction into Africa. Population structure and differentiation of D. citri populations from China, Ethiopia, Kenya, Tanzania, and the USA were assessed using 10 microsatellite loci. Additionally, five new complete mitogenomes of D. citri collected in China, Ethiopia, Kenya, Tanzania, and the USA were analyzed in the context of publicly available sequences. Genotype data grouped the D. citri populations from Kenya and Tanzania in one cluster, and those from Ethiopia formed a separate cluster. The two genetic clusters inferred from genotype data were congruent with mitochondrial sequence data. The mitogenomes from Kenya/Tanzania/China had 99.0% similarity, and the Ethiopia/USA had 99.9% similarity. In conclusion, D. citri populations in eastern Africa have different sources, as the Kenyan and Tanzanian populations probably originated from southeastern Asia, while the Ethiopian population most probably originated from the Americas., Competing Interests: The authors declare no conflict of interest., (© 2022 The Authors. Ecology and Evolution published by John Wiley & Sons Ltd.)

The utilization of insect-based diets to improve gastrointestinal function and gut health in poultry is gaining global attention as a promising feed additive. The objective of this study was to determine the optimal inclusion level of the full-fat black soldier fly larvae (BSFL) and Desmodium intortum (DI) in broiler chicken diets and to evaluate their impact on the microbial community in the gut. The bacterial communities were characterized using Oxford nanopore sequencing of the full-length bacterial 16S rRNA gene. Four dietary treatments, T1 (25% DI + 75% BSFL), T2 (50% DI + 50% BSFL), T3 (75% DI + 25% BSFL) and T4 (100% fishmeal + 0% DI + BSFL), were fed to the broiler chickens for a period of 42 days. Out of the 395,034 classified reads analyzed, the most predominant phyla identified across all the four dietary treatments were Firmicutes (94%), Bacteroidetes (3%), and Proteobacteria (2%). The T1 diet showed the highest alpha diversity and richness according to the Chao1 and Shannon indices. Beta diversity assessment revealed a significant influence of diet on the abundance of the microbiome. There was an increase in beneficial lactic acid bacteria with increasing inclusion of BSFL in the diets. Our findings strongly support the inclusion of BSFL into poultry diet as a promising protein source to reshape the gut microbiota for improved gut health, immune response, and food safety.

Currently, no data exist on the utilization of the newly described cricket species ( Scapsipedus icipe ) meal as additive in food products, though they have high protein (57%) with 88% total digestibility as well as a variety of essential amino acids. This article presents the first report on the effects of processing techniques and the inclusion of cricket meal (CM) on the nutrient and antinutrient properties of four porridge products compared to a popularly consumed commercial porridge flour (CPF). Porridge enriched with CM had significantly higher protein (2-folds), crude fat (3.4-4-folds), and energy (1.1-1.2-folds) levels than the CPF. Fermented cereal porridge fortified with CM had all three types of omega-3 fatty acids compared to the others. The vitamin content across the different porridge products varied considerably. Germinated cereal porridge with CM had significantly higher iron content (19.5 mg/100 g). Zinc levels ranged from 3.1-3.7 mg/100 g across the various treatments. Total flavonoid content varied significantly in the different porridge products. The phytic acid degradation in germinated and fermented porridge products with CM was 67% and 33%, respectively. Thus, the fortification of porridge products with cricket and indigenous vegetable grain powder could be considered an appropriate preventive approach against malnutrition and to reduce incidences in many low-and middle-income countries.

The use of endophytic fungi has dramatically increased plant performance through the enhancement of plant protection against abiotic and biotic stressors. We previously demonstrated that the endophytic fungus Trichoderma asperellum M2RT4 improves tomato defenses against the tomato leafminer Tuta absoluta through the reduction of oviposition, leafmining, pupation, and adult emergence. However, the underlying mechanism by which the presence of this endophytic fungus within tomato host plant affects T. absoluta host selection and life-history traits is unknown. We tested the behavioral responses of T. absoluta in Y-tube olfactometer bioassays and found that females preferred non-inoculated tomato plants against those inoculated by endophytes. Additionally, T. absoluta females were not attracted to non-inoculated infested nor to inoculated-infested tomato plants. Chemical analysis revealed the emission of methyl salicylate in inoculated tomato plant and an increase in the amounts of monoterpenes emitted from non-inoculated infested plants. Additionally, we found that upon herbivory, T. asperellum M2RT4 modulates tomato plant chemistry through the production of ( Z )-jasmone thus activating both salicylic and jasmonic acid defense pathways. Further, T. absoluta females were attracted to monoterpernes including α-pinene, 2-carene, and β-phellandrene but repelled by methyl salicylate. Methyl salicylate could therefore be considered as a good semiochemical-based candidate for sustainable T. absoluta management using a "push-pull" approach. However, in dose-response bioassays, females of T. absoluta did not show any preference to the four component-blend (α-pinene, 2-carene, β-phellandrene, and methyl salicylate). ( Z )-jasmone-treated tomato leaflets significantly reduced the leafmining activity of the pest at the concentration of 10 ng/μL and causing the highest larval mortality rate (83%) with the shortest LT 50 (1.73 days) 7 days post-treatment. T. asperellum M2RT4 effect on herbivore performance was then ( Z )-jasmone-mediated. These findings expand our understanding of how the endophytic fungus T. asperellum M2RT4 could mediate chemical interactions between T. absoluta and its host plant which are potentially important for development of environmentally friendly T. absoluta management programs., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Agbessenou, Akutse, Yusuf and Khamis.)

