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The present study was intended to screen the wild crustaceans for co-infection with Infectious Hypodermal and Hematopoietic Necrosis Virus (IHHNV) and White Spot Syndrome Virus (WSSV) in Andaman and Nicobar Archipelago, India. We screened a total of 607 shrimp and 110 crab samples using a specific polymerase chain reaction, and out of them, 82 shrimps (13.5%) and 5 (4.5%) crabs were found positive for co-infection of IHHNV and WSSV. A higher rate of co-infection was observed in Penaeus monodon and Scylla serrata than other shrimp and crab species. The nucleotide sequences of IHHNV and WSSV obtained from crab in this present study exhibited very high sequence identity with their counterparts retrieved from various countries. Histopathological analysis of the infected shrimp gill sections further confirmed the eosinophilic intra-nuclear cowdry type A inclusion bodies and basophilic intra-nuclear inclusion bodies characteristics of IHHNV and WSSV infections, respectively. The present study serves as the first report on co-infection of WSSV and IHHNV in Andaman and Nicobar Archipelago, India and accentuates the critical need for continuous monitoring of wild crustaceans and appropriate biosecurity measures for brackishwater aquaculture.
Kandasamy Saravanan; Jayasimhan Praveenraj; Rajendran Kiruba-Sankar; Varsha Devi; Utpal Biswas; Thangaraj Kumar; Arun Sudhagar; Mansour El-Matbouli; Gokhlesh Kumar. Co-Infection of Infectious Hypodermal and Hematopoietic Necrosis Virus (IHHNV) and White Spot Syndrome Virus (WSSV) in the Wild Crustaceans of Andaman and Nicobar Archipelago, India. Viruses 2021, 13, 1378 .
AMA StyleKandasamy Saravanan, Jayasimhan Praveenraj, Rajendran Kiruba-Sankar, Varsha Devi, Utpal Biswas, Thangaraj Kumar, Arun Sudhagar, Mansour El-Matbouli, Gokhlesh Kumar. Co-Infection of Infectious Hypodermal and Hematopoietic Necrosis Virus (IHHNV) and White Spot Syndrome Virus (WSSV) in the Wild Crustaceans of Andaman and Nicobar Archipelago, India. Viruses. 2021; 13 (7):1378.
Chicago/Turabian StyleKandasamy Saravanan; Jayasimhan Praveenraj; Rajendran Kiruba-Sankar; Varsha Devi; Utpal Biswas; Thangaraj Kumar; Arun Sudhagar; Mansour El-Matbouli; Gokhlesh Kumar. 2021. "Co-Infection of Infectious Hypodermal and Hematopoietic Necrosis Virus (IHHNV) and White Spot Syndrome Virus (WSSV) in the Wild Crustaceans of Andaman and Nicobar Archipelago, India." Viruses 13, no. 7: 1378.
Hepatopancreatic microsporidiosis (HPM) is an infectious shrimp disease caused by the microsporidian Enterocytozoon hepatopenaei (EHP). In recent years, the widespread occurrence of EHP poses a significant challenge to the shrimp aquaculture industry. Early, rapid and accurate diagnosis of EHP infection is very much essential for the control of HPM crop-related losses. Loop-mediated isothermal amplification (LAMP) is a robust, sensitive, cost-effective disease diagnostic technique. Here, we demonstrate an improved, simple, closed-tube, colorimetric EHP LAMP diagnostic assay. LAMP assay was illustrated with the specific EHP spore wall protein (SWP) gene primers. Naked eye visual detection of LAMP amplicons was achieved using Hydroxy naphthol blue (HNB) or Phenol red dye without opening the tubes. This LAMP assay is efficient in detecting the EHP pathogen in all clinical samples include shrimp hepatopancreas, FTA card samples, feces, pond water, and soil. Also, the elution of EHP DNA from FTA cards was demonstrated within 17 min using a simple dry bath. In clinical evaluation, the visual LAMP assay established 100% diagnostic sensitivity and 100% diagnostic specificity. The visual LAMP assay is rapid, can detect the EHP pathogen within 40 min using a simple dry bath, and does not require any expensive instruments and technical proficiency. In conclusion, this visual LAMP protocol is a user-friendly, specific assay that can be conceivably operated at the farm-site/ resource-limited settings by the farmer himself with simple equipment.
