Spatiotemporal distribution of Flavobacterium psychrophilum and ayu Plecoglossus altivelis in rivers revealed by environmental DNA analysis

Outbreaks of bacterial cold-water disease (BCWD), caused by Flavobacterium psychrophilum, are widespread in Japan, especially among ayu Plecoglossus altivelis. There are few investigations of F. psychrophilum in river water, and its seasonal distribution has not been clarified. We aimed to identify the spatiotemporal dynamics of F. psychrophilum and ayu to provide information that is useful for establishing a countermeasure for BCWD. Quantitative analysis of environmental DNA (eDNA) was used to clarify the year-round dynamics of ayu and F. psychrophilum. We sampled river water from the Nagara and Ibi rivers in Japan, and conducted monthly water sampling and eDNA quantification. Changes in the eDNA concentration of ayu were consistent with the known life histories of the fish. There was a strong negative correlation between the eDNA concentration of F. psychrophilum and water temperature, suggesting a strong dependence of F. psychrophilum dynamics in the river on water temperature. Furthermore, relatively high eDNA concentrations were recorded for both organisms in early summer and fall, suggesting that ayu is infected with F. psychrophilum during these seasons when experiencing up- and downmigration, respectively.

     

Cross-protection of a live-attenuated Flavobacterium psychrophilum immersion vaccine against novel Flavobacterium spp. and Chryseobacterium spp. strains

For salmonid producers, a common threat is Flavobacterium psychrophilum. Recent advancements in bacterial coldwater disease (BCWD) management include the development of a live-attenuated immersion vaccine that cross-protects against an array of F. psychrophilum strains. Emerging family Flavobacteriaceae cases associated with clinical disease have been increasing, including pathogenic isolates of Flavobacterium spp. and Chryseobacterium spp. The cross-protective ability of a live-attenuated F. psychrophilum vaccine was determined against three virulent Flavobacteriaceae isolates. Juvenile rainbow trout were vaccinated, developed high F. psychrophilum-specific antibody titres and were challenged with Chryseobacterium spp. isolates (S25 and T28), a Flavobacterium sp. (S21) isolate, a mixed combination of S21:S25:T28, and a standard virulent F. psychrophilum CSF259-93 strain. Results demonstrated strong protection in the CSF259-93 vaccinated group (relative per cent survival (RPS)=94.44%) when compared to the relevant CSF259-93 controls (p < .001). Protection was also observed for vaccinated fish challenged with the S21:S25:T28 mix (RPS = 85.18%; p < .001). However, protection was not observed with the S21, S25 or T28 isolates alone. Analysis of whole-cell lysates revealed differences in protein banding by SDS-PAGE, but conserved antigenic regions by Western blot in S25 and T28. Results demonstrate that this live-attenuated vaccine provided protection against mixed flavobacterial infection and suggest further benefits against flavobacteriosis.     

Antimicrobial susceptibility of Flavobacterium psychrophilum isolates from the United Kingdom

Routine application of antimicrobials is the current treatment of choice for rainbow trout fry syndrome (RTFS) or bacterial coldwater disease (BCWD) caused by Flavobacterium psychrophilum. In this study, the antimicrobial susceptibilities of 133 F. psychrophilum isolates, 118 of which were from the UK, were evaluated by broth microdilution and disc diffusion methods following VET04‐A2 and VET03‐A guidelines of Clinical and Laboratory Standards Institute (CLSI), respectively. Isolates were categorized as wild type (fully susceptible, WT) or non‐wild type (NWT) using normalized resistance interpretation (NRI)‐determined cut‐off values (CO_WT). Broth microdilution testing showed that only 12% of UK isolates were WT to oxolinic acid (MIC CO_WT ≤ 0.25 mg/L) and 42% were WT for oxytetracycline (MIC CO_WT ≤ 0.25 mg/L). In contrast, all the isolates tested were WT (MIC CO_WT ≤ 2 mg/L) for florfenicol, the main antimicrobial for RTFS control in the UK. Disc diffusion‐based CO_WT values were ≥51 mm for 10 μg amoxicillin, ≥44 mm for 30 μg florfenicol, ≥30 mm for 2 μg oxolinic acid and ≥51 mm for 30 μg oxytetracycline. There was a high categorical agreement between the classifications of the isolates by two testing methods for florfenicol (100%), oxytetracycline (93%) and oxolinic acid (99%).     

