‘Becoming a species by becoming a pest’ or how two maize pests of the genus Ostrinia possibly evolved through parallel ecological speciation events

New agricultural pest species attacking introduced crops may evolve from pre‐existing local herbivores by ecological speciation, thereby becoming a species by becoming a pest. We compare the evolutionary pathways by which two maize pests (the Asian and the European corn borers, ACB and ECB) in the genus Ostrinia (Lepidoptera, Crambidae) probably diverged from an ancestral species close to the current Adzuki bean borer (ABB). We typed larval Ostrinia populations collected on maize and dicotyledons across China and eastern Siberia, at microsatellite and mitochondrial loci. We found only two clusters: one on maize (as expected) and a single one on dicotyledons despite differences in male mid‐tibia morphology, suggesting that all individuals from dicotyledons belonged to the ABB. We found evidence for migrants and hybrids on both host plant types. Hybrids suggest that field reproductive isolation is incomplete between ACB and ABB. Interestingly, a few individuals with an ‘ABB‐like’ microsatellite profile collected on dicotyledons had ‘ACB’ mtDNA rather than ‘ABB‐like’ mtDNA, whereas the reverse was never found on maize. This suggests asymmetrical gene flow directed from the ACB towards the ABB. Hybrids and backcrosses in all directions were obtained in no‐choice tests. In laboratory conditions, they survived as well as parental strain individuals. In Xinjiang, we found ACB and ECB in sympatry, but no hybrids. Altogether, our results suggest that reproductive isolation between ACB and ABB is incomplete and mostly prezygotic. This points to ecological speciation as a possible evolutionary scenario, as previously found for ECB and ABB in Europe.     

Scanning the European corn borer (Ostrinia spp.) genome for adaptive divergence between host-affiliated sibling species

It has recently been shown that the European corn borer, a major pest of maize crops, is actually composed of two genetically differentiated and reproductively isolated taxa, which are found in sympatry over a wide geographical range in Eurasia. Each taxon is adapted to specific host plants: Ostrinia nubilalis feeds mainly on maize, while O. scapulalis feeds mainly on hop or mugwort. Here, we present a genome scan approach as a first step towards an integrated molecular analysis of the adaptive genomic divergence between O. nubilalis and O. scapulalis. We analysed 609 AFLP marker loci in replicate samples of sympatric populations of Ostrinia spp. collected on maize, hop and mugwort, in France. Using two genome scan methods based on the analysis of population differentiation, we found a set of 28 outlier loci that departed from the neutral expectation in one or the other method (of which a subset of 14 loci were common to both methods), which showed a significantly increased differentiation between O. nubilalis and O. scapulalis, when compared to the rest of the genome. A subset of 12 outlier loci were sequenced, of which 7 were successfully re‐amplified as target candidate loci. Three of these showed homology with annotated lepidopteran sequences from public nucleotide databases.     

Homology of olfactory receptor neuron response characteristics inferred from hybrids between Asian and European corn borer moths (Lepidoptera: Crambidae)

First generation hybrid males from crosses between the Asian corn borer (ACB), Ostrinia furnacalis, and the “univoltine Z-strain” European corn borer (ECB), Ostrinia nubilalis, were examined with respect to behavioral and physiological responses to ACB and ECB pheromones. The hybrid males often flew to the pheromone of ECB Z-strain, but very rarely to the ACB pheromone. We mapped the tuning profiles of each ORN of the F₁ hybrids with respect to the relevant pheromone components and a common behavioral antagonist by employing differential cross-adaptation and varying doses of the ligands. In the trichoid sensilla of F₁ hybrid males, the three co-compartmentalized ORNs produced spikes that were very difficult to distinguish by size, unlike the parental populations. Comparing the responses to ACB and ECB components at different doses reveals overlapping profiles similar to males of both parental types, but more responsiveness to the ECB pheromone components. We were unable to detect any differences in the ORN tuning profiles when comparing males with different behavioral phenotypes. While the two ECB pheromone races have similar ORN tuning properties that are different from those in ACB, the spike-amplitude patterns of ECB E-strain and ACB have greater homology when compared to ECB Z-strain.     