Wild swarms of the long-horned grasshoppers Ruspolia differens (Serville) which are widely harvested for consumption and sale in Africa are seasonal and unsustainable, hence the need for innovative ways of artificially producing the insects. We investigated the development, survival, and reproduction of R. differens in the laboratory on diets mixed with host plants [Digitaria gayana Kunth, Cynodon dactylon (L.) and Megathyrsus maximus Jacq (Poales: Poaceae); Ageratum conyzoides L. (Asterales: Asteraceae)] identified from guts of their wild conspecifics with a view to developing a suitable diet for artificial mass rearing of the edible insect. A standard diet comprising ground black soldier fly, Hermetia illucens L. (Diptera: Startiomyidae) larvae, soybean flour, maize flour, vitamin premix, and ground bones was tested for rearing R. differens as a control against the same ingredients incorporated with individual powders of the different host plants. Whereas R. differens developed more slowly in the diet mixed with D. gayana than in the control diet; its development was faster in the diet mixed with C. dactylon. Mortalities of R. differens in host plant-based diets were 42.5-52.5%, far lower than in the control diet with 71% mortality. The insects raised on the diet mixed with M. maximus laid approximately twice more eggs compared to R. differens fecundities from the rest of the diets. However, inclusion of host plants in the diets had no detectable influence on R. differens adult weight and longevity. These findings support inclusion of specific host plants in artificial diets used for mass rearing of R. differens to enhance its survival, development, and fecundity., (© The Author(s) 2022. Published by Oxford University Press on behalf of Entomological Society of America.)

Tuta absoluta is one of the most devastating pests of Solanaceae crops in Africa. We previously demonstrated the efficacy of Metarhizium anisopliae isolates ICIPE 18, ICIPE 20 and ICIPE 665 against adult T. absoluta. However, adequate strain selection and accurate spatial prediction are fundamental to optimize their efficacy and formulations before field deployment. This study therefore assessed the thermotolerance, conidial yield and virulence (between 15 and 35 °C) of these potent isolates. Over 90% of conidia germinated at 20, 25 and 30 °C while no germination occurred at 15 °C. Growth of the three isolates occurred at all temperatures, but was slower at 15, 33 and 35 °C as compared to 20, 25 and 30 °C. Optimum temperatures for mycelial growth and spore production were 30 and 25 °C, respectively. Furthermore, ICIPE 18 produced higher amount of spores than ICIPE 20 and ICIPE 665. The highest mortality occurred at 30 °C for all the three isolates, while the LT 50 values of ICIPE 18 and ICIPE 20 were significantly lower at 25 and 30 °C compared to those of ICIPE 665. Subsequently, several nonlinear equations were fitted to the mortality data to model the virulence of ICIPE 18 and ICIPE 20 against adult T. absoluta using the Entomopathogenic Fungi Application (EPFA) software. Spatial prediction revealed suitable locations for ICIPE 18 and ICIPE 20 deployment against T. absoluta in Kenya, Tanzania and Uganda. Our findings suggest that ICIPE 18 and ICIPE 20 could be considered as effective candidate biopesticides for an improved T. absoluta management based on temperature and location-specific approach., (© 2021. The Author(s).)

