Crayfish Cherax Quadricarinatus Research Articles

The Optimum Lipid Level for the Juvenile Redclaw Crayfish Cherax quadricarinatus: Practical Diets with Soybean Oil as the Lipid Source.

As a new species in aquaculture, the lipid nutrition requirement for the juvenile redclaw crayfish Cherax quadricarinatus on a dietary basis on a practical formula needs to be evaluated accurately. In this study, the optimum dietary lipid level was explained by analyzing the growth performance, antioxidant state, lipid metabolism, and gut microbiota of C. quadricarinatus after an eight-week cultivation trial. Six diets with different soybean oil levels (named L0, L2, L4, L6, L8, and L10) were fed to C. quadricarinatus (11.39 ± 0.28 g). The results indicated that the specific growth rate and weight gain of crayfish fed the L4 and L6 diets were significantly higher than those of the other groups (P < 0.05). By the analysis of a second-order polynomial regression model according to growth performance (weight gain rate), the optimum lipid level in a practical diet for juvenile C. quadricarinatus was 9.67%. The survival, condition factor, and hepatosomatic index of crayfish were not significantly affected by dietary oil levels (P > 0.05). As the level of dietary lipids increased, the total antioxidant capacity and glutathione peroxidase activity in serum showed a tendency to rise and then fall and the enzyme activity was highest in crayfish fed the L6 diet. Gut lipase and pepsin activities showed the highest value in crayfish fed the L6 diet. There was no significant difference in acetyl-CoA carboxylase and carnitine palmitoyltransferase-1 contents in crayfish among all the groups (P > 0.05). The relative abundance of Proteobacteria in the phylum and Citrobacter in the genus showed a significant decrease in crayfish of the L10 diet, while the relative abundance of Firmicutes in the phylum markedly increased compared to that of the other groups (P < 0.05). In summary, the results indicated that the 10.39% (L6 diet) dietary lipid level could induce better growth performance, antioxidant ability, and digestive enzyme activity. Most of the fatty acid composition of muscle is not closely related to the fatty acid composition of the diet. Moreover, the composition and diversity of the gut microbiota of C. quadricarinatus were changed by high dietary lipid levels.

Differences in Intestinal Microbial Composition between Red Claw Crayfish (Cherax quadricarinatus) and Red Swamp Crayfish (Procambarus clarkii) Cultured in Pond

Intestinal microbiota communities participate in several metabolic processes in the host, and are highly correlated to digestion, nutrition, growth, and immunity. However, the intestinal microbiota of aquatic invertebrates is poorly understood, especially in freshwater crayfish. In this study, the intestinal microbiota of two important freshwater economic aquaculture species, the invasive species, the red swamp crayfish (Procambarus clarkii, Pc), and the introduced species, the red claw crayfish (Cherax quadricarinatus, Cq), were investigated. The results showed that the community richness and diversity of Pc were higher than those of Cq, which might be one of the reasons that Pc have stronger environmental adaptability than Cq. Five core phyla were identified in the Pc group, including Proteobacteria (26.92%), Tenericutes (25.73%), Bacteroidetes (25.12%), Firmicutes (14.03%), and RsaHF231 (8.02%), and three phyla were detected in the Cq group, including Tenericutes (67.35%), Proteobacteria (25.98%), and Firmicutes (4.69%). In Pc and Cq groups, Proteobacteria exhibited significant differential abundance between males and females. In addition, Vibrio coralliilyticus were found particularly in the intestine of Cq. This study provides information on intestinal microbiota differences of Pc and Cq, contributing to the development of new dietary formulations and providing significance to future aquaculture.

EDTA-Na&lt;sub&gt;2&lt;/sub&gt; - anticoagulant for hemolymph of australian red claw crayfish (cherax quadricarinatus)

