Aquatic Animal Diseases Research Articles

Gentamicin Combined With Hypoionic Shock Rapidly Eradicates Aquaculture Bacteria in vitro and in vivo.

Bacterial pathogens are a major cause of infectious diseases in aquatic animals. The abuse of antibiotics in the aquatic industry has led to the proliferation of antibiotic resistance. It is therefore essential to develop more effective and safer strategies to increase the efficacy and extend the life span of the antibiotics used in aquaculture. In this study, we show that six aquaculture bacterial pathogens (i.e., Aeromonas hydrophila, Vibrio alginolyticus, Edwardsiella tarda, Streptococcus iniae, Vibrio harveyi, and Vibrio fluvialis) in the stationary phase can be rapidly killed after immersion in gentamicin- or neomycin-containing, ion-free solutions for a few minutes. Such hypoionic shock treatment enhances the bacterial uptake of gentamicin in an ATP-dependent manner. Importantly, we demonstrate, as a proof of concept, that gentamicin under hypoionic shock conditions can effectively kill A. hydrophila in vivo in a skin infection model of zebrafish (Danio rerio), completely curing the infected fish. Given that pathogenic bacteria generally adhere to the skin surface and gills of aquatic animals, our strategy is of potential significance for bacterial infection control, especially for small-scale economic fish farming and ornamental fish farming. Further, the combined treatment can be completed within 5 min with a relatively small volume of solution, thus minimizing the amount of residual antibiotics in both animals and the environment.

Measures of diagnostic precision (repeatability and reproducibility) for three test methods designed to detect spring viremia of carp virus

Spring viremia of carp virus (SVCV) is a rhabdovirus of the Sprivivirus genus and the etiological agent of an internationally regulated aquatic animal disease in several fish species, including koi carp Cyprinus carpio L. The virus has a complex lifecycle with both acute and persistent stages of infection and can cause high mortality in affected populations. In this study, the diagnostic repeatability (within laboratory agreement) and reproducibility (between laboratory agreement) of 3 tests were investigated to assess their fitness as SVCV diagnostic tools. The tests, reverse transcription quantitative polymerase chain reaction (RT-qPCR) assays targeting either the SVCV glycoprotein (Q1G) or nucleoprotein (Q2N) genes and virus isolation by cell culture (VI), were performed in a blinded study with four Canadian laboratories. Test panels consisted of duplicate sets of 100 tissue samples collected from 3 SVCV prevalence populations of koi: a low-prevalence negative reference population (n = 20 fish) as well as moderate- (n = 50 fish) and high-prevalence (n = 30 fish) populations of koi experimentally infected with SVCV. The Q1G and Q2N tests were performed with kidney tissue in 3 laboratories and with brain tissue in 1 laboratory whereas pools of kidney, spleen and gill tissues were tested with the VI assay in 2 laboratories. Agreement of binary results was evaluated using the observed proportion of agreement, Cohen’s kappa and Gwet’s agreement coefficient (AC1) whereas the concordance correlation coefficient (ccc) and Bland Altman’s limit of agreement were used to evaluate agreement of the RT-qPCR continuous data. Gwet’s AC1 provided a more stable estimate of agreement than Cohen’s kappa. Overall, high repeatability (AC1, 0.78−0.90) and reproducibility (AC1, 0.74−0.89) were observed for the Q1G and Q2N tests when kidney tissue was used. Lower agreement estimates of repeatability (AC1, 0.54−0.77) and reproducibility (AC1, 0.50−0.80) were obtained for the VI test. RT-qPCR reproducibility was low with kidney-brain tissue pairs (AC1, 0.09−0.46) and high with inter-test pairs of brain (AC1, 0.76−0.86) or kidney tissue (0.75−0.86). Tissue-specific differences in virus load affected test precision and informed final tissue selection. Repeatability (ccc, 0.94−0.97) and reproducibility (ccc, 0.91−0.97) estimates of agreement for paired continuous data from the RT-qPCR assays were similarly high with kidney tissue and lower with paired brain (ccc, 0.15−0.83) and kidney-brain tissues (ccc, 0.01−0.55). The high precision of Q1G and Q2N with kidney tissue suggests that the tests are performing similarly and are suitable candidates for assessment of their diagnostic accuracy.

