Perkinsus olseni in the short neck yellow clam, Paphia malabarica (Chemnitz, 1782) from the southwest coast of India.

Parasites of the genus Perkinsus predominantly infect bivalves, and two species among them, P. olseni and P. marinus, are notifiable to OIE. P. olseni infections are known to cause extensive damage to wild as well as farmed bivalves globally with enormous implications to its fishery. Consequent to the initiation of a surveillance programme for aquatic animal diseases in India, Perkinsus infections were observed in many species of bivalves. The present paper describes P. olseni infections in the short neck yellow clam, Paphia malabarica from the southwest coast of India. Diagnosis of the parasite was carried out using Ray's Fluid Thioglycollate Medium culture, histology, in-situ hybridisation and molecular taxonomy. Pathology of infection and development of zoospores is also described. This forms the first report of a P. olseni infection in P. malabarica. High prevalence and intensity of infection of Perkinsus in clams raises concerns, as clam reserves in this geographical area sustain fisheries and the livelihoods of local fishing communities.

Morphological and molecular characterization of Chloromyxum argusi n. sp. (Myxosporea) infecting the urinary bladder of Scatophagus argus Linnaeus 1766 (Scatophagidae) from the southwest coast of India.

The present paper describes a new species of Chloromyxum infecting the urinary bladder of the estuarine fish, Scatophagus argus, from the southwest coast of India. The parasite exhibited an overall prevalence of 41.93%; the prevalence is influenced by host size and seasons. Mature spores are subspherical, measure 9.40 ± 0.66 by 9.32 ± 0.87 µm, and are characterized by the presence of sutural and extra-sutural ridges, binucleated sporoplasm, and a pair of caudal extensions. Four pyriform, unequal polar capsules with raised polar filament discharge pores and ribbon-like polar filaments are present. Polar filament coils numbered four to five in large polar capsules and three in small polar capsules. Pansporoblast is irregular with granulated cytoplasm and has fine villosites on its surface. Plasmodia are spherical/irregular with monosporic and polysporic forms. In molecular and phylogenetic analysis, the myxosporean stands out with a high bootstrap value and was positioned as a sister branch of Chloromyxum kurisi. In view of the morphologic, morphometric, and molecular differences with the existing species of Chloromyxum, and considering the differences in hosts and geographic locations, the present species is treated as new and the name Chloromyxum argusi n. sp. is proposed.

Morphometric and molecular characterisation of Tenuiproboscis keralensis n. sp. infecting marine and brackish water fishes from the south-west coast of India with a note on morphological plasticity.

A new species of acanthocephalan infecting marine and brackish water fishes from the south-west coast of India is described. The parasite belongs to the genus Tenuiproboscis, and the fish hosts include Lutjanus argentimaculatus, L. ehrenbergii, Siganus javus, Epinephelus malabaricus, E. coioides, Scatophagus argus, Parascolopsis aspinosa, Caranx ignobilis, Gerres filamentosus and Lates calcarifer. The parasite inhabits mid- and hindgut regions and is characterised by an elongated, cylindrical, bulbous and posteriorly tapering metasoma and a claviform proboscis having 14-15 rows of 14-15 hooks each. Females larger than males, measured 3898.16-10,318.00 µm (6430.00 ± 1417.30) in length and 458.93-1435.68 µm (929.81 ± 250.39) in width. Males measured 3234.89-8644.20 µm (5729.50 ± 1176.60) in length and 388.30-1584.61 µm (795.88 ± 184.12) in width. Parasites recovered from different host species showed morphological/morphometric variations. However, principal component analysis (PCA) revealed significant overlapping of characters indicating their similarities. Proboscis profiling based on variations in size and position of hooks also yielded similar results. Further, in molecular phylogenetic analysis, parasites from different fish hosts formed a monophyletic clade with strong bootstrap support, again indicating their conspecific nature. These morphological/morphometric variations can be ascribed to differences in host species. Morphology and morphometrics in combination with PCA, proboscis profiling and molecular analysis suggest the present acanthocephalan parasite is different from other described species of Tenuiproboscis. Hence, it is considered as a new species and named T. keralensis n. sp. Prevalence, intensity and abundance of the parasite in different hosts are also discussed.

Molecular and morphological descriptions of Ceratomyxa collarae n. sp. and Ceratomyxa leucosternoni n. sp. from marine ornamental fishes of Indian waters.

