Publications by authors named "Omkar Byadgi"

14 Publications

  • Page 1 of 1

Innate immune-gene expression during experimental amyloodiniosis in European seabass (Dicentrarchus labrax).

Vet Immunol Immunopathol 2021 Apr 21;234:110217. Epub 2021 Feb 21.

Section of Animal and Veterinary Sciences, Department of Agricultural, Food, Environmental and Animal Sciences (DI4A), University of Udine, 33100, Udine, Italy.

The ectoparasite protozoan Amyloodinium ocellatum (AO) is the causative agent of amyloodiniosis in European seabass (ESB, Dicentrarchus labrax). There is a lack of information about basic molecular immune response mechanisms of ESB during AO infestation. Therefore, to compare gene expression between experimental AO-infested ESB tissues and uninfested ESB tissues (gills and head kidney) RNA-seq was adopted. The RNA-seq revealed multiple differentially expressed genes (DEG), namely 679 upregulated genes and 360 downregulated genes in the gills, and 206 upregulated genes and 170 downregulated genes in head kidney. In gills, genes related to the immune system (perforin, CC1) and protein binding were upregulated. Several genes involved in IFN related pathways were upregulated in the head kidney. Subsequently, to validate the DEG from amyloodiniosis, 26 ESB (mean weight 14 g) per tank in triplicate were bath challenged for 2 h with AO (3.5 × 10/tank; 70 dinospores/mL) under controlled conditions (26-28 °C and 34‰ salinity). As a control group (non-infested), 26 ESB per tank in triplicate were also used. Changes in the expression of innate immune genes in gills and head kidney at 2, 3, 5, 7 and 23 dpi were analysed using real-time PCR. The results indicated that the expression of cytokines (CC1, IL-8) and antimicrobial peptide (Hep) were strongly stimulated and reached a peak at 5 dpi in the early infestation stage, followed by a gradual reduction in the recovery stage (23 dpi). Noticeably, the immunoglobulin (IgM) expression was higher at 23 dpi compared to 7 dpi. Furthermore, in-situ hybridization showed positive signals of CC1 mRNA in AO infested gills compared to the control group. Altogether, chemokines were involved in the immune process under AO infestation and this evidence allows a better understanding of the immune response in European seabass during amyloodiniosis.
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http://dx.doi.org/10.1016/j.vetimm.2021.110217DOI Listing
April 2021

Red mark syndrome (RMS) in farmed rainbow trout: First report of outbreaks in Bosnia and Herzegovina.

J Fish Dis 2021 May 21;44(5):627-631. Epub 2021 Jan 21.

School of Biosciences and Veterinary Medicine, University of Camerino, Matelica, Italy.

Red mark syndrome (RMS) is a non-lethal inflammatory skin disorder spreading in farmed adult rainbow trout (Oncorhynchus mykiss) and reported worldwide. The aetiology is still uncertain, but positive correlation was found between Midichloria-like organism and RMS-affected fish. Here, we describe the first cases of RMS in Bosnia and Herzegovina. The outbreaks under study occurred in two intensive farms during the late winter and spring of 2020. Affected fish showed signs of disease ascribable to RMS, confirmed by pathological and molecular examination.
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http://dx.doi.org/10.1111/jfd.13336DOI Listing
May 2021

Transcriptome Analysis of Tomonts Revealed Basic Information on the Major Potential Virulence Factors.

Genes (Basel) 2020 10 24;11(11). Epub 2020 Oct 24.

Section of Animal and Veterinary Sciences, Department of Agricultural, Food, Environmental and Animal Sciences (DI4A), University of Udine, 33100 Udine, Italy.

The ectoparasite protozoan (AO) is the etiological agent of amyloodiniosis in European seabass () (ESB). There is a lack of information about basic molecular data on AO biology and its interaction with the host. Therefore, de novo transcriptome sequencing of AO tomonts was performed. AO trophonts were detached from infested ESB gills, and quickly becoming early tomonts were purified by Percoll density gradient. Tomont total RNA was processed and quality was assessed immediately. cDNA libraries were constructed using TruSeq Stranded mRNA kit and sequenced using Illumina sequencer. CLC assembly was used to generate the Transcriptome assembly of AO tomonts. Out of 48,188 contigs, 56.12% belong to dinophyceae wherein had 94.61% similarity among dinophyceae. Functional annotations of contigs indicated that 12,677 had associated GO term, 9005 with KEGG term. The contigs belonging to dinophyceae resulted in the detection of several peptidases. A BLAST search for known virulent factors from the virulence database resulted in hits to Rab proteins, AP120, Ribosomal phosphoprotein, Heat-shock protein70, Casein kinases, Plasmepsin IV, and Brucipain. Hsp70 and casein kinase II alpha were characterized in-silico. Altogether, these results provide a reference database in understanding AO molecular biology, aiding to the development of novel diagnostics and future vaccines.
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http://dx.doi.org/10.3390/genes11111252DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7692099PMC
October 2020

Resistant and susceptible rainbow trout (Oncorhynchus mykiss) lines show distinctive immune response to Lactococcus garvieae.

