Host factors associated with the detection of Aeromonas salmonicida and Yersinia ruckeri in Ontario, Canada government fish hatcheries

This paper presents an epidemiological investigation of Ontario Ministry of Natural Resources Fish Health Laboratory data from 1981 to 1997, to determine whether fish species and age were associated with lot-level detection of Aeromonas salmonicida and Yersinia ruckeri in hatchery fish. In stepwise logistic regression, the species brook trout and back-cross (lake trout crossed with the hybrid “splake”) were more likely to test A. salmonicida-positive compared to all other species reared in the hatcheries. Similarly, the species brook trout was significantly more likely to test Y. ruckeri-positive compared to all other species. For both pathogens, the 1–5-month age group was associated significantly with detection. These findings suggest that purposive sampling of higher-risk fish lots could increase the likelihood of detecting both study pathogens.     

Rearing unit-level factors associated with bacterial gill disease treatment in two Ontario, Canada government salmonid hatcheries

Early-rearing salmonids in Ontario Ministry of Natural Resources (OMNR) fish hatcheries have been consistently affected by bacterial gill disease (BGD) (causative agent: Flavobacterium branchiophilum) for many years. Separate retrospective epidemiological investigations of BGD treatments at two OMNR fish hatcheries (Hatcheries A and B) for the 1999 production year were conducted using on-site hatchery records. Both investigations were carried out at the rearing unit-level, with early-rearing (<9 months of age) “tank-lot” as the unit of analysis to identify unique fish populations over time. Multivariable repeated measures logistic regression models were created for both hatchery datasets, controlling for lot-level and species effects. For Hatchery A, the species brook trout (Salvelinus fontinalis) and brown trout (Salmo trutta) were significantly associated with BGD treatment, as well as lower water exchange rate, and higher feeding and mortality percentages during the 2 weeks previous to BGD treatment. At Hatchery B, the species brook trout (S. fontinalis) and splake (Salvelinus namaycush × S. fontinalis) were significantly associated with BGD treatment, as well as lower individual fish weights and treatment for BGD during the previous week. These results emphasize the importance of water quality, feeding rate, fish size and prior mortality on the development of BGD. Significant hatchery and species effects were evident, and future observational research on BGD must account for these factors in their design and analysis.     

The first report of Gyrodactylus salaris Malmberg, 1957 (Platyhelminthes, Monogenea) on Italian cultured stocks of rainbow trout (Oncorhynchus mykiss Walbaum)

The monogenean Gyrodactylus salaris Malmberg, 1957 is considered one of the most important parasites of wild salmonids in the European Community due to the heavy ecological and economical damage it has inflicted on Atlantic salmon (Salmo salar) parr populations. Rainbow trout (Oncorhynchus mykiss) is susceptible to G. salaris and can act as a suitable carrier host and, consequently, its trade in EU territory is restricted in relation to the status of “recognized free” zones. Despite the economic importance of rainbow trout farming in Italy, information on the Italian gyrodactylid fauna is lacking and prior to this study, G. salaris had not been officially reported. During a routine health examination of farmed rainbow trout stock throughout Central and Northern Italy in 2004–2005, five fish farms were found to be infected with G. salaris alongside three other gyrodactylids. Morphological and molecular characterisation confirmed the presence of G. salaris, Gyrodactylus teuchis Lautraite, Blanc, Thiery, Daniel et Vigneulle, 1999 and Gyrodactylus derjavinoides Malmberg, Collins, Cunningham et Jalali, 2007, while Gyrodactylus truttae Gläser, 1974 was identified by morphological analysis only. The findings from this study extend the distribution of G. salaris within Europe and highlight the importance of the rainbow trout trade in its dissemination.     

