December 1988 issue of DVM magazine featured this Vaccine Roundtable discussion.

Safety, efficacy heart of vaccine use; experts discuss pros, cons

Modified live virus veterinary vaccines developed during the past two decades have been very effective in reducing the incidence of viral diseases in many species of domestic animals. Now, an increasing number of new, inactivated virus vaccines are being produced and more will be available in the future.
     Discussing the advantages and disadvantages of both types of vaccines in our exclusive roundtable, co-sponsored by Diamond Scientific, and hold at the Salk Institute in San Diego, are; Dr. Ted Rude, founder of Veterinary Consulting Services in Hudson, Wis., a consulting firm for the veterinary biologic and pharmaceutical industry in pathology, toxicology, forensic veterinary medicine, biological development and vaccination programs in domestic animals and poultry. He is a graduate of the University of Pennsylvania and a diplomate, American College of Veterinary Pathologists; 
Dr. Ian Tizard is professor and head of the Department of Veterinary Microbiology and Parasitology at Texas A&M University. He received his DVM degree from the University of Edinburgh and his Ph.D from Cambridge University; 
Dr. Gunter Siegl is head of the Department of Virology at the University of Bern, Switzerland, where he oversees experimental virology, diagnostic virology as well as AIDS research. He received his Ph.D degree from the University of Munich; 
Dr. Ran Schultz is professor and chairman of the Department of Pathobiological Sciences at the University of Wisconsin Madison School of Veterinary Medicine and holds joint appointments in the Medical School and the Department of Veterinary Science, College of Agricultural and Life Sciences. His undergraduate and graduate training in bacteriology biochemistry, immunology and pathology were done at Pennsylvania State University; 
Dr. Jonas Salk is founding director and distinguished professor in international health sciences at the Salk Institute for Biological Studies in San Diego Calif. He received his MD degree from New York University College of Medicine in 1939. Along with his lifelong biological research especially polio he has held professor and fellow positions at the University of California-San Diego, University of Pittsburgh School of Medicine, University of Michigan School of Public Health, Department of Epidemiology. His brother is a veterinarian; 
Dr. Larry Swango is associate professor of Virology in the Department of Pathobiology, College of Veterinary Medicine, Auburn University. His primary area of research has been with canine virus diseases and vaccines. He received his DVM degree from Oklahoma State University. 

Rude: What is the rationale for the use of modified I live virus vaccines in domestic animals and why have they been so successful during the past two decades in reducing the incidence of various diseases in domestic animals?

Swango: Certainly, modified live virus vaccines have made a very valuable contribution in the control of infectious diseases in domestic animals. One of their strong points has been the simulation of a natural infection and immunity in response to replication of the virus within a host.
     We can go back perhaps more than two decades with the introduction of rabies vaccines for a little historical background. At that time, rabies was a disease of great significance in the United States and other developing countries. We needed something to control it and the live virus vaccines that were initially used were very effective in controlling the disease. As we began to control the disease, we then became aware of vaccine-induced disease in which we could then switch our emphasis to increasing safety to further attenuation. One of the strong points of the modified live virus vaccine is the fact that in the animal, it stimulates immunity based on replication of a virus which stimulates an infection with a non-virulent strain of virus.

Siegl: I would just add that one of the advantages of modified live virus vaccines is the fact you usually need to vaccinate only once with the virus. In general, you also need a relatively small dose of vaccine. The modified live virus contained in it replicates in the animal and induces an immune response comparable to the one obtained after exposure to the wild-type virus.

Salk: Well, there are advantages and disadvantages as already mentioned, and I suppose it's appropriate to consider modified live virus vaccines in the evolutionary development of the ultimate which would be not merely to simulate nature, but to improve upon nature. So perhaps we should look upon this discussion, and this point, about advantages and disadvantages relative to where we stand in our science and technology.

Schultz: I believe the history on modified live viral vaccines, especially in veterinary medicine, stands as an example of how effective and how safe certain modified live products really are. Dr. Swango mentioned the example of rabies. I think the examples of feline and canine distemper and canine adenovirus are equally good examples of effective modified live viral vaccines in small animal medicine. We also have examples in large animal medicine In which we can show both safety and efficacy for these products because they do produce protective immune responses, and that is what we're attempting to get from our modified live products.
Protective immunity is a very different concept from simply inducing a serologic response. As the discussion continues I think we can get into the details of why and how a modified live product can provide a protective response, humoral and cellular, local and systemic, whereas we may not be able to get a protective response from certain inactivated or sub-unit or synthetic products.

Tizard: But having said that, it appears, as Dr. Sulk said, that the production of vaccines is an evolutionary process and in the past, it has proven easier to make modified live vaccines than inactivated ones. They overcome a lot of shortcomings in terms of adjuvants, in terms of ways of preparing antigen, and most economically, in terms of antigen dosage. You can produce them very cheaply. One could argue, however, that they are not as devoid of side effects as we would like to think. When, we started to control diseases like canine distemper, those side effects were minor concerns. While we certainly have to give credit to modified live veterinary vaccines for having had a tremendous impact in controlling disease in this country, they do represent a fairly basic technology that can be improved upon. Many of the disadvantages of inactivated products have now been overcome.

Rude: Dr. Swango, you mentioned something about vaccine-induced disease and there are some reports about disease in vaccinated animals. Since you are primarily in canine vaccine research, what modified live virus vaccines is the canine area are most likely to do this and how serious is the problem?

