Selection Signal Analysis Reveals Hainan Yellow Cattle Are Being Selectively Bred for Heat Tolerance.

Hainan yellow cattle are indigenous Zebu cattle from southern China known for their tolerance of heat and strong resistance to disease. Generations of adaptation to the tropical environment of southern China and decades of artificial breeding have left identifiable selection signals in their genomic makeup. However, information on the selection signatures of Hainan yellow cattle is scarce. Herein, we compared the genomes of Hainan yellow cattle with those of Zebu, Qinchuan, Nanyang, and Yanbian cattle breeds by the composite likelihood ratio method (CLR), Tajima's D method, and identifying runs of homozygosity (ROHs), each of which may provide evidence of the genes responsible for heat tolerance in Hainan yellow cattle. The results showed that 5210, 1972, and 1290 single nucleotide polymorphisms (SNPs) were screened by the CLR method, Tajima's D method, and ROH method, respectively. A total of 453, 450, and 325 genes, respectively, were identified near these SNPs. These genes were significantly enriched in 65 Gene Ontology (GO) functional terms and 11 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways (corrected p < 0.05). Five genes-Adenosylhomocysteinase-like 2, DnaJ heat shock protein family (Hsp40) member C3, heat shock protein family A (Hsp70) member 1A, CD53 molecule, and zinc finger and BTB domain containing 12-were recognized as candidate genes associated with heat tolerance. After further functional verification of these genes, the research results may benefit the understanding of the genetic mechanism of the heat tolerance in Hainan yellow cattle, which lay the foundation for subsequent studies on heat stress in this breed.

Neuropeptide hormone bursicon mediates female reproduction in the whitefly, Bemisia tabaci (Hemiptera: Aleyrodidae).

Bursicon, a neuropeptide hormone comprising two subunits-bursicon (burs) and partner of burs (pburs), belongs to the cystine-knot protein family. Bursicon heterodimers and homodimers bind to the lucine-rich G-protein coupled receptor (LGR) encoded by rickets to regulate multiple physiological processes in arthropods. Notably, these processes encompass the regulation of female reproduction, a recent revelation in Tribolium castaneum. In this study we investigated the role of burs/pburs/rickets in mediating female vitellogenesis and reproduction in a hemipteran insect, the whitefly, Bemisia tabaci. Our investigation unveiled a synchronized expression of burs, pburs and rickets, with their transcripts persisting detectable in the days following eclosion. RNAi-mediated knockdown of burs, pburs or rickets significantly suppressed the transcript levels of vitellogenin (Vg) and Vg receptor in the female whiteflies. These effects also impaired ovarian maturation and female fecundity, as evidenced by a reduction in the number of eggs laid per female, a decrease in egg size and a decline in egg hatching rate. Furthermore, knockdown of burs, pburs or rickets led to diminished juvenile hormone (JH) titers and reduced transcript level of Kruppel homolog-1. However, this impact did not extend to genes in the insulin pathway or target of rapamycin pathway, deviating from the results observed in T. castaneum. Taken together, we conclude that burs/pburs/rickets regulates the vitellogenesis and reproduction in the whiteflies by coordinating with the JH signaling pathway.

Unexpected worker mating and colony-founding in a superorganism.

The emergence of caste-differentiated colonies, which have been defined as 'superorganisms', in ants, bees, and wasps represents a major transition in evolution. Lifetime mating commitment by queens, pre-imaginal caste determination and lifetime unmatedness of workers are key features of these animal societies. Workers in superorganismal species like honey bees and many ants have consequently lost, or retain only vestigial spermathecal structures. However, bumble bee workers retain complete spermathecae despite 25-40 million years since their origin of superorganismality, which remains an evolutionary mystery. Here, we show (i) that bumble bee workers retain queen-like reproductive traits, being able to mate and produce colonies, underlain by queen-like gene expression, (ii) the social conditions required for worker mating, and (iii) that these abilities may be selected for by early queen-loss in these annual species. These results challenge the idea of lifetime worker unmatedness in superorganisms, and provide an exciting new tool for the conservation of endangered bumble bee species.

Functional analysis of a NPC1 gene from the whitefly, Bemisia tabaci (Hemiptera: Aleyrodidae).

