Thangapalam Jawahar Abraham

Detection of goldfish haematopoietic necrosis herpes virus (Cyprinid herpesvirus-2) with multi-drug resistant Aeromonas hydrophila infection in goldfish: First evidence of any viral disease outbreak in ornamental freshwater aquaculture farms in India

This outbreak report details of a mortality event where Cyprinid herpes virus-2 (CyHV-2) was detected in association with multidrug-resistant Aeromonas hydrophila infection in goldfish, Carassius auratus, from commercial farms. The goldfish exhibited large scale haemorrhages on the body, fins and gills, lepidorthosis, necrosed gills, protruded anus and shrunken eyes. White nodular necrotic foci in spleen and kidneys were noticed, along with necrosis and fusion of gill lamellae. Transmission electron microscopy of affected tissues revealed the presence of mature virus particles. Involvement of CyHV-2 was confirmed by PCR, sequencing and observed cytopathic effect in koi carp fin cell line along with experimental infection study. A bacterium isolated from the internal organs of affected fish was found to be pathogenic Aeromonas hydrophila having resistance to more than 10 classes of antibiotics. We postulate that CyHV-2 was the primary etiological agent responsible for this outbreak with secondary infection by A. hydrophila. The experimental infection trials in Labeo rohita and koi carp by intraperitoneal challenge with CyHV-2 tissue homogenates failed to reproduce the disease in those co-cultured fish species. This is the first report of a viral disease outbreak in organised earthen ornamental fish farms in India and bears further investigation.

Pathology of Edwardsiella tarda infection in African catfish, Clarias gariepinus (Burchell 1822), fingerlings

Abstract Edwardsiella tarda is one of the serious fish pathogens infecting both cultured and wild fish species. This study aimed to assess the phenotypic characterization and pathogenicity of E. tarda isolated from Clarias gariepinus (Burchell) with dropsy and histopathological alterations. The causative agent was identified with Vitek 2, and its pathogenicity was determined by intramuscular injection. The challenged catfish exhibited vertical hanging, frothing, excess mucus production, listing, swollen abdomen, anorexia, fin and tail rot, and reddish operculum. The LD50 of E. tarda PBB and PBP strains was found to be 8.52 × 106 and 1.68 × 107 cells fish-1, respectively. Histopathological observations on catfish infected naturally revealed lymphocyte infiltration in muscle and focal necrosis, hyperplasia, edema, and swelling of the gill lamellar epithelium. The kidney of diseased fish exhibited ischemic type tubulopathy, necrosis of nephritic tubules, hyperplastic hematopoietic tissue, rupture of the tubular basement membrane, hydropic dystrophy of nephritic cells, neutrophil infiltration, fibrinoid necrosis of nephretic tubules, hemosiderin deposition, and edema. The liver sections revealed lymphocyte infiltration, dilation of hepatic sinusoids, expansion of space between hepatic sinusoids, and focal necrosis. The inflammatory responses observed in kidney and liver in the present study were presumably suppuration and were attributed to the potential virulence factors of E. tarda.

Rapid Detection of Flavobacterium columnare Infection in Fish by SpeciesspecificPolymerase Chain Reaction

This study describes rapid detection of Flavobacterium columnare induced columnaris disease in cultured freshwater fish, viz., Labeo rohita, Ctenopharyngodon idella, Puntius sp. and Anabas testudineus by species-specific polymerase chain reaction. Gill discolouration, yellow necrotic areas, white patches on gill, saddle back and erosion of scales were the prominent clinical signs in all diseased fish, except Puntius sp., which had typical signs of ulcer at the base of dorsal fin. Of the nine disease cases, eight were found columnaris positive through culture independent species-specific PCR. The two sets of F. columnare specific primers such as ColF, ColR and Col72F, Col1260R yielded amplicons of around 675 bp and 1000 bp, respectively in all positive samples. Phylogenetically, the nucleotide sequences of the positive samples namely, C1 and RG1 formed monophyletic group with F. columnare, thus confirmed the infection as columnaris.

ANTIBIOTIC RESISTANT MOTILE AEROMONADS INDUCED SEPTICEMIA IN PHILIPPINE CATFISH Clarias batrachus (Linnaeus, 1758) FINGERLINGS

Philippine catfish, Clarias batrachus farming is receiving much attention in recent years so also the diseases in catfish aquaculture. During the disease surveillance in catfish farms, mortalities were observed in C. batrachus fingerlings in a nursery and this report describes the bacteriological and histopathological observations made on the diseased catfish. The gross and clinical signs observed were lethargy, anorexia, vertical movement, skin erosion, operculum erosion, pale gills, reddening of mouth, fin haemorrhage, red lateral line and distended abdomen. The bacteria isolated from the kidney were identified phenotypically as species belonging to classical motile aeromonad group (Aeromonas caviae, A. hydrophila, A. jandaei and A. sobria) and newly described aeromonad group (A. aquariorum, A. fluvialis and A. rivuli). Multiple antibiotic resistance (MAR) index was in the range of 0.3-0.8. These antibiotic resistant motile aeromonads caused septicaemia and 100% mortality. Histologically, haemocyte infiltration, necrosis, inflammation of epidermal tissue, rough epidermal layer and fibrosis in muscle tissue, and loss of typical tubular epithelial lining, necrosis of tubular tissue, inflammation of epithelial layer, cellular and nuclear hypertrophy, pycnotic nuclei, karyolysis and hypoplastic haematopoietic tissue in the kidney of diseased catfish were noted. The inflammatory responses observed in the kidney of C. batrachus were indicative of suppurative infection.

