鳢流行性溃疡综合征病原分离鉴定与病理形态学观察

为寻找冬春季鳢(Channa maculata)烂身病的病原,应用组织压片、组织切片、扫描电镜技术、HE 常规染色法和 Grocott 六胺银染色法、霉菌分离纯化及 ITS 的序列分析等对冬春季患疑似溃疡综合征的烂身病鳢进行了病原学与病理学研究。病变组织压片观察到大量直径 10~20 μm,分枝较少、纤细菌丝。扫描电镜观察到肌肉组织中延伸出大量的纤细的丝状真菌。患病鳢皮肤和肌肉表现为变性、坏死与炎性细胞浸润,溃疡灶肌肉内可见大量的慢性肉芽肿结节和炎性细胞浸润。结节基本结构由类上皮细胞和多核巨细胞、炎性细胞、霉菌菌丝(横断面呈圆形,斜断或纵断呈丝状)组成,霉菌位于结节中央。Grocott 六胺银染色观察到结节中有大量棕色菌体。无菌分离培养可观察到典型丝状霉菌;霉菌在灭菌池塘水 20℃过夜, 12 h 后可观察原代孢子群形成,真菌 ITS 序列分析表明其与侵袭丝囊菌(Aphanomyces invadans)同源性为 100%。侵袭丝囊霉菌(A. invadans)是杂交鳢溃疡综合症病的主要病原,为鱼类流行病学调查和疾病防治的深入研究奠定基础。     

First identification and characterization of Streptococcus iniae obtained from tilapia (Oreochromis aureus) farmed in Mexico

This is the first study to isolate, identify and characterize Streptococcus iniae as the causative disease agent in two tilapia (Oreochromis aureus) populations. The populations were geographically isolated, of distinct origins, and did not share water sources. Affected fish showed various external (e.g., exophthalmia and cachexia, among others) and internal (e.g., granulomatous septicaemia and interstitial nephritis, among others) signs. All internal organ samples produced pure cultures, two of which (one from each farm, termed S‐1 and S‐2) were subjected to biochemical, PCR and 16S rRNA sequencing (99.5% similarity) analyses, confirming S. iniae identification. The two isolates presented genetic homogeneity regardless of technique (i.e., RAPD, REP‐PCR and ERIC‐PCR analyses). Pathogenic potentials were assessed through intraperitoneal injection challenges in rainbow trout (Oncorhynchus mykiss) and zebrafish (Danio rerio). Rainbow trout mortalities were respectively 40% and 70% at 10⁴ and 10⁶ CFU per fish with the S‐1 isolate, while 100% mortality rates were recorded in zebrafish at 10² and 10⁴ CFU per fish with the S‐2 isolate. The obtained data clearly indicate a relationship between intensified aquaculture activities in Mexico and new disease appearances. Future studies should establish clinical significances for the tilapia industry.     

First record of Tetramicra brevifilum in lumpfish (Cyclopterus lumpus,L.)

A microsporidian species with 98.3–98.4% nucleotide identity to Tetramicra brevifilum (Journal of Fish Diseases, 3 , 1980, 495) was diagnosed in lumpfish (Cyclopterus lumpus, L.) broodstock held at a breeding and rearing facility in western Ireland. The fish were wild‐caught from the west coast of Ireland, and the first case was diagnosed one year after capture. Clinical signs included severe bloating, lethargy, exophthalmos, anorexia, white patches on the cornea and externally visible parasitic cysts on skin and fins. Necropsy revealed severe ascites, white nodules and vacuoles in all the internal organs and partial liquefaction of the skeletal muscle. On histological examination, microsporidian xenomas were observed in all internal organs, the skin, skeletal muscle, gills and the eyes. The microsporidian species was identified by molecular analysis and transmission electron microscopy. This is the first record of T. brevifilum infecting lumpfish, and the disease is considered to be of potential significance to the rising aquaculture industry of this species.     

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.     

High efficacy of white spot syndrome virus replication in tissues of freshwater rice‐field crab,Paratelphusa hydrodomous (Herbst)

An attempt was made to determine the replication efficiency of white spot syndrome virus (WSSV) of shrimp in different organs of freshwater rice‐field crab, Paratelphusa hydrodomous (Herbst), using bioassay, PCR, RT‐PCR, ELISA, Western blot and real‐time PCR analyses, and also to use this crab instead of penaeid shrimp for the large‐scale production of WSSV. This crab was found to be highly susceptible to WSSV by intramuscular injection. PCR and Western blot analyses confirmed the systemic WSSV infection in freshwater crab. The RT‐PCR analysis revealed the expression of VP28 gene in different organs of infected crab. The indirect ELISA was used to quantify the VP28 protein in different organs of crab. It was found that there was a high concentration of VP28 protein in gill tissue, muscle, haemolymph and heart tissue. The copy number of WSSV in different organs of infected crab was quantified by real‐time PCR, and the results revealed a steady increase in copy number in different organs of infected crab during the course of infection. The viral inoculum prepared from different organs of infected crab caused significant mortality in tiger prawn, Penaeus monodon (Fabricius). The results revealed that this crab can be used as an alternate host for WSSV replication and production.     