Niemann-Pick C (NPC) disease is a neurodegenerative disorder related to cellular sterol trafficking and mutation of NPC1 gene is the main cause for this disease. The function of NPC1 have been reported in a few insects but rarely studied in hemipterans. In the present study, we investigate the function of NPC1 in a hemipteran pest, the whitefly Bemisia tabaci. It was found that B. tabaci had only one NPC1 homolog (BtNPC1), in contrast to two homologs in many other insects. BtNPC1 was ubiquitously expressed at all developmental stages and body parts of whiteflies, with the highest level in adult abdomen, and the expression of BtNPC1 was induced by cholesterol feeding. To further investigate the function of BtNPC1, leaf-mediated RNA interference experiments were carried out. Results showed that knockdown of BtNPC1 led to reduced survival of whiteflies, as well as reduced fecundity. Moreover, knockdown of BtNPC1 affected the development and metamorphosis of whitefly nymphs. Taken these together, we conclude that BtNPC1 played a crucial role in sterol-related biological processes of B. tabaci and might be used as an insecticide target for development of novel pest management approaches.

Divergence and convergence of gut microbiomes of wild insect pollinators.

Pollination services provided by wild insect pollinators are critical to natural ecosystems and crops around the world. There is an increasing appreciation that the gut microbiota of these insects influences their health and consequently their services. However, pollinator gut microbiota studies have focused on well-described social bees, but rarely include other, more phylogenetically divergent insect pollinators. To expand our understanding, we explored the insect pollinator microbiomes across three insect orders through two DNA sequencing approaches. First, in an exploratory 16S amplicon sequencing analysis of taxonomic community assemblages, we found lineage-specific divergences of dominant microbial genera and microbiota community composition across divergent insect pollinator genera. However, we found no evidence for a strong broad-scale phylogenetic signal, which we see for community relatedness at finer scales. Subsequently, we utilized metagenomic shotgun sequencing to obtain metagenome-assembled genomes and assess the functionality of the microbiota from pollinating flies and social wasps. We uncover a novel gut microbe from pollinating flies in the family Orbaceae that is closely related to Gilliamella spp. from social bees but with divergent functions. We propose this novel species be named Candidatus Gilliamella eristali. Further metagenomes of dominant fly and wasp microbiome members suggest that they are largely not host-insect adapted and instead may be environmentally derived. Overall, this study suggests selective processes involving ecology or physiology, or neutral processes determining microbe colonization may predominate in the turnover of lineages in insect pollinators broadly, while evolution with hosts may occur only under certain circumstances and on smaller phylogenetic scales. IMPORTANCE Wild insect pollinators provide many key ecosystem services, and the microbes associated with these insect pollinators may influence their health. Therefore, understanding the diversity in microbiota structure and function, along with the potential mechanisms shaping the microbiota across diverse insect pollinators, is critical. Our study expands beyond existing knowledge of well-studied social bees, like honey bees, including members from other bee, wasp, butterfly, and fly pollinators. We infer ecological and evolutionary factors that may influence microbiome structure across diverse insect pollinator hosts and the functions that microbiota members may play. We highlight significant differentiation of microbiomes among diverse pollinators. Closer analysis suggests that dominant members may show varying levels of host association and functions, even in a comparison of closely related microbes found in bees and flies. This work suggests varied importance of ecological, physiological, and non-evolutionary filters in determining structure and function across largely divergent wild insect pollinator microbiomes.

A ferritin protein is involved in the development and reproduction of the whitefly, Bemisia tabaci.

Ferritins are conserved iron-binding proteins that exist in most living organisms and play an essential role in the maintenance of cellular iron homeostasis. Although ferritin has been studied in many species, little is known about its role in the whitefly, Bemisia tabaci. In this study, we identified an iron-binding protein from B. tabaci and named it BtabFer1. The full-length cDNA of BtabFer1 is 1,043 bp and encodes a protein consisting of 224 amino acids with a deduced molecular weight of 25.26 kDa, and phylogenetic analysis shows that BtabFer1 is conserved among Hemiptera insects. The expression levels of BtabFer1 in different developmental stages and tissues were analyzed by real-time PCR, and results showed that BtabFer1 was ubiquitously expressed at all developmental stages and in all examined tissues. The RNAi-mediated knockdown of BtabFer1 caused a significant reduction in survival rate, egg production, and egg hatching rate of whiteflies. Knockdown of BtabFer1 also inhibited the transcription of genes in the juvenile hormone signal transduction pathway. Taken together, these results suggest that BtabFer1 plays a critical role in the development and reproduction of whiteflies. This study can broaden our understanding of ferritin in insect fecundity and development, as well as provide baseline data for future studies.

