Twenty Asian sea bass Lates calcarifer from a floating cage in Bt. Tambun, Penang were examined for the presence of parasitic gill copepod, Lernanthropus latis. The prevalence of L. latis was 100% with the intensity of infection ranging from 1 to 18 parasites per host or 3.75 of mean intensity. Female parasites having oblong cephalothorax and egg-strings were seen mainly on the entire gill of examined Asian sea bass. The infected gill of Asian sea bass was pale and had eccessive mucus production. Under light and scanning electron microscopies (SEM), L. latis was seen grasping or holding tightly to the gill filament using their antenna, maxilla and maxilliped. These structures are characteristically prehensile and uncinate for the parasite to attach onto the host tissue. The damage was clearly seen under SEM as the hooked end of the antenna was embedded into the gill filament. The parasite also has the mandible which is styliform with eight teeth on the inner margin. The pathological effects such as erosion, haemorrhages, hyperplasia and necrosis along the secondary lamellae of gill filaments were seen and more severe at the attachment site. The combined actions of the antenna, maxilla and maxilliped together with the mandible resulted in extensive damage as L. latis attached and fed on the host tissues.
Acute hepatopancreatic necrosis disease (AHPND) is an important shrimp disease of economic importance which causes mass mortality of cultivated penaeid shrimps in Southeast Asian countries, Mexico and South America. This disease was originally caused by Vibrio parahaemolyticus (VPAHPND) which is reported to harbour a transferable plasmid carrying the virulent PirAB-like toxin genes (pirABvp). However, little is known about the pathogenicity of VPAHPND. To extend our understanding, comparative genomic analyses was performed in this study to identify the genetic differences and to understand the phylogenetic relationship of VPAHPND strains. Seven Vibrio parahaemolyticus strains (five VPAHPND strains and two non-VPAHPND strains) were sequenced and 31 draft genomes of V. parahaemolyticus were retrieved from NCBI database and incorporated into the genomic comparison to elucidate their genomic diversity. The study showed that the genome sizes of the VPAHPND strains were approximately 5 Mbp. Ten sequence types (STs) were identified among the VPAHPND strains using in silico-Multilocus Sequence Typing analysis (MLST) and ST 970 was the predominant ST. Phylogenetic analysis based on MLST and single nucleotide polymorphisms (SNP) showed that the VPAHPND strains were genetically diverse. Based on the comparative genomic analysis, several functional proteins were identified from diiferent categories associated with virulence-related proteins, secretory proteins, conserved domain proteins, transporter proteins, and phage proteins. The CRISPR analysis showed that VPAHPND strains contained less number of CRISPRs elements than non-VPAHPND strains while six prophages regions were identified in the genomes, suggested the lack of CRISPR might promote prophage insertion. The genomic information in this study provide improved understanding of the virulence of these VPAHPND strains.
In Southeast Asia, a new disease called scale drop disease (SDD) caused by a novel Megalocytivirus (SDDV) has emerged in farmed Asian sea bass (Lates calcarifer) in Singapore, Malaysia and Indonesia. We received samples from an Eastern Thai province that also showed gross signs of SDD (loss of scales). Clinical samples of 0.2-1.1 kg L. calcarifer collected between 2016 and 2018 were examined for evidence of SDDV infection. Histopathology was similar to that in the first report of SDDV from Singapore including necrosis, inflammation and nuclear pyknosis and karyorrhexis in the multiple organs. Intracytoplasmic inclusion bodies were also observed in the muscle tissue. In a density-gradient fraction from muscle extracts, TEM revealed enveloped, hexagonal megalocytiviral-like particles (~100-180 nm). By PCR using primers derived from the Singaporean SDDV genome sequence, four different genes were amplified and sequenced from the Thai isolate revealing 98.7%-99.9% identity between the two isolates. Since viral inclusions were rarely observed, clinical signs and histopathology could not be used to easily distinguish between SDD caused by bacteria or SDDV. We therefore recommend that PCR screening be used to monitor broodstock, fry and grow-out fish to estimate the current impact of SDDV in Southeast Asia and to prevent its spread.
We investigated the effects of four iso-nitrogenous (40% crude protein) and iso-caloric (17.6 kJ g(-1)) diets with different dietary carbohydrate levels (15%, 20%, 25% and 30%) on the growth performance, feed utilization efficiency, body composition and liver histology of Malaysian mahseer (Tor tambroides) fingerlings in a 10-week feeding trial. Fish (initial weight of 0.8?0.1 g; initial total length 4.2?0.1 cm) were fed twice daily at 4% body mass. Dietary carbohydrate level had significant effects (P<0.05) on weight gain, SGR (specific growth rate), FCR (feed conversion rate), PER (protein efficiency rate), survival percentage and all nutrient retention values (PRV, LRV, CRV, ERV). Protein, carbohydrate and gross energy composition of the fish body were also significantly differed (P<0.05) among treatments. Liver histology showed mild hepatic steatosis and hypertrophy for fishes receiving a higher dietary carbohydrate inclusion. In general, treatments with 20% and 25% dietary carbohydrate levels produced better growth results compared to the rest of the treatments. Using a second-order polynomial regression analysis model, the optimal dietary carbohydrate level of 23.4% was estimated for mahseer fingerlings. ?