Fish Shellfish Immunol 2020 Oct 13;105:457-468. Epub 2020 Jul 13.

Istituto Zooprofilattico Sperimentale del Piemonte, Liguria e Valle d'Aosta (IZSPLV), via Bologna 148, 10154, Torino, TO, Italy.

Lactococcosis is one of the main bacterial diseases affecting rainbow trout (Oncorhynchus mykiss), with significant economic and sanitary repercussion. Vaccination and antibiotic treatments are commonly used to prevent and control the infection outbreaks; however, these strategies have some drawbacks including limited coverage, handling costs, induction of antibiotic resistance and chemical residues in the environment. Selective breeding programs represent a promising complementary approach for increasing fish disease resistance in commercial farms and some immunological parameters may be tentatively used as indirect indicators for this purpose. The present study investigated for the first time some innate and adaptive immune responses in two groups of rainbow trout derived from selected lines (susceptible and resistant) showing a different "in field" phenotypical resistance to Yersinia ruckeri, Flavobacterium branchiophilum, F. psychrophilum, and Ichthyophthirius multifiliis, after an immersion-dilution based exposure to Lactococcus garvieae carried out in controlled experimental conditions. Twenty-six resistant and twenty-six susceptible female rainbow trout (mean body weight 80 g, 9 months aged, F5 generation) were obtained from an intensive farm considered L. garvieae free and were exposed to the pathogen. Moreover, 10 resistant and 10 susceptible fish were used as uninfected controls. After 5 days, blood and tissue samples were collected for immunological analyses. A significantly higher serum and mucus lysozyme activity was recorded in resistant rainbow trout compared to susceptible fish (P ≤ 0.05), both before and after exposure to L. garvieae. Similarly, respiratory burst activity of head kidney leukocytes resulted more intense in resistant fish (P ≤ 0.05), suggesting that phagocytes could more quickly activate their microbicidal mechanisms to counteract the bacterial spread. Resistant group displayed also an up-regulation of immunoglobulins M (IgM), major histocompatibility complex II (MHC-II) and interleukin 8 (IL-8) gene expression (P ≤ 0.05) and a significantly higher blood lymphocytes count (P ≤ 0.05), highlighting their potential better ability to trigger the recruitment of defensive cells and the initiation of specific immune processes such as antigen presentation to CD4 T lymphocytes and IgM synthesis. The results herein presented might be useful for the identification of immunological markers to be used as indirect indicators in rainbow trout selective breeding programs.
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http://dx.doi.org/10.1016/j.fsi.2020.06.040DOI Listing
October 2020

Expression of infection-related immune response in European sea bass (Dicentrarchus labrax) during a natural outbreak from a unique dinoflagellate Amyloodinium ocellatum.

Fish Shellfish Immunol 2019 Jan 25;84:62-72. Epub 2018 Sep 25.

Section of Animal and Veterinary Sciences, Department of Agricultural, Food, Environmental and Animal Sciences, University of Udine, 33100, Udine, Italy.

In the Mediterranean area, amyloodiniosis represents a major hindrance for marine aquaculture, causing high mortalities in lagoon-type based rearing sites during warm seasons. Amyloodinium ocellatum (AO) is the most common and important dinoflagellate parasitizing fish, and is one of the few fish parasites that can infest several fish species living within its ecological range. In the present study, A. ocellatum was recorded and collected from infected European sea bass (Dicentrarchus labrax) during a summer 2017 outbreak in north east Italy. Histological observation of infected ESB gill samples emphasized the presence of round or pear-shaped trophonts anchored to the oro-pharingeal cavity. Molecular analysis for small subunit (SSU) rDNA of A. ocellatum from gill genomic DNA amplified consistently and yielded 248 bp specific amplicon of A. ocellatum, that was also confirmed using sequencing and NCBI Blast analysis. Histological sections of ESB gill samples were addressed to immunohistochemical procedure for the labelling of ESB igm, inos, tlr2, tlr4, pcna and cytokeratin. Infected gills resulted positive for igm, inos, pcna and cytokeratin but negative to tlr-2 and tlr-4. Furthermore, ESB immune related gene response (innate immunity, adaptive immunity, and stress) in the course of A. ocellatum infection using quantitative polymerase chain reaction (qpcr) for infected gills and head kidney was analysed. Among the twenty three immune related gene molecules tested, cc1, il-8, il-10, hep, cox-2, cla, cat, casp9, and igt were significantly expressed in diseased fish. Altogether, these data on parasite identification and expression of host immune-related genes will allow for a better understanding of immune response in European sea bass against A. ocellatum and could promote the development of effective control measures.
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http://dx.doi.org/10.1016/j.fsi.2018.09.069DOI Listing
January 2019

