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Pearl gentian grouper is a new aquacultural hybrid resulted from breeding of female tiger grouper (Epinephelus fuscogutattus) and male giant grouper (E. lanceolatus). Our preliminary study found that pearl gentian grouper exhibits less susceptible to the primary infection of Cryptocaryon irritans, which is an important parasitic ciliate in marine aquaculture, indicated that pearl gentian grouper might own a strong innate immune system. Complement system play key roles in innate immunity, whether pearl gentian grouper's complement component contribute for the defensing against the C. irritans infection remain unclear. In the present study, we found that C. irritans can be immobilized by untreated serum but not heat-treated serum from pearl gentian grouper, suggested that the heat-labile components in serum are responsible for the immobilization of C. irritans. Moreover, we cloned and characterized the encoding sequence of pearl gentian grouper complement C3 (PGC3), a key component in complement system. We also found that the expression level of PGC3 was increased in infected grouper serum when compared with that of control grouper. Furthermore, the binding of PGC3 on the surface of C. irritans trophonts located on the grouper skin was detected. These data suggested that pearl gentian grouper's complement system indeed play roles in the immune response against the C. irritans infection.

The adoption of new investigative strategies based on environmental DNA (eDNA) can be used to monitor parasites, associated bacterial microbiomes, and physical-chemical parameters in fish farms. In this study, we used the economically important and globally distributed fish ciliate parasite Cryptocaryon irritans as a model to understand the parasite abundance and potential drivers of its presence in marine fish farms. Environmental (rainfall) and physical-chemical (temperature, oxygen, salinity, pH) data collected from a marine fish farm in Hong Kong were analyzed together with the eDNA approach targeting C. irritans abundance based on digital droplet PCR and 16S metagenomics to determine associations and triggers between parasites and specific bacterial groups. Rainfall and temperature demonstrated positive associations with high abundance of C. irritans (eDNA) at the studied marine fish cage farm. However, rainfall was the only parameter tested that demonstrated a significant association with parasite eDNA, indicating that the raining season is a risky period for fish farmers in Hong Kong. Coraliomargarita was the bacterial genus with the most significant relationship with low abundance of C. irritans in water. Understanding the environmental triggers of ciliate parasites propagation and associated bacterial microbiome could elucidate new insights into environmental control, microbial management, and promote the reduction of chemical use in marine fish farms.

Ciliates are fundamental components of microzooplankton, with important ecological roles. However, ciliate communities are particularly difficult to monitor using conventional morphological approaches. New molecular tools, such as DNA metabarcoding, can facilitate the study of these communities. This study used high-throughput sequencing to examine the diversity and seasonal dynamics of ciliate communities in four estuarine ecosystems in the South China Sea from June 2019 to March 2020. The amplification of the V4 region of 18S rDNA using ciliate-specific primers identified a total of 1645 amplicon sequence variants (ASVs), corresponding to 13 ciliate classes, 97 families, and 157 genera. The dominant species across all four sampling stations were spirotrichs (including choreotrichs, oligotrichs, and stichotrichs), oligohymenophorean scuticociliates, litostomateans Didinium, and prostomateans Cryptocaryon. Significant differences in ciliate diversity and community composition in the four stations were mainly due to differences in rare, rather than abundant, ASVs. Analysis of the ciliate communities and seasonal patterns in their composition revealed that variations in habitat and environmental conditions have a greater effect than seasonal changes on community composition.

Large-scale transcription studies have revealed numerous lncRNAs (long non-coding RNAs). lncRNAs have been proposed to participate in the regulation of a diverse range of biological processes, including transcriptional regulation. Although lncRNAs have attracted increasing attention, the studies in large yellow croaker (Larimichthys crocea) are still rare, and they lack systematic analysis. In this study, 101 RNA-seq datasets varied in ages, sexes, and tissues were retrieved from the NCBI database to generate a comprehensive catalog of large yellow croaker transcriptome database. A set of 14,599 high-confidence lncRNAs from 13,673 loci were identified and characterized. Furthermore, RNA-seq datasets obtained from the infection of C. irritans were employed to investigate the differential expression pattern of lncRNAs and analyze potential biological functions. A total of 77 differentially expressed lncRNAs targeting to 567 protein-coding genes were identified by using expression analysis. Several immune genes, including TLR5, CD2AP, and MMP9, were highlighted. With GO enrichment and KEGG pathway analysis, the immune-related terms or pathways were enriched. This study created a comprehensive dataset of lncRNAs for large yellow croaker, which would be helpful for the researches of functional roles of lncRNAs in large yellow croaker.

