OREGON STATE UNIVERSITY

Calicivirus Information

01/05/1996

CSIRO Australian Animal Health Laboratory Statement put out by Dr. Keith Murray, Head, AAHL, dated 13 December 1995. As a News Release --This news release can be found at the following address:

gopher://commsum.its.CSIRO.au:70/100/commnews/rabbits/rb9516.txt

Response by Dr. Alvin W. Smith, Head, Laboratory for Calicivirus Studies, Oregon State University, dated 20 December 1995.


CSIRO statement #1. CSIRO virologists today corrected misinformation from an overseas veterinarian about rabbit calicivirus posing a threat to humans and other species.

Response: There was no misinformation to correct in the news release that came from Dr. Alvin Smith at Oregon State University's Laboratory for Calicivirus Studies at the College of Veterinary Medicine. Perhaps there are some who misunderstood or misquoted.

CSIRO statement # 2. The Australian scientists said that Alvin Smith of Oregon State University had confused different groups of the Calicivirus family. Some members of one group of caliciviruses can infect several species, though this is a natural ability of that virus and does not come about through mutation, as claimed by Alvin Smith.

Response: There was no confusion. The clear and simple statement is "That Caliciviruses as a group can and do cross species barriers." There are many published examples proven by research on San Miguel/Vesicular Exanthema caliciviruses, Feline caliciviruses, and hepatitis E caliciviruses . Cross species transmission is suspect but not proven for small round structured and perhaps Sapporo calicivirus. If rabbit calicivirus does not cross species lines it could be the exception. That hope is poorly founded because of the likelihood of rabbit calicivirus having emerged in rabbits from some other unknown reservoir species, thus, conventional wisdom would suggest that 10 years ago the virus had already crossed species boundaries into rabbits. Rabbits would be a new host species and that is why it is so deadly to rabbits. CSIRO statement #3. According to Dr. Keith Murray, Head of AAHL, "Rabbit calicivirus is found in more than forty countries around the world. There is no evidence that rabbit calicivirus has infeccted any animal other than European rabbits nor is ther any evidence that it affects humans."

Response: Statements of "no evidence was found" regarding the occurance of a specific event is the standard jargon sometime used for the purpose of implying that an event will not or cannot occur when proof does not exist. What tests were used? How sensitive and how specific are the tests? What populations of animals were sampled? How adequate was the sample size? How massive was the exposure in terms of virus numbers? Where in the life cycle of each species tested were they examined, i.e., the very young, the very old, the disease stressed, the environmentally stressed, the pregnant, etc.? The same questions need to be answered in terms of human exposure to rabbit calicivirus before statements of "no evidence" carry any meaning.

CSIRO statement #4. "In a three year study at AAHI, 28 different species of animals were inoculated with large doses of the virus. None of those animals became infected with the virus or became sick from it."

Response: The studies on 28 species is an excellent start but should be interpreted as showing that the research is as yet inadequate to fully define the host/parasite relationships of rabbit calicivirus in nature. Because the rabbit is such and unusually susceptible host for this virus, the 1000 rabbit lethal dose50 (1000 RLD50) of virus given to the other 28 test species seems unusually small. Doses in the 1,000,000 range might be better indications. In fact the 1000 RLD50 is so few virus particles that the test animals were said to not even develop antibody. Why were these challenge doses so purposefully small? If the 28 species tested truely cannot become infected then why not give 1 million or 10 million or more doses. These are much more in the dose range that will occur under conditions of natural exposure with millions of rabbits dying. Please do not forget the probability of this being a virus which has spilled out of some other reservoir species into rabbits.

CSIRO statement #5. Alvin Smith's press release contained several factual errors and ignores major developments over the past five years in our understanding of rabbit calicivirus.

Response: If one is stating that errors were reported these should be carefully spelled out rather than alluded to. Otherwise, honest disagreements or differing interpretations never get carefully argued on a basis of available evidence.

CSIRO statement #6. "It is important to understand that viruses are not classified by the symptoms they cause,"said Dr. Harvey Westbury, a veterinary virologist at CISRO Australian Animal Health Laboratory (AAHL).

Response: Dr. Westbury's reasons for studying viruses is interesting. I do not believe viruses are simply studied to be filed into some catalogue on the basis of sequence data alone, but instead the viruses we both are interested in are studied because of the symptoms they cause, the disease they produce and the species they infect. Sequence data does not in its present infancy define these factors and neither is sequence data reliably predictive of a virus's future capabilities. The sequences are constantly shifting, especially with small RNA viruses and this will continue to confound a scientific community that does not use this information in its proper context.

