Red Swamp Crayfish Procambarus Clarkii Research Articles

Distribution and diet of recovering Eurasian otter (Lutra lutra) along the natural-to-urban habitat gradient (river Segura, SE Spain)

In the catchment of the River Segura (SE Spain), ongoing Eurasian otter (Lutra lutra) recovery and concurrent spread of human settlements are resulting in increasing otter records into urban areas. We analysed otter diet along a natural-to-urban gradient, with the main aim of assessing to which extent man-driven variation in food availability affects its feeding habits and how this secretive mustelid can thrive in apparently unsuitable habitats. From 2016 to 2019, we collected 521 spraints, monitoring 37 sampling stations (mean length ± SD = 0.5 ± 0.3 km). In natural areas, the percent cover of undisturbed banks was the highest, with the lowest number of water abstraction systems, whilst water pollution was the highest in urban areas. Introduced red swamp crayfish (Procambarus clarkii) and fish formed the bulk of otter diet. The two most preyed fish, Luciobarbus sclateri and Pseudochondrostoma polylepis, were among the most abundant species in the river, confirming the generalist feeding behaviour of the otter. The contribution of non-native fish was on average low, while invasive crayfish exceeded the overall importance of fish in all habitats except urban areas. Overall, diet data did not allow attesting food as a limiting resource for otters in urban areas. In contrast, marking intensity varied consistently with the natural-to-urban gradient (from 0.31 to 0.65 spraints/100 m in urban- and natural areas, respectively), suggesting that human disturbance (i.e. habitat modification and water pollution) may play a major role in shaping otter distribution.

High feeding level alters physiological status but does not improve feed conversion efficiency and growth performance of juvenile red swamp crayfish Procambarus clarkii (Girard, 1852)

Red swamp crayfish, Procambarus clarkii (Girard, 1852), aquaculture has developed rapidly in recent years. However, limited knowledge about the response of physiological processes to different feeding levels has hindered the expansion of scientific feeding strategies. The present study was conducted to evaluate the growth performance (specific growth rate: SGRw, survival, and moulting frequency), feed utilisation (feeding rates, feed conversion ratio: FCR, waste production ratio: WPR), and physiological response variables to different feeding levels (1%, 2%, 3%, 4%, 5%, and 6% body weight per day, BW day−1) of juvenile P. clarkii for 60 days. Each treatment was replicated ten times, with 40 juveniles per treatment. No significant differences were observed in the growth performance of P. clarkii fed at 3%, 4%, 5%, and 6% BW day−1. However, they exhibited a significantly higher feeding rate, and FCR and WPR values when fed at 6% BW day−1. Among the physiological variables, activities of digestive enzymes (trypsin, lipase, and AKP) and antioxidant enzymes (SOD, CAT, and MDA) were significantly correlated with feeding levels; the Pearson correlation coefficients of these enzymes were 0.82, 0.57, 0.39, 0.66, 0.70, and −0.38, respectively. The biochemical indicators of nutrient status (total protein, total cholesterol, triglyceride, and GOT and PK activities) increased with increasing feeding levels, as revealed by the Self-Organizing Map. The two-way ANOVA revealed that P. clarkii fed at 5% and 6% BW day−1 had significantly higher trypsin and PK activities, and total protein and total cholesterol contents, whereas when fed at 6% BW day−1, it was exposed to oxidative stress. When the physiological variables were grouped by principal component analysis (PCA), they varied over experimental dates. P. clarkii exhibited higher trypsin, lipase, and MDA activities on the 20th day, but lower triglyceride content and GOT activity than on the 40th day. This study indicated that P. clarkii did not benefit from high feeding level (6% BW day−1), providing an incentive for farmers to reduce dietary input and production costs, thus promoting the sustainability of P. clarkii aquaculture.

