Polyunsaturated fatty acids stimulate immunity and eicosanoid production in Drosophila melanogaster.

Eicosanoids are a class of molecules derived from C20 polyunsaturated fatty acids (PUFAs) that play a vital role in mammalian and insect biological systems, including development, reproduction, and immunity. Recent research has shown that insects have significant but lower levels of C20 PUFAs in circulation in comparison to C18 PUFAs. It has been previously hypothesized in insects that eicosanoids are synthesized from C18 precursors, such as linoleic acid (LA), to produce downstream eicosanoids. In this study, we show that introduction of arachidonic acid (AA) stimulates production of cyclooxygenase, lipoxygenase, and cytochrome P450-derived eicosanoids. Downstream immune readouts showed that LA stimulates phagocytosis by hemocytes, while both LA and AA stimulate increased antimicrobial peptide production when D. melanogaster is exposed to a heat-killed bacterial pathogen. In totality, this work identifies PUFAs that are involved in insect immunity and adds evidence to the notion that Drosophila utilizes immunostimulatory lipid signaling to mitigate bacterial infections. Our understanding of immune signaling in the fly and its analogies to mammalian systems will increase the power and value of Drosophila as a model organism in immune studies.

Comprehensive Proteomics Analysis of the Hemolymph Composition of Sugar-Fed Aedes aegypti Female and Male Mosquitoes.

In arthropods, hemolymph carries immune cells and solubilizes and transports nutrients, hormones, and other molecules that are involved in diverse physiological processes including immunity, metabolism, and reproduction. However, despite such physiological importance, little is known about its composition. We applied mass spectrometry-based label-free quantification approaches to study the proteome of hemolymph perfused from sugar-fed female and male Aedes aegypti mosquitoes. A total of 1403 proteins were identified, out of which 447 of them were predicted to be extracellular. In both sexes, almost half of these extracellular proteins were predicted to be involved in defense/immune response, and their relative abundances (based on their intensity-based absolute quantification, iBAQ) were 37.9 and 33.2%, respectively. Interestingly, among them, 102 serine proteases/serine protease-homologues were identified, with almost half of them containing CLIP regulatory domains. Moreover, proteins belonging to families classically described as chemoreceptors, such as odorant-binding proteins (OBPs) and chemosensory proteins (CSPs), were also highly abundant in the hemolymph of both sexes. Our data provide a comprehensive catalogue of A. aegypti hemolymph basal protein content, revealing numerous unexplored targets for future research on mosquito physiology and disease transmission. It also provides a reference for future studies on the effect of blood meal and infection on hemolymph composition.

Pleiotropic immunoregulation by growth-blocking peptide in Ostrinia furnacalis.

Insects rely on their innate immune system to eliminate pathogenic microbes. As a system component, cytokines transmit intercellular signals to control immune responses. Growth-blocking peptide (GBP) is a member of the stress-responsive peptide family of cytokines found in several orders of insects, including Drosophila. However, the physiological role of GBP in defence against pathogens is not thoroughly understood. In this study, we explored the functions of GBP in a lepidopteran pest, Ostrinia furnacalis. Injection of recombinant O. furnacalis GBP (OfGBP) precursor (proGBP) and chemically synthesised GBP significantly induced the transcription of antimicrobial peptides (AMPs) and other immunity-related genes including immune deficiency (IMD) and Dorsal. The level of OfGBP mRNA was upregulated after bacterial infection. Knockdown of OfGBP expression led to a decrease in IMD, Relish, MyD88 and Dorsal mRNA levels. OfGBP induced phenoloxidase activity and affected hemocyte behaviours in O. furnacalis larvae. In summary, GBP is a potent cytokine, effectively regulating AMP synthesis, melanization response and cellular immunity to eliminate invading pathogens.

Differential molecular responses of hemolymph and hepatopancreas of swimming crab, Portunus trituberculatus, infected with Ameson portunus (Microsporidia).