Temporal and spatial dynamics of white spot syndrome virus in the Chinese mitten crab,Eriocheir sinensis

White spot syndrome virus (WSSV) is the most severe viral pathogen to the crustacean aquaculture industry worldwide. Recently, serious WSSV outbreaks caused catastrophic losses in the Chinese mitten crab, Eriocheir sinensis in Jiangsu Province, Eastern China. However, to date, little is known about its infection mechanism in the new natural host. This study aimed to reveal the temporal and spatial dynamics of WSSV in E. sinensis. The slow viral growth in the early stage of infection was the light infection stage (from 0 to 24 hpi), and the exponential growth stage that followed was the logarithmic phase (from 24 to 72 hpi). The viral growth curve ended with the plateau phase (from 72 to 144 hpi) which demonstrated a consistent high level of viral load and accompanied heavy crab mortality. The viral load increased as time progressed with similar growth curves, however, at different degrees. The viral copy numbers of tissues at different time intervals, analysed using one‐way analysis of variance (anova ), showed significant differences between tissues at all time points (P < 0.05). Infection was detectable as early as 6 hpi in all the tissues screened. The severity of infection was found to be maximum in gill and pleopods, which could be recommended for diagnostic testing. This study might provide important data to analyse theoretically the interaction between WSSV and the host.     

Atlantic salmon smolts in the Irish Sea: First evidence of a northerly migration trajectory

Results from an acoustic telemetry study revealed for the first time a northerly migration route for Atlantic salmon (Salmo salar L.) smolts leaving the east coast of Ireland. Atlantic salmon smolts were tagged in spring 2019 in the Castletown and Boyne rivers. Three tagged smolts registered on disparate deep‐water offshore marine receivers as they travelled northwards out of the Irish Sea through the North Channel. One fish had migrated an estimated 250 km in a period of 32 days. The remaining two individuals were detected on receivers located off the Northern Ireland coast, further corroborating the northward migration of salmon smolts through the Irish Sea.     

Co‐Infection with Pseudomonas anguilliseptica and Delftia acidovorans in the European eel,Anguilla anguilla (L.): a case history of an illegally trafficked protected species

Inspections by customs agents at Barcelona airport discovered 420 kg of contraband glass eels prepared for shipment to Hong Kong. After confiscation of these animals by police, they were transported to holding facilities to be maintained until after a judicial hearing. Upon arrival, they were separated into two groups and held under ambient flow‐through conditions in fresh water. During their captivity period, several peaks in mortality occurred and multiple bacterial strains were isolated from moribund animals. Sequencing of 16S rDNA was used to determine specific identity of the isolates. An initial isolation of Pseudomonas anguilliseptica was treated with oxytetracycline. A subsequent isolation of Delftia acidovorans proved resistant to oxytetracycline and was treated with gentamicin in combination with sulphadiazine–trimethoprim. Once the health condition of the animals was stabilized, they were partitioned into groups and subsequently released as part of a restocking effort for the species following the guidelines of Regulation (EC) 1100/2007 (Anon 2007). This represents the first record for both bacterial species in the host Anguilla anguilla in the Spanish Mediterranean.     

Antagonism against fish pathogens by cellular components and verification of probiotic properties in autochthonous bacteria isolated from the gut of an Indian major carp,Catla catla (Hamilton)

Probiotic potential of the autochthonous bacteria in catla, Catla catla has been evaluated through determination of antagonistic activity (in vitro) of the cellular components of gut bacteria against seven fish pathogens. Altogether 208 strains were isolated, inhibitory activity of the isolates was evaluated through cross‐streaking and 16 primarily selected antagonistic strains were confirmed using the double‐layer method. Four bacteria that showed antagonism against ≥4 pathogens were selected as putative probiotics. The intracellular, extracellular, whole‐cell and heat‐killed cell components exhibited bactericidal activity against the pathogens. In addition, the selected strains were capable of producing different extracellular enzymes, competent to grow in intestinal mucus and could tolerate diluted bile juice. Analysis of 16SrRNA partial gene sequence revealed that both the strains CC1FG2 and CC1FG4 were Bacillus methylotrophicus (KF559344 and KF559345), while the isolates CC1HG5 and CC2HG7 were Bacillus subtilis subsp. spizizenii (KF559346) and Enterobacter hormaechei (KF559347) respectively. Bio‐safety evaluation through intra‐peritoneal injection of the isolates did not induce any pathological signs or mortalities in C. catla. The study confirmed probiotic properties of autochthonous gut bacteria in C. catla and demonstrated potential for using them as bio‐control agents. However, in vivo studies are essential to explore their efficacy in the commercial aquaculture.     

