Antimicrobial activity of partially characterized analytes from Bacillus pumilus(B2)

This study was carried out to characterize the antimicrobial substance produced by the strain of Bacillus pumilus (B2) obtained from Novozymes Biologicals Inc. to compare its antimicrobial activity by agar well diffusion assay and bacteriocin activity assay via critical dilution method against seven different strains of Vibrio spp., specifically V. alginolyticus (A01), V. cholerae (C01), V. fluvialis (F01, F02), V herveyii (H), V. mimicus (M01), V. parahaemolyticus (P01) and V. vulnificus (V01, V02). All Vibrio spp. were isolated from the hemolymph and intestine of the white faeces disease‐infected moribund pacific white‐leg shrimp Litopenaeus vannamei (Boone 1931) and one strain (V. harveyi) from its diseased postlarva. The cell‐free neutralized supernatant (CFNS) of B2 showed moderate thermo‐stability being stable up to 70°C for 60 min with, however, reducing activity above 80°C for 20 min. B2 antimicrobials showed a stable activity within the pH ranging from 6 to 10 at room temperature and at 4°C, while residual antimicrobial activity of crude CFNS showed tolerance to salinity up to 7% of sodium chloride below 4°C. No B2 activity was obtained while exposed to proteolytic enzyme, such as proteinase k and pepsin, while its activity kept stable exposed to lipase. Initial B2 characterization for antimicrobial substance in CFNS revealed proteinaceous in nature owing to activity loss against proteolytic enzymes and no lipid moiety activity against lipase, which could be categorized as bacteriocin‐like inhibitory substance having potential application against several strains of Vibrio spp. in aquaculture.     

Multilocus sequence analysis of Vibrionaceae isolated from farmed amberjack and the development of a multiplex PCR assay for the detection of pathogenic species

Since 2011, high mortality rates and symptoms consistent with vibriosis have been observed in farmed amberjack (Seriola dumerili) in Japan. To identify 41 strains isolated from diseased amberjack, a multilocus sequence analysis using nine concatenated genes (ftsZ, gapA, gyrB, mreB, pyrH, recA, rpoA, topA and 16S rRNA) was conducted. Twenty‐seven strains were identified as Vibrio harveyi, suggesting an epidemic of V. harveyi infection in amberjack farms. Other strains were identified as Vibrio anguillarum, Vibrio owensii and Photobacterium damselae subsp. damselae. To develop an efficient diagnostic method for vibriosis in amberjack, a multiplex PCR system was developed to identify V. anguillarum, V. harveyi and P. damselae subsp. damselae. The method successfully discriminated between these three bacterial species, with amplification products of 350 bp for V. anguillarum, 545 bp for V. harveyi and 887 bp for P. damselae subsp. damselae and can be used for diagnosis in aquaculture farms.     

Prevalence and antimicrobial resistance of vibrios of human health significance in inland saline aquaculture areas

This study was conducted to evaluate the prevalence, potential pathogenicity and antimicrobial resistance of Vibrio isolates from 65 soil/water/fish samples collected from inland saline aquaculture areas. Depending on the sample type, presumptive Vibrio counts ranged from 2.50 to 6.16 log₁₀ CFU/ml (or/g). Among the 119 confirmed Vibrio isolates, Vibrio cholerae was found to most dominant (91.6%) and it was detected in all the samples from inland saline areas. Seven other Vibrio spp. including Vibrio parahaemolyticus and Vibrio vulnificus were also detected. Except one O139 serotype, rest of the V. cholerae isolates were found belonging to non‐O1/non‐O139 serogroups. None of the V. cholerae isolate was found positive for ctx gene. Antimicrobial susceptibility testing against 7 commonly used antibiotics revealed highest resistance (50.4%) against ampicillin. Very high intermediate resistance (87.4%) was also observed against erythromycin. Contrary to previous studies, high susceptibility (>70%) to chloramphenicol, nalidixic acid, tetracycline and trimethoprim was observed in Vibrio isolates obtained in present study. Almost 20% of Vibrio isolates were resistant to two or more antibiotic classes with multiple antibiotic resistance (MAR) index value of ≥0.28. Presence of V. cholerae isolates with very high MAR index value of 0.85 also suggested that these multidrug‐resistant environment isolates could serve as reservoir of antibiotic‐resistant genes in aquatic systems. The presence of multiple drug resistance vibrios in emerging inland saline aquaculture systems emphasizes the need for their routine monitoring for developing the risk assessment and mitigation strategies.     

