DNA metabarcoding uncovers the diet of subterranean rodents in China.

OBJECTIVE: A type of rodent called a zokor causes great harm to agriculture and forestry production due to its large and sophisticated diet. As this rodent lives subterrane for most of its life, researchers know little about its dietary habits. Further understanding of its diet is important for developing green and sustainable control strategies for the zokor. METHODS: Chloroplast trnL gene and internal transcription spacer 1 primers were selected for high-throughput sequencing of stomach contents of captured zokor by DNA metabarcoding. RESULTS: A total of 25 zokors were selected, the food list of 32 families, 80 genera, and 154 species was obtained. At the family level, it was found that zokors mainly fed on Asteraceae, Poaceae, Rosaceae, Pinaceae, Brassicaceae, and Apiaceae. At the genus level, zokors mainly fed on Echinops, Littledalea, Artemisia, Picea, Cirsium, and Elymus. The diet alpha diversity of Eospalax cansus was slightly higher than that of Eospalax cansus (P>0.05). The zokor's diet is highly phconsistent with the resources of its habitat. Most food choices tend to be the same between the two zokors. They fed primarily on Calamagrostis, Cirsium, Echinops, Medicago, Sanguisorba, and Taraxacum. Zokors mainly fed on the roots of perennial herbs, which are important source of energy. CONCLUSION: High-throughput sequencing-based DNA metabarcoding technology has effectively revealed the diet of zokors and indicated that zokors are food generalists.

The associations between intestinal bacteria of Eospalax cansus and soil bacteria of its habitat.

BACKGROUND: Intestinal bacteria of mammal can be influenced by many factors, environmental bacteria is an important factor. However, there are few studies on the interactions between environmental bacteria and intestinal bacteria in wild mammals. To explore the associations between the intestinal bacteriome and the related environmental bacteriome, the intestinal bacterial communities of Eospalax cansus at three different sites and the bacterial communities of the surrounding soil (outside and inside the cave) at each site were investigated by 16S rRNA sequencing. RESULTS: The composition and structure between zokor intestinal bacteria and related soil bacteria were distinct, and the soil of zokor habitat harbored significantly higher diversity than that of zokor intestinal bacteria. We have found that host factors may be more important than environmental factors in shaping intestinal bacteriome. In addition, it was found that the relative abundances of shared OTUs between zokors and related soil were significantly negatively related. These shared OTUs were present in the soil at relatively low abundance. However, these shared OTUs between zokors and soil were affiliated with diverse bacterial taxa, and they were related to the degradation of complex carbohydrates. CONCLUSIONS: These results suggested that the zokor gut may mainly select for low-abundance but diverse soil bacteria, which may be a host- specific choice for zokor to meet the needs of its phytophagous dietary.

Functional differentiation related to decomposing complex carbohydrates of intestinal microbes between two wild zokor species based on 16SrRNA sequences.

BACKGROUND: The intestinal microbes in mammals play a key role in host metabolism and adaptation. As a subterranean rodent, zokor digs tunnels for foraging and mating. These digging activities of zokors increase the energy expenditure relative to their aboveground counterparts. However, relatively little is known regarding intestinal microbes of zokor and how they make full use of limited food resources underground for high energy requirements. RESULTS: Eospalax cansus and Eospalax rothschildi had distinct intestinal microbes. Although the composition of intestinal microbes is similar in two species, the proportion of bacterium are distinctly different between them. At phylum level, 11 phyla were shared between two species. Firmicutes and Bacteroidota were two dominant microbes in both of two species, while Eospalax cansus have a significantly high proportion of Firmicutes/Bacteroidota than that of Eospalax rothschildi. At genus level, norank_f_Muribaculaceae were dominant microbes in both of two zokor species. The relative abundance of 12 genera were significantly different between two species. Some bacterium including unclassified_f__Lachnospiraceae, Lachnospiraceae_NK4A136_group, Ruminococcus and Eubacterium_siraeum_group associated with cellulose degradation were significantly enriched in Eospalax cansus. Although alpha diversity was with no significant differences between Eospalax cansus and Eospalax rothschildi, the intestinal microbes between them are significant distinct in PCoA analysis. We have found that trapping location affected the alpha diversity values, while sex and body measurements had no effect on alpha diversity values. PICRUSt metagenome predictions revealed significant enrichment of microbial genes involved in carbohydrate metabolism in Eospalax cansus rather than Eospalax rothschildi. CONCLUSIONS: Our results demonstrate that Eospalax cansus harbor a stronger ability of fermentation for dietary plants than Eospalax rothschildi. The stronger ability of fermentation and degradation of cellulose of intestinal microbes of Eospalax cansus may be a long-time adaptation to limited food resources underground.