Two novel species of Ceratomyxa infecting marine ornamental fishes from Indian waters are described. Marine ornamentals, Chaetodon collare and Chaetodon decussatus, collected from Vizhinjam, along the southwest coast of India and Acanthurus leucosternon collected from Lakshadweep islands of Arabian Sea revealed Ceratomyxa infections in their gall bladders. Mature spores of Ceratomyxa from Chaetodon collare and Chaetodon decussatus were elongate and slightly crescentic, with rounded ends, and measured 5.20 ± 0.32 µm in length and 16.32 ± 1.29 µm in thickness. Polar capsules spherical, equal in size and measured 2.23 ± 0.16 µm long and 2.24 ± 0.20 µm wide. Posterior angle measured 157.75 ± 8.650. Principle Component Analysis and molecular analysis using partial SSUrDNA sequences showed the isolates from these two hosts to be identical. Morphological, morphometric and molecular analysis using partial SSUrDNA sequences revealed the taxonomic novelty of isolates and are hence treated as Ceratomyxa collarae n. sp. Mature spores of Ceratomyxa from Acanthurus leucosternon were elongate, slightly tapering with rounded ends, and measured 7.34 ± 0.92 µm in length and 24.37 ± 2.34 µm in thickness. Shell valves were equal, joined by a narrow suture line. Polar capsules spherical in shape, equal in size, 2.59 ± 0.32 µm long and 2.46 ± 0.32 µm wide, and polar filament measured 18.68 ± 2.54 µm. Based on morphological, morphometric and molecular analyses, the present species of Ceratomyxa is distinct, considered as a new species and named Ceratomyxa leucosternoni n. sp. The paper also discusses the prevalence of the recovered parasites and host specificity of Ceratomyxa collarae n. sp.

First report of viral nervous necrosis-induced mass mortality in hatchery-reared larvae of clownfish, Amphiprion sebae Bleeker.

Frequent mortality was observed in the larval rearing facility of marine clownfish, Amphiprion sebae Bleeker. Mortality resulted in 80% loss of larval stock. Moribund larvae showed clinical signs typical of viral nervous necrosis, such as uncoordinated, corkscrew-like swimming behaviour, hypersensitivity to stimuli, darkening of body and assembly into large groups, similar to bunches of grapes. The aetiology of the disease was confirmed by gross observation of clinical signs, histopathology and molecular diagnosis. Histological studies revealed severe vacuolation in the brain and in the bipolar and ganglion layers of the eye. Molecular diagnosis by reverse transcription-polymerase chain reaction (RT-PCR) specific to piscine nodavirus yielded a positive result. The partial nucleotide sequences of the PCR-amplified fragment were 97-98% similar to other betanodavirus isolates reported globally and more closely aligned with red-spotted grouper nervous necrosis virus (RGNNV). This is the first report of susceptibility of clownfish, A. sebae, to betanodavirus and the presence of the RGNNV in India.

Mortality due to viral nervous necrosis in zebrafish Danio rerio and goldfish Carassius auratus.

Heavy mortality was observed in an experimental lot of 2 ornamental fish species, zebrafish Danio rerio (F. Hamilton, 1822) and goldfish Carassius auratus (Linnaeus, 1758). The fishes showed typical symptoms of viral nervous necrosis before death. Gross morphological examination revealed no visible lesions except in the brain, visible as a creamy opaque patch through the dorsal side of the head. Parasitic and bacteriological analysis revealed no pathogenic agents. Histopathological analysis revealed severe vacuolation in the brain and spinal cord of the samples. A fragment within the variable region of genomic RNA2 of betanodavirus was amplified from the samples by reverse transcription polymerase chain reaction using specific primers designed previously. The analysis suggests that the observed mortality in the fishes was due to betanodavirus infection. This is the first report of natural infection of betanodavirus in laboratory fishes causing viral nervous necrosis leading to mortality. The observation is alarming, as the ornamental fish culture and trade is being popularized in India where the fatal disease may cause severe setbacks in the industry. It emphasizes the need for quarantine and control strategies to prevent the spread of the virus and outbreak of the disease.

Genomic classification of betanodavirus by molecular phylogenetic analysis of the coat protein gene.

The classification of betanodavirus into four species was reviewed including newer and well-characterised isolates. Six major clusters were identified, four of which were similar to the classic species. Two single isolate clusters were worth consideration as new species.

Genetic characterization of betanodavirus isolates from Asian seabass Lates calcarifer (Bloch) in India.

Betanodavirus has been detected in Asian seabass in India. Molecular characterization of the isolates on the basis of the full-length viral RNA2 sequence was performed. Subsequent phylogenetic analysis with sequences from members of the four species in the genus Betanodavirus revealed that the present isolates are closely related to members of the species Redspotted grouper nervous necrosis virus. The analysis also revealed that the RNA2 sequence was not responsible for acute symptoms in seabass. This is the first attempt to characterize Indian isolates of fish nodaviruses, and the result will be useful for devising specific control and health-management strategies for this virus.