Development and evaluation of a multiplex PCR assay for simultaneous detection of Flavobacterium psychrophilum, Yersinia ruckeri and Aeromonas salmonicida subsp. salmonicida in culture fisheries

Bacterial cold water disease, enteric red mouth disease and frunculosis are the common bacterial diseases of fish worldwide. The etiologic agents of these diseases are Flavobacterium (F.) psychrophilum, Yersinia (Y.) ruckeri and Aeromonas (A.) salmonicida subsp. salmonicida, respectively. In this study, a multiplex polymerase chain reaction (m-PCR) method with YER8/10-Fer3/4-FP1/3 primer pairs which can identify these fish pathogens simultaneously was developed and optimized. In optimized conditions, neither false specific nor nonspecific amplification occurred. The detection limits of the m-PCR method using DNA extracts from dilutions of pure cultures of bacteria were 35 pg for Y. ruckeri and F. psychrophilum and 70 pg for A. salmonicida subsp. salmonicida. It was determined that 15 CFU Y. ruckeri and F. psychrophilum and 30 CFU A. salmonicida subsp. salmonicida could be detected by m-PCR developed using genomic DNA extracted from dilutions of the suspensions. The detection limits in the presence of tissue debris were 125 CFU for Y. ruckeri and F. psychrophilum and 250 CFU for A. salmonicida subsp. salmonicida. In conclusion, we submit that the m-PCR method developed and optimized in this study can be used for accurate and rapid identification of these bacteria.     

Infections of Edwardsiella tarda among Brook Trout in Quebec

Abstract

Acute bacterial septicemia is commonly diagnosed in brook trout Salvelinus fontinalis of Quebec, Canada. The agents most commonly isolated include Aeromonas salmonicida (furunculosis), Aeromonas hydrophila (motile aeromonad septicemia), and Pseudomonas species. Septicemia in brook trout caused by the gram-negative bacterium Edwardsiella tarda was diagnosed for the first time in the province of Quebec from two different fish farms producing stock for fee fishing establishments. Affected fish displayed nonspecific lesions associated with bacterial septicemia including hemorrhages on the gills and viscera and exophthalmia. Stress-associated immunosuppression due to an increase in summer water temperatures and lack of precipitation were considered as primary causes of these disease outbreaks.     

Effects of Salinity on Yersinia ruckeri Infection of Rainbow Trout and Brown Trout

Abstract

Juvenile rainbow trout Oncorhynchus mykiss and brown trout Salmo trutta acclimated to freshwater or salinities of 9.0‰ or less were exposed to Yersinia ruckeri, the bacterial pathogen that causes enteric redmouth disease (ERM). Both species of fish were kept in the same recirculating systems after bacterial exposure. Rainbow trout mortality was significantly (P < 0.05) different in each salinity: 96.5% in freshwater, 89.5% in water of 1.1‰ salinity, 81.3% in 3.0‰ salinity, and 75.0% in 9.0‰ salinity (model SE = 1.0). All deaths occurred between 3 and 12 d after exposure to Y. ruckeri. Only 2.3% of brown trout in all salinities died, and differences among treatments were not significant. For both fish species, Y. ruckeri was isolated from liver, spleen, and trunk kidney of fish dying during this experiment, and lesions of rainbow trout were consistent with ERM. Yersinia ruckeri was not isolated from brown trout surviving for 21 d after bacterial exposure but was isolated from 3 of 24 surviving rainbow trout; a polymerase chain reaction assay detected the DNA of Y. ruckeri in 3 additional rainbow trout survivors. Neither the lesions of fish with ERM nor the percentage of surviving fish subclinically infected with Y. ruckeri was affected by salinity. Bacterial growth in vitro was not affected by low (≤9.0‰) salinity; however, bacterial adhesion to polystyrene was significantly reduced as salinity increased. Although mortality caused by Y. ruckeri was significantly lower for rainbow trout in water with slightly increased salinity, none of the salinities tested was effective in preventing serious losses caused by this pathogen in recirculating systems.     