The redclaw crayfish: A prominent aquaculture species with invasive potential in tropical and subtropical biodiversity hotspots

AbstractThe redclaw crayfish (Cherax quadricarinatus; hereafter redclaw), native to northern Australia and southern New Guinea, is among the largest freshwater decapods. It matures early and is considered highly prolific as females may lay over one thousand eggs in a single clutch. Despite generally preferring slow‐moving streams in its native range, it has a wide environmental tolerance, making it capable of establishing populations when introduced to a wide range of other conditions and habitats. These biological and ecological features render it a highly suitable and popular species for aquaculture worldwide, being the second most important crayfish species economically (after the red swamp crayfish Procambarus clarkii). Adding to that, its unique coloration fuels demand and value among aquarium enthusiasts, making it attractive for the aquarium pet trade. Today, redclaw is widely translocated (67 countries/territories) and various established wild populations (22 countries) have been reported on every continent except Antarctica. Information on its potential or observed impacts, however, is sparse and often anecdotal. To address this gap, this comprehensive review compiles all available information on this species, covering its taxonomy and description, biology and ecology, native and non‐native ranges accompanied with documented introduction pathways. Built upon these, we conducted biological and socio‐economic classification and species distribution modelling. We reveal a lack of thorough impact assessments for this species despite sufficient indications of major observable impacts at local scales. We call attention to the importance of managing the use of this prominent introduced species in aquaculture and aquarium pet trade.

Evaluation of the nutritional quality of edible tissues (muscle and hepatopancreas) of cultivated Procambarus clarkii using biofloc technology

A 60-day study was conducted to evaluate the growth performance, nutritional quality of edible tissues (muscle and hepatopancreas) of red swamp crayfish (Procambarus clarkii) cultivated using commercial diet feeding (diet group) and biofloc technology (biofloc group). The commercial diet used in the biofloc group was only 60 % of that in the diet group. Glucose and wheat bran were used as additional carbon sources in the biofloc group. The results showed bioflocs was characterized by sufficient protein content but lower lipid content, however, the growth performance of crayfish in the biofloc group were significantly better at the end of the experiment. Besides, the total lipid content in hepatopancreas of crayfish in the biofloc group was higher. Moreover, saturated fatty acid (STA) content in hepatopancreas and monounsaturated fatty acid (MUFA) content in muscle were higher in the biofloc group, while MUFA content in muscle was significantly lower. No significant difference was observed in total amino acid (TAA), essential amino acid (EAA) and essential amino acids index (EAAI) in muscle in both groups. In conclusion, crayfish could be better cultivated using biofloc technology. However, the lipid deficiency and nutritional imbalance of bioflocs would affect the lipid nutritional composition in edible tissues of crayfish.

A sex‐reversing factor: insulin‐like androgenic gland hormone in decapods

AbstractA large number of decapod crustaceans with sexual dimorphism in body size, most of which, are important economic animals. Besides, some of them such as the red swamp crayfish Procambarus clarkii possess strong invasive ability and would damage ecosystem environment. Thereby, monosex culture of decapods would contribute to not only increasing yield and profits, but also reducing the risks of biological invasion. Insulin‐like androgenic gland hormone was reported to be a key regulator in sex differentiation. All‐male populations in Macrobrachium rosenbergii were successfully produced through insulin‐like androgenic gland hormone manipulation. In addition to sex differentiation, insulin‐like androgenic gland hormone was reported to be involved in male secondary sex characteristics maintenance, growth, metabolism, morphology change, ovary development and other functions. Considering its important application value for decapods, insulin‐like androgenic gland hormone has attracted much attention in recent years. This hormone was reported to have the structure and function similar to androgenic gland hormone of isopods. In this study, the origin of name of androgenic gland hormone and insulin‐like androgenic gland hormone was investigated, and the recent research on structure, function, application and molecular regulation mechanism of insulin‐like androgenic gland hormone of decapods was reviewed.

