Ranched Southern Bluefin Tuna Research Articles

Dynamics of Cardicola spp. Infection in Ranched Southern Bluefin Tuna: First Observation of C. orientalis at Transfer.

Aporocotylid blood flukes Cardicola forsteri and C. orientalis are an ongoing health concern for the Southern Bluefin Tuna (Thunnus maccoyii, SBT) industry, where infections can lead to morbidity and mortality in ranched SBT populations. This study compared blood fluke infection in SBT from two companies during the 2021 ranching season. Both companies administered the same dosage of praziquantel approximately 5 weeks after transfer, feeding with frozen baitfish daily; the only difference in the company's practices was that the pontoons were located 2.5 km apart. Infection severity was measured as prevalence and intensity by quantifying adult C. forsteri in SBT heart and copy numbers of C. forsteri and C. orientalis ITS-2 DNA in SBT heart and gills. Data from the 2018 and 2019 harvests of SBT were used to make comparisons with 2021 harvest data. Cardicola orientalis was detected at transfer and no longer detected after treatment with praziquantel. Cardicola spp. were present in 83% of sampled SBT in 2021. Both companies demonstrated similar patterns of infection, and Company A had higher prevalence and intensity of Cardicola spp. infection. Based on C. forsteri ITS-2 DNA, infection intensity at harvest was significantly greater for both companies in 2021 when compared to 2018 and 2019. Continued monitoring of Cardicola spp. in SBT and improvements in diagnostics contribute to our understanding of Cardicola spp. epizootiology and the detection of changes in treatment efficacy.

Effects of company and season on blood fluke (Cardicola spp.) infection in ranched Southern Bluefin Tuna: preliminary evidence infection has a negative effect on fish growth.

Aporocotylid blood flukes Cardicola forsteri and C. orientalis are an ongoing health concern for Southern Bluefin Tuna (SBT), Thunnus maccoyii, ranched in Australia. Therapeutic application of praziquantel (PZQ) has reduced SBT mortalities, however PZQ is not a residual treatment therefore reinfection can occur after the single treatment application. This study documents the epidemiology of Cardicola spp. infection in ranched SBT post treatment over three ranching seasons (2018, 2019 and 2021). Infection prevalence (percentage of SBT affected) and intensity (parasite load) was determined by adult fluke counts from heart, egg counts from gill filaments and the use of specific quantitative polymerase chain reaction (qPCR) for detection of C. forsteri and C. orientalis ITS-2 DNA in SBT hearts and gills. SBT Condition Index decreased as intensity of Cardicola spp. DNA in SBT gills increased, suggesting blood fluke infection had a negative effect on SBT growth (Spearman's r = -0.2426, d.f. = 138, p = 0.0041). Prevalence and intensity of infection indicated PZQ remained highly effective at controlling Cardicola spp. infection in ranched SBT, 10 years after PZQ administration began in this industry. Company A had the highest prevalence and intensity of Cardicola spp. infection in 2018, and Company G had the highest in 2019. No consistent pattern was seen in 2021. Overall, intensity of infection did not increase as ranching duration increased post treatment. Results from this study improve our knowledge of the biology of blood flukes and helps the SBT industry to modify or design new blood fluke management strategies to reduce health risks and improve performance of SBT.

Prevalence and Intensity of Cardicola spp. Infection in Ranched Southern Bluefin Tuna and a Comparison of Diagnostic Methods.