Exclusion of Vibrio spp. by an antagonistic marine actinomycete Streptomyces rubrolavendulae M56

A marine actinomycete Streptomyces rubrolavendulae M56 isolated from the sediments of Bay of Bengal and displaying biogranulation property was used for the study. The strain showed antagonistic property against vibrios, the opportunistic pathogens in aquaculture. The efficacy of the biogranules of actinomycete M56 in competitive exclusion of Vibrio spp. was tested both in vitro and in vivo. Streptomyces rubrolavendulae M56 biogranules could significantly exclude the pathogenic Vibrio spp. in co‐culture experiments (in vitro). In vivo exclusion of Vibrio spp. in a Penaeus monodon postlarval rearing system was evaluated by treatment of the rearing water with biogranules of S. rubrolavendulae M56. The experiments proved that S. rubrolavendulae M56 biogranules could reduce the pathogenic Vibrio spp., while maintaining total heterotrophic bacterial count. Therefore, the actinomycete biogranules (M56) can be used as a promising alternative to antibiotics in the shrimp larval production system which is often affected by vibriosis.     

Antiparasitic activity of the medicinal plant Dillenia suffruticosa against the marine leech Zeylanicobdella arugamensis (Hirudinea) and its phytochemical composition

Zeylanicobdella arugamensis (Annelida: Hirudinea), a marine parasitic leech, is currently affecting different species of cultured groupers, hybrid groupers, snappers and sea bass in Malaysia. Dillenia suffruticosa (Dilleniaceae), a medicinal plant found in Sabah, has been selected in our experiment to kill the leeches as a natural control method. The leech‐infested hybrid groupers were collected from aquaculture facilities, and the isolated leeches were challenged against methanol extract of D. suffruticosa leaves. The experiment was carried out using various concentrations of the extracts such as 25, 50 and 100 mg/ml. The methanol extract showed significant antiparasitic activity against Z. arugamensis with 100% mortality at a concentration of 100 mg/ml in 14.39 ± 3.75 min., followed by 50 and 25 mg/ml in 32.97 ± 9.29 and 41.77 ± 5.40 min., respectively. The phytochemical composition of the extract was determined using GC‐MS analysis to understand the nature of the principal compounds responsible for its antiparasitic properties. The leaves of D. suffruticosa demonstrated the presence of different bioactive compounds of various natures with varying percentages. Thus, it could be revealed that the methanol extract of D. suffruticosa mainly contains vital phytochemical compounds and showed an effective antiparasitic activity against the harmful leeches of hybrid groupers.     

Comparative assessment of Vibrio virulence in marine fish larvae

Vibrionaceae infections are a major obstacle for marine larviculture; however, little is known about virulence differences of Vibrio strains. The virulence of Vibrio strains, mostly isolated from vibriosis outbreaks in farmed fish, was tested in larval challenge trials with cod (Gadus morhua), turbot (Scophthalmus maximus) and halibut (Hippoglossus hippoglossus) using a multiwell dish assays with single‐egg/larvae cultures. The strains differed significantly in virulence as some caused a high mortality of larva reaching 100% mortality after a few days, while others had no or only marginal effects on survival. Some Vibrio strains were pathogenic in all of the larva species, while some caused disease only in one of the species. Twenty‐nine of the Vibrio anguillarum strains increased the mortality of larvae from at least one fish species; however, pathogenicity of the strains differed markedly. Other Vibrio species had no or less pronounced effects on larval mortalities. Iron uptake has been related to V. anguillarum virulence; however, the presence or absence of the plasmid pJM1 encoding anguibactin did not correlate with virulence. The genomes of V. anguillarum were compared (D. Castillo, P.W. D'Alvise, M. Middelboe & L. Gram, unpublished data) and most of the high‐virulent strains had acquired virulence genes from other pathogenic Vibrio.     