Ameson portunus (Microsporidia) has caused serious economic losses to the aquaculture industry of swimming crab, Portunus trituberculatus. The hemolymph and hepatopancreas are the main immune organs of P. trituberculatus, and the main sites of A. portunus infection. Elucidating the response characteristics of hemolymph and hepatopancreas to microsporidian infection facilitates the development of microsporidiosis prevention and control strategy. This study performed comparative transcriptomic analysis of hemolymph (PTX/PTXA) and hepatopancreas (PTG/PTGA) of P. trituberculatus uninfected and infected with A. portunus. The results showed that there were 223 and 1309 differentially expressed genes (DEGs) in PTX/PTXA and PTG/PTGA, respectively. The lysosome pathway was significantly enriched after the invasion of the hemolymph by A. portunus. Also, immune-related genes were all significantly up-regulated in the hemolymph and hepatopancreas, suggesting that the invasion by A. portunus may activate host immune responses. Unlike hemolymph, antioxidant and detoxification-related genes were also significantly up-regulated in the hepatopancreas. Moreover, metabolism-related genes were significantly down-regulated in the hepatopancreas, suggesting that energy synthesis, resistance to pathogens, and regulation of oxidative stress were suppressed in the hepatopancreas. Hemolymph and hepatopancreas have similarity and tissue specificity to microsporidian infection. The differential genes and pathways identified in this study can provide references for the prevention and control of microsporidiosis.

Complete genome of the new bacilliform virus that causes Milky Hemolymph Syndrome in Chionoecetes bairdi (Rathbun, 1924).

The genome of a new member of the Nimaviridae family has been sequenced. The Chionoecetes bairdi bacilliform virus (CbBV) causes Milky Hemolymph Syndrome (MHS) in Chionoecetes bairdi populations of the Pacific coast of Kamchatka. The CbBV genome is represented by double-stranded DNA with a length of 245,567 nucleotides containing 120 ORFs. Of these, 85 proteins had significant matches in the NCBI database, and 57 genes encoded capsid, envelope, tegument and nonstructural proteins. Comparative analysis of the genomes of CbBV and a number of representatives of the class nuclear arthropod large DNA viruses (NALDVs) made it possible to isolate 49 evolutionarily conserved orthologue core genes. Among them, 5 were multicopy genes, and 44 were single-copy genes. There were ancestral genes characteristic of all Naldaviricetes - per os infectivity complex genes, one DNA polymerase gene and one thymidylate synthase gene. Phylogenetic analysis of representatives of the Nimaviridae family revealed that the CbBV and Chionoecetes opilio bacilliform virus (CoBV) form an independent clade within the family separate from the clade containing WSSV strains. This is supported by data on the order and arrangement of genes in the genomes of nimaviruses that were identical within each clade but differed between them. In addition, a high identity of the genomes and proteomes of CbBV and CoBV (approximately 99%) was shown, and their identity with WSSV strains was no more than 33%. The data on the structure of the genome of the new virus that causes MHS in C. bairdi indicate that it belongs to the family Nimaviridae, genus Whispovirus. Thus, the CbBV infecting the commercially important species of Tanner crab in populations of the Pacific coast of Kamchatka is the second "wild" representative of replicating nimaviruses whose genome has been characterized after the CoBV that causes MHS in C. opilio in populations of the Sea of Japan. The discovery of a new member of the family that infects decapods indicates the prevalence of nimaviruses in marine ecosystems. The information obtained is important for understanding the evolution of representatives of the class of nuclear arthropod large DNA viruses. The discovery of a new nimavirus that causes MHS in Chionoecetes crabs, in contrast to the white spot syndrome (WSS) caused by WSSV strains, makes it relevant to identify two variants and possibly species within the family, namely, WSSV and Milky Hemolymph Syndrome virus (MHSV).

Hemolymph metabolism of black soldier fly (Diptera: Stratiomyidae), response to different supplemental fungi.

