Variation in encapsulation sensitivity of Cotesia sesamiae biotypes to Busseola fusca

Many braconid wasp species inject polydnaviruses to overcome their host's immune system. In the species Cotesia sesamiae, two biotypes exist that differ in their ability to develop in the host Busseola fusca. The biotype from coastal Kenya is infected with Wolbachia and is not able to develop in larvae of B. fusca, whereas the uninfected inland biotype of this wasp can develop in B. fusca. The genetic transmission of the developmental ability was studied through a series of genetic crosses and superparasitization experiments. The Wolbachia infection of the coastal type did not play a role in the encapsulation response of the host. Experiments show that the polydnaviruses of the wasps could not prevent the encapsulation of the coastal parasitoid eggs. Most likely, larval characteristics such as surface proteins played a more important role in the encapsulation response of the host even in the presence of a functional polydnavirus.     

Courtship behaviour of the African Maize Stem Borer: Busseola fusca (Fuller) (Lepidoptera: Noctuidae) under laboratory conditions

Busseola fusca (Fuller) (Lepidoptera: Noctuidae) is the most important African stem borer damaging maize and sorghum. Chemical mediators play an essential role in all life cycle of this moth, especially for mating recognition and host plant choice. The female sex pheromone, courtship and mating behaviours act on the reproductive isolation within insect populations. B. fusca courtship behaviour was studied to decipher each step that could account as a process for reproductive isolation. B. fusca males and females presented a very simple and fast courtship behaviour, without any particular events or male pheromone emission.     

Susceptibility of Eldana saccharina (Lepidoptera: Pyralidae), Busseola fusca and Sesamia calamistis (Lepidoptera: Noctuidae) to Bacillus thuringiensis Cry toxins and potential side effects on the larval parasitoid Cotesia sesamiae (Hymenoptera: Braconidae)

Laboratory trials were conducted to determine the efficacy of four Bacillus thuringiensis (Bt) Cry toxins at five different concentrations (0.016, 0.08, 0.4, 2, 10 μg/mL) for controlling three lepidopteran stem-borer species (i.e., the pyralid Eldana saccharina and the noctuids Busseola fusca and Sesamia calamistis) as well as to evaluate their indirect effect on the braconid larval parasitoid Cotesia sesamiae. In addition, larvae from the treatments above, after having been parasitized, were either fed a contaminated (group 1) or a toxin-free (group 2) diet and compared with a control (i.e., parasitized larvae which have never fed on Bt-toxin). All Bt Cry toxins induced larval feeding inhibition. Compared with the control, significant mortality resulted at all concentrations and for all species of Lepidoptera. Cry1Ab was the most toxic with 10 days post treatment mortalities ranging from 81% in B. fusca and S. calamistis to 100% in E. saccharina. In contrast, Cry1Ac had comparatively low toxicity particularly for B. fusca and S. calamistis (e.g., respectively, 54 and 74% mortality at 10 days at the highest concentration). In the toxin-treated group 1, percentages of C. sesamiae cocoon-producing moth larvae were higher compared to group 2 and the control, whereas clutch size was higher than in group 2 but similar to the control. In both groups, 0.016, 0.08 μg/mL CrylAc yielded lower female-biased sex ratios than the control. It was suggested that paralyses of the moth larvae caused by the toxin may have facilitated parasitization leading to higher parasitism and clutch size.     

