Bacillary Necrosis Research Articles

Sàng lọc các chủng vi khuẩn lactic từ ruột một số loài cá da trơn có tiềm năng sử dụng làm probiotic

The research was conduted to isolate and screen acid lactic bacteria which had antimicrobial activity against pathogens Edwarsiella ictaluri and Aeromonas hydrophila causing Bacillary necrosis pangasius (BNP) and Hemorrhagic disease in cultured Tra catfish in the gastroinstestinal tracts of wild freshwater catfish. As the result, 96 strains of lactic acid bacteria (LAB) were selected which had inhibitory activity against indicator bacteria Escherichia coli with 29 strains isolated from Stripped catfish (Pangasius hypophthalmus (Sauvage, 1878)), 24 strains from Mystus nemurus (Valenciennes, 1839), 21 strains from Pangasius larnaudii (Bocourt, 1866), 12 strains from Clarias macrocephalus (Gunther, 1864) and 8 strains from Pangasius larnaudii (Bocourt, 1866). The result of gram staining and biochemical characteristic tests illustrated most of selected strains of lactic acid bacteria were gram-positive; spherical, oval, short rod or long rod shape; negative oxidase and negative catalase. Result from determination of antagonist activity and bacteriocin producing, there were 46 strains showed the inhibitory activity against A. hydrophia, 42 strains could inhibit E. ictaluri which were Gram-possitive, spherical or rod in shape and e specially, three strains of obtained LAB presented the antibacterial activities (produced bacteriocin) which all of them inhibited A.hydrophia and only one of three strains could inhibit E. ictaluri when the culture supernatants were neutralized to pH 6.5-7.0. Nucleotide sequences of their 16sRNA showed 100% (548/548bp) and 99% (551/552bp) homology to Lactobacillus reuteri HFI-LD5 CL2, CL 2.20, respectively and 100% (539/539bp) homology to Lactobacillus fermentum JCM- 1173 CT3.7.

Genetic diversity of culturable Vibrio in an Australian blue mussel Mytilus galloprovincialis hatchery

Bacillary necrosis associated with Vibrio species is the common cause of larval and spat mortality during commercial production of Australian blue mussel Mytilus galloprovincialis. A total of 87 randomly selected Vibrio isolates from various stages of rearing in a commercial mussel hatchery were characterised using partial sequences of the ATP synthase alpha subunit gene (atpA). The sequenced isolates represented 40 unique atpA genotypes, overwhelmingly dominated (98%) by V. splendidus group genotypes, with 1 V. harveyi group genotype also detected. The V. splendidus group sequences formed 5 moderately supported clusters allied with V. splendidus/V. lentus, V. atlanticus, V. tasmaniensis, V. cyclitrophicus and V. toranzoniae. All water sources showed considerable atpA gene diversity among Vibrio isolates, with 30 to 60% of unique isolates recovered from each source. Over half (53%) of Vibrio atpA genotypes were detected only once, and only 7 genotypes were recovered from multiple sources. Comparisons of phylogenetic diversity using UniFrac analysis showed that the culturable Vibrio community from intake, header, broodstock and larval tanks were phylogenetically similar, while spat tank communities were different. Culturable Vibrio associated with spat tank seawater differed in being dominated by V. toranzoniae-affiliated genotypes. The high diversity of V. splendidus group genotypes detected in this study reinforces the dynamic nature of microbial communities associated with hatchery culture and complicates our efforts to elucidate the role of V. splendidus group bacteria in vibriosis.

An evaluation of fish health-management practices and occupational health hazards associated with Pangasius catfish (Pangasianodon hypophthalmus) aquaculture in the Mekong Delta, Vietnam

This study aimed to evaluate the current status on the use of probiotics, disinfectants and antimicrobials in hatcheries, nurseries and grow-out farms producing Pangasius catfish (Pangasianodon hypophthalmus) in the Mekong Delta, Vietnam. Semi-structured interviews were conducted with 83 aquaculture enterprises (15 hatcheries, 32 nurseries and 36 grow-out farms). Farmers reported use of a total of 24 different antimicrobials, e.g. for treatment of bacillary necrosis and motile aeromonad septicaemia, and a variety of disinfectants, probiotics and nutritional supplements. In contrast to small-scale farmers, all large-scale grow-out farmers studied were certified and therefore had higher levels of formal education and specialized aquaculture training to diagnose and treat diseases. All farmers prepared their own medicated feed with a high risk of treatment failure, negative environmental impact from released antimicrobials and resistance development. Small-scale farmers were at particular occupational health risks when handling antimicrobials and other chemicals, e.g. mixing medicated feed with bare hands. There is an urgent need to improve knowledge and use innovative approaches, e.g. private-public partnerships, to assure a prudent use of chemicals, to improve capacity and access to disease diagnosis, particularly for small-scale grow-out farmers and nurseries. Efforts to control use of antimicrobials in aquaculture should be coordinated with the livestock and human health sectors taking an One-Health approach.