The black soldier fly, Hermetia illucens L. (Diptera: Stratiomyidae), is commonly used for organic waste recycling and animal feed production. However, the often inadequate nutrients in organic waste necessitate nutritional enhancement of black soldier fly larvae, e.g., by fungal supplementation of its diet. We investigated the amino acid composition of two fungi, Candida tropicalis (Castell.) Berkhout (Saccharomycetales: Saccharomycetaceae) and Pichia kudriavzevii Boidin, Pignal & Besson (Saccharomycetales: Pichiaceae), from the black soldier fly gut, and commercial baker's yeast, Saccharomyces cerevisiae Meyen ex E.C. Hansen (Saccharomycetales: Saccharomycetaceae), and their effects on larval growth and hemolymph metabolites in fifth-instar black soldier fly larvae. Liquid chromatography-mass spectrometry was used to study the effect of fungal metabolites on black soldier fly larval metabolism. Amino acid analysis revealed significant variation among the fungi. Fungal supplementation led to increased larval body mass and differential metabolite accumulation. The three fungal species caused distinct metabolic changes, with each over-accumulating and down-accumulating various metabolites. We identified significant alteration of histidine metabolism, aminoacyl-tRNA biosynthesis, and glycerophospholipid metabolism in BSF larvae treated with C. tropicalis. Treatment with P. kudriavzevii affected histidine metabolism and citrate cycle metabolites, while both P. kudriavzevii and S. cerevisiae treatments impacted tyrosine metabolism. Treatment with S. cerevisiae resulted in down-accumulation of metabolites related to glycine, serine, and threonine metabolism. This study suggests that adding fungi to the larval diet significantly affects black soldier fly larval metabolomics. Further research is needed to understand how individual amino acids and their metabolites contributed by fungi affect black soldier fly larval physiology, growth, and development, to elucidate the interaction between fungal nutrients and black soldier fly physiology.

Polyester Microfibers Exposure Modulates Mytilus galloprovincialis Hemolymph Microbiome.

Microplastic (MP) contamination in the aquatic environment is a cause of concern worldwide since MP can be taken up by different organisms, altering different biological functions. In particular, evidence is accumulating that MP can affect the relationship between the host and its associated microbial communities (the microbiome), with potentially negative health consequences. Synthetic microfibers (MFs) represent one of the main MPs in the marine environment, which can be accumulated by filter-feeding invertebrates, such as bivalves, with consequent negative effects and transfer through the food chain. In the mussel Mytilus galloprovincialis, polyethylene terephthalate (PET) MFs, with a size distribution resembling that of an MF released from textile washing, have been previously shown to induce multiple stress responses. In this work, in the same experimental conditions, the effects of exposure to PET-MF (96 h, 10, and 100 µg/L) on mussel hemolymph microbiome were evaluated by 16S rRNA gene amplification and sequencing. The results show that PET-MF affects the composition of bacterial communities at the phylum, family and genus level, with stronger effects at the lowest concentration tested. The relationship between MF-induced changes in hemolymph microbial communities and responses observed at the whole organism level are discussed.

ERK/CREB and p38 MAPK/MMP14 Signaling Pathway Influences Spermatogenesis through Regulating the Expression of Junctional Proteins in Eriocheir sinensis Testis.

The hemolymph-testis barrier (HTB) is a reproduction barrier in Crustacea, guaranteeing the safe and smooth process of spermatogenesis, which is similar to the blood-testis barrier (BTB) in mammals. The MAPK signaling pathway plays an essential role in spermatogenesis and maintenance of the BTB. However, only a few studies have focused on the influence of MAPK on crustacean reproduction. In the present study, we knocked down and inhibited MAPK in Eriocheir sinensis. Increased defects in spermatogenesis were observed, concurrently with a damaged HTB. Further research revealed that es-MMP14 functions downstream of ERK and p38 MAPK and degrades junctional proteins (Pinin and ZO-1); es-CREB functions in the ERK cascade as a transcription factor of ZO-1. In addition, when es-MMP14 and es-CREB were deleted, the defects in HTB and spermatogenesis aligned with abnormalities in the MAPK. However, JNK impacts the integrity of the HTB by changing the distribution of intercellular junctions. In summary, the MAPK signaling pathway maintains HTB integrity and spermatogenesis through es-MMP14 and es-CREB, which provides insights into the evolution of gene function during barrier evolution.