In the scope of mitigating the negative impacts of pesticide use and managing greenhouse whiteflies, Trialeurodes vaporariorum sustainably, 16 endophytic fungal isolates from five different genera ( Beauveria , Trichoderma , Hypocrea , Bionectria , and Fusarium ) were screened for their ability to colonise two preferred host plant species, namely, tomato ( Solanum lycopersicum L.) and French bean ( Phaseolus vulgaris L.), through seed inoculation. Seven and nine isolates were endophytic to P. vulgaris and S. lycopersicum , respectively, where significant differences in the endophytic colonisation rates were observed among the fungal isolates in P. vulgaris and its plant parts, with a significant interaction between the isolates and plant parts in S. lycopersicum . Hypocrea lixii F3ST1, Trichoderma asperellum M2RT4, Trichoderma atroviride F5S21, and T. harzianum KF2R41 successfully colonised all the plant parts of both hosts and therefore were selected and further evaluated for their endophytic persistence, effect on plant growth, and pathogenicity to T . vaporariorum adults and F1 progeny. The four endophytes remained in both host plants for the 5-week assessment with varied colonisation rates related to the strong interaction with the time, isolates, and plant parts in both hosts. The effect of the same endophytes on the different host growth parameters varied in P. vulgaris and S. lycopersicum , with T. asperellum M2RT4 not boosting the growth in both host plants while T . atroviride F5S21 resulted in enhanced shoot biomass in S. lycopersicum . T. atroviride F5S21 and T . harzianum KF2R41 inoculated S. lycopersicum plants and H. lixii F3ST1, T. asperellum M2RT4, and T . harzianum KF2R41 inoculated P. vulgaris plants had significantly lower oviposition, while nymph development in both hosts was significantly prolonged in all the endophytically-colonised plants. The endophytes H. lixii F3ST1 and T. asperellum M2RT4 significantly reduced the longevity/survival of the exposed T. vaporariorum adults and the progeny in both S. lycopersicum and P. vulgaris . The findings demonstrate the attributes of the various endophytes in host plant growth promotion as well as their effects on the life-history parameters of T. vaporariorum and could consequently be developed as potential endophytic fungal-based biopesticides for the sustainable management of the pest in S. lycopersicum and P. vulgaris cropping systems., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Paradza, Khamis, Yusuf, Subramanian, Ekesi and Akutse.)

The pathogenicity of dry conidia and fungal suspensions of 16 entomopathogenic fungal isolates (10 Metarhizium anisopliae and six Beauveria bassiana ) was evaluated against adults and second instar nymphs of the greenhouse whitefly, Trialeurodes vaporariorum respectively. All the tested isolates were pathogenic to T. vaporariorum and caused mortality of 45-93% against the adults and 24-89% against the nymphs. However, M. anisopliae strains showed higher virulence to both developmental stages as compared to B. bassiana strains. The three most virulent isolates that caused high mortalities in adults were M. anisopliae ICIPE 18, ICIPE 62 and ICIPE 69, with cumulative mortalities of 82, 91 and 93%, and median lethal times (LT 50 ) of 5.20, 5.05 and 4.78 days, respectively. These isolates were further assessed for spore acquisition and retention by the adult insects at 0, 24, 48 and 72 h after exposure to dry conidia spores. There was no significant difference among isolates on their acquisition by the insects, although the effect of time on the number of spores retained by each insect was significant. For M. anisopliae ICIPE 62 and ICIPE 69, spore number was significantly higher immediately after exposure at 0 h than at 24, 48 and 72 h, whereas for M. anisopliae ICIPE 18, the spore number remained constant for all the days. The infected "donor" insects were able to horizontally transmit the acquired spores to uninfected "recipient" insects causing high mortality rates in both donor and recipient groups. Metarhizium anisopliae ICIPE 7, ICIPE 18 and ICIPE 62 were the most virulent isolates against the nymphs in aqueous formulation during the first screening with >80% mortality. However, in 2% (v/v) oil formulations at 1 × 10 8 conidia/ml, canola formulated ICIPE 62, ICIPE 18 and olive formulated ICIPE 18 were the most effective, resulting in 87.8, 88.1 and 99.4% nymphal mortalities respectively and with lower LT 50 . Oil formulations significantly enhanced the efficacy and virulence of the isolates against the nymphs compared to aqueous formulations., Competing Interests: The authors declare no conflict of interest., (© 2021 The Author(s).)

Insect seminal fluid, the non-sperm component of the ejaculate, comprises a variegated set of molecules, including, but not limited to, lipids, proteins, carbohydrates, salts, hormones, nucleic acids, and vitamins. The identity and functional role of seminal fluid proteins (SFPs) have been widely investigated, in multiple species. However, most of the other small molecules in insect ejaculates remain uncharacterized. Metabolomics is currently adopted to deepen our understanding of complex biological processes and in the last 15years has been applied to answer different physiological questions. Technological advances in high-throughput methods for metabolite identification such as mass spectrometry and nuclear magnetic resonance (NMR) are now coupled to an expanded bioinformatics toolbox for large-scale data analysis. These improvements allow for the processing of smaller-sized samples and for the identification of hundreds to thousands of metabolites, not only in Drosophila melanogaster but also in disease vectors, animal, and agricultural pests. In this review, we provide an overview of the studies that adopted metabolomics-based approaches in insects, with a particular focus on the reproductive tract (RT) of both sexes and the ejaculate. Progress in the field of metabolomics will contribute not only to achieve a deeper understanding of the composition of insect ejaculates and how they are affected by endogenous and exogenous factors, but also to provide increasingly powerful tools to decipher the identity and molecular interactions between males and females during and after mating., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Scolari, Khamis and Pérez-Staples.)

