Stable composition of gut microbiome in the Asian ladybeetle Coccinella septempunctata reared on natural and artificial diets.

The Asian ladybeetle, Coccinella septempunctata, is an important insect of predatory natural enemy, which has a strong control effect and application prospects for aphids, whiteflies, mealybugs, and other small-sized pests of agriculture and forestry crops. Gut microbiota composition posed impacts on development of insects. In order to clarify the effect of artificial feed feeding on the intestinal microbial species and structure of C. septempunctata, we compared the intestinal microbial flora of C. septempunctata reared on bean aphids and artificial food for 15 days. Results show that Proteobacteria was the dominant component in all groups at phylum level, Rhodobacter, Methylovigula, Burkholderia, and Bradyrhizobium were the dominant bacteria among all groups at genus level. As to the differences in bacterial community structure and diversity, there is no significant difference between Shannon index and Simpson index, the principal components analysis of the bacterial communities, and the samples were roughly distributed in different regions. After 15 days of feeding, artificial diet did not significantly reduce the microbial diversity of the gut of C. septempunctata compared to the aphid group, and there was no significant effect on the abundance of dominant flora in the gut of C. septempunctata, C. septempunctata gut has a similar core microbiota. This study clarifies the effects in intestinal microbial diversity and composition structure of the C. septempunctata with artificial diet, and provides a theoretical basis for understanding the intestinal microorganisms and optimizating the artificial diet of C. septempunctata.

Chromosome-level genome assembly of the giant ladybug Megalocaria dilatata.

The giant ladybug Megalocaria dilatata (Fabricius) is a potential biocontrol agent and a valuable model for coccinellid genomics and evolutionary biology. However, the lack of a reference genome for M. dilatata has impeded further explorations into its evolution and constrained its use in pest management. Here, we assembled and annotated a high-quality, chromosome-level genome of M. dilatata. The resulting assembly spans 772.3 Mb, with a scaffold N50 of 72.48 Mb and a GC content of 34.23%. The Hi-C data aided in anchoring the assembly onto 10 chromosomes ranging from 43.35 to 108.16 Mb. We identified 493.33 Mb of repeat sequences, accounting for 63.88% of the assembled genome. Our gene prediction identified 25,346 genes, with 81.89% annotated in public protein databases. The genome data will provide a valuable resource for studying the biology and evolution of Coccinellidae, aiding in pest control strategies and advancing research in the field.

The Adaptive Evolution in the Fall Armyworm Spodoptera frugiperda (Lepidoptera: Noctuidae) Revealed by the Diversity of Larval Gut Bacteria.

Insect gut microbes have important roles in host feeding, digestion, immunity, development, and coevolution with pests. The fall armyworm, Spodoptera frugiperda (Smith, 1797), is a major migratory agricultural pest worldwide. The effects of host plant on the pest's gut bacteria remain to be investigated to better understand their coevolution. In this study, differences in the gut bacterial communities were examined for the fifth and sixth instar larvae of S. frugiperda fed on leaves of different host plants (corn, sorghum, highland barley, and citrus). The 16S rDNA full-length amplification and sequencing method was used to determine the abundance and diversity of gut bacteria in larval intestines. The highest richness and diversity of gut bacteria were in corn-fed fifth instar larvae, whereas in sixth instar larvae, the richness and diversity were higher when larvae were fed by other crops. Firmicutes and Proteobacteria were dominant phyla in gut bacterial communities of fifth and sixth instar larvae. According to the LDA Effect Size (LEfSe) analysis, the host plants had important effects on the structure of gut bacterial communities in S. frugiperda. In the PICRUSt2 analysis, most predicted functional categories were associated with metabolism. Thus, the host plant species attacked by S. frugiperda larvae can affect their gut bacterial communities, and such changes are likely important in the adaptive evolution of S. frugiperda to host plants.

Complete Mitochondrial Genomes and Phylogenetic Positions of Two Longicorn Beetles, Anoplophora glabripennis and Demonax pseudonotabilis (Coleoptera: Cerambycidae).

Anoplophora glabripennis (Motschulsky, 1854) and Demonax pseudonotabilis Gressitt & Rondon, 1970 are two commonly found longicorn beetles from China. However, the lack of sufficient molecular data hinders the understanding of their evolution and phylogenetic relationships with other species of Cerambycidae. This study sequenced and assembled the complete mitochondrial genomes of the two species using the next-generation sequencing method. The mitogenomes of A. glabripennis and D. pseudonotabilis are 15,622 bp and 15,527 bp in length, respectively. The mitochondrial gene content and gene order of A. glabripennis and D. pseudonotabilis are highly conserved with other sequenced longicorn beetles. The calculation of nonsynonymous (Ka) and synonymous (Ks) substitution rates in PCGs indicated the existence of purifying selection in the two longicorn beetles. The phylogenetic analysis was conducted using the protein-coding gene sequences from available mitogenomes of Cerambycidae. The two species sequenced in this study are, respectively, grouped with their relatives from the same subfamily. The monophyly of Cerambycinae, Dorcasominae, Lamiinae, and Necydalinae was well-supported, whereas Lepturinae, Prioninae, and Spondylidinae were recovered as paraphyletic.