Insects as a Prospective Source of Biologically Active Molecules and Pharmaceuticals-Biochemical Properties and Cell Toxicity of Tenebrio molitor and Zophobas morio Cell-Free Larval Hemolymph.

Insects are of great interest as novel sources of alternative proteins and biologically active compounds, primarily anticancer agents. Protein-rich insect larval hemolymph is a prospective candidate for pharmaceutical and food industry-related research. In this study, selected biochemical properties and cell toxicity of larval hemolymph from two mealworm species, Tenebrio molitor and Zophobas morio, were analyzed. Total proteins and carbohydrates, antioxidant capacity, and the level of lipid peroxidation were determined. Human cancer (U-87) and normometabolic (MRC-5) cells were treated with different concentrations of larval hemolymph proteins, and the effects on cell viability were assayed 24, 48, and 72 h after treatments. Z. morio hemolymph was shown to be richer in total proteins, showing a higher antioxidant capacity and lipid peroxidation level than T. molitor hemolymph, which was richer in total carbohydrates. Cytotoxicity assays showed that T. molitor and Z. morio hemolymphs differently affect the viability of U-87 and MRC-5 cells in cell type-, dose-, and time-dependent manners. Hemolymph from both species was more cytotoxic to U-87 cells than to MRC-5 cells, which was particularly prominent after 48 h. Additionally, a more potent cytotoxic effect of Z. morio hemolymph was observed on both cell lines, likely due to its higher antioxidant capacity, compared to T. molitor hemolymph.

Development and Evaluation of a Shrimp Virus (IHHNV)-Mediated Gene Transfer and Expression System for Shrimps.

An efficient gene transfer and expression tool is lacking for shrimps and shrimp cells. To solve this, this study has developed a shrimp DNA virus-mediated gene transfer and expression system, consisting of insect Sf9 cells for viral packaging, the shrimp viral vector of pUC19-IHHNV-PH-GUS and the baculoviral vector of Bacmid or Bacmid-VP28 encoding the shrimp WSSV envelope protein VP28. The pUC19-IHHNV-PH-GUS vector was constructed by assembling the genomic DNA of shrimp infectious hypodermal and hematopoietic necrosis virus (IHHNV), which has shortened inverted terminal repeats, into a pUC19 backbone, and then an expression cassette of baculoviral polyhedron (PH) promoter-driven GUS (ß-glucuronidase) reporter gene was inserted immediately downstream of IHHNV for proof-of-concept. It was found that the viral vector of pUC19-IHHNV-PH-GUS could be successfully packaged into IHHNV-like infective virions in the Sf9 cells, and the gene transfer efficiency of this system was evaluated and verified in three systems of Sf9 cells, shrimp hemolymph cells and tissues of infected shrimps, but the GUS expression could only be detected in cases where the viral vector was co-transfected or co-infected with a baculovirus of Bacmid or Bacmid-VP28 due to the Bacmid-dependence of the PH promoter. Moreover, the packaging and infection efficiencies could be significantly improved when Bacmid-VP28 was used instead of Bacmid.

The circadian clock is required for rhythmic lipid transport in Drosophila in interaction with diet and photic condition.

Modern lifestyle is often at odds with endogenously driven rhythmicity, which can lead to circadian disruption and metabolic syndrome. One signature for circadian disruption is a reduced or altered metabolite cycling in the circulating tissue reflecting the current metabolic status. Drosophila is a well-established model in chronobiology, but day-time dependent variations of transport metabolites in the fly circulation are poorly characterized. Here, we sampled fly hemolymph throughout the day and analyzed diacylglycerols (DGs), phosphoethanolamines (PEs) and phosphocholines (PCs) using LC-MS. In wild-type flies kept on sugar-only medium under a light-dark cycle, all transport lipid species showed a synchronized bimodal oscillation pattern with maxima at the beginning and end of the light phase which were impaired in period01 clock mutants. In wild-type flies under constant dark conditions, the oscillation became monophasic with a maximum in the middle of the subjective day. In strong support of clock-driven oscillations, levels of the targeted lipids peaked once in the middle of the light phase under time-restricted feeding independent of the time of food intake. When wild-type flies were reared on full standard medium, the rhythmic alterations of hemolymph lipid levels were greatly attenuated. Our data suggest that the circadian clock aligns daily oscillations of DGs, PEs, and PCs in the hemolymph to the anabolic siesta phase, with a strong influence of light on phase and modality.