Factors other than metalloprotease are required for full virulence of French Vibrio tubiashii isolates in oyster larvae.

Vibrio tubiashii is a marine pathogen isolated from larval and juvenile bivalve molluscs that causes bacillary necrosis. Recent studies demonstrated the isolation of this species in a French experimental hatchery/nursery affecting Crassostrea gigas spat in 2007. Here, using larvae of C. gigas as an interaction model, we showed that the French V. tubiashii is virulent to larvae and can cause bacillary necrosis symptoms with an LD50 of about 2.3 × 10(3) c.f.u. ml(-1) after 24 h. Moreover, complete or gel permeation HPLC fractionated extracellular products (ECPs) of this strain appeared toxic to larvae. MS-MS analysis of the different ECP fractions revealed the existence of an extracellular metalloprotease and other suspected virulence factors. This observation is also supported by the expression level of some potential virulence factors. The overall results suggest that the pathology caused by the French V. tubiashii in C. gigas oysters is caused by a group of toxic factors and not only the metalloprotease.

Early interactions of Edwardsiella ictaluri, with Pangasianodon catfish and its invasive ability in cell lines

Commercial Pangasianodon catfish production is heavily impacted by Bacillary Necrosis of Pangasius (BNP) caused by Edwardsiella ictaluri. This study aimed to investigate the early bacterium-host interactions following immersion challenge and to compare the retrieved data with the invasion ability of the used isolates in fish cell lines. Firstly, Pangasianodon hypophthalmus fingerlings were challenged via immersion using E. ictaluri isolate HO2 or 223. At different times post inoculation, fish were sacrificed and gill and internal organ samples were taken for bacteriological, histological and immunohistochemical evaluation. The bacterial load was higher for fish inoculated with isolate HO2 compared with 223. Histological and immunohistochemical analysis revealed multifocal necrotic areas in kidney, spleen and liver of HO2 inoculated fish at 72 h post inoculation with short rod-shaped immunoperoxidase positive bacteria clustered inside cells respectively. Bacteria especially were present in the gills and intestinal tract of HO2 inoculated fish, suggesting the gastrointestinal tract and gills act as portals of entry. Following, the ability of HO2, 223 and four additional isolates to invade a Chinook salmon embryo cell line, a fat head minnow cell line and a rainbow trout liver cell line was tested. All E. ictaluri isolates were invasive in all cell lines albeit at different degrees. Isolate HO2 was highly invasive in all cell lines with a significantly higher invasion capacity than isolate 223 in the Chinook salmon embryo cell line. A correlation between in vivo virulence and in vitro invasiveness hence is suggested although further studies are needed to confirm this hypothesis.

Application of Biocontrol Techniques to Prevent Bacillary Necrosis in Larval Oysters Caused by Vibrio Infection

Application of Biocontrol Techniques to Prevent Bacillary Necrosis in Larval Oysters Caused by Vibrio Infection

Experimental challenge studies in Vietnamese catfish, Pangasianodon hypophthalmus (Sauvage), exposed to Edwardsiella ictaluri and Aeromonas hydrophila

The two main diseases in the pangasius catfish industry are bacillary necrosis of Pangasianodon (BNP) and motile aeromonas septicaemia (MAS), where the aetiological agents have been identified as Edwardsiella ictaluri and Aeromonas hydrophila, respectively. In this study, apparently healthy Pangasianodon hypophthalmus were exposed to E. ictaluri, A. hydrophila or both bacterial species by intraperitoneal injection or immersion. There were 20 fish per treatment group, and the bacterial isolates used for the study were recovered from natural infections of BNP or MAS in farmed Vietnamese P. hypophthalmus. The results of the experimental infections mimicked the natural disease outbreaks reported from these pathogens in P. hypophthalmus. Furthermore, it was clearly demonstrated that E. ictaluri was only recovered from the fish exposed to the bacterium and not recovered from the animals receiving A. hydrophila.