Swango: Probably some of our least attenuated canine distemper vaccines, when used in a very young animal that does not have a fully developed immune system, are where some of our greatest risks lie. We have some very good distemper vaccines. Modified live distemper vaccines, when used in the animal at an age for which they were developed and intended to be used, are relatively safe. But when used in a younger animal in which the immune system has not developed to the point of being able to respond and contain the modified live virus antigen, then we do get vaccine induced disease.

Rude: In your research experience, Dr. Swango, what particular vaccines can you cite as causing vaccine-induced diseases?

Swango: Canine distemper in, again, very young puppies. Certain strains of modified live canine distemper virus can cause disease in very young puppies If we broaden our perspective of disease to indicate and to include any pathological condition, not necessarily the typical disease, then we would have to refer back to a very low rate of pathologic change associated with canine adenovirus Type I as a modified live virus vaccine.

Tizard: And you talking about " blue eyes?"

Swango: "Blue eye" and perhaps renal lesions in some instances. Those would be adverse effects, a disadvantage of the vaccine, but not causing the typical disease. There has been a great deal of interest in the last eight to 10 years about modified live canine parvo vaccines causing disease in dogs. I have yet to see a modified live canine Parvo virus cause disease in a dog.

Rude: When you talk about younger dogs, Dr. Swango, can you be more specific on week of age in these dogs?

Swango: Any animal under 4 weeks of age, but extending even on to 6 to 8-weeks of age in some instances. Now this may be a function of genetics of that breed or that bloodline, but we have some indication that even in 8 to 8-week-old puppies the distemper vaccine may occasionally induce disease. There may be some con-current factors, other infections or something that contributes to that, but especially in puppies under 4 weeks of age.

Rude: There have been reports in the literature about chronic canine distemper and old dog encephalitis. Chronic canine distemper is usually seen in young dog 2 to 3 years old, whereas old dog encephalitis may be seen in dogs 5 years of age or older. Many of these dogs have been vaccinated as puppies with modified live virus canine distemper vaccines. Do you believe there may be a connection between old dog encephalitis or chronic distemper and the use of these vaccines?

Swango: The difficulty of reproducing those chronic infections experimentally has complicated our interpretation and assessment of the role the distemper vaccine may be playing there I do know that this distemper virus can be isolated from lymphoid tissue of vaccinated puppies, in some instances a much as six weeks after vaccination. It's difficult sometimes to prove that it's a distemper vaccine strain of virus rather than a virulent field strain.
     In some recent isolations that we have made, in which it was very easy to isolate the virus in cell cultures that normally do not support replication of field strains, we're inclined to think that certainly we can get some persistence of the view in the vaccinated animal. Now whether this contributes to the chronic distemper and the question you're raising, I'm not sure.

Tizard: In making a modified live vaccine you make it for an animal that you assume is immunologically normal. Essentially you're doing a balancing act; taking an agent and attenuating it so that in the immunologically competent host, (the normal animal), it will not cause disease. The trouble is, you cannot assume that every animal you vaccinate is functioning immunologically normal. There will be a proportion of any population that is not immunologically 100 percent. This can immediately tilt an animal towards disease susceptibility.
     In addition, a vaccine may not cause freak disease itself. It may cause mild immunosuppression.

Salk: Discussion, even at this early stage, brings me back to the days when I heard, in 1936, that it was not possible to immunize against a virus disease with a killed virus vaccine We've come a long way and now we're on the threshold of being able to eliminate vaccine risks. There is a tendency to want to eliminate all vaccine associated disease both in the human realm and in the veterinary, realm. I think we're appropriately reevaluating and reexamining the question as to whether any risk is necessary. Can we not do so with killed virus vaccine* and even improve upon nature by combining many viruses into a single vaccine using the adjuvants that make this possible? There is a problem of vaccine-associated disease that cannot be avoided.

Rude: Dr. Swango, world you like to comment?

Swango: Dr. Tizard made a comment that I think may require some response at this point. It is true, we can't be assured that 100 percent of the animals will respond to vaccinations. I'd like to extend his comment to an area that I feel, in veterinary medicine, we have perhaps been remiss in leading our profession as well as the public to believe that if we use Vaccines, we can guarantees 100 percent success in immunization. Then when then is a problem that develops in a vaccinated animal, people an greatly concerns.
     So I think some of the going Dr. Tizard mentioned are certainly paramount to this, but I feel that we somehow need to redirect an emphasis that just because we vaccinate, we cannot assure immunity. There is a difference between vaccination and immunization.

Rude: Does shedding of modified live virus vaccine viruses from vaccinated animals have the potential to cause disease in non-vaccinated contact animals of the same species and/or different species?

Salk: Well, it's self-evident that this is what happens with polio. I have not seen evidence that it is of significant advantage. There's a disadvantage in the U.S. because of vaccine associated disease. In the developing countries live virus vaccines often fail because they don't always protect those who are directly vaccinated due to inhibitors in the intestinal tract. In the U.S., vaccine virus is the principal cause of continuing polio.