The First Mitochondrial Genomes of the Family Haplodiplatyidae (Insecta: Dermaptera) Reveal Intraspecific Variation and Extensive Gene Rearrangement.

Haplodiplatyidae is a recently established earwig family with over 40 species representing a single genus, Haplodiplatys Hincks, 1955. The morphology of Haplodiplatyidae has been studied in detail, but its molecular characters remain unclear. In this study, two mitogenomes of Haplodiplatys aotouensis Ma & Chen, 1991, were sequenced based on two samples from Fujian and Jiangxi provinces, respectively. These represent the first mitogenomes for the family Haplodiplatyidae. The next-generation sequencing method and subsequent automatic assembly obtained two mitogenomes. The two mitogenomes of H. aotouensis were generally identical but still exhibit a few sequence differences involving protein-coding genes (PCGs), ribosomal RNA (rRNA) genes, control regions, and intergenic spacers. The typical set of 37 mitochondrial genes was annotated, while many transfer RNA (tRNA) genes were rearranged from their ancestral locations. The calculation of nonsynonymous (Ka) and synonymous (Ks) substitution rates in PCGs indicated the fastest evolving nd4l gene in H. aotouensis. The phylogenetic analyses supported the basal position of Apachyidae but also recovered several controversial clades.

Similar Gut Bacterial Microbiota in Two Fruit-Feeding Moth Pests Collected from Different Host Species and Locations.

Numerous gut microbes are associated with insects, but their composition remains largely unknown for many insect groups, along with factors influencing their composition. Here, we compared gut bacterial microbiota of two co-occurring agricultural pests, the peach fruit moth (PFM), Carposina sasakii, and the oriental fruit moth (OFM), Grapholita molesta, collected from different orchards and host plant species. Gut microbiota of both species was mainly composed of bacteria from Proteobacteria, followed by Firmicutes. The two species shared bacteria from the genera Pseudomonas, Gluconobacter, Acetobacter, and Pantoea. When we compared two pairs of PFM and OFM populations collected from the same host species and the same orchard, there is no difference in alpha and beta diversity in gut microbiota. When we compared gut microbiota of the same species and host plant from different orchards, alpha and beta diversity was different in populations of PFM collected from two pear orchards but not in other comparisons. Our study suggests that the two pests share many features of gut microbiota and environment in orchards is a main factor influencing their gut microbiota.

The highly rearranged mitochondrial genomes of three economically important scale insects and the mitochondrial phylogeny of Coccoidea (Hemiptera: Sternorrhyncha).

The mitochondrial genomes (mitogenomes) of scale insects are less known in comparison to other insects, which hinders the phylogenetic and evolutionary studies of Coccoidea and higher taxa. Herein, the complete mitogenomes of Unaspis yanonensis, Planococcus citri and Ceroplastes rubens were sequenced for Coccoidea. The 15,220-bp long mitogenome of U. yanonensis contained the typical set of 37 genes including 13 PCGs, 22 tRNA genes and two rRNA genes; the 15,549-bp long mitogenome of P. citri lacked the tRNA gene trnV; the 15,387-bp long mitogenome of C. rubens exhibited several shortened PCGs and lacked five tRNA genes. The mitochondrial gene arrangement of the three mitogenomes was different from other scale insects and Drosophila yakuba. Most PCGs used standard ATN (ATA, ATT, ATC and ATG) start codons and complete TAN (TAA or TAG) termination codons. The ND4L had the highest evolutionary rate but COX1 and CYTB were the lowest. Most tRNA genes had cloverleaf secondary structures, whereas the reduction of dihydrouridine (DHU) arms and TψC arms were detected. Tandem repeats, stem-loop (SL) structures and poly-[TA]n stretch were found in the control regions (CRs) of the three mitogenomes. The phylogenetic analyses using Bayesian inference (BI) and maximum likelihood methods (ML) showed identical results, both supporting the inner relationship of Coccoidea as Coccidae + (Pseudococcidae + Diaspididae).

Mitogenomic features of the citrus leafminer, Phyllocnistis citrella (Lepidoptera: Gracillariidae) and the related mitogenomic phylogeny.

The complete mitochondrial genome of the citrus leafminer, Phyllocnistis citrella (Lepidoptera: Gracillariidae) was sequenced for the first time, which was 15,416 bp in length and with an A + T content of 81.9%. The common set of 37 genes (13 PCGs, 22 tRNA genes, and two rRNA genes) were all annotated. Most PCGs had standard ATN start codons and TAN stop codons. Most tRNA genes exhibited typical cloverleaf secondary structures, whereas the dihydrouridine (DHU) arm of trnSer (AGN) was reduced. The mitochondrial phylogenetic analyses consistently recovered the relationships within Gracillariidae.

[EST-SSR identification, markers development of Ligusticum chuanxiong based on Ligusticum chuanxiong transcriptome sequences].