Risk of invasiveness of non-native aquatic species in the eastern Mediterranean region under current and projected climate conditions

Human-induced biological introductions pose a major threat to global biodiversity, and this is especially frequent in the eastern Mediterranean region, which is a globally important biodiversity hotspot area of high conservation value. To predict at which level introduced species in this region might become invasive under current and projected climate conditions, 232 non-native aquatic organisms were screened using the Aquatic Species Invasiveness Screening Kit. Based on receiver operative characteristic curve analysis, thresholds were identified to distinguish between low, medium and high risk species. The “top invasive” (very high risk) species identified were: brown bullhead Ameiurus nebulosus, blue crab Callinectes sapidus, gibel carp Carassius gibelio, Philippine catfish Clarias batrachus, Chinese mitten crab Eriocheir sinensis, bluespotted cornetfish Fistularia commersonii, silver carp Hypophthalmichthys molitrix, silver-cheeked toadfish Lagocephalus sceleratus, half-smooth golden pufferfish Lagocephalus spadiceus, Suez pufferfish Lagocephalus suezensis, signal crayfish Pacifastacus leniusculus, fathead minnow Pimephales promelas, channeled applesnail Pomacea canaliculata, red swamp crayfish Procambarus clarkii, devil firefish Pterois miles and European catfish Silurus glanis. The risk of being invasive of more than half of the screened species increased after taking global warming predictions into account, and several species considered to be globally invasive (cf. high risk) were classified as posing only a medium risk for the eastern Mediterranean region. Region-specific risk screenings, as implemented in this study, are therefore essential for setting priorities in preventative management for the conservation of key biodiversity hotspots and the optimal allocation of resources in view of full risk assessment for the species identified as (very) high risk.

Evaluation of dissolved carbon dioxide to stimulate emergence of red swamp crayfish Procambarus clarkii (Decapoda: Cambaridae) from infested ponds

Evaluation of dissolved carbon dioxide to stimulate emergence of red swamp crayfish Procambarus clarkii (Decapoda: Cambaridae) from infested ponds

Effects of co-occurrence of invading Procambarus clarkii and Pomacea canaliculata on Vallisneria denseserrulata-dominated clear-water ecosystems: a mesocosm approach

Single invaders often substantially alter ecosystems, but the potential impacts by multiple invaders remain understudied. The golden apple snail (Pomacea canaliculata) and the red swamp crayfish (Procambarus clarkii) are two widespread coinvaders. To test the effects of co-occurrence of the two species on a clear-water macrophyte state, we performed a 20-day experiment in 16 mesocosms (4 contained only two snails, 4 contained only two crayfishes, 4 contained two snails and two crayfishes, and 4 controls). Comparing with the control, the leaf length, number, and biomass of Vallisneria denseserrulata had decreased in the snail-only and crayfish-only treatments. In the crayfish-only treatment, total nitrogen, total phosphorus and total suspended solids had increased compared with the control, while they did not differ between the snail-only and the control treatment. Nutrients and turbidity concentrations did not differ between the snail + crayfish and the crayfish-only treatment, and plant biomass did not differ between the snail + crayfish and the snail-only treatment. These findings suggest that golden apple snails mainly affected the lake ecosystem by plant grazing, while red swamp crayfishes disturbed the sediment by increasing nutrients in the water and through resuspension. These snail and crayfish together had mainly additive effects on macrophyte and the physico-chemical variables studied.

Predictor complexity and feature selection affect Maxent model transferability: Evidence from global freshwater invasive species