Virulence of Flavobacterium psychrophilum isolates in rainbow trout Oncorhynchus mykiss (Walbaum)

Flavobacterium psychrophilum, the causative agent of bacterial cold‐water disease (BCWD) in freshwater‐reared salmonids, is also a common commensal organism of healthy fish. The virulence potential of F. psychrophilum isolates obtained from BCWD cases in Ontario between 1994 and 2009 was evaluated. In preliminary infection trials of rainbow trout juveniles, significant differences (0% to 63% mortality) in the virulence of the 22 isolates tested were noted following intraperitoneal injection with 10⁸ cfu/fish. A highly virulent strain, FPG 101, was selected for further study. When fish were injected intraperitoneally with a 10⁶, 10⁷ or 10⁸ cfu/fish of F. psychrophilum FPG 101, the 10⁸ cfu/fish dose produced significantly greater mortality (p < 0.05). The bacterial load in spleen samples collected from fish every 3 days after infection was determined using rpoC quantitative polymerase chain reaction amplification and by plate counting. Bacterial culture and rpoC qPCR were highly correlated (R² = 0.92); however, culture was more sensitive than the qPCR assay for the detection of F. psychrophilum in spleen tissue. Ninety‐seven per cent of the asymptomatic and the morbid fish had splenic bacterial loads of <2.8 log₁₀ gene/copies and >3.0 log₁₀ gene copies/reaction, respectively, following infection with 10⁸ cfu/fish.     

Genetic parameters of growth and resistance to Polydora ciliata in the pearl oyster Pinctada fucata

The pearl oyster Pinctada fucata is an economically important oyster cultured for pearl production in China. In this study, we used an animal model to estimate the variance components and genetic parameters of growth traits and a sire‐dam threshold model to estimate the resistance of P. fucata to polychaete Polydora ciliata. The results showed that the heritability estimates were 0.3850 ± 0.1421 for shell width, 0.3478 ± 0.1292 for body weight and 0.3387 ± 0.1316 for resistance to P. ciliata, and significantly different from zero (P < 0.05). Both phenotypic and genetic correlations were high between shell width and body weight (0.935 and 0.883 respectively) (P < 0.01), but low between resistance and shell width (0.260 and 0.233 respectively)/body weight (0.152 and 0.083 respectively) (r < 0.30; P > 0.05). The higher heritabilities for shell width, body weight and resistance traits suggested that the three traits can be genetically improved through selective breeding, but the low correlation between growth and resistance traits suggested that the resistance trait can be effectively improved through index selection or through hybridization of two specialized lines, i.e. a high‐resistant line and a high growth line.     

Multilocus sequence typing (MLST) analysis of California Flavobacterium psychrophilum reveals novel genotypes and predominance of CC‐ST10 in California salmonid hatcheries

The Gram‐negative bacterium, Flavobacterium psychrophilum, is endemic to California, USA, where it is an important pathogen in salmonid aquaculture, especially in rainbow trout (Oncorhynchus mykiss). Disease outbreaks caused by F. psychrophilum in rainbow trout fingerlings can approach 90% mortality, resulting in millions of dollars of economic losses annually. The focus of this study was to investigate the genetic diversity of 49 F. psychrophilum isolates collected from disease outbreaks in 17 salmonid hatcheries in California, USA, from 2015 to 2018 using multilocus sequence typing. Results suggest California F. psychrophilum isolates are diverse, representing 11 distinct sequence types (STs), three of which were previously undescribed. Still, the majority of genotyped isolates (n = 41) belonged to a single clonal complex (CC), CC‐ST10, which is the largest CC worldwide and has been linked to disease outbreaks on several continents. Results of this study provide evidence of marked intraspecific genetic diversity of F. psychrophilum from California. The biological significance of this genetic variability is unclear but could have implications for future vaccine development and treatments. Further studies investigating the virulence, antigenic, and antimicrobial susceptibility profiles of F. psychrophilum are warranted to better understand the epizootiology of this pathogen in the Western United States.     