A cross-sectional survey was used to estimate the prevalence of infections with the Infectious spleen and kidney necrosis virus (ISKNV, Megalocytivirus), nervous necrosis virus (NNV, Betanodavirus), and infestations with ectoparasites during the rainy season in juvenile grouper (Epinephelus spp.) farmed in Aceh, Indonesia. The survey was intended to detect aquatic pathogens present at 10% prevalence with 95% confidence, assuming 100% sensitivity and specificity using a sample size of 30 for each diagnostic test. Eight populations of grouper from seven farms were sampled. Additional targeted sampling was conducted for populations experiencing high mortality. Infection with NNV was detected at all farms with seven of the eight populations being positive. The apparent prevalence for NNV ranged from 0% (95% CI: 0&ndash, 12) to 73% (95% CI: 54&ndash, 88). All of the fish tested from the targeted samples (Populations 9 and 10) were positive for NNV and all had vacuolation of the brain and retina consistent with viral nervous necrosis (VNN). Coinfections with ISKNV were detected in five populations, with the highest apparent prevalence being 13% (95% CI: 4&ndash, 31%). Trichodina sp., Cryptocaryon irritans and Gyrodactylus sp. were detected at three farms, with 66% to 100% of fish being infested. Hybrid grouper sourced from a hatchery were 5.4 and 24.9 times more likely to have a NNV infection and a higher parasite load compared to orange-spotted grouper collected from the wild (p &lt, 0.001). This study found that VNN remains a high-impact disease in grouper nurseries in Aceh, Indonesia.

(Plectropomus leopardus) or grouper coral trout is one kind of commodities that have high economic value. As marine fisheries commodities are much in demand for large-scale farmers or household level will require broodstock who have qualified to produce quality eggs and a good seed. qualified to produce quality eggs and a good seed. The purpose of research is to gain knowledge in maintenance activities of the broodstock grouper coral trout on a concrete tank by comparing theory and application in the field. Maintenance engineering of coral trout grouper broodstock includes the preparation of concrete tanks, broodstock selection, water quality management, feeding, spawning and harvesting of eggs, and disease. Research in Balai Besar Penelitian dan Pengembangan Budidaya Laut (BBPPBL) Gondol. Penyabangan Village, Subdistrict Gerokgak, Buleleng, Bali. On 18th January – 18th February 2016. Methods of work used in this research is descriptive data retrieval methods include primary data and secondary data. Data retrieval is done by means of active participation, observation, interviews and literature study. Results of Field Work Practice is the broodstock of grouper coral trout are maintained as much 217 with an average weight of 512.9 kg and an average length of 56.1 cm. Maintenance of broodstock grouper coral trout performed in container maintenance the round-shaped sized 100 m3 with a depth of 3 m with inlet and outlet Water management is performed by using a system is running water that flowing water as much as 300% -400%, the quality of sea water showing the value of 34.9 ppt salinity, DO 6.0 ppm, pH 8, and the temperature of 29 ˚C. Feed given in the form trash fish as much as 7-10 kg a day. Grouper coral trout spawning is done naturally by the male and female parent ratio of 1: 2, which produce eggs an average 2,252,292 items. During maintenance activities parasites invade have been found. Parasites invade the broodstock of grouper coral trout is Benedenia sp., Cryptocaryon sp., And viral infections Viral Nervous Necrosis (VNN)