鲤鱼杀鲑气单胞菌的分离鉴定及耐药性分析

为探究鲤鱼病原菌的生物学特性及耐药状况,从患病鲤鱼中分离到菌株CS126,对其进行形态学观察、生理生化性质测定、16SrDNA序列分析及药敏性测定。结果显示,菌株CS126为革兰氏阴性菌,不产生吲哚、不具有动力性,分解半乳糖、甘露醇,VP试验、赖氨酸脱羧酶、氧化酶阴性,七叶苷阳性。16SrDNA序列长度为1 459bp,GenBank登录号为KJ942580,与GenBank中杀鲑气单胞菌的相似性高达100%,进化树显示与杀鲑气单胞菌杀鲑亚种聚为一分支。这表明菌株CS126为杀鲑气单胞菌杀鲑亚种。该菌株对菌必治、诺氟沙星、氟苯尼考等8种药物高度敏感,但对氨苄西林、氨苄西林/舒巴坦、甲氧卞胺嘧啶和磺胺异恶唑等耐药。本试验结果为鱼类杀鲑气单胞菌的快速鉴定及鱼类疾病诊治提供参考依据。     

Journal of Aquatic Animal Health: Guide for Authors 2014

Abstract

Brook trout Salvelinus fontinalis were treated with single 60-min static baths of 250 mg formalin/L, 3% NaCl, and 15 mg Chloramine-T/L to evaluate the efficacy of these compounds against external infections of Aeromonas salmonicida. Prevalence of A. salmonicida was significantly lower in brook trout treated with Chloramine-T than among those treated with formalin or salt. Further laboratory tests substantiated the therapeutic value of a single treatment of ChloramineT (15 mg/L) against A. salmonicida. In two experiments, viable counts of A. salmonicida in mucus did not vary among replicate groups of treated brook trout, but the counts for treated fish were significantly (P < 0.05) lower than those for untreated controls. In vitro tube dilution assays indicated that mean minimum inhibitory concentrations of Chloramine-T for 10 isolates of A. salmonicida were 9.0 mg/L for 1 h and 2.25 mg/L, for 24 h. In field trials at the White River National Fish Hatchery (Bethel, Vermont), the pathogen was detected principally as an external infection of juvenile Atlantic salmon Salmo solar maintained in two culture ponds. In one pond, the bacterium accounted for 100% of the total distribution of tnicroflora isolated from mucus. Seven days after treatment with Chloramine-T, A. sahnonicida accounted for 11% of the total bacterial counts identified from these fish. In the second pond, A. salmonicida composed 3% of the counts of bacteria isolated from the mucus of fish before treatment but was not isolated after treatment.     

Application of the California mastitis test in intramammary Streptococcus agalactiae and Staphylococcus aureus infections of camels (Camelus dromedarius) in Kenya

A study was conducted on 207 lactating camels in six herds in Kenya to evaluate the California mastitis test (CMT) for the detection of intramammary infections (IMIs) caused by Streptococcus agalactiae and Staphylococcus aureus and to investigate the prevalence of both the pathogens in the camel udder. IMI with S. agalactiae was found in 12% of all camels sampled. IMI with S. aureus was present in 11% of all camels sampled. The herd-level prevalence of IMI varied between 0 and 50% for S. agalactiae and between 0 and 13% for S. aureus. Longitudinal observations over 10–12 months confirmed persistent infections for both pathogens. Observations in one herd suggested that camel pox was a contributing factor in spreading and exacerbating S. agalactiae udder infections.The CMT had quarter-level sensitivities of 77 and 68% for S. agalactiae and S. aureus in camels, respectively. The CMT specificities were 91% for both the pathogens.     