虹鳟源嗜冷黄杆菌的分离鉴定及致病性研究

为确定患病虹鳟的病原,本实验从患病鱼溃烂肌肉中分离到2株细菌,分别命名为CH06和CH07,经回归感染证实分离菌株为导致此次虹鳟患病的病原菌,并进一步对其形态特征、理化特性、分子特征、血清型及耐药性进行分析。结果显示,CH06和CH07株在TYES琼脂平板上呈煎蛋状外观,产黄色素,氧化酶和过氧化氢酶呈阳性,能水解明胶和酪蛋白,不能水解淀粉,不能利用果糖、半乳糖和七叶苷等。16S rRNA比对结果显示,CH06和CH07株与嗜冷黄杆菌模式株NBRC 15942的同源性分别为99.35%和99.42%。综合菌株理化和分子特性确定CH06和CH07株为嗜冷黄杆菌。利用多重PCR方法鉴定CH06和CH07株的血清型均为1型(Fd型);多位点序列分型(MLST)分析表明,CH06和CH07株的基因型分别为ST-12和ST-78型,且均属于CC-ST10克隆型。人工感染结果显示,CH06和CH07株对虹鳟幼鱼具有较高致病性,其半致死浓度(LD₅₀)分别为7.1×10⁵和1.1×10⁵ CFU/mL,攻毒剂量与临床病症出现时间呈反比,从人工感染实验鱼的肌肉、脾脏等组织中可重新分离到嗜冷黄杆菌。组织病理变化显示,病鱼肝细胞肿胀,空泡变性,部分肝细胞溶解坏死,细胞核溶解消失;脾脏充血、出血,淋巴细胞减少,红细胞和含铁血黄素增多;肌纤维间隙增宽、断裂、弯曲不齐,部分肌细胞肌浆溶解呈蜂窝状。CH06和CH07株对10种抗菌药物的耐药谱略有不同,均对氨苄西林和甲氧苄啶-磺胺甲噁唑敏感;CH06株对恩诺沙星、氟苯尼考等耐药,而CH07株对恩诺沙星和氟苯尼考中度敏感。本研究首次报道了我国虹鳟源嗜冷黄杆菌的分离鉴定及生物学特性,以期为虹鳟细菌性冷水病的防控提供科学依据。     

Effects of body weight on the growth and physiology of Salmo salar L. during smoltification

To study the effects of body weight on growth and physiological response mechanism of S. salar, fish of three average body weights were selected: 30.25 ± 2.12 g (SW1), 24.27 ± 1.59 g (SW2) and 18.05 ± 2.46 g (SW3). Condition factor, daily weight gain and feed conversion efficiency were significantly higher in the SW2 group compared with the other treatment groups (P < 0.05). Food conversion ratio decreased with increasing body weight, and weight and specific growth rate of body length were highest in the SW2 group (P < 0.05). Larger fish (SW1 and SW2) had higher haemoglobin concentration and number of red blood cells. The number of white blood cells, lymphocytes and neutrophils were greatest in the SW3 group (P < 0.05), suggesting that these fish had strong resistance to disease. The content of total protein, albumin and urea were significantly higher in the SW2 group compared with other groups (P < 0.05). The concentration of total cholesterol and triglyceride increased with increasing body size (P < 0.05). Hormone (T3 and T4) levels first increased significantly and then decreased in fish of various sizes (P < 0.05), and the concentration of cortisol rose, fell and then rose again in SW1 and SW2 groups. Light affected the endocrine activity of S. salar by influencing the growth performance of the fish, and smaller sized fish exhibited decreased smoltification rate and a reduced success rate of smoltification. Based on these findings, S. salar weighing about 24 g were chosen to begin the process of smoltification.     

Microencapsulation of a putative probiotic Enterobacter species,C6‐6, to protect rainbow trout,Oncorhynchus mykiss (Walbaum), against bacterial coldwater disease

Flavobacterium psychrophilum is the causative agent of bacterial coldwater disease (BCWD), which has a major impact on salmonid aquaculture globally. An Enterobacter species, C6‐6, isolated from the gut of rainbow trout, Oncorhynchus mykiss (Walbaum), has been identified as a potential probiotic species providing protection against BCWD. This study examined the effects of alginate microencapsulation on the protective efficacy of C6‐6 against BCWD in vivo when administered to rainbow trout fry orally or by intraperitoneal (IP) injection. Viable C6‐6 bacteria were microencapsulated successfully, and this process (microencapsulation) did not significantly deteriorate its protective properties as compared to the administration of non‐microencapsulated C6‐6 bacteria. Both oral and IP delivery of C6‐6 achieved significantly better protection than control treatments that did not contain C6‐6 bacteria. The highest relative percent survival (RPS) resulted from IP delivery (71.4%) and was significantly greater than the highest oral RPS (38.6%). Successful intestinal colonization was not critical to protective effects of C6‐6. The study showed that C6‐6 administration, with or without encapsulation, was a viable choice for protecting fry from BCWD especially when administered intraperitoneally.     

