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December 11, 2013

Vietnam

EMS—The Perfect Killer—A Webinar

 

On December 10, 2013, from Vietnam, the Global Aquaculture Alliance (GAA) aired a one-hour webinar titled “Early Mortality Syndrome: Managing the Perfect Killer”.  It featured a thirty-minute, 53-slide show narrated by GAA President Dr. George Chamberlain and was followed by a thirty-minute question and answer session in which Chamberlain and the University of Arizona’s Dr. Donald Lightner answered questions about the current status of the EMS epidemic.

 

Following are some excerpts from the webinar:

 

EMS, early mortality syndrome, also known as AHPNS, acute hepatopancreatic necrosis syndrome, is an epidemic disease that damages the digestive system of shrimp and causes mortality, often within thirty days of stocking.  EMS was first detected in China in 2009.  It moved to Vietnam in 2010 and then jumped to peninsular Malaysia in 2011 and East Malaysia (the northern part of the island of Borneo) in 2011.  It landed in Thailand in 2012, and then, remarkably, jumped all the way to Mexico in 2013.  Now there is news of an outbreak of EMS in India, although it has not been officially confirmed, yet.

 

 

How has EMS affected the global shrimp farming industry?  In 2013, the global production of shrimp declined by about 15% from 2011 levels, but considering that the industry was expected to grow by about five percent a year, production was actually 23 percent below market expectations.  In rough numbers, global production of farmed shrimp is about four million metric tons a year.  If it were down 25%, that’s a million metric tons.  A million metric tons is a billion kilograms.  Assuming a value of five dollars a kilogram, that’s a five billion dollar loss to the industry at the farm gate.

 

 

One of the first advances in EMS research was identifying the clinical signs of the disease.  This was important because farmers were calling any disease that occurred shortly after stocking “EMS”.  After the clinical signs were identified, an acute phase was recognized in which a toxin was released, causing the hepatopancreas to dysfunction and slough off tubal epithelial cells.  The acute phase lead to a terminal phase in which opportunistic bacteria colonized the damaged hepatopancreas, often leading to mortality.  The next step was to determine if the disease could be transmitted to other shrimp.

 

In the early period of EMS research, there was tremendous speculation about the possible cause of the disease.  Some were convinced that it was a virus; others though it was a toxin.  Toxic algae, crusticides and soil toxins were all suggested as possible causes.  All of them proved to be dead ends.  Trying to transmit the disease using frozen tissue from dead animals failed.  Then in the summer of 2012, the Responsible Aquaculture Foundation and the World Bank initiated a mission to Vietnam to investigate the disease at hatcheries and farms.  That mission was tremendously helpful in providing a lot of information about the progression of the disease and getting samples of infected animals, and it subsequently led to the breakthrough when Vietnam’s Dr. Loc succeeded in transmitting the disease using fresh tissue.  The fresh tissue was offered to healthy shrimp for five days and induced the typical clinical signs of EMS pathology.  It only worked when stomach tissue from EMS-infected shrimp was fed to healthy shrimp, which lead to the conclusion that EMS was an infectious disease caused by a bacterial agent found in the stomach of sick shrimp.  The pathogen colonizes the shrimp’s stomach and releases a toxin that damages the shrimp’s hepatopancreas.

 

Further work at Dr. Lightner’s lab at the University of Arizona succeeded in identifying the bacterial pathogen as Vibrio parahaemolyticus.  His work also demonstrated that the strain of V. parahaemolyticus that infected shrimp does not include human toxins, meaning that it does not affect people.  Further work lead to the development of a PCR diagnostic test for identifying the disease in hatchery and farm animals.  The diagnostic test is expected to become available for private sector as early as January 2014.

 

At this point, it’s valuable to look at an analogous bacterial disease caused by a luminescent Vibrio that essentially shut down the shrimp farming industry in the Philippines in the early 1990s called “Vibrio harveyi”.  Some very relevant lessons have been learned from that research.  For example, in the hatchery, Vibrio harveyi was found to colonize shrimp eggs.  It became very important to rinse the eggs, so that when the nauplii hatched, the nauplii would have a better chance of survival.  This means that controls in the hatchery will probably be very important in preventing EMS from being passed from broodstock to postlarvae.