Purpose: selection of anticoagulants based on EDTA-Na2 and methods of their use for working with the hemolymph of the Australian red claw crayfish (C. quadricarinatus)Materials and methods. For the study, different-sized males and females of the australian red-clawed crayfish (Cherax quadricarinatus (Von Martens, 1868)) were used. EDTA-Na2 was used as an anticoagulant, the concentrationof which was 4%. A 2 ml syringe with a 23G needle for hemolymph removal was pre-washed with anticoagulant remaining in the needle and the needle cone (about 1/3 of the volume of the needle cone was filled with a solution). The experiment is presented in the following series: 1. differences between the total hemocytes number (THC) and the proportion of granulocytes in native hemolymph and treated with a small amount of EDTA-Na2; 2. differences in the content of total hemolymph protein (THP) in native and treated with a small amount of EDTA-Na2 hemolymph; 3. changes in THC and the proportion of granulocytes in hemolymph treated with a small amount the amount of EDTA-Na2 immediately after sampling and a day later. To carry out the work, samples of 20 individuals were used, while for the first two series, two samples of hemolymph were taken from each, and one from the third. Syringes with hemolymph treated with anticoagulant were stored in the refrigerator at a temperature of 8.5 °C. Additionally, similar studies were carried out on differences in the THC and proportion of granulocytes in the hemolymph immediately after sampling with treatment with a multicomponent anticoagulant consisting of 4 g of EDTA-Na2, 3 g of sodium citrate, 2 g of glucose and 1 g of NaCl per 100 ml of distilled water. Hemolymph was taken with a syringe from the ventral sinus. Three indicators were determined: the total number of hemocytes (THC), the proportion of granulocytes and the total protein content (THC). THC and proportion of granulocytes was determined in the Goryaev chamber under a light microscope. THP was determined by the refractometric method.Results. The study revealed significant differences in the proportion of granulocytes (p&lt;0.05), which are 32% more in the anticoagulant treated than in the native hemolymph, which can be explained by the uniform distribution of all types of hemocytes in the sample. There were no significant differences in the remaining indicators of all series. Studies of hemolymph treated with a multicomponent anticoagulant showed a low level of hemocyte preservation (p&lt;0.05) and the proportion of granulocytes compared to the indicators immediately after selection, therefore, they are not reflected in the work. Conclusion. The data obtained indicate that there is a possibility of using four percent EDTA-Na2 when working with hemolymph. The anticoagulant prevents the formation of gel and the rapid destruction of hemocytes, promotes the uniform distribution of cells in the Goryaev chamber and allows the use of a refractometer in determining the total blood protein. The use of an anticoagulant contributes to the preservation of hemocytes during the day, at a level that allows the use of EDTA-Na2 in practice. At the same time, work should continue on the development of methods for storing and transporting blood treated with an anticoagulant.

A Deacetylase CqSIRT1 Promotes WSSV Infection by Binding to Viral Envelope Proteins in Cherax quadricarinatus.

Sirtuin 1 (SIRT1), a member of the class III lysine deacetylases, exhibits powerful functional diversity in physiological processes and disease occurrences. However, the potential molecular mechanism underlying the role of SIRT1 during viral infection in crustaceans is poorly understood. Herein, SIRT1 was functionally characterized from the red claw crayfish Cherax quadricarinatus, which possesses typically conserved deacetylase domains and strong evolutionary relationships across various species. Moreover, gene knockdown of CqSIRT1 in crayfish haematopoietic tissue (Hpt) cell culture inhibited white spot syndrome virus (WSSV) late envelope gene vp28 transcription. In contrast, enhancement of deacetylase activity using a pharmacological activator promoted the replication of WSSV. Mechanically, CqSIRT1 was co-localized with viral envelope protein VP28 in the nuclei of Hpt cells and directly bound to VP28 with protein pulldown and co-immunoprecipitation assays. Furthermore, CqSIRT1 also interacted with another two viral envelope proteins, VP24 and VP26. To the best of our knowledge, this is the first report that WSSV structural proteins are linked to lysine deacetylases, providing a better understanding of the role of CqSIRT1 during WSSV infection and novel insights into the basic mechanism underlying the function of lysine deacetylases in crustaceans.

Estimates of heritability and genetic correlation for growth traits at harvest in redclaw crayfish, Cherax quadricarinatus