Histopathology and Wound Healing in Oxytetracycline Treated Oreochromis niloticus (L.) Against Aeromonas hydrophila Intramuscular Challenge

Antibiotics are very important tools for the control of fish bacterial diseases. Yet, there are strict regulations controlling the use of antibiotics in aquaculture. This study assessed the efficacy oxytetracycline dihydrate (OTC) at a dose of 2 g, 4 g, 6 g and 8 g/100 pounds fish/day against Aeromonas hydrophila challenge in Nile tilapia Oreochromis niloticus as well as the histopathological alterations in kidney and muscle, and wound healing. The commercial pellet feed was top dressed with OTC using 5 ml vegetable oil as a binder. The fish were injected intramuscularly with A. hydrophila at ≈1 × 108 cells/fish and then fed OTC feeds at 2% of their body weight for 10 days. The fish fed with 8g OTC/100 pounds fish/day recorded the lowest mortality (3.33%). The untreated fish recorded 8.33% mortality. Histologically, the kidney tissues of O. niloticus exhibited nephropathy and glomerulopathy. The kidney of OTC fed groups had improved organization of nephritic tubules and glomerulus. The muscle tissues exhibited haemocyte infiltration with mild necrosis initially, followed by melanization and disrupted muscle bundles. The results demonstrated that the OTC treatment for 10 days could bring out improved functioning of fish kidneys that carry infectious agents. Within 3 days of OTC therapy, tissue reddening and inflammation subsided with the formation of black scar. Full recovery of normal skin architecture was reached within 26-31 days post-injection. Based on the results, prudent use of 4-8 g OTC/100 pounds fish/day is recommended for the control of A. hydrophila infection in O. niloticus.

Meningoencephalitis in farmed monosex Nile tilapia (Oreochromis niloticus L.) caused by Streptococcus agalactiae

Abstract Aquaculture of tilapia is a new research venture in India. With intensification in farming practices, tilapia are increasingly susceptible to bacterial infections. This article describes the isolation and identification of pathogenic bacteria from cultured monosex Nile tilapia, Oreochromis niloticus (L.), that experienced moderate to severe mortalities in West Bengal, India between September and August 2014 and histopathological alterations in various organs. Gram-positive diplococci, identified as Streptococcus agalactiae with Streptococcus identification kits and 16S rDNA sequencing analysis, were isolated from the brain, operculum, and kidney. Other bacteria from the kidney were identified as Aeromonas sobria, A. caviae, Klebsiella pneumoniae ssp. pneumoniae, Escherichia coli, and Enterobacter cloacae. Staphylococcus epidermis was isolated from opercular hemorrhages. Histological sections of the infected tilapia brain revealed meningoencephalitis and granulomatous lesions. Sections from other organs indicated congestion, hemorrhagic and hyperplastic cells, necrosis, vacuolation, hemosiderin deposition, hypertrophic nuclei, melanomacrophage aggregation, and ruptured veins. This report is the first description of S. agalactiae as a primary pathogen causing meningoencephalitis in cultured tilapia in India.

Association of Aeromonas salmonicida subsp. achromogenes in the haemorrhagic blister of cultured carp Cyprinus carpio in West Bengal, India

Abstract Objective To study the association of Aeromonas salmonicida subsp. achromogenes in the haemorrhagic blister of cultured exotic carp Cyprinus carpio in West Bengal, India, its cellular fatty acid composition and antibiotic sensitivity. Methods The phenotypic characterization and antibiotic sensitivity of bacterial flora of diseased fish were performed. The whole cellular fatty acid composition of Aeromonas salmonicida ( A. salmonicida ) was determined by MIS Sherlock automatic identification system. Results Aeromonas hydrophila, A. salmonicida and Pseudomonas alcaligenes were isolated from the haemorrhagic blister. The fatty acid methyl esters pattern confirmed the A. salmonicida strain as A. salmonicida subsp. achromogenes . Unsaturated fatty acid 16:1 w7c/16:1 w6c (39.09%), saturated fatty acid 16:0 (26.84%) and the mono-unsaturated fatty acids 18:1 w7c (8.89%) and 16:1 iso I/14:0 3OH (8.49%) were the most common fatty acids, which accounted for 83.31% of the total fatty acids. A. salmonicida subsp. achromogenes was highly susceptible to broad spectrum antibiotics such as chloramphenicol, ciprofloxacin, co-trimoxazole, gentamycin, nitrofurantoin and oxytetracycline. Conclusions The finding on the presence of A. salmonicida in carps necessitates molecular level study on establishing the prevalence of this bacterium in Indian aquaculture systems especially on its free-living viable but non-culturable state.