Genomic features of Pseudomonas sp. RGM2144 correlates with increased survival of rainbow trout infected by Flavobacterium psychrophilum

This study evaluated the probiotic potential of the biofilm formed by the strain Pseudomonas sp. RGM2144 on rainbow trout survival. When challenged with the fish pathogen Flavobacterium psychrophilum, Pseudomonas sp. RGM2144 increased rainbow trout survival to 92.7 ± 1.2% (control: 35.3 ± 9.5%, p < .0001). The draft genome of Pseudomonas sp. RGM2144 is 6.8 Mbp long, with a completeness 100% and a contamination of 0.4%. The genome contains 6122 protein-coding genes of which 3564 (~60%) have known functions. The genome and phylogeny indicate that Pseudomonas sp. RGM2144 is a new species in the Pseudomonas genus, with few virulence factors, plasmids, and genes associated with antimicrobial resistance, suggesting a non-pathogenic bacterium with protective potential. In addition, the genome encodes for 11 secondary metabolite biosynthetic gene clusters that could be involved in the inhibition of F. psychrophilum. We suggest that Pseudomonas sp. RGM2144 may be applied as a probiotic in salmonid fish farming.     

Effect of poly‐β‐hydroxybutyrate on growth,enzyme activity and intestinal microbial community of Chinese mitten crab,Eriocheir sinensis (Milne‐Edwards) juveniles

Poly‐β‐hydroxybutyrate (PHB) is microbial carbon and energy storage polymer, which can be degraded into water‐soluble β‐hydroxybutyric acid in the gastrointestinal tract of aquatic animals. A 60‐day culture experiment was performed using Chinese mitten crab, Eriocheir sinensis (Milne‐Edwards) juveniles with an average initial body weight of 0.74 ± 0.06 g which were fed a diet supplemented with 0%, 0.5%, 1%, 3% or 5% PHB. A PHB dietary supplementation of 1% and 3% significantly improved the body weight gain, moulting frequency and concomitantly reduced 2nd–3rd moulting intervals of the crabs (P < 0.05). The dietary PHB level positively related to hepatopancreatic pepsin, trypsin and lipase activity (P < 0.05). Increasing the dietary PHB also improved total superoxide dismutase activity, but reduced alkaline phosphatase and acid phosphatase activity in the serum of hemolymph (P < 0.05). A 16S rRNA gene analysis by denaturing gradient gel electrophoresis indicated that PHB supplementation led to a significantly higher range‐weighted richness, diversity and evenness of the gut bacterial community when dosed at 3% in the feed. The beneficial effects of PHB are discussed in terms of immune defense, metabolism and gut microbiota of the crabs.     

Effect of dietary supplementation of brewers yeast and nucleotide singularly on growth,survival and vibriosis resistance on juveniles of the gastropod spotted babylon (<Emphasis Type="Italic">Babylonia areolata</Emphasis>)

This study was undertaken to evaluate the use of brewers yeast and nucleotides as a growth promoter and to provide vibriosis resistance for the juveniles of gastropod spotted babylon (Babylonia areolata). Juvenile spotted babylon (0.3 g initial weight) were randomly distributed at a density of 50 snails in 45-L aquaria and fed a basic diet (40% crude protein) containing two incremental levels of 1 and 2% brewers yeast and nucleotides singularly for 4 months. After the feeding trial, snails from each treatment were challenged by pathogenic bacteria Vibrio alginolyticus given by intramuscular injection and kept under observation for 5 days to record clinical signs and daily mortality rates. Results indicated that the snails fed with diets supplemented with brewers yeast or nucleotides exhibited significantly greater growth than those fed the basic diet (P < 0.05) and significantly better food conversion ratios compared to snails fed the basic diet (P < 0.05). These results indicated that dietary supplementation of brewers yeast or nucleotides, at least at the tested dosages, enhanced spotted babylon growth. Supplementing the diet with 1% brewers yeast promises to provide appropriate resistance to V. alginolyticus.     