Taxonomy and phylogenetic relationship of zokors.

Zokor (Myospalacinae) is one of the subterranean rodents, endemic to east Asia. Due to the convergent and parallel evolution induced by its special lifestyles, the controversies in morphological classification of zokor appeared at the level of family and genus. To resolve these controversies about taxonomy and phylogeny, the phylogenetic relationships of 20 species of Muroidea and six species of zokors were studied based on complete mitochondrial genome and mitochondrial Cytb gene, respectively. Phylogeny analysis of 20 species of Muroidea indicated that the zokor belonged to the family Spalacidae, and it was closer to mole rat rather than bamboo rat. Besides, by investigating the phylogenetic relationships of six species of zokors, the status of two genera of Eospalax and Myospalax was affirmed because the two clades differentiated in phylogenetic tree represented two types of zokors, convex occiput type and flat occiput type, respectively. In addition, the two origins in Eospalax were found diverged at 3.71 million years ago (Ma) based on estimation of divergence time. It is suggested that the climate and ecology changes caused by the Qinghai-Tibet Plateau uplift event in 3.6 Ma led to the inner divergence of Eospalax. The intraspecific phylogenetic relationships of partial zokors were well resolved, the two clades of Eospalax cansus represented two geographical populations, respectively, and the divergent pattern of Eospalax baileyi was characterized by allopatric divergence spatially. In this study, we explored the taxonomic status and phylogenetic relationships of Myospalacinae at the molecular level. These works would be significant to understanding the evolutionary process and to clarify the mechanism of differentiation of Myospalacinae.

The complete mitogenome of Eucryptorrhynchus brandti (Harold) (Insecta: Coleoptera: Curculionidae).

The complete mitochondrial genome of Eucryptorrhynchus brandti (Harold) (Insecta: Coleoptera: Curculionidae) were reconstructed from whole-genome Illumina Hiseq 2000 sequencing data with an average coverage of 1406.7X. The circular genome is 15,122 bp in length, consisting of 13 protein-coding genes (PCGs), 21 transfer RNAs (tRNAs), two ribosomal RNAs (rRNAs) and one D-loop or control region. The tRNA-Ile gene was not found in the mitochondrial genome, as is identical to two other curculionidae species, i.e. Sphenophorus sp. (GU176342) and Naupactus xanthographus (GU176345). All PCGs initiated with ATN codons, except for the ND1 started with TTG. Two PCGs (COI and ND4) have an incomplete stop codon T. Two PCGs (ND4L and ND1) harbor the stop codon TAG, while all other PCGs terminated with the TAA codon. The nucleotide composition is highly asymmetric (38.7% A, 14.4% C, 9.2% G and 37.8% T) with an overall AT content of 76.5%.

Analysis of Inter-Individual Bacterial Variation in Gut of Cicada Meimuna mongolica (Hemiptera: Cicadidae).

