B. Le Ru

Phylogeography and population genetics of the maize stalk borer Busseola fusca (Lepidoptera, Noctuidae) in sub‐Saharan Africa

The population genetics and phylogeography of African phytophagous insects have received little attention. Some, such as the maize stalk borer Busseola fusca, display significant geographic differences in ecological preferences that may be congruent with patterns of molecular variation. To test this, we collected 307 individuals of this species from maize and cultivated sorghum at 52 localities in West, Central and East Africa during the growing season. For all collected individuals, we sequenced a fragment of the mitochondrial cytochrome b. We tested hypotheses concerning the history and demographic structure of this species. Phylogenetic analyses and nested clade phylogeographic analyses (NCPA) separated the populations into three mitochondrial clades, one from West Africa, and two — Kenya I and Kenya II — from East and Central Africa. The similar nucleotide divergence between clades and nucleotide diversity within clades suggest that they became isolated at about the same time in three different refuges in sub‐Saharan Africa and have similar demographic histories. The results of mismatch distribution analyses were consistent with the demographic expansion of these clades. Analysis of molecular variance (amova) indicated a high level of geographic differentiation at different hierarchical levels. NCPA suggested that the observed distribution of haplotypes at several hierarchical levels within the three major clades is best accounted for by restricted gene flow with isolation by distance. The domestication of sorghum and the introduction of maize in Africa had no visible effect on the geographic patterns observed in the B. fusca mitochondrial genome.

Electrically recorded feeding behaviour of cassava mealybug on host and non‐host plants

The feeding behaviour of cassava mealybug (Phenacoccus manihoti) was analysed by the electrical penetration graph technique (EPG, DC‐system) and by light microscopy. This study confirmed a typical phloem‐feeding behaviour with an aphid‐like predominance of extracellular pathways of stylets. Similarities of EPGs from mealybugs with those of aphids and whiteflies allowed adoption of standard pattern labelling. The main differences with aphid EPGs consisted of fewer but longer intracellular punctures (pd duration of 20 s vs 5–7 s for aphids), longer times to the first phloem‐ingestion period and a lower motility of stylets within the phloem searching process. Comparison of the feeding behaviour on 6 plant types, including two occasional hosts (Talinum and poinsettia), one cassava hybrid and three common cassava cultivars (Manihot esculenta), allowed differentiation of penetration profiles. Intracellular punctures were shorter on occasional hosts, on which phloem finding and subsequent ingestion were markedly delayed although a great variability within plants persisted. Chemical analysis of some secondary plant substances revealed that none of the plants tested contained detectable amounts of alkaloids, and that cyanides were restricted to true hosts (cassavas and Manihot hybrid). Levels of total flavonoids did not differ between hosts and non‐hosts, in contrast to those of phenolic acids. Preliminary correlations with EPG patterns are discussed.

Importance of plant physical cues in host acceptance for oviposition by Busseola fusca

Plant and surrogate stems exhibiting specific combinations of physical cues were used to determine which plant‐related stimuli influence the oviposition of Busseola fusca (Fuller) (Lepidoptera: Noctuidae). The number of eggs and egg batches laid per female increased with an increase in diameter of both natural and artificial stems. Direct observations of the oviposition behaviour (walking, antennating, and sweeping with the ovipositor) indicated that the female moths preferred oviposition supports with a large diameter and non‐pubescent or smooth surfaces over pubescent or rough ones. Pubescence and rough surfaces significantly affected the behavioural steps leading to oviposition by interfering with the ovipositor sweep process necessary to find a suitable oviposition site. Furthermore, more eggs and egg batches were laid on soft than rigid supports. The rigidity of the support affected the proper insertion of the ovipositor for egg deposition. Our results underline the importance of physical stimuli in B. fuscas choice of an oviposition site, which may facilitate the identification of potential host plants or preferred oviposition sites on a plant for this species.