Globally, the expansion of livestock and fisheries production is severely constrained due to the increasing costs and ecological footprint of feed constituents. The utilization of black soldier fly (BSF) as an alternative protein ingredient to fishmeal and soybean in animal feed has been widely documented. The black soldier fly larvae (BSFL) used are known to voraciously feed and grow in contaminated organic wastes. Thus, several concerns about their safety for inclusion into animal feed remain largely unaddressed. This study evaluated both culture-dependent sequence-based and 16S rDNA amplification analysis to isolate and identify bacterial species associated with BSFL fed on chicken manure (CM) and kitchen waste (KW). The bacteria species from the CM and KW were also isolated and investigated. Results from the culture-dependent isolation strategies revealed that Providencia sp. was the most dominant bacterial species detected from the guts of BSFL reared on CM and KW. Morganella sp. and Brevibacterium sp. were detected in CM, while Staphylococcus sp. and Bordetella sp. were specific to KW. However, metagenomic studies showed that Providencia and Bordetella were the dominant genera observed in BSFL gut and processed waste substrates. Pseudomonas and Comamonas were recorded in the raw waste substrates. The diversity of bacterial genera recorded from the fresh rearing substrates was significantly higher compared to the diversity observed in the gut of the BSFL and BSF frass (leftovers of the rearing substrates). These findings demonstrate that the presence and abundance of microbiota in BSFL and their associated waste vary considerably. However, the presence of clinically pathogenic strains of bacteria in the gut of BSFL fed both substrates highlight the biosafety risk of potential vertical transmission that might occur, if appropriate pre-and-postharvest measures are not enforced., Competing Interests: MS was employed by the company Hermetia Baruth GmbH. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Shumo, Khamis, Ombura, Tanga, Fiaboe, Subramanian, Ekesi, Schlüter, van Huis and Borgemeister.)

The African citrus psyllid Trioza erytreae is one of the major threats to citrus industry as the vector of the incurable disease known as huanglongbing (HLB) or citrus greening. The psyllid invaded the northwest of the Iberian Peninsula 6 years ago. The invasion alarmed citrus growers in the Mediterranean basin, the largest citrus producing area in Europe, which is still free of HLB. Before our study, no research had been carried out on the genetic diversity of T. erytreae populations that have invaded the Iberian Peninsula and the archipelagos of the Macaronesia (Madeira and the Canary Islands). In this study, combining microsatellites markers and mtDNA barcoding analysis, we characterize the genetic diversity, structure and maternal relationship of these new invasive populations of T. erytreae and those from Africa. Our results suggest that the outbreaks of T. erytreae in the Iberian Peninsula may have derived from the Canary Islands. The populations of T. erytreae that invaded Macaronesia and the Iberian Peninsula are likely to have originated from southern Africa. We anticipate our results to be a starting point for tracking the spread of this invasive pest outside of Africa and to be important for optimizing contingency and eradication plans in newly invaded and free areas., (© 2021. The Author(s).)

The promotion of edible insects, including saturniid caterpillars as potential food source is widely gaining momentum. They are adequately rich in nutrients such as proteins, amino acids, fatty acids, and micronutrients. Despite saturniids being a traditional food source with economic benefits, information on their diversity, host plants and their potential distribution in Africa are lacking, which this study seeks to address. Edible saturniids and their host plants were characterized using specific primers (LepF1/LepR1 and 3F&#95;KIM&#95;F/1R&#95;KIM&#95;R, respectively). Maximum entropy (MaxENT) and GARP (genetic algorithm for ruleset production) models were used to characterize the potential distribution of commonly consumed saturniids under current and future climate scenarios. Seven species of saturniids were recorded from 11 host plants in Kenya: Gonimbrasia zambesina , Gonimbrasia krucki , Bunaea alcinoe , Gonimbrasia cocaulti , Gonimbrasia belina , Gynanisa nigra and Cirina forda . Two morphotypes of G. zambesina and B. alcinoe were recorded. These saturniid caterpillars occur twice a year except for G. cocaulti . Predictive models revealed that tropical and subtropical regions were potentially suitable for B. alcinoe and C. forda. The information generated from this study would be important to guide conservation efforts and their sustainable utilization as food in Africa.