National Surveillance Programme For Aquatic Animal Diseases – A Stepping Stone For Establishing Disease Governance System In India

The main driver for National Surveillance Programme for Aquatic Animal Diseases (NSPAAD) in India is the threat of diseases to the growth of aquaculture. Keeping this in consideration, the Department of Fisheries (DoF), Government of India (GoI) is implementing NSPAAD involving 31 collaborating centres in 19 states, and this programme is being coordinated by ICAR-NBFGR, Lucknow. The programme has been successful in developing a network of aquatic animal health laboratories; diagnostic capability for detection of OIE/NACA-listed and emerging pathogens; mechanism for first time confirmation of new diseases; sending alerts/advisories to stakeholders; and providing scientific advice to the farmers. NSPAAD has helped in strengthening passive disease surveillance, which is evident by detection of seven new pathogens. It has enabled DoF in better understanding of disease situation in the country, and meeting international disease reporting obligations. Currently, DoF is contemplating to further strengthen the disease surveillance programme.

A preliminary assessment of indirect impacts on aquaculture species health and welfare in Scotland during COVID-19 lockdown.

COVID-19 led to sudden changes in human activities, mainly due to restrictive measures required to supress the virus. We assess the preliminary evidence for impacts on animal health and welfare in Scottish aquaculture, a key economic activity in remoter areas of the country. We summarise the industry structure, explore pathways of vulnerability to aquatic animal disease within a One Health framework that may be accentuated by impacts of COVID-19, and use basic routine data collection on the key welfare indicators of salmon mortality and parasitic sea lice counts. The indicators were published on schedule and provide no evidence of gross impact on health and welfare, at least for salmon, during the period of intensive lockdown restrictions in Scotland. Longer term effects cannot be ruled out and we do not assess impacts on the economic or social aspects of aquaculture production.

Establishment of recombinase polymerase amplification technology for detecting Viral Hemorrhagic Septicemia Virus (VHSV) in cultured aquatic animals

We established a recombinase polymerase amplification (RPA) assay for the rapid detection of viral hemorrhagic septicemia virus (VHSV) using primers designed to the N gene in this species. Optimization experiments showed that the optimal amplification temperature of the RPA assay was 37 °C and that the reaction could be completed within only 15 min. Specificity tests showed that the RPA assay did not exhibit any cross-reactivity with other infective fish viruses. Sensitivity tests further showed that the detection limit of the new RPA assay was 8.3 copies/μL, indicating that this assay was more sensitive than traditional RT-PCR method. The method was verified by detecting VHSV in 1924 batches of samples collected from domestic and imported fishes. The detection results were consistent with that of traditional RT-PCR, and the specificity and sensitivity of the method met the detection requirements for aquatic animal diseases. Collectively, our findings indicate that the novel RPA assay is fast, simple, specific, sensitive, and has significant potential for the clinic, on-site testing, and a wide range of other applications.

