Identification and characterization of motile Aeromonas spp. isolated from farmed Nile tilapia (Oreochromis niloticus) in the Philippines.

AIMS: Motile Aeromonas septicaemia (MAS) caused by motile Aeromonas species is an important disease in farmed freshwater fish due to intensification of culture and improper farm practices. This study characterized and profiled motile Aeromonas species recovered from clinically sick tilapia farmed in the Philippines, with a view to identifying targeted disease prevention and control measures against MAS in farmed tilapia species. METHODS AND RESULTS: Sixteen isolates from diseased farmed Nile tilapia were identified as Aeromonas veronii (n = 14), Aeromonas caviae (n = 1), and Aeromonas dhakensis (n = 1). Five biochemical profiles using API 20E were exhibited by the A. veronii strains giving an unreliable identification. A high level of agreement was observed in identifying the Aeromonas strains using 16S rRNA and rpoD gene sequencing, although the latter has a higher discriminatory value. Three or more virulence genes dominated by cytotoxic enterotoxin act and aerolysin aer were detected. Different genotypes based on virulence gene clustering suggested varied mechanisms used by Aeromonas to colonize and infect or to mutualistically co-exist with the fish. Acquired multiple antibiotic resistance was found in a single A. veronii isolate. All were susceptible to enrofloxacin, oxolinic acid, florfenicol, and chloramphenicol. Tetracycline and sulfonamide resistances and class 1 integron were detected in three A. veronii isolates. CONCLUSION: Several strains of motile aeromonads, especially A. veronii, which have varied genotypes based on virulence, biochemical profile, and antibiotic resistance, are involved in MAS in natural disease outbreaks in farmed Nile tilapia in the Philippines.

Qualitative Behavioural Assessment as a welfare indicator for farmed Atlantic salmon (Salmo salar) in response to a stressful challenge.

Animal welfare assessments have struggled to investigate the emotional states of animals while focusing solely on available empirical evidence. Qualitative Behavioural Assessment (QBA) may provide insights into an animal's subjective experiences without compromising scientific rigor. Rather than assessing explicit, physical behaviours (i.e., what animals are doing, such as swimming or feeding), QBA describes and quantifies the overall expressive manner in which animals execute those behaviours (i.e., how relaxed or agitated they appear). While QBA has been successfully applied to scientific welfare assessments in a variety of species, its application within aquaculture remains largely unexplored. This study aimed to assess QBA's effectiveness in capturing changes in the emotional behaviour of Atlantic salmon following exposure to a stressful challenge. Nine tanks of juvenile Atlantic salmon were video-recorded every morning for 15 min over a 7-day period, in the middle of which a stressful challenge (intrusive sampling) was conducted on the salmon. The resultant 1-min, 63 video clips were then semi-randomised to avoid predictability and treatment bias for QBA scorers. Twelve salmon-industry professionals generated a list of 16 qualitative descriptors (e.g., relaxed, agitated, stressed) after viewing unrelated video-recordings depicting varying expressive characteristics of salmon in different contexts. A different group of 5 observers, with varied experience of salmon farming, subsequently scored the 16 descriptors for each clip using a Visual Analogue Scale (VAS). Principal Components Analysis (correlation matrix, no rotation) was used to identify perceived patterns of expressive characteristics across the video-clips, which revealed 4 dimensions explaining 74.5% of the variation between clips. PC1, ranging from 'relaxed/content/positive active' to 'unsettled/stressed/spooked/skittish' explained the highest percentage of variation (37%). QBA scores for video-clips on PC1, PC2, and PC4 achieved good inter- and intra-observer reliability. Linear Mixed Effects Models, controlled for observer variation in PC1 scores, showed a significant difference between PC1 scores before and after sampling (p = 0.03), with salmon being perceived as more stressed afterwards. PC1 scores also correlated positively with darting behaviours (r = 0.42, p < 0.001). These results are the first to report QBA's sensitivity to changes in expressive characteristics of salmon following a putatively stressful challenge, demonstrating QBA's potential as a welfare indicator within aquaculture.

Investigating the Effect of an Oxytetracycline Treatment on the Gut Microbiome and Antimicrobial Resistance Gene Dynamics in Nile Tilapia (Oreochromis niloticus).

