Effects of Aminomethylphosphonic Acid on the Transcriptome and Metabolome of Red Swamp Crayfish, Procambarus clarkii.

Red swamp crayfish, Procambarus clarkii (P. clarkii), is an important model crustacean organism used in many types of research. However, the effects of different doses of aminomethylphosphonic acid (AMAP) on the transcriptome and metabolites of P. clarkii have not been explored. Thus, this study investigated the molecular and metabolic mechanisms activated at the different exposure dosages of AMAP in P. clarkii to provide new insights into the strategies of P. clarkii in response to the high concentrations of AMAP in the environment. In the present study, the P. clarkii were divided into three groups (control group; low-dosage AMAP exposure; high-dosage AMAP exposure), and hepatopancreatic tissue samples were dependently taken from the three groups. The response mechanisms at the different dosages of AMAP were investigated based on the transcriptome and metabolome data of P. clarkii. Differentially expressed genes and differentially abundant metabolites were identified in the distinct AMAP dosage exposure groups. The genes related to ribosome cell components were significantly up-regulated, suggesting that ribosomes play an essential role in responding to AMAP stress. The metabolite taurine, involved in the taurine and hypotaurine metabolism pathway, was significantly down-regulated. P. clarkii may provide feedback to counteract different dosages of AMAP via the upregulation of ribosome-related genes and multiple metabolic pathways. These key genes and metabolites play an important role in the response to AMAP stress to better prepare for survival in high AMAP concentrations.

Color Perspectives in Aquatic Explorations: Unveiling Innate Color Preferences and Psychoactive Responses in Freshwater Crayfish.

Color preference assay is a test for an animal's innate and adaptive response to differentiate colors and can be used as an endpoint for psychoactive activity evaluation. Several color preference test methods in aquatic animals that can be used to perform behavioral screening have been established. However, the color preference test conditions have yet to be extensively studied and standardized in aquatic invertebrates. This study aimed to replicate and optimize the previously published method to evaluate the potential color preference in freshwater crayfish based on four different approaches: species, life stages, sex, and pharmaceutical exposure. Using the optimized setup, two crayfish species display color preferences to some specific colors. P. clarkii displays more dominant color preference behavior than C. quadricarinatus in terms of color preference ranking and index. P. clarkii prefers the red color compared to other colors (red > green > blue > yellow), while C. quadricarinatus dislikes yellow compared to other colors (blue = green = red > yellow). Since P. clarkii has a more obvious color index ranking and several advantages compared to C. quadricarinatus, we conducted further tests using P. clarkii as an animal model. In the juvenile and adult stages of P. clarkii, they prefer red and avoid yellow. However, the juvenile one did not display a strong color preference like the adult one. Different sex of crayfish displayed no significant differences in their color preference responses. In addition, we also evaluated the potential effect of the antidepressant sertraline on color preference in P. clarkii and found that waterborne antidepressant exposure can significantly alter their color preference. This fundamental information collected from this study supports the crayfish color preference test as a good behavioral test to address environmental pollution.

The effect of a polystyrene nanoplastic on the intestinal microbes and oxidative stress defense of the freshwater crayfish, Procambarus clarkii.