Influence of Dietary Glycyrrhetinic Acid Combined with Different Levels of Lipid on Growth,Body Composition,and Cortisol of Juvenile Channel Catfish,Ictalurus punctatus

Glycyrrhetinic acid (GA), the main active constituent of Glycyrrhiza glabra roots, is extensively used as food additives and herbal medicines. This study was carried out to evaluate the effects of different dietary levels of GA on growth and body composition of channel catfish (average weight, 1.10 ± 0.02 g). Eight diets were formulated to contain two levels of lipid (6.0 and 8.0%), and four levels of GA (0, 0.3, 0.6, and 0.9 g/kg). The experiments lasted for 60 d. Results showed that weight gain tended to decrease as dietary GA levels increased (P > 0.05) with the lowest found in fish fed 0.9 g/kg dietary GA (P < 0.05). Feed intake increased significantly as dietary GA concentrations increased up to 0.6 g/kg (P < 0.05) and viscera/body ratio, condition factor, hepatosomatic index, and intraperitoneal fat ratio decreased significantly as GA concentration increased (P < 0.05). Lipid content of whole body, viscera, and liver decreased significantly (P < 0.01) as dietary GA increased, whereas moisture content showed an opposite trend. The cortisol in fish fed 0.3 and 0.6 g/kg GA was not significantly higher than that of fish without GA supplementation, but glucose in the former two groups was higher (P < 0.05). Results of this study indicate 0.3 and 0.6 g/kg GA had a positive influence in reducing body lipid and IFR deposition in channel catfish without significant side effects on growth. The beneficial effect may be due to extending endogenous cortisol half‐life and increasing the duration of action.     

Dietary amino acid <Emphasis Type="SmallCaps">l</Emphasis>-threonine requirement of fingerling Indian catfish, <Emphasis Type="Italic">Heteropneustes fossilis</Emphasis> (Bloch) estimated by growth and biochemical parameters

An eight-week feeding experiment was conducted to quantify the dietary threonine requirement of young catfish, Heteropneustes fossilis (9.20 ± 0.85 cm, 3.60 ± 0.45 g) using isonitrogenous and isoenergetic diets [40% crude protein (CP); 4.28 kcal g/100 g, gross energy (GE)] containing casein, gelatin and l-crystalline amino acids. Six dietary treatments supplemented with graded levels of l-threonine (0.50, 0.75, 1.00, 1.25, 1.50 and 1.75 g per 100 g, dry diet), in gradations of 0.25 g per 100 g dry diet were formulated. Fish were randomly stocked, in triplicate groups, in 55-l indoor polyvinyl flow-through circular tanks and fed experimental diets at 4% of their body weight divided over two equal feedings at 08:00 and 16:00 hours. Feeding schedule and ration size were worked out prior to the start of the feeding trial. Live weight gain (263%), feed conversion ratio (FCR) (1.35) and protein efficiency ratio (PER) (1.85) were significantly higher (P < 0.05) in fish fed a diet containing 1.25% dietary threonine. However, second-degree polynomial regression analysis of live weight gain, FCR, PER and body protein deposition data indicated the dietary threonine requirement to be 1.37, 1.26, 1.23 and 1.24 g per 100 g of dry diet, respectively. Whole-body moisture decreased significantly (P < 0.05) with the increase of dietary concentration up to 1.25%. A significantly (P < 0.05) higher protein content was evident in fish fed a diet containing 1.25% threonine. Body fat increased significantly (P < 0.05) with the increase of dietary concentration and was found to be highest at a 1.75% threonine concentration. A significantly (P < 0.05) higher ash content was reported at the 0.50 and 0.75% threonine levels. Body protein deposition was also found to be significantly (P < 0.05) higher at the 1.25% threonine level, followed by the 1.50% threonine level. Based on these results, it is recommended that the diet for fingerling H. fossilis should contain threonine at a level of 1.27 g per 100 g of dry diet, corresponding to 3.17 g per 100 g of dietary protein for optimum growth and efficient feed utilization. No mortality was observed during the experiment.     

