Emerging diseases in Tilapia: Future prevention methods will alleviate the economic threat C. Komar*, L. Labrie, J. Ng, B. Sheehan, W. Enright and Z. Tan Intervet Norbio Singapore Cedric.komar@intervet.com Good afternoon to everyone and thanks to the organizers and to phil Klesius During the last few years, diseases in tilapia have become a threatening reality 20 minutes is a short time, and I want to take the opportunity of this talk to expose you the concept of fish vaccination without going into too much details.

Outline One observation: diseases are now a reality in tilapia farming One piece of advice: identify and understand diseases present in your fish farm One important question: what should be farmer’s expectation to a fish vaccine A few practical aspects of vaccination in tilapia First illustrate some of the many diseases causing problems in tilapia

Streptococcosis

Columnaris

Francisella spp.

Iridovirus

Parasitic diseases monogenean Oodinium sp.

Tilapia used to be considered as a hardy or disease-resistant fish But this is changing…

Major Diseases Affecting Tilapia during the Farming Cycle Phase Hatchery Sex-reversal Nursery Pre-grow-out Grow-out 0 g 1 g 10 g 100 g 1 kg Nodavirus? Flavobacterium columnare Saprolegnia & Branchiomyces Pathogen Trichodina & Dactylogyrus Edwardsiella tarda After several years of sampling all over the world, here is a table of major diseases we have found in tilapia This is a general table and presence of such or such disease might vary from one farm to the other Therefore , it is very important that one’s be very aware of diseases present in his farm Francisella spp. Iridovirus Streptococcus spp. (Sa; Si) Nocardia seriolae Note: importance of the disease is roughly in proportion to the size of the arrow bars

Outline One observation: diseases are now a reality in tilapia farming One piece of advice: identify and understand diseases present in your fish farm One important question: what should be farmer’s expectation to a fish vaccine A few practical aspects of vaccination in tilapia

Know your enemy : how to evaluate the health status of your farming operation In other words, it is critical to know and understand your enemy in order to avoid to get killed

Random sampling is not the ideal method to study disease If a disease causes 30% mortality over 12 months The probability to find 1 sick fish in a random sample of 10 fish is <1% At any given moment and even in an outbreak situation, most of the fish are healthy and it is virtually impossible to isolate any pathogens from most of the population.

How to get an accurate picture of the diseases present in your farm? Even during a disease outbreak, most fish in the population are healthy Sample moribund fish with clear external signs Samples should be processed by a professional laboratory Therefore, it is critical to have a good sampling of the diseased population. Only this type of sampling will give a good understanding of what type of disease is present in the farm

Bacterial isolation Bacterial isolation from internal organs is an important part of diagnosis

Bacterial identification and antibiogram Antibiotic Mar 05 Chloramphenicol R Nitrofurantoin S Tetracycline Flumequine Neomycine Trimethoprim - sulfamethoxazole Ampicilline AMoxicilline Gentamicine Rifampicine Kanamycine Perfloxacine Norfloxacine It allows bacterial id that can be used for vaccine development or antibiogram and antibiotic selection

PCR Analysis for Viral Disease Nodavirus positive at 426 bp Iridovirus positive at 568bp For common virus, PCR is an easy and rapid method of diagnosis

Electromicroscopy Iridovirus 500 nm Electromicroscopy is necessary for novel virus or virus for which pcr primers are not avalaible 500 nm Iridovirus

Histopathology H&E x20: lateral muscle H&E x400: eye H&E x200: brain Histopathology can give a clue on pathologies that can not be diagnosed by other methods H&E x400: eye H&E x200: brain

Factors in disease expression Nutrition It is important to keep in mind that diseases are rarely caused by one cause only They are multifactorial and clearly depend on the environment of the fish For instance, nutrition (quality and quantity) plays a clear role in the expression of diseases

Factors in disease expression Fish density Micro-organisms Fish density and micro-organisms present in the water are other important factors to consider An interesting parabol that can be drawn is the one of SARS: this disease first started in HK where the population density is 6,000 inhabitants per sqkm and not in

Factors in disease expression Fish density Micro-organisms In Mississippi where there are only 3!

