Comparative pathogenicity of Nocardia seriolae in Nile tilapia (Oreochromis niloticus), milkfish (Chanos chanos) and Asian seabass (Lates calcarifer).

Nocardiosis, caused by Nocardia seriolae, has been a prominent disease in Southeast Asian aquaculture in the last three decades. This granulomatous disease reported in various fish species is responsible for significant economic losses. This study investigated the pathogenicity of N. seriolae in three cultured species in Taiwan: Nile tilapia (omnivore), milkfish (herbivore) and Asian seabass (carnivore). Administration of an infective dose of 1 × 106 CFU/ fish in tilapia, seabass and milkfish demonstrated mortalities of 100%, 90% and 75%, respectively. Additionally, clinical signs namely, granuloma and lesions displayed varying intensities between the groups and pathological scores. Polymerase chain reaction (PCR) amplification specific for N. seriolae was confirmed to be positive (432 bp) using NS1/NG1 primers. Post-mortem lesions revealed the absence of granulomas in tilapia and milkfish and their presence in the seabass. Interestingly, the gut in tilapia showed an influx of eosinophils suggesting its role during the acute stages of infection. However, post-challenge, surviving milkfish exhibited granulomatous formations, while surviving seabass progressed toward healing and tissue repair within sampled tissues. Overall, in conclusion, these results demonstrate the versatility in the immunological ability of individual Perciformes to contain this pathogen as a crucial factor that influences its degree of susceptibility.

An epidemiological analysis of Nocardia seriolae isolated from a wide range of aquatic animals in Taiwan, based on their genotype and enzymatic activity.

Chronic disease following Nocardia seriolae infection in a wide range of aquatic animals has been reported in many Asian countries and recently in America and Mexico. This study aimed to investigate the epidemiological relationship among N. seriolae isolates in Taiwan by investigating their genotype and enzymatic activities. A total of 66 strains isolated from 14 known and four unknown host fish from five sites in Taiwan were characterized using five combined methods. High genotypic diversity was recognized among the isolates with 10 pulsotypes being identified from the pulsed-field gel electrophoresis method and 21 reptypes from the repetitive extragenic palindromic amplification method; however, no natural plasmids were detected in this bacterial population. Pulsotypes A8 and RI analysed by PFGE and repPCR, respectively, were found to be predominant within five sites in Taiwan over 17 years of isolation. Enzymatically, the majority of isolates displayed high leucine arylamidase, ß-glucosidase and α-glucosidase activities but were negative for lipase, α-galactosidase, ß-glucuronidase, N-acetyl-glucosaminidase, α-mannosidase and α-fucosidase activities. We identified a strong association between genotype and enzymatic activity since the majority of pulsotypes displayed the same type of enzymatic profile. This study provides comprehensive and potential epidemiological data, which will aid the fish farming activities and prevention method development.

Nocardia seriolae cell wall lipids: An effective protective mechanism in resistance and virulence.

Piscine nocardiosis, caused by Nocardia seriolae, is a refractory granulomatous disease in South-East Asian aquaculture. This study investigates the virulence of nocardial lipids essential for pathogenesis among Actinomycetes. Petroleum ether (PE) was used to selectively delipidate two groups of N. seriolae, namely, live cell (LC) and killed cell (KC); resulting in delipidated live cell (DLC) and delipidated killed cell (DKC), respectively. Changes post-delipidation on genus characteristics, such as loss in acid-fast nature and resistance to lysozyme were observed. Transmission electron microscopy revealed notable changes in the lipid layer. Additionally, Lates calcarifer, Asian seabass intraperitoneally injected with LC and DLC had mortality rates of 90% and 50%, respectively, with the latter exhibiting a delay in mortality. Reverse-transcription quantitative PCR (RT-qPCR) analysis of host cytokines from the spleen and head kidney showed delipidation contributed to the induction of an immune response with increased transcriptional levels of interferon-γ (ifn-γ). Histopathological samples collected on day 7 post-inoculation displayed a varied granulomatous response between the treatment groups and scored for pathological changes. These findings affirm that the virulence of the lipids remains independent of the living state of the cell, significantly altering the immune and granulomatous responses in L. calcarifer to N. seriolae.