Characterization of two bacteriophages for specific treatment of biofilm formed by a Vibrio sp. isolated from an abalone farm

Vibrio harveyi and related vibrios are potential pathogens causing luminous vibriosis in marine aquaculture systems. In this study, two lytic phages P4A and P4F isolated using Vibrio strains B4A and B4F as indicator bacteria, respectively, were isolated from seawater of an abalone farm. Vibrio strain B4F belongs to the Harveyi clade of the genus Vibrio and was found to cause mortality of abalones in laboratory microcosms. Both phages were able to lyse Vibrio strain B4F. Electron microscopy revealed that phage P4A had an icosahedral head while P4F possessed an elongated hexagonal head. Both phages belong to the family Siphoviridae with long non‐contractile tails. Restriction endonuclease analysis indicated that both phages were double‐stranded DNA viruses and the genome sizes of P4A and P4F were estimated to be about 49 and 44 kb respectively. One‐step growth curves revealed that these two phages exhibited distinct latent periods, exponential periods and burst sizes by infecting the same Vibrio strain B4F. Both phages were able to significantly reduce Vibrio population density in biofilm formed by Vibrio strain B4F on the surface of polyethylene film. It is suggested that these two phages may be promising candidates as biocontrol agents of infections caused by Vibrio strains belonging to the Haveyi clade in marine aquaculture systems.     

Effect of probiotics on immune competence of giant freshwater prawn Macrobrachium rosenbergii

This study evaluates the effect of probiotics, Zymetin, on the immune efficiency of Macrobrachium rosenbergii juvenile against pathogenic Vibriospp. and Aeromonasspp. The experiment was conducted in glass aquaria with same level of feeding under different treatments, that is, negative control (Cn), positive control with Vibrio spp. (Cv) and Aeromonasspp. (Ca), prawn juveniles fed with probiotics (Zymetin) at 5 g/kg of feed but without pathogen (T1), probiotic fed prawn challenged with Vibrio spp. (T2) and Aeromonasspp. (T3). The results demonstrated that T1 showed higher survival rate, total haemocyte count, non‐granular haemocyte count, phagocytic activity, and clearance efficiency compared to other treatment groups. In contrast, decreased number of small and large granular haemocyte was observed in T1. Despite that, THC was found to be significantly different (p < 0.05) among all the controls and lowest was in Cv, followed by Ca. Besides, phagocytic activity and clearance efficiency against Vibrio spp. and Aeromonasspp. decreased significantly in Cn, Ca, and Cv fed M. rosenbergiijuvenile, while the values were found to be higher in T1 followed by T2 and T3, in that order. In addition, the challenge test showed an increasing trend of total and beneficial bacterial density as well as declining trend of some harmful bacteria in the water and gut of prawn in all the groups tested (p < 0.05).     

Impact of a reduced water salinity on the composition of Vibrio spp. in recirculating aquaculture systems for Pacific white shrimp (Litopenaeus vannamei) and its possible risks for shrimp health and food safety

Tropical shrimp, like Litopenaeus vannamei, in land‐based recirculating aquaculture systems (RAS) are often kept at low water salinities to reduce costs for artificial sea salt and the amount of salty wastewater. Although these shrimp are tolerant against low salinities, innate immunity suppression and changes in the microbial composition in the water can occur. As especially Vibrio spp. are relevant for shrimp health, alterations in the species composition of the Vibrio community were analysed in water from six RAS, run at 15‰ or 30‰. Additionally, pathogenicity factors including pirA/B, VPI, toxR, toxS, vhh, vfh, tdh, trh, flagellin genes and T6SS1/2 of V. parahaemolyticus were analysed. The Vibrio composition differed significantly depending on water salinity. In RAS at 15‰, higher numbers of the potentially pathogenic species V. parahaemolyticus, V. owensii and V. campbellii were detected, and especially in V. parahaemolyticus, various pathogenicity factors were present. A reduced salinity may therefore pose a higher risk of disease outbreaks in shrimp RAS. Because some of the detected pathogenicity factors are relevant for human health, this might also affect food safety. In order to produce healthy shrimp as a safe food for human consumption, maintaining high water salinities seems to be recommendable.     