Ligusticum chuanxiong is a well-known traditional Chinese medicine plant. The study on its molecular markers development and germplasm resources is very important. In this study, we obtained 24 422 unigenes by assembling transcriptome sequencing reads of L. chuanxiong root. EST-SSR was detected and 4 073 SSR loci were identified. EST-SSR distribution and characteristic analysis results showed that the mono-nucleotide repeats were the main repeat types, accounting for 41.0%. In addition, the sequences containing SSR were functionally annotated in Gene Ontology (GO) and KEGG pathway and were assigned to 49 GO categories, 242 KEGG pathways, among them 2 201 sequences were annotated against Nr database. By validating 235 EST-SSRs,74 primer pairs were ultimately proved to have high quality amplification. Subsequently, genetic diversity analysis, UPGMA cluster analysis, PCoA analysis and population structure analysis of 34 L. chuanxiong germplasm resources were carried out with 74 primer pairs. In both UPGMA tree and PCoA results, L. chuanxiong resources were clustered into two groups, which are believed to be partial related to their geographical distribution. In this study, EST-SSRs in L. chuanxiong was firstly identified, and newly developed molecular markers would contribute significantly to further genetic diversity study, the purity detection, gene mapping, and molecular breeding.

Mitochondrial genomes of the hoverflies Episyrphus balteatus and Eupeodes corollae (Diptera: Syrphidae), with a phylogenetic analysis of Muscomorpha.

The hoverflies Episyrphus balteatus and Eupeodes corollae (Diptera: Muscomorpha: Syrphidae) are important natural aphid predators. We obtained mitochondrial genome sequences from these two species using methods of PCR amplification and sequencing. The complete Episyrphus mitochondrial genome is 16,175 bp long while the incomplete one of Eupeodes is 15,326 bp long. All 37 typical mitochondrial genes are present in both species and arranged in ancestral positions and directions. The two mitochondrial genomes showed a biased A/T usage versus G/C. The cox1, cox2, cox3, cob and nad1 showed relatively low level of nucleotide diversity among protein-coding genes, while the trnM was the most conserved one without any nucleotide variation in stem regions within Muscomorpha. Phylogenetic relationships among the major lineages of Muscomorpha were reconstructed using a complete set of mitochondrial genes. Bayesian and maximum likelihood analyses generated congruent topologies. Our results supported the monophyly of five species within the Syrphidae (Syrphoidea). The Platypezoidea was sister to all other species of Muscomorpha in our phylogeny. Our study demonstrated the power of the complete mitochondrial gene set for phylogenetic analysis in Muscomorpha.

Effects of Transgenic Bt Rice on Nontarget Rhopalosiphum maidis (Homoptera: Aphididae).

The effects of three transgenic Bacillus thuringiensis (Bt) rice lines, KMD1, KMD2, and G8-7, on biological parameters and population dynamics of nontarget insect, Rhopalosiphum maidis (Fitch) (Homoptera: Aphididae), were investigated in the laboratory and field. No significant differences were found between Bt and non-Bt rice lines for aphid survival. The developmental time of R. maidis that fed on KMD1 and KMD2 did not differ significantly from those of the individuals feeding on the parental variety Xiushui11, but significantly prolonged developmental time was observed on G8-7 as compared with its parental variety Xiushui110. Aphid fecundity was significantly higher on Bt than on parental rice. A 2-yr field survey indicated that Bt rice did not significantly affect the population dynamics of R. maidis in comparison with non-Bt rice. Additionally, guttation droplets of Bt rice and aphids feeding on Bt rice were analyzed for presence of Cry1Ab using ELISA. No Cry1Ab protein was found in aphid adults feeding on Bt rice lines both in the laboratory and field. By using the guttation droplets from the top of rice seedlings, we designed a novel method to collect phloem sap, and found that relatively low concentrations were detected in the guttation droplets from Bt rice lines. In conclusion, although the Bt rice lines tested in this study stimulate the fecundity of R. maidis, the aphid population density did not increase in Bt rice fields.

Multi-generation effects of Bt rice on Anagrus nilaparvatae, a parasitoid of the nontarget pest Nilapavarta lugens.

Little is known about the potential cumulative long-term effects of transgenic crops on nontarget organisms. In the present laboratory study, the potential cumulative effects of transgenic Bacillus thuringiensis (Bt) rice on parasitoids in successive generations were observed for an egg parasitoid, Anagrus nilaparvatae parasitizing eggs of Nilaparvata lugens (Stål) (Hemiptera: Delphacidae) feeding on Bt rice. Enzyme-linked immunosorbent assay test confirmed that Cry1Ab insecticidal protein could be detected in newly eclosed parasitoid adults. However, no significant effect on the fecundity of Anagrus nilaparvatae Pang et Wang (Hymenoptera: Mymaridae) was observed between Bt and non-Bt rice. Developmental times of both genders of A. nilaparvatae parasitizing host eggs laid in Bt (KMD1 and KMD2) rice lines were significantly prolonged from first generation to second generation, but not always prolonged from third generation to 11th generation as compared with the control rice line. Furthermore, the sex ratio of A. nilaparvatae progeny from the first generation to 11th generation in three rice lines was not significantly different. In general, our results suggested that the effect of Bt rice on this parasitoid could be negligible.