Population Structure, Demographic History, and Adaptation of Giant Honeybees in China Revealed by Population Genomic Data.

There have been many population-based genomic studies on human-managed honeybees (Apis mellifera and Apis cerana), but there has been a notable lack of analysis with regard to wild honeybees, particularly in relation to their evolutionary history. Nevertheless, giant honeybees have been found to occupy distinct habitats and display remarkable characteristics, which are attracting an increased amount of attention. In this study, we de novo sequenced and then assembled the draft genome sequence of the Himalayan giant honeybee, Apis laboriosa. Phylogenetic analysis based on genomic information indicated that A. laboriosa and its tropical sister species Apis dorsata diverged ∼2.61 Ma, which supports the speciation hypothesis that links A. laboriosa to geological changes throughout history. Furthermore, we re-sequenced A. laboriosa and A. dorsata samples from five and six regions, respectively, across their population ranges in China. These analyses highlighted major genetic differences for Tibetan A. laboriosa as well as the Hainan Island A. dorsata. The demographic history of most giant honeybee populations has mirrored glacial cycles. More importantly, contrary to what has occurred among human-managed honeybees, the demographic history of these two wild honeybee species indicates a rapid decline in effective population size in the recent past, reflecting their differences in evolutionary histories. Several genes were found to be subject to selection, which may help giant honeybees to adapt to specific local conditions. In summary, our study sheds light on the evolutionary and adaptational characteristics of two wild giant honeybee species, which was useful for giant honeybee conservation.

Sacbrood Virus: A Growing Threat to Honeybees and Wild Pollinators.

Sacbrood virus (SBV) is one of the many viruses that infect both the Western honeybee (Apis mellifera) and the Eastern honeybee (Apis cerana). Recently, the interspecies transmission of SBV has been discovered, especially among wild pollinators. This newly discovered evolutionary occurrence regarding SBV indicates a much wider host range than previously believed, causing further concern about the future sustainability of agriculture and the resilience of ecosystems. Over the past few decades, vast numbers of studies have been undertaken concerning SBV infection in honeybees, and remarkable progress has been made in our understanding of the epidemiology, pathogenesis, transmission, and manifestations of SBV infection in honeybees and other pollinators. Meanwhile, some methods, including Chinese medicine, have been established to control and prevent sacbrood disease in A. cerana in Asian countries. In this review, we summarize the existing knowledge of SBV and address the gaps in the knowledge within the existing literature in the hope of providing future directions for the research and development of management strategies for controlling the spread of this deadly disease.

Chromosome-scale genome assembly of the high royal jelly-producing honeybees.

A high royal jelly-producing strain of honeybees (HRJHB) has been obtained by successive artificial selection of Italian honeybees (Apis mellifera ligustica) in China. The HRJHB can produce amounts of royal jelly that are dozens of times greater than their original counterparts, which has promoted China to be the largest producer of royal jelly in the world. In this study, we generated a chromosome-scale of the genome sequence for the HRJHB using PacBio long reads and Hi-C technique. The genome consists of 16 pseudo-chromosomes that contain 222 Mb of sequence, with a scaffold N50 of 13.6 Mb. BUSCO analysis yielded a completeness score of 99.3%. The genome has 12,288 predicted protein-coding genes and a rate of 8.11% of repetitive sequences. One chromosome inversion was identified between the HRJHB and the closely related Italian honeybees through whole-genome alignment analysis. The HRJHB's genome sequence will be an important resource for understanding the genetic basis of high levels of royal jelly production, which may also shed light on the evolution of domesticated insects.

A Survey of Common Pathogens of Apis spp. in Wild Non-Cave Honeybees in Southwest China.

Honeybees are crucial pollinators with significant ecologic value. The decline of wild honeybee populations has been recognized and documented during recent decades. However, the health status of wild non-cave Apis spp., including giant and dwarf honeybees, remains generally unknown. We investigated eight common viruses and five bacterial or fungal pathogens in four wild non-cave honeybee species at 11 locations in Southwest China. As a result, Melissococcus plutonius, the pathogenic agent of European foulbrood, was detected in all the species, and the sequences were identical to the pathogen in managed cave honeybees. Only one virus, black queen cell virus (BQCV), was positive in one dwarf species, Apis florea, in our study. The positive BQCV infected three A. florea colonies in Guangxi Province, with distinct sequences from this virus reported in cave honeybees or in the same host in the nearby Yunnan Province. Although our results indicated a low pathogenic level of common diseases in the wild non-cave Apis spp. in Southwest China, the conservation of these wild pollinators is of importance in light of the noticeable decline in populations and the irreplaceable position of pollination.