Divergence in behaviour between the European corn borer,Ostrinia nubilalis,and its sibling species Ostrinia scapulalis: adaptation to human harvesting?

Divergent adaptation to host plant species may be the major mechanism driving speciation and adaptive radiations in phytophagous insects. Host plants can differ intrinsically in a number of attributes, but the role of natural enemies in host plant specialization is often underappreciated. Here, we report behavioural divergence between the European corn borer (ECB, Ostrinia nubilalis) and its sibling species Ostrinia scapulalis, in relation to a major enemy: humans. Harvesting maize imposes selective mortality on Ostrinia larvae: those located above the cut-off line of the stalk face almost certain death. We show that ECB larvae diapause closer to the ground than those of O. scapulalis, which is sympatric but feeds mainly on weeds. The difference in diapause height results from genetically determined differences in geotactic behaviour. ECB larvae descend towards the ground specifically at harvest time, increasing their chances of surviving harvesting by about 50 per cent over O. scapulalis larvae. Natural enemies appear as a major driver of host-plant specialization in this example, stressing the need to consider ‘tri-trophic’ ecological niches to understand insect diversification. Our results also strongly suggest that geotaxis evolved as a singular instance of behavioural resistance in a major agricultural pest.     

Olfactory neuron responsiveness and pheromone blend preference in hybrids between Ostrinia furnacalis and Ostrinia nubilalis (Lepidoptera: Crambidae)

The olfactory receptor neuron (ORN) and behavioral responses of hybrids between the Asian corn borer (ACB), Ostrinia furnacalis, and the E-strain European corn borer (ECB(E)), Ostrinia nubilalis were examined and compared to the parental populations. In hybrids and both parents, the large-spike-size ORN was capable of responding to all four pheromone components of ACB and ECB, despite differences in which compounds elicited the greatest spike frequency in each population. There was a small-spiking ORN more narrowly tuned to the minor pheromone components in both ACB and ECB(E). In hybrids the homologous small-spiking ORN was tuned primarily to the ECB(E) minor pheromone component, with some responsiveness to the ACB minor component. Both species and all the hybrids had an intermediate spike-size ORN tuned primarily to their common behavioral antagonist. Dominance of responsiveness to the ECB(E) versus the ACB minor pheromone component on the small-spiking ORN may explain the greater tendency of hybrids to fly upwind to the ECB(E) pheromone blend than the ACB blend. This finding points toward a distinct evolutionary role for this ORN in allowing a pheromone shift.     

Introgression between divergent corn borer species in a region of sympatry: Implications on the evolution and adaptation of pest arthropods

The Asian corn borer, Ostrinia furnacalis, and European corn borer, O. nubilalis (Lepidoptera: Crambidae), cause damage to cultivated maize in spatially distinct geographies and have evolved divergent hydrocarbons as the basis of sexual communication. The Yili area of Xinjiang Uyghur Autonomous Region in China represents the only known region where O. furnacalis has invaded a native O. nubilalis range, and these two corn borer species have made secondary contact. Genetic differentiation was estimated between Ostrinia larvae collected from maize plants at 11 locations in Xinjiang and genotyped using high‐throughput SNP and microsatellite markers. Maternal lineages were assessed by direct sequencing of mitochondrial cytochrome c oxidase subunit I and II haplotypes, and a high degree of genotypic diversity was demonstrated between lineages based on SNP genotypes. Furthermore, historical introgression was predicted among SNP genotypes only at sympatric locations in the Yili area, whereas in Xinjiang populations only O. furnacalis haplotypes were detected and no analogous introgressed genotypes were predicted. Our detection of putative hybrids and historical evidence of introgression defines Yili area as a hybrid zone between the species in normal ecological interactions and furthermore, might indicate that adaptive traits could spread even between seemingly divergent species through horizontal transmission. Results of this study indicate there may be a continuum in the degree of reproductive isolation between Ostrinia species and that the elegance of distinct and complete speciation based on modifications to the pheromone communication might need to be reconsidered.     