Antibiotics play a vital role in aquaculture where they are commonly used to treat bacterial diseases. However, the impact of antibiotic treatment on the gut microbiome and the development of antimicrobial resistance in Nile tilapia (Oreochromis niloticus) over time remains to be fully understood. In this study, fish were fed a single treatment of oxytetracycline (100 mg/kg/day) for eight days, followed by a 14-day withdrawal period. Changes in the distal gut microbiome were measured using 16S rRNA sequencing. In addition, the abundance of antimicrobial resistance genes was quantified using real-time qPCR methods. Overall, the gut microbiome community diversity and structure of Nile tilapia was resilient to oxytetracycline treatment. However, antibiotic treatment was associated with an enrichment in Plesiomonas, accompanied by a decline in other bacteria taxa. Oxytetracycline treatment increased the proportion of tetA in the distal gut of fish and tank biofilms of the treated group. Furthermore, the abundance of tetA along with other tetracycline resistance genes was strongly correlated with a number of microbiome members, including Plesiomonas. The findings from this study demonstrate that antibiotic treatment can exert selective pressures on the gut microbiome of fish in favour of resistant populations, which may have long-term impacts on fish health.

Qualitative Behavioral Assessment in Juvenile Farmed Atlantic Salmon (Salmo salar): Potential for On-Farm Welfare Assessment.

There is a growing scientific and legislative consensus that fish are sentient, and therefore have the capacity to experience pain and suffering. The assessment of the welfare of farmed fish is challenging due to the aquatic environment and the number of animals housed together. However, with increasing global production and intensification of aquaculture comes greater impetus for developing effective tools which are suitable for the aquatic environment to assess the emotional experience and welfare of farmed fish. This study therefore aimed to investigate the use of Qualitative Behavioral Assessment (QBA), originally developed for terrestrial farmed animals, in farmed salmon and evaluate its potential for use as a welfare monitoring tool. QBA is a "whole animal" approach based on the description and quantification of the expressive qualities of an animal's dynamic style of behaving, using descriptors such as relaxed, agitated, lethargic, or confident. A list of 20 qualitative descriptors was generated by fish farmers after viewing video-footage showing behavior expressions representative of the full repertoire of salmon in this context. A separate, non-experienced group of 10 observers subsequently watched 25 video clips of farmed salmon, and scored the 20 descriptors for each clip using a Visual Analog Scale (VAS). To assess intra-observer reliability each observer viewed the same 25 video clips twice, in two sessions 10 days apart, with the second clip set presented in a different order. The observers were unaware that the two sets of video clips were identical. Data were analyzed using Principal Component (PC) Analysis (correlation matrix, no rotation), revealing four dimensions that together explained 79% of the variation between video clips, with PC1 (Tense/anxious/skittish-Calm/mellow/relaxed) explaining the greatest percentage of variation (56%). PC1 was the only dimension to show acceptable inter- and intra-observer reliability, and mean PC1 scores correlated significantly to durations of slow and erratic physical movements measured for the same 25 video clips. Further refinements to the methodology may be necessary, but this study is the first to provide evidence for the potential of Qualitative Behavioral Assessment to serve as a time-efficient welfare assessment tool for juvenile salmon under farmed conditions.

The Complex Influences on How We Care for Farmed Fish.

As a veterinarian and academic in aquaculture, in my personal experience, most farmers are concerned for their animals and want to take good care of them. There has been substantial improvement in the welfare of farmed fish in recent decades, but improvements have been inconsistent across culture systems and species. Where there has been a lack of progress, it is not simply due to the more obvious barriers, for example, lack of clear messages, lack of effective dissemination, or cost of implementation. Why have the good intentions of farmers and research by academics failed to improve the care of many farmed fish? The reasons would appear to be complex; however, human behavioral theory (this term is used to differentiate from animal ethology) offers both a conceptual framework and practical guidelines for improving the care of fish by influencing the behavior of farmers. Here, I present some background context and apply human behavioral theory to examples of on-farm care of fish.

Factors affecting variation in mortality of marine Atlantic salmon Salmo salar in Scotland.