Proteomics and Immune Response Differences in Apis mellifera and Apis cerana Inoculated with Three Nosema ceranae Isolates.

Nosema ceranae infects midgut epithelial cells of the Apis species and has jumped from its original host A. cerana to A. mellifera worldwide, raising questions about the response of the new host. We compared the responses of these two species to N. ceranae isolates from A. cerana, A. mellifera from Thailand and A. mellifera from France. Proteomics and transcriptomics results were combined to better understand the impact on the immunity of the two species. This is the first combination of omics analyses to evaluate the impact of N. ceranae spores from different origins and provides new insights into the differential immune responses in honeybees inoculated with N. ceranae from original A. cerana. No difference in the antimicrobial peptides (AMPs) was observed in A. mellifera, whereas these peptides were altered in A. cerana compared to controls. Inoculation of A. mellifera or A. cerana with N. ceranae upregulated AMP genes and cellular-mediated immune genes but did not significantly alter apoptosis-related gene expression. A. cerana showed a stronger immune response than A. mellifera after inoculation with different N. ceranae isolates. N. ceranae from A. cerana had a strong negative impact on the health of A. mellifera and A. cerana compared to other Nosema isolates.

Segmental pairs of dermal secretory cells release proteins into the hemolymph at the larval-pupal molt.

At each molt of Manduca, the large dermal secretory cells expel the protein contents of their vacuoles into the hemocoel. The constellation of proteins expelled at the last larval-pupal molt, however, differs qualitatively from those proteins released at earlier larval-larval molts. Secretory cells at the two stages not only have different lectin staining properties but also have different proteins that separate on two-dimensional gels. Numerous physiological changes accompany the termination of the last larval instar, including increased chitin synthesis, diminished oxygen delivery, and reduced humoral immunity. Secretion of trehalase that is essential for chitin synthesis and the release of hypoxia up-regulated protein to ameliorate oxygen deprivation help ensure normal transition from larva to pupa. Proteins released by dermal secretory cells at this last molt could supplement the diminished immune defenses mediated by fat body and hemocytes at the end of larval life. Additional immune defenses provided by dermal secretory cells could help ensure a safe transition during a period of increased vulnerability for the newly molted pupa with its soft, thin cuticle and reduced mobility.

Hemolymph Node - An Immunomorphlogical Organ: Modeling the Hemolymph Node by Allografting Renal Tissue in the Rat.

There is no authoritative characterization of the attributes of the hemolymph node (HLN) since Gibbes' first description in 1884. Early reports showed that HLN are found near the kidney in human and animals with the feature of numerous erythrocytes in sinuses. Subsequent studies mainly focused on anatomy and histology, such as the source, distribution, and quantity of erythrocytes in sinuses. Recent articles mentioned that the emergence of HLN was related to immunity, but there was no strong evidence to support this hypothesis. Therefore, it is still uncertain whether the HLN is an organ of anatomy, histology, or immunology. It has been found that the development of HLN could be elicited in the parathymic area by stimuli such as Escherichia coli, allogeneic breast cancer cells, and renal tissue that were injected/transplanted into the tail of rats in our pilot studies. In this study, the model of the HLN was established by transferring allogeneic renal tissue in the rat. Intrasinusoidal erythrocytes of the node were the component for producing a red macroscopic appearance, while macrophage-erythrocyte-lymphocyte rosettes were the major immunomorphological changes, reflecting the immune activity against the invasion of the allogeneic tissue within the node. Therefore, the HLN is an immunomorphological organ.

Purification of phenoloxidase from Haliotis discus hannai and its anti-inflammatory activity in vitro.