Seasonal changes in secondary compounds in the phloem sap of cassava in relation to plant genotype and infestation by Phenacoccus manihoti (Homoptera : Pseudococcidae)

A previous study on the population dynamics of the cassava mealybug, Phenacaccus manihofi (Matile-Ferrero), on cassava has shown that populations increase during the dry season. The aim of the present studies was to determine whether these important changes in the pest population observed throughout the . seasons in the field in the Congo, could be correlated with seasonal variations in the level of secondary compounds in the phloem, implicated in cassava plant resistance to P. manihofi. Our results showed that cassava was in drought stress conditions during the dry season (foliar area decreased). The combination of infestation and dry stress factors was clearly visible during the d r ~ season. In fact, the decrease in leaf water potential together with infestation by P. manihofi was clearly observed at the end of the rainy season and during the dry season. The levels of the secondary compounds, rutin (an unfavourable substance to P. manihofi) decreased in the less resistant genotype, and cyanide (a phagostimulant substance) increased in each genotype in the dry season. These results may partially explain the important population changes in P. manihofi during the seasons.

Differences in mate acceptance and host plant recognition between wild and laboratory‐reared Busseola fusca (Fuller)

The present study was aimed at characterizing differences in mate acceptance and host plant recognition between Busseola fusca (Lep.: Noctuidae) reared for several generations under laboratory conditions and wild conspecifics, directly collected from maize stems in the field. The mating success was significantly higher in laboratory reared when compared with the wild B. fusca population. Oviposition on artificial stems was significantly higher for laboratory‐reared insects than for the wild ones. Moreover, unlike adults of the wild strain, laboratory‐reared B. fusca showed no preference to oviposit on surrogate stems impregnated with maize extracts. Long‐range attraction to the host plant was significantly lower for laboratory‐reared insects. Furthermore, the polymerase chain reaction‐restriction fragment length polymorphism (PCR‐RFLP) analysis of mitochondrial DNA showed that the wild type and laboratory populations belonged to the same genetic strain, indicating that these differences between the populations were mainly phenotypic.

Phylogeography in continuous space: coupling species distribution models and circuit theory to assess the effect of contiguous migration at different climatic periods on genetic differentiation in Busseola fusca (Lepidoptera: Noctuidae)

Current population genetic models fail to cope with genetic differentiation for species with large, contiguous and heterogeneous distribution. We show that in such a case, genetic differentiation can be predicted at equilibrium by circuit theory, where conductance corresponds to abundance in species distribution models (SDMs). Circuit‐SDM approach was used for the phylogeographic study of the lepidopteran cereal stemborer Busseola fuscaFüller (Noctuidae) across sub‐Saharan Africa. Species abundance was surveyed across its distribution range. SDMs were optimized and selected by cross‐validation. Relationship between observed matrices of genetic differentiation between individuals, and between matrices of resistance distance was assessed through Mantel tests and redundancy discriminant analyses (RDAs). A total of 628 individuals from 130 localities in 17 countries were genotyped at seven microsatellite loci. Six population clusters were found based on a Bayesian analysis. The eastern margin of Dahomey gap between East and West Africa was the main factor of genetic differentiation. The SDM projections at present, last interglacial and last glacial maximum periods were used for the estimation of circuit resistance between locations of genotyped individuals. For all periods of time, when using either all individuals or only East African individuals, partial Mantel r and RDA conditioning on geographic distance were found significant. Under future projections (year 2080), partial r and RDA significance were different. From this study, it is concluded that analytical solutions provided by circuit theory are useful for the evolutionary management of populations and for phylogeographic analysis when coalescence times are not accessible by approximate Bayesian simulations.

