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Dan Fegan, Owner of The Shrimp List
Whitespot as a Model for EMS/AHPNS, and Some Notes about The List
Dan Fegan (email@example.com), owner and creator of The Shrimp List, rarely intervenes or post comments to The List. On June 25, 2013, however, he posted one of the longest entries that has ever appeared on The List.
Here it is in its entirety, exactly as he presented it:
Apologies for the long posting. I have been following a lot of the discussion off and on since the start but haven’t had the time or the inclination to post as I tend to be so far behind my input wouldn’t be relevant by the time I got it written down! However, I would like to offer some perspective from someone who has been dealing with shrimp disease at different levels for what seems (actually is…) a lifetime. I don’t say that to build myself up but in recognition that dealing with disease requires a wide range of skills and expertise that can only be obtained by working as a team with different people over a long period. I have benefited in my career from working with, and learning from, people who know little about shrimp but a lot about animal diseases, how they spread and how to develop biosecurity programmes. Having a 30-year (OK, 29-year) experience in this field, I have seen a lot of things that work, many that don’t and quite a few that used to work but don’t work anymore. It is in this vein that I will offer my tuppence (two pence or two cents for the linguistic nitpickers in the audience…!) worth.
First point is that we need to stop confusing EMS, a syndrome whose diagnosis is based purely on a (changing) pattern of early mortality, and AHPNS, a syndrome whose diagnosis is based on a histopathological demonstration of specific symptoms that appear to be associated with presence of a Vibrio (either V. harveyi or V. parahaemolyticus according to the pre-publication abstract in DAO) in affected shrimp.
From discussions with various people, the number of samples of EMS-affected shrimp that are submitted for confirmatory diagnosis is very small. This is perfectly understandable as farmers don’t generally waste time and money doing lab tests to confirm that their shrimp were sick or dead. However, it does not mean that every single case of EMS has the same cause or is due to AHPNS. It could be 1%, 100% or anything in between but without data, we are only guessing.
Although we can’t assess what proportion of EMS shrimp are sent for confirmation of AHPNS, we should be able to evaluate the proportion of shrimp samples submitted that prove to have AHPNS versus those with other conditions. I would assume that this proportion might vary over time but I have heard figures for AHPNS confirmation varying from around 10% of submitted samples up to around 50% from different places and times. It would be useful if we did have this kind of data available so we could make some estimate of the proportion of EMS samples that prove to be due to AHPNS. At the very least it would avoid panic and over-reaction, both of which have, in my opinion, played a significant part in the development of the current problem.
As I have said many times in various forums (fora?), we need to use epidemiological methods in tandem with histopathology and the use of diagnostic tools to be able to develop any understanding of how diseases spread, the various factors or “causes” involved and how these can be controlled. Disease in farming systems is a population issue and epidemiology is focused on understanding the patterns of disease within a population and between populations (ponds, farms, regions, countries). It was through applying epidemiological and pathology approaches (and a hefty dose of luck) that we were able to progress so quickly in the case of earlier diseases such as WSSV.
Establishing a clear, unequivocal diagnosis of a disease is fundamental to conducting epidemiological investigations. In the case of WSSV, we were lucky because the diagnostic tool was clear and cheap – the presence of white spots on the carapace. This meant that we could very quickly differentiate a “case” from a “non-case” and look for patterns in the “cases”. We were even more fortunate when that pattern suggested that outbreaks of WSSV appeared to be related to PL source (ponds supplied with PL from the same batch or hatchery had a higher likelihood of coming down with WSSV). Thus we were able to begin to adopt limited control measures even before we knew what was the causative organism. The virus was not identified until about 6 months later but we had by then begun to use various methods (only stocking with PL from previously unaffected hatcheries, using stress tests to kill weaker PL etc) to try to control the risk.
Once we knew it was a virus, that narrowed down the field and when the researchers at Mahidol University developed the first PCR primers for the virus, we could then go out and look for it in various places. The first and most obvious use was to screen PL before stocking. This threw up a particular challenge common to any diagnostic method – sample size. When taking a sample from a large population, the likelihood of finding at least one infected individual depends on the sample size and the prevalence of the infection in that population. However, many people were confused by the fact that PCR can detect very few copies of a virus into thinking that this meant it would pick up any virus present. Although PCR can be very sensitive in detecting copies of the virus (or its DNA), what is important in identifying the risk of introducing the disease is the number of infected individuals.