Single-step PCR differentiation of Cotesia sesamiae (Cameron) and Cotesia flavipes Cameron (Hymenoptera: Braconidae) using polydnavirus markers

Cotesia sesamiae (Cameron) and Cotesia flavipes Cameron (Hymenoptera: Braconidae) are the main larval parasitoids of cereal stemborers in sub-Saharan Africa. Cotesia sesamiae is endemic to eastern and southern Africa, while C. flavipes was introduced into the region for biological control against the exotic lepidopteran Chilo partellus (Swinhoe) (Lepidoptera: Crambidae). The two are sibling parasitoids, difficult to distinguish morphologically. The introduced insect could potentially lead its African biotype to extinction because of their similar ecological niche. In order to distinguish the two species, multiplex primer-specific and PCR-RFLP tests were developed. Rapid identification of the two species was possible using primer-specific tests on DNA extracts as well as on pieces of tissue in a single PCR step followed by gel electrophoresis. The CRV1 gene of the polydnavirus, a symbiont to the wasps, was used as the marker. The results show that the morphological identifications, validated by molecular tests, are accurate in 93% of cases.     

Differential expression of the CrV1 haemocyte inactivation-associated polydnavirus gene in the African maize stem borer Busseola fusca (Fuller) parasitized by two biotypes of the endoparasitoid Cotesia sesamiae (Cameron)

Polydnaviruses are rarely studied for their natural variation in immune suppressive abilities. The polydnavirus harboring braconid Cotesia sesamiae, a widespread endoparasitoid of Busseola fusca and Sesamia calamistis in sub-Saharan Africa exists as two biotypes. In Kenya, the western biotype completes development in B. fusca larvae. However, eggs of the coastal C. sesamiae are encapsulated in this host and ultimately, no parasitoids emerge from parasitized B. fusca larvae. Both biotypes develop successfully in S. calamistis larvae. Encapsulation activity by B. fusca larvae towards eggs of the avirulent C. sesamiae was detectable six hours post-parasitization. The differences in encapsulation of virulent and avirulent strains were associated with differences in nucleotide sequences and expression of a CrV1 polydnavirus (PDV) gene, which is associated with haemocyte inactivation in the Cotesia rubecula/Pieris rapae system. CrV1 expression was faint or absent in fat body and haemolymph samples from B. fusca parasitized by the avirulent C. sesamiae, which exhibited encapsulation of eggs. Expression was high in fat body and haemolymph samples from both B. fusca and S. calamistis larvae parasitized by the virulent C. sesamiae, encapsulation in the former peaking at the same time points as CrV1 expression in the latter. Non synonymous difference in CrV1 gene sequences between virulent and avirulent wasp suggests that variations in B. fusca parasitism by C. sesamiae may be due to qualitative differences in CrV1-haemocyte interactions.     

Multiple Parasitism by Cotesia sesamiae and Cotesia flavipes (Hymenoptera: Braconidae) on Busseola fusca (Lepidoptera: Noctuidae)

Busseola fusca (Fuller) is one of the most important pest of cereals in sub-Saharan Africa. Cotesia sesamiae (Cameron) is the predominant parasitoid attacking B. fusca larvae in many parts of Africa. An exotic parasitoid, Cotesia flavipes Cameron, was introduced into Kenya in 1993 for the control of Chilo partellus (Swinhoe). Laboratory studies indicated that although C. flavipes would search for, and attack B. fusca , it was not able to complete its development in this host. The aim of the present study was to investigate the outcome of multiple parasitism of B. fusca by the two Cotesia species. The study showed that when both parasitoid species stung a B. fusca larva at the same time, both parasitoids emerged from more than half of the host larvae, C. flavipes alone emerged from 17%, and C. sesamiae alone emerged from 9%. When the larvae were parasitized by C. sesamiae first, and then 2 h later by C. flavipes , and vice versa, most of the progeny were C. flavipes . However, when B. fusca larvae were stung by C. sesamiae three days before oviposition by C. flavipes , significantly more C. sesamiae emerged from the larvae. When C. flavipes oviposited first, no larvae produced C. flavipes only. The interaction of parasitoids and the host immune system, and the implications of these results for the biological control of stem borers in East Africa are discussed.     