Survival of eastern oysters Crassostrea virginica from three lines following experimental challenge with bacterial pathogens

Shellfish production is often affected by bacterial pathogens that cause high losses in hatcheries and nurseries. We evaluated the relative survival of larvae and juveniles of 3 Crassostrea virginica oyster lines: (1) GHP, a Rhode Island line; (2) NEHY, a line resistant to dermo and multinucleated sphere X diseases; and (3) FLOWERS, a line resistant to Roseovarius oyster disease, experimental challenge with Vibrio spp. isolates RE22 and RE101, causative agents of bacillary necrosis in Pacific oyster larvae, and the type strain of Roseovarius crassostreae, causative agent of Roseovarius oyster disease. All of the isolates were able to induce significant mortalities in oyster larvae and juveniles. Susceptibility to bacterial challenge in larvae was significantly higher at 25 degrees C than at 20 degrees C. Susceptibility decreased with oyster age; mean survival time ranged from 24 h in oyster larvae to more than 6 wk in juveniles. Significant differences in susceptibility to bacterial challenge were observed between oyster lines; NEHY was the most resistant line overall. Extracellular products (ECPs) from Vibrio sp. RE22 and R. crassostreae, as well as viable bacteria, were toxic to hemocytes from the 3 oyster lines, suggesting that ECPs are involved in pathogenesis and that external and mucosal barriers to infection are major contributors to resistance to bacterial challenge. These protocols will be useful in the elucidation of mechanisms of bacterial pathogenesis and resistance to infection in oysters.

Comparative susceptibility of veliger larvae of four bivalve mollusks to a Vibrio alginolyticus strain.

The susceptibility of 7 d old veliger larvae of the scallops Argopecten ventricosus and Nodipecten subnodosus, the penshell Atrina maura, and the Pacific oyster Crassostrea gigas to a pathogenic strain of Vibrio alginolyticus was investigated by challenging the larvae with different bacterial concentrations in a semi-static assay. The results indicate that the larvae of the 2 scallop species are more susceptible to the V. alginolyticus strain than those of the oyster and the penshell. Signs of the disease were similar to bacillary necrosis described in previous work. Interspecies differences in susceptibility to pathogens are discussed.

Bacillary necrosis in farmed Pangasius hypophthalmus (Sauvage) from the Mekong Delta, Vietnam

This is the first description of an economically important disease of intensively reared Pangasius hypophthalmus from the Mekong Delta. Major presenting signs included an increase in mortality that would not respond to treatment, and at necropsy, the presence of numerous white necrotic and pyogranulomatous foci throughout the viscera, notably in spleen, liver and kidney. Despite the confounding presence in earlier samples of numerous, primarily renal, myxosporean parasites, the common denominator amongst these and samples from eight subsequently sampled sites was the presence of moderate‐to‐large numbers of bacilli in all typical lesions. These bacteria were subsequently isolated and tentatively identified as a Bacillus sp. Pilot studies experimentally reproduced similar lesions approximately 7 days following intraperitoneal injection of the bacteria.

Inhibitory Effects of Ovoglobulins on Bacillary Necrosis in Larvae of the Pacific Oyster, Crassostrea gigas

In order to develop an alternative method to antibiotics for preventing bacillary necrosis in bivalve mollusc larvae, we examined the effects of ovoglobulins (proteins derived from the whites of hens’ eggs) on the survival of larvae of the Pacific oyster Crassostrea gigas. The pathogenic Vibrio tubiashii (ATCC 19106) was used to infect larvae of the Pacific oyster. V. tubiashii showed strong pathogenicity to oyster larvae, causing 100% mortality after experimental exposure for 24 h at a concentration of 105 cfu (colony-forming units)/ml. In contrast, the addition of ovoglobulins at a concentration of 10 μg/ml to larval oysters, challenged with V. tubiashii at 105 cfu/ml, led to a marked increase in larval survival of 96.5% at 24 h after infection. The V. tubiashii culture supernatant was also shown to be pathogenic to larval oysters; however, its pathogenicity was completely inhibited by the addition of 10 μg/ml of ovoglobulins. Larval oysters infected by V. tubiashii showed typical symptoms of bacillary necrosis including anomalous swimming and detachment of cilia and/or vela. In contrast, live larvae were actively motile, and their cilia and vela were not necrotized in the ovoglobulins-added group. The addition of ovoglobulins clearly suppressed the growth of V. tubiashii in gelatin–sea water broth, but the number of viable V. tubiashii 24 h after incubation did not decrease to the initial dose level. Findings obtained in this study indicate that ovoglobulins almost completely protect larval oysters from V. tubiashii infection by nonbactericidally inhibiting the growth of V. tubiashii without affecting survival of the oysters.