Intestinal bacterial community plays a crucial role in the nutrition, development, survival, and reproduction of insects. When compared with other insects with piercing-sucking mouthparts, the habitats of cicada nymphs and adults are totally different. However, little is known about the differences in the gut bacterial communities in the nymphs and adults within any cicada species. The diversity of bacteria in the gut of nymphs and adults of both genders of Meimuna mongolica (Distant) was studied using the denaturing gradient gel electrophoresis (DGGE) method. Few inter-individual variations among gut microbiota were observed, suggesting that M. mongolica typically harbors a limited and consistent suite of bacterial species. Bacteria in the genera Pseudomonas and Enterobacter were the predominant components of the gut microflora of M. mongolica at all life stages. Bacteria of Pantoea, Streptococcus, and Uruburuella were also widespread in the cicada samples but at relatively lower concentrations. The relative stability and similarity of the PCR-DGGE patterns indicate that all individuals of this cicada species harbor a characteristic bacterial community which is independent from developmental stages and genders. Related endosymbionts that could be harbored in bacteromes of cicadas were not detected in any gut samples, which could be related to the cicada species and the distribution of these endosymbionts in the cicada cavity, or due to some of the possible limitations of PCR-DGGE community profiling. It is worthwhile to further address if related cicada endosymbiont clades distribute in the alimentary canals and other internal organs through diagnostic PCR using group-specific primer sets.

Composition and diversity analysis of the gut bacterial community of the Oriental armyworm, Mythimna separata, determined by culture-independent and culture-dependent techniques.

The intestinal bacteria community structure and diversity of the Oriental armyworm, Mythimna separata (Walker) (Lepidoptera: Noctuidae), was studied by analysis of a 16S rDNA clone library, denaturing gradient gel electrophoresis,and culture-dependent techniques. The 16S rDNA clone library revealed a bacterial community diversity comprising Cyanobacteria, Firmicutes, Actinobacteria, Gracilicutes and Proteobacteria, among which Escherichia coli (Migula) (Enterobacteriales: Enterobacteriaceae) was the dominant bacteria. The intestinal bacteria isolated by PCR-denaturing gradient gel electrophoresis were classified to Firmicutes, Proteobacteria, and Gracilicutes, and E. coli was again the dominant bacteria. The culture-dependent technique showed that the intestinal bacteria belonged to Firmicutes and Actinobacteria, and Staphylococcus was the dominant bacteria. The intestinal bacteria of M. separata were widely distributed among the groups Cyanobacteria, Firmicutes, Actinobacteria, Gracilicutes, Proteobacteria, and Gracilicutes. 16S rDNA clone library, denaturing gradient gel electrophoresis, and culture-dependent techniques should be integrated to obtain precise results in terms of the microbial community and its diversity.

The gut bacteria associated with Camponotus japonicus Mayr with culture-dependent and DGGE methods.

The bacterial composition and distribution in the different gut regions of Camponotus japonicus were investigated using both culture-dependent method and culture-independent method of polymerase chain reaction and denaturing gradient gel electrophoresis (PCR-DGGE). Five different bacterial strains were isolated using culture-dependent method, and they all belong to the phylum Firmicutes, including three genera of bacteria Bacillus, Paenibacillus, and Enterococcus. Bacillus cereus and Enterococcus mundtii were found in the midgut; Paenibacillus sp. was isolated from the hindgut; and the other two Bacillus spp. were isolated from the crop. Twelve distinct DGGE bands were found using PCR-DGGE method, and their sequences blasting analysis shows that they are members of the Proteobacteria and the Firmicutes, respectively, including three genera (Pseudomonas, Candidatus Blochmannia, Fructobacillus) and one uncultured bacterium, in which Pseudomonas was the most dominant bacteria group in all the three gut regions. According to the DGGE profile, the three gut regions had very similar gut communities, and all the DGGE bands were presented in the midgut and hindgut, while just two bands representing Blochmannia were not present in the crop. The results of our study indicate that the gut of C. japonicus harbors several other bacteria besides the obligate endosymbionts Blochmannia, and more work should be carried on to verify if they are common in the guts of other Camponotus ants.