亚洲玉米螟与欧洲玉米螟混生区的研究

生殖隔离试验、形态鉴定、网室内性信息素活性反应和食性试验的结果表明 :张家口苍耳中的玉米螟与新疆伊宁的欧洲玉米螟同种 ,而为害玉米的优势种与广东阳山的亚洲玉米螟同种。张家口为害玉米的种群中 ,有部分雄蛾对欧洲玉米螟性信息素及苍耳中的雌处女蛾具有反应。苍耳和玉米中的种群在玉米、谷子、高梁、苍耳、草和大马蓼等 6种寄主上均可产卵和取食为害 ,并能正常完成生活史。研究结果证实张家口为亚洲玉米螟和欧洲玉米螟混生区 ,而且在玉米中很可能有少量欧洲玉米螟与亚洲玉米螟混生。     

Resistance monitoring of field populations of the corn borers Sesamia nonagrioides and Ostrinia nubilalis after 5 years of Bt maize cultivation in Spain

Approximately 22 000 hectares (5% of the total maize growing area) of transgenic maize expressing the Cry1Ab toxin from Bacillus thuringiensis (Bt maize) have been planted annually in Spain since 1998. Changes in the susceptibility to Cry1Ab of Spanish populations of the Mediterranean corn borer (MCB), Sesamia nonagrioides (Lefebvre) (Lepidoptera: Noctuidae), and the European corn borer (ECB), Ostrinia nubilalis (Hübner) (Lepidoptera: Crambidae), were assessed by annual monitoring on Bt maize fields. No increase in resistance was detected in the MCB populations from Ebro, Albacete, and Badajoz, nor in the ECB populations from Ebro and Badajoz during the period 1999–2002. The susceptibility of the MCB population from Madrid fluctuated from year to year, but a gradual trend towards higher levels of tolerance was not observed. Laboratory selection assays for eight generations yielded selected strains of MCB and ECB that were 21‐ and 10‐fold significantly more tolerant to Cry1Ab than the corresponding unselected strains, respectively. Nevertheless, none of the field‐collected or laboratory‐selected larvae were able to survive on Bt maize. Considering these data, no consistent shifts in susceptibility were found for Spanish populations of MCB nor ECB after 5 years of Bt maize cultivation, but systematic field monitoring needs to be continued.     

Host specialization involving attraction,avoidance and performance,in two phytophagous moth species

Host specialization plays a key role in the extreme diversification of phytophagous insects. Whereas proximate mechanisms of specialization have been studied extensively, their consequences for species divergence remain unclear. Preference for, and performance on hosts are thought to be a major source of divergence in phytophagous insects. We assessed these major components of specialization in two moth species, the European corn borer (ECB) and the Adzuki bean borer (ABB), by testing their oviposition behaviour in different conditions (choice or no‐choice set‐ups) and their performances, by reciprocal transplant at the larval stage on the usual host and an alternative host plant. We demonstrated that both ABB and ECB have a strong preference for their host plants for oviposition, but that relative larval performances on the usual host and an alternative host differed according to the experiment and the trait considered (weight or survival). Finally, we show for the first time that the preference for maize in ECB conceals a strong avoidance of mugwort. The differences in performance, attraction and avoidance between ECB and ABB are discussed in the light of the underlying mechanisms and divergence process.     

Evidence of olfactory antagonistic imposition as a facilitator of evolutionary shifts in pheromone blend usage in Ostrinia spp. (Lepidoptera: Crambidae)

Olfactory receptor neuron (ORN) response was measured to assess why some males ("rare males") of the Asian corn borer (ACB), Ostrinia furnacalis, have a broad behavioral response to fly upwind to both the ACB and the European corn borer (ECB), Ostrinia nubilalis, pheromone blends. We performed single-sensillum electrophysiological recordings on ACB males that had been behaviorally assessed for upwind flight response to the ACB blend [60:40 (Z)-12-tetradecenyl acetate (Z12-14:OAc) to (E)-12-tetradecenyl acetate (E12-14:OAc)], as well as to ECB (Z-strain) and ECB (E-strain) blends [3:97 and 99:1 (Z)-11-tetradecenyl acetate (Z11-14:OAc) to (E)-11-tetradecenyl acetate (E11-14:OAc)]. Sensilla from all types of males had large- and small-spike-sized ORNs responding strongly to Z12- or E12-14:OAc, but weakly to Z11- and E11-14:OAc. In the majority of males ("normal males") that flew upwind only to the ACB blend, Z11-14:OAc elicited responses in an intermediate spike-sized ORN associated with behavioral antagonism that is mainly tuned to (Z)-9-tetradecenyl acetate (Z9-14:OAc). In the rare-type ACB males that flew to both the ACB and ECB pheromone blends, Z11-14:OAc did not stimulate this ORN. Increased responsiveness to ancestral pheromone components by ORNs associated with behavioral antagonism could be instrumental in reproductive character displacement, or in reinforcement and reproductive isolation during speciation by helping to increase assortative mating between males and females in derived populations that use novel sex pheromone blends.     