Schultz: Dr. Salk, is this the vaccine virus itself that, when spread, is causing problems? If that's the case, then we have the spread of a vaccine virus that really is not attenuated.
     In veterinary medicine, we have some examples of truly attenuated vaccine viruses that spread, but the spread of the vaccine viruses leads to herd immunity which Is a very positive effect within the animal population. We, as well as others, have demonstrated that when spread of the vaccine occurs, there is no reversion to virulence. In fact, some of these viruses remain truly attenuated throughout the animals lifetime because they are viruses that become latent and can be reactivated and can be shown to be truly attenuated five to 10 years after an animal was vaccinated.

Salk: What you just described is not my understanding of herd immunity. That's direct Immunization and that ought to be clarified.

Schultz: In this case, it is herd immunity. It's within a herd or cattle. So in the context of herd immunity in veterinary medicine, it's real. With regard to 'herd immunity' in human medicine, it may not apply.

Siegl: I just want to go back to your final question and to what Dr. Salk said. It has been shown that shedding of virus, especially in the polio virus field, can mean selection of new variant viruses which spread and cause disease. I would say the basic principle, as soon as a live virus is being applied to an individual of a certain species, in that out of the mass of various mutant, as I tried to point out before, you are going to have an opportunity to select for one viral mutant which is able to replicate even in the presence of some sort of immunity. That new selected view can spread within the same species or in a different species.

Tizard: I can think of one veterinary example and that's laryngotracheitis in poultry. Vaccines used in the Northeast appear to be pretty good vaccines and are essentially avlrulent. If vaccinated carrier birds are moved, say to the Southwest, these vaccinated birds are a source of infection for the naive flocks sail cause clinical Iaryngotracheitis.

Swango: Your original question had to do with, as I recall, modified live viruses shedding from vaccinated animals and affecting not only animals within the species for which it was intended, but other species. I think one example perhaps relates to canine distemper virus, again, and I'll just touch an this briefly.
     I think Dr. Max Appel at Cornell, has shown, a number of years ago that within a population of virus particles of a modified live distemper vaccine there was a certain small percentage of that population that was still virulent. So in an animal in which there could be selection for enhancement of replication of that small population, then I think we could have reversion of virulence. Maybe in an immunosuppressed or immunodeficient young dog we might have expression of that type of reversion of virulence with transmission to other animals and perhaps not causing a typical, classical, acute distemper, but maybe the chronic infection within the species of what I refer to as strains of low virulence.
     We have evidence for this type of phenomena when distemper vaccine was used in a different species. A single passage of a modified live virus canine distemper vaccine through a gray fox, in which it was virulent resulted is reversion of virulence for the dog. The virus isolated back from The virus isolated back from the gray fox was no longer attenuated for the dog. So that touches, at least, on the involvement of other species and this problem of shedding and reversion of virulence.

Rude: What are the chances of mutant viruses developing in animals vaccinated with modified live virus vaccines; in contact animals of the same species or other species?

Siegl: I would say the chances for mutations are quite well understood at present. You have a chance of mutation at about 10-3 to 10-4 for RNA viruses. Rates an about 100 to 1,000 times lower for DNA viruses, but considering the fact that effective replication of a virus produces hundreds of thousands or millions of viruses in the cells of an organism there is always a great number of mutants around.
     Of course, most of these mutant are not able to survive in that surrounding because they are defective mutations. Nevertheless, we have to consider the fact that this is a Huge pool of genetic material from which new variants can be selected, and as has been pointed out before, it is clear. Especially in the polio virus field, that one passage through the human gut can result in selection of quite a number of new variants.
     Another situation favoring the appearance of mutations not directly related to vaccination, is persistent infection. We have been talking about persistent infection in context with canine distemper virus. It might be the result of vaccination that as soon as persistent infection is established, the incidence of mutations increases considerably. In addition, we have the possibility of recombination between wild-type virus and attenuated viruses. This might at least to my understanding, produce one of the main problems once we get to the point that we can engineer so-called safe attenuated vaccine. We will hardly be able to exclude the possibility that viruses from such products will recombine with wild-type virus under field conditions. My concern at the moment is the theoretical considerations Nevertheless, there are practical examples available to stress the reality of this problem.

Salk: The object of this exercise, it seems to me, is how can we produce biological products with the maximum amount of safety and efficacy? As a theoretician, one can imagine how that can be done. In the meantime, one has to be altogether pragmatic when the disease is of far greater risk than the methods that an employed in their control.

Rude: We have talked about mutations of viruses in animals. What are the chances of mutant viruses developing in cell cultures associated with the production of modified live virus vaccine? The second part of this question is what are the chances of harmful, contaminating viruses or adventitions viruses being present in modified live virus vaccines?

Schultz: I think that we have plenty of examples where we can demonstrate that, in cell culture, we have failed to attenuate. I think in part, we're getting back to an earlier, more primitive age. Some veterinary vaccine were not made from cloned viruses. Instead, an unknown population of virulent viruses were passed in cell culture and we developed something that was called a modified live attenuated vaccine, but sometime it was a modified live virulent vaccine. I think that if we would come up into the 1980s with the current technology, we would relook at some of the things we've done in the '50's, '60's and '70's.
     So yes, there is that opportunity with modified live product, to have mutations or variations that can act very differently among species. That's the reason we should not use any product, biologic or drug, in a species in which it has not been tested; especially wildlife species.
With regards to contaminating agents within cell cultures the most common contaminating agent is one of the viruses, of greatest problem to the cattle industry a togavirus, called bovine virus diarrhea virus (BVDV). It is very difficult to maintain any cell lines free of that virus and, furthermore. It's even more difficult to detect the contaminating non-cytopathic BVD when it's present.
     A second common example of an adventitious agent is another group of viruses of concern to the veterinary community the Parvoviruses, particularly the porcine parvovirus which gets into tissue culture from contaminated trypsin. So yes, we must be ever aware of vaccine contaminants. We must also be aware of mycoplasma contamination and every other contaminant that can end up in cell culture that have product like fetal bovine serum and trypsin used on them.