Characterization of eight microsatellite loci in the maize stalk borer Busseola fusca Fuller, 1901 (Lepidoptera: Noctuidae)

The noctuid stem borer Busseola fusca occurs throughout sub‐Saharan Africa, where it is considered as a major pest of corn and sorghum. To understand B. fusca phylogeography, we isolated and characterized eight microsatellite loci in this species. We investigated these loci for polymorphism on a subset sample of individuals from different geographic populations. All loci were polymorphic, showing between four and 10 alleles. Cross‐species amplification of four of these loci was tested on 11 other noctuid species.     

Phylogeographic pattern and regional evolutionary history of the maize stalk borer Busseola fusca (Fuller) (Lepidoptera: Noctuidae) in sub-Saharan Africa

Busseola fusca (Fuller) (Lepidoptera: Noctuidae) is one of the major cereal pests in sub-Saharan Africa. Previous phylogeographic investigations on samples collected in Kenya, Cameroon and West-Africa showed the presence of three main clades (W, KI, KII) originated from populations isolated in West and East Africa around one million years ago. Demographic and phylogenetic analyses suggested that this event was followed by local demographic expansion and isolation by distance. These hypotheses were tested by a more comprehensive sampling across B. fusca’s geographic range in Africa. Comparisons of sequences of partial mitochondrial DNA gene (cytochrome b) from 489 individuals of 98 localities in southern, central, eastern and western African countries confirmed the presence of the three main clades. Phylogenetic, F-statistics, demographic parameters and nested clade phylogeographic analyses confirmed that the clades experienced geographic and demographic expansion with isolation by distance after their isolation in three refuge areas. The geographic range of clade KII, already known from East to Central sub-Saharan Africa was extended to Southern Africa. Mismatch distribution analysis and the negative values of Tajima’s D index are consistent with a demographic expansion hypothesis for these three clades. Significant genetic differentiations were revealed at various hierarchical levels by analysis of molecular variance (AMOVA). Hypotheses about the geographic origin of the three main clades are detailed.     

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.     

Reproductive compatibility of several East and West African Cotesia sesamiae (Hymenoptera: Braconidae) populations and their crosses and backcrosses using Sesamia calamistis (Lepidoptera: Noctuidae) as the host

The relative importance of the braconid Cotesia sesamiae, a gregarious larval parasitoid of lepidopteran stemborers, varies greatly with region in Africa; while the most common parasitoid of noctuid stemborers in eastern Africa, it is rare in western Africa. Thus, several strains of C. sesamiae from Kenya are envisaged for introduction into western Africa. The present study investigates the reproductive compatibility between four populations of C. sesamiae from West Africa and Kenya with the noctuid Sesamia calamistis as the host using reciprocal crosses as well as backcrosses of hybrid females with males of the parental populations. Searching time of the male for the female and mating period varied significantly with couple and ranged between 0.78-1.9 min and 3.4-12.8 s, respectively. Crosses that involved females from inland Kenya (KI) did not yield any female offspring. However, backcrosses of hybrid female bearing a KI male genome with a KI male yielded both female and male offspring. Thus, there was a partial reproductive incompatibility between KI and West African populations which suggested that the latter were infected with Wolbachia sp. However, this should not affect the efficacy of a population introduced from East into West Africa, as there is a high degree of sib-mating in this gregarious parasitoid species. It was concluded that the regional differences in the relative importance of C. sesamiae was due to differences in the insect and plant host range of the different populations.     

Sexual dimorphism of antennal,tarsal and ovipositor chemosensilla in the African stemborer,Busseola fusca (Fuller) (Lepidoptera: Noctuidae)

The number and distribution of chemosensilla located on different organs of Busseola fusca (Fuller) (Lepidoptera: Noctuidae) males and females are described based on observations using scanning electron microscopy, selective staining with silver nitrate, and gustatory electrophysiological recording. The antennae and the fifth tarsomere of the prothoracic legs of both sexes bear chemosensilla: uniporous chaetica and multiporous trichoidea sensilla. However, there is a sexual dimorphism in the number and size of sensilla on these organs. The distal part of the ovipositor has uniporous gustatory chemosensilla of the chaetica type. The involvement of these sensilla in oviposition site selection by B. fusca is discussed.     