Genetic mapping of two components of reproductive isolation between two sibling species of moths,Ostrinia nubilalis and O. scapulalis

We report the quantitative trait loci (QTL) mapping of reproductive isolation traits between Ostrinia nubilalis (the European corn borer) and its sibling species O. scapulalis (the Adzuki bean borer), focusing on two traits: mating isolation (mi) and pheromone production (Pher). Four genetic maps were generated from two backcross families, with two maps (one chromosomal map and one linkage map) per backcross. We located 165–323 AFLP markers on these four maps, resulting in the identification of 27–31 linkage groups, depending on the map considered. No-choice mating experiments with the offspring of each backcross led to the detection of at least two QTLs for mi in different linkage groups. QTLs underlying Pher were located in a third linkage group. The Z heterochromosome was identified by a specific marker (Tpi) and did not carry any of these QTLs. Finally, we considered the global divergence between the two sibling species, distortions of segregation throughout the genome, and the location and effect of mi and Pher QTLs in light of the known candidate genes for reproductive isolation within the genus Ostrinia and, more broadly, in phytophagous insects.     

Physiological adaptation of the Asian corn borer Ostrinia furnacalis to chemical defenses of its host plant, maize

A number of gramineous plants such as maize contain cyclic hydroxamic acids (cHx) that are toxic to many herbivores. Among the Ostrinia species found in Japan, the Asian corn borer Ostrinia furnacalis is the only one that utilizes maize, a gramineous plant. We used O. furnacalis and two congeners, Ostrinia scapulalis and Ostrinia latipennis, to obtain insights into the physiological adaptation of O. furnacalis to cHx. When an artificial diet containing a low concentration (0.3 mg/g diet) of cHx was fed to the larvae of O. furnacalis and O. scapulalis, larval growth and survival were significantly less affected in O. furnacalis than O. scapulalis. An artificial diet containing a high level (0.7 mg/g diet) of cHx was found to severely retard the growth of both species, albeit to different degrees. In an assay in vitro, homogenate of the digestive tract of O. furnacalis larvae degraded cHx more rapidly than that of O. scapulalis or O. latipennis. The degradation was found to be enzymatic and dependent on a cofactor, UDP-glucose, suggesting that UDP-glucosyltransferase or other UDP-glucose-dependent enzymes were involved. This enzymatic adaptation probably has enabled O. furnacalis to utilize plants containing cHx.     