The parasitic blood flukes Cardicola forsteri and C. orientalis are an ongoing health concern for Southern Bluefin Tuna Thunnus maccoyii (SBT) ranched in Australia. In this study we compared the effect of treatment, company, and ranching year on blood fluke infections in ranched SBT. SBT were sampled during the 2018 and 2019 ranching seasons from praziquantel (PZQ) treated pontoons and untreated pontoons managed by two companies. Severity of infection was diagnosed by several criteria including adult fluke counts from hearts, egg counts from gill filaments and the use of specific quantitative polymerase chain reaction (qPCR) for detection of C. forsteri and C. orientalis ITS-2 DNA in SBT hearts and gills. PZQ treatment remains highly effective against C. forsteri infection. Prevalence and intensity of Cardicola spp. infection was lower in 2019 than 2018 for Company A in treated pontoons at week 12 and week 17 of ranching, and lower for Company A than Company B in untreated pontoons at month 5 of ranching. Results indicate re-infection may be less likely in the environment near Company A pontoons, and consistent years of treatment may have lowered the parasite load in the environment. qPCR demonstrated higher sensitivity when comparing diagnostic methods for C. forsteri in heart, and higher specificity when comparing diagnostic methods for Cardicola spp. in gills. Continuing to monitor blood fluke infections in ranched SBT can help to detect changes in drug efficacy over time and help industry to develop a best practice for treatment.

The Southern Bluefin Tuna Mucosal Microbiome Is Influenced by Husbandry Method, Net Pen Location, and Anti-parasite Treatment.

Aquaculture is the fastest growing primary industry worldwide. Marine finfish culture in open ocean net pens, or pontoons, is one of the largest growth areas and is currently the only way to rear high value fish such as bluefin tuna. Ranching involves catching wild juveniles, stocking in floating net pens and fattening for 4 to 8 months. Tuna experience several parasite-induced disease challenges in culture that can be mitigated by application of praziquantel (PZQ) as a therapeutic. In this study, we characterized the microbiome of ranched southern Bluefin Tuna, Thunnus maccoyii, across four anatomic sites (gill, skin, digesta, and anterior kidney) and evaluated environmental and pathological factors that influence microbiome composition, including the impact of PZQ treatment on microbiome stability. Southern bluefin tuna gill, skin, and digesta microbiome communities are unique and potentially influenced by husbandry practices, location of pontoon growout pens, and treatment with the antiparasitic PZQ. There was no significant relationship between the fish mucosal microbiome and incidence or abundance of adult blood fluke in the heart or fluke egg density in the gill. An enhanced understanding of microbiome diversity and function in high-value farmed fish species such as bluefin tuna is needed to optimize fish health and improve aquaculture yield. Comparison of the bluefin tuna microbiome to other fish species, including Seriola lalandi (yellowtail kingfish), a common farmed species from Australia, and Scomber japonicus (Pacific mackerel), a wild caught Scombrid relative of tuna, showed the two Scombrids had more similar microbial communities compared to other families. The finding that mucosal microbial communities are more similar in phylogenetically related fish species exposes an opportunity to develop mackerel as a model for tuna microbiome and parasite research.

The effect of different treatment strategies on Cardicola spp. (Trematoda: Aporocotylidae) infection in ranched Southern Bluefin Tuna (Thunnus maccoyii) from Port Lincoln, South Australia

The aporocotylids Cardicola forsteri and C. orientalis are considered one of the most significant health concerns for Southern Bluefin Tuna (SBT) ranched off Port Lincoln, South Australia. In this study we compared the effects of different treatment strategies on blood fluke infections in ranched SBT by sampling from one untreated and two praziquantel treated pontoons progressively throughout the 2018 season. Severity of infection was assessed relative to a number of criteria including adult fluke counts from hearts, egg counts from gill filaments and the use of specific quantitative polymerase chain reaction (qPCR) for detection of C. forsteri and C. orientalis ITS-2 DNA in SBT hearts and gills. Cardicola forsteri was the dominant species detected in this study, and intensity of C. forsteri infection in SBT was significantly higher in the untreated pontoon than the two treated pontoons from week 8 of ranching. Cardicola orientalis in SBT was rarely detected, and no significant differences were seen in mortalities or condition of SBT between pontoons. Comparing diagnostic methods for C. forsteri in SBT heart demonstrated qPCR as the gold standard. This study provides important insights into the continued efficacy of praziquantel on blood fluke infections in ranched SBT.