Seaweeds: A potent source of antimicrobial drugs for aquaculture industry

The purpose of this review is to elucidate the antimicrobial effect of seaweed extracts as a dietary supplement in aquatic animals. Aquaculture, a future of food producing sector, plays an important role in global food security, elimination of mal nutrition, and provides livelihood and employment over 59 million people around the world. Though, one side global food fish production and consumption shows an increasing trend, on the other side due to the over exploitation of available marine resources and emergence of deadly bacterial and viral disease in culture practices, the sector is leading to a great adversity which can bridge a gap between fish production and consumption. Nowadays there are several chemicals and drugs have been used to treat the diseased aquatic animals. Antibiotic usage in aquaculture practices, to mitigate the pathogenic intruders, is unsustainable and its application makes the bacteria to evolve resistance and their consumption leads to food vulnerability in human and animal. Presently, the global fish production sector is moving towards the sustainable and eco-friendly solutions to mitigate the emerging deadly intruders. However, there are many sustainable alternatives in practice such as, bacteriophage therapy, vaccine, RNA interference, use of short-chain fatty acids and polyhydroxyal kanoates to inhibit the growth of bacterial intruders, use of immune stimulants, probiotics, prebiotics, symbiotics, quorum sensing inhibition, and also the use of seaweed extract in the place of antibiotics and chemicals considered as an effective and sustainable alternative in fighting against the diseases in aquatic animals. Extracts derived from seaweeds, exerts numerous benefits over the cultivable aquatic animal which includes, promotion of growth and enhancement of the immune and other physiological performances. The present review gives an insight to the use of bio-active compounds derived from seaweeds and its beneficial effects to improve the production and immune status in aquatic animals.

Ornamental fish: a potential source of pathogenic and multidrug-resistant motile Aeromonas spp.

Aeromonas spp. are ubiquitous bacteria that cause diseases in fish and other aquatic animals. They are the natural inhabitants of different aquatic environments, such as freshwater, brackishwater and marinewater. Extrinsic stressors, such as crowding, unhygienic handling, poor water quality, polluted feeding and inadequate nutrition, can predispose fish to Aeromonas infection. In ornamental fish, motile Aeromonas spp. are known as aetiological agents of motile aeromonad infections, which cause significant mortality in fish and economic loss in the ornamental fish industry. The existence of different virulence factors leads to the virulence potential of motile Aeromonas spp. There are several antimicrobials used to treat bacterial infections in ornamental fish. However, the extensive use of antimicrobials in the ornamental fish industry causes multidrug resistance. This article reviewed a multitude of virulence factors that are related to the ornamental fish-borne Aeromonas pathogenicity and the antimicrobial resistance determinants related to the multidrug resistance phenotypes of motile Aeromonas spp. in ornamental fish.

Synergistic Effects of Medicinal Plants Carica papaya, Telferia ocidentalis and Oxytetracyline against some Pathogens Isolated from Clarias Gariepinus

The problem of fish diseases may be due to the development of antibiotic-resistant bacteria and there is an increasing awareness to determine the efficacy of plants as an alternative to treat microbial diseases of aquatic animals. Antibacterial and antifungal synergistic evaluation of methanolic and ethanolic extracts of Fluted Pumpkin Leaves (FPL), Pawpaw Leaves (PL) and combination with oxytetracycline were investigated using the agar cup diffusion and broth diffusion techniques. The extracts were tested against 8 clinical strains of Gram-positive and Gram-negative bacteria and a fungal. Minimum Inhibitory Concentration (MIC) of methanolic and ethanolic extracts of pumpkin, pawpaw leaves and combination with oxytetracycline were determined using standard methods. Data were analyzed using descriptive statistics. The extracts displayed higher antimicrobial properties to the Gram-positive and G negative bacteria and these plants inhibited the growth of all the microorganisms used in the study when combine with oxytetracycline at 10 mg/ml and 30 mg/ml. The diameter of the zone of inhibition of methanolic and ethanolic extracts of fluted pumpkin and pawpaw leaves when combine with oxytetracycline (10 mg/ml and 30 mg/ml) were higher than single application and combination of the two plants with oxytetracycline. The combination of extracts of each plant with oxytetracycline was found to be the most effective combination in retarding microbial growth of all tested pathogenic bacteria The results of MIC of methanolic and ethanolic extracts of pawpaw leaves, fluted pumpkin leaves and oxytetracycline at 10 mg/ml and 30 mg/ml range between 212.5 µg/ml - 850 µg/ml while the control doesn’t show any antimicrobial properties in terms of zone of inhibition and minimum inhibitory concentration. The results observed in the pumpkin and pawpaw leaves and combination with oxytetracycline can provide additive or synergistic inhibitory effects making them more effective as antimicrobial agents against fish pathogens.