二种交配型球孢白僵菌对亚洲玉米螟的生态控制作用

利用生物种间互做关系抑制农业害虫的暴发是生物防治的重要手段。为探讨二种交配型内共生球孢白僵菌与玉米之间的互惠关系及其形成的共生体在亚洲玉米螟控制中的生态效应,以玉米为宿主植物,以球孢白僵菌孢子悬浮液进行灌根,在温室内构建了二种交配型(MAT1-1-1型,B5;MAT1-2-1型,B2)球孢白僵菌-玉米共生体,并研究了共生体对玉米的生长、对亚洲玉米螟的产卵选择和幼虫发育及其对球孢白僵菌生物学特性的影响。结果显示:通过叶片离体培养、ITS基因和交配型基因MAT检测,均能检测到白僵菌的内生定殖;MAT1-2-1型B2菌株定殖检出率高,MAT1-1-1型B5菌株在混合型接种中定殖有优势。回收后的球孢白僵菌菌落直径和毒力无显著性变化,但其产孢量都显著提高其中回收B5处理组来源菌株的产孢量提高最显著。接种过球孢白僵菌的玉米植株地上部生长速度、生物量和地下根系生物量均优于对照组,其中根系干重明显增加,而地上植株干重也相对增加。MAT1-1-1型菌株B5对共生体玉米植株地上高度促生长贡献明显;MAT1-2-1型菌株B2对共生体玉米植株地下干重增加贡献明显。总体上球孢白僵菌内生定殖对玉米地下根系生物量影响大于对地上植株生物量的影响。在产卵选择性试验中,各处理组亚洲玉米螟的产卵量显著少于对照组。共生体对亚洲玉米螟产卵具有明显的趋避作用,MAT1-2-1型菌株B2对产卵的趋避作用明显,而MAT1-2-1型菌株B5的趋避作用较弱。在人工接种幼虫的试验中,处理组回收的亚洲玉米螟幼虫存活率均显著低于对照组,其中,B5组回收幼虫的存活率最低,仅为38.33%;处理组的化蛹率与对照组差异不显著,但B5组的回收幼虫化蛹率显著低于B2组和对照组,仅为34.77%,这说明MAT1-1-1型B5菌株对玉米螟幼虫发育抑制最明显。上述结果表明,不同交配型球孢白僵菌内生定殖效率有差异,在经过内生定殖后在产孢量方面有显著性提高,两个交配型菌株在联合应用时具有协同增效作用;两个交配型菌株均能够通过内生定殖与玉米形成共生体并促进玉米植株的生长,这显示球孢白僵菌和玉米之间已经建立具有互惠关系的共生体。这种共生体通过趋避亚洲玉米螟产卵、抑制幼虫存活和降低化蛹率等方面的潜力虽然不一样,但都有助于对亚洲玉米螟的可持续生态防治,也证明了共生体的建成有效提高了玉米的生态适应性,为利用球孢白僵菌内共生性实施亚洲玉米螟防控提供了新思路。     

A recombination suppressor contributes to ecological speciation in OSTRINIA moths

Despite unparalleled access to species'' genomes in our post-genomic age, we often lack adequate biological explanations for a major hallmark of the speciation process—genetic divergence. In the presence of gene flow, chromosomal rearrangements such as inversions are thought to promote divergence and facilitate speciation by suppressing recombination. Using a combination of genetic crosses, phenotyping of a trait underlying ecological isolation, and population genetic analysis of wild populations, we set out to determine whether evidence supports a role for recombination suppressors during speciation between the Z and E strains of European corn borer moth (Ostrinia nubilalis). Our results are consistent with the presence of an inversion that has contributed to accumulation of ecologically adaptive alleles and genetic differentiation across roughly 20% of the Ostrinia sex chromosome (~4 Mb). Patterns in Ostrinia suggest that chromosomal divergence may involve two separate phases—one driving its transient origin through local adaptation and one determining its stable persistence through differential introgression. As the evolutionary rate of rearrangements in lepidopteran genomes appears to be one of the fastest among eukaryotes, structural mutations may have had a disproportionate role during adaptive divergence and speciation in Ostrinia and in other moths and butterflies.     

Allozyme differentiation among nine populations of the corn borer (Ostrinia) in China

To test the hypothesis of the migration of the corn borer, the allozymes of nine populations of the corn borer(Ostrinia) in China were checked using verticalslab polyacrylamide gel electrophoresis. Eight loci of six allozymes were analyzed. The mean of the genetic identities among the nine populations calculated from the allele frequencies was 0.99068, much closer than that of other species and geographical populations. The mean (0.97955) of the genetic identities between the XJYN population (Ostrinia nubilalis Hübner, collected from Yining, Xinjiang Autonomous Region) and each of the other eight populations (O. furnacalis Guenée) was significantly smaller than that between the pairs of the eight populations (0.99386;t test,P<0.01). Although the population XJYN clearly deviates from the other eight populations in the dendrogram, the relationship of the two species of corn borer was very close. It is possible that the speciation of corn borer may have resulted from single-gene substitutions.     