Annual variability of infection with Cardicola forsteri and Cardicola orientalis in ranched and wild southern bluefin tuna (Thunnus maccoyii)

Infections by blood flukes from the genus Cardicola are the primary health concern in ranched bluefin tuna. This study explores annual variability of infection by C. forsteri and C. orientalis in ranched southern bluefin tuna (SBT) from Port Lincoln, South Australia, as well as in wild SBT caught at Pedra Branca, Tasmania. A hydrolysis probe-based qPCR assay was developed and validated for the quantification of C. orientalis, C. forsteri, and C. opisthorchis ITS2 rDNA. Having identified and confirmed C. forsteri and C. orientalis as the two species known to infect SBT the assay was used to quantify their ITS2 rDNA in SBT heart and gill samples from 2013 to 2015. Significant annual variability of infection in ranched SBT hearts was identified and C. orientalis was only detected in the 2013 samples. This study also presents the first detection of C. orientalis in wild SBT. Significance–Validation of a hydrolysis probe-based qPCR assay for the detection and quantification of C. orientalis, C. forsteri, and C. opisthorchis ITS2 rDNA–First detection of C. orientalis in wild SBT–Significant annual variability in detection of Cardicola forsteri and Cardicola orientalis in ranched SBT heart from 2013, 2014, and 2015 harvests

Branchial Pathomorphology of Southern Bluefin Tuna Thunnus maccoyii (Castelnau, 1872) Infected by Helminth and Copepodan Parasites

Three metazoan parasites, a monogenean Hexostoma thynni and two species of copepods Pseudocycnus appendiculatus and Euryphorus brachypterus are known to parasitize the gills of ranched southern bluefin tuna (SBT) and other tuna species. However, there is no detailed information describing the pathological response to infection by these parasites in this species. Wild southern bluefin tuna Thunnus maccoyii (approximately 3 years of age), captured and towed to a grow-out site in the waters immediately south of Port Lincoln, South Australia were subsequently sampled (n = 10) monthly from March until August 2004 during commercial harvest operations. Longitudinal sections of gill hemibranchs with attached parasites were excised and fixed for routine histology and immunohistochemistry. Reference samples were also collected from fish displaying no signs of parasitism or other grossly observable anomalies. Two morphologically distinct granulocytes were observed and putatively identified as eosinophils and mast cells. Pathology was localized to filaments upon and immediately adjacent to parasite attachment sites. Branchial cellular responses, adjunct to the attachment of H. thynni by its opisthaptoral clamps, included hyperplasia and inflammation resulting in structural remodeling of branchial tissues. Inflammatory infiltrates were often dominated by putative eosinophils and lymphocytes when parasitized by H. thynni and P. appendiculatus. Gill associated lymphoid tissue infiltrated the lamellar regions particularly in response to helminth infection. A variable response ranging from hemorrhage with minor hyperplasia or fibroplasia and eosinophilic inflammation to a barely discernible change was seen for gill sections harboring P. appendiculatus and E. brachypterus. The magnitude of the host response to attachment by the latter was congruent with attachment proximity and parasite load. On the basis of the host responses reported here and the low intensity of infection observed in other associated studies these gill ectoparasites are currently considered a low risk for wild and ranched adult SBT.

Effect of ranching time on melanomacrophage centres in anterior kidney and spleen of Southern bluefin tuna, Thunnus maccoyii

Melanomacrophage centres (MMCs) are common in the haemopoietic tissue of fish and are an important component of the humoral immune response. Increasingly they are being studied as a biomarker of environmental stress in both wild and aquaculture fish stocks. Computer image analysis of histological sections of the head kidney and spleen of ranched Southern bluefin tuna (SBT) was used to investigate changes in MMCs over various stages of the ranching cycle. Histochemical pigment differentiation indicated haemosiderin, lipofuscin and melanin were present in MMCs of both the kidney and spleen, with kidney MMCs being highly positive for lipofuscin and melanin and spleen having high levels of haemosiderin and a lower level of melanin than kidney. The MMC area in the kidney of wild sampled SBT ranged from 0.41 ± 0.13% to 2.95 ± 0.26% and that in the spleen ranged from 0.35 ± 0.09% to 2.75 ± 0.86%. The MMC area in the spleen of SBT increased over the latter stages of ranching and at harvest compared to SBT sampled in the wild and at the end of tow. There was no statistically significant difference in the MMC area in the kidney of ranched SBT sampled at various stages. SBT appear to have a higher MMC area in haemopoietic organs than numerous other fish species, perhaps providing a greater capacity to deal with ‘wear and tear’ and increased immune-competence.