Tuta absoluta (Meyrick) has become a serious menace to sustainable production of tomato in Kenya. A survey was conducted between April 2015 and June 2016 to determine its distribution, abundance, infestation, and damage levels on tomato, and associated natural enemies. Trap counts of T. absoluta moths were recorded in all surveyed 29 counties, which indicated its nationwide distribution irrespective of altitude. Tuta absoluta was present in both open fields and greenhouses. The highest moth/trap/day was 115.38 ± 15.90. Highest leaf infestation was 92.22% and the highest number of mines and larvae per leaf were 3.71 ± 0.28 and 2.16 ± 0.45, respectively. Trap captures in terms of moth/trap/day were linearly and positively related to leaf infestations in open fields (R2 = 0.81) and greenhouses (R2 = 0.61). Highest fruits' infestation and damage were 60.00 and 59.61%, respectively, while the highest number of mines per fruit was 7.50 ± 0.50. Nesidiocoris tenuis (Reuter) and Macrolophus pygmaeus (Rambur) were identified as predators of T. absoluta larvae. Nine species of larval parasitoids were recovered from infested foliage, with a combined parasitism of 7.26 ± 0.65%. Hockeria species was the most dominant (31.25%) and accounted for 12.88 ± 1.47% parasitism. Two species of larval parasitoids, Hockeria and Necremnus were obtained from sentinel plants with an average parasitism of 1.13 ± 0.25. The overall abundance and parasitism rates of recovered natural enemies were low to effectively control the field populations of T. absoluta. These findings form the basis of researching and developing effective and sustainable management strategies for the pest.

Whiteflies (Hemiptera: Aleyrodidae) are devastating agricultural pests of economic importance vectoring pathogenic plant viruses. Knowledge on their diversity and distribution in Kenya is scanty, limiting development of effective sustainable management strategies. The present study is aimed at identifying whitefly pest species present in Kenya across different agroecological zones and establish predictive models for the most abundant species in Africa. Whiteflies were sampled in Kenya from key crops known to be severely infested and identified using 16S rRNA markers and complete mitochondrial genomes. Four whitefly species were identified: Aleyrodes proletella, Aleurodicus dispersus, Bemisia afer and Trialeurodes vaporariorum, the latter being the most dominant species across all the agroecology. The assembly of complete mitogenomes and comparative analysis of all 13 protein coding genes confirmed the identities of the four species. Furthermore, prediction spatial models indicated high climatic suitability of T. vaporariorum in Africa, Europe, Central America, parts of Southern America, parts of Australia, New Zealand and Asia. Consequently, our findings provide information to guide biosecurity agencies on protocols to be adopted for precise identification of pest whitefly species in Kenya to serve as an early warning tool against T. vaporariorum invasion into unaffected areas and guide appropriate decision-making on their management.

The sustainable utilization of black soldier fly (BSF) for recycling organic waste into nutrient-rich biomass, such as high-quality protein additive, is gaining momentum, and its microbiota is thought to play important roles in these processes. Several studies have characterized the BSF gut microbiota in different substrates and locations; nonetheless, in-depth knowledge on community stability, consistency of member associations, pathogenic associations, and microbe-microbe and host-microbe interactions remains largely elusive. In this study, we characterized the bacterial and fungal communities of BSF larval gut across four untreated substrates (brewers' spent grain, kitchen food waste, poultry manure, and rabbit manure) using 16S and ITS2 amplicon sequencing. Results demonstrated that substrate impacted larval weight gain from 30 to 100% gain differences among diets and induced an important microbial shift in the gut of BSF larvae: fungal communities were highly substrate dependent with Pichia being the only prevalent genus across 96% of the samples; bacterial communities also varied across diets; nonetheless, we observed six conserved bacterial members in 99.9% of our samples, namely, Dysgonomonas , Morganella , Enterococcus , Pseudomonas , Actinomyces , and Providencia . Among these, Enterococcus was highly correlated with other genera including Morganella and Providencia . Additionally, we showed that diets such as rabbit manure induced a dysbiosis with higher loads of the pathogenic bacteria Campylobacter . Together, this study provides the first comprehensive analysis of bacterial and fungal communities of BSF gut across untreated substrates and highlights conserved members, potential pathogens, and their interactions. This information will contribute to the establishment of safety measures for future processing of BSF larval meals and the creation of legislation to regulate their use in animal feeds., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Tanga, Waweru, Tola, Onyoni, Khamis, Ekesi and Paredes.)