Vibrio splendidus biovar II as the causative agent of bacillary necrosis of Japanese oyster Crassostrea gigas larvae.

Recurrent outbreaks of a disease leading to mass mortalities in an oyster (Crassostrea gigas) hatchery located in western Japan were investigated. The disease occurred regularly in 2- to 8-d-old larvae and has been experimentally controlled in the hatchery by treating the larval rearing water with streptomycin, without ascertaining the etiological agent. The signs of the disease and the course of infection resembled bacillary necrosis reported in oysters and other bivalve molluscs in the USA and Europe. Quantitative and qualitative examinations of the bacterial flora of hatchery samples including source water, broodstock, larval feed and larvae revealed a very high total bacterial load and presumptive vibrios in diseased larvae. Further, the bacterial profile revealed that Vibrio spp. constituted approximately 60 to 95% of the bacteria isolated from infected larvae and most isolates were identified as V. splendidus biovar II and V. harveyi, suggesting their possible role in the disease. However, experimental challenges proved the pathogenicity of V. splendidus II. Several isolates of V. splendidus II from infected larvae were highly pathogenic, producing 100% mortality at levels of 10(5) cfu ml-1 in 24 h, while isolates from other sources demonstrated a low degree of virulence. Detection of V. splendidus II from broodstock, especially in the gonad of a few breeders, suggests the probability that broodstock could be the source and route of transmission of this pathogen.

Pathogenicity of Vibrio splendidus biovar II, the causative bacterium of bacillary necrosis of Japanese oyster larvae.

The potential pathogenicity of strains of Vibrio splendidus biovar II, which were isolated from bacillary necrosis of triploid larvae of Pacific oyster Crassostrea gigas in a hatchery in western Japan, was investigated. The course of experimental infection with virulent strains of V. splendidus biovar II was very rapid in 5-day-old veliger larvae with disease signs apparent within 6 to 12 h after exposure of larvae at doses of 104 to 106 CFU/ml, and mortalities up to 100% were recorded in 24 h at 105 and 106 CFU/ml. Although all the tested stages of larvae were experimentally infected by virulent strains of V. splendidus biovar II, a later stage (17 days posthatching) was less susceptible than the earlier stages of development. Diploid and triploid larvae were almost equally susceptible to this pathogen.Extracellular products and intracellular components of the strains were lethal to larvae, but their lethality did not correlate with the virulence of live cultures. These results suggest that the ability to elaborate toxins is not the only virulence factor in the pathogenicity of V. splendidus biovar II. The ability of this pathogen to bring about significant mortalities in oyster larvae at densities of 104 CFU/ml and its long survival in seawater make this pathogen a potential threat to larval oyster productions in hatchery systems.

Role of Low-Molecular-Weight Ciliostatic Toxins in Vibriosis of Bivalve Mollusks

Abstract Toxins play a major role in bacillary necrosis of hatchery-reared bivalve larvae. Two principal types of toxin of Vibrio spp. have been identified: A proteinase (molecular weight = 40,000) that degrades connective tissue and at least one ciliostatic toxin (molecular weight = 500–1,000). The ciliostatic toxin is stable at 100°C for 10 min and probably accounts for the cessation of feeding and swimming in the early stages of bacillary necrosis. Ciliostatic toxins were produced by 17 of 20 Vibrio spp. that are pathogenic to fish or that were isolated from diseased mollusks and included Vibrio tubiashii, V. anguillarum, and V. alginolyticus. Of 53 Vibrio and Aeromonas isolates not associated with bivalve infections, only 15 produced a ciliostatic toxin.