Factors affecting abundance and prevalence of blood fluke, Cardicola forsteri, infection in commercially ranched southern bluefin tuna, Thunnus maccoyii, in Australia

A survey of blood fluke, Cardicola forsteri, infection in ranched southern bluefin tuna, Thunnus maccoyii, was undertaken over three farming seasons, from March 2004 to September 2006. Analyses of covariance and logistic regression were used to explore the effects of company, year, season, time in culture, and condition index on intensity, abundance and prevalence of blood fluke infection. Average prevalence of blood fluke infection was 62.64% over the period of the survey. Average intensity was 6.20 (±0.57) fluke per infected host and the average abundance was 3.70 (±0.57) fluke per host. Year did not influence mean intensity or abundance although a significant decrease in prevalence in 2005 was evident. Tuna harvested in winter had a significantly greater abundance and prevalence of blood fluke than the tuna harvested in autumn. No effect of intensity or abundance of infection was observed on the condition of the infected tuna. A universal factor in explaining variation in C. forsteri intensity, abundance and prevalence was company. Differences in infection levels between tuna from different companies may be related to differences in husbandry measures employed on each farm, or due to different average sizes of tuna farmed by each of the companies, or due to the location of the operations.

Inter-annual and intra-annual variability in blood variables and parasitic loads of wild Thunnus maccoyii

The health of southern bluefin tuna (SBT), Thunnus maccoyii, in the Great Australian Bight was assessed. Samples were obtained over 3 years (2010–2012) and over five individual schools in 2012. Several blood variables and parasite loads were measured and used as indicators of SBT health. The health of wild SBT varied among years and among schools sampled within the same month. Haemoglobin and the humoral immune parameters lysozyme and alternative complement activity explained the majority of the variation. Haemoglobin concentration, lysozyme activity, alternative complement activity, and lactate were lower in wild SBT sampled in 2010 than in 2011 and 2012. The extent to which the health differences in wild SBT may affect ranched SBT is currently unknown. Benchmark values were set for various blood parameters and parasite loads in wild SBT.

SYBR, TaqMan, or both: Highly sensitive, non-invasive detection of Cardicola blood fluke species in Southern Bluefin Tuna (Thunnus maccoyii)

Three species of blood fluke from the genus Cardicola are known to parasitize and cause disease in Bluefin Tunas – C. forsteri, C. orientalis, and C. opisthorchis. Although initially believed to be separated by geography and host specificity, recent identification of at least two Cardicola spp. concurrently present within all three Bluefin species has raised questions concerning pathogenicity, relative abundance, and distribution of these parasites within Bluefin populations. Here, we present sensitive and differential real-time qPCR nucleic acid detection of these Cardicola spp. by targeting the ITS2 region of the parasite rDNA for PCR amplification. A limit of sensitivity of 1–5 genome copy equivelents was achieved for each of the three Cardicola species tested without cross-species or host genomic amplification. Similar sensitivity was further achieved in the presence of up to 20ng/μL non-target host gDNA using SYBR Green chemistry alone, or in the presence of up to 160ng/μL host gDNA through the utilization of a TaqMan probe common-reporter detection system. These methods were subsequently used to positively identify both C. forsteri and C. orientalis DNA in preserved samples of serum, gill, and heart from ranched Southern Bluefin Tuna Thunnus maccoyii. Both methods were more sensitive for positively and differentially identifying the presence of Cardicola spp. than either histological or heart-flush microscopy techniques previously employed, and also possess the ability to be applied in non-lethal blood sampling of these highly valued fish. This is the first report for rapid and differential molecular quantitative detection of Cardicola, and opens the potential for effective monitoring of infection in cultured bluefin populations. Further, it is anticipated that the use of SYBR Green for melt-curve analyses in conjunction with a common-reporter TaqMan assay will present a flexible, accurate, and cost-effective approach for differential detection of a variety of other pathogens in future.