Identification and characterization of the pathogen associated with skin ulcer syndrome in lined seahorse,Hippocampus erectus

The lined seahorse, Hippocampus erectus, is an Atlantic species, which mainly inhabits shallow sea beds or coral reefs. Seahorse has become very popular in China due to its wide use in traditional Chinese medicine (TCM). However, the aquaculture of the lined seahorse has been threatened by a variety of diseases, especially bacterial infections. Skin ulcer syndrome becomes more and more common in the culture of seahorses, leading to skin ulcer, liver erosion and minority tail rotting. These diseased seahorses die in only 72 hr, causing huge economic losses in aquaculture. Pathogen HMT‐1 was isolated from the lined seahorses with skin ulcer syndrome, which was identified as Vibrio harveyi. The median lethal dose (LD₅₀) of HMT‐1 was 2.88 × 10⁸ cfu/ml. The pathogenic mechanism of HMT‐1 was preliminarily studied, and the pathological changes of the diseased lined seahorses were also investigated. Moreover, the activities of ACP, complement 3, SOD and LZM at different time post injection were measured by commercial kits. Our findings can provide reliable reference for diagnosis and treatment in the future.     

Changes in the gut microbiome of the Chinese mitten crab (Eriocheir sinensis) in response to White spot syndrome virus (WSSV) infection

Intestinal microorganisms play important roles in maintaining host health, but their functions in aquatic animal hosts have yet to be fully elucidated. The Chinese mitten crab, Eriocheir sinensis, is one such example. We attempted to identify the shift of gut microbiota that occurred in response to infection of white spot syndrome virus (WSSV), an emerging viral pathogen in the crab aquaculture industry. The microbiota may exert some control over aspects of the viral pathogenesis. We investigated the changes in composition and structure of the crab gut microbiome during various WSSV infection stages of 6 h post‐infection (hpi) and 48 hpi, using a 16S rRNA approach on the MiSeq Illumina sequencing platform. Four phyla (Firmicutes, Proteobacteria, Tenericutes and Bacteroidetes) were most dominant in the gut of E. sinensis regardless of the WSSV infection stages. However, further analysis revealed that over 12 bacterial phyla, 44 orders and 68 families were significantly different in abundance at various states of WSSV infection. Several intriguing aspects of E. sinensis gut bacteria that had not been previously reported were also uncovered, such as class Mollicutes was dominant here, but absent in crabs from Yangtze River estuary and Chongming Islands. Overall, this study provided the first evidence that changes in gut microbiome were closely associated with the severity of WSSV infection and that indicator taxa could be used to evaluate the crab health status.     

Evidence for the presence of white spot syndrome virus (WSSV) and monodon baculovirus (MBV) in wild Penaeus monodon (Fabricius) broodstock,in the southeast coast of India

A survey on the presence of the viruses of two economically significant diseases, white spot syndrome virus (WSSV) and monodon baculovirus (MBV) in wild‐collected Penaeus monodon broodstock, was conducted during different seasons of the year in two major coastal areas of southeast India. The broodstock were collected along the coast of Tamil Nadu and Andhra Pradesh during summer, premonsoon, monsoon and post‐monsoon seasons for three consecutive years. A total of 7905 samples were collected and subjected to MBV screening, and 6709 samples that were screened as MBV negative were diagnosed for WSSV. MBV was detected using rapid malachite green staining and WSSV by nested polymerase chain reaction. Prevalence data of the viruses were analysed using the EpiCalc 2000 program at 95% confidence interval. Samples collected from the Andhra Pradesh coast displayed a slightly higher prevalence of WSSV and MBV infection than those collected from Tamil Nadu, although this difference was not statistically significant (P > 005). In addition, it was found that the prevalence of both WSSV and MBV infections fluctuated according to season. Data on prevalence of these viruses in broodstock would be useful to develop strategies for shrimp health management along the southeast coast of India.     

Changing salmon: An analysis of body mass,abundance, survival,and productivity trends across 45 years in Puget Sound

Pacific salmon and trout (Oncorhynchus spp., Salmonidae) of the Puget Sound region of Washington State, USA, have experienced recent and longer‐term (multidecadal) variability in abundance while supporting robust fisheries. As part of the post‐season salmon management process, population‐specific estimates of total adult abundance to Puget Sound (Strait of Juan de Fuca) for pink (O. gorbuscha), chum (O. keta), coho (O. kisutch), sockeye (O. nerka), and Chinook (O. tshawytscha) salmon and steelhead trout (O. mykiss) are calculated annually. We compiled annual estimates of body mass, abundance and survival of hatchery‐ and naturally produced salmon from 1970 to 2015 to compare spatial and temporal patterns across species. Average weights of adult salmon and steelhead returning to Puget Sound, with the exception of coho salmon, have decreased since the 1970s. Temporal trends in abundance, survival and productivity varied by species and origin (hatchery vs. naturally produced). Generally, abundance and survival rates of natural‐origin species decreased whereas those of hatchery‐produced species did not, which is in contrast with other studies' general conclusions of decreasing survival among Puget Sound salmonids. Species diversity has decreased in recent years, with salmonids that rely on a short freshwater rearing phase in the natural environment (hatchery‐produced fish and naturally produced pink and chum) representing >90% of total returns in most years. This new information reveals patterns of body size, abundance, survival and productivity across species, life history and rearing type over the past 45 years and, in doing so, demonstrates the strength in multidecadal, multifactor time series to critically evaluate salmonid species.