European corn borer injury effects on lignin, carbon and nitrogen in corn tissues

Plant herbivores often stimulate lignin deposition in injured plant tissue, but it is not known whether corn (Zea mays L.) reacts to European corn borer (ECB, Ostrinia nubilalis Hubner) injury in this manner. Bt (Bacillus thuringiensis) genetic modification is also reported to affect lignin in corn. This study evaluated the effects of ECB injury and the Bt gene on the chemical composition and decomposition of corn tissues. Eight near isolines (Bt and NBt) were grown in pots and half were infested with ECB. The experiment was repeated in 2 years. ECB injury increased the lignin concentration in corn leaves in one of 2 years and lowered the C:N ratio in injured stems. Lignin concentration in leaves was greater in Bt than NBt corn in 1 year and Bt stems had greater N concentration than NBt stems in 1 year of the 2 year study. ECB injury affected the composition of lignin-derived phenols, however ECB infested and non-infested stems lost the same amount of mass after 5 months in buried field litterbags. In conclusion ECB injury and the Bt gene had subtle effects on the chemical composition of corn tissue, which did not alter the short-term decomposition of corn residues.     

Genetic relationships between resistance to stalk-tunneling by the European corn borer and cell-wall components in maize population B73×B52

The objective of this study was to assess the relationships among quantitative trait loci (QTL) detected for European corn borer (ECB) tunneling and cell-wall components (CWC) neutral detergent fiber (NDF), acid detergent fiber (ADF), and acid detergent lignin (ADL) content in leaf-sheath and stalk tissues in a maize recombinant inbred line population derived from inbred lines B73 and B52. Most of the QTL for ECB resistance (10/13) were at QTL positions for one or more CWC. Of the 12 QTL for NDF and ADF in leaf-sheaths, five for each trait were at or near QTL for ECB tunneling. Four of these five QTL for NDF and ADF mapped to common locations. Four of the eight leaf-sheath ADL QTL were detected in the same genomic regions as ECB QTL. For stalk tissue, four regions contained common/overlapping QTL for ECB tunneling, NDF, and ADF. Six such regions were observed for stalk ADL and ECB tunneling. Seven of the ten QTL associated with both CWC and ECB tunneling contributed to the negative correlations observed between these traits, while relatively few QTL effects were positively correlated. This suggests that while CWC contribute to ECB resistance in this population, other mechanisms and other genes also are involved. Several QTL contributing to the negative correlations between ECB tunneling and CWC in the leaf-sheaths mapped to similar positions as QTL detected in tropical maize populations for resistance to leaf-feeding by Diatraea grandiosella Dyar and Diatraea saccharalis Fabricus. These regions may contain genes involved in the synthesis of cellulose, hemicellulose, and lignin in the leaf-blades and leaf-sheaths of maize.     

Evaluation of the European Union Maize Landrace Core Collection for resistance to Sesamia nonagrioides (Lepidoptera: Noctuidae) and Ostrinia nubilalis (Lepidoptera: Crambidae)

Two corn borer species are the principal maize insect pests in Europe, the European corn borer, Ostrinia nubilalis (Hübner), and the pink stem borer, Sesamia nonagrioides (Lefebvre). Hence, it would be advisable to evaluate the European maize germplasm for corn borer resistance to generate European varieties resistant to corn borer attack. The creation of the European Union Maize Landrace Core Collection (EUMLCC) allowed the screening of most of the variability for European corn borer resistance present among European maize local populations from France, Germany, Greece, Italy, Portugal, and Spain, testing a representative sample. The objective of this study was the evaluation of stem and ear resistance of the EUMLCC to European corn borer and pink stem borer attack. Trials were made at two Spanish locations that represent two very different maize-growing areas. Populations that performed relatively well under corn borer infestation for stem and ear damage were 'PRT0010008' and'GRC0010085', among very early landraces; 'PRT00100120' and 'PRT00100186', among early landraces; 'GRC0010174', among midseason landraces; and 'ESP0070441', among late landraces. Either the selection that could have happen under high insect pressure or the singular origin of determined maize populations would be possible explanations for the higher corn borer resistance of some landraces. Landraces 'PRT0010008', 'FRA0410090', 'PRT00100186', and 'ESP0090214' would be selected to constitute a composite population resistant to corn borers and adapted to short season, whereas populations 'ESP0090033', 'PRT00100530', 'GRC0010174', and 'ITA0370005' would be used to make a resistant composite adapted to longer season.