Fortunately, all of the statistical information has been worked out for livestock when testing animal herds for disease presence (not absence which is a whole different ball game). Based on these tables, a sample of 150 individuals is enough to detect a disease present at a prevalence of 2% with 95% confidence. However, it was more common (cheaper) to take a sample of 30 individuals which drops the prevalence detection to 10%. That carries the risk of missing batches with lower prevalences but some work conducted in Thailand showed that this was enough to significantly reduce the commercial risk of disease, especially in the dry season when there were fewer contributing factors. Unfortunately many labs reported a negative PCR test as “free from WSSV” rather than “if WSSV is present it is at a prevalence of less than 2%” and so there was, and remains, confusion over the value of PCR as a tool for testing PL. The corollary is that not every infection results in clinical disease so the mere fact of a positive PCR result does not guarantee that disease will happen but that the risk of disease is higher.
The researchers at Mahidol screened a wide variety of possible vectors and found that there were several crustaceans that could carry the virus without getting sick. For the record, the majority of these were crabs and other shrimp species, especially the small, clear mysid shrimps that often occurred in ponds in large numbers. Bearing in mind that PCR is a test for a fragment of DNA, we were also aware that some of these PCR positives could be non-viable and so challenge tests were carried out to confirm some of these. I remember in one farm we found oysters, mussels and fish testing positive following an outbreak but when we tested in more detail, the oysters and mussel gills were positive and the gills and stomach of the fish. Feeding these tissues to healthy shrimp did not result in any infection and so we concluded that the PCR positives were due to the presence of fragments of degraded DNA.
If I recall, we didn’t really consider copepods a major risk for WSSV in Asia as managing the larger vectors and PL reduced the risk sufficiently to make the business viable. The small size of farms and ponds made a policy of eliminating vectors using chemical treatments viable and there were generally fairly limited populations of other crustaceans such as crabs. I would not underestimate the success of this approach with monodon at the time when we had no SPF stocks and WSSV was a more common and significant risk. Of course, this approach carried its own risks but the measures taken by the Thai industry in particular at that time meant that Thailand suffered less, and for a shorter time, than many other countries. I would emphasise that the intent at the time was to kill macro-crustaceans, not plankton and there was a lot of nonsense talked at the time about using chemicals to “sterilize” the water and eliminate the bacteria and viruses. Anyone with a modicum of knowledge about water treatment would realize that the methods used were far from adequate to do this in an open pond system (and in fact many times the crustaceans would also survive). The later use of short half-life insecticides to target only the crustacean population was also much less hazardous from both an environmental and human health standpoint.
Vector control was much more problematic in Latin America due to the large size of the farms and ponds, and the sizeable populations of wild crustaceans in the farm environment. This meant that wild vectors were a much more significant risk than in Asia and so PL screening on its own was not enough to significantly reduce the risk. At that time, Granjas Marinas (I think) conducted some trials using different mesh nets to trap different sizes of plankton and then checked the plankton by PCR to see if it could be screened out. Again if I recall correctly, they found that the main PCR positive plankton was in the fraction larger than 250 microns and so recommended screening incoming water to remove these larger plankton (which contained crustacean larvae as well as copepods). This was a fairly cheap and practical but probably less effective solution than eliminating the vectors as in Asia.
One thing that I believe was very important in Thailand at that time was the formation of a White Spot Task Force bringing together representatives of the public (Department of Fisheries, BIOTEC, Universities) and private (Farmers groups, feed companies, processors) sectors to develop a common approach to the disease issue as it was evolving. As a group, we identified what was known and more importantly what was not known about the disease and discussed what needed to be done. Several research gaps were identified and research proposals developed to cover these. Sources of funding were obtained, again from public and private funds and a common statement developed describing the disease, what was known about it and importantly what measures might or would not work to solve it. The Department of Fisheries developed this into a brochure that was funded and distributed by the private sector. This approach meant that there was a common objective and that there was limited duplication of effort in working towards a solution for the problem. Everyone accepted that it was not “someone else’s responsibility” and that the common good of the industry was more important than the individual competitive position of the various private sector players.