Immobilization antigens (i-antigens) are surface membrane proteins that are widely recognized to be the ideal candidates as vaccines antigens for immunization against Cryptocaryon irritans. In this study, we cloned a putative i-antigen gene from C. irritans, which was expressed in all three stages of the C. irritans life-cycle, and localized primarily to the cell surface. The recombinant GDCI3 i-antigen was expressed and purified using the free-living ciliate, Tetrahymena thermophila as an expression system. The purified recombinant protein was recognized by rabbit anti-C. irritans antiserum and was capable of eliciting immobilizing antibodies in rabbits and fish suggesting that the antigen itself was correctly folded. Following immunization and parasite challenge, groupers vaccinated with, recombinant GDCI3 i-antigen had a 25% cumulative percent survival rate compared to 8.3% for controls. Both non-specific and parasite-specific IgMs were generated in fish following immunization, with the levels of both increasing following challenge. Parasite-specific IgM in mucus could only be elicited after challenge of the GDCI3 i-antigen vaccinated groupers. To our knowledge, this is the first report using the Tetrahymena expression system to generate C. irritans i-antigens and investigate their use for fish vaccination.

The ciliated protozoan Cryptocaryon irritans infects a wide range of marine fish and causes the highly lethal white spot disease. This parasite possesses three morphologically and physiologically distinct life stages: an infectious theront, a parasitic trophont, and an asexually reproductive tomont. In the past few years, several attempts have been made to help elucidate how C. irritans transforms from one stage to another using transcriptomic or proteomic approaches. However, there has been no research studying changes in transcription profiles between different time points of a single C. irritans life stage-the development of this parasite. Here we use RNA-seq and compare gene expression profiles of theront cells collected by 1 and 10 hrs after they emerged from tomonts. It has been shown that infectivity of theront cells declines 6-8 hours post-emergence, and we used this characteristic as a physiological marker to confirm the aging of theront cells. We identified a total of 41 upregulated and 90 downregulated genes that were differentially expressed between young and aging theront cells. Using Blast2Go to further analyze functions of these genes, we show that genes related to energy production are downregulated, but quite surprisingly many genes involved in transcription/translation processes are upregulated. We also show that expression of all nine detectable agglutination/immobilization antigen genes, with great sequence divergence, is invariably downregulated. Functions of other differentially expressed genes and indications are also discussed in our study.

During expeditions to Curacao in August and October of 2013, a large number of fish infected with dermal parasites was observed. Infected individuals pre- sented black spots and white blemishes on their skin and fins that were easily observed by divers, and which have been associated with infections by trematodes, turbel- larians, and protozoans (Cryptocaryon). In order to com- pare rates of infection across localities in the Caribbean, we conducted visual censuses of reef fish communities along 40 m 2 belt transects in Belize (n=35), Curacao (n=82), and Mexico (n=80) over a 4-week period. Three affected individuals were recorded in Belize, 75 in Curacao, and none in Mexico. Approximately 68 % of the infected individuals in Curacao were surgeonfishes (Acanthuridae). There was no correlation between inci- dence of infection and species abundance (r 2 =0.03), or with functional traits (diet, mobility, schooling behavior, or position in the water column). The causes of the strik- ingly high incidence of dermal parasites in Curacao and its consequences remain unknown. However, considering that parasites with complex life cycles have several hosts throughout their lives, and that past disease outbreaks have had severe consequences on communities of the Caribbe- an, we caution that coral reef ecosystems of Curacao should be closely monitored.

Background Cryptocaryon irritans is an obligate parasitic ciliate protozoan that can infect various commercially important mariculture fish species and cause high lethality and economic loss. Current methods of controlling this parasite with chemicals or antibiotics are widely considered to be environmentally harmful. Piscidins with broad spectrum antibacterial, antifungal and antiviral activities were found to have potent activity against C. irritans. Little, however, has been understood about the killing mechanisms of piscidins in parasites. Results In total, 57.12, 50.44, 55.86 and 47.87 million raw reads were generated from untreated theront and trophont, and piscidin (Lc-pis) treated theront and trophont libraries, respectively. After de novo assembly, 966,609 unigenes were generated with an average length of 420 bp: among these, 618,629 unigenes showed identity with sequences in one or more databases, with some showing to be significantly manipulated by Lc-pis treatment. The species classification showed that more than 25.8% unigenes from trophonts were homologous to the large yellow croaker (Larimichthys crocea) and less than 3.8% unigenes from theronts were matched. The homologous unigenes demonstrated that the tissue from host could exist in trophonts and might be transported to parasite via vesicular transports. Our analysis showed that regulatory transcripts were involved in vesicular trafficking. Among transcripts induced by Lc-pis, most genes up-regulated in treated and untreated theronts were involved in cell migration and apoptosis related pathways. Few transcripts were found to be down-regulated in treated and untreated trophonts related to cell structure and migration after treatment. Conclusions This is the first transcriptome analysis of C. irritans exposed to Lc-pis, which enhanced the genomic resources and provided novel insights into molecular mechanisms of ciliates treated by cationic antimicrobial peptide. Our comprehensive transcriptome analysis can facilitate the identification of potential drug targets and vaccines candidates for controlling this devastating fish pathogen. Electronic supplementary material The online version of this article (10.1186/s12864-018-4565-5) contains supplementary material, which is available to authorized users.