CSIRO statement #7. "Viruses are grouped together and named on characteristics such as their genetic makeup and shape - not on the infections they cause. You cannot generalize about the different infections caused by caliciviruses, as Alvin Smith has done," explained Dr. Westbury.

CSIRO statement #8. Over the years, to facilitate information exchange, scientists have developed protocols to classify viruses as they have for animals, plants and bacteria. Thousands of viruses that infect vertebrate animals are placed in 18 "families"; Caliciviridae is one of those families.

Response: Genetic make-up is one way of grouping viruses, their morphology, and/or other functional and physicochemical characteristics are also ways of grouping viruses. In fact all the viruses grouped into the family Caliciviridae were first placed in this taxon based on morphology and physicochemical characteristics. Sequence data is useful in examining specific differences within this group but was not used to establish the Caliciviridae group. Generalizing about and extrapolating known information within a group of viruses is an extremely useful tool used by all established and reputable researchers when they are asked to predict or guess about the characteristics of a less well studied virus within that group. Often times these extrapolations are the basis for new hypotheses to be tested. In fact, Australian researchers were doing just that when they began testing 28 non rabbit species for their susceptibility to rabbit calicivirus. Certainly they did not look at the published genomic sequence of rabbit calicivirus and determine beyond a shadow of a doubt that this virus infects only rabbits.

CSIRO statement #9. The Caliciviridae can be subdivided into five groups of viruses based on their genetic makeup and strategy for replication. One of those groups contains San Miguel Sealion calicivirus that can infect pigs, seals, and some fish. This virus is unusual because it can infect more than one species, a peculiarity that was quickly recognized. It is a virus that CSIRO scientists have worked with at AAHL. However this virus has not mutated to infect other animals. It has always been known to have a wide host range.

Response: The information repeated here on the Caliciviridae being grouped into 5 groups was reported to Dr. Westbury on 8 November 1995 by Dr. David Matson and was primarily work which came out of Dr. Matson's laboratory. I saw no credits given for this information to one of my collaborators, Dr. David Matson. One should also note that I am co-author of the cited research which was presented this past October in San Francisco at a scientific symposium with and international audience. The statements regarding the San Miguel Sealion virus are incorrect in several ways. Although it is a group whose host range has been studied more intensively than others, members of other calicivirus groups are not host specific. Furthermore, viruses of the San Miguel grouping were said to naturally infect only swine from 1932 to 1972 (a 40 year period) and were said not to infect humans. In 1985 the first example of human caliciviral disease from members of this group was reported at the national ASM meeting and published as a proceedings abstract. To say that host non-specificity was "a peculiarity that was quickly recognized" is misleading in two ways. It is not a peculiarity, rather it appears to be a rule that becomes established as one properly examines the Caliciviridae and point two, decades of error repeated as scientific dogma by U.S. officials declaring host specificity for this group of caliciviruses was displaced grudgingly and only under pressure of overwhelming evidence to the contrary. In fact in this country there is still strong resistance to lumping San Miguel viruses and Vesicular exanthema viruses together, yet CSIRO scientists had no trouble doing so. CSIRO scientists claim expertise with the San Miguel; viruses, yet they don't say which one (there are 17 diverse serotype with the specific San Miguel Sealion designation) and all of them were first isolated and characterized at Dr. Alvin Smith's Laboratory for Calicivirus Studies. Copies of these viruses have been provided to Pirbright in the U.K., Plum Island and the NADL in the U.S. and because of the international restrictions, Dr. Smith knows of no other movement of these agents, nor has he authorized their release without proper agreements being put into place. One wonders why CSIRO scientists would say the San Miguel viruses have not mutated. There will be a viable mutant occur in a ratio of about 1 per 1,000,000 small RNA viruses reproduced. The statement that it has always been known to have a wide host range is untrue. Historical scientific records show quite the opposite. Host diversity was not known until proven by the Laboratory for Calicivirus Studies working intensively for two decades and this story is still evolving. Such efforts yet remain for the rabbit calicivirus. A start would be to identify its natural reservoir species in nature as has been done for other caliciviruses . As a footnote on the groupings of the Caliciviridae , Sapporo calicivirus which causes gastroenteritis in humans is more closely related genetically to the San Miguel viruses than it is to the human Norwalk calicivirus which also causes gastroenteritis. The rabbit calicivirus is genetically more closely related to feline calicivirus (FCV) than FCV is to the San Miguel viruses, yet the FCV is not host specific. An even more distantly related calicivirus, the Human Hepatitis E virus will infect both monkeys and swine, thus it is not host specific. In short, as one studies the variants within each group the group sizes tend to increase and their characteristics overlap with each other.

CSIRO statement #10."By comparison, rabbit calicivirus has a very narrow host range, as confirmed by exhaustive tests at AAHL," Dr. Westbury said.

