Materials and Methods: A batch of newly hatched hybrid grouper fry (Epinephelus fuscoguttatus × Epinephelus lanceolatus) were followed from the larval stage to market size. Samples of the hybrid groupers, water, live feed, and artificial fish pellets were collected periodically from day 0 to 180 in the hybrid grouper hatchery. Reverse transcription-polymerase chain reaction (RT-PCR) and nested PCR amplifications were carried out on VNN-related sequences. The phylogenetic tree including the sampled causative agent of VNN was inferred from the coat protein genes from all known Betanodavirus species using Molecular Evolutionary Genetics Analysis (MEGA). Pearson's correlation coefficient values were calculated to determine the strength of the correlation between the presence of VNN in hybrid grouper samples and its associated risk factors.
Results: A total of 113 out of 146 pooled and individual samples, including hybrid grouper, water, and artificial fish pellet samples, demonstrated positive results in tests for the presence of VNN-associated viruses. The clinical signs of infection observed in the samples included darkened skin, deformation of the backbone, abdominal distension, skin lesions, and fin erosion. VNN was present throughout the life stages of the hybrid groupers, with the first detection occurring at day 10. VNN-associated risk factors included water temperature, dissolved oxygen content, salinity, ammonia level, fish size (adults more at risk than younger stages), and life stage (age). Detection of VNN-associated viruses in water samples demonstrated evidence of horizontal transmission of the disease. All the nucleotide sequences found in this study had high nucleotide identities of 88% to 100% to each other, striped jack nervous necrosis virus (SJNNV), and the reassortant strain red-spotted grouper NNV/SJNNV (RGNNV/SJNNV) isolate 430.2004 (GenBank accession number JN189932.1) (n=26). The phylogenetic analysis showed that quasispecies was present in each VNN-causing virus-positive sample, which differed based on the type of sample and life stage.
Conclusion: This study was the first to confirm the existence of a reassortant strain (RGNNV/SJNNV) in hybrid groupers from Malaysia and Southeast Asia. However, the association between the mode of transmission and the risk factors of this virus needs to be investigated further to understand the evolution and potential new host species of the reassortant strain.
Materials and Methods: A. hydrophila was biochemically identified and subjected to antibiotic susceptibility tests. The isolate was then intraperitoneally injected into red hybrid tilapia, and the mortality, clinicopathological changes, and LD50 were determined up to 240 h post-infection (hpi).
Results: The isolate demonstrated multiple antibiotic resistances (MAR) toward amikacin, ampicillin, cefotaxime, amoxicillin, trimethoprim-sulfamethoxazole, erythromycin, and streptomycin, with a MAR index of 0.5. The experimental infection of A. hydrophila at 105 CFU/mL in the red hybrid tilapia resulted in 100% mortality at 240 hpi. The LD50 was determined at 1.1×104 CFU/mL. Infected fish demonstrated occasional erratic swimming patterns, localized hemorrhages and depigmentation on the body and operculum areas, fin erosion, enlargement of the gall bladder, and hemorrhage in internal organs. Microscopic observation of infected fish revealed brain congestion, tubular necrosis, and glomerular shrinkage in the kidneys, necrosis of hepatocytes, and congestion of blood vessels in the liver.
Conclusion: The high virulence of A. hydrophila to the red hybrid tilapia emphasizes the importance of active, on-going monitoring of its prevalence in Malaysian tilapia farming.
METHODS: In this study, we extracted retrospective data from Malaysian surveillance database. Analysis was carried out using WHONET software focusing noncholera Vibrio spp. including Vibrio parahaemolyticus, Vibrio vulnificus, Vibrio fluvialis, Vibrio alginolyticus, Vibrio hollisae (Grimontia hollisae), Vibrio mimicus, Vibrio metschnikovii, and Vibrio furnissii.
RESULTS: Here, we report the first distribution and prevalence of these species isolated in Malaysia together with the antibiotic sensitivity profile based on the species. We found that V. parahaemolyticus is the predominant species isolated in Malaysia. Noticeably, across the study period, V. fluvialis is becoming more prevalent, as compared to V. parahaemolyticus. In addition, this study also reports the first isolation of pathogenic V. furnissii from stool in Malaysia.
CONCLUSION: These data represent an important step toward understanding the potential emergence of noncholera Vibrio spp. outbreaks.
METHODOLOGY/PRINCIPAL FINDINGS: We performed a literature review, whole genome sequencing on 145 GBS isolates collected from six Southeast Asian countries, and phylogenetic analysis on 7,468 GBS sequences including 227 variants of ST283 from humans and animals. Although almost absent outside Asia, ST283 was found in all invasive Asian collections analysed, from 1995 to 2017. It accounted for 29/38 (76%) human isolates in Lao PDR, 102/139 (73%) in Thailand, 4/13 (31%) in Vietnam, and 167/739 (23%) in Singapore. ST283 and its variants were found in 62/62 (100%) tilapia from 14 outbreak sites in Malaysia and Vietnam, in seven fish species in Singapore markets, and a diseased frog in China.
CONCLUSIONS: GBS ST283 is widespread in Southeast Asia, where it accounts for a large proportion of bacteraemic GBS, and causes disease and economic loss in aquaculture. If human ST283 is fishborne, as in the Singapore outbreak, then GBS sepsis in Thailand and Lao PDR is predominantly a foodborne disease. However, whether transmission is from aquaculture to humans, or vice versa, or involves an unidentified reservoir remains unknown. Creation of cross-border collaborations in human and animal health are needed to complete the epidemiological picture.