Immune-Related Functional Differential Gene Expression in Koi Carp () after Challenge with .

Int J Mol Sci 2018 Jul 20;19(7). Epub 2018 Jul 20.

Department of Veterinary Medicine, College of Veterinary Medicine, National Pingtung University of Science and Technology, Pingtung 91201, Taiwan.

In order to understand the molecular basis underlying the host immune response of koi carp (), Illumina HiSeq 2000 is used to analyze the muscle and spleen transcriptome of koi carp infected with (). assembly of paired-end reads yielded 69,480 unigenes, of which the total length, average length, N50, and GC content are 70,120,028 bp, 1037 bp, 1793 bp, and 45.77%, respectively. Annotation is performed by comparison against various databases, yielding 42,229 (non-redundant protein sequence (NR): 60.78%), 59,255 (non-redundant nucleotide (NT): 85.28%), 35,900 (Swiss-Prot: 51.67%), 11,772 (clusters of orthologous groups (COG): 16.94%), 33,057 (Kyoto Encyclopedia of Genes and Genomes (KEGG): 47.58%), 18,764 (Gene Ontology (GO): 27.01%), and 32,085 (Interpro: 46.18%) unigenes. Comparative analysis of the expression profiles between bacterial challenge fish and control fish identifies 7749 differentially expressed genes (DEGs) from the muscle and 7846 DEGs from the spleen. These DEGs are further categorized with KEGG. Enrichment analysis of the DEGs and unigenes reveals major immune-related functions, including up-regulation of genes related with Toll-like receptor signaling, complement and coagulation cascades, and antigen processing and presentation. The results from RNA-Seq data are also validated and confirmed the consistency of the expression levels of seven immune-related genes after 24 h post infection with qPCR. Microsatellites (11,534), including di-to hexa nucleotide repeat motifs, are also identified. Altogether, this work provides valuable insights into the underlying immune mechanisms elicited during bacterial infection in koi carp that may aid in the future development of disease control measures in protection against .
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http://dx.doi.org/10.3390/ijms19072107DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6073842PMC
July 2018

Transcriptome analysis of immune response against Vibrio harveyi infection in orange-spotted grouper (Epinephelus coioides).

Fish Shellfish Immunol 2017 Nov 20;70:628-637. Epub 2017 Sep 20.

Department of Veterinary Medicine, College of Veterinary Medicine, National Pingtung University of Science and Technology, No. 1, Shuefu Road, Neipu, Pingtung 91201, Taiwan; International Degree Program of Ornamental Fish Science and Technology, International College, National Pingtung University of Science and Technology, No. 1, Shuefu Road, Neipu, Pingtung 91201, Taiwan. Electronic address:

Vibrio harveyi is a gram-negative bacterium reported as found in many aquaculture species. To increase knowledge of the immune response against V. harveyi, in this study we performed transcriptome analysis of head kidney and spleen in orange-spotted grouper (Epinephelus coioides) at 1 and 2 days post-infection (dpi), using the Illumina sequencing platform. After de novo assembly, a total of 79,128 unigenes was detected with an N50 of 2511 bp. After alignments with sequences recorded in the major databases (NT, NR, Swiss-Prot COG, KEGG, Interpro and GO), based on sequence similarity, 61,208 (77.4%) of the unigene total could be annotated using at least one database. Comparison of gene expression levels between V. harveyi and a control group at each time point revealed differentially expressed genes (DEGs) (P < 0.05): a total of 7918 (5536 upregulated and 2282 downregulated genes) from head kidney at 1 day post infection (dpi), 4260 (1444 upregulated and 2816 downregulated genes) from head kidney at 2 dpi, 7887 (4892 upregulated and 2995 downregulated genes) from spleen at 1 dpi, and 8952 (7388 upregulated and 1564 downregulated genes) from spleen at 2 dpi. The DEGs were mainly annotated into signal transduction and immune system categories, based on the KEGG database. The DEGs were enriched in immune-related pathway functions, NOD-like receptor signaling pathways, Toll-like receptor signaling pathways, NF-κB signaling pathways, and Jak-STAT signaling pathways. Additionally, we selected several DEGs and validated their expression level by RT-qPCR. The data generated in this study may provide a valuable resource for further immune response research and offer improved strategies against V. harveyi infection in teleost fishes.
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http://dx.doi.org/10.1016/j.fsi.2017.09.052DOI Listing
November 2017