The current study assessed genetic parameters of three growth traits in redclaw crayfish Cherax quadricarinatus, including body weight (BW), body length (BL) and carapace length (CL), and explored how the number of families affected the heritability estimates of these traits. A total of 350 families were produced, and seven family levels containing 50, 100, 150, 200, 250, 300 and 350 families respectively were designed. After a 170 d communally rearing period for 350 families, males had significantly higher BW and BL averages (P < 0.001), but significantly lower CL average than females (P < 0.001). Heritability estimates of BW, BL and CL based on 350 families were 0.138 ± 0.051, 0.123 ± 0.048 and 0.073 ± 0.035, with common environmental effects of 0.049 ± 0.024, 0.053 ± 0.023 and 0.039 ± 0.017, respectively. There were strong positive genetic correlations (0.863 ± 0.089–0.970 ± 0.028) between these growth traits. Among different family levels, heritability estimates ranged from 0.030 ± 0.090 to 0.138 ± 0.051, 0.010 ± 0.091 to 0.123 ± 0.048, and 0 to 0.073 ± 0.035, and common environmental effects ranged from 0.045 ± 0.101 to 0.092 ± 0.046, 0.053 ± 0.023 to 0.100 ± 0.047, and 0.038 ± 0.043 to 0.062 ± 0.033, for BW, BL and CL, respectively. Only heritability estimates of BW and BL based on 300 and 350 families, and that of CL based on 350 families, were significantly different from zero (P < 0.05), which suggested that larger numbers of families could contribute to more accurate estimation of heritability. The results indicate that there exists some genetic variation for our selection population to improve growth in C. quadricarinatus. However, the future program for redclaw crayfish needs to include crossing with outside stocks to replenish genetic variation of selection population.

Exploration of the functional properties of red cuticle patch of redclaw crayfish (Cherax quadricarinatus) by histological and transcriptomic analysis

The redclaw crayfish (Cherax quadricarinatus) is being farmed on an increasing scale in various tropical and subtropical regions. The most distinctive feature of the adult male redclaw crayfish is the red cuticle patch on the lateral cuticle of the chelae. It is an important indicator of the health status, reproduction, and resistance to predators of this species, but little is known about its functional properties. In this study, we used redclaw crayfish (45.35 ± 16.74 g) as experimental subjects. Histological comparison of the red cuticle patch on male redclaw crayfish chelae with the cuticle on female redclaw crayfish chelae revealed that the red cuticle patch lacked exocuticle. High performance liquid chromatography analysis demonstrated that the concentration of free astaxanthin in the red cuticle patch was much higher than that in the chelae of female redclaw crayfish. Further analysis of transcriptome data from epithelial tissue of the chelae revealed that genes related to chela structure were highly expressed in the red cuticle patch of males. Immunity-related genes were not specifically expressed in the red cuticle patch, and immune- and growth-related genes that require calcium ion for activation were down-regulated in the patch. Our results indicated that regulation of genes related to astaxanthin binding was not a single up- or down-regulation mode. This study demonstrated structural differences in the chela cuticle between male and female redclaw crayfish and revealed the molecular basis for the functional properties of the red cuticle patch.

Impact of low-cost diets on maturation of the red claw crayfish Cherax quadricarinatus: An integrative approach during a long-term study

The onset of sexual maturity represents a complex step in animal life involving morphological, physiological, and behavioral changes. From this point onward, the energy income from food will be involved in three energetic-demanding-competing processes: metabolism-homeostasis, growth, and reproduction. Hence, in cultured species formulated feeds need to cover all nutritional requirements of the growing phase and the additional cost demanded by reproduction. Two low-cost diets, AD1: 0.56 and 2 CE: 0.64 U$S kg−1 manufactured with local supplies specifically formulated for Cherax quadricarinatus, were evaluated in a prolonged assay with females at the transition from juvenile to mature. Protein content was 440,2 and 380 g kg−1 and lipid content was 134,8 and 114,9 g kg−1 for AD1 and AD2, respectively. Females were obtained in the laboratory from our reproductive stock and were fed daily at 1.5% of their weight/day. The female mean initial total length was 6.5 ± 0.6 cm and mean body mass was 6.8 ± 0.7 g, which represents juveniles at the beginning of vitellogenesis. The experiment ended after three molting events when females achieved the full mature condition (⁓16–20 g). Somatic indices, growth, feed efficiency, biochemical composition of the ovary and hepatopancreas, ovarian growth, and histological alterations in hepatopancreas were analyzed. Both diets demonstrated that the high demands of both somatic and ovarian growth in maturating females were not covered, contrasting with previous results in early juveniles. This highlights the enormous energetic cost of vitellogenesis at the start of sexual maturity and the differential nutrients demand during crayfish growth. Biochemical analysis demonstrated a lower oocyte quality due to the experimental diets, as well as decreased reserves both of lipids and carbohydrates and a possible impact on carotenoids content. The hepatopancreas also showed partial histological damage and differences in its biochemical content. Although further analysis is required, lower embryo survival, hatching rate, and early performance of juveniles are expected in progeny from females fed with both diets. The low-cost diets tested are useful as a baseline from which modified formulations can be considered to achieve a more adequate composition enhancing somatic and gonad growth as well as feed efficiency in maturing females.