Siegl: We did a lot of work isolating porcine parvovirus out of cell cultures. We analyzed 43 cell cultures of different origin. From 38 of these cultures, we could isolate parvovirus. About 32 isolates were porcine parvovirus and some of the other were of rodent origin. For two viruses we don't yet know the natural host.

Tizard: I think it's difficult to overestimate the importance of these contaminants. There an a number of examples of diseases being spread because of this.
For instance, reticuloendotheliosis was introduced into Australia with Marek's disease vaccines for poultry. This problem, is intrinsic to modified live products and can be readily overcome through the use of inactivating agents.

Swango: Yes, I agree, and certainly when we're at in the 1980s approaching 1990, we have technology that will greatly improve that. But I think Dr. Salk could take us back to the early days of that vaccine and inactivating agents did not inactivate a contaminant. Unless we know the contaminating agent is there to conduct surveillance on it, we cannot be assured that inactivation will inactivate all contaminating agents.

Salk: Apropos of that, you're referring to the SV40 virus which was a contaminant in monkey kidney cell cultures. The last thing in the world that one would want to do now is to make vaccines out of the tissues of monkeys that come from the jungle. That was a learning experience you might say. At this point, we are much more sophisticated and can use continuously propagating cell lines, such as the Vero cell. The SV40 story alerts us how to avoid these things in the future.

Schultz: One more comment is, if we don't know what the contaminating agent is or if it is introduced after inactivation has occurred, we still have the potential for the introduction of a live agent, even in an inactivated product. So we've come a long way, but we've got a tong way to go.

Tizard: Except it's much more common, of course, using an inactivated product to add preservatives which will minimize that.

Schultz: Nucleic acids are quite resistant to many of the inactivating agents or the preservative and the nucleic acid, the genetic material of the virus is often all that is necessary under certain sets of circumstances, to get infection in the vaccinate.

Salk: This is free advice, You might just use some gamma rediation on the final product.

Rude: What do you think the average veterinarian should know or be told about the advantages and disadvantages of modified live virus vaccines?

Salk: There's one unique situation. That has to do with polio, because there's a choice. The argument is offered that the live virus vaccine is more advantageous from the public point of view and the killed virus vaccine from the individual point of view. I cannot make that kind of distinction because I know that with a killed virus vaccine one can control the disease effectively from both the public and individual point of view. If the physician, in this instance, were fully informed, he could make a choice because with the killed virus vaccine we can eliminate the disease altogether, both vaccine-induced and naturally-occurring disease. This unique situation is possible since we can provide a clear-cut choice.

Swango: We have a difference in animal medicine compared to human medicine.
Perhaps this could be illustrated best by some of our diagnostic procedures.
     In dealing with poultry flocks we've got sick birds. Instead of immediately trying to treat all the birds sometimes it is prudent to go ahead and euthanize some of those living birds so that we can get a more accurate, specific pathologic diagnosis. The same is true of swine. So we do have a difference here. 
     With respect to what do we tell the veterinarian, let me suggest that we need to do everything we can to minimize risk to make products as safe and as effective as possible. However, I think the veterinarian or the animal owner should be aware that anytime a product is administered there is a risk be it biological or pharmaceutical. 

Schultz: I think that all of us here would probably go on record as saying that are would find useful and essential, both modified live and inactivated vaccines in veterinary medicine. At least I would hope that we would go on record as saying that because there are certain disease conditions for which there an no satisfactory inactivated vaccines currently available.
     I think that's an important point that the veterinary practitioner needs to understand. Both modified live and inactivated vaccines have a place in veterinary practice. To get back to the issue Dr. Salk brought up, if we have a choice and we truly have the same efficacy in both products, then I think that everyone at this table would opt for the inactivated vaccine because of the potential safety aspect: I don't think there is anyone here who would disagree with that.

Rude: We're going to be talking about safety and I think it's been brought out that any biologic can create some problems in vaccinated animals.

Schultz: I brought that out earlier. It you're going to put an exogenous agent, whether it be biologic or drug, into an animal, including humans, you've got to realize the potential danger, whether it's an aspirin you're going to take 10 minutes from now because you have a headache or whether it's an inactivated vaccine.

Rude: We have been discussing modified live virus vaccines and also talked briefly about inactivated virus vaccines. Now I would like to discuss inactivated virus vaccines in more depth. What do you believe is the primary advantage associated with the use of inactivated virus vaccines?

Swango: From my perspective the primary advantage would be and this is partly theoretical is safety; to minimize the risk of a live agent that could replicate and undergo these theoretical changes we have talked about. That would be the primary advantage.

Schultz: We are talking in terms of inactivated viral vaccines and it might be important to point out again to the practitioner that we're probably talking about whole unit and sub-unit virus vaccines. We probably should also be talking about synthetic vaccines, so I think that the people know that inactivated only refers to whole virus and in veterinary medicine inactivated or killed viral vaccines are still just whole inactivated virus with which we've had some poor experiences. We also need to bring this to the level of current development. Are we, in this context, talking about sub-unit or synthetic or just whole inactivated product?