Examples of proper reporting for evaluation (Stage 2 validation) of diagnostic tests for diseases listed by the World Organisation for Animal Health.

Reporting and design standards are key indicators of the quality of diagnostic accuracy (validation) studies but, with the exception of aquatic animal diseases and paratuberculosis in ruminants, there is limited guidance for designing these studies in animals. There is, therefore, a need for generic guidelines that are based on disease characteristics, such as mode of transmission, latent period and pathogenesis. Comprehensive, clear and transparent reporting of primary test accuracy studies for diseases listed by the World Organisation for Animal Health (OIE) has value for the end users of diagnostic tests and, ultimately, for decision-makers, who require systematic reviews and meta-analysis of multiple tests for specified diseases and testing purposes. The recent publication of reporting standards for Bayesian latent class models, to analyse test-accuracy data from naturally occurring disease events, fills an important gap as these methods are being increasingly used for OIE-listed diseases. Adherence to design and reporting standards, as well as to guidelines, helps to ensure that research funding for test validation studies is used appropriately and that the strengths and limitations of single tests or test combinations are made clear to test users. The authors provide a review of key points that are often overlooked or misinterpreted in test validation studies, as well as two concrete examples of good practice for use as a reference point for future studies.

APPLICATION OF COMPARATIVE GENOME IN AQUACULTURE DISEASES DIAGNOSIS

The sustainability of aquaculture industry is critical both for global food security and economic welfare. However, the massive wealth of pathogenic bacteria poses a key challenge to the development of a sustainable bio-control method. Recent advances in genome sequencing study combined with pan-genome analysis can be an efficacious management applied to numerous aquatic pathogens. Thus, routine comparative genome analyses of aquatic pathogens will deduce the phylogenomic diversity and possible evolutionary trendsof aquatic bacterial pathogen strains, elucidate the mechanisms of pathogenesis, as well as estimate patterns of pathogen transmission across epidemiological scales. This study also reviews comparative pan-genome analysis with a particular focus on controlling aquatic diseases, especially for: (i) re-identifying the previously misidentified strain with high accuracy and discovering novel isolates that may be associated with high rateof fish mortalities, (ii) developing routine pan-PCR based on highly informative identified genetic targets that are capable of distinguishing all the clinical isolates, and finally (iii) studying the multivalent vaccine following reverse vaccinology towards the prevention of numerous aquatic animal diseases.

Fabrication, Characterization, and Application of CuO Nano Wires as Electrode for Ammonia Sensing in Aqueous Environment Using Extended Gate-FET

This article draws attention to monitoring water-based diseases of human beings and other aquatic animals caused by a high level of ammonia present in water. A flexible sensor for measuring the level of ammonia in water or aqueous solution has been fabricated and tested. Most of the conventional sensors are either not stable or suitable for the measurement of underwater ammonia. Because of this, we intend to make a modest attempt to propose a low-cost, foldable, stable, and reliable Copper oxide (CuO) nanowire electrode-based ammonia sensitive extended gate field-effect transistor (EG-FET). In this article, CuO nanowire has been synthesized at low temperatures for realizing an extended gate for field-effect-transistor (FET). The device has been characterized for aqueous ammonia sensing at different levels of concentrations of ammonia. The sensor exhibits very high linearity [>99%] along with high sensitivity (>65 [mV]/[mg/L] and $1.62~\mu \text{A}^{1}/2$ / [mg/L]). The sensor performance has also been studied under the bending condition showing an excellent sensing performance and linearity without any performance degradation. The various characterizations have been done to validate the crystalline, uniform, and dense formation of CuO nanowires using a high-resolution scanning electron microscope (SEM) and tunnel microscope TEM. The device is well suited to measure the NH3 content (from 1.5 mg/L (or ppm) to 7.5 mg/L) in an aqueous solution.