Redclaw crayfish (Cherax quadricarinatus) responds to Vibrio parahaemolyticus infection by activating toll and immune deficiency signaling pathways and transcription of associated immune response genes

In invertebrates, several genes controlled by the Toll and immunodeficiency (IMD) signaling cascade are altered during microbial infection. However, little is known about the expression patterns of innate immune response genes in red-clawed crayfish (Cherax quadricarinatus). In the present study, the transcription of five genes was assessed in C. quadricarinatus challenged with Vibrio parahaemolyticus (V. parahaemolyticus). The expression of Relish, Toll-like receptor (TLR), tumor necrosis factor receptor-related factor 6 (TRAF6), Akirin, and IMD in different tissues and at different time points after infection were assessed. In addition, the Relish gene was amplified, the protein conformation of the Relish gene was predicted, and gene expression changes associated with antimicrobial peptide production in C. quadricarinatus were analyzed using RNA interference (RNAi). During V. parahaemolyticus infection, the transcripts of the above five genes were significantly increased in the hepatopancreas of C. quadricarinatus (P < 0.05). In contrast, TLR was significantly downregulated in muscle tissue at the initial stage of infection (P < 0.05); TRAF6 and IMD were significantly down-regulated throughout infection (P < 0.05); Akirin transcripts had the lowest abundance at 24 h post-infection; Relish, IMD and Akirin genes were significantly up-regulated in gill tissue at the early stage of infection (P < 0.05); only TRAF6 was significantly up-regulated at 6, 24 and 48 h after infection. The Relish gene of C. quadricarinatus is closely related to the Exopalaemon carinicauda. When the Relish gene was knocked down by RNAi, the V. parahaemolyticus challenge showed that the mortality rate of the RANi group was significantly higher than that of the NC group; pathological sections showed that the hepatopancreatic tissue damage was the most severe 12 h after the interference; and the interference significantly inhibited IRF4, NF-κB, ALF, laccase, SOD1, and lectin genes. Therefore, it can be hypothesized that the Toll and IMD pathways are activated in C. quadricarinatus in response to bacterial infection and that genes associated with these pathways are differentially transcribed in different tissues. This study provides insights into the Toll and IMD signaling pathways and the spatiotemporal expression of key genes regulating bacterial infection resistance in C. quadricarinatus.

Nanocalcium of Pila ampullacea Shell incorporated into Feed on Molting and Growth Performance of Crayfish Cherax quadricarinatus

Freshwater snail Pila ampullacea shell can be used as alternative for calcium source. The shell has been known as solid waste and it also has been a major problem in the agriculture field. The present study aims to determine the effect of nanocalcium made from P.ampullacea shell to improve the molting and growth performance of red claw crayfish Cherax quadricarinatus. A laboratory study was conducted for four weeks of feeding trials using 120 freshwater crayfish C. quadricarinatus. A total of four different levels of nanocalcium regimes including 0%, 2%, 4%, and 6% were applied for experimental treatments and each treatment regime consisted of three replicates. Data was collected on molting frequency, absolute weight growth, specific growth rate, survival rate, food conversion ratio, and water quality parameters. The results suggests that dietary nanocalcium treatments were significantly different in absolute weight growth, specific growth rate, and molting frequency, survival rate. However, the nanocalcium of P.ampullacea shell showed no significant difference in feed conversion ratio. The highest absolute weight growth, specific growth rate, molting frequency, survival rate was obtained from 2% of dietary P.ampullacea nanocalcium.