Identification and functional analysis of isopentenyl pyrophosphate isomerase genes in the whiteflies Bemisia tabaci (Hemiptera: Aleyrodidae).

The juvenile hormone (JH) plays a vital role in the regulation of a number of physiological processes, including development, reproduction, and ovarian maturation. Isopentenyl pyrophosphate isomerase (IPPI) is a key enzyme in the biosynthetic pathway of JH. In this study, we identified an isopentenyl pyrophosphate isomerase protein from Bemisia tabaci and named it BtabIPPI. The open reading frame (ORF) of BtabIPPI is 768 bp and encodes a protein of 255 amino acids that contains a conserved domain of the Nudix family. The temporal and spatial expression profiles showed that BtabIPPI was highly expressed in the female adults.RNA interference (RNAi)-mediated silencing of BtabIPPI reduced JH titers and the relative expression of vitellogenin receptor (VgR) and JH signaling pathway genes, resulting in a dramatic reduction in fecundity and hatchability. These results indicate that the BtabIPPI gene plays an important role in the female fecundity of B. tabaci. This study will broaden our understanding of the function of IPPI in regulating insect reproduction and provide a theoretical basis for targeting IPPI for pest control in the future.

A Hepatocyte Nuclear Factor BtabHNF4 Mediates Desiccation Tolerance and Fecundity in Whitefly (Bemisia tabaci).

Hepatocyte nuclear factor 4 (HNF4) is essential for glucose homeostasis and lipid metabolism in insects. However, little is known about the role of HNF4 in whiteflies. In the present study, we identified a hepatocyte nuclear factor protein from Bemsia tabaci (Diptera: Drosophilidae) and named it BtabHNF4. The full-length of BtabHNF4 was 3,006 bp, encoding a sequence of 434 amino acids that contains a conserved zinc-finger DNA-binding domain (DBD) and a well-conserved ligand-binding domain (LBD). The temporal and spatial expression showed that BtabHNF4 was highly expressed in the female adult stage and abdominal tissues of B. tabaci. A leaf-mediated RNA interference method was used to explore the function of BtabHNF4 in whiteflies. Our results showed that the knockdown of BtabHNF4 influences the desiccation tolerance, egg production, and egg hatching rate of whiteflies. Additionally, BtabHNF4 silencing significantly inhibited the expression level of vitellogenin. These results expand the function of HNF4 and pave the way for understanding the molecular mechanisms of HNF4 in regulating multiple physiological processes.

Transcriptome Profiling Reveals a Novel Mechanism of Antiviral Immunity Upon Sacbrood Virus Infection in Honey Bee Larvae (Apis cerana).

The honey bee is one of the most important pollinators in the agricultural system and is responsible for pollinating a third of all food we eat. Sacbrood virus (SBV) is a member of the virus family Iflaviridae and affects honey bee larvae and causes particularly devastating disease in the Asian honey bees, Apis cerana. Chinese Sacbrood virus (CSBV) is a geographic strain of SBV identified in China and has resulted in mass death of honey bees in China in recent years. However, the molecular mechanism underlying SBV infection in the Asian honey bee has remained unelucidated. In this present study, we employed high throughput next-generation sequencing technology to study the host transcriptional responses to CSBV infection in A. cerana larvae, and were able to identify genome-wide differentially expressed genes associated with the viral infection. Our study identified 2,534 differentially expressed genes (DEGs) involved in host innate immunity including Toll and immune deficiency (IMD) pathways, RNA interference (RNAi) pathway, endocytosis, etc. Notably, the expression of genes encoding antimicrobial peptides (abaecin, apidaecin, hymenoptaecin, and defensin) and core components of RNAi such as Dicer-like and Ago2 were found to be significantly upregulated in CSBV infected larvae. Most importantly, the expression of Sirtuin target genes, a family of signaling proteins involved in metabolic regulation, apoptosis, and intracellular signaling was found to be changed, providing the first evidence of the involvement of Sirtuin signaling pathway in insects' immune response to a virus infection. The results obtained from this study provide novel insights into the molecular mechanism and immune responses involved in CSBV infection, which in turn will contribute to the development of diagnostics and treatment for the diseases in honey bees.