Transcriptome analysis of grey mullet (Mugil cephalus) after challenge with Lactococcus garvieae.

Fish Shellfish Immunol 2016 Nov 5;58:593-603. Epub 2016 Oct 5.

Department of Veterinary Medicine, College of Veterinary Medicine, National Pingtung University of Science and Technology, Pingtung, 91201, Taiwan, ROC. Electronic address:

Grey mullet (Mugil cephalus) is an economically important fish species in Taiwan mariculture industry. Moreover, grey mullet are common hosts of a bacterial infection by Lactococcus garvieae. However, until now the information related to the immune system of grey mullet is unclear. Therefore, to understand the molecular basis underlying the host immune response to L. garvieae infection, Illumina HiSeq™ 2000 was used to analyse the head kidney and spleen transcriptome of infected grey mullet. De novo assembly of paired-end reads yielded 55,203 unigenes. Comparative analysis of the expression profiles between bacterial challenge fish and control fish identified a total of 7192 from head kidney and 7280 in spleen differentially expressed genes (P < 0.05), including 4211 upregulated genes and 2981 downregulated genes in head kidney, while in spleen 3598 genes were upregulated and 3682 downregulated. A significant enrichment analysis of these differentially expressed genes (DEG) in spleen and head kidney revealed major immune-related pathways, including complement and coagulation cascades, Toll-like receptor signalling, and antigen processing and presentation. Moreover, selected DEGs were validated using qPCR. Altogether, the results obtained on immune-related genes may allow for a better understanding of immunity in grey mullet to Lactococcus garvieae, carrying out detailed functional analysis of these genes and developing strategies for efficient immune protection against infections in grey mullet.
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http://dx.doi.org/10.1016/j.fsi.2016.10.006DOI Listing
November 2016

Identification, Molecular Cloning of IL-1β and Its Expression Profile during Nocardia seriolae Infection in Largemouth Bass, Micropterus salmoides.

Int J Mol Sci 2016 Oct 1;17(10). Epub 2016 Oct 1.

Department of Veterinary Medicine, College of Veterinary Medicine, National Pingtung University of Science and Technology, Pingtung 912, Taiwan.

In the present study, IL-1β cDNA was identified and analyzed from largemouth bass (). Full length IL-1β mRNA was obtained using Rapid Amplification of cDNA Ends (RACE), which contains 78 bp 3'-UTR, a 455 bp 5'-UTR, and an open reading frame (ORF) of 702 bp coding for 233 amino acid residues. The molecular weight and theoretical isoelectric point of largemouth bass IL-1β protein was predicted to be 26.7 kDa and 6.08 respectively. A largemouth bass IL-1β phylogenetic analysis showed a close relation to the IL-1βs of striped trumpeter (), Chinese perch (), and Japanese sea bass (). Peptidoglycan upregulated IL-1β in the spleen and head kidney, while lipopolysaccharide upregulated detectable levels of IL-1β in the spleen only. Largemouth bass, challenged with (1.0 × 10⁶ cfu/mL), showed a significant increase in IL-1β at 3 and 5 days post infection (dpi) in the spleen, while in the head kidney significant expression was found at 2 and 3 dpi, peaking at 3 dpi. Furthermore, tumor necrosis factor α (TNF-α) showed significantly higher expression in the spleen at 3 and 5 dpi, and in the head kidney at 1 and 3 dpi, with expression decreasing at 5 dpi in both tissues.
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http://dx.doi.org/10.3390/ijms17101670DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5085703PMC
October 2016

De Novo Transcriptome Analysis of Differential Functional Gene Expression in Largemouth Bass (Micropterus salmoides) after Challenge with Nocardia seriolae.