Immunization of rainbow trout Oncorhynchus mykiss (Walbaum) with a crude lipopolysaccharide extract from Flavobacterium psychrophilum

Control methods for Flavobacterium psychrophilum are limited and oftentimes ineffective; hence, research efforts have focused on vaccine development. This study tested the hypothesis that a crude lipopolysaccharide (LPS) extract from F. psychrophilum will elicit a protective immune response in rainbow trout Oncorhynchus mykiss (Walbaum) against F. psychrophilum challenge. Rainbow trout (mean weight, 3 g) were immunized intraperitoneally with the following treatment and control preparations: 10 μg of crude LPS with or without Freund's complete adjuvant (FCA), 25 μg of crude LPS with or without FCA and saline with or without FCA. Immunization of fish with 10 or 25 μg of crude LPS/FCA resulted in significant antibody responses against F. psychrophilum using ELISA with a whole‐cell lysate as the coating antigen, but only minimal levels of protection were conferred following F. psychrophilum challenge at 14 weeks post immunization. Western blot analyses demonstrated that fish exhibited antibodies specific for low‐molecular mass proteins present in the crude LPS extract, but did not exhibit antibodies specific for F. psychrophilum LPS. The results indicate that higher immunization doses and/or the use of an alternative extraction method that yields larger LPS molecules (23–70 kDa) may be necessary to elicit specific antibody responses against F. psychrophilum LPS.     

The use of fenugreek seed meal in fish diets: growth performance,haematological and biochemical parameters,survival and stress resistance of common carp (Cyprinus carpio L.)

The present study was conducted to investigate the effects of fenugreek seed meal (FKSM) on growth performance, blood haematological and biochemical factors, survival and stress resistance of the common carp (2.46 ± 0.06 g) fingerlings. Four practical diets containing 0 (control), 0.5%, 1% and 1.5% FKSM were used to feed fish. Fish were fed on the tested diets at a rate of 3% of body weight for 8 weeks. Results indicated that fish fed the control diet had a significantly lower weight gain, specific growth rate and condition factor compared to the other treatments (P < 0.05). Feed conversion ratio decreased significantly in fish fed diets containing FKSM. There were no significant differences in glucose levels, red blood cell count, haematocrit and haemoglobin between treatment groups and the control group (P > 0.05). Significantly lower white blood cells count was observed in fish treatment with FKSM when compared with the control group (P < 0.05). The cholesterol level in group fed supplemented diet by 1% fenugreek significantly lower than other groups (P < 0.05). Total protein levels were significantly higher in all treatments compared to the control (P < 0.05). No mortality was recorded during the feeding trial. In salinity stress experiment, highest survival rate belong to fish fed supplemented diet by 1% fenugreek. These results indicate that FKSM can be considered as a beneficial dietary supplement for improving the growth performance and blood indices of common carp fingerling.     

Dietary Protein Level Affects the Growth Performance of Large Male Genetically Improved Farmed Tilapia,Oreochromis niloticus,Reared in Fertilized Freshwater Cages

An 8‐wk experiment was conducted to evaluate the effects of dietary protein on the growth performance, body composition, and serum biochemical indices of large male genetically improved farmed tilapia (GIFT), Oreochromis niloticus, reared in fertilized freshwater cages. Six semi‐purified diets were formulated, containing 18.05, 22.39, 25.97, 31.62, 35.97, and 39.89% protein (designated as P18, P22, P26, P30, P34, and P38, respectively). Each diet was fed to randomly selected fish (initial mean weight 216.7 g) in four cages. At the end of the experiment, the final body weights varied significantly (P < 0.05), ranging from 520 to 580 g. Fish receiving the P30 diet had the highest final weight value (580.5 ± 5.4 g). Weight gains of fish fed the P26 and P30 diets were significantly higher (P < 0.05) than those fed the P18 and P38 diets. The apparent feed efficiency of fish fed the P18 diet was significantly lower (P < 0.05) than that of fish in the other groups except the P22 group (P > 0.05). The apparent protein efficiency ratio decreased gradually with increasing dietary protein. Whole‐body moisture and ash showed no significant difference (P > 0.05), but protein and fat content were significantly affected by dietary protein level (P < 0.05). No significant differences were found for serum total protein, triglyceride, total cholesterol, glutamic‐oxaloacetic transaminase, and glutamic‐pyruvic transaminase levels among different diets (P > 0.05); only serum glucose level was affected by dietary protein levels (P < 0.05). Second‐order polynomial regression of weight gain suggested 29.3% dietary protein is adequate content for maximum growth of large male GIFT, O. niloticus.     