Marine sponges are sessile invertebrates that can be found in temperate, polar and tropical regions. They are known to be major contributors of bioactive compounds, which are discovered in and extracted from the marine environment. The compounds extracted from these sponges are known to exhibit various bioactivities, such as antimicrobial, antitumor and general cytotoxicity. For example, various compounds isolated from Theonella swinhoei have showcased various bioactivities, such as those that are antibacterial, antiviral and antifungal. In this review, we discuss bioactive compounds that have been identified from marine sponges that showcase the ability to act as antibacterial, antiviral, anti-malarial and antifungal agents against human pathogens and fish pathogens in the aquaculture industry. Moreover, the application of such compounds as antimicrobial agents in other veterinary commodities, such as poultry, cattle farming and domesticated cats, is discussed, along with a brief discussion regarding the mode of action of these compounds on the targeted sites in various pathogens. The bioactivity of the compounds discussed in this review is focused mainly on compounds that have been identified between 2000 and 2020 and includes the novel compounds discovered from 2018 to 2021.
In this study, potential probiotic strains were isolated from fermented pickles based on antagonistic activity against two shrimp pathogens (Vibrio harveyi and Vibrio parahaemolyticus). Two strains L10 and G1 were identified by biochemical tests, followed by16S ribosomal RNA gene sequence analysis as Bacillus subtilis, and characterized by PCR amplification of repetitive bacterial DNA elements (Rep-PCR). Subsequently, B. subtilis L10 and G1 strains were tested for antibacterial activity under different physical conditions, including culture medium, salinity, pH and temperature using the agar well diffusion assay. Among the different culture media, LB broth was the most suitable medium for antibacterial production. Both strains showed the highest level of antibacterial activity against two pathogens at 30 °C and 1.0% NaCl. Under the pH conditions, strain G1 showed the greatest activity against V. harveyi at pH 7.3-8.0 and against V. parahaemolyticus at pH 6.0-8.0, whereas strain L10 showed the greatest activity against two pathogens at pH 7.3. The cell-free supernatants of both strains were treated with four different enzymes in order to characterize the antibacterial substances against V. harveyi. The result showed considerable reduction of antibacterial activity for both strains, indicating the proteinaceous nature of the antibacterial substances. A wide range of tolerance to NaCl, pH and temperature was also recorded for both strains. In addition, both strains showed no virulence effect in juvenile shrimp Litopenaeus vannamei. On the basis of these results and safety of strains to L. vannamei, they may be considered for future challenge experiments in shrimp as a very promising alternative to the use of antibiotics.
Culture of Asian seabass, Lates calcarifer (Bloch) is a popular aquaculture activity in Malaysia. This fish is in high demand and fetches a good price in the local market. The seed for this fish is commercially produced by induced spawning in hatcheries. However, the seed supply is affected by frequent mass mortality of larvae aged between 15 and 60 dph. The clinical signs shown by the affected larvae include lethargy, loss of appetite, uncoordinated swimming, unusual spiral movement pattern and dark coloration. Histological examination of brain and eye of the affected specimens revealed extensive cell vacuolation in larvae aged 15-25 dph. Partial nucleotide sequence of the nervous necrosis virus coat protein gene of the affected larvae showed 94.0-96.1% homology to the nucleotide sequences of coat protein gene from nervous necrosis virus isolated from other countries in the Southeast Asia and Australia. This study provides scientific evidence based on molecular technique that many episodes of mass mortality in seabass larvae in Sabah is associated with the viral nervous necrosis. Because no effective treatment has been reported for this infection, stringent biosecurity measures must be adopted for exclusion of the pathogen from the culture system.