SUMMARYAquaculture, including both freshwater and marine production, has on a world scale exhibited one of the highest growth rates within animal protein production during recent decades and is expected to expand further at the same rate within the next 10 years. Control of diseases is one of the most prominent challenges if this production goal is to be reached. Apart from viral, bacterial, fungal and metazoan infections it has been documented that protozoan parasites affect health and welfare and thereby production of fish in marine aquaculture. Representatives within the main protozoan groups such as amoebae, dinoflagellates, kinetoplastid flagellates, diplomonadid flagellates, apicomplexans, microsporidians and ciliates have been shown to cause severe morbidity and mortality among farmed fish. Well studied examples areNeoparamoeba perurans, Amyloodinium ocellatum, Spironucleus salmonicida, Ichthyobodo necator, Cryptobia salmositica, Loma salmonae, Cryptocaryon irritans, Miamiensis avidusandTrichodina jadranica. The present report provides details on the parasites’ biology and impact on productivity and evaluates tools for diagnosis, control and management. Special emphasis is placed on antiprotozoan immune responses in fish and a strategy for development of vaccines is presented.

The protozoan parasite Cryptocaryon irritans is responsible for significant economic losses of commercially farmed marine fishes. In this study we report, for the first time, C. irritans infection in the seawater-adapted guppy Poecilia reticulata. We describe a simple method for experimental propagation and maintenance of the infection in a laboratory, without apparent loss of infectivity, for 15 months. Fish were gradually adapted to increasing concentrations of artificial seawater by successive transfer to incremental steps of 10 ppt every 2 days and then transferred to disease propagation aquaria (salinity 30 ± 2 ppt) where the infection was maintained. Between 2- 3 fish were held at any one time in each aquaria, each fish being replaced once it succumbed to the infection and died. Gross observations of heavily infected fish revealed a uniform distribution of trophonts visible on the skin and fins as white surface lesions or ‘spots’ after 4-5 days. Death of the host usually occurred after 7-8 days. Heavily infected fish were fixed and processed using routine histological techniques and histological examination revealed infective trophonts, round or pyriform in shape, invading the fish epithelium as well as larger, feeding trophonts located beneath the epithelium. A thickening of the epithelial layers and a proliferation of mucous secreting cells was evident around embedded parasites.

6 páginas, 1 tabla, During the paat 5 years there has been considerable development of lurbot (Scophthalmus maximus L.) farming in Galicia (Notthwestern Spain ). This development caused a concomitant increase of pathological problems in this species. In this study, the preliminary results of a microbiological survey (comprisingbacreria, virusand pamsires) in an ongrowingturbot farm aregiven.The bacteria more frequently isolated belong 10 the genus Vibrio ( V. splendidus-V. pelagius), and wirh less fr& quency IO Pseudumonar. Srreptocorcus and Staphylococcus. Birnaviruses (IPN-like virus) were irelated from only two samples. The flagellate Costia sp., lhe ciliates Trichodino up. and Cryptacwyon sp.. the microrporidian Tetmmicra breeijitwn and Ihe cestode Bofhriocephalus scorpii all occurred in low prevalence., This work was supported by grants Mar-89-0270 from the Comision Investigacion Cientifica y Tecnica and 70708888 from the Xunta de Galicia