Efficacy of a polyvalent injectable vaccine against Flavobacterium psychrophilum administered to rainbow trout (Oncorhynchus mykiss L.)

Flavobacterium psychrophilum is one of the most important pathogens affecting cultured rainbow trout (Oncorhynchus mykiss). Recent information from UK salmonid farms showed country‐wide distribution of genetically and serologically divergent clones, which has hampered the development of a vaccine for rainbow trout fry syndrome. The current study assessed the efficacy of an injectable polyvalent vaccine containing formalin‐inactivated F. psychrophilum in rainbow trout. The vaccine was formulated with an oil adjuvant (Montanide ISA 760VG) or formalin‐killed cells alone. Duplicate groups of trout (60 ± 13 g) were given phosphate‐buffered saline or vaccine formulated with Montanide by intra‐peritoneal (i.p.) injection and challenged by intra‐muscular (i.m.) injection with a homologous and a heterologous isolate of F. psychrophilum at 525 degree days post‐vaccination (dd pv). Significant protection was achieved in vaccinated fish (p = 0.0001, RPS 76% homologous, 88% heterologous). Efficacy of the adjuvanted vaccine was also demonstrated by heterologous challenge at 1155 dd pv resulting in 100% protection, whereas survival in the un‐adjuvanted group was not significantly different from control fish. Levels of specific antibody at 1155 dd pv, as measured by ELISA, were significantly higher in the fish vaccinated with adjuvant when compared with unvaccinated fish.     

The relationship between channel catfish female body weight and relative fecundity and fry production when induced to ovulate with carp pituitary extract and fertilized with blue catfish sperm

ABSTRACT

Little is known regarding the correlation between channel catfish female body weight and relative fecundity and fry production when the females are induced to ovulate with carp pituitary extract and fertilized with blue catfish sperm. A total of 145 female channel catfish were used in the early, middle, and late spawning season. Female body weight of channel catfish had negative correlations with relative fecundity (r = ?0.33 and ?0.25, P < .05; r = ?0.23, P > .05) and before the late spawning season with fry/kg (r = ?0.21, P > .05; r = ?0.30, P < .05). Eggs/kg female body weight and fry/kg female body weight followed a parallel pattern throughout the season, illustrating the logical relationship between relative fecundity and fry output. The size of females used is a consideration to maximize hybrid catfish embryo production during the early and middle spawning season but not late in the spawning season. However, these correlations are relatively small even though significant; thus a dramatic change in fry production would not be predicted. Since reproductive relationships can change during the spawning season, conclusions and recommendations could be flawed if data are not collected periodically during an entire spawning season.     

Zebrafish (Danio rerio) as an animal model for bath infection by Flavobacterium psychrophilum

Flavobacterium psychrophilum is the causative agent of bacterial cold-water disease and rainbow trout syndrome in freshwater salmonid fish worldwide, generating injuries and high mortality rates. Despite several studies on this bacterium, the infection mechanism remains unknown due to limitations in the employed animal models. In this work, we propose using zebrafish (Danio rerio) as a model for studying bacterial pathogenicity. To substantiate this proposal, zebrafish infection by F. psychrophilum strain JIP 02/86 was characterized. Zebrafish larvae were infected using the bath method, and morphological changes and innate immune system activation were monitored using transgenic fish. Salmonid-like infection phenotypes were observed in 4.74% of treated larvae, as manifested by fin, muscle and caudal peduncle damage. Symptomatic and dead larvae accounted for 1.35% of all challenged larvae. Interestingly, infected larvae with no infection phenotypes showed stronger innate immune system activation than specimens with phenotypes. A failure of function assay for myeloid factor pu.1 resulted in more infected larvae (up to 43.5%), suggesting that low infection rates by F. psychrophilum would be due to the protective actions of the innate immune system against this bacterium in zebrafish larvae. Our results support the use of zebrafish as an infection model for studying F. psychrophilum. Furthermore, the percentage of infected fish can be modulated by disturbing, to varying extents, the differentiation of myeloid cells. Using this evidence as a starting point, different aspects of the infection mechanism of F. psychrophilum could be studied in vivo.