AbstractAimEcological niche models (ENMs) are widely used to address urgent real‐world problems such as climate change effects or invasive species; however, the generality of models when projected through space and/or time, that is transferability, remains a key challenge. Here, we explored the effects of complex predictors and feature selection on ENM transferability in a widely employed algorithm, Maxent, using five globally invasive freshwater species as case studies.LocationGlobal.MethodsWe modelled the global distributions of five notorious freshwater invasive species (African sharptooth catfish Clarias gariepinus, Mozambique tilapia Oreochromis mossambicus, American bullfrog Lithobates catesbeianus, red swamp crayfish Procambarus clarkii, and Australian redclaw crayfish Cherax quadricarinatus), using three predictor datasets of varying complexities derived from two commonly used climatic data sources (WorldClim and IPCC) and three methods of model tuning that differentially incorporated feature selection. Spatially explicit transferability assessments were then conducted using a suite of evaluation metrics previously used to quantify Maxent model performance.ResultsWe show that in the absence of detailed biological knowledge of focal species, simpler predictor datasets produce models that are more accurate than those calibrated using comprehensive “bioclimatic” datasets. Additionally, we find that tuning models for both optimal regularization parameters as well as feature‐class combinations led to the greatest increases in transferability and geographic niche conservatism. Results indicate a tenuous link between model transferability and Akaike's information criterion corrected for small sample sizes (AICc), suggesting that the indiscriminate use of AICc as an estimate of model parsimony may lead to erratic model performance.Main conclusionsOur findings demonstrate that methodological considerations can drastically affect the reliability of spatial and possibly temporal projections, which has severe implications when ENMs are used to infer species’ niches, and quantify ecological or evolutionary change across impacted landscapes.

Transcriptomic analysis of Procambarus clarkii affected by \u201cBlack May\u201d disease

Each year from April to May, high mortality rates are reported in red swamp crayfish (Procambarus clarkii) cultured in Jiangsu and other regions, in China, and this phenomenon has come to be known as “Black May” disease (BMD). Therefore, in order to investigate the possible causes of this disease, this study gathered BMD-affected P. clarkii samples and performed transcriptome analysis on hepatopancreas, gill, and muscle tissues. A total of 19,995,164, 149,212,804, and 222,053,848 clean reads were respectively obtained from the gills, muscle, and hepatopancreas of BMD-affected P. clarkii, and 114,024 unigenes were identified. The number of differentially expressed genes (DEGs) in gill, muscle, and hepatopancreas was 1703, 964, and 476, respectively. GO and KEGG enrichment analyses of the DEGs were then conducted. Based on KEGG pathway enrichment analysis, the most significantly differentially expressed pathways were mainly those involved with metabolism, human disease, and cellular processes. Further analysis of the significantly DEGs revealed that they were mainly related to the mitochondrial-mediated apoptosis pathway and that the expression of these DEGs was mostly down-regulated. Moreover, the expression of genes related to immune and metabolism-related pathways was also significantly down-regulated, and these significantly-inhibited pathways were the likely causes of P. clarkii death. Therefore, our results provide a basis for the identification of BMD causes.

Enhanced growth performance, immune responses, immune-related gene expression and disease resistance of red swamp crayfish (Procambarus clarkii) fed dietary glycyrrhizic acid

Glycyrrhizic acid (GA) is an effective antiviral agent and is widely used in humans and animals. However, while the impact of disease on crustaceans is increasingly serious, research on the application of GA in crustaceans is very rare. The effects of GA on the growth performance, immune response, immune-related gene expression and disease resistance of crayfish were studied. A total of 900 healthy and equal weight crayfish were randomly divided into six groups for a feeding experiment, which lasted 60 days: the GA25, GA50, GA75, GA100 and GA150 groups were fed a basal diet supplemented with 25, 50, 75, 100 and 150 mg·kg−1 GA, respectively, and the control group was fed a basal diet. The results showed that the crayfish in the groups supplemented with 75 to 100 mg·kg−1 GA had an increased final body weight (FBW), weight gain (WG) and specific growth rate (SGR) and decreased feed conversion ratio (FCR) compared with the controls (P < 0.05). The total hemocyte count (THC) and phenoloxidase (PO) content in the hemolymph of crayfish in the groups supplemented with 75 to 100 mg·kg−1 GA were significantly increased compared with those in the control group. Additionally, the total antioxidant capacity (T-AOC) and the total superoxide dismutase (T-SOD), glutathione peroxidase (GPX), acid phosphatase (ACP) and alkaline phosphatase (AKP), and lysozyme (LZM) in the hemolymph and hepatopancreas of crayfish were significantly increased, while the content of malondialdehyde (MDA) and reactive oxygen species (ROS) were significantly decreased in the groups supplemented with 50 to 150 mg·kg−1 GA compared with the control group (P < 0.05). Furthermore, supplementation with 50 to 100 mg·kg−1 GA downregulated the expression of bax and upregulated the expression of bax inhibitor-1 (bi-1), toll-like receptor (tlr), lectin, nuclear factor kappa-B (NF-κB) and heat shock protein 70 (hsp70) (P < 0.05). In addition, supplementation with 50–150 mg·kg−1 GA could decrease the mortality of crayfish infected with white spot syndrome virus (WSSV). These results showed that growth performance, immune responses and immune-related gene expression can be improved by adding GA at the optimal dose of 50–150 mg·kg−1 in crayfish.