Release and establishment of Cotesia flavipes Cameron (Hymenoptera: Braconidae) an exotic parasitoid of Chilo partellus (Swinhoe) (Lepidoptera: Crambidae) in East and Southern Africa

Cotesia flavipes Cameron (Hymenoptera: Braconidae) was first imported into Kenya in 1991 from Pakistan for control of Chilo partellus (Swinhoe) (Lepidoptera: Crambidae). First releases were made at the Kenya coast in 1993 and a few recoveries of the parasitoid were made the following year. Additional foreign exploration for C. flavipes was conducted in central India in 1996 and 1998, which resulted in additional importation of the parasitoid for subsequent releases in eastern and southern Africa. Region-wide releases commenced with releases in Mozambique in 1996, Somalia in 1997 and Uganda in 1998. By 2005, many releases had been made in 10 countries in East and Southern Africa with establishment being reported in all of them except Eritrea but including Ethiopia where releases had never been made. This paper describes the progress made in the release and establishment of C. flavipes in East and Southern Africa and quantifies the rate of spread from the initial release sites in Kenya.     

Multiparasitism by the pupal parasitoids, Xanthopimpla stemmator (Hymenoptera: Ichneumonidae) and Pediobius furvus (Hymenoptera: Eulophidae) on two African cereal stemborers, Chilo partellus (Lepidoptera: Crambidae) and Busseola fusca (Lepidoptera: Noctuidae)

The outcome of multiparasitism by two pupal parasitoids, the exotic solitary ichneumonid Xanthopimpla stemmator and the indigenous gregarious eulophid Pediobius furvus, was studied using the invasive crambid stem borer Chilo partellus and the indigenous noctuid Busseola fusca as hosts. Two parasitism sequences were observed, where X. stemmator oviposited before P. furvus, and vice versa. In addition, the effect of time between first and second parasitism on parasitoid emergence, development, sex ratio, and number of offspring was assessed. For most treatments, time interval between parasitism had no significant effect on the percentage if pupae producing either P. furvus or X. stemmator. In general, X. stemmator outcompeted P. furvus irrespective of the order of oviposition, time interval between ovipositions and host species. Further studies, especially on host finding capacity of the two parasitoid species are required to determine the competitiveness of the two species under field conditions.     

Mitochondrial DNA phylogeography of the Cotesia flavipes complex of parasitic wasps (Hymenoptera: Braconidae)

The Cotesia flavipes species complex of parasitic wasps are economically important worldwide for the biological control of lepidopteran stem borers. The complex currently comprises three species: Cotesia flavipes Cameron, C. sesamiae (Cameron) and C. chilonis (Matsumura) (Hymenoptera: Braconidae), which appear morphologically similar. Despite their economic importance, little is known about the genetic diversity and phylogeography of these parasitoids. Differences in the biology of geographic populations have generally been interpreted as genetic divergence among strains, but direct genetic evidence is lacking. In Australia, several stem borer pests in neighbouring countries have been identified as significant threats to the sugar industry. However, the status of C. flavipes in Australia is unknown. To examine the genetic variation among worldwide populations of the C. flavipes complex and investigate the status of the Australian C. flavipes-like species, partial sequence data were generated for mitochondrial gene regions, 16S rRNA and COI. Parsimony, minimum evolution and Bayesian analyses based on 21 geographic populations of the complex and four outgroups supported the monophyly of the complex and the existence of genetically divergent populations of C. flavipes and C. sesamiae. The geographically isolated Australian haplotypes formed a distinct lineage within the complex and were ~3.0% divergent from the other species. The results indicated that historical biogeographic barriers and recent biological control introductions play an important role in structuring lineages within these species. This study provides a phylogeographical context for examining adaptive evolution and host range within biologically divergent strains of the C. flavipes complex.     