Int J Mol Sci 2016 Aug 11;17(8). Epub 2016 Aug 11.

Department of Veterinary Medicine, College of Veterinary Medicine, National Pingtung University of Science and Technology, Pingtung 91201, Taiwan.

Largemouth bass (Micropterus salmoides) are common hosts of an epizootic bacterial infection by Nocardia seriolae. We conducted transcriptome profiling of M. salmoides to understand the host immune response to N. seriolae infection, using the Illumina sequencing platform. De novo assembly of paired-end reads yielded 47,881 unigenes, the total length, average length, N50, and GC content of which were 49,734,288, 1038, 1983 bp, and 45.94%, respectively. Annotation was performed by comparison against non-redundant protein sequence (NR), non-redundant nucleotide (NT), Swiss-Prot, Clusters of Orthologous Groups (COG), Kyoto Encyclopaedia of Genes and Genomes (KEGG), Gene Ontology (GO), and Interpro databases, yielding 28,964 (NR: 60.49%), 36,686 (NT: 76.62%), 24,830 (Swissprot: 51.86%), 8913 (COG: 18.61%), 20,329 (KEGG: 42.46%), 835 (GO: 1.74%), and 22,194 (Interpro: 46.35%) unigenes. Additionally, 8913 unigenes were classified into 25 Clusters of Orthologous Groups (KOGs) categories, and 20,329 unigenes were assigned to 244 specific signalling pathways. RNA-Seq by Expectation Maximization (RSEM) and PossionDis were used to determine significantly differentially expressed genes (False Discovery Rate (FDR) < 0.05) and we found that 1384 were upregulated genes and 1542 were downregulated genes, and further confirmed their regulations using reverse transcription quantitative PCR (RT-qPCR). Altogether, these results provide information on immune mechanisms induced during bacterial infection in largemouth bass, which may facilitate the prevention of nocardiosis.
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http://dx.doi.org/10.3390/ijms17081315DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5000712PMC
August 2016

Evaluation of biofilm of Aeromonas hydrophila for oral vaccination of Channa striatus.

Fish Shellfish Immunol 2014 Dec 22;41(2):581-5. Epub 2014 Sep 22.

Aquatic Health Management Laboratory, Department of Aquaculture, College of Fisheries, Karnataka Veterinary, Animal and Fisheries Sciences University, Mangalore 575002, Karnataka, India.

Our laboratory has developed a biofilm oral vaccine of Aeromonas hydrophila, which has given significantly higher antibody agglutination titre and protection in herbivorous carps and omnivorous walking catfish compared to that with free cell vaccine. Against this background, in the present study A. hydrophila biofilm oral vaccine was evaluated in Channa striatus, a carnivorous fish model. The fish was fed with biofilm (BF) and free cell (FC) of A. hydrophila vaccine at 10(10) cells/g fish/day for 20 d. Serum antibody production monitored with a monoclonal antibody based ELISA for 60 day post vaccination. Significantly higher antibody titre was recorded with BF compared to that with FC. Furthermore, BF vaccinated fish upon challenge with A. hydrophila at 10(9) cfu/ml had significantly higher relative per cent survival (88) than that with FC (29.6).
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http://dx.doi.org/10.1016/j.fsi.2014.09.021DOI Listing
December 2014

The effect of TLR9 agonist CpG oligodeoxynucleotides on the intestinal immune response of cobia (Rachycentron canadum).

J Immunol Res 2014 2;2014:273284. Epub 2014 Jun 2.

Department of Tropical Agriculture and International Cooperation, National Pingtung University of Science and Technology, Pingtung 91201, Taiwan.

Cytosine-guanine oligodeoxynucleotide (CpG ODN) motifs of bacterial DNA are recognized through toll-like receptor 9 (TLR9) and are potent activators of innate immunity. However, the interaction between TLR9 and CpG ODN in aquatic species has not been well characterized. Hence, cobia TLR9 isoform B (RCTLR9B) was cloned and its expression and induction in intestine were investigated. RCTLR9B cDNA consists of 3113bp encoding 1009 amino acids containing three regions, leucine rich repeats, transmembrane domain, and toll/interleukin-1 receptor (TIR) domain. Intraperitoneal injection of CpG ODN 2395 upregulated RCTLR9 A and B and MyD88 and also induced the expressions of Mx, chemokine CC, and interleukin IL-1 β . Cobia intraperitoneally injected with CpG ODN 1668 and 2395 had increased survival rates after challenge with Photobacterium damselae subsp. piscicida. In addition, formulation of CpG ODN with formalin-killed bacteria (FKB) and aluminum hydroxide gel significantly increased expressions of RCTLR9 A (50 folds) and B (30 folds) isoforms at 10 dpi (CpG ODN 1668) and MyD88 (21 folds) at 6 dpv (CpG ODN 2395). Subsequently, IL-1 β increased at 6 dpv in 1668 group. No histopathological damage and inflammatory responses were observed in the injected cobia. Altogether, these results facilitate CpG ODNs as an adjuvant to increase bacterial disease resistance and efficacy of vaccines in cobia.
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http://dx.doi.org/10.1155/2014/273284DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4060301PMC
February 2015