Unbalanced population structure and reliance on intraspecific predation by largemouth bass in an agricultural pond with no available prey fish

The carnivorous largemouth bass Micropterus salmoides has been introduced to many regions and poses a threat to native aquatic organisms due to predation, resulting in a significant decrease in biodiversity. Under conditions of prey scarcity, the largemouth bass is thought to consume smaller conspecifics. However, little is known about this species’ dietary reliance on intraspecific predation and its impact on the population structure. We investigated the length–frequency distribution of largemouth bass and estimated their reliance on intraspecific predation using stable carbon and nitrogen isotopes in an agricultural pond with no available prey fish. Almost all individual largemouth bass were collected via the pond drainage system. Within the largemouth bass population, the dominant length class was composed of fish measuring 100–160 mm standard length (SL), whereas few individuals measuring < 100 mm SL (first-year juveniles) were collected. The results from a Stable Isotope Analysis in R mixing model indicated that all four length classes (105–146, 165–180, 216–246, and 300–337 mm SL) of largemouth bass fed mainly on juvenile conspecifics and on red swamp crayfish (Procambarus clarkii). First-year juveniles could have been predated by other individuals of various sizes. The dominant 100–160 mm SL length class may have experienced a weak predation pressure when they were in the first-year juvenile class. Consequently, the largemouth bass population was unbalanced, with few first-year juveniles (< 100 mm SL) and dominance of a 100–160 mm SL length class.

Recycling biofloc waste as novel protein source for crayfish with special reference to crayfish nutritional standards and growth trajectory

Screening of novel feedstuffs, that too for data-deficient (nutritionally) animals, is somewhat ambiguous or problematic. Through systematic meta-analyses, the present study formulated most up-to-date crayfish nutritional standards, against which a recyclable waste (biofloc biomass, BM) from intensive aquaculture systems was assessed as a novel protein source. Growth trajectory dependencies and thermal growth coefficient qualifying for good growth in crayfish (TGC 0.5–0.64 units) were benchmarked. Using these standards and a 7-week growth trial, BM's suitability as a novel protein source for red swamp crayfish Procambarus clarkii was evaluated through its graded inclusions in a commercial feed. Results suggest that BM can elevate growth at 33–66% inclusion in existing feed formulations. Beyond 66% inclusion, BM can deteriorate growth in crayfish due to high ash content (exceeding physiological limit > 14%), arginine deficiency (~ 14–20% lower than an optimum requirement), and insufficient non-protein energy: protein ratio (3.7 cal mg−1). Arginine is perhaps the most critical amino acid in dietary protein for crayfish, and deficient in BM. Although no critical bioaccumulation levels of heavy metals were breached by feeding 100% BM to crayfish, a mineral and heavy metal (Hg) stress seemed plausible. Crayfish raised solely on biofloc may not realize full growth potential.