Correlation of humoral immune response in southern bluefin tuna, T. maccoyii, with infection stage of the blood fluke, Cardicola forsteri.

The blood fluke, Cardicola forsteri, is a prevalent infection in ranched southern bluefin tuna. This project aimed to define the timing and intensity of the various developmental stages of C. forsteri within southern bluefin tuna as well as to relate infection to host pathology and immune response. Archival samples from several cohorts of T. maccoyii sampled from 2008 to 2010 were used in this study. The prevalence and intensity of C. forsteri infection was described using heart flushes and histological examination. Humoral immune response, i.e. C. forsteri specific antibody, lysozyme activity, and alternative complement activity, was also described. Based on the validated and detailed C. forsteri infection timeline, relationships between infection events, physiological response, and diagnosis were proposed. Immune response developed concurrently with C. forsteri infection, with the majority of physiological response coinciding with commencing egg production. Further research is needed to confirm the origin of C. forsteri antigen which is responsible for immune response development and how T. maccoyii immune response works against infection. To aide this research, further diagnostic methods for confirmation of infection need to be developed.

Moving Cages Further Offshore: Effects on Southern Bluefin Tuna, T. maccoyii, Parasites, Health and Performance

The effects of offshore aquaculture on SBT health (particularly parasitic infections and haematology) and performance were the main aim of this study. Two cohorts of ranched Southern Bluefin tuna (SBT) (Thunnus maccoyii) were monitored throughout the commercial season, one maintained in the traditional near shore tuna farming zone and one maintained further offshore. SBT maintained offshore had reduced mortality, increased condition index at week 6 post transfer, reduced blood fluke and sealice loads, and haematological variables such as haemoglobin or lysozyme equal to or exceeding near shore maintained fish. The offshore cohort had no Cardicola forsteri and a 5% prevalence of Caligus spp., compared to a prevalence of 85% for Cardicola forsteri and 55% prevalence for Caligus spp. near shore at 6 weeks post transfer. This study is the first of its kind to examine the effects of commercial offshore sites on farmed fish parasites, health and performance.

Effects of immunostimulants on ranched southern bluefin tuna Thunnus maccoyii: immune response, health and performance

Ranched southern bluefin tuna Thunnus maccoyii were fed baitfishes supplemented with vitamins (predominantly E and C) or vitamins and immunostimulants, nucleotides and β-glucans, over 12 weeks after transfer and monitored for enhancement in immune response, health and performance through their 19 week grow-out period. Fish from two different tows were sampled separately at three different sampling points: at transfer to grow-out pontoons, at 8 weeks post-transfer and at harvest, 19 weeks post-transfer. Lysozyme activity was enhanced during vitamin supplementation compared to control fish. Performance (i.e. survival, condition index and crude fat), health (i.e. blood plasma variables including pH, osmolality, cortisol, lactate and glucose) and alternative complement activity were not commonly improved through diet supplementation. There were some tow-specific improvements in performance through vitamin supplementation including survival, selected parasite prevalence and intensity, and alternative complement activity. Immunostimulant supplementation also showed a tow-specific improvement in plasma cortisol level. Tow-specific responses may suggest that life history, previous health condition and husbandry can affect the success of vitamin and immunostimulant enhancement of immune response, health and performance of ranched T. maccoyii.