PARN Knockdown in Cell Lines Results in Differential and Cell-Specific Alterations in the Expression of Cancer-Associated mRNAs.

Ribonucleases (RNases) is the collective term used for the group of enzymes that are involved in mRNA degradation. The shortening of the poly (A) tail through deadenylation is the preferred mechanism of degradation of most eukaryotic mRNAs and poly (A)-specific ribonuclease (PARN) is the most important player in deadenylation.  Besides its primarily role in mRNA stability, PARN is also involved in several non-conventional functions. It is conceivable that a decreased RNase activity can alter the stability of cancer-associated mRNAs and this alteration may be differential in cells of different origin. METHODS: The effects of siRNA-mediated knockdown of PARN on the post-transcriptional expression of 16 oncogenes and 18 tumor suppressor genes in cells derived from different lineages (NCI-H460 and NCI-H522; lung cancer) and (HEK-293; kidney) were investigated. Further, the effects of PARN depletion on proliferation and death of the lung cancer cells were investigated. RESULTS: Quantitative real time PCR analysis revealed an cell-specific alteration in the expression of the target onco and tumor suppressor genes upon PARN depletion, differently, for cells derived from different lineages. The tumor suppressor genes showed a consistent pattern of down regulation upon PARN depletion in all the three cell types tested. In contrast, the expression of oncogenes was not consistent; while some oncogenes showed overexpression in HEK 293 cells, the majority of them were downregulated in the lung cancer cells. Further, PARN depletion did not alter the proliferation of lung cancer cells, which was in contrast to previous reports. CONCLUSION: The results of this study reveal that PARN deficiency leads to an altered stability of cancer-associated mRNA, distinctly, in cells of different lineages. Despite previous reports suggesting a potential therapeutic role of PARN in cancer, our results suggest that PARN may not be an important biomarker, particularly in lung cancer.

First report on genetic characterization, cell-surface properties and pathogenicity of Lactococcus garvieae, emerging pathogen isolated from cage-cultured cobia (Rachycentron canadum).

The diseased cage-cultured cobia (Rachycentron canadum) displayed clinical signs, haemorrhagic eyes, dorsal darkness and gross pathological lesions, enlargement of spleen and liver. Haemorrhages were found in brain, heart and liver with cumulative mortality rates ranging from 20% to 50%. Extensive congestion in the heart, liver, spleen, kidney and brain was observed histopathologically. Epicarditis and meningitis were also revealed in diseased cobia. All isolates recovered from the organs (liver, spleen, head kidney, posterior kidney, brain and muscle) of cobia were found to be gram-positive, non-motile, ovoid cocci, short-chain-forming (diplococci) and α-haemolytic. The API 32 strep system together with the polymerase chain reaction assay for species-specific primers (pLG1 and pLG2) and the internal transcribed spacer (ITS) region (G1 and L1 primers) confirmed all four selected isolates as Lactococcus garvieae. Partial 16S rDNA nucleotide sequence (~1,100 bp) of one representative L. garvieae isolate AOD109191 (GenBank accession number, MW328528.1) shared 99.9% identities with the 16S rDNA nucleotide sequence of L. garvieae (GenBank accession numbers: MT604790.1). Transmission electron microscopy (TEM) evaluation of one representative L. garvieae isolate (AOD109191) and the results of multiplex PCR did not reveal the presence of the capsular gene cluster (CGC), thus categorizing the isolate as the KG+ phenotype. Capsule staining and TEM observations confirmed the presence of a hyaluronic acid-like capsule, a possible virulence factor in KG+ phenotype L. garvieae isolates. The pathogenic potential of the representative isolate (AOD109191) was assessed through intraperitoneal injection challenges in cobia. The gross lesions and histopathological changes found in experimentally infected cobia were similar to those seen in naturally infected fish. This is the first report that confirms L. garvieae-induced 'warm water lactococcsis' can cause outbreaks of diseases in cage-cultured cobia.