The Phylogeny and Pathogenesis of Sacbrood Virus (SBV) Infection in European Honey Bees, Apis mellifera.

RNA viruses that contain single-stranded RNA genomes of positive sense make up the largest group of pathogens infecting honey bees. Sacbrood virus (SBV) is one of the most widely distributed honey bee viruses and infects the larvae of honey bees, resulting in failure to pupate and death. Among all of the viruses infecting honey bees, SBV has the greatest number of complete genomes isolated from both European honey bees Apis mellifera and Asian honey bees A. cerana worldwide. To enhance our understanding of the evolution and pathogenicity of SBV, in this study, we present the first report of whole genome sequences of two U.S. strains of SBV. The complete genome sequences of the two U.S. SBV strains were deposited in GenBank under accession numbers: MG545286.1 and MG545287.1. Both SBV strains show the typical genomic features of the Iflaviridae family. The phylogenetic analysis of the single polyprotein coding region of the U.S. strains, and other GenBank SBV submissions revealed that SBV strains split into two distinct lineages, possibly reflecting host affiliation. The phylogenetic analysis based on the 5'UTR revealed a monophyletic clade with the deep parts of the tree occupied by SBV strains from both A. cerane and A. mellifera, and the tips of branches of the tree occupied by SBV strains from A. mellifera. The study of the cold stress on the pathogenesis of the SBV infection showed that cold stress could have profound effects on sacbrood disease severity manifested by increased mortality of infected larvae. This result suggests that the high prevalence of sacbrood disease in early spring may be due to the fluctuating temperatures during the season. This study will contribute to a better understanding of the evolution and pathogenesis of SBV infection in honey bees, and have important epidemiological relevance.

Genomic and transcriptomic analysis of the Asian honeybee Apis cerana provides novel insights into honeybee biology.

The Asian honeybee Apis cerana is one of two bee species that have been commercially kept with immense economic value. Here we present the analysis of genomic sequence and transcriptomic exploration for A. cerana as well as the comparative genomic analysis of the Asian honeybee and the European honeybee A. mellifera. The genome and RNA-seq data yield new insights into the behavioral and physiological resistance to the parasitic mite Varroa the evolution of antimicrobial peptides, and the genetic basis for labor division in A. cerana. Comparison of genes between the two sister species revealed genes specific to A. cerana, 54.5% of which have no homology to any known proteins. The observation that A. cerana displayed significantly more vigilant grooming behaviors to the presence of Varroa than A. mellifera in conjunction with gene expression analysis suggests that parasite-defensive grooming in A. cerana is likely triggered not only by exogenous stimuli through visual and olfactory detection of the parasite, but also by genetically endogenous processes that periodically activates a bout of grooming to remove the ectoparasite. This information provides a valuable platform to facilitate the traits unique to A. cerana as well as those shared with other social bees for health improvement.

High Royal Jelly-Producing Honeybees (Apis mellifera ligustica) (Hymenoptera: Apidae) in China.

China is the largest producer and exporter of royal jelly (RJ) in the world, supplying >90% of the global market. The high production of RJ in China is principally owing to the high RJ-producing lineage of honeybees (Apis mellifera ligustica Spinola, 1806) established by beekeepers in the 1980s. We describe the development of high royal jelly-producing honeybees and the management of this lineage today. Previous research and recent advances in the genetic characterization of this lineage, and the molecular markers and mechanisms associated with high RJ production are summarized. The gaps in our knowledge and prospects for future research are also highlighted.

Sonication inhibited browning but decreased polyphenols contents and antioxidant activity of fresh apple (malus pumila mill, cv. Red Fuji) juice.

Enzyme browning is the main challenge in the preparation of fresh apple juice. The influence of sonication on browning, as well as polyphenols and antioxidant activity of fresh apple juice was investigated. It was found that ultrasound can inhibit the browning of fresh apple (Malus pumila Mill, cv. Red Fuji) juice, but decreased the contents of total phenolic content (TPC), total flavonoid content (TFC) and chlorogenic acid and reduced the antioxidant activity. On the whole, ultrasound technology cannot be used to the antibrowning of fresh apple (Malus pumila Mill, cv. Red Fuji) juice.