Efficiency assessment of the use of compound feeds with protein different levels for juveniles Australian red claw crayfish in RAS conditions

The aim of the work is to investigate the effect of compound feeds with different levels of protein on the growth of juveniles of the Australian red claw crayfish Cherax quadricarinatus. Used methods: To solve this problem, two experiments were carried out in the aquariums of aquaculture invertebrate department at VNIRO: the first was on juveniles weighing 0.017 ± 0.004 g and 0.87 ± 0.08 mm long; the second one was on juveniles weighing 0.31 ± 0.14 g and 23.7 ± 3.9 mm long. The duration of the experiments was 30 and 40 days, respectively. Feeding of juveniles was carried out with three variants of compound feed recipes with an estimated protein content of 25 %, 35 % and 45 %. The main part of the formulations consisted of components of plant origin. Tetra Wafer Mix food for fish and crustaceans (Germany) with a protein content of 45 % was used as a control. The duration of the experiments was 30 and 40 days, respectively. Feeding of juveniles was carried out with three variants of compound feed recipes with an estimated protein content of 25 %, 35 % and 45 %. The main part of the formulations consisted of components of plant origin. Tetra Wafer Mix food for fish and crustaceans (Germany) with a protein content of 45 % was used as a control. Newness: Development of a feed formulation for rearing juveniles of the Australian red claw crayfish in recirculating aquaculture systems (RAS). Results: These experiments showed that the growth rate of juvenile Australian red claw crayfish is directly dependent on the proportion of protein in the feed. It was established that the relative content of protein in the muscles of crayfish depends on its content in compound feed. An analysis of our own and published data indicates that feeds with protein content below 35 % cannot be recommended for growing of juveniles’ Australian red claw crayfish. Feeds with a protein content of 45 % are optimal for the growth of juveniles. It was established that the effectiveness of compound feeds depends not only on the protein content and energy value, but also on their taste attractiveness. Practical significance: The obtained results will be used in the development of mixed feed for juveniles Australian red claw crayfish.

Cloning and expression of the thioredoxin gene in Cherax quadricarinatus (Von Martens, 1868) (Decapoda: Astacidea: Parastacidae) under bacterial stress

Abstract As a member of the thioredoxin (Trx) system, the Trx1 gene plays essential roles in the pathogenesis of various diseases. The function of Trx in bacterial infections in the freshwater crayfish Cherax quadricarinatus (Von Martens, 1868), however, has not been clarified. We cloned thioredoxin 1 (CqTrx1) from C. quadrangularis, and its expression was investigated. The results showed that the coding sequence of the CqTrx1 gene was 318 bp, encoding a polypeptide of 105 amino acids, contains the conserved motif CGPC (Cys-Gly-Pro-Cys), and was 75.24% and 66.67% identical to sequences of the shrimps Macrobrachium rosenbergii (De Man, 1879) and Fenneropenaeus chinensis (Osbeck, 1765), respectively. The CqTrx1 transcript existed in all organs tested, with the highest expression level in the intestinal tract and the lowest expression level in the gonads. Under the stress of Vibrio algolyticus, V. parahaemolyticus, or Aeromonas hydrophila, the expression level of CqTrx in the gills was 3.1, 4, and 7 times that in the control group at 3, 9, and 24 h, respectively, indicating that CqTrx1 plays an essential role in shrimp antibacterial immunity.

Growth performance, physiological and antioxidant capacity responses to dietary fish meal replacement with insect meals for aquaculture: A case study in red claw crayfish ( Cherax quadricarinatus )

A 60-day feeding trial was conducted to evaluate the effects of yellow mealworm (YM) (Tenebrio molitor) (9%, 18%, 27% and 36%) and black soldier fly larvae (BSFL) (Hermetia illucens) (7%, 14%, 21% and 28%) meal replacing fish meal (FM) on growth performance and antioxidant capacity of red claw crayfish (Cherax quadricarinatus). Nine diets, including control, were fed with quadruplicate of 20 crayfish (0.20 ± 0.05 g). The highest weight gain (WG) and lowest feed conversion ratio were observed in crayfish fed YM18 and BSFL14. The highest superoxide dismutase activity and the lowest malondialdehyde content were detected in the hepatopancreas of the crayfish fed YM18 and BSFL14. Compared with the control group, crayfish fed BSFL14 had higher levels of glutathione peroxidase, catalase acid and alkaline phosphatase and the lowest malondialdehyde (MDA) content in hemocytes. The mRNA expression of CqBAF, HSP70 and MnSOD in the hepatopancreas of crayfish showed an increasing trend. Based on the second-order polynomial regression analysis of WG, the optimal dietary replacement level of FM with YM meal and BSFL was determined to be 21.9% and 17.1% for juvenile C. quadricarinatus.

