Strategies for Vaccinating Mares, Foals, and Weanlings

The primary goals of vaccination programs for pregnant mares are preventing diseases that could affect the mare or her fetus and maximizing the level of colostral antibodies that will be passively absorbed by the neonatal foal after nursing, which provides the foal with protection against diseases that pose a risk during the first few months of life. Additional considerations in the selection of vaccines for use in pregnant mares include the safety to the mare and fetus, the influence of pregnancy on vaccine responses, and the potential for interference between multiple vaccines administered simultaneously. Maintenance of consistent vaccination protocols for broodmares will maximize the uniformity of maternal antibody protection provided to the foal. Maternal antibodies may, however, substantially interfere with the response of foals to many vaccinal antigens and may complicate priming of the immune system of the foal against important pathogens. Documentation that maternal antibody interference substantially impacts the responses of young foals to many of the antigens commonly included in equine vaccination programs has led to revision of the traditional recommendation to begin primary immunization of foals at 3 months of age or earlier [1]. In addition to inhibiting the primary response of foals to many antigens, including influenza, tetanus, equine herpes virus type 1 (EHV-1), equine herpes virus type 4 (EHV-4), Eastern equine encephalomyelitis (EEE), Western equine encephalomyelitis (WEE), and rabies, maternal antibodies have also been shown to interfere with the ability of foals to mount appropriate responses to inactivated influenza vaccines administered later in life.

Although this phenomenon does not constitute true immunotolerance, it is nevertheless a cause for concern. For influenza and diseases that are of low risk during the first year of life, it is advisable to delay primary immunization until foals are at least 6 months of age to minimize the likelihood that maternal-antibody interference will compromise protection induced by the primary series. Maternal-antibody interference renders it difficult or impossible to adequately immunize foals against certain high-risk diseases that foals are likely to be exposed to during the first 6 months of life such as EHV-4, EHV-1, and, in the southeastern states, EEE. For these diseases, it is rational to narrow the "window of susceptibility" as much as possible by commencing vaccination at less than 6 months of age, although maternal antibodies will likely interfere with the response of many foals to these antigens. Additional doses of vaccine are typically added to the primary immunization series to counter this effect. In contrast to the experience with most other inactivated vaccines, the inactivated West Nile Virus (WNV) vaccine seems to be capable of priming the immune response of 3-month-old foals born to antibody-positive mares, making it possible to protect foals against WNV during the high-risk summer and fall months of their first year of life. Regardless of the age when foals are first vaccinated and the inactivated vaccine used (with the exception of rabies), at least three doses of vaccine, rather than the two recommended by most vaccine manufacturers, seem to be necessary for adequate primary immunization [1].

1. Introduction

Much of the research performed in the areas of equine infectious disease and immunology during the past 50 yrs has been directed toward developing vaccines as aids to the prevention of important infectious diseases. Vaccination of horses is now widely practiced and constitutes an important part of integrated infectious-disease control programs on breeding farms where the predominant focus is on broodmares, foals, weanlings, and yearlings. Fully licensed or conditionally licensed vaccines are now available in North America as aids to the prevention of tetanus, the viral encephalitides (Eastern equine encephalomyelitis [EEE], Western equine encephalomyelitis [WEE], Venezuelean equine encephalomyelitis [VEE]), West Nile virus (WNV) infection, influenza, equine herpes virus type 1 (EHV-1) infection, equine herpes virus type 4 (EHV-4) infection, strangles, rabies, equine viral arteritis (EVA), Potomac horse fever (PHF), Type B botulism, rotavirus infection, and equine protozoal myeloencephalopathy (EPM). Label directions for use of these vaccines typically include recommendations regarding the schedule for primary immunization and revaccination, but only a handful of the available products provide any recommendations regarding use in pregnant mares or the minimum age at which to initiate the primary vaccination series in foals.

Similarly, the published literature addressing these issues is sparse. This paucity of information has led to the development of vaccination protocols for mares and foals that are not based on either label directions or published research data. It is, therefore, important to consider the potential influence of the many pre-partum and postpartum interactions that occur between the mare and her foal when establishing the overall goals of the vaccination program and the specific vaccination protocols that are necessary to achieve these goals. Ideally, vaccination schedules for each age group and type of horse should be coordinated so that they maximally benefit all groups.