Isolation,molecular characterization and antimicrobial susceptibility of Aeromonas spp. obtained from Tiger Grouper (Epinephelus fuscoguttatus) and Marble Goby (Oxyeleotris marmoratus) fish in Sabah,Malaysia

Aeromonads are ubiquitous in aquatic environments and have been implicated in fish and human infections. In this study, we isolated, studied antimicrobial susceptibility patterns and screened the existence of 15 virulence genes in aeromonads from two famously consumed fish species—seven marine Tiger Grouper (Epinephelus fuscoguttatus) and eight freshwater Marble Goby (Oxyeleotris marmoratus) from the aquaculture hatchery in Sabah, Malaysia. A total of 30 aeromonads (17 A. caviae, 9 A. rivuli, 4 A. dhakensis) were identified using PCR targeting GCAT gene, rpoD‐restriction fragment length polymorphism and multi‐locus phylogenetic analysis. All 30 strains were resistant to amoxicillin and cephalothin and five strains were multidrug‐resistant. Nine virulence genes (lip, ela, eno, fla, aerA, hylA, dam, alt and ser) present in A. dhakensis, suggesting the virulence potential of this species as a fish pathogen. This study offers as a baseline for future studies in monitoring and managing these two fish in aquaculture industry.     

Use of molasses as a carbon source during the nursery rearing of Farfantepenaeus brasiliensis (Latreille, 1817) in a Biofloc technology system

The present work evaluated the use of molasses as a carbon source during the nursery rearing of Farfantepenaeus brasiliensis in a Biofloc technology system. During a 30 days trial, three replicate tanks were randomly assigned to the following treatments: 1.) molasses (with molasses addition) and 2.) control (without molasses addition). Bacteriological analysis was used to quantify the abundance of presumptive Vibrio spp. between control and molasses treatment. The concentration of this microorganism was lower in molasses compared with control. For the immunological analysis, shrimp haemolymph was collected to determine the total haemocyte count and the total protein concentration. The immunological results were not different between treatments. The performance results of shrimp reared with molasses addition showed that the survival rate (88.87 ± 6.36), the mean final weight (1.22 ± 0.38) and the specific growth rate (0.0309 ± 1.06) were significantly higher compared with control (80.5 ± 2.42; 1.03 ± 0.13; 0.0256 ± 0.97 respectively). Moreover, the addition of molasses contributed to the maintenance of water quality and lower concentration of presumptive Vibrio spp. The control presented an unstable variation of Vibrio spp. reaching values of 80 × 10² CFU/ml, while the highest result of molasses was 20 × 10² CFU/ml, confirming the beneficial effects of molasses addition.     

Identification and pathogenicity study of emerging fish pathogens Acinetobacter junii and Acinetobacter pittii recovered from a disease outbreak in Labeo catla (Hamilton, 1822) and Hypophthalmichthys molitrix (Valenciennes, 1844) of freshwater wetland in West Bengal,India

The disease outbreaks in aquaculture system of wetlands are the major cause of fish mortality. Among various bacterial septicaemic diseases, fish mortality caused by Acinetobacter spp. is recently reported in different fish species. Fish disease outbreak was investigated in a wetland of West Bengal, India to identify the aetiological factors involved. The moribund fish were examined and subjected to bacterial isolation. Two bacterial causative agents were identified as Acinetobacter junii and Acinetobacter pittii by biochemical characterization and 16S rRNA gene amplification. Both the isolates were oxidase‐negative, nitrate‐negative, catalase‐positive and indole‐negative. The molecular identification using 16S rRNA gene sequencing and phylogenetic tree analysis further confirmed the two Acinetobacter spp. with 97%–99% similarity. The antibiotic resistance patterns of these two bacteria revealed that both of them were resistant to β‐lactam, cefalexin, cephalothin, amoxyclav, cefuroxime, cefadroxil, clindamycin, vancomycin and penicillin. In addition, A. pittii was also resistant to other antibiotics of cephams group such as ceftazidime and cefotaxime. In the challenge experiment, both A. junii and A. pittii were found to be pathogenic with LD₅₀ of 1.24 × 10⁵ and 1.88 × 10⁷ cfu/fish respectively. Histopathological examination of gill, liver and kidney revealed prominent changes supporting bacterial septicaemia. The investigation reports for the first time on concurrent infection by A. junii and multidrug‐resistant (MDR)‐A. pittii as emerging fish pathogens to cause severe mortality in Labeo catla and Hypophthalmichthys molitrix in a freshwater wetland.     