Variability in Pheromone Communication Among Different Haplotype Populations of Busseola fusca

The relationship between pheromone composition and mitochondrial haplotype clades was investigated by coupling DNA analyses with pheromone identification and male mate searching behavior among different geographic populations of Busseola fusca. The within-population variations in pheromone blend were as great as those observed between geographic populations, suggesting that the female sex pheromone blend was not the basis of reproductive isolation between the geographic clades. Furthermore, while data from wind tunnel experiments demonstrated that most of the tested males were sensitive to small variations in pheromone mixture, there was considerable within-population variability in the observed response. The study identified a new pheromone component, (Z)-11-hexadecen-1-yl acetate, which when added to the currently used three-component synthetic blend resulted in significantly higher traps catches. The new recommended blend for monitoring flight phenology and for timing control measures for optimal efficacy of B. fusca is (Z)-11-tetradecen-1-yl acetate (62%), (E)-11-tetradecen-1-yl acetate (15%), (Z)-9-tetradecen-1-yl acetate (13%), and (Z)-11-hexadecen-1-yl acetate (10%).

Importance of the experimental setup in research on attractiveness of odours in moths: an example with Busseola fusca

Wind tunnel and Y‐tube olfactometer studies are useful for demonstrating the responsiveness of male moths to sex pheromones. However, in the cereal stem borer, Busseola fusca (Fuller) (Lepidoptera: Noctuidae), wind tunnel bioassays yielded poor results when the behavioural responses of females to plant odours were tested. We demonstrated that for B. fusca females, the Y‐tube olfactometer was better suited for measuring the attractiveness of plant odours, compared with the wind tunnel. In particular, we showed B. fusca preference for odours of a host over a non‐host plant species using this apparatus. Behavioural responses for B. fusca after host finding are also discussed.

Influence of plant silicon in Busseola fusca (Lepidoptera: Noctuidae) larvae - Poaceae interactions.

The noctuid stem borer Busseola fusca is an important pest of maize and sorghum in Sub-Saharan Africa. The presence of this species occurred mostly on cultivated than on wild habitats. Busseola fusca is oligophagous having a narrow range of a wild grass species. This might be due, in part, to differences in silicon (Si) content in plant tissues between cultivated and wild grasses. In the present study, we have tested this hypothesis by studying the survival and the relative growth rate (RGR) expressed as daily weight gains of B. fusca larvae on maize and six wild host plants, mostly present in the natural habitat where B. fusca occurred, and correlated with their Si contents. Survival and RGR of B. fusca larvae were considerably higher on maize and wild sorghum than on the other grass species, and they were negatively related to plant Si content. This was corroborated with results on RGR from artificial diets amended with increasing levels of Si. In addition, if Si was added to maize growing substrate B. fusca larval growth was significantly reduced confirming the involvement of Si in B. fusca larvae - Poaceae interactions. The results provide insight into the possible mechanisms of oligophagy of B. fusca and provide a correlative support for a physical role of plant endogenous Si in impeding feeding of B. fusca larvae.

Sex pheromone composition and reproductive isolation in two Busseola species (Lepidoptera: Noctuidae) in Kenya

Closely related species provide an interesting and useful model to understand mate communication diversification. Pre-mating isolation is the result of selection processes and is achieved by utilization of different sex pheromones, but also by different ecological and life traits such as geographic distribution, host-plant specialization and allochrony of sexual behavior. Here, we report sex pheromone identifications and pre-mating isolation of two closely related species of stemborers: Busseola segeta and the yet undescribed Busseola nov. sp., developing on different host-plants and in different geographical locations in Kenya. GC–MS analyses showed that the two species produced the same pheromone components, but in slight different ratios. The respective synthetic pheromone mixtures tested in the field were attractive for males of each species. The low inter-specific variation in the pheromone mixture would allow possible interbreeding in case of accidental introduction of both species in the same environment. Under laboratory conditions, allochrony of the mating period is total and might prevent interbreeding if confirmed in natural conditions. This point would ensure the reproductive isolation between the two species in case of modifications of ecological factors. This study and the chemical structures of the identified pheromone components clearly demonstrate that these Busseola species are closely related although they differ in morphology.