Cell-Surface-Associated Properties of Fish Pathogenic Bacteria

Abstract

Pathogenicity assays showed that 33 of 42 potentially pathogenic strains of bacteria tested were virulent to rainbow trout Oncorhynchus mykiss. Regardless of their degree of virulence to fish, strains of motile Aeromonas, A. salmonicida, and Vibrio anguillarum were moderately hydrophobic. Only 46 and 25°10 of the strains were able to hemagglutinate human and trout erythrocytes, respectively. Hydrophobicity and hemagglutination were practically absent in isolates of Yersinia ruckeri. A notable number of the strains positively adhered to salmonid (51%) and nonsalmonid (55%) fish cells. Whereas the treatment of the bacteria with proteinase K or trypsin did not decrease the hydrophobicity of the isolates, within motile Aeromonas and A. salmonicida species, strains with both protease-sensitive and -resistant hemagglutinating and adhesive abilities occurred. The effects of heat and sugars on hemagglutinating and hydrophobic properties varied within all bacterial groups. Although treatment of strains with D-mannose or L-fucose had distinct effects on adhesiveness according to the bacterial species and the cell system used, none of the heat-treated (80°C for 15 min) bacteria lost their capacity to adhere to cultured fish cells. The results showed that there was no direct relationship between any of the cell surface properties analyzed and the degree of virulence of the strains.     

Newly emerged nulliparous Culicoides imicola Kieffer (Diptera: Ceratopogonidae) with pigmented abdomen

The method of segregating nulliparous and parous females of Culicoides spp. based on the presence of burgundy-red pigment inside the abdominal wall of parous Culicoides midges, is used worldwide. Out of 320 females of Culicoides imicola trapped by emergence traps, set over an artificial breeding site for 10 and 24 days, 73 (22.8%) showed a red-pigmentation despite the fact that they were nulliparous. This finding indicated that 23% of the “parous” females that are examined for the presence of arboviruses and other pathogens or for age-grading purposes, are actually old nulliparous females, which had no chance of acquiring pathogens. This bias in parous rate distorts upward the calculation of vectorial capacity.     

Seroprevalence of antibodies to Encephalitozoon cuniculi and Encephalitozoon intestinalis in humans and animals

The presence of antibodies against Encephalitozoon cuniculi (E. cuniculi) and Encephalitozoon intestinalis (E. intestinalis) was examined in 215 samples from humans and in 488 samples from five different species of domestic and companion animals in Slovakia. The 215 human samples and samples from 90 swine, 123 non-infected cattle (cattle), 24 cattle infected with bovine leukosis virus (BLV-positive cattle), 140 sheep and 111 dogs were examined by the enzyme-linked immunosorbent assay (ELISA). Samples with serum titres 1:200 or higher were considered as positive. Specific anti-E. cuniculi antibodies were found in humans (0.9%), swine (52%), cattle (2%), sheep (9%) and dogs (15%) except for the BLV-positive cattle at the titre of 1:200. The titre of 1:400 was detected only in humans (0.5%). The presence of specific anti-E. intestinalis antibodies at the titre of 1:200 was confirmed in humans (6%), swine (51%), cattle (11%), BLV-positive cattle (13%) and dogs (6%) but not in sheep. The anti-E. intestinalis antibodies reached the 1:400 in humans (1%), swine (4%) and BLV-positive cattle (17%). The presence of specific anti-E. intestinalis antibodies at the titre of 1:600 was observed only in one swine (1%). Significant differences were observed in animals at titres 1:200 and 1:400 (chi-squared test: p < 0.0001) for both pathogens and in humans only for E. cuniculi at the titre of 1:400 (chi-squared test: p < 0.0075).     

Natural infection by Cryptosporidium sp., Giardia sp. and Eimeria leuckarti in three groups of equines with different handlings in Rio de Janeiro, Brazil