The effects of the herbicides terbuthylazine and metazachlor at environmental concentration on the burrowing behaviour of red swamp crayfish

Despite their low concentrations in many aquatic environments, evidence exists to suggest that herbicides do affect non-target organisms. Given that burrowing is a primary life-history trait in crayfish, herbicides could potentially have serious negative effects on these ecologically important freshwater macroinvertebrates. In this study, we exposed the red swamp crayfish Procambarus clarkii to terbuthylazine (a triazine) and metazachlor (a chloroacetanilide) at an environmental concentration of 2.0 μg/L for 28 days, and then observed their burrowing behaviour for two days. The metazachlor-exposed males excavated a greater number of burrows than the other tested groups, with comparable depths and volumes relative to individual specimen weight. The relative depth and volume of female burrows were identical in all groups. The natural habit of female crayfish of constructing deeper burrows than males was marginally significant in the control and META groups but was not significant for relative volume. The hypothesized adverse effects of chronic exposure to real environmental concentrations of herbicides were not documented in terms of either relative depth or volume. However, the increased number of burrows in metazachlor-exposed animals may mean that this invasive species will cause greater damage to embankments and river banks. The mechanisms behind these effects require closer study.

Effects of dietary protein levels on growth, muscle composition, digestive enzymes activities, hemolymph biochemical indices and ovary development of pre-adult red swamp crayfish (Procambarus clarkii)

This study was conducted to evaluate the effect of dietary protein levels on growth performance, muscle composition, digestive enzymes activities, hemolymph biochemical indices and ovary development of pre-adult red swamp crayfish Procambarus clarkii (initial weight = 12.86 ± 0.10 g). Six isoenergetic diets (fish meal, soybean meal, gluten, and shrimp powder as main protein sources) were formulated to contain graded levels of 20.70 %, 24.70 %, 30.56 %, 35.77 %, 41.12 % and 44.64 % crude protein. Each diet was assigned to triplicate groups of 15 experimental crayfish (10 female and 5 male) for 8 weeks. Our results showed that weight gain (WG) increased with dietary protein supplementation from 20.7% to 35.77%, and then decreased with further increases in dietary protein. Broken-line regression analysis based on WG showed that the optimal dietary protein level for maximum growth performance of crayfish was 39.60 %. Feed conversion ratio (FCR) exhibited an opposite trend as observed in WG, with the lowest in crayfish fed the 30.56 % or 35.77 % protein diet. Protein efficiency ratio (PER) in crayfish fed the 41.12 % or 44.64 % protein diet was significantly lower than that in crayfish fed the 20.70–30.56 % protein diet. Hepatosomatic index (HSI) value of crayfish decreased with dietary protein supplementation and achieved minimum values at 35.77 % or 41.12 % protein diet. The highest crude protein content of tail muscle was found in crayfish fed the diet containing 35.77 % or 41.12 % protein. Activities of protease and lipase in both hepatopancreas and intestine were significantly affected by dietary protein levels, while amylase activity showed no significant difference. Dietary protein also significantly induced the contents of hemolymph total protein, albumin, globulin, glucose, total cholesterol and low-density lipoprotein, along with aspartate transaminase and alanine transaminase activities. Moreover, histological analysis revealed that the ovary development of most female crayfish fed the 35.77–41.12 % protein diet were at stage V, which had a higher frequency of mature oocytes than that of females fed other protein diets. Thus, these results provide the theoretical basis for feed formulation to determine the appropriate protein requirement of pre-adult red swamp crayfish.