Databases of site production have an important role to play in the investigation and understanding of diseases, since they store valuable amounts of disease and management data. Diseases pose an important constraint to economic expansion of aquaculture. They are dependent on the complex interacting factors of pathogen, environment, and host, and the causes of death can be related to nutritional, environmental, and genetic factors of the host or infectious agents. We examined the drivers of mortality from a single site-production database, which represented one-third of Scottish farmed salmon Salmo salar L. production in 2005, to determine whether mortality 'benchmarking' data could be generalised across sites and production cycles. We show that farm mortality records play an important role in studying mortality losses and identifying of management problems in production. We found that mortalities varied across the months of the year and with the time of year of initial stocking. Production cycles that started in the third quarter of the year had the highest mortality overall. Furthermore, we found site-to-site variation in mortality that may have been caused by either random occurrence of epidemics and environmental events or other local effects.

Human Streptococcus agalactiae strains in aquatic mammals and fish.

BACKGROUND: In humans, Streptococcus agalactiae or group B streptococcus (GBS) is a frequent coloniser of the rectovaginal tract, a major cause of neonatal infectious disease and an emerging cause of disease in non-pregnant adults. In addition, Streptococcus agalactiae causes invasive disease in fish, compromising food security and posing a zoonotic hazard. We studied the molecular epidemiology of S. agalactiae in fish and other aquatic species to assess potential for pathogen transmission between aquatic species and humans. METHODS: Isolates from fish (n = 26), seals (n = 6), a dolphin and a frog were characterized by pulsed-field gel electrophoresis, multilocus sequence typing and standardized 3-set genotyping, i.e. molecular serotyping and profiling of surface protein genes and mobile genetic elements. RESULTS: Four subpopulations of S. agalactiae were identified among aquatic isolates. Sequence type (ST) 283 serotype III-4 and its novel single locus variant ST491 were detected in fish from Southeast Asia and shared a 3-set genotype identical to that of an emerging ST283 clone associated with invasive disease of adult humans in Asia. The human pathogenic strain ST7 serotype Ia was also detected in fish from Asia. ST23 serotype Ia, a subpopulation that is normally associated with human carriage, was found in all grey seals, suggesting that human effluent may contribute to microbial pollution of surface water and exposure of sea mammals to human pathogens. The final subpopulation consisted of non-haemolytic ST260 and ST261 serotype Ib isolates, which belong to a fish-associated clonal complex that has never been reported from humans. CONCLUSIONS: The apparent association of the four subpopulations of S. agalactiae with specific groups of host species suggests that some strains of aquatic S. agalactiae may present a zoonotic or anthroponotic hazard. Furthermore, it provides a rational framework for exploration of pathogenesis and host-associated genome content of S. agalactiae strains.

Draft genome sequence of a nonhemolytic fish-pathogenic Streptococcus agalactiae strain.

Streptococcus agalactiae is a significant Gram-positive bacterial pathogen of terrestrial and aquatic animals. A subpopulation of nonhemolytic strains which appear to be pathogenic only for poikilotherms exists. We report here the first draft genome sequence of a nonhemolytic S. agalactiae isolate recovered from a diseased fish.

Mortality and fish welfare.

Mortality has received insufficient attention as a fish welfare topic. Here, we aim to prompt fish farming stakeholders to discuss fish mortalities in relation to welfare. Mortality in farmed fish populations is due to a variety of biotic and abiotic causes, although it is often difficult to differentiate between underlying and immediate causes of mortality. Most mortality appears to occur during episodes associated with disease outbreaks and critical periods (in development or production). Most causes of mortality can be assumed to be associated with suffering prior to death. As mortality rates in farmed fish populations are suspected to rank amongst the highest in commonly farmed vertebrate species, mortality should be a principal fish welfare issue. Long-term mortality rates can be used as a retrospective welfare performance indicator and short-term mortality rates as an operational welfare indicator. Scrutiny of mortality records and determining causes of death will enable action to be taken to avoid further preventable mortality. The welfare performance of fish farms should only be judged on levels of predictable and preventable mortality. Fish farmers will already be monitoring mortality due to commercial and legal requirements. As profitability in fish farming is directly linked to survival, confronting mortality should ultimately benefit both fish and farmers.

A prospective longitudinal study of "Candidatus arthromitus"-associated rainbow trout gastroenteritis in the UK.