Bacillary necrosis in hatcheries of Ostrea edulis in Spain

Two hatcheries of flat oyster ( Ostrea edulis) located in the North-West of Spain, which were affected by massive larval mortalities during summer months, were monitored in order to look for the causative organism(s) and the source(s) of the pathogens. Qualitative and quantitative analyses of the bacterial flora, conducted in different parts of the hatcheries, revealed that the total bacterial count (TVC) and the presumptive vibrios (PVC) increased during the spring, following the rise of temperature, up to the start of the outbreaks. Very high TVC levels were found in the phytoplanktonic food of broodstock and larvae; these bacterial populations were composed mainly of Flavobacterium and Pseudomonas but no vibrios were detected. Whereas the number of vibrios in the incoming water was effectively reduced by filtration and U.V. light, Vibrio strains were detected as part of the normal flora in the broodstock and larval tanks favoured by the conditions of nutrients and temperature provided in the hatcheries. The fact that vibrios were present in the gonads of hatchery-conditioned broodstock but not in wild oysters seems to indicate that these opportunistic pathogens can invade and proliferate in the oyster tissues, and subsequently infect the larvae. The agents responsible for the vibriosis outbreaks in both hatcheries resembled Vibrio tubiashii which produces lethal exotoxins for larvae and the typical symptoms of bacillary necrosis. Chloramphenicol was the most effective drug for controlling the disease.

Soft-shell clam, Mya arenaria, a convenient laboratory animal for screening pathogens of bivalve mollusks.

Attempts to introduce infectious or foreign material into oysters and other bivalve mollusks usually involve force or trauma because of immediate, prolonged adduction of the tightly closing valves. The soft-shell clam, Mya arenaria, is unable to seal its valves completely and relaxes readily, exposing soft tissue and a large siphon. This species is free from fouling organisms and is readily available at all seasons in the New England and mid-Atlantic areas. Suspensions of five strains of Vibrio sp. that cause bacillary necrosis in larval and juvenile bivalve mollusks were injected into the heart, siphon tissue, and the incurrent and excurrent siphon lumina of soft-shell clams. All vibrio strains caused significant mortality, usually within 2 days. Heaviest losses resulted from heart and excurrent siphon injections. No mortality occurred in control clams injected with seawater, broth, Serratia sp., and Escherichia coli. The soft-shell clam appears to be a useful animal for testing the pathogenicity of marine microorganisms for bivalve mollusks.

Soft-Shell Clam, Mya arenaria, a Convenient Laboratory Animal for Screening Pathogens of Bivalve Mollusks

Attempts to introduce infectious or foreign material into oysters and other bivalve mollusks usually involve force or trauma because of immediate, prolonged adduction of the tightly closing valves. The soft-shell clam, Mya arenaria, is unable to seal its valves completely and relaxes readily, exposing soft tissue and a large siphon. This species is free from fouling organisms and is readily available at all seasons in the New England and mid-Atlantic areas. Suspensions of five strains of Vibrio sp. that cause bacillary necrosis in larval and juvenile bivalve mollusks were injected into the heart, siphon tissue, and the incurrent and excurrent siphon lumina of soft-shell clams. All vibrio strains caused significant mortality, usually within 2 days. Heaviest losses resulted from heart and excurrent siphon injections. No mortality occurred in control clams injected with seawater, broth, Serratia sp., and Escherichia coli. The soft-shell clam appears to be a useful animal for testing the pathogenicity of marine microorganisms for bivalve mollusks.

Marine vibrios associated with bacillary necrosis, a disease of larval and juvenile bivalve mollusks.

The etiological agents of bacillary necrosis, a disease of larval and juvenile bivalve mollusks, are designated members of the genus Vibrio.

Bacillary necrosis, a disease of larval and juvenile bivalve mollusks. I. Etiology and epizootiology.

Tubiash, Haskell S. (U.S. Bureau of Commercial Fisheries, Milford, Conn.), Paul E. Chanley, and Einar Leifson. Bacillary necrosis, a disease of larval and juvenile bivalve mollusks. I. Etiology and epizootiology. J. Bacteriol. 90:1036-1044. 1965.-Lethal bacterial infections of a variety of hatchery-spawned bivalve mollusk larvae and juveniles have been studied. The symptoms of the disease and the course of the infection are described. Four biotypes and five antigenic types of bacteria, pathogenic for the larvae of five species of bivalve mollusks, were isolated and described in some detail. All are gram-negative motile rods. Comparative studies were made of a fairly large number of similar bacteria isolated from presumably normal marine fauna. None of these was pathogenic for the bivalve larvae nor did they have antigens in common with the pathogenic group. The four biotypes had a number of characteristics in common that rarely were present in other cultures from marine fauna. Several antibiotic preparations proved to be of value in the treatment and control of the infection.

Bacillary Necrosis of the Liver: A Disease of the Unborn Lamb

Bacillary Necrosis of the Liver: A Disease of the Unborn Lamb