 

Now let’s jump to a little practical experience with EMS through some information shared by Agrobest, a large shrimp farm in Malaysia, managed by Noriaki Akazawa, who was part of GOAL 2013 GAA meeting in Paris, France, in October 2013.  Mr. Akazawa described how Agrobest steadily increased its giant tiger shrimp (Penaeus monodon) production, but when it switched to white shrimp (P. vannamei), production increased even more rapidly, jumping to 11,000 metric tons by the end of 2009.  Then, in early 2010, the farm was hit by early mortality syndrome, which basically cut its production in half.  The outbreak at Agrobest was associated with a batch of postlarvae from a particular hatchery.  Other outbreaks at the farm were associated with dense blue-green algal blooms that triggered very high pH levels in the ponds.  By carefully managing the ponds to control algal blooms, bottom sludge and water quality, Agrobest was able to slowly recover and increase production to 14 mertic tons per hectare.  But then, in the middle of 2013, the farm was hit with a second outbreak of EMS and had to develop new management solutions.  Trials at Agrobest showed that tiger shrimp were never infected by EMS, even when they were raised in ponds adjacent to ponds with infected white shrimp.  The farms primary source of postlarvae is from wild broodstock, many of which are infected with monodon baculovirus and other diseases.  The farm found it difficult to buy enough clean tiger shrimp postlarvae to meet its production goals.  A reliable source of pathogen free monodon postlarvae is needed.

 

The CP Group has done an enormous amount of work on EMS, and it has established a challenge center where it can test a variety of treatments on shrimp that are intentionally exposed to the disease.  Dr. Lightner’s lab has a similar challenge center.  The work at CP and Dr. Lightner’s lab is leading to a vast accumulation of information about possible treatments for EMS.  One of the things that they learned is that the Vibrio parahaemolyticus pathogen grows very fast.  In fact, it can quickly out compete other bacteria.  When shrimp ponds are chlorinated before stocking to eliminate bacteria, V. parahaemolyticus can bounce back much faster than competing bacteria, so the disease can be worse after chlorination—if there isn’t immediate use of an appropriate probiotic to restore the diversity of the microbial community.  It’s also a colonizer and tends to stick to surfaces.  It’s found on the bottom of ponds and produces a very potent toxin, but the colonies don’t release the toxin until they reach a certain density.  They have a quorum sensing capability, which means they only release their toxins when they reach a density of log five or six.  That’s ten to the fifth or ten to the sixth colony forming units, and they do it all at once.  They don’t release the toxin until they can do major damage, giving them the name “The Perfect Killer”.

 

The CP Group has uncovered some interesting facts about EMS’s likes and dislikes.  For example, EMS does not like low salinities.  At salinities of less that five parts per thousand, EMS is not a problem, so those farms that are fortunately enough to be operating in nearly fresh water seem to have no problems with EMS.  At salinities above five parts per thousand, EMS becomes progressively more of a problem up to ten parts per thousand when it’s pretty much at full strength.  Temperature is another factor that affects EMS.  It likes hot weather and warm water, and multiplies much faster when it gets them.  It seems to be indifferent to pH, but high pH levels favor dense blue-green algae blooms and EMS does well in those blooms.  It also likes pond fertilizers, like urea, nitrogen, carbon and phosphorous, that are used to stimulate the growth of plankton in the pond.  The nutrients may actually inadvertently stimulate the growth of the EMS pathogen.  It likes water that has been flowing in and out of a concentration of ponds better than ground water or water pumped directly from the sea.  EMS is more of a problem in areas where farms share the same surface water, especially where multiple farms are using the same tributary for the intake and discharge of their water.

 

The CP Group also found that shrimp raised in cages that were off the pond bottom were much less susceptible to EMS.  The pathogen likes pond bottoms because it attaches to surfaces.  If shrimp are kept off the bottom, they have less exposure to the disease.  Then there are certain organisms that tend to concentrate the pathogen.  For example, bryozoans, organisms that live in small calcareous tubes in the bottom of the pond, filter the water and tend to concentrate the pathogen in their bodies.  When the shrimp eat the bryozoans, they can get a lethal dose of the pathogen.

 

In Thailand, the CP Group has found that the disease has been most prevalent in the intermediate zone, not in the coastal area where water is taken directly from the sea and not in inland areas where the salinities are lower, but in the area between the two where the water is brackish and many farms are pumping and discharging water from the same source.  That’s where the load of the EMS pathogen is the greatest and where the most damaging outbreaks occur.

 

In terms of managing this disease, we have to rethink our sanitary and hygiene strategies because this disease is caused by bacteria, not a virus.  With viral diseases, we must control the host—and the pathogen—because the pathogen doesn’t replicate outside the host.  For example, with the whitespot virus, if we prevent crustacean carriers from entering the pond then we know that the free viral particles will deactivate with in a matter of a few days.  This is not the case with the EMS pathogen.  It thrives in the water, outside of the host.