To detect Cryptosporidium sp., Giardia sp. and Eimeria leuckarti in horses, fecal samples were collected from three different handling horse groups from the state of Rio de Janeiro, Brazil. Group A was composed of “Mangalarga Marchador” pure breed horses, Group B was formed by horses of a Military Corporation and Group C by stray horses captured by the Center of Zoonosis Control Paulo Dacorso Filho. A total of 396 fecal samples were collected, 212 samples from Group A, 154 samples from Group B and 30 from Group C. The material was submitted to the centrifugation - flotation technique and staining by the safranin-methylene blue technique and analyzed. Oocysts of Cryptosporidium sp. were identified in 0.75% of the samples (n = 3); cysts of Giardia sp. in 0.5% (n = 2) and oocysts of E. leuckarti in 0.5% (n = 2). One case of E. leuckarti in group A and one of Cryptosporidium sp. in group B were observed. In group C were observed two cases of Cryptosporidium, two of Giardia and one of E. leuckarti,. Horses of group C were more parasitized by the three protozoans than animals from the other groups (p < 0.01). It was possible to verify that factors related to the animals, like host individual susceptibility and sanitary factors may influence the occurrence of natural infections by gastrointestinal protozoans, although the age did not have influence. This study reports, for the first time, the occurrence of Cryptosporidium sp., Giardia sp. and E. leuckarti in equines of the State of Rio de Janeiro.     

Fast detection of Vibrio species potentially pathogenic for mollusc

Vibrio tasmaniensis, Vibrio splendidus and Vibrio neptunius species were distributed worldwide and associated with aquaculture and have been reported as the cause of diseases in aquatic organisms. Polyphasic analyses for bacterial identification are not feasible for routine diagnostic because of the time involved. The aim of this study is to design three PCR primer sets that can assist with fast detection of these species. They were designed from the 16S ribosomal RNA gene, and PCR conditions were found. Each PCR test successfully identified all the tested strains of each target species. The combined specificity of V. tasmaniensis and V. splendidus primer sets offered the best coverage (86%) in terms of separating target organisms from other related species. The primer set of V. tasmaniensis showed a lower sensitivity limit (500 fg of DNA) than the V. splendidus set (1 pg) and both sets gave positive amplification using homogenized tissues from inoculated clams, with 10² and 10⁴ cfu/g of clam, respectively. The primer set of V. neptunius was highly specific, showing only cross-reaction with V. parahaemolyticus species from 44 tested species. Its sensitivity limit was 100 pg of DNA. A small number of biochemical tests were proposed concurrently with the PCR to differentiate the cross-reacting bacteria. The time of detection of the three tested species was reduced and the further affected animals can be diagnosed in a rapid fraction of time. The detection of virulent strains of V. tasmaniensis pointed to the risk of mollusc culture outbreaks.     

The Andean hog-nosed skunk Conepatus chinga Molina, 1782 as a new definitive host for Spirometra erinacei Faust, Campbell & Kellog, 1929

This report describes the finding of Spirometra erinacei Faust, Campbell & Kellog, 1929 (Cestoda, Diphyllobothridae) infecting the small intestine of two Andean hog-nosed skunks (Conepatus chinga Molina, 1782), collected from the locality “Abra La Raya”, at Cusco, Peru. Four cestodes were studied and identified as S. erinacei. This is the first report showing that the Andean hog-nosed skunk is one of the natural hosts for this parasite.     

Detection of the causative agent of furunculusis, Aeromonas salmonicida in salmonids of the Krka River

In this paper we describe the bacterial community associated with salmonids from the Krka River. Diversity analysis demonstrated that majority of the recovered bacteria were related to Aeromonadaceae group. Bacterial analysis also revealed the presence of Shigella spp. and Pseudomonas fluorescens. Isolation of Aeromonas salmonicida from trout, presents first isolation of this bacteria Croatian rivers.     

Occurrence and Distribution of Aeromonas salmonicida Infections on Swedish Fish Farms, 1951–1987

Abstract

The occurrence of typical and atypical Aeromonas salmonicida infections on Swedish fish farms was surveyed, and the methods for controlling the disease are described here. This review is based on data for a period of 36 years from fish health control studies conducted by the National Veterinary Institute and the Salmon Research Institute. The epizootiology and routes of transmission of the pathogens are described, and policy and methods for disease control are discussed. The distribution of atypical A. salmonicida infections was entirely different from that of typical infections. There was also circumstantial evidence of different routes of transmission for typical and atypical A. salmonicida infections. Although subclinical carriers of atypical A. salmonicida occurred among Atlantic salmon Salmo salar and rainbow trout Oncorhynchus mykiss on fish farms, acute disease outbreaks were most frequent among Arctic char Salvelinus alpinus and brown trout Salmo trutta. On the other hand, acute outbreaks of typical A. salmonicida infections were frequent among Atlantic salmon and rainbow trout.     