Background: Insect growth regulators (IGRs) can control insect vector populations by disrupting growth and development in juvenile stages of the vectors. We previously identified and described the curry tree (Murraya koenigii (L.) Spreng) phytochemical leaf extract composition (neplanocin A, 3-(1-naphthyl)-L-alanine, lumiflavine, terezine C, agelaspongin and murrayazolinol), which disrupted growth and development in Anopheles gambiae sensu stricto mosquito larvae by inducing morphogenetic abnormalities, reducing locomotion and delaying pupation in the mosquito. Here, we attempted to establish the transcriptional process in the larvae that underpins these phenotypes in the mosquito., Methods: We first exposed third-fourth instar larvae of the mosquito to the leaf extract and consequently the inherent phytochemicals (and corresponding non-exposed controls) in two independent biological replicates. We collected the larvae for our experiments sampled 24 h before peak pupation, which was 7 and 18 days post-exposure for controls and exposed larvae, respectively. The differences in duration to peak pupation were due to extract-induced growth delay in the larvae. The two study groups (exposed vs control) were consequently not age-matched. We then sequentially (i) isolated RNA (whole larvae) from each replicate treatment, (ii) sequenced the RNA on Illumina HiSeq platform, (iii) performed differential bioinformatics analyses between libraries (exposed vs control) and (iv) independently validated the transcriptome expression profiles through RT-qPCR., Results: Our analyses revealed significant induction of transcripts predominantly associated with hard cuticular proteins, juvenile hormone esterases, immunity and detoxification in the larvae samples exposed to the extract relative to the non-exposed control samples. Our analysis also revealed alteration of pathways functionally associated with putrescine metabolism and structural constituents of the cuticle in the extract-exposed larvae relative to the non-exposed control, putatively linked to the exoskeleton and immune response in the larvae. The extract-exposed larvae also appeared to have suppressed pathways functionally associated with molting, cell division and growth in the larvae. However, given the age mismatch between the extract-exposed and non-exposed larvae, we can attribute the modulation of innate immune, detoxification, cuticular and associated transcripts and pathways we observed to effects of age differences among the larvae samples (exposed vs control) and to exposures of the larvae to the extract., Conclusions: The exposure treatment appears to disrupt cuticular development, immune response and oxidative stress pathways in Anopheles gambiae s.s larvae. These pathways can potentially be targeted in development of more efficacious curry tree phytochemical-based IGRs against An. gambiae s.s mosquito larvae.

Efforts to recycle organic waste using black soldier fly (BSF) larvae into high-quality alternative protein ingredients in animal feeds and organic fertilizers have gained momentum worldwide. However, there is limited information on waste manipulation to increase nutrient retention for enhanced larval performance and frass fertilizer quality. In the present study, brewer's spent grain with a carbon to nitrogen (C/N) ratio of 11 (control) was amended with sawdust to obtain substrates with C/N ratios of 15, 20, 25 and 30. The effects of substrate C/N ratios on BSF larval yield, waste degradation, biomass conversion efficiency, compost maturity and nutrient levels of frass fertilizer were evaluated. Substrates amended with sawdust did not significantly affect waste degradation efficiency and biomass conversion rates of BSF larvae. The wet and dried larval yields were significantly higher for substrates with C/N ratio of 15 compared to the other amended substrates. An amended substrate with C/N ratio of 15 enhanced nutrients uptake by BSF larvae, and increased nitrogen (N) and phosphorus retention in frass compost by 21 and 15%, respectively. Compost maturation time was shortened to five weeks, as indicated by the stable C/N ratios and high seed germination indices. This study has demonstrated that the amendment of the substrate with sawdust to C/N ratio of 15 could generate compost with desirable nutrients for use as high-quality fertilizer for organic farming., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Blood transfusion is a common practice in hospitals. In many diseases, transfusion of red blood cells can be a life-saving measure. Knowing the blood group phenotypes of the donors in any population is important to improve health care services and to better serve patients. We aimed to determine the blood group phenotypes of the regular donors visiting the main blood donation center in Sharjah, United Arab Emirates (UAE). A total of 500 regular blood donors were included, and blood group antigens were tested using serologic methods. Major blood group systems included in this testing were ABO, Rh, Kell, Kidd, Duffy, Lewis, Lutheran, MNS, and P1PK. The most prevalent ABO/D blood type was group O, D+ (29.6%), with the most prevalent phenotype in the Rh blood group system reported as R 1 r (34.4%). In the Kell system, the K-k+ phenotype was reported in 92.4 percent of individuals tested. The rare Duffy null phenotype, Fy(a-b-), was reported in 11.0 percent of the cohort. Most donors (99.0%) were Lu(b+). In the Lewis system, the phenotype Le(a-b+) was seen in 61.8 percent of the donors. MNS system phenotypes were comparable to the reports from Saudi Arabia and India, with M+N+S-s+ being the most prevalent (21.2%). The prevalence of many major blood group phenotypes in the UAE blood donor population showed similarities to results from populations in neighboring countries, but other phenotype prevalences were unique to this population. This finding can be used to build a blood group registry for the local population that can better serve hospitals' transfusion requirements., Blood transfusion is a common practice in hospitals. In many diseases, transfusion of red blood cells can be a life-saving measure. Knowing the blood group phenotypes of the donors in any population is important to improve health care services and to better serve patients. We aimed to determine the blood group phenotypes of the regular donors visiting the main blood donation center in Sharjah, United Arab Emirates (UAE). A total of 500 regular blood donors were included, and blood group antigens were tested using serologic methods. Major blood group systems included in this testing were ABO, Rh, Kell, Kidd, Duffy, Lewis, Lutheran, MNS, and P1PK. The most prevalent ABO/D blood type was group O, D+ (29.6%), with the most prevalent phenotype in the Rh blood group system reported as R 1 r (34.4%). In the Kell system, the K–k+ phenotype was reported in 92.4 percent of individuals tested. The rare Duffy null phenotype, Fy(a–b–), was reported in 11.0 percent of the cohort. Most donors (99.0%) were Lu(b+). In the Lewis system, the phenotype Le(a–b+) was seen in 61.8 percent of the donors. MNS system phenotypes were comparable to the reports from Saudi Arabia and India, with M+N+S–s+ being the most prevalent (21.2%). The prevalence of many major blood group phenotypes in the UAE blood donor population showed similarities to results from populations in neighboring countries, but other phenotype prevalences were unique to this population. This finding can be used to build a blood group registry for the local population that can better serve hospitals’ transfusion requirements.