So…why the history lesson on WSSV? Well, mainly because I can see history repeating itself with the current situation. During the “developmental stage” of getting to grips with WSSV, there was a lot of speculation, opinion and heated argument. Farmers getting upset because researchers took samples and didn’t find a cure straight away. People throwing around words and concepts with limited knowledge of what they actually mean (this is in the historical context before anyone takes umbrage – I have listened to a lot of people over the years discussing Vibrios and viruses when they don’t know if they grow on trees or arrive in a taxi!). Numerous people arriving out of the blue with “cures” and “solutions” that turn out to be nothing of the sort but cost a lot of time and expense to find out.
Unlike WSSV, we do not have a good “case definition” to separate EMS from AHPNS in the field. We know that “early mortality” can occur for many reasons so this is hardly a definition. That makes it impossible to determine the common factors and patterns in the epidemiology of the disease at this point in time. As a result, we are trying to work with reports based on a claim of EMS with the assumption that it is AHPNS – a dangerous assumption. Looking at the reports of when and where outbreaks took place in Asia, it was speculated at one point that it could be associated with factors moving from North (China) to south (Malaysia) over a roughly 2-year period. Inshore currents were one possibility and the fact that Thailand was affected later could be explained by the currents in the Gulf of Thailand. However, there could be other explanations/interpretations based on migratory patterns of birds, broodstock transfers and shipping of maturation feeds around the region. This was speculation that fit some of the information (not “facts”) at the time. How true any of it might be is still open.
Similarly, there has been a lot of speculation that broodstock and genetic selection could be contributing factors. I am not convinced by this as we have seen mortalities in shrimp from almost every broodstock supplier regardless of their genetic selection. I find it hard to believe that every single selection programme, regardless of their breeding goals or level of selection pressure would end up with the same problem.
Overuse of probiotics in hatcheries has also been proposed as a cause, or at least a contributing factor. Again, I am not sure that this is so clear as the same lot from a single hatchery can show early mortality in one farm but not in another, and PL from different hatcheries on the same farm can also show early mortality. It simply doesn’t seem to be so clear cut. However, I have some concerns over the way that probiotics are used in Asia as they are largely unregulated in their development and use. Poor quality control in manufacture and storage of probiotic bacteria could result in strain variation and mutations that could pose a risk and the incubation of probiotics in open tanks at the side of the pond carries a high-risk of promoting the growth of potentially harmful bacteria, especially when untreated pond water is used for the uncubation.
Fortunately, Don Lightner’s teams work has already moved us ahead considerably by identifying the possible pathogenic agent. I have never believed that it is a researchers job to solve our problems in the industry – that is for us to do. What they can do is to provide us with clues or tools that allow us to do a better job. If they, or anyone, can provide us with an unequivocal test for the causative agent of AHPNS, it will be a massive boost for two reasons. One, it will bring down the cost and complexity of diagnosing AHPNS so we can reduce the EMS/AHPNS confusion. Secondly, and equally importantly, it will allow us to look for the sources and possible vectors of the pathogen. Once we know that, then we can start to develop ways to manage them either through improved biosecurity or targeted interventions to reduce the risk of shrimp being infected or developing a disease once they are infected.
A couple of points on the list discussion before I finish. I have a feeling that the hyper-critical tone and occasional personal attacks will cause people to back off from contributing to the discussion for fear of ridicule which would be a great loss to the openness we have had on the list since it began. Everyone in their own way is trying to help and deserves the respect that engenders. I may not agree with much of what is said – to me the proof and the truth will be clear when we see a solution or an approach that is clearly shown to reduce the risk(s) associated with EMS and/or AHPNS. Until then, I can only read, listen and try to gain a better understanding of the various pieces of information that are available or presented and accept or refuse them as I see fit from my point of view. The large number of people on the list who are “lurking” (horrible term) or passively assessing the information provided by Daniel, Nelson and others in the light of their own experience should not be underestimated. It may well be that one of them will develop the solution, who knows?
Finally, I have noticed a big increase in the number of trade journalists joining the list or using information that appears on the list. I think that this should make us take a little more time to consider what we post as careless posting could unintentionally increase the confusion and panic. We owe it to them and our industry to focus as much as we can on hard facts since, as we all know, many people’s livelihoods depend on this business and careless rumours could cost jobs.
Oh, and PS – can people posting try to cut off the long strings of previous comments when sending a mail. Maybe keep just the immediate post to which you are responding. This will make the list easier to read. Thanks!
Sources: The Shrimp List (a mailing list for shrimp farmers). Subject: Thoughts on Shrimp Disease Investigations and the EMS Situation. June 25, 2013. 2. Bob Rosenberry, Shrimp News International, June 25, 2013.
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