Immunogenicity study of OmpU subunit vaccine against Vibrio mimicus in yellow catfish, Pelteobagrus fulvidraco

The outer membrane protein U (OmpU) is a conserved outer membrane protein in a variety of pathogenic Vibrio species and has been considered as a vital protective antigen for vaccine development. Vibrio mimicus (V. mimicus) is the pathogen causing ascites disease in aquatic animals. In this study, the prokaryotically expressed and purified His-tagged OmpU of V. mimicus (His-OmpU) was used as a subunit vaccine. The formalin inactivated V. mimicus, purified His tag (His-tag), and PBS were used as controls. The vaccinated yellow catfish were challenged with V. mimicus at 28 days post-vaccination, and the results showed that the His-OmpU and inactivated V. mimicus groups exhibited much higher survival rates than the His-tag and PBS groups. To fully understand the underlying mechanism, we detected the expression levels of several immune-related genes in the spleen of fish at 28 days post-vaccination and 24 h post-challenge. The results showed that most of the detected immune-related genes were significantly upregulated in His-OmpU and inactivated V. mimicus groups. In addition, we performed the serum bactericidal activity assay, and the results showed that the serum from His-OmpU and inactivated V. mimicus groups exhibited much stronger bactericidal activity against V. mimicus than those of His-tag and PBS groups. Finally, the serum agglutination antibody was detected, and the antibody could be detected in His-OmpU and inactivated V. mimicus groups with the antibody titers increasing along with the time post-vaccination, but not in His-tag or PBS group. Our data reveal that the recombinant OmpU elicits potent protective immune response and is an effective vaccine candidate against V. mimicus in yellow catfish.

Defining the aetiology of amoebic diseases of aquatic animals: trends, hurdles and best practices.

Disease caused by parasitic amoebae impacts a range of aquatic organisms including finfish, crustaceans, echinoderms and molluscs. Despite the significant economic impact caused in both aquaculture and fisheries, the aetiology of most aquatic amoebic diseases is uncertain, which then affects diagnosis, treatment and prevention. The main factors hampering research effort in this area are the confusion around amoeba taxonomy and the difficulty proving that a particular species causes specific lesions. These issues stem from morphological and genetic similarities between cryptic species and technical challenges such as establishing and maintaining pure amoeba cultures, scarcity of Amoebozoa sequence data, and the inability to trigger pathogenesis under experimental conditions. This review provides a critical analysis of how amoebae are commonly identified and defined as aetiological agents of disease in aquatic animals and highlights gaps in the available knowledge regarding determining pathogenic Amoebozoa.

OIE actions on emerging diseases of aquatic animals - The example of tilapia lake virus

OIE actions on emerging diseases of aquatic animals - The example of tilapia lake virus

Cost-effectiveness of longitudinal surveillance for Piscirickettsia salmonis using qPCR in Atlantic salmon farms (Salmo salar) in Chile.

Costs of diagnostic testing including sample collection, sampling frequency and sample size are an important consideration in the evaluation of the economic feasibility of alternative surveillance strategies for detection of infectious diseases in aquatic animals. In Chile, Piscirickettsia salmonis is the primary reason for antibiotic treatments in farmed Atlantic salmon. In 2012, a surveillance and control programme for piscirickettsiosis was established with an overall goal of reducing antibiotic use. The present study estimated the cost-effectiveness of different sampling frequencies and sample sizes to achieve at least 95% confidence of early detection of P.salmonis at the netpen and farm levels using a validated qPCR test. We developed a stochastic model that incorporated variability in test accuracy, within-pen prevalence and sampling costs. Our findings indicated that the current piscirickettsiosis surveillance programme based on risk-based sampling of five moribund or dead fish from 2 to 3 netpens is cost-effective and gives a high probability of detection of P.salmonis in Atlantic salmon farms in Chile at both the netpen and farm levels. Results from this study should incentivize salmon farmers to establish cost-effective strategies for early detection of P.salmonis infection and the application of this approach to other highly infectious diseases.

