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Proposed Guidelines For

White Spot Disease Prevention & Treatment

Proposed Guidelines ForWhite Spot Disease Prevention & Treatment

Compiled by

Cawangan Penyelidikan Akuakultur & Industri

April 1999

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Prevention in hatcheries

?? Locate hatchery away from the shrimp farm.

A hatchery located near a shrimp farm is more likely to be infected during an epizootic in grow-outponds. The source of infection can be from untreated discharge water contaminated with thevirus or through terrestrial or aquatic animals. To prevcent air and water-borne transmission, thehatchery must be located far away from the grow-out ponds and have a closed system.

?? Follow good sanitary practice.

The hatchery must have a foot-bath filled with 200 ppm iodophore, lysol or any other suitabledisinfectant at rhe entrance. Anyone entering the hatchery must strictly practice good sanitaryprocedures, that is disinfect the feet and hands before entering the facility. Items regularly usedin the tanks, such as glassware and nets, must be also disinfected prior to and after use,otherwise they could be a source of infection to other tanks. The tools must be dipped in

disinfectant, such as iodophore (200 ppm) or calcium hypochlorite (200 ppm), for 5 minutes. Forconvenience there should be a few containers for disinfection and the disinfectant must bechanged frequently.

?? Treat water before use.

Water used in the hatchery must be disinfected in order to eradicate the virus and other harmfulmicrooganisms. Seawater must be filtered (sand filter/net filter), precipitated overnight,disinfected with 30 ppm calcium hypochlorite (containing 65% active chlorine) for 12 hours,neutralised by soda (Na

2S

2O

3, at 30 gm/,m

3) and aerated prior to use. Filtered water can also be

treated with ultraviolet radiation or ozonated.

?? Select virus-free broodstock.

Vertical transmission of WSD can occur from broodstock to offspiring through infected oocytes.Thus, it is important to select virus-free broodstock by screening with a sensitive diagnostictechnique such as nested polymerase chain reaction (PCR). If the broodstock spawns beforethey are screened for the virus, remedial action must be taken to screen the released eggs andlate PLs.

?? Do not import broodstock and larvae.

Many exotic disease are introduced through imported broodstock or PLs. Just one recklessintroduction can cause disease problems resulting in severe losses to the industry. Introductionof animals from neighbouring countries, even if done under proper inspection and quarantine,may not be 100% risk-proof. Similarly, movement of shrimp between different regions within acountry must also be minimised to avoid the spread of the disease.

?? Prevent infection during transportation and maintenance.

During transportation, use the original water in which the shrimp were caught. Water from othersources may be contaminated with the virus. The water used in the tanks to maintain thebroodstock must also be treated to ensure that it is virus-free.

?? Maintain broodstock from different sources in separate tanks.

In order to prevent contamination between shrimps from different sources, different batchesshould be maintained separately. Eggs released from the different batches of broodstock shouldalso be segregated.

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?? Do not use trash fish to feed broodstock.

Many species of trash fish, namely, crabs, shrimps and clams,either act as mechanical carriers of

the virus or are infected with the virus. Thus feeding trash fish to broodstock could be a source ofWSD infection. Similarly, frozen products must also be avoided since the virus does not die afterfreezing. To avoid the possibility of infection from trash fish, it must be steamed before it is fed tothe broodstock.

?? Wash eggs to minimise infection.

Usually, the virus infects the ovarian tissues during the late infection stage. In other words, theoocytes may not be infected even if all other tissues or organs are infected. In such cases, thereleased eggs are still good and only the surface of the eggs may be contaminated from thewater containing virus-tainted fecal matter and discharged tissues. Thus, removing the eggs fromthe contaminated tank and washing them properly can significantly reduce the chances ofinfection. Washing is carried out be collecting the released eggs in a fine net and dipping them in

5 ppm calcium hypochlorite for 5 min: dipping for 10 minutes in 1 ppm Egg Cleanser

?

; and finallywashing with clean seawater for 5 min before releasing the eggs into the hatching tank.

?? Use immunostimulants to enhance resistance to diseases.

Use of immunostimulants such as, -1, 3-glucan, peptidoglycan, lipopolysaccharides and, IPS?in

feed may enhance the non-specific defences of shrimps against pahogens for a short durationand reduce mortalities due to secondary infection.

?? Optimise stocking density and maintain good water quality.

Stocking larvae at lower will minimise the stress on the animals, thus giving them a higherresistance against pathogens and higher chances of survival. It is a good proactice to initialy

stock larvae at 150 000 200 000 nauplii/m

3

. The health status of shrimp also depends on thewater quality. Good water quality will help to reduce stress and minimise the occurrence ofdiseases. In a hatchery, attention must also be given to minimise temperature fluctuations andreduce organic matter. Excess feed and dead shrimp must be removed regularly. Otherparameters such as salinity, pH, dissolved oxygen, and ammonia must also be monitoredregularly and maintained at optimum levels.

?? Use chemicals to minimise horizontal transmission.