Improving the technology of growing crustaceans the example of redclaw crayfish Cherax quadricarinatus

The urgency of research with crustaceans in aquaculture is due to the significant demand for the consumption of delicacies of aquatic organisms, including redclaw (Cherax quadricarinatus). Production of freshwater crustaceans provides high profits. Significant volumes of cultivation of these objects on an industrial scale will contribute to the development of aquaculture as a sector of the economy of Ukraine. The scientific novelty is that for the first time in the aquarium-pool complex of a higher education institution a study was conducted on the artificial cultivation and maintenance of australian redclaw crayfish (Cherax quadricarinatus) with the improvement of the technological scheme of cultivation. For the first time in the technological scheme of cultivation, a shelter for crustaceans was used, where the main elements are separator plates and polymer pipes. The proposed design for sheltering crustaceans reduces their death during cultivation, improves the overall functional state of the body after changing the chitinous cover (molting), prevents cannibalism and minimizes stress in intensive reproduction and cultivation technology, optimizes the maintenance of the artificial shelter. The experiment involved testing the technological process of growing crustaceans using structures for their shelter, studying the growth rate and monitoring the hydrochemical regime. The practical significance of the study will help to improve the technology of artificial cultivation of australian redclaw with the prevention of cannibalism.

A comprehensive analysis based on histopathology, metabolomics and transcriptomics reveals molecular regulatory mechanisms of the red claw crayfish (Cherax quadricarinatus) in response to acute hepatopancreatic necrosis disease (AHPND)

The maintenance of homeostasis is central for the survival of organisms under exposure to various types of stress, toxicants, and pathogens, and small metabolites often play key roles in this process. Vibrio parahaemolyticus is one of the main pathogens causing acute hepatopancreatic necrosis disease in crustaceans. Here, we characterized changes in the histopathology of the hepatopancreas, gene expression, and the metabolism of small molecules in Cherax quadreicarinatus challenged with V. parahaemolyticus. Susceptible crayfish were characterized by hemorrhage in the hepatopancreatic interstitium, irregular contraction of the stellate structures in the lumen, the partial loss of luminal structures, and increased vacuolization; asymptomatic crayfish exhibited a small amount of hemorrhage, partial contraction of the stellate structures, and no significant increase in the number of vacuoles. V. parahaemolyticus challenge also resulted in significant differences in the accumulation of metabolites such as adenosine triphosphate, adenosine diphosphate, inosine triphosphate, glycodeoxycholic acid (hydrate), phosphatidylcholine, deoxyguanosine, arginine, and adenosine. Transcriptome analysis revealed a total of 3678 differentially expressed genes that were mainly enriched in the following Kyoto Encyclopedia of Genes and Genomes pathways: phagosome, starch and sucrose metabolism, apoptosis, cysteine and methionine metabolism, Vibrio cholerae infection, oxidative phosphorylation, Toll and IMD signaling pathways, arginine biosynthesis, and other signaling pathways. A comprehensive analysis showed that Lysosomal, citric acid cycle (TCA cycle), vitamin digestion and absorption, bile secretion, glycolysis/gluconeogenesis, oxidative phosphorylation, tryptophan metabolism, pyrimidine metabolism, purine metabolism, pyruvate metabolism, ascorbate and aldehyde metabolism, NOD-like receptor signaling pathway, AMPK signaling pathway, PI3K-Akt signaling pathway, cAMP signaling pathway, calcium signaling pathway, and other signaling pathways were also significantly enriched under V. parahaemolyticus challenge. Our results indicate that C. quadricarinatus maintains its homeostasis under challenge with V. parahaemolyticus by activating multiple signaling pathways that mediate the repair of hepatopancreatic damage.

Prevalence of temnocephalids on cultured and wild Cherax quadricarinatus in Malaysia

The introduction of the crayfish Cherax quadricarinatus into new areas of Malaysia resulted in the introduction of new parasites that harmed other aquatic organisms. A study was conducted to determine the prevalence and sites specificity of temnocephalids in wild and cultured Cherax quadricarinatus. In this study, 33 wild (Melaka) and 32 cultured (Johor) C. quadricarinatus were sampled in January 2020. The prevalence of the parasite was determined using dissecting microscopes and their sites specificity were compared. The prevalence (P) and mean intensity (MI) of temnocephalids in wild crayfish were 93.94% and 35.13%, whereas in the cultured crayfish were 100% and 18.31%, respectively. The temnocephalid species were identified as Diceratocephala sp., Craspedella sp., and Decadidymus sp. The prevalence of adults and eggs of Decadidymus sp. was found to be significant in cultured C. quadricarinatus (p < 0.05). Significant differences were observed by adult Diceratocephala sp. and Craspedella sp. on the cephalothorax and walking legs, and in the gills and gill covers of wild C. quadricarinatus, respectively. These findings showed that the cultured C. quadricarinatus had a higher temnocephalid infestation compared to the wild C. quadricarinatus. These different temnocephalid species occupied different niche areas, but factors accounting for their distribution on C. quadricarinatus need further investigation.