Aeromonas veronii associated with red gill disease and its induced immune response in Macrobrachium nipponense

Aeromonas veronii is a freshwater bacterium associated with many diseases in aquatic animals. In this study, the pathogenic of A. veronii was isolated from diseased Macrobrachium nipponense showing signs of weakness, poor appetite and red gills in some farms in Jiangsu province, China. The pathogenicity of A. veronii to M. nipponense was determined by challenge experiments. Results of artificial challenge showed A. veronii NJ1 was pathogenic to M. nipponense, and the LD50 was 4.5 × 106 CFU/ml at 7 d post infection. Detection virulence factors of the A. veronii NJ1 showed that A. veronii NJ1 produced caseinase, lipase, amylase, lecithinase, gelatinase and haemolysin. Detection of virulence-related genes showed that A. veronii NJ1 was positive for aerolysin (aer), outer membrane protein (ompA), adhesin (aha), flagellin (flaA, flgH, flgM, flgN), lipoidase (lip), cytotonic enterotoxins (act, alt, ast), serine protease (ser), DNases (exu), haemolysin (hlyA), type IV pilus (tapA) and fimbrillin (fim). Antibiotic sensitivity testing indicated A. veronii NJ1 was susceptible to quinolones, but resistant to penicillins. Additionally, the immune-related gene expression in M. nipponense was evaluated, and the results showed that anti-lipopolysaccharide factor (ALF1, ALF2, ALF3), Cusrin1, Cusrin2, Cusrin3, Haemocyanin, i-lysozyme and Prophenoloxidase were significantly upregulated from 6 to 24 hr after A. veronii infection. The results of our study suggested that A. veronii was an aetiological element in the mass mortalities of M. nipponense and this study provided preliminary insights into the diversity in the immune response of M. nipponense to the A. veronii infection.

Animal Coronaviruses in the Light of COVID-19.

Coronaviruses are extremely susceptible to genetic changes due to the characteristic features of the genome structure, life cycle and environmental pressure. Their remarkable variability means that they can infect many different species of animals and cause different disease symptoms. Moreover, in some situations, coronaviruses might be transmitted across species. Although they are commonly found in farm, companion and wild animals, causing clinical and sometimes serious signs resulting in significant economic losses, not all of them have been classified by the World Organization for Animal Health (OIE) as hazardous and included on the list of notifiable diseases. Currently, only three diseases caused by coronaviruses are on the OIE list of notifiable terrestrial and aquatic animal diseases. However, none of these three entails any administrative measures. The emergence of the SARS-CoV-2 infections that have caused the COVID-19 pandemic in humans has proved that the occurrence and variability of coronaviruses is highly underestimated in the animal reservoir and reminded us of the critical importance of the One Health approach. Therefore, domestic and wild animals should be intensively monitored, both to broaden our knowledge of the viruses circulating among them and to understand the mechanisms of the emergence of viruses of relevance to animal and human health.

Analysis of Streptococcal Infection and Correlation with Climatic Factors in Cultured Tilapia Oreochromis spp. in Taiwan