Identification and expression analysis of cobia (Rachycentron canadum) Toll-like receptor 9 gene.

Fish Shellfish Immunol 2014 Feb 27;36(2):417-27. Epub 2013 Dec 27.

Department of Tropical Agriculture and International Cooperation, National Pingtung University of Science and Technology, Pingtung, Taiwan. Electronic address:

Cobia culture is hindered by bacterial infection (Photobacterium damselae subsp. piscicida) and in order to study the effect of P. damselae subsp. piscicida challenge and CpG ODN stimulation on cobia Toll like receptor 9 (RCTLR9), we used PCR to clone RCTLR9 gene and qRT-PCR to quantify gene expression. The results indicated that RCTLR9 cDNA contains 3141 bp. It encodes 1047 amino acids containing 16 typical structures of leucine-rich repeats (LRRs) including an LRRTYP, LRRCT and a motif involved in PAMP binding was identified at position 240-253 amino acid. Broad expression of RCTLR9 was found in larval, juvenile and adult stages irrespective of the tissues. In larval stage, RCTLR9 mRNA expression decreased at 5 d and then increased at 10 dph. At juvenile stage cobia, the expression was significantly high (p < 0.05) in spleen and intestine compared to gill, kidney, liver and skin. However, at adult stage, the significant high expression was found in gill and intestine. Cobia challenged with P. damselae subsp. piscicida showed significant increase in RCTLR9 expression at 24 h post challenge in intestine, spleen and liver, while in kidney the expression was peak at 12 h and later it decreased at 24 h. The highest expression was 40 fold increase in spleen and the lowest expression was ∼3.6 fold increase in liver. Cobia stimulated with CpG oligonucleotides showed that the induction of these genes was CpG ODN type and time dependent. In spleen and liver, CpG ODNs 1668 and 2006 injected group showed high expression of RCTLR9, IL-1β, chemokine CC compared to other groups. Meanwhile, CpG ODN 2006 has induced high expression of IgM. The CpG ODNs 2395 have induced significant high expression of Mx in spleen and liver. These results demonstrates the potential of using CpG ODN to enhance cobia resistance to P. damselae subsp. piscicida infection and use as an adjuvant in vaccine development.
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http://dx.doi.org/10.1016/j.fsi.2013.12.017DOI Listing
February 2014

Development and standardization of a monoclonal antibody-based rapid flow-through immunoassay for the detection of Aphanomyces invadans in the field.

J Vet Sci 2013 30;14(4):413-9. Epub 2013 Jun 30.

Aquatic Health Management Laboratory, Department of Aquaculture, College of Fisheries, Karnataka Veterinary, Animal and Fisheries Sciences University, Mangalore 575002, India.

A monoclonal antibody-based flow-through immunoassay (FTA) was developed using a nitrocellulose membrane placed on the top of adsorbent pads enclosed in a plastic cassette with a test zone at the center. The FTA could be completed within 10 min. Clear purple dots against a white background indicated the presence of Aphanomyces (A.) invadans. The FTA limit of detection was 7 μg/mL for A. invadans compared to 56 μg/mL for the immunodot. FTA and polymerase chain reaction (PCR) could detect A. invadans in fish tissue homogenates at a 10(-11) dilution compared to a 10(-8) dilution by immunodot. In fish suffering from natural cases of epizootic ulcerative syndrome (EUS) collected from Mangalore, India, FTA and PCR could detect A. invadans in 100% of the samples compared to 89.04% detected by immunodot. FTA reagents were stable and produced expected results for 4 months when stored at 4~8°C. This rapid test could serve as simple and cost-effective on-site screening tool to detect A. invadans in fish from EUS outbreak areas and in ports during the shipment of live or frozen fish.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3885734PMC
http://dx.doi.org/10.4142/jvs.2013.14.4.413DOI Listing
August 2014
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