Seroprevalence of Toxoplasma gondii infection in sheep and alpacas (Llama pacos) in Chile

Serum samples from 408 sheep from different regions of Chile and 447 alpacas (Llama pacos) from the north of the country were tested for Toxoplasma gondii antibodies. The indirect haemagglutination test (IHAT) was used in both species and the indirect immunofluorescence test (IIFT) was also used on the sheep samples in order to compare the performance of the tests in that species. In both tests, titers ≥1:16 were considered diagnostically significant. Sera from 49 sheep (12%) were positive to T. gondii antibodies by the IHAT. When using the IIFT, 114 sheep sera (28%) were positive. The different results obtained in sheep sera between the tests were significant (p<0.0001). No differences were observed between geographical locations or sex of the sampled sheep regarding serological detection of T. gondii antibodies in sheep. As expected, adult sheep showed higher T. gondii reactivity than young sheep (p=0.0008). The corrected prevalence of toxoplasmosis in alpaca was 16.3% (32 positive out of 447). The rather low prevalence in alpacas may be associated with their extensive management as well as the extreme climatic conditions of The Andes which apparently would not be favorable for the transmission of the parasite.     

Prevalence and age-related infection of Cryptosporidium suis, C. muris and Cryptosporidium pig genotype II in pigs on a farm complex in the Czech Republic

A total of 413 pig faecal samples were collected from pre-weaners (119), starters (131), pre-growers (123) and sows (40) from a farm with a closed breeding system segmented into two breeding complexes and a growing complex in the region of Vysočina, Czech Republic and screened for the presence of Cryptosporidium using staining methods and genotyping (SSU rRNA). Cryptosporidium oocysts were detected by microscopy in the faeces of 21.1% of the samples (87/413). Sequence analyses and RFLP identified C. suis in 44, Cryptosporidium pig genotype II in 23 and C. muris in 2 samples. No mixed infections were found.Pigs under 7 weeks of age were infected with C. suis only. Cryptosporidium pig genotype II was found in animals from 7 weeks of age. No relationship was found between diarrhoea and any Cryptosporidium infection in any of the different age groups (P < 0.05). The pre-weaned pigs shed significantly more Cryptosporidium oocysts than older pigs and it was associated with C. suis infection.     

In Vitro Antimicrobial Activity of Sarafloxacin against Clinical Isolates of Bacteria Pathogenic to Fish

Abstract

An in vitro susceptibility assay of sarafloxacin (A-56620), a new 4-quinolone, was performed against five important bacterial species that are pathogenic to fish. A collection of 44 clinical isolates and five corresponding type strains were included in the study. The objectives were to determine the minimal inhibitory concentrations (MICs) of sarafloxacin by a drug microdilution method and to compare the MIC values at two different temperatures, 4 and 15°C. Sarafloxacin was active against all species tested and showed the following mean MIC values at 15 and 4°C, respectively, against the bacterial pathogens investigated: Aeromonas salmonicida subspecies salmonicida, 0.029 and 0.045 μg/mL; atypical A. salmonicida, 0.053 and 0.041 μg/mL; Vibrio anguillarum, 0.085 and 0.054 μg/mL; V. salmonicida, 0.125 and 0.123 μg/mL; and Yersinia ruckeri, 0.023 and 0.031 μg/mL. The MICs ranged from 0.0025 μg/mL (or less) for two strains of A. salmonicida salmonicida to 0.32 μg/mL for one strain of atypical A. salmonicida and one strain of V anguillarum. A decrease in antimicrobial activity was observed as the incubation temperature was lowered from 15 to 4°C; however, no significant statistical difference between the measured values was demonstrated.