Tilapia (Oreochromis spp.), a prominent warm water food fish, is one of the major fish species grown in the aquaculture industry in south-east Asia. Tilapia can tolerate adverse water quality and other stressors, like diverse salinity and fluctuation of pH value, better than most other commercial aquaculture species. Environmental fluctuations are one of the main factors that affect the outbreak of infectious diseases in cultured tilapia. Cultured tilapia in Taiwan appears to be more susceptible to infections caused by Streptococci during the summer season. The present study emphasizes the Streptococcus spp. infection in tilapia in Taiwan and is the first study on the analysis of the potential impact of climate change on streptococcal infection in cultured tilapia in Asia. The data collected from the treatment and diagnosis system (TDS) of the aquatic animal diseases database from 2006 to 2015 were used to analyze the endemic streptococcal infection and the effect of climatic factors. Based on the results, the factor, average atmospheric pressure, is negatively correlated to streptococcal infection, while the other three, including average temperature, ultraviolet (UV) index, and rainfall, are positively correlated to streptococcal infection. A multivariate logistic regression model with these four factors was also built. When the average temperature is above 27.0 °C, the average atmospheric pressure is lower than 1005.1 hPa, or the UV index is above 7.2, the percentage of cumulated positive farms from all submitted tilapia cases was more than 50%. In addition, within 3 days of rain, rainfall is relevant to the occurrence of Streptococcus in tilapia. Using TDS to alert the occurrence of streptococcal infection in tilapia can be a very useful tool for veterinary aquatic animal inspection stations, and reducing economic losses and labour costs in aquatic agriculture.

Aquaculture Health Management Practices in Zambia: Status, Challenges and Proposed Biosecurity Measures

Aquaculture is an important industry in Zambia which is contributing to economic development, food security and employment in the country. Significant increase in aquaculture production has been recorded recently, although its full potential is still far from being realized. Because of this, efforts are being made by the Zambian government to make the industry a more viable venture to expand its production. As these efforts continue to make a positive impact, and aquaculture production continue to increase, diseases of aquatic animals are likely to emerge and threaten the sustainability of the industry. Here, we review the status and challenges associated with aquaculture health management practices in Zambia as well as proposed biosecurity measures farmers may adopt to minimize the likelihood of fish disease outbreaks. The paper has established that despite the country having been spared from aquaculture diseases so far, there is lack of appropriate measures to prevent and control disease outbreaks. Overall, aquaculture producers in Zambia may benefit significantly from adopting available biosecurity measures to prevent disease outbreaks in their enterprises.

Identifying Potential Polymicrobial Pathogens: Moving Beyond Differential Abundance to Driver Taxa.

It is now recognized that some diseases of aquatic animals are attributed to polymicrobial pathogens infection. Thus, the traditional view of "one pathogen, one disease" might mislead the identification of multiple pathogens, which in turn impedes the design of probiotics. To address this gap, we explored polymicrobial pathogens based on the origin and timing of increased abundance over shrimp white feces syndrome (WFS) progression. OTU70848 Vibrio fluvialis, OTU35090 V. coralliilyticus, and OTU28721 V. tubiashii were identified as the primary colonizers, whose abundances increased only in individuals that eventually showed disease signs but were stable in healthy subjects over the same timeframe. Notably, the random Forest model revealed that the profiles of thethree primary colonizers contributed an overall 91.4% of diagnosing accuracy of shrimp health status. Additionally, NetShift analysis quantified that the three primary colonizers were important "drivers" in the gut microbiotas from healthy to WFS shrimp. For these reasons, the primary colonizers were potential pathogens that contributed to the exacerbation of WFS. By this logic, we further identified a few "drivers" commensals in healthy individuals, such as OUT50531 Demequina sediminicola and OTU_74495 Ruegeria lacuscaerulensis, which directly antagonized the three primary colonizers. The predicted functional pathways involved in energy metabolism, genetic information processing, terpenoids and polyketides metabolism, lipid and amino acid metabolism significantly decreased in diseased shrimp compared with those in healthy cohorts, in concordant with the knowledge that the attenuations of these functional pathways increase shrimp sensitivity to pathogen infection. Collectively, we provide an ecological framework for inferring polymicrobial pathogens and designing antagonized probiotics by quantifying their changed "driver" feature that intimately links shrimp WFS progression. This approach might generalize to the exploring disease etiology for other aquatic animals.