Roles of a small GTPase Sar1 in ecdysteroid signaling and immune response of red swamp crayfish Procambarus clarkii

Secretion-associated and ras-related protein 1 (Sar1) is a small GTPase that plays an important role in the transport of protein coated with coat protein complex II vesicles. However, its alternative roles in the biological processes of Procambarus clarkii remain unclear. Here, a sar1 gene (named as Pc-sar1) with an open reading frame of 582 bp from P. clarkii was identified. Pc-sar1 was expressed in all examined tissues with highest expression levels in muscle, which was determined by real-time PCR and western blotting. After the induction of lipopolysaccharide (LPS) and polycytidylic acid (Poly I: C), the transcriptional levels of Pc-sar1 differed in hepatopancreas, gill, muscle and intestine. In contrast, the expression of Pc-sar1 was upregulated by 20-hydroxyecdysone in these four tissues. In addition, the RNA interference of Pc-sar1 significantly affected the expression levels of immune and hormone-related genes. These results indicate that Pc-sar1 is involved in the innate immune response and ecdysteroid signaling pathway.

Silicone stressed response of crayfish (Procambarus clarkii) in antioxidant enzyme activity and related gene expression.

Organosilicon has been widely used in various fields of industry and agriculture due to its excellent properties, such as high and low temperature resistance, flame retardant, insulation, radiation resistance and physiological inertia. However, organosilicon toxicity in aquatic animals is seldom known. In this research, two typical silicone or silane coupling agents (KH-560 (3-Glycidoxypropyltrimethoxysilane) and KH-570 (3-Methacryloxypropyltrimethoxysilane)) were used in a hydroponic experiment to evaluate the effects on survival rate, antioxidant response and gene expression in red swamp crayfish (Procambarus clarkii). Crayfishes were grown in black aquaculture boxes containing different concentrations (0, 10, 100 and 1000mgL-1) of KH-560 and KH-570 for 72h, and then crayfish samples were harvested and separated into tissues of carapace, gill and muscle for analysis. The results showed that silicone significantly increased malondialdehyde (MDA) content in muscle by 17%-38% except for the treatment of 100mgL-1 KH-570, and reduced the survival rate of crayfish. Additionally, silicone KH-570 increased the activities of superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD) by 15%-31%, 17%-35%, and 9%-46%, as well as the contents of ascorbate (AsA) and glutathione (GSH) by 19%-31%, and 23%-29% respectively, in muscle tissue, and similar results occurred in KH-560. In the carapace, however, SOD activity was significantly decreased at high concentrations level of both silicone treatments. Moreover, silicon (Si) content was higher in the abdominal muscle of crayfish after silicone treatment. Assay of gene expression showed an obvious increasing expression of antioxidant related genes (Sod1, Sod2, Cat1, Cat2, and Pod1, Pod2) under silicone stress. The above results suggested that silicone caused an obvious stress response in crayfish in both biochemical and molecular levels.

Molecular characterization of Rab5 and Rab6, and their involvement in innate immunity in red swamp crayfish Procambarus clarkii

Rab5 and Rab6 are members of the Rab GTPases protein family, and they play important roles in regulating trafficking organelles, especially in phagosome formation. In this study, full-length cDNA sequences of Rab5 and Rab6 were cloned from crayfish Procambarus clarkii, which were designated as PcRab5 and PcRab6, respectively. The full-length cDNA of PcRab5 is 1367 bp and contains an open reading frame (ORF) of 639 bp encoding 212 amino acids with an estimated molecular weight of 23.0 kDa. The full-length cDNA of PcRab6 is 1097 bp, and ORF is 636 bp encoding 211 amino acids with 23.7 kDa. The deduced amino acid sequence of PcRab5 displayed high identity to that of Penaeus vannamei (97.17%), and PcRab6 shared high similarity to that of Macrobrachium rosenbergii (99.05%). According to the results of multiple sequence alignment and phylogenetic analysis, the guanine-base binding motif (G), phosphate/magnesium-binding motif (PM) and Rab family motifs (RabF) of PcRab5 and PcRab6 are high conserved among different species, however, the N- and C-terminal regions were hypervariable. Besides, the mRNA transcripts of PcRab5 and PcRab6 were widely detected in various tissues of P. clarkii, with the highest expression levels in hemocytes and gill. The gene expression levels of PcRab5 and PcRab6 were significantly induced in hepatopancreas, gill, and intestines of P. clarkii after WSSV and Aeromonas hydrophila infection, respectively. RNA interference experiment was designed to further investigate the roles of PcRab5 and PcRab6 during pathogen infection. The results showed that the mortality rate of crayfish was influenced by silenced PcRab5 and PcRab6 after challenged by WSSV or A. hydrophila. These results suggested that PcRab5 and PcRab6 were involved in bacterial and viral infection in the immune response of P. clarkii