The edible long-horned grasshopper Ruspolia differens (Serville) is widely distributed and consumed in sub-Saharan Africa. Efficient mass rearing of the edible grasshopper is critical to ensure their sustainable supply for food and nutritional security. Hence, we investigated the effect of temperature on development, survival and reproduction of R. differens under six constant (15, 20, 25, 30, 32 and 35 °C) and fluctuating temperatures. Using Insect Life Cycle Modeling software we fitted, linear and non-linear models to R. differens development, mortality, longevity, and fecundity. The best-fitted functions were compiled for each life stage to yield a phenology model, which was stochastically simulated to estimate the life table parameters. We used the process-based climatic phenology models, and applied establishment risk index (ERI) and generation index (GI) in a geographic information system to map the potential distribution of R. differens under current and future climates. At optimum temperatures of 30-32 °C, egg incubation period was 14-15 days and the developmental time was shortest at 52.5-58 days. Lowest nymphal mortality (3.4-13%) and the highest female fecundity was obtained at 25-30 °C. The optimum temperature for the reproduction ranged between 27 and 30 °C. Most simulated lifetable parameters were at their maximum at 28 °C. Predictive models showed that countries in the East, Central, West, Southern and the Horn of Africa were suitable for establishment of R. differens under current climate scenarios (2000). However, by 2050, climatically suitable areas for the establishment of R. differens were predicted to shrink in the West, Southern and the Horn of Africa than its current distribution. We predict up to three generations per year for R. differens in sub-Saharan Africa under current scenarios which can increase to 4 under future scenarios. The optimum rearing temperatures identified can guide optimization of mass rearing of R. differens., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Endophytic fungi live within plant tissues without causing any harm to the host, promote its growth, and induce systemic resistance against pests and diseases. To mitigate the challenging concealed feeding behavior of immature stages of Tuta absoluta in both tomato (Solanum lycopersicum) and nightshade (Solanum scabrum) host plants, 15 fungal isolates were assessed for their endophytic and insecticidal properties. Twelve isolates were endophytic to both host plants with varied colonization rates. Host plants endophytically-colonized by Trichoderma asperellum M2RT4, Beauveria bassiana ICIPE 706 and Hypocrea lixii F3ST1 outperformed all the other isolates in reducing significantly the number of eggs laid, mines developed, pupae formed and adults emerged. Furthermore, the survival of exposed adults and F1 progeny was significantly reduced by Trichoderma sp. F2L41 and B. bassiana isolates ICIPE 35(4) and ICIPE 35(15) compared to other isolates. The results indicate that T. asperellum M2RT4, B. bassiana ICIPE 706 and H. lixii F3ST1 have high potential to be developed as endophytic-fungal-based biopesticide for the management of T. absoluta.

Fall armyworm, Spodoptera frugiperda (Lepidoptera: Noctuidae), is a key invasive pest of maize and other crops in Africa. Entomopathogenic fungi play an important role in regulating the immature stages of this invasive pest as opposed to synthetic pesticides that are hazardous to human, environment and biodiversity. To tackle the adult stage of the pest (the moth) and to improve on the application strategy of the fungal-based biopesticides, this study evaluated the effect of various entomopathogenic fungi isolates on S. frugiperda moths. Twenty-two isolates (16 Metarhizium anisopliae and 6 Beauveria bassiana) were screened in the laboratory to assess their pathogenicity and virulence against S. frugiperda moths. The compatibility of the most pathogenic isolates with S. frugiperda pheromone FALLTRACT lure, the horizontal transmission of the inoculum among S. frugiperda moths, and the effect on oviposition were also determined under laboratory conditions. All 22 fungal isolates screened were pathogenic to the moths, but the mortality varied significantly among the isolates (P < 0.0001) seven days post-treatment. Beauveria bassiana ICIPE 621 and M. anisopliae ICIPE 7 outperformed all the other isolates by causing 100% mortality of the moths with the lowest LT 50 values of 3.6 ± 0.1 and 3.9 ± 0.0 days, respectively. Both isolates were also found compatible with FALLTRACT lure, as the lure had no effect on the conidial germination in the laboratory. Male and female moths were able to horizontally transmit conidia of both fungal isolates to untreated moths, causing high mortality of S. frugiperda in 'donor' and 'recipient' groups. In addition, the oviposition, hatchability of eggs and longevity of larvae were significantly affected on the fungal infected females. Although single moths still retained high conidial numbers 72 h post-inoculation, the number of conidia decreased with time. These results suggest that ICIPE 7 and ICIPE 621 could be used in combination with S. frugiperda pheromone in an autodissemination approach to suppress S. frugiperda population., (Copyright © 2020 Elsevier Inc. All rights reserved.)