Strain and dose infectivity of Vibrio parahaemolyticus: the causative agent of early mortality syndrome in shrimp

Diseases of shrimp have contributed to billions of dollars of economic loss in the aquaculture industry. Newly emerging strains of the bacterium Vibrio parahaemolyticus produce a condition in shrimp called early mortality syndrome or acute hepatopancreatic necrosis disease. Three different V. parahaemolyticus strains were evaluated for their respective pathogenicity on shrimp, Litopenaeus vannamei, when the bacterial strains were grown under various laboratory conditions prior to inoculating shrimp. For each trial, feed was inoculated with a known concentration of bacteria and then fed to the shrimp. The early mortality syndrome strain of V. parahaemolyticus was the most lethal resulting up to 100% mortality within 24 h after being introduced to shrimp via a single feeding. The other two strains of Vibrio, one isolated from the environment and the other from a human clinical case, resulted in 0% and 30% mortality within 96 h respectively. The concentration of the early mortality syndrome strain of V. parahaemolyticus that the shrimp were exposed to directly correlated with mortality rate, which allowed for lethal or sublethal short‐term disease challenge assays to be established. Infiltration of haemocytes was also evident in the midgut caeca of shrimp infected with the early mortality syndrome strain of V. parahaemolyticus, which has not been previously reported.     

Vibrio harveyi (VirB11) recombinant vaccine development against vibriosis in orange‐spotted grouper (Epinephelus coioides)

Despite significant improvements in aquaculture to compensate wild catch, disease organisms have thrived in limiting its national and global potential. Using antibiotics, in a bid to remedy the havoc, has given rise to complications, attracting attention to disease prevention by immune enhancement against diseases. Grouper production has been inhibited for the threats of bacterial infection, particularly of Vibrio origin. Considering the rise in vibriosis cases, improved vaccines are necessary; moreover, recombinant vaccines, the choice for trial in the present experiment have been effective and more specific in improving immunity. The current work deals with grouper immune system enhancement with a recombinant vaccine developed from VirB11 gene in Vibrio harveyi. VirB11 was cloned in V. harveyi for recombinant vaccine development against vibriosis in orange‐spotted grouper (Epinephelus coioides). As indicated by the results, recombinant VirB11 protein showed effectiveness in conferring protection against vibriosis with observable specific antibody response in enzyme‐linked immunosorbent assay (ELISA) analysis; a significant increase (p < 0.05) in antibody levels was observed after a week and after 8 weeks post‐vaccination. From the weeks post‐vaccination, log2 (antibody titres) in the sera of vaccinated groups reached a peak of 14.2 at week 5 in the vaccinated group in comparison with a peak of approximately 5 and 2 in adjuvant and PBS controls. As indicated by the challenge results, 90% relative survival was observed in vaccinated group and 13% relative survival in control group I (adjuvant control). The cumulative performance of protein concludes VirB11 commendable for recombinant vaccine development.     