Salk: Let's use the concept of non-infectious.

Rude: That's a good term. Instead of saying 'inactivated,' we will use the term 'non-infectious' vaccines

Salk: Well, one has already been mentioned, its non-infectious character, reducing those risks that are associated with an infectious agent. Another advantage I see is the capacity to include immunogens for a large number of diseases at the same time, so as to reduce the number of vaccine administrations. This will be of increasing advantage in human medicine, and in veterinary medicine requiring the use of adjuvants as potentiating agents for that purpose. It allows the use of smaller quantities of the immunogen, producing more powerful and longer lasting effects.

Siegl: When we are using non-infectious vaccines, we can have these vaccines highly controlled with respect to purity. We can purify them and take out as far as possible all material then might generate adverse effects.

Tizard: The only other advantage could think of In addition to the primary one which is safety, is that commonly they're rather easier to store.

Siegl: There's one additional advantage. Application of such a vaccine by chance to another species is not very likely to do any harm.

Schultz: That raises an important point. We have had the idea for years that vaccines, if they don't do any good, won't cause harm. I think that's another concept the veterinarian has to get away from because whether it be modified live or non-infectious, there is the potential to cause harm! We've seen a good example of that within the last couple of year with some non-infectious products that were causing a great deal of harm, i.e. hypersensitivity reactions both generalized and systemic causing untoward disease namely immune mediated disease which we had never seen in that species prior to the introduction of a non-infectious vaccine.

Rude: In the past, inactivated or as we now say, non-infectious, virsus vaccines have been criticized for inducing low levels of immunity of rather short duration in vaccinated animals. The early killed canine parvovirus vaccines were an example, How do the newer non-infectious virus vaccines perform in this regard?

Tizard: I think that's a fair statement you just made, that indeed at least in veterinary medicine the non-infectious products have been poorer vaccines than the modified live ones. One of the reasons for our enthusiasm for modified live products has been that they have been somewhat better vaccines.
     In addition, of course, we have also needed to incorporate considerably more antigen is a non-infectious product than in a modified live product in order to get enough antigen into the dose and this had an adverse effect; on the price. Until we recognized that we had to enhance the immunogenicity of these non-living products. They were secondary products that could not compete with the modified living ones.

Rude: When you talk about enhancing the immunogenicity of non-living products you are referring to adjuvants How Important are adjuvants in non-infectious virus vaccines?

Tizard: They make a whole range of non-infectious, noninfectious living products possible that otherwise would not be possible. This is especially true as we move into genetically-engineered products, into sub-unit products, into synthetic products where relatively small quantities of relatively poorly antigenic material are used. As Dr. Salk pointed out, if you want to do better than nature you're going to have to add something to that product in order to get a superior immune response. An immune response that is at least of a size and duration similar to the currently available modified live virus vaccines. This all centers on the successful use of adjuvants.

Siegl: I would like to raise two points. The first is that, with virology proceeding, we certainly have made progress towards production of more antigen at lower cost. I'm thinking of canine parvovirus vaccine where we can now grow the virus to high concentrations. This possibility has been denied before, when most laboratories failed to produce high concentrations of antigen in cell culture. From that time stems the difficulty of immunizing dogs with a killed or non-infectious parvovirus vaccine.
     The second point is in context with genetically engineered vaccines, sub-unit vaccines and so on. We now begin to understand how the immune system works. We know what antigenic determinants must be present to trigger the immune system at its various levels and there's a huge possibility for the future to produce good and effective non-infectious vaccines.

Schultz: I think one of the things that the veterinary community is having problems with again, as it had problems with for years, is considering antibody responses to mean protective immunity.
     When we develop new, inactivated or non-infectious viruses and as we add new adjuvants the principle measure of immunity, unfortunately that the companies are using are antibody titers to the virus. Some or these titers are higher than we've ever seen before but the vaccines are no more efficacious against challenge with the disease organism than were the older, non-infectious products that didn't induce or induced a just detectable antibody response. So we have to be very careful that we understand the mechanism of protective immunity and realize it's not always evident by measuring an antibody titer.
     It veterinary biologic companies will keep that in mind and will slop using antibody titers as a marketing ploy which they have used for year to the veterinary practitioner, most of whom feel antibody titers an synonymous with protective immunity, we might better serve our clients and their animals.

Salk: I think this deserves the attention that you're giving it. I have tried to determine what the correlates of immunity were. In the case of influenza. we found that it related to keep of antibody at the time of infection. The reason is that the portal of entry and of pathology in the same. That's where the block of infection has to occur.   
     In the case of polio, the minimum requirement for immunity to disease, i.e., paralysis, is merely the presence of immunologic memory. We do not require persistence of detectable antibody. The presence of memory can be determined by giving a dose of vaccine and observing a maximum response within seven days.|
     In the case or polio a single effective dose can produce lifelong immunity because memory is durable. In the case of influenza you need two doses in order to induce the anamnestic response and in addition, you need an adjuvant. For polio, you don't. We've come a long way in understanding the mechanism of immunity, and therefore, how to design an immunogen to produce protection against clinical disease, i.e., protection against paralysis, be it paralysis of the immune system or paralysis of the central nervous system or wherever find of pathology it may be, hepatitis, etc. The immunogen needs to be formulated to produce the desired effect.