T. Sathish Kumar; K. Radhika; J. Joseph Sahaya Rajan; M. Makesh; S.V. Alavandi; K.K. Vijayan. Closed-tube field-deployable loop-mediated isothermal amplification (LAMP) assay based on spore wall protein (SWP) for the visual detection of Enterocytozoon hepatopenaei (EHP). Journal of Invertebrate Pathology 2021, 183, 107624 .
AMA StyleT. Sathish Kumar, K. Radhika, J. Joseph Sahaya Rajan, M. Makesh, S.V. Alavandi, K.K. Vijayan. Closed-tube field-deployable loop-mediated isothermal amplification (LAMP) assay based on spore wall protein (SWP) for the visual detection of Enterocytozoon hepatopenaei (EHP). Journal of Invertebrate Pathology. 2021; 183 ():107624.
Chicago/Turabian StyleT. Sathish Kumar; K. Radhika; J. Joseph Sahaya Rajan; M. Makesh; S.V. Alavandi; K.K. Vijayan. 2021. "Closed-tube field-deployable loop-mediated isothermal amplification (LAMP) assay based on spore wall protein (SWP) for the visual detection of Enterocytozoon hepatopenaei (EHP)." Journal of Invertebrate Pathology 183, no. : 107624.
The emerging microsporidian parasite Enterocytozoon hepatopenaei (EHP), the causative agent of hepatopancreatic microsporidiosis, has been widely reported in shrimp-farming countries. EHP infection can be detected by light microscopy observation of spores (1.7 × 1 μm) in stained hepatopancreas (HP) tissue smears, HP tissue sections, and fecal samples. EHP can also be detected by polymerase chain reaction (PCR) targeting the small subunit (SSU) ribosomal RNA (rRNA) gene or the spore wall protein gene (SWP). In this study, a rapid, sensitive, specific, and closed tube visual loop-mediated isothermal amplification (LAMP) protocol combined with FTA cards was developed for the diagnosis of EHP. LAMP primers were designed based on the SSU rRNA gene of EHP. The target sequence of EHP was amplified at constant temperature of 65 °C for 45 min and amplified LAMP products were visually detected in a closed tube system by using SYBR™ green I dye. Detection limit of this LAMP protocol was ten copies. Field and clinical applicability of this assay was evaluated using 162 field samples including 106 HP tissue samples and 56 fecal samples collected from shrimp farms. Out of 162 samples, EHP could be detected in 62 samples (47 HP samples and 15 fecal samples). When compared with SWP-PCR as the gold standard, this EHP LAMP assay had 95.31% sensitivity, 98.98% specificity, and a kappa value of 0.948. This simple, closed tube, clinically evaluated visual LAMP assay has great potential for diagnosing EHP at the farm level, particularly under low-resource circumstances.
Sathish Kumar T; Navaneeth Krishnan A; Joseph Sahaya Rajan J; Makesh M; Jithendran K. P; Alavandi S. V; Vijayan K. K. Visual loop-mediated isothermal amplification (LAMP) for the rapid diagnosis of Enterocytozoon hepatopenaei (EHP) infection. Parasitology Research 2018, 117, 1485 -1493.
AMA StyleSathish Kumar T, Navaneeth Krishnan A, Joseph Sahaya Rajan J, Makesh M, Jithendran K. P, Alavandi S. V, Vijayan K. K. Visual loop-mediated isothermal amplification (LAMP) for the rapid diagnosis of Enterocytozoon hepatopenaei (EHP) infection. Parasitology Research. 2018; 117 (5):1485-1493.
Chicago/Turabian StyleSathish Kumar T; Navaneeth Krishnan A; Joseph Sahaya Rajan J; Makesh M; Jithendran K. P; Alavandi S. V; Vijayan K. K. 2018. "Visual loop-mediated isothermal amplification (LAMP) for the rapid diagnosis of Enterocytozoon hepatopenaei (EHP) infection." Parasitology Research 117, no. 5: 1485-1493.