The widespread generation and accumulation of plastic waste has become a globally recognized problem. However, there are limited reports on the adverse effects of nanomaterials on freshwater crustaceans. This study tested the acute effects of different concentrations (0, 5, 10, and 20 mg/L) after 48 h exposure of 75 nm polystyrene nanoplastic on intestinal microbes, and oxidative stress parameters of freshwater crayfish, Procambarus clarkii. High-throughput sequencing analysis revealed the richness, diversity, and composition of intestinal microbiota in P. clarkii exposed to polystyrene nanoplastic. At the genus level, abundances of Lactobacillus, Faecalibaculum, Niveibacterium, and Candidatus Bacilloplasma were significantly different. The reduced abundance of Lactobacillus could affect the balance of intestinal microbes through quantitative disadvantage, which may lead to reduced immunity of P. clarkii. Streptococcus salivarius, Clostridium butyricum and Lachnospiraceae bacterium10-1 in intestinal tract reached maximum abundance at a polystyrene concentration of 20 mg/L. The increase in the number of some pathogenic bacteria may upset the balance of intestinal microorganisms through the number of dominance, and the decrease in the relative abundance of lactic acid bacteria. Probiotics, such as Lactobacillus salivarius, Lactobacillus murinus, Lactobacillus gasseri, Lactobacillus reuteri, Lactobacillus iners AB-1, and Lactobacillus crispatus in the intestinal tract reached the lowest value at a concentration of 10 mg/L. The reduced abundance of Lactobacillus can affect the balance of intestinal microbes through quantitative disadvantage, which may lead to reduced immunity in P. clarkii. At nanoplastic 10 mg/L, the relative abundance of intestinal pathogens increased, while the relative abundance of lactic acid bacteria and other probiotics decreased. With increases in nanoplastic concentrations, the values of glutathione (GSH), superoxide dismutase (SOD), acid phosphatase (ACP), lysozyme (LZM), alkaline phosphatase (AKP), peroxidase (POD), glutathione peroxidase (GPX), and protein carbonylation were significantly changed. Our data suggested that Lactobacillus may play an adjunctive role in the treatment of oxidative stress in P. clarkii exposed to 75 nm polystyrene. This study represents an important step towards a better understanding of the toxic effects of nanoplastics on aquatic crustaceans.

A novel Dicistro-like virus discovered in Procambarus clarkii with "Black May" disease.

The peak period of morbidity and death in cultured Procambarus clarkii is around May each year and is called the "Black May" disease. The pathogen causing "Black May" disease is believed to be a white spot syndrome virus (WSSV). In 2018, a significant number of P. clarkii died in the pond culture of Xinglong Township, Xuyi County. Two sampling tests on the affected pond showed that, in addition to WSSV, a novel Dicistro-like virus (PcDV) was present. Genomic sequence analysis indicated that this new virus belongs to the Dicistroviridae family, Picornaviridaes order. A high number of spherical particles were detected in gill tissues of P. clarkii with "Black May" disease by electron microscopy, a finding consistent with viruses from the Picornaviridaes order. From October 2018 to September 2019, we took monthly samples from Hubei, Jiangsu and Anhui provinces, and tested for the presence of PcDV and WSSV in P. clarkii. The detection rates of PcDV in P. clarkii peaked from April to June, consistent with the onset of the "Black May" disease. In conclusion, we believe that the discovery of PcDV will provide new research directions for investigating the pathogens causing "Black May" disease in P. clarkii.

A novel ferritin gene from Procambarus clarkii involved in the immune defense against Aeromonas hydrophila infection and inhibits WSSV replication.

Ferritin is a protein related to the storage of iron and widely distributed in animals. It participates in many biological process, including antioxidation, cell activation, angiogenesis, regulation of iron metabolic balance and immune defense. In the present study, a novel ferritin gene was identified from red swamp crayfish Procambarus clarkii, with a cDNA sequence encoding a predicted 221 amino-acid residues. The ferritin protein contains a 19-residue signal peptide and 145-residue classic ferritin domain. The NJ phylogenetic analysis showed PcFer clustered with other crustacean peptides. The recombinant PcFer protein was produced and purified in E. coli, and the anti-rabbit polyclonal antibody was obtained. The rPcFer exhibited iron binding activity at a dose-dependent effect. The qPCR and western blot analysis revealed that PcFer was highly expressed in hemocytes, hepatopancreas, and gills. After challenged with WSSV and Aeromonas hydrophila, the mRNA and protein expression patterns of PcFer were significantly up-regulated in hemocytes and hepatopancreas. dsRNA interfering technique was utilized to silence the expression of PcFer gene. The WSSV copy number in PcFer silenced shrimp was much higher than that in the control group. The present study indicated that PcFer was involved in the immune defense against WSSV and Aeromonas hydrophila, and might inhibit WSSV replication in P. clarkii. These results will provide important data support for further study of the functional role of the ferritin gene.