Molecular markers for hemocyte subpopulations in crayfish Cherax quadricarinatus

Semigranular cells (SGCs) and granular cells (GCs) are two dominant groups of circulating hemocytes in crayfish Cherax quadricarinatus. Molecular markers are required for the clear classification of the hemocytes and the research of their function and differentiation. In this study, we compared the protein content of GCs and SGCs by using two workflows: one-dimensional gel electrophoresis followed by LC-MS/MS and in-solution digestion of cell lysate followed by LC-MS/MS. Cell type-specific proteins were identified, and their expression in SGCs and GCs was further investigated by RT-PCR, Western blotting, and immunofluorescence analysis. Three molecular markers for GCs (peroxinectin, a mannose-binding protein, and prophenoloxidase-activating enzyme 2a) and three molecular markers for SGCs (a vitelline membrane outer layer protein I-like protein, a C-type lectin, and a peptidase) were identified. The application of some of the markers in Eriocheir sinensis was also analyzed. These molecular markers are useful tools for the research of crustaceans hemocytes.

Decapod iridescent virus 1: An emerging viral pathogen in aquaculture

AbstractDecapod iridescent virus 1 (DIV1) is an emerging viral pathogen that seriously threatens crustacean farming and has caused enormous economic losses in recent years. This virus was originally isolated and identified from diseased crayfish Cherax quadricarinatus and shrimp Litopenaeus vannamei in Fujian and Zhejiang, China, in 2014 by two research groups. In 2019, the Executive Committee of the International Committee on Taxonomy of Viruses approved DIV1 as a member of the new genus Decapodiridovirus in the family Iridoviridae. DIV1 is icosahedral in shape and contains a single linear double‐stranded DNA genome of approximately 165 kb. DIV1 can infect many host species, including freshwater and seawater crustaceans, but infected hosts do not show unique pre‐clinical signs, which increases the difficulties in the diagnosis and treatment of this disease. Increasing progress has been made towards understanding many aspects of this virus, including its classification, genomic features, transmission mode, host range, pathogenicity, clinical symptoms, histopathology, detection method and host immunity in response to infection. In this review, we provide an overview of recent progress towards understanding the epidemiology and pathobiology of disease caused by DIV1 and highlight the future research prospects in control and prevention strategies.

CqFlightless-I promotes the depolymerization of actin cytoskeleton to reduce white spot syndrome virus infection in Cherax quadricarinatus

As a member of the actin-associated protein gelsolin family, Flightless-I (FliI) is involved in promoting the depolymerization and rearrangement of the actin cytoskeleton, thus regulating a variety of biological processes, including cell proliferation, differentiation, apoptosis, and immune response. However, the molecular regulation of FliI is poorly understood in its regulation on viral infection in crustaceans. Previously, we found that the transcript of CqFliI was up-regulated in a differentially expressed transcriptome library of the hematopoietic tissue (Hpt) cells from red claw crayfish Cherax quadricarinatus at 1 h post white spot syndrome virus (WSSV) infection. To reveal the role of CqFliI in WSSV infection in crustacean, the open reading frame of CqFliI gene was cloned with 3870 bp, which encoded 1289 amino acids with eleven leucine-rich repeat (LRR) domains and six gelsolin-like (GEL) domains. Tissue distribution analysis showed that CqFliI was widely expressed in all the tested tissues with the highest expression in muscle, followed by a high expression in Hpt. In addition, the mRNA expression of CqFliI was significantly up-regulated in Hpt cells at 1, 3, 6, 12 and 24 h post WSSV infection. Importantly, the internalization of WSSV was clearly increased at an early infection stage of 1 hpi in Hpt cells by CqFliI gene silencing; meanwhile, the expression of viral immediate early gene IE1 and late viral envelope protein gene VP28 were both significantly increased after gene silencing of CqFliI in Hpt cells post WSSV infection, indicating a key role of CqFliI in reducing WSSV infection. Furthermore, protein pull-down assay revealed that CqFliI could bind with cytoskeletal β-actin through its GEL domains, rather than LRR domains; and the cytoskeleton of HeLa cells was obviously depolymerized after overexpression of GEL domain of CqFliI, implying that CqFliI might reduce the internalization of WSSV probably also the intracellular transport mediated by cytoskeleton via inhibiting actin polymerization. These findings may contribute to further understanding of WSSV infection mechanism which may provide more antiviral targeting design against WSSV disease.