Geographical influences on content of 10-hydroxy-trans-2-decenoic acid in royal jelly in China.

The content of 10-hydroxy-trans-2-decenoic acid (10-HDA), a marker compound in royal jelly (RJ), is the most important criterion in grading RJ for commercial trade and varies with its origin. To identify the effect of geographical origin on 10-HDA content in RJ, 138 samples were collected from 19 provinces of China (divided into three groups) produced by either Apis mellifera ligustica Spinola, 1806 or a hybrid of A. m. ligustica and Apis mellifera carnica Pollman, 1879 and analyzed for moisture, sugar, crude protein, ash, acid, and 10-HDA concentration. The results show that RJ from western China has a significantly higher 10-HDA level (2.01 +/- 0.05%) than those from northeastern (1.87 +/- 0.05%) and eastern (1.75 +/- 0.03%) China. RJ secreted by hybrid bees contained more 10-HDA (1.89 +/- 0.03%) than that secreted by A. m. ligustica (1.78 +/- 0.03%). The 10-HDA content of RJ produced during flowering of rape (Brassica campestris L.), lime (Tilia amurensis Ruprecht), and vitex (Vitex negundo L. variety heterophylla (Franch.) Rehder) was 1.92, 1.80, and 1.68%, respectively. The results would be helpful during the process of price determination of RJ by providing some basis of geographical, bee strain, and botanical information for commercial trade.

[Phylogenetic relationships of honey bees (Hymenoptera: Apinae) inferred from mitochondrial and nuclear DNA sequences].

The sequences of mitochondrial ND2, CO2, and 16S rRNA genes and nuclear ITPR gene were obtained from 22 samples of 5 Apis species from China. The characteristics of the sequences and the pairwise distances among species were analyzed. Phylogenetic trees were reconstructed for Apis species using maximum parsimony, neighbor-joining and maximum likelihood methods together with the sequences of the other 4 Apis species downloaded from GenBank. Results supported that Apis species were divided into three major clusters: dwarf bees (A. florea and A. andreniformis), giant bees (A. dorsata and A. laboriosa), and cavity-nesting bees (A. mellifera, A. cerana, A. koschevnikovi, A. nigrocinta, and A. nuluensis). The dwarf honey bees were confirmed as basal lineage. Our study also revealed a high level of genetic divergence between A. dorsata from Hainan Island and China mainland.

Transcriptome comparison between honey bee queen- and worker-destined larvae.

Caste differentiation in the female honey bee is one of the most intriguing polyphenism phenomena. This developmental switch depends on the differential expression of entire suites of the genes involved in the larval fate between the queens and workers. In this study, we compared the transcriptome differences between full-sister queen- (QL) and worker-destined larvae (WL) using high-throughput RNA-Seq. QL and WL at fourth (L4) and fifth instar (L5) were used to prepare four libraries and to generate 50,191,699 (QL4), 57,628,541 (WL4), 56,613,619 (QL5), and 58,626,829 (WL5) usable reads, which were assembled into groups of 7,952, 7,993, 7,971, and 8,023 genes, respectively. The transcriptome changes were investigated using the DEGs Package (DEGseq), which resulted in more than 4,500 differentially expressed genes (DEGs) between the castes. Eight of the DEGs were verified by quantitative real-time RT-PCR (qRT-PCR), and the results supported our sequencing data. All of the DEGs were analysed using Web Gene Ontology Annotation Plot (WEGO) and then mapped using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. These results suggest that over 70% of the DEGs in each instar were more highly expressed in QL than in WL, possibly suggesting that the QL genes had higher transcriptional activity than the WL genes during differentiation. The same gene set is active (but differentially expressed) in both castes, which in turn result in dimorphic females. The L4 stage is a very active gene expression period for both QL and WL before their pupal stage. The activity of the mTOR (a target of rapamycin) encoding gene in the mTOR signalling pathway is higher in QL4 than in WL4, and this difference was no longer present by the L5 feeding stage. The genes down-stream of mTOR maintained this change at the L5 stage. These results could contribute to an in-depth study of the candidate genes during honey bee caste differentiation and improve our current understanding of the polyphenism phenomenon in insects.