Tizard: May I make a more general comment following this discussion an immunology? Veterinary vaccines, while remarkably successful in controlling disease, have had major areas of failure where they have not been impressively successful. The two I can think of are control of diarrhea diseases and the control of respiratory disease in large animals.
     Much of thin stems from our lack of knowledge about the things we were talking about earlier, In many cases, we succumbed to the ideas that the appearance of antibodies is sufficient to cause protection. Although there has been a tremendous growth of knowledge of the exact mechanism of defense in many respiratory and enteric diseases, until we know as much about them as we do about viruses such a distemper, we maybe are doomed to continue our unimpressive performance.

Salk: Apropos of the remarks I made earlier about Influenza where there is a short incubation period, you don't have the advantage of the anamnestic effect after exposure as in the prevention against disease as with rabies, hepatitis, polio, measles, etc. For respiratory disease generally and enteric disease, became of portal of entry, you do have to have antibody at the portal of entry and at a higher level than you would for the other diseases because for respiratory and enteric disease the degree of immunity locally in proportional to the level of antibody. It would be possible to develop improved vaccines by the use of adjuvants a we've been able to show for influenza in humans. There are remedies. We need to understand the mechanisms and the differences to formulate the different vaccines.

Rude: This rationale for the use of adjuvants is primarily to enhance antibody-mediated and cell-mediated immune responses. Would you give an explanation about what we mean by antibody-mediated immune responses and cell-mediated immune responses?

Schultz: With regard to cellular immunity sometimes referred to as cell-mediated immunity, we have to include phagocytic processes with macrophages and T-cell cytotoxic lymphocyte responses. I would also include natural effect or natural cell-mediated cytotoxicity as cellular immunity and delayed type hypersensitivity. There is a transitional form of immunity called antibody dependent cell-mediated cytotoxicity (ADCC) that require both cells and antibody for protective immunity to disease. Then there is the classical humoral immunity in which antibody alone provides protection.
     It's important to understand that for certain viral agents and perhaps for most, both cellular and humoral immunity play a role in protection against clinical disease. However, to my knowledge, the only specific immune factor that can truly prevent infection with a virus is antibody. It's probably rare that we achieve that level of protection, but I just gave an example of it in canine distemper when. if you look at maternal antibody which in the only thing available to prevent infection is the antibody that's present as a result of transfer in the colostrum, then you get some ides of how important humoral or antibody-mediated protection is. That puppy with maternal antibody cannot be infected: is able to resist challenge with a particular agent. Distemper is an example where antibody alone will work to provide solid protection against infection.

Rude: Dr. Salk, you've had experience with the polio virus in this regard. Would you like to comment?

Salk: In polio, the immune response that's required is not the same. It's altogether humoral, and in fact, it is so efficient that all you need is the capacity to respond rapidly. The only individuals who become paralyzed are those who have a sluggish immune response to the intestinal component of their infection. The immune response, as evidenced by the appearance of antibody in the blood, is too late to prevent the viremia that occurs. The purpose of immunization is simply to accelerate the responsiveness sufficiently which can be done by inducing enough memory.
     In the case of influenza, as I mentioned earlier, memory is not enough. There you also need antibody for preventing the infection.
     With a disease like AIDS the virus may very well be intracellular at the time of infection. Here it maybe possible to protect against disease but not against infection. Here the cellular component may be important. Both cellular and humoral factors an necessary for certain diseases, of which AIDS is an example and tuberculosis and many other bacterial diseases and parasitic infestations as well. Consequently. the immunogens that are needed have to be designed appropriately in each case.
     In the care of bacterial disease, such a diphtheria or tetanus, the antitoxin alone is altogether sufficient. It is important for practitioners to understand these differences and in that way anticipate the limitations as well as opportunities of a given immunogen.

Rude: Do you believe the development of new types of adjuvants, that further enhance both antibody-mediated and cell-mediated immune responses or cellular immunity is important?

Salk: Yes, so far as adjuvants are concerned, some of them can produce immunopathic effects as well as beneficial effects. In the case of complete Freund's adjuvant (mineral oil plus non-infectious mycobacterium tuberculosis bacilli), for example, when combined with a central nervous system component (i.e. myelin basic protein), you can actually produce experimental allergic encephalomyelitis. If you were to use incomplete Freunds' adjuvant (mineral oil without non-Infectious mycobacterium tuberculosis bacilli), you could actually prevent the induction of allergic encephalomyelitis by immunizing the animal with incomplete Freund's adjuvant
     This in an Illustration of the need to be circumspect about the nature of the adjuvant that you employ so as see to enhance the beneficial as compared to the undesirable effect. It's for this reason that there's a great misunderstanding about the usefulness of Freund's adjuvants. Freund's incomplete adjuvant is very useful and does not cure the detrimental effects that can be associated with the use of Freund's complete adjuvant. We were able to show this with influenza vaccine some 35 years ago,
     The application of that principle can now be further studied by the development of adjuvants designed specifically to produce particular desire effects, depending on which class of antibody you want to produce and whether cell-mediated or humoral immunity. We're entering a new phase, a new science you might say, in the development and use of adjuvants. This, I predict, is the next breakthrough in the evolution of vaccinology. 