Response: It may be true that rabbit calicivirus has a narrow host range compared to other selected virus types but that provides no assurance that it will not and can not infect other species including humans.

CSIRO statement #11. "Moreover, rabbit calicivirus is found in over forty countries and has spread from Asia to Europe and northern Africa. In over ten years, it has killed millions of farmed and wild European rabbits and there has been close contact between people and diseased rabbits.

CSIRO statement #12. No transmission of the virus to humans, or illness related to the virus has been reported in the scientific or medical literature. "Blood tests have not revealed the presence of antibodies of the virus in humans who have had contact with the virus or with diseased rabbits," he emphasized.

Response: The disclaimer for possible human infection has numerous red flags. How can one assume that if disease occurs it will be recognized and reported in the scientific and medical literature. Every community has a preponderance of disease cases, especially transient diseases that go without definitive laboratory diagnosis and this also goes for the human population working with rabbits that are potentially calicivirus infected. Consider the respiratory diseases, diarrheas, spontaneous abortions, hepatitis, viral myocarditis, herpes-virus like mouth lesions and hand-foot and mouth lesions of children etc. that are clinically diagnosed each year without a single laboratory test to prove definitive etiology. Caliciviruses of various pathotype have been implicated in all of such disease conditions in various animals and some of them in humans, yet caliciviruses as a cause of such illnesses are rarely suspected and less often looked for. A so-called "blood test" that did not reveal human antibodies against the rabbit calicivirus is no guarantee that there has been no infection. How many people were looked at, how good was the test. One might suspect that the tests are not adequate to pick up all antibodies if they occur. Otherwise, there might be a bit of antibody in people who have been exposed to massive viral doses even if infection did not actually occur. Furthermore, there are documented cases of animals having been calicivirus infected that shed the virus for weeks but did not develop any detectable antibody using both type specific neutralizing tests and group specific immunoblot techniques or Enzyme Linked Immunoadsorption assays. These are highly sensitive tests for antibodies.

CSIRO statement #13. The genetic material of all living things can change from, or mutate, from generation to generation. Viruses are no different but this does not mean that they'll jump species. A good example is the myxoma virus which has mutated, killing fewer rabbits, but still only infects rabbits.

Response: With myxomavirus, the Australian scientists are experts but the lesson from myxoma (a DNA virus much less prone to mutation than the RNA calicivirus) is not that it has apparently remained confined to rabbits of genus Oryctolagus. Historically it has been proven to have not only jumped species but also moved from one genus to another. Its origins were said to be the New World (Uruguay and Brazil) in the genus Sylvilagus i.e., wild rabbits. When it jumped genera to the European species it became deadly. Genetically adapting the virus to new growth or host species conditions has, for years, been the basis for vaccine production, not only for Myxoma vaccines but for many other viruses as well. The argument that viruses do mutate from generation to generation but that this will somehow preclude them from adapting to infect new host species is lost on me. For nearly all viruses this genetic adaptation through mutation has been shown many times both inside the laboratory and under conditions of natural exposure in nature.

CSIRO statement #14. Dr. Westbury also rebutted comments attributed to him in the Sydney Morning Herald. "Rabbit calicivirus has not changed at all since imported to Australia in 1991. The genetic make up of rabbit calicivirus strains in Europe can differ by 1 or 2%. However, this difference does not mean that the biological activity of the virus has changed.

Response: It is encouraging to see that once again "biological activity of the virus" becomes important after the rather pointed remarks in CSIRO's paragraphs 6 through 9 that would lead one to believe that "biological activity' was an archaic way of looking at viruses. It is also important to examine the conceptual flaws underlying the CSIRO pronouncement that the genetic make-up of the virus has changed only (1-2%) from the European ancestral virus. If one looks only at highly susceptible Oryctolagus rabbits where virus copy numbers may reach tens of billion and then at the rabbit livers as an organ of choice to harvest large numbers of virus for genomic sequence comparison, is it surprising that the virus populations found there have shown only a 1-2% change in base sequence? After all these virus populations first had to retain the genetic windows which allow them to replicate in a single species (the rabbit) and then in a specific organ (the liver). These are powerful selective forces which help insure that of the viruses tested the predominating number will retain like genomic characteristics. Finally, one demonstrates likeness to some predetermined sequence by using sequence homologies and Polymerase chain amplifications which, in themselves, are powerful selectors for identifying only the predominating viral species in this population of potentially thousands of genetic variants. It is this population of variants that the debate is over. It is this population of mutant viruses that will ultimately shift the rabbit calicivirus to a new species, if such occurs, and it is this population that in the past has served the family Caliciviridae so well in terms of moving the virus between species across great phylogenetic distances.