 

How do we move forward?  Let’s review some of the management practices that are working.  One is that larger shrimp seem to be less affected by the disease.  It’s not that they are immune; it’s that the feeding behavior of early stage shrimp is different from that of older shrimp.  Older shrimp are less exposed to the disease because their eating behavior is different.  Therefore, one technique is to use raceways or nurseries to head start the shrimp before releasing them into ponds.  Another technique is to stock postlarvae into cages in the pond and then release them when they get larger.  Another is to use a different species.  Giant tiger shrimp seem to be less affected by EMS than white shrimp.  It’s not that they are immune.  Challenge trials show that when vannamei and monodon are fed a dose of EMS, they both die, but when they’re both put into ponds the giant tiger shrimp seem less affected.  Once again this may be related to feeding behavior.  This could lead to a resurgence in giant tiger shrimp farming.  Another technique is rearing tilapia with shrimp.  This is reminiscent of the strategy of raising shrimp and tilapia together to avoid the whitespot virus.

 

There’s a lot of interest in the use of bioflocs to diversify the microbial community in shrimp ponds.  There have not been controlled studies on this, but during the World Bank/Responsible Aquaculture Foundation mission to Vietnam, we heard lots of anecdotal evidence about bioflocs making shrimp less susceptible to EMS, which makes sense because presumably the many competing species of bacteria in a biofloc pond would tend to displace the EMS pathogen.

 

There have also been reports from China that small intensive ponds with little sludge and salinities reduced to four parts per thousand have been successful with stocking densities of up to 450 postlarvae per square meter.  High yields are being achieved, even in the presence of EMS.  There is also some work being done on using recirculating system to avoid EMS.  Another technique is to breed disease resistant shrimp.  This technique has been successful in producing shrimp that are resistant to Taura, IMNV and whitespot.  There does seem to be a genetic component to EMS tolerance, which means breeding for EMS resistance would probably work.

 

 

Question and Answer Session

 

During the question and answer session, Dr. Lightner answered a number of questions:

 

Question: What is the toxin that is killing the shrimp?

 

Dr. Lightner: My laboratory is working on that.  We don’t really know what the toxin is at this time.  We’re still working on it.  We think it’s a protein, but that’s about all we know, so far.

 

Question: Does India have EMS?

 

Dr. Lightner: My lab has been working with agencies in India and trying to confirm whether or not India has EMS.  Unfortunately, most of the samples that were received to date were so contaminated with the whitespot virus that we have not been able to confirm that India has EMS.

 

Question: There’s been some talk about the possible presence of a bacterial phage or plasmid affecting the virulence of Vibrio parahaemolyticus.  Is the Vibrio strain alone enough to cause EMS, or is the presence of another pathogen, for example, a bacterial phage, also required?

 

Dr. Lightner: We were very excited when we found a bacteria phage associated with Vibrio parahaemolyticus.  We thought that it might be related to its virulence and EMS, however, it turned out not to be the case, so some of the early announcements that were made by my laboratory were incorrect.

 

We are also working with things called “contigs” because we don’t know if plasmids or other kinds of transmissible material are capable of causing EMS.  The PCR test for EMS is based on something that we call “contig-89”, and we think the commercial version of that test will be ready for use as early as January 2014.  We have detected EMS in hatchery postlarvae from some countries, but EMS does not seem to cause disease in hatcheries.

 

Question: Is one strain of Vibrio parahaemolyticus causing EMS, or are several strains causing it?

 

Dr. Lightner: We are finding some very minor differences among the strains of V. parahaemolyticus that are causing EMS.  The differences are very, very minor.  They all key out to V. parahaemolyticus, but they do differ slightly in their PCR reaction when we use different primers to different parts of the extrachromosomal genes.  There appears to be just one strain that is causing the global epidemic, but within that strain, there are some very minor differences.

 

 

Other Information from the Question and Answer Session

 

• It’s not likely that Vibrio parahaemolyticus is being transmitted in growout feeds.  The feeds are heat-treated and are not likely to carry the disease.

 

• Repeated trials show that frozen tissue failed to transmit the disease to healthy shrimp, but maybe even more relevant is some work done with oysters that were infected with Vibrio parahaemolyticus and V. vulnificus.  The combination of freezing and holding oysters in cold storage for three weeks reduced the bacteria to undetectable levels.  Imported frozen shrimp that has been in transit for three weeks should be safe.

 

• The primary reasons that Indonesia does not have EMS is that it restricts the import of live shrimp products.  The transport of live animals from one country to another is probably why EMS is now in five or six countries.

 

Sources: 1. Early Mortality Syndrome: Managing the Perfect Killer, a webinar organized by the Global Aquaculture Alliance and sponsored by SeafoodSource.com.  Ho Chi Minh City, Vietnam.  December 10–13, 2013.  2. Bob Rosenberry, Shrimp News International, December 11, 2013.

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