Using the combined gene approach and multiple analytical methods to improve the phylogeny and classification of Bombus (Hymenoptera, Apidae) in China.

Bumble bees are vital to our agro-ecological system, with approximately 250 species reported around the world in the single genus Bombus. However, the health of bumble bees is threatened by multiple factors: habitat loss, climate change, pesticide use, and disease caused by pathogens and parasites. It is therefore vitally important to have a fully developed phylogeny for bumble bee species as part of our conservation efforts. The purpose of this study was to explore the phylogenetic relationships of the dominant bumble bees on the Tibetan plateau and in northern China as well as their placement and classification within the genus Bombus. The study used combined gene analysis consisting of sequence fragments from six genes, 16S rRNA, COI, EF-1α, Argk, Opsin and PEPCK, and the phylogenetic relationships of 209 Bombus species were explored. Twenty-six species, including 152 gene sequences, were collected from different regions throughout China, and 1037 gene sequences representing 183 species were obtained from GenBank or BOLD. The results suggest that the 209 analyzed species belong to fifteen subgenera and that most of the subgenera in Bombus are monophyletic, which is in accordance with conventional morphology-based classifications. The phylogenetic trees also show that nearly all subgenera easily fall into two distinct clades: short-faced and long-faced. The study is the first to investigate the phylogenetic placement of Bombus turneri (Richards), Bombus opulentus Smith, Bombus pyrosoma Morawitz, Bombus longipennis Friese, Bombus minshanensis Bischoff, and Bombus lantschouensis Vogt, all of which are widely distributed throughout different regions of China. The knowledge and understanding gained from the findings can provide a molecular basis to accurately classify Bombus in China and to define strategies to conserve biodiversity and promote pollinator populations.

Dynamic Changes of Gut Microbial Communities of Bumble Bee Queens through Important Life Stages.

Bumble bees are important pollinators in natural and agricultural ecosystems. Their social colonies are founded by individual queens, which, as the predominant reproductive females of colonies, contribute to colony function through worker production and fitness through male and new queen production. Therefore, queen health is paramount, but even though there has been an increasing emphasis on the role of gut microbiota for animal health, there is limited information on the gut microbial dynamics of bumble bee queens. Employing 16S rRNA amplicon sequencing and quantitative PCR, we investigate how the adult life stage and physiological state influence a queen's gut bacterial community diversity and composition in unmated, mated, and ovipositing queens of Bombus lantschouensis We found significant shifts in total gut microbe abundance and microbiota composition across queen states. There are specific compositional signatures associated with different stages, with unmated and ovipositing queens showing the greatest similarity in composition and mated queens being distinct. The bacterial genera Gilliamella, Snodgrassella, and Lactobacillus were relatively dominant in unmated and ovipositing queens, with Bifidobacterium dominant in ovipositing queens only. Bacillus, Lactococcus, and Pseudomonas increased following queen mating. Intriguingly, however, further analysis of unmated queens matching the mated queens in age showed that changes are independent of the act of mating. Our study is the first to explore the gut microbiome of bumble bee queens across key life stages from adult eclosion to egg laying and provides useful information for future studies of the function of gut bacteria in queen development and colony performance.IMPORTANCE Bumble bee queens undergo a number of biological changes as they transition through adult emergence, mating, overwintering, foraging, and colony initiation including egg laying. Therefore, they represent an important system to understand the link between physiological, behavioral, and environmental changes and host-associated microbiota. It is plausible that the bumble bee queen gut bacteria play a role in shaping the ability of the queen to survive environmental extremes and reproduce, due to long-established coevolutionary relationships between the host and microbiome members.