Experimental evaluation of the culture performance of three exotic and a ‘local’ Giant Freshwater Prawn (Macrobrachium rosenbergii,De Man 1879) culture strain in Fiji

Giant freshwater prawn (GFP; Macrobrachium rosenbergii) aquaculture has expanded rapidly since 1990. Most local culture industries, however, have developed in an unsystematic way. Fiji has a small culture industry producing the ‘Anuenue’ strain; however, performance of this strain has never been systematically evaluated. Recently, some Fijian farmers have reported declines in stock productivity. The current project evaluated the relative performance of three exotic strains with different genetic backgrounds from Malaysia, Indonesia and Vietnam, against the ‘local’ strain in Fiji in a 4 × 3 replicated pond trial experiment. A total of 5827 prawns were harvested after 143 days growout. Individual growth rate and relative survival of the Fiji strain were not statistically different from any of the introduced strains, but Vietnam strain was superior to that of the Malaysia strain. Genetic diversity showed significant differences in variability among strains, with the Malaysian strain displaying the lowest genetic diversity. Indonesia strain showed that females were reaching maturation earlier than other strains and were smaller in size. This study suggests that Malaysian and Indonesian strains would constitute a poor choice for Fiji, whereas the Vietnam strain consistently performed well on all criteria measured. High variation among replicate ponds within strains unfortunately confounded among‐strain variation.     

Formulated diet alters gut microbiota compositions in marine fish Nibea coibor and Nibea diacanthus

Nibea, an economical marine fish, is generally fed on trash fish (the low‐value fish), which can cause high feed costs and waste pollutions in high‐density aquaculture. To assess the effect of formulated diet on the gut microbiota in Nibea coibor and Nibea diacanthus, denaturing gradient gel electrophoresis, clone libraries analysis and Illumina sequencing of the 16S ribosomal RNA gene were used in this study. Two Nibea fishes were both dominated by Gammaproteobacteria (especially Photobacterium) and shared a set of gut microbiota, including Bacilli, Mollicutes, Alphaproteobacteria and Fusobacteriia. Statistical analyses revealed that formulated diet led to lower feed conversion ratio (p < 0.001), lower abundance of Vibrio (p = 0.040) and infectious diseases pathways (p = 0.001), higher abundance of polysaccharide‐degrading bacterium Cellvibrio (p = 0.006) in two Nibea species, with higher weight gain rate (p = 0.023) and microbial diversity (Shannon, p = 0.049 and Simpson, p = 0.044) and more carbohydrate metabolism (p = 0.020) observed in N. coibor. The distribution and correlation network of 17 potential short‐chain fatty acid producing bacteria were obtained and visualized in all treatment groups. The results reveal that formulated diet has beneficial effects on the gut microbial ecology in two Nibea fishes, which suggests the possibility of replacing trash fish diet with formulated diet in Nibea aquaculture.     

Effects of Pediococcus pentosaceus supplementation on growth performance,intestinal microflora and disease resistance of white shrimp,Litopenaeus vannamei

Litopenaeus vannamei is economically important shrimp species in worldwide aquaculture. This study was conducted to assess the effect of different levels of probiotic Pediococcus pentosaceus (PP) on growth performance, feed utilization, digestive enzyme activity, intestinal microflora count and body composition of L. vannamei. Four diets containing different concentrations [0 (PP0), 10⁶ (PPI), 10⁷ (PPII) and 10⁸ (PPIII) CFU/g] of PP were formulated. After 8 weeks feeding trial, the experimental shrimps were challenged with Vibrio anguillarum and noted the surveillance. At the end of the feeding trial, the obtained results revealed a significant increase (p < .05) in final body weight, final length, weight gain (WG), survival rate, protease and amylase activities, lactobacillus sp. and Bacillus sp. intestinal count, total haemocyte counts (THC) and lysozyme activity in PPIII group when compared with the other groups. Similarly, WG, amylase activity, Bacillus sp. count, THC and lysozyme activity were significantly enhanced in case of PPII compared to the control group (p < .05). Interestingly, FCR and Vibrio sp. counts were significantly decreased in case of PPIII group when compared to the other groups (p < .05). Also, significant level of surveillance was noted in the challenging test with V. anguillarum. These results suggested that the P. pentosaceus improved the growth performance, digestive enzyme activity, immunity and tolerance against V. anguillarum of L. vannamei.     