The ciliated protozoan parasite Cryptocaryon irritans infecting marine fishes in Taiwan is described. Developmental characteristics and sequences of the ribosomal DNA regions such as part of 18 S, the entire first internal transcribed spacer, and part of 5.8 S of various Taiwan isolates of C. irritans were investigated. A total of 5 isolates was obtained from different fish-host species and localities, the majority from cultured fish species. C. irritans from Taiwan is able to shift its developmental characteristics, i.e. from non-adherent to adherent tomonts, from individualistic to aggregate-forming tomonts, from infection of the gills only to infection of the gills and body. Thus, it is not possible to classify strains of C. irritans on the basis of these parameters. Premature tomonts that developed from dead fishes were able to produce theronts that could infect fish host. Isolates from Pingtung and the USA had identical nucleotide sequences while an isolate from Malaysia was identical to an Israel isolate. Percentage variation among pairs of Taiwan isolates showed a higher degree of variation than isolate sequences listed in GenBank. Sequence analysis revealed highly aberrant isolates in Taiwan, and a phylogenetic tree distinguished a marine and a low-salinity variant. C. irritans from marine fishes in Taiwan, therefore, display some characteristics not previously reported. Since manipulation of salinity in brackishwater ponds and marine cage sites is not feasible, there is a need to develop new strategies for the control and prevention of cryptocaryoniasis.
Strains of Aeromonas hydrophila isolates from skin lesions of the common freshwater fish, Telapia mossambica, were screened for the presence of plasmid DNA by agarose gel electrophoresis and tested for susceptibility to 10 antimicrobial agents. Of the 21 fish isolates examined, all were resistant to ampicillin and sensitive to gentamycin. Most isolates were resistant to streptomycin (57%), tetracycline (48%) and erythromycin (43%). While seven of 21 isolates harboured plasmids, with sizes ranging from 3 to 63.4 kilobase pair (kb), it was only possible to associate the presence of a plasmid with antibiotic resistance (ampicillin and tetracycline) in strain AH11. Both the plasmid and the associated antimicrobial resistance could be transferred to an Escherichia coli recipient by single-step conjugation at a frequency of 4.3 x 10(-3) transconjugants per donor cell.
Examination of 35 barramundi (Lates calcarifer) from aquaculture cages in Setiu Wetland, Malaysia, revealed a single fish infected with three Henneguya spp. (Cnidaria: Myxosporea). Characterization of the infections using tissue tropism, myxospore morphology and morphometry and 18S rDNA sequencing supported description of three new species: Henneguya setiuensis n. sp., Henneguya voronini n. sp. and H. calcarifer n. sp. Myxospores of all three species had typical Henneguya morphology, with two polar capsules in the plane of the suture, an oval spore body, smooth valve cell surfaces, and two caudal appendages. Spores were morphometrically similar, and many dimensions overlapped, but H. voronini n. sp. had shorter caudal appendages compared with H. calcarifer n. sp. and H. setiuensis n. sp. Gross tissue tropism distinguished the muscle parasite H. calcarifer n. sp. from gill parasites H. setiuensis n. sp. and H. voronini n. sp.; and these latter two species were further separable by fine-scale location of developing plasmodia, which were intra-lamellar for H. setiuensis n. sp. and basal to the filaments for H. voronini n. sp. small subunit ribosomal DNA sequences distinguished all three species: the two gill species H. setiuensis n. sp. and H voronini n. sp. were only 88% similar (over 1708 bp), whereas the muscle species H. calcarifer n. sp. was most similar to H. voronini n. sp. (98% over 1696 bp). None of the three novel species was more than 90% similar to any known myxosporean sequence in GenBank. Low infection prevalence of these myxosporeans and lack of obvious tissue pathology from developing plasmodia suggested none of these parasites are currently a problem for barramundi culture in Setiu Wetland; however additional surveys of fish, particularly at different times of the year, would be informative for better risk assessment.
The vp28 gene encoding an envelope protein (28 kDa) of white spot syndrome virus (WSSV) was amplified from WSSV-infected tiger shrimp that originated from Malaysia. Recombinant VP28 protein (r-28) was expressed in Escherichia coli and used as an antigen for preparation of monoclonal antibodies (MAbs). Three murine MAbs (6F6, 6H4 and 9C10) that were screened by r-28 antigen-based enzyme-linked immunosorbent assay (ELISA) were also able to recognize viral VP28 protein as well as r-28 on Western blot. Three non-overlapping epitopes of VP28 protein were determined using the MAbs in competitive ELISA; thus, an antigen-capture ELISA (Ac-ELISA) was developed by virtue of these MAbs. Ac-ELISA can differentiate WSSV-infected shrimp from uninfected shrimp and was further confirmed by a polymerase chain reaction (PCR) and Western blot. Approximately 400 pg of purified WSSV sample and 20 pg of r-28 could be detected by Ac-ELISA, which is comparable in sensitivity to PCR assay but more sensitive than Western blot in the detection of purified virus. Hemolymph and tissue homogenate samples collected from a shrimp farm in Malaysia during December 2000 and July 2001 were also detected by Ac-ELISA and PCR with corroborating results.