Biology of Cotesia ( = Apanteles) telengai (Hymenoptera: Braconidae) on its natural host Agrotis segetum (Lepidoptera: Noctuidae)

The host suitability of Agrotis segetum Denis & Schiff., A. ipsilon Hufn., Spodoptera littoralis Boisd, S. exigua Hub., Mythimna loreyi Duponchel and Mamestra oleracea L. for the gregarious braconid Cotesia ( = Apanteles) telengai Tobias was determined under laboratory conditions. The parasitoid only completed its development in larvae of A. segetum. The percentage of successfully parasitized larvae and the mean duration of C. telengai egg‐larval period were inversely related to the age of the host or host instar at parasitization. The mean number of parasitoids which emerged per parasitized larva was positively correlated with the larval age. The sex ratio was consistently high (ca. eight males to one female), independent of the host instar parasitized, as compared to 1:1 as observed frequently in field populations of this wasp. The females of C. telengai were active and produced offspring at temperatures of 15, 20, 25 and 30° C. However, the mean percentage of parasitized larvae increased from 13.1 to 72% and the mean progeny per parasitoid female increased from 14.7 to 129.4 parasitoids, both significant, when the experimental temperature was raised from 15 to 30°C, while their mean development time decreased from 75.5 to 19.2 days. At 25°C, the virgin and mated females continued oviposition until days 16 and 17, with a lifetime total of progeny of 397.6 (SD ±224.7) and 611.1 (SD± 128.8) parasitoids respectively, reaching a maximum of 64.3 and 99.2 on day 2 respectively.     

Host Suitability of Four Cereal Stem Borers (Lepidoptera: Crambidae, Noctuidae) for Different Geographic Populations of Cotesia sesamiae (Cameron) (Hymenoptera: Braconidae) in Kenya

This study focused on the suitability of four species of cereal stem borers for the development of five geographic populations of Cotesia sesamiae (Cameron). C. sesamiae, an indigenous larval parasitoid of gramineous stem borers, is widespread in Africa. Four stem borers, Chilo partellus (Swinhoe), Chilo orichalcociliellus Strand (Lepidoptera: Crambidae), Busseola fusca Fuller, and Sesamia calamistis Hampson (Lepidoptera: Noctuidae), were offered to C. sesamiae for oviposition. Parasitoid individuals originated from five locations in Kenya. Biological parameters such as developmental time, percentage parasitism, progeny production, mortality of immature parasitoids, and proportion of female progeny were compared across host species. The two populations from western Kenya developed well on B. fusca. However, populations from the coast and the Eastern Province could not successfully parasitize B. fusca. With the exception of B. fusca, the percentage of hosts successfully parasitized by the different C. sesamiae populations was not different. The size of the host appeared to be an important factor influencing the development and reproductive potential of the parasitoid. We conclude that the different parasitoid populations were adapted to location-specific characteristics. Parasitoid–host compatibility must be evaluated before release for better establishment and colonization.     

Responses of the stem borer larval endoparasitoid Cotesia flavipes (Hymenoptera: Braconidae) to plant derived synomones: Laboratory and field cage experiments

Laboratory and field cage experiments investigated the response of females of the stem borer larval endoparasitoid Cotesia flavipes to two synthetic synomone components, the terpenoid (E)-β-farnesene and the green leaf volatile, (Z)-3-hexenyl acetate, both compounds identified previously in headspace volatiles of maize plants damaged by stem borer (Chilo partellus). In dose response tests performed in a Y-tube olfactometer, parasitoids were significantly more attracted to the arms bearing 10 or 15 µg of (Z)-3-hexenyl acetate and (E)-β-farnesene than to the control arm. (E)-β-Farnesene was as attractive as the essential oil from the plant Hemizygia petiolata (Lamiaceae) rich in the same compound (80% relative amount). The plant essential oil elicited responses from females of the parasitoid comparable to those elicited by two positive controls, stem borer larval frass and adult parasitoid diet (20% honey solution), tested in the laboratory assays. In field cage trapping experiments, captures in traps baited with the terpenoid, the plant essential oil, (Z)-3-hexenyl acetate and the control of 20% honey solution, were not significantly different relative to captures in unbaited traps. Addition of the green leaf volatile (Z)-3-hexenyl acetate to the plant essential oil to yield a 1:1 two-component blend captured significantly more female parasitoids than traps baited with either of the two components alone. The results show that blends of green leaf volatiles and sesquiterpenoids may have potential in monitoring C. flavipes populations in the field.     