Sea lice infections of wild fishes near ranched southern bluefin tuna (Thunnus maccoyii) in South Australia

In contrast with sea lice infestations of other farmed fishes, attached larval stages of sea lice on ranched southern bluefin tuna (Thunnus maccoyii) are rarely detected. In this study, we monitored sea lice on ranched T. maccoyii and surveyed wild fishes adjacent to ranching sea cages over a 3-month period in early 2009. Prevalence of the adult Caligus chiastos on tuna within a day of arrival at the ranching site was 10%; prevalence then increased significantly and peaked almost 25days later to 75%; by harvest (after a further 18–28days), prevalence decreased significantly to 0%. We collected and examined a total of 502 wild fishes outside T. maccoyii sea cages, comprising 307 Degen's leatherjackets (Thamnaconus degeni), 136 yellowtail horse mackerel (Trachurus novaezelandiae), 31 sand trevally (Pseudocaranx wrighti), 10 West Australian salmon (Arripis truttacea), 6 Port Jackson sharks (Heterodontus portusjacksoni), and a single blue mackerel (Scomber australasicus) and pilchard (Sardinops sagax); we also examined an additional 10 pilchards that were collected from the centre of Spencer Gulf and stored fresh in a T. maccoyii feed bin. Of these potential hosts, we identified adult C. chiastos only from Degen's leatherjackets; of the many larvae also occurring on this host, molecular comparison of five specimens analysing cytochrome C oxidase I region of mitochondrial DNA and five specimens analysing partial D1–D2 domains of 28S rDNA confirmed that these were C. chiastos. In contrast with the decline in infections of C. chiastos on ranched T. maccoyii near the end of March, on Degen's leatherjackets there was a significant increase in prevalence and abundance over the study period, with a peak prevalence of 97.14% and a mean abundance reaching 11.17 lice per fish near the end of April. The percentage of chalimus larvae on Degen's leatherjackets increased over the study period, ranging from 0% near the start of sampling to over 93% on the final sample date. We also recorded additional copepod infestations, including Orbitacolax williamsi on Degen's leatherjackets, Caligus sp. on sand trevally, and Dissonus nudiventris on Port Jackson sharks. We conclude that Degen's leatherjacket, which is a major scavenger of excess tuna feed, is likely to contribute to sea lice infestations of T. maccoyii.

Concurrent epizootic hyperinfections of sea lice (predominantly Caligus chiastos) and blood flukes ( Cardicola forsteri) in ranched Southern Bluefin tuna

Peaks in epizootics of sea lice (mostly Caligus chiastos) and blood flukes ( Cardicola forsteri) among Southern Bluefin tuna ( Thunnus maccoyii) appear to coincide with the onset of an increased mortality. The mortality event occurs 6–12 weeks after T. maccoyii have been transferred into static ranching pontoons from the wild. However, to date available data on parasite occurrence before commercial harvesting begins, are scant. This research gathered epizootiological data from weeks 4 to 13 post-transfer, for 153 T. maccoyii sampled from two research and four commercial pontoons. Counts of both parasites in the research pontoons reached levels far heavier than previously documented in ranched T. maccoyii. For sea lice, the prevalence in most pontoons was 100%; the highest intensity reached 495 individuals, and mean counts at the peak of the infection exceeded 265 lice per fish. Almost all of the 5407 individual lice counted were identified as adult C. chiastos (89.44% female, 10.14% male); adult females of two other species were also present, C. amblygenitalis (0.13%), in addition to an undescribed species, C. sp. (0.04%). Lice counts were correlated positively with gross eye pathology scores ( r s,151 df = 0.3394, p = 0.0000), negatively correlated with condition index ( r s,151 df = −0.5396, p = 0.0000), and positively correlated with plasma cortisol ( r s,131 df = 0.3906, p = 0.0000) and glucose ( r s,131 df = 0.2240, p = 0.0096). For the blood fluke, prevalences were less uniform than those of sea lice, with lower rates of infection at the beginning (ranging from 10% to 40%), reaching 100% mid-study, and declining again (40% in one pontoon). The highest intensity reached 441 individual flukes. Fluke counts were negatively correlated with plasma haemoglobin ( r s,151 df = −0.2436, p = 0.0051) and positively with lysozyme ( r s,151 df = 0.3013, p = 0.0019). Fluke counts were also correlated with sea lice counts ( r s,150 df = 0.3143, p = 0.0000). Peaks in these epizootics occurred near the onset of elevated mortalities, which started after 7 weeks of ranching.