Comparison of Edwardsiella ictaluri isolates from different hosts and geographic origins

The intraspecific variability of E. ictaluri isolates from different origins was investigated. Isolates were recovered from farm‐raised catfish (Ictalurus punctatus) in Mississippi, USA, tilapia (Oreochromis niloticus) cultured in the Western Hemisphere and zebrafish (Danio rerio) propagated in Florida, USA. These isolates were phenotypically homologous and antimicrobial profiles were largely similar. Genetically, isolates possessed differences that could be exploited by repetitive‐sequence‐mediated PCR and gyrB sequence, which identified three distinct E. ictaluri genotypes: one associated with catfish, one from tilapia and a third from zebrafish. Plasmid profiles were also group specific and correlated with rep‐PCR and gyrB sequences. The catfish isolates possessed profiles typical of those described for E. ictaluri isolates; however, plasmids from the zebrafish and tilapia isolates differed in both composition and arrangement. Furthermore, some zebrafish and tilapia isolates were PCR negative for several E. ictaluri virulence factors. Isolates were serologically heterogenous, as serum from a channel catfish exposed to a catfish isolate had reduced antibody activity to tilapia and zebrafish isolates. This work identifies three genetically distinct strains of E. ictaluri from different origins using rep‐PCR, 16S, gyrB and plasmid sequencing, in addition to antimicrobial and serological profiling.     

Outbreak of mass mortality of yearling groupers of Epinephelus (Perciformes,Serranidae) associated with the infection of a suspected new enteric Sphaerospora (Myxozoa: Myxosporea) species in South China Sea

A suspected new enteric Sphaerospora species was believed to be directly associated with the mass mortality of yearling groupers of Epinephelus spp. in South China. The epizootic generally emerged from late September to late April of the following year. The infection prevalence and mortality rate were significantly negatively correlated with fish size. Clinical signs included anorexia, cachexia and extrusion of white pulp‐like substance from anus after gentle pressure on the abdomen. Upon necropsy, severe intestinal oedema, thin and transparent intestinal wall, swollen spleen, kidney and gall bladder could be observed. Wet preparation of the infected samples showed large amount of typical disporous plasmodia of the genus Sphaerospora, but no mature spores were observed. Epidemiological investigation showed that this parasite exclusively infected Epinephelus groupers. Histopathologically, this species mainly infected the epithelium of intestine and kidney tubules and caused severe epithelia sloughing and the collapse of intestinal villus. Interestingly, this enteric myxosporidiosis did not cause severe emaciation of infected fish for mass mortality usually emerged within 2–3 days after appearance of clinical signs. The species was most genetically related to Sphaerospora fugu (89% sequence identity) and phylogenetically positioned within marine Sphaerospora lineage. This is the first report of enteric sphaerosporosis of groupers.     

Aeromonas hydrophila and Aeromonas veronii cause motile Aeromonas septicaemia in the cultured Chinese sucker,Myxocyprinus asiaticus

In August 2017, a serious disease causing high mortality occurred in a Myxocyprinus asiaticus aquaculture farm. According to necropsy findings, bacteriology experiments and phylogenetic analysis based on 16S rRNA, cpn60, gyrB and rpoB genes and concatenated alignment sequences (cpn60, gyrB and rpoB genes), two isolates, that is, BBAh1 and BBAv1, were identified as Aeromonas hydrophila and Aeromonas veronii respectively. Artificial infection experiments were carried out, showing that the BBAh1 and BBAv1 strains can cause similar symptoms and have LD50 values of 1.93×10⁵ and 8.77×10⁵ cfu/g respectively. In addition, some virulent genes coding for AerA, Alt, Ast, AscV, AexT, AspA, HlyA, OmpA, Lip and FlaA were detected in the two strains. Furthermore, BBAh1 and BBAv1 showed the same sensitivities to 28 antibiotics, some of which, such as cefotaxime, aztreonam, ceftriaxone and tetracycline, may be used to control the disease. However, the strains were also resistant to many antibiotics. These results provide a scientific reference for the diagnosis, prevention and treatment of motile Aeromonas septicaemia caused by A. hydrophila or A. veronii in cultured Chinese sucker.