CDNA Characterization and Expression of Selenium-Dependent CqGPx3 Isoforms in the Crayfish Cherax quadricarinatus under High Temperature and Hypoxia.

Glutathione peroxidase 3 (GPx3) is the only extracellular selenoprotein (Sel) that enzymatically reduces H2O2 to H2O and O2. Two GPx3 (CqGPx3) cDNAs were characterized from crayfish Cherax quadricarinatus. The nerve cord CqGPx3a isoform encodes for a preprotein containing an N-terminal signal peptide of 32 amino acid residues, with the mature Sel region of 192 residues and a dispensable phosphorylation domain of 36 residues. In contrast, the pereiopods CqGPx3b codes for a precursor protein with 19 residues in the N-terminal signal peptide, then the mature 184 amino acid residues protein and finally a Pro-rich peptide of 42 residues. CqGPx3 are expressed in cerebral ganglia, pereiopods and nerve cord. CqGPx3a is expressed mainly in cerebral ganglia, antennulae and nerve cord, while CqGPx3b was detected mainly in pereiopods. CqGPx3a expression increases with high temperature and hypoxia; meanwhile, CqGPx3b is not affected. We report the presence and differential expression of GPx3 isoforms in crustacean tissues in normal conditions and under stress for high temperature and hypoxia. The two isoforms are tissue specific and condition specific, which could indicate an important role of CqGPx3a in the central nervous system and CqGPx3b in exposed tissues, both involved in different responses to environmental stressors.

The effect of ethanol on some hematological parameters of the australian red claw crayfish (Cherax quadricarinatus)

Purpose:to study the effect of ethanol on the parameters of THC, the percentage of granulocytes and total protein in the hemolymph of the Red claw crayfish (Cherax quadricarinatus).Materials and methods.The object of this experiment was 26 males of the Australian red-clawed crayfish (Cherax quadricarinatus) weighing from 23 to 83 g. The individuals were evenly divided into two experimental groups - with an injection of ethanol and a control group without an injection of 13 crayfish for each group. The injection dose was 2515 mg per 100 g of body weight. A day after the introduction of ethanol, hemolymph was taken with a syringe from the ventral sinus, the syringe was pre-washed with a 4% EDTA-Na2 solution. Three parameters were determined: the total hemocyte count (THC), percent granulocytes and percent total protein content. Counting of hemocytes and determination of granulocytes were performed in a Goryaev chamber under a light microscope. The total protein was determined by the refractometric method.Results.Differences in THC and total protein between the groups were statistically unreliable (p&gt;0,05). THC in the experimental group is 36% more than in the control group. The total protein after the introduction of ethanol actually increased by 0,7%, and relatively by 14%. There were statistically different indicators of the proportion of granulocytes (p&lt;0,05) - the average value of 33,1% in the experimental group versus 24,5% in the control group. A reliable (p&lt;0,05) strong feedback was revealed between the total protein and the mass of individuals in both experimental groups, while in the experimental group there is a visible shift in the values of dependent hemolymph indicators towards an increase in smaller individuals.Conclusion.A single injection of ethyl alcohol with a dosage of 2515 mg per 100 g of body weight into the hemolymph of C. quadricarinatus does not cause significant changes in the THC and total protein after 24 hours. At the same time, the proportion of granulocytes actually increases by 9%, relative to 37%. This may indicate that granulocytes are involved in the formation of cancer defense mechanisms when exposed to toxic substances. The effect of different dosages of ethanol injections and the duration of its effect on hematological parameters requires additional consideration. It is necessary to investigate its effect on other indicators, such as the pH and buffer capacity of the hemolymph, the concentration of hemocyanin, glucose, lactates and calcium.