Factors in disease expression Biosecurity Water quality parameters Stress level Living in a confined environment is a stressful. Diseases such as cholera which are considered to be rare in developed country will reappear if standard of living suddenly decrease, water become contaminated and people have to live in a confined environment. We have to keep in mind that similar patterns will happen to our fish and diseases are a consequence of the intensification of the system

Factors in disease expression Stress level Critical level of stress not reached: system ok Feeding As long as we keep all parameters related to the fish in some kind of equilibirum, the system is ok and we can be relatively free from diseases.. Fish: density/quality/size/handling Micro-organisms Water quality

Factors in disease expression Stress level Critical level of stress reached: system suboptimal Feeding Fish However, if there is disrupture in one or more of the parameters, the system becomes suboptimal Micro-organisms Water quality

Factors in disease expression Stress level Critical level of stress breached: situation of “disease” Feeding Fish And we can enter in a situation of diseases Micro-organisms Water quality

Fish health management Therapeutic treatment Antimicrobials Antibiotics Antiparasitics Antifungals Prevention – good health management Biosecurity/quarantine/exclusion (Responsible movement of live AA) Selection Sanitation Nutrition and feed management Vaccination Clearly, fish health management depends on a number of factors that include biosecurity, etc…

Outline One observation: diseases are now a reality in tilapia farming One piece of advice: identify and understand diseases present in your fish farm One important question: what should be farmer’s expectation to a fish vaccine A few practical aspects of vaccination in tilapia Vaccination is one of these parameters, what shall we expect from that?

The concept of vaccination “Vaccines are preparations of antigens derived from pathogenic organisms, rendered non-pathogenic by various means, which will stimulate the immune system in such a way as to increase the resistance to disease from subsequent infection by a pathogen.” A.E. Ellis, 1988 Feeding Fish Micro-organisms Water quality

The concept of vaccination Pathogens (bacteria, viruses…) Vaccination Infection Diseased Resistance to infection Vaccination does not eradicate diseases, it does not solve problems of biosecurity and water quality parameters and if we reach the limits of the system, disease will still be present. However it gives more freedom in the system if we know what are its limits. Feeding Fish Mortality Treatment Healthy Micro-organisms Water quality

The salmon industry is an excellent example of the benefits of vaccination: 95% survival

Development of Fish Vaccines Epidemiology and disease investigation Disease Information Epidemiology and disease investigation Aetiology of Disease Characterization of Isolates Definition of Disease Challenge Model Devel. Laboratory phase Vaccine Development Vaccine Production Lab Vaccination Tests Normally 5 – 8 years in R&D Pre-Licensing Studies Field and licencing phase Licensing Marketing Preparation Product Introduction

Outline One observation: diseases are now a reality in tilapia farming One piece of advice: identify and understand diseases present in your fish farm One important question: what should be farmer’s expectation to a fish vaccine A few practical aspects of vaccination in tilapia

Intervet Norbio Singapore Pte. Ltd. (operated January 2000) Seriola spp. Marine fish Tilapia Dedicated to disease research and product development for aquatic animal health in the warmwater environment

Norvax®Strep Si Onset and duration immersion

Norvax® Strep Si Onset and duration injection Data from the field have shown full protection for 14 months

Sea bass: S. iniae infections Immersion (0.5 – 10 g) Injection (15 – 100 g)

Choice of application Comparison of application methods

Vaccination window Phase Pathogen Vaccination window Nodavirus? Hatchery Sex-reversal Nursery Pre-grow-out Grow-out 0 g 1 g 10 g 100 g 1 kg Nodavirus? Flavobacterium columnare Saprolegnia & Branchiomyces Pathogen Trichodina & Dactylogyrus Edwardsiella tarda Francisella spp. Iridovirus Streptococcus spp. (Sa; Si) Nocardia seriolae Vaccination window Note: importance of the disease is roughly in proportion to the size of the arrow bars

Important aspects of vaccination Vaccination window: fish must be vaccinated before to be in contact with the disease Vaccine stimulates the specific immune system: no cross-protection between S. iniae and S. agalactiae in laboratory challenges

1 skilled person can inject 1,000 fish in one hour A team of 4 injectors working 8 hours a day can vaccinate 30,000 fish per day

Take home message Prevention is better than cure Vaccination is a tool for better health management and not a substitute for other important health management measures (biosecurity, good husbandry practices, etc…) It has been a successful disease management method in the salmon industry for the last 15 years. Vaccination in tilapia will help the sustainable development of this industry Vaccination works like an insurance: You pay an insurance policy in case that you will get sick. Fish must be vaccinated before they are in contact with the disease As for an insurance policy, vaccination is specific to certain pathogens Vaccination against Streptococcus iniae will protect against this specific disease only

Acknowledgements Support from the Intervet team and many collaborators throughout the world is highly appreciated. Thank you!