Bioaccumulation of estrogenic hormones and UV-filters in red swamp crayfish (Procambarus clarkii)

Estrogenic hormones and organic ultraviolet-filters (UV-filters) have attracted increased attention as endocrine disrupting chemicals (EDCs) due to their potent estrogenicity and widespread occurrence in the environment. This study investigated the accumulation of three estrogenic hormones and five UV-filters in red swamp crayfish (Procambarus clarkii). Exposure experiments were conducted for 42 days with a mixture of EDCs at two environmentally-relevant design concentrations (i.e., 500 and 5000 ng L−1). The aqueous-phase EDC concentrations decreased over time and were re-established every two days. Within 14 days of exposure, the five UV-filters were measured at 2.2 to 265 ng g−1 (dry weight) in crayfish tail tissue. Only one estrogenic hormone, 17β-estradiol, was detected in the crayfish at 10.4–13.5 ng g−1. No apparent changes were observed for EDC concentrations in the tail tissue over the next four weeks of exposure. The apparent bioaccumulation factors for the EDCs ranged from 23 L (kg tail tissue, dry weight)−1 for 4-methylbenzylidene camphor to 1050 L (kg tail tissue, dry weight)−1 for 2-ethylhexyl-4-methoxycinnamate. EDC input was stopped after 42 days, and the more hydrophobic UV-filters (i.e., octocrylene, 2-ethylhexyl-4-methoxycinnamate, homosalate) were found to be persistent throughout a 14-d elimination period. A lyticase-assisted yeast estrogen screen demonstrated that the residual estrogenic activity of water samples aligned with (or was lower than) predictions from targeted chemical analysis. These results suggest that the transformation products did not contribute significant estrogenicity, although further analysis of endocrine disruption outcomes in crayfish is recommended.

Diagnosis of co-infection with white spot syndrome virus and Aeromonas veronii in red swamp crayfish Procambarus clarkii

Red swamp crayfish Procambarus clarkii is an economically important species that is widely cultivated throughout China. With the application of high-density culture, P. clarkii is usually infected by different pathogens in water. Co-infection with multiple pathogens was common, which has resulted in greater economic loss than single infections. In this study, we investigated a case of co-infected crayfish with bacterial and viral pathogens at a local farm. Firstly, a bacterial strain was isolated from the gills of infected crayfish. According to the results of 16S rRNA sequence and analysis of the Biolog Gen III MicroPlate system, the isolated bacterial strain was finally identified as Aeromonas veronii. The isolated A. veronii exhibited significant virulence to crayfish, with an LD50 value of 6.04 × 106 CFU/crayfish. As a result, drug sensitivity tests showed that the isolated A. veronii was completely resistant to ampicillin, penicillin, and compound sulfamethoxazole tablets, and highly sensitive to seven kinds of antibiotics. Furthermore, white spot syndrome virus (WSSV) was also detected from the gill tissue of diseased crayfish by polymerase chain reaction. After experimentally co-infected with A. veronii and WSSV, the healthy P. clarkii presented similar clinical and histopathological signs with natural infected P. clarkii. The mortality of the co-infected group was 100%, which was higher than that in the group single infected with A. veronii (70%) or WSSV (83.3%). In summary, a case of co-infection with WSSV and A. veronii from P. clarkii was comprehensively studied in this study. The results will supply an effective diagnosis strategy for the co-infection, and provide therapy reference to prevent and control P. clarkii disease.