There is a paucity of information on the edible grasshoppers and their host plants in East Africa. This study adopted morphological and molecular analysis to identify edible grasshoppers in Kenya and Uganda. The associated host plants were identified through molecular analysis of the gut contents of the grasshoppers. The cytochrome b and 16s gene primers were used for grasshopper DNA analysis; while matK gene primers were used for plant DNA analysis. All long-horned grasshoppers sampled were identified as Ruspolia differens (Serville) (Orthoptera: Tettigonidae); whereas short-horned grasshoppers were identified as Acanthacris ruficornis (Fabricius) (Orthoptera: Acrididae) and Cyrtacanthacris tatarica (L.) (Orthoptera: Acrididae). Host plants of A. ruficornis were Achyranthes aspera (L.), Centella virgata L.f. Drude, Digitaria gayana (Kunth), Galinsoga quadriradiata Ruiz and Pavon, and Triumfetta pilosa Roth; whereas those of C. tatarica were Alysicarpus rugosus (Willd.) DC and Teramnus uncinatus (L.) SW. Host plants of R. differens were Ageratum conyzoides (L.), Citrus depressa Hayata, Cynodon dactylon (L.), D. gayana, Eragrostis mexicana Hornem, Eucalyptus saligna SM., Indigofera arrecta Hochst. ex A. Rich., Persicaria nepalensis (L.), and Sorghum halepense (L.). Information on the host plants of edible grasshoppers can help in the development of their mass rearing protocols., (© The Author(s) 2020. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Black soldier fly frass fertilizer (BSFFF) is increasingly gaining momentum worldwide as organic fertilizer. However, research on its performance on crop production remains largely unknown. Here, we evaluate the comparative performance of BSFFF and commercial organic fertilizer (SAFI) on maize (H513) production. Both fertilizers were applied at the rates of 0, 2.5, 5, and 7.5 t ha -1 , and 0, 30, 60, and 100 kg nitrogen (N) ha -1 . Mineral fertilizer (urea) was also applied at 0, 30, 60 and 100 kg N ha -1 to establish the N fertilizer equivalence (NFE) of the organic fertilizers. Maize grown in plots treated with BSFFF had the tallest plants and highest chlorophyll concentrations. Plots treated with 7.5 t ha -1 of BSFFF had 14% higher grain yields than plots treated with a similar rate of SAFI. There was a 27% and 7% increase in grain yields in plots treated with 100 kg N ha -1 of BSFFF compared to those treated with equivalent rates of SAFI and urea fertilizers, respectively. Application of BSFFF at 7.5 t ha -1 significantly increased N uptake by up to 23% compared to the equivalent rate of SAFI. Likewise, application of BSFFF at 100 kg N ha -1 increased maize N uptake by 76% and 29% compared to SAFI and urea, respectively. Maize treated with BSFFF at 2.5 t ha -1 and 30 kg N ha -1 had higher nitrogen recovery efficiencies compared to equivalent rates of SAFI. The agronomic N use efficiency (AE N ) of maize treated with 2.5 t ha -1 of BSFFF was 2.4 times higher than the value achieved using an equivalent rate of SAFI. Also, the AE N of maize grown using 30 kg N ha -1 was 27% and 116% higher than the values obtained using equivalent rates of SAFI and urea fertilizers, respectively. The NFE of BSFFF (108%) was 2.5 times higher than that of SAFI. Application rates of 2.5 t ha -1 and 30 kg N ha -1 of BSFFF were found to be effective in improving maize yield, while double rates of SAFI were required. Our findings demonstrate that BSFFF is a promising and sustainable alternative to commercial fertilizers for increased maize production., (Copyright © 2020 Beesigamukama, Mochoge, Korir, Fiaboe, Nakimbugwe, Khamis, Subramanian, Dubois, Musyoka, Ekesi, Kelemu and Tanga.)