Use of some commercial products such as Happy Larvae? (at a dosage of 50 g/m3) applied from

mysis stage onwards in rearing water has shown satisfactory results in minimising horizontaltransmission among larvae.

?? Treat hatchery effluent.

The hatchery must have a treatment pond where the effluent can be treated before it isdischarged. The water must be treated with disinfectant. Discharge of untreated effluent, whichmay contain WSD virus or other harmful pathogens, will contaminate the surrounding waters, andhave a negative effect on nearby hatcheries. It is typical for a hatchery to suffer from persistentdisease problem from their own doing, because the untreated contaminated water that wasdischarged is drawn back by the same hatchery for further use.

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Prevention of WSD in grow-out pond

?? Prepare pond properly.

After harvest, the sludge left in the pond contains a high load of organic matter, toxic compoundsbacteria, parasites, viral particles, as well as many WSD virus carriers that are harmful to theshrimp. If these pernicous substances are not removed throughtly, the shrimp will suffer andbecome morbid easily during the next cycle. For example, flushing out the sludge with high-pressure water from the pond bottom without treatment will spread pathogens easily. Effectivepond preparation involves the application of 100 ppm CaO (burnt lime) after harvest, exposure ofthe pond bottom to sunlight until it dries completely, removal of the top soil of the pond bottom orploughing of the soil and mixing again with lime, and clearing up of the harmful animals especially

the virus carriers such as crabs and fish that burrow into the ground.

?? Use closed system.

Closed system here means that the water must be pre-treated to kill the virus and otherorganisms that can be carriers of the virus before the water is used in the grow-out ponds. Aclosed system is strongly recommended to avoid direct use of seawater so as to reduce thechances of viral infection. Do not use the traditional open system, since WSD is widespreadalong the coastal waters. Water must be disinfected by treating with at least 30 ppm calciumhypochlorite and then left for 3-4 days prior to use.

?? Eradicate virus carriers and practise sanitary measures.

Aquatic animals such as wild shrimp, crabs, mysids, copepods and other small crustaceans arecarriers of WSD virus. Some fish and clams are mechanical carriers of the virus. These

animals must be screened out by using a bag net of 60-80 meshes/cm2, placed at the inlet of thereservoir. Free virus particles that are present in the seawater, can be killed by applying 30 ppmcalcium hypochlorite. Larger animals, such as crabs, frogs, mudskippers and snakes, must bekept out of the farm complex by installing a fine net that encloses the ponds or the whole farm.Stray dogs should also kept out of the farm since they can get into different ponds and spread thevirus.

Farm workers must practice sanitary procedures when using nets or other tools and shoulddisinfect feet and hands as well. Spray created by paddle wheels and strong wind is anotherpossible source of virus transmission, which can be minimised by using submerged aerators.

??Stock virus-free postlarvae.

Before stocking, the PLs must be sent to a reliable laboratory for WSD detection using nestedPCR. If PLs are infected, the hatchery operator must destroy them. Never release the infectedPLs into the sea or sell them to shrimp farmers.

?? Importation of PLs from foreign countries is prohibited.

Presently, Southeast Asian countries are facing severe multiple disease problems, and WSD is ofmajor concern. Careless introduction of PLs without quarantine could bring in more diseaseproblems.

?? Do not feed trash fish and its frozen products.

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This aspect has been discussed earlier, since trash fish could be a possible source of the virus.Trash fish should be steamed or boiled before it is fed to shrimp.

?? Optimise stocking density.

Lower stocking density reduces stress on the shrimp thereby increasing their resistance topathogens. The ideal stocking density should not exceed 25-30 PLs/m

2.

?? Apply probiotics and maintain good water qual ity.

Towards the end of the year, the cooler weather with heavy rainfall in most Southeast Asiancountries usually results in sudden change of water quality parameters and causes algae crash,

which may trigger a sudden viral explosion and increase the incidence of WSD. It is shown thatthe probiotic bacteria-water system is more stable than the algae-water system. Based on thisevidence, use of probiotics does seem to help control the occurrence of WSD. In addition, theproliferation of greater quantities of beneficial bacteria among the microbes in the pond will inhibitthe growth and multiplication of other pathogenic microorganisms. Please note that probiotics arenot the magic formula for everything. Other water quality parameters such as pH, DO, salinity,alkalinity, ammonia and turbidity of water must also be maintained at optimum levels.

?? Low salinity reduces WSD infection.

Field investigations revealed that the incidence and severity of WSD was lower when shrimpwere cultured in lower salinities. It is believed that lower salinity may prevent the occurrence ofthe disease. In-depth research is being undertaken to investigate the optimum salinity range fromWSD virus propagation.

?? Monitor shrimp health with PCR screening.

During the culture period, it is necessary to monitor the disease status routinely. Samples shouldbe collected every fortnight and sent to a reliable laboratory for PCR screening. The PCRtechnique will detect early infection and enable the farm to adapt a suitable strategy to minimiselosses. Diagnosis relying only on the appearance of white spots does not help as by then theshrimps will die within a few days resulting in severe losses.

?? Polyculture shrimp with fish.