Haliotis discus hannai, a food with a high protein content, is widely consumed in Asian countries. It is known to have antioxidant, anticancer, and antibacterial effects. Since the biological significance of H. discus hannai hemolymph has not been widely studied, the objective of the present study was to purify phenoloxidase (PO) and investigate its immunological effects on human colonic epithelial cells. PO was purified through ammonium sulfate precipitation and one step column chromatography. The molecular weight of the protein was about 270 kDa. When PO was mixed with Gram-negative bacteria-derived lipopolysaccharide (LPS) at various ratios (10:1-1:10, w/w), the amount of residual LPS was reduced. PO at concentrations up to 200 µg/mL was not cytotoxic to HT-29 cells. The inflammatory response induced by LPS in HT-29 cells was regulated when the concentration of PO was increased. With increasing concentration of PO, production levels of pro-inflammatory cytokines, cytokines associated with hyperimmune responses such as IL4, IL-5, and INF-γ, and prostaglandin 2 (PGE2) were regulated. It was thought that simultaneous treatment with PO and LPS anti-inflammatory effects in HT-29 cells showed by regulating the ERK1/2-mediated NF-κB pathway. Results of this study suggest that H. discus hannai hemolymph is involved in the regulation of Gram-negative bacteria-related inflammatory immune responses in human colonic epithelial cells.

Digestive enzyme activity and macromolecule content in the hemolymph of differentially adapted Lymantria dispar L. populations after short-term increases in ambient temperature.

Global, unpredictable temperature increases have strong effects on all organisms, especially insects. Elucidating the effects of short-term temperature increases on midgut digestive enzymes (α-glucosidase, lipase, trypsin, and leucine aminopeptidase - LAP) and metabolic macromolecules in the hemolymph (proteins, lipids, and trehalose) of phytophagous pest larvae of Lymantria dispar is important for general considerations of insect adaptation to a warming climate and potential pest control options. We also wanted to determine whether the different adaptations of L. dispar populations to environmental pollution might affect their ability to cope with heat stress using larvae from the undisturbed, Kosmaj forest and disturbed, Lipovica forest. Heat treatments at 28 °C increased α-glucosidase activity in both larval populations, inhibited LAP activity in larvae from the polluted forest, and had no significant effect on trypsin and lipase activities, regardless of larval origin. The concentration of proteins, lipids, and trehalose in the hemolymph of larvae from the disturbed forest increased, whereas the population from the undisturbed forest showed only an increase in proteins and lipids after the heat treatments. Larval mass was also increased in larvae from the undisturbed forest. Our results suggest a higher sensitivity of digestive enzymes and metabolism to short-term heat stress in L. dispar populations adapted to pollution in their forest habitat, although climate warming is not beneficial even for populations from unpolluted forests. The digestive and metabolic processes of L. dispar larvae are substantially affected by sublethal short-term increases in ambient temperature.

Proton gradient mediates hemolymph trehalose influx into aphid bacteriocytes.

Aphids harbor proteobacterial endosymbionts such as Buchnera aphidicola housed in specialized bacteriocytes derived from host cells. The endosymbiont Buchnera supplies essential amino acids such as arginine to the host cells and, in turn, obtains sugars needed for its survival from the hemolymph. The mechanism of sugar supply in aphid bacteriocytes has been rarely studied. It also remains unclear how Buchnera acquires its carbon source. The hemolymph sugars in Acyrthosiphon pisum are composed of the disaccharide trehalose containing two glucose molecules. Here, we report for the first time that trehalose is transported and used as a potential carbon source by Buchnera across the bacteriocyte plasma membrane via trehalose transporters. The current study characterized the bacteriocyte trehalose transporter Ap_ST11 (LOC100159441) using the Xenopus oocyte expression system. The Ap_ST11 transporter was found to be proton-dependent with a Km value ≥700 mM. We re-examined the hemolymph trehalose at 217.8 mM using a fluorescent trehalose sensor. The bacteriocytes did not obtain trehalose by facilitated diffusion along the gradient across cellular membranes. These findings suggest that trehalose influx into the bacteriocytes depends on the extracellular proton-driven secondary electrochemical transporter.