Schultz: I didn't mention anything about adjuvants when you asked the previous question, but just very briefly, to build on what Dr. Salk has indicated in veterinary medicine we're doing to have to be very careful about the adjuvants we use. There is currently a great fervor and desire to get into the addition of many cytokines which include both lymphokines and monokines, without really having any knowledge about their effect on the whole body system. We're adding such things as interleukins to vaccines without knowing what the interleukines are going to do to the animal and to the immune system and perhaps they may be generating immunologic diseases.
     We have to add a note of caution at this point. Since these adjuvants and these products are so poorly understood, we need to enter into using them with the hope of better understanding and if ,in fact, we don't have that understanding, we need to get the results from their use into the hands of the scientific community so we can better explore the potential effects that they might have on an immunogen and on the immune system.

Tizard: And yet, the use of poorly understood adjuvants has a long end distinguished history in vaccinology. We've been using alum since the 1920s and are still not sure how it works. It's also fair to say that we've been very conservative is our use of adjuvants. To the best of my knowledge, alum is still the only adjuvant used in human vaccines. In veterinary medicine there are a few examples of saponin or DEAE dextrose being used.

Schultz: Newer adjuvants are in use now in veterinary vaccines.

Tizard: Yes, but in veterinary medicine the problems caused by adjuvants relate not only to safety and efficacy but also to contamination of meat and meat products.

Siegl: I would say a complete new field is developing. At its scientific level there is an especially huge effort being made to understand how all these adjuvants are working and what is really the active substance. We might get away from using the old mycobacterial extract and go down to some few amino acid or peptides or phosphorylated peptides to see how these work. In addition, as far as I see it, there is a field developing on how you can apply these adjuvants in the best possible way, that they really produce the effect you like.
     For example, adding these adjuvants to the vaccines directly or blending them in might not be as beneficial as applying the adjuvant is the form of liposomes or something like that.

Swango: Just a general comment about the adjuvants and where we're at. I think this certainly is a new ere that we're entering in which, in the next few years, we're going to see a lot of information being developed on how some of these factors are beginning to operate. The only additional comment is just to underscore what has been alluded to and stated here by Dr. Schultz and Tizard. In food animal medicine, we have to be very careful about residues of products in animals following vaccination, so we may be confronting a different problem in that facet of veterinary medicine than what we will be in companion animal medicine or human medicine.

Rude: Earlier, when we were talking about modified live virus vaccines, we talked about virus shedding. Some of you mentioned there might be some benefits associated with shedding of modified live vaccine viruses from vaccinated animals. Would you characterize the lack of viral shedding as a disadvantage which might be associated with non-infectious vaccines when they are used to immunize a herd of cattle or a litter of puppies?

Swango: With respect to canine parvo we have a unique situation in the passive immunity of that disease in which, as puppies are, losing their maternally derived antibody, they reach a point at which they become susceptible to infection with virulent virus earlier than what they can be immunized with vaccine, so we have a window of increased susceptibility but not immunizable and we have variation of titers within a litter. Everybody is not going to be on the same scale, so it is
conceivable and in fact it has been demonstrated that on a given day we may have one individual in a group that can respond to vaccine, either killed or live. If we're using a live virus vaccine, that animal may be shedding the virus, depending on the vaccine that's been used a few days following vaccination at a time when its litter mates become susceptible or immunizable and hence, become infected and respond to that live virus vaccine.
     So, in essence, with certain live virus vaccines, one can immunize one individual and in turn immunize the entire group. whereas with the killed vaccine that cannot happen.
     Earlier, I think it was Dr. Schultz who said we needed to be careful in what we communicate to practicing veterinarians because we have gone through an era where there was a lot of promotional emphasis pointing out all the adverse effects of virus shedding from vaccinated animals lymphopenia. Most of these comments that had been perpetrated on our profession were not based on factual data. They were based on erroneous assumption. Canine parvovirus
is not immuno-suppressive, period, even though virulent virus, in certain vaccines, do cause lymphopenia following vaccination. It is not immuno-suppressive. Shedding of the vaccine virus from canine parvovirus has not been demonstrated by anyone with definitive data to cause a problem and, therefore a lot of the promotional emphasis stressing lack of shedding of virus lack of
lymphopenia, or in one or two cases, killed vaccine, was off base. It only generated a lot of fear response among dog owners as well as veterinarians about the use of these vaccines. That's one advantage to a live virus vaccine in the case.

Tizard: While what Dr. Swango is saying almost certainly true, it is also in sense anti-scientific to simply put an a virulent strain into a population and hope that by doing this it will transmit to the unprotected individuals. I would have thought that it would be much more appropriate to use a slightly different technology such as a slow release vaccine to in fact, ensure that animals get protected in a more formal and organized manner.

Swango: I agree with Dr. Tizard team the standpoint of approaching a better solution to a problem, but when we've got a dog developing a parvovirus disease. we cannot wait for our scientists develop a slow release product..

Tizard: I support that.

Salk: There are two things I'd like to comment on. One has to do with the herd effect as I understood it. To me, it means that if you immunize a fraction of the population that the rest of the population will be protected. Vaccinating a segment of the population protests the unvaccinated segment, Is that true?

Tizard: Yes. That's true.

Salk: We have shown that in influenza and in polio. That's what I mean by herd effect..

Swango: We can show you that for rabies control as well in animals.