Culturing shrimp with fish in the same pond can reduce the occurrence of white spot disease, asfish prey upon the virus carriers, such as small crustaceans, without becoming infected. This will

gradually destroy and reduce the amount of the virus. The species and quantity of the fishstocked in the pond should be compatible with shrimp culture. Possible species which can becultured with black tiger shrimp are Tilapia spp., Chanos chanos(milk fish) and Mugil sp. (greymullet).

?? Enhance non-specific immune response by oral supplements.

Shrimps lack a specific defense system with almost no immunological memory but have a well-developed non-specific defense mechanism. Hence, any enhancement of its defences againstpathogens can only be for a short time through increase of phagocytic and associated enzymaticactivity (pro-phenol oxidase system). Immunostimulants incorparated into feed, as describedearlier, may help in enhancing resistance of shrimps. In addition, nutritional supplements, (such

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as vitamin C) and herbal preparations, (eg Phyllanthus spp, Calotrpis gigantica) may also help inenhancing the shrimps defences.

?? Select disease-resistant species.

UPM observations revealed that the banana shrimp Penaeus merguiensis has a higherresistance to WSD than Penaeus monodon. In many cases, epidemiological data has shownthat banana shrimps did not show remarkable white spots and mortality while P. monodonbecame moribund and succumbed to severe mortal ity in the same pond. Histopathologystudies also revealed lower severity of pathological changes in banana shrimp. Thus, if WSD ispersistent in a certain locality it is recommend that banana shrimp be cultured for a few cycles inplace of P. monodon. This is practical since the hatchery technology for this species is alsoavailable.

?? Treat pond effluent.

The pond effluent must be treated before it is discharged into the sea. The treatment pond is anessential part of the modern shrimp farming system in which several procedures are carried outnamely, disinfection (if necessary), sedimentation, oxidation, physical filtration and biologicalfiltration (accompl ished by cultivation of algae, clams and the filter-feeding or omnivorous fishes)to reduce the amount of organic matter and pathogenic microorganisms. Treatment of effluentwill help in preventing the spread of disease and also improve the water quality. If there is a

severe outbreak of WSD, use 40 ppm calcium hypochlorite or 1 ppm Dipterex? to kill thediseased shrimp and disinfect the water. The water with the dead shrimp should be dischargedonly after 7 days. After discharging the water, 100 ppm CaO is applied again to disinfect theremaining decomposed shrimp on the pond bottom, as it also is the first step for pondpreparation.

Minimising losses caused by WSD

There is no successful treatment to overcome WSD. However, appropriate measures can betaken to minimise the losses. Measures to be taken are based on the severity of infection,mortal ity rate, and other factors such as growth rate, and output-input ratio. Once WSD infectionis detected, the shrimp and the pond water must not be discharged into the sea. The followingare the possible scenarios and actions required:

?? Detection of WSD infection by PCR in the first month after stocking.

If the shrimps are lethargic or anorexic, cease growing, show clinical signs of white spot and startdyingwithin 10 days of the PCR test, kill the shrimps by using 40 ppm calcium hypochlorite or 1

ppm Dipterex? . Keep the carcass and the water for 7 days before discharging.

Generally, if white spot infection is confirmed by PCR twice in the first month of culturing, it isbetter to stop the culture since it will be difficult to achieve satisfactory yield with diseased shrimp.

?? Detection of WSD infection by PCR in the second month after stocking.

(a) The culture can be continued if the shrimp do not show abnormal activity, are active, feedingwill, exhibiting normal growth, do not have clinical signs of white spot and are not dying. In

such cases, application Iodine-12? at 0.3 ppm (repeated application at 3-4 days interval), orformalin at 70 ppm (every day) or BKC (80%) at 1 ppm in grow-out ponds is recommended.The function of these chemicals is to prevent the horizontal transmission of the virus among

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the shrimps in the pond, and also to irritate and force the weak shrimp, which are infected, toswim near the water surface so that they can be removed.

(b) If the shrimp health condition deteriorates, iethe shrimp stop feeding, the FCR is rather high,shrimp show clincal signs of white spots and start dying, and single step PCR testing ispositive, then catch as many diseased shrimps as possible by using a big net, and disinfectthe pond water using 40 ppm calcium hypochlorite or 1 ppm Dipterex? and leave for 7 daysbefore discharging.

?? Detection of WSD infection by PCR in third or fourth month.

If possible infection is detected by PCR in the third or fourth month, follow the same proceduresas suggested for the second month.

An important reminder: do not release the diseased shrimp and the pond water when the shrimpare severely infected with WSD virus. Even when disinfectant is used to kill the shrimps, the

pond water and dead shrimp cannot be released soon after the death of shrimp, because thevirus is viable for a longer time within the nuclei of deeper tissues of the shrimp body even afterits death. Thus, the dead carcass must be kept in the pond for about 7 days to let the shrimpdecay and to allow the virus to completely die by autolysis. Release of freshly killed shrimp intoopen water will result in the spread of the disease and its persistent in the same farm!