Prevalence and effects of a parasitic trematode on the blue mussel, Mytilus edulis, in the Boston Harbor.

The blue mussel, Mytilus edulis, is a keystone species in the North Atlantic that plays critical roles in nutrient cycling, water filtration, and habitat creation. Blue mussel populations have declined significantly throughout the North Atlantic due to various factors, including habitat loss, pollution, increasing water temperature, and parasites. One parasite is Proctoeces maculatus, a digenetic trematode, which uses M. edulis as an intermediate host. This parasite causes reduced growth, castration, and death in mussels. The range of P. maculatus has expanded northward from Cape Cod, MA to Maine which may be associated with rising temperatures in the Gulf of Maine. To evaluate the negative impacts of P. maculatus on mussels, we analyzed its infections in M. edulis throughout the Boston Harbor, MA. P. maculatus was present in every population and time point analyzed, with approximately 50% of mussels in the harbor infected. The parasite reduced gonadal development in infected mussels, which could lead to decreased fecundity. Severe P. maculatus infections induced a stress response, indicated by increased HSP70 expression. We developed a non-destructive hemolymph-based assay to determine if mussels are infected with P. maculatus, thus speeding up the evaluation process and eliminating the need to sacrifice individuals. With P. maculatus' continued expansion northward, more mussel populations will be under threat from this parasite.

Milky hemolymph syndrome (MHS) associated with a virus in tanner crab Chionoecetes bairdi off the Pacific coast of Kamchatka.

Milky hemolymph syndrome (MHS) caused by a bacilliform virus (BV) was found in tanner crab Chionoecetes bairdi in the shelf zone of Kamchatka. The prevalence of the disease varied from 0.18 to 1.02%. A total lesion of the cells of the interstitial connective tissue and the connective tissue component of all internal organs was noted, which was expressed in the hypertrophy of their nuclei. In addition, hypertrophy of fixed phagocytes and circulating hemocytes was noted. Ultrastructural analysis of the tissues confirmed that in the interstitial connective tissue of pathologically altered organs, virus particles of two morphotypes were found - rod-shaped and globular. In the cytoplasm of infected cells, bands of microtubules formed near where viral particles were concentrated. In the area of contacts at the poles of microtubules, successive stages of the transformation of rod-shaped viruses into globular viruses was observed. The bacilliform virus that infects C. bairdi is structurally very similar to CoBV found in Chionoecetes opilio. Structural features are characteristic of representatives of fam. Nimaviridae. The molecular data obtained suggest that the virus causing MHS in C. bairdi is systematically very close, if not identical, to CoBV.

Molecular Cloning and Characteristics of a Lectin from the Bivalve Glycymeris yessoensis.

C-type lectins (CTLs) are a family of carbohydrate-binding proteins that mediate multiple biological events, including adhesion between cells, the turnover of serum glycoproteins, and innate immune system reactions to prospective invaders. Here, we describe the cDNA cloning of lectin from the bivalve Glycymeris yessoensis (GYL), which encodes 161 amino acids and the C-type carbohydrate recognition domain (CRD) with EPN and WND motifs. The deduced amino acid sequence showed similarity to other CTLs. GYL is a glycoprotein containing two N-glycosylation sites per subunit. N-glycans are made up of xylose, mannose, D-glucosamine, 3-O-methylated galactose, D-quinovoses, and 3-O-methylated 6-deoxy-D-glucose. The potential CRD tertiary structure of the GYL adopted CTL-typical long-form double-loop structure and included three disulfide bridges at the bases of the loops. Additionally, when confirming the GYL sequence, eight isoforms of this lectin were identified. This fact indicates the presence of a multigene family of GYL-like C-type lectins in the bivalve G. yessoensis. Using the glycan microarray approach, natural carbohydrate ligands were established, and the glycotope for GYL was reconstructed as "Galß1-4GlcNAcß obligatory containing an additional fragment", like a sulfate group or a methyl group of fucose or N-acetylgalactosamine residues.