Salk: I don't doubt that. You reduce the number of disseminator and therefore the potential recipients are protected. For every disease there is a threshold beyond which you need not go to have a desired effect. That's one way of dealing with the problem that you were referring to. You don't have to shed virus in order to produce a herd effect. That's not what I understand to be the herd effect. That is spreading by shedding. We have used incomplete Freund's adjuvant to protect antigens from maternal anti-body.

Schultz: I'll use canine parvovirus in this specific example. We've been working on the problem of overcoming maternal immunity to a variety of agents to about 15 years now and with regard to canine parvovirus specifically we have used incomplete Freund's adjuvant. We are using liposomes 'with both intact virions and make DNA. We have also used viral vectored vaccines. I can categorically state that Incomplete Freund's adjuvants and other adjuvants will not overcome this suppression by maternal immunity to canine parvovirus 
     Even when we give incomplete or complete Feund's adjuvant with Vaccine to animals that we would consider to be just at the border of being susceptible to immunization (maternal antibody is very low) in our hands that happens to be about 1:40 HI titer canine parvovirus adjuvant will not make the vaccine immunize I don't know why the addition of adjuvant will not work but we can show that just by increasing the antigenic mass we can overcome a 1:40 maternal titer. I'd like to go on record that we have not found any commercial canine parvovirus vaccine that will overcome maternal immunity even at levels as low as I:40 to 1:80 titer. So we're dealing in canine parvovirus disease with this "window of vulnerability" where the puppy becomes susceptible to infection with virulent virus two to four weeks before the current vaccines are able to actively immunize which may not be until 16, I8 or 20 weeks. The only way that I know of overcoming that problem will be by increasing the antigenic mass in the vaccine not by adding an adjuvant.

Salk: You cannot possibly do magic by the use of an adjuvant unless the amount of antigen is adequate, so that is clear. The other point I was going to raise was whether or not the immunogen is a carbohydrate immunogen because, under those circumstances there is delay.

Schultz: No. this is a protein.

Rude: I have always believed, a Dr. Salk has mentioned, that an adjuvant, such as Freund's incomplete adjuvant. protects a non-infections virus from being neutralized by maternal antibodies. I have also believed that the slow release of a non-infectious virus from an adjuvant depot may induce an active immune response if maternal anti-bodies have decreased to non-protective levels. Based on what you have stated. Dr. Schultz, this does not happen?

Schultz: In practice it does not happen. The vaccines that we currently have available with adjuvants do not immunize animals with maternal antibody more effectively than the modified live vaccines on the market. The reason for the inability of the inactivated vaccines to immunize are not clear but may relate to the adjuvant being used or the lack of an adequate antigenic mass. Thus, although your statement is theoretically correct, field experience would
suggest that adjuvanted non-infectious vaccines do not effectively immunize animals with maternal immunity.
     Likewise, we're not seeing an anamnestic response from most modified live or non-infectious products. So this yearly vaccination, for example, for distemper, cause an anamnestic response in less than 1 percent of vaccinated dogs.

Rude: If a non-infectious virus vaccine is used for the initial vaccination, should animals be revaccinated with a non-infectious or modified live virus vaccine, or dose it matter?

Tizard: It doesn't matter but the USDA license is issued for a matched set of vaccines.

Siegl: Several concepts have been developed which call for revaccination with a modified live vaccine. In my opinion, this is nothing but a compromise to come to meet both the adherents of non-infectious and of live vaccines. Personally. I can see no need for using modified live virus for the revaccination and, anyway. I favor the use of non-infectious vaccines whenever possible.

Schultz: If an animal is vaccinated with a non-infectious vaccine it should, if necessary, be revaccinated with a non-infectious vaccine, since the immune response from the original vaccination would likely interfere with a modified live vaccine. For the most effective immunity when modified live vaccines and non-infectious products are used in the vaccination programs the modified live vaccine should be used fist, followed by the non-infectious vaccine. This is done for two reasons.
     First, the modified live vaccine usually provides more effective and complete immunity, and secondly, the non-infectious vaccine is more likely to provide an anamnestic response. Non-infectious vaccine fail to stimulate certain aspects of cellular and local immunity, therefore, if the modified live vaccine is used first, these forms of immunity will present to protect against disease.

"If a non-infectious virus vaccine is used for the initial vaccination, should animals be revaccinated with a non-infectious or modified live virus vaccine?"     Dr. Ted Rude

"We're entering a new phase, a new science, you might say, in the development and use of adjuvants. This, I predict, is the next breakthrough in the evolution of vaccinology. " Dr. Jonas Salk

"Both modified live and inactivated vaccines have a in a veterinary practice... if we have a choice, and we truly have the same efficacy in both products, I think everyone at this table would opt for the inactivated vaccine because of the potential safety aspect."
Dr. Ron Schultz

" We now begin to understand how the immune system works. We know that antigenic determinants must be present to trigger the immune system at its various levels and there's a huge possibility for the future to produce good and effective, non- infectious vaccines."          Dr. Gunter Siegl

We somehow need to redirect emphasis that just because we vaccinate, we cannot ensure immunity, There is a difference between vaccination immunization. 
Dr. Larry Swango

"'While we certainly have to give credit to modified live veterinary vaccines for having had a tremendous impact in controlling disease in this country, they do represent a fairly basic technology that can be improved upon."      
Dr. Tizard