Host plants of the herbivorous insect Mythimna separata (Lepidoptera: Noctuidae) affect its susceptibility to parasitism by the larval parasitoid Cotesia kariyai (Hymenoptera: Braconidae)

Previous studies have shown that the food plant species consumed by herbivorous insects affect their parasitism by parasitoid wasps or flies, but this phenomenon has only been observed in a limited number of systems. Here, we investigate how feeding on different plant species affects the survival of the rice armyworm Mythimna separata (Walker) (Lepidoptera: Noctuidae) and its successful parasitism by the larval endoparasitoid Cotesia kariyai (Watanabe) (Hymenoptera: Braconidae). Survival rate and pupal weight of unparasitised M. separata fed with daikon Raphanus sativus L. var. longipinnatus was lower than with maize Zea mays L. In addition, caterpillars fed with daikon were less susceptible to parasitism by C. kariyai than those fed with maize, and parasitised M. separata larvae fed with daikon had higher survival rates than those fed with maize. These results indicate that while daikon is not an optimal food for unparasitised caterpillars, it may protect the larvae from death by C. kariyai parasitism.     

Dispersal behavior of the parasitic wasp Cotesia urabae (Hymenoptera: Braconidae): A recently introduced biocontrol agent for the control of Uraba lugens (Lepidoptera: Nolidae) in New Zealand

Estimates of the dispersal range of a recently introduced biocontrol agent in its new environment are vital to understanding its relative searching capacity, and to foresee the maximum area that could be covered in a release event. In New Zealand, the solitary endoparasitoid Cotesia urabae Austin and Allen (Hymenoptera: Braconidae) was first released in January 2011 as a biological control agent for the gum leaf skeletoniser, Uraba lugens Walker (Lepidoptera: Nolidae). The objective of this study was to utilize an experimental approach to quantify the dispersal behavior of one generation of C. urabae. In our experiment, which used sentinel larvae as target hosts, parasitoids dispersed up to 20 m away from the release point but parasitism was highest within 5 m of the release site. A high level of parasitism was observed at the release tree (87.6%) which suggests that most of the females released may have stayed there. According to the dispersal model developed from the data collected, Cotesia would be able to disperse up to 53 m in one release event. In addition, significant differences were found between the different directions tested for dispersal, showing a clear downwind effect on dispersal suggesting that wind has a direct effect on the dispersal behavior of C. urabae in the field.     

Suitability ofBusseola fuscaandSesamia calamistis(Lepidoptera: Noctuidae) for the Development of Two Populations ofCotesia sesamiae(Hymenoptera: Braconidae) in Kenya

The suitability ofSesamia calamistisHampson andBusseola fusca(Fuller) for the development of two geographical populations ofCotesia sesamiae(Cameron) was examined in the laboratory. One population of the parasitoid was collected from the coast of Kenya and the other from the inland. Both populations of the parasitoid could develop onS. calamistis.OnB. fusca,the inland population ofC. sesamiaewas able to develop, while the population from the coastal area of Kenya was encapsulated. Mating studies revealed that the two parasitoid populations were partially reproductively isolated. Unidirectional incompatibility, possibly caused by theWolbachiainfection, was observed when males from the infected coastal population were mated with females from the uninfected inland population.