Rainbow trout gastroenteritis (RTGE) is an emerging disease of farmed rainbow trout (Oncorhynchus mykiss) reported in Croatia, France, Italy, Spain and the UK. The impact of RTGE at the site-level varies, and daily mortalities of 0.5-1% are common. Gross lesions include severe enteritis with congestion and oedema, and the segmented filamentous bacterium "Candidatus arthromitus" has been suggested as a possible aetiological agent. This paper presents findings from a prospective longitudinal epidemiology study that took place from June to November 2006 in 12 RTGE-positive UK sites, examining the impact and presentation of the disease, spread pattern, risk factors for disease presence and severity and the efficacy of current control strategies. RTGE was present in a total of 164 production units across the 12 sites, representing 39% of all stocked units. Fish losses due to RTGE totalled 61.4tonnes. The plotting of epidemic curves revealed a propagating epidemic pattern in 51% affected units, suggesting RTGE is infectious. This was supported by risk analysis of fish transfers and site layouts, which revealed an increased risk of becoming RTGE positive after contiguity or fish transfer to and from a previously affected unit. Conditional multivariate logistic regression analysis identified eight variables significantly associated with the presence of RTGE. Seven variables were risk factors, including higher stocking densities, mortalities due to predation or handling, higher water temperature, higher daily feed input and contiguity or fish movement to and from a previous case. One variable was protective: the absence of an aeration system in a unit. General linear modelling identified a significant association of mean feed input per fish during an outbreak with RTGE cumulative mortality. Finally, great variability was observed with regards to the control strategies targeted to RTGE in these sites, suggesting an absence of a common strategy. In feed NaCl treatments apparently presented a palliative effect if given to affected fish during the clinical outbreaks, but they did not prevent recurrence.

Prevalence of zoonotic trematodes in fish from a Vietnamese fish-farming community.

The prevalence of fish-borne zoonotic trematode (FZT) metacercariae was investigated in fish farmed by rural households in Nghe An Province, located in northern Vietnam. In total, 716 fish, including tilapia (Oreochromis niloticus) and 6 carp species, i.e., grass carp (Ctenopharyngodon idellus), bighead carp (Aristichthys nobilis), mrigal (Cirrhinus mrigala), common carp (Cyprinus carpio), silver carp (Hypophthalmychthys molitrix), and roha (Labeo rohita), collected from 53 fish farms were examined. The overall prevalence of FZT metacercariae was 44.6%, ranging from 12.5% to 61.0% in fish species collected from grow-out ponds, which are the production system for growing fish from fingerling size to market size. The overall prevalence was 43.6% in fingerlings cultured in nurseries, ranging from 7.4% to 62.8% for different fish species. The FZT species recovered were heterophyids and echinostomatids and included Haplorchis pumilio, H. taichui, H. yokogawai, Centrocestus formosanus, Stellantchas musfalcatus, and Echinochasmus japonicus, all of which are intestinal flukes in humans, other mammals, and birds. This is the first report of H. yokogawai and E. japonicus in fish in Vietnam, and the first record for S. falcatus in northern Vietnam. Although a previous cross-sectional survey of the people living in these fish farm households revealed a very low prevalence of FZTs (< 1%), our results demonstrate that intestinal flukes are common in farmed fish in this area, suggesting that reservoir hosts such as dogs, cats, and pigs are more important in sustaining the life cycles of these flukes in fish farms than human hosts. This has implications for the effectiveness of control programs focused mainly on treatment of humans.

Pair-level approximations to the spatio-temporal dynamics of epidemics on asymmetric contact networks.

The process of infection during an epidemic can be envisaged as being transmitted via a network of routes represented by a contact network. Most differential equation models of epidemics are mean-field models. These contain none of the underlying spatial structure of the contact network. By extending the mean-field models to pair-level, some of the spatial structure can be contained in the model. Some networks of transmission such as river or transportation networks are clearly asymmetric, whereas others such as airborne infection can be regarded as symmetric. Pair-level models have been developed to describe symmetric contact networks. Here we report on work to develop a pair-level model that is also applicable to asymmetric contact networks. The procedure for closing the model at the level of pairs is discussed in detail. The model is compared against stochastic simulations of epidemics on asymmetric contact networks and against the predictions of the symmetric model on the same networks.