2. Considerations for Use of Vaccines in Broodmares

While vaccination programs for pleasure, show, and performance horses typically focus on prevention of diseases that pose a risk to the individual horse and it's herd mates, vaccination protocols for broodmares must also consider protection of the fetus, both prenatally and during the first few months of postnatal life. Both EHV-1 and EVA are capable of inducing fetal loss, abortion, or the birth of infected live but severely compromised foals. Thus, these two diseases command special attention in vaccination programs for broodmares. Capitalizing on the protective effect of maternal antibodies passively transferred to the foal through colostrum constitutes an additional major focus of vaccination programs for broodmares. This goal is typically accomplished by administration of booster vaccines to mares during the last 2 months of gestation to maximize levels of specific colostral antibodies directed against diseases that pose a risk to the foal during the first few weeks of life. Maintaining consistent vaccination protocols for mares will maximize the likelihood that a uniformly high level of colostral antibody transfer and passive protection will be achieved within the foal crop. While intranasally administered vaccines may afford good protection to the mare, they are typically less effective than parenterally administered inactivated vaccines in stimulating high levels of circulating IgG, the isotype that is passively transferred to the foal in highest concentration. Parenterally administered vaccines are, therefore, preferred over intranasally administered vaccines for vaccination of mares during late gestation.

Of the many antigens available for inclusion in vaccines for use in horses, tetanus, EHV-1, and WNV are typically considered to be "core" diseases against which all broodmares in North America should be vaccinated. In addition, inclusion of EEE and WEE antigens is strongly recommended in the many states and provinces in which these two arboviral diseases are endemic. Similarly, vaccination of mares against rabies is indicated in endemic areas. The high horse traffic and high concentration of foals and young horses that typify many breeding farms contribute to a high risk of exposure to contagious respiratory diseases, including EHV-4, EHV-1, influenza, and strangles. Inclusion of influenza and EHV-4 antigens in vaccination protocols for broodmares is, therefore, routinely practiced, and the addition of strangles vaccines is frequently recommended when conditions of significant risk are anticipated. Vaccination of mares against EVA before breeding may be indicated when they are to be bred to a known or suspected EVA-carrier stallion, either by natural cover or by artificial insemination. While protection of the broodmare and fetus or herd mates against the abortigenic effects of EHV-1 or EVA is the primary goal underlying inclusion of these antigens in vaccination protocols for broodmares, protection of the foal features at least as prominently as protection of the mare in the rationale for vaccinating mares against tetanus, WNV, EEE, WEE, rabies, influenza, EHV-4, and strangles. Inclusion of rotavirus and botulism vaccines in protocols for pregnant mares is directed almost exclusively at protecting the young foal against these diseases.

Consideration of vaccine safety in broodmares must take into account risks to the pregnancy and safety to the fetus. Potential adverse effects of vaccines on pregnancy are difficult to document, even when large numbers of mares are used, unless obvious problems occur. Because fetal organogenesis occurs early in gestation and this period is also characterized by substantial embryonic loss even in normal mares, it is sound practice to avoid administering vaccines to mares during the first 60 days of gestation unless conditions of imminent risk prevail. Few vaccines carry specific label recommendations for use in pregnant mares and little published data exists to document the safety of equine vaccines during pregnancy. In addition to the two available EHV-1 vaccines [a,b] marketed for use in pregnant mares as an aid to prevention of EHV-1 abortion, only one other fully licensed vaccine [c] for prevention of Type B botulism in foals and one conditionally licensed vaccine [d] for prevention of rotavirus infection in foals carry label recommendations for use in pregnant mares.

Although not specifically labeled for administration during pregnancy, widespread use in practice over many years has failed to document that any of the inactivated vaccines currently marketed for use in horses pose an unacceptable risk to pregnant mares. Therefore, pregnant mares are routinely vaccinated with inactivated vaccines directed against tetanus, EEE, WEE, WNV, influenza, EHV-4, strangles, and, to a lesser extent, PHF, rabies, and VEE. Similarly, adverse impacts on pregnancy have not been documented for modified live intranasally administered strangles and influenza vaccines or the modified live parenterally administered EHV-1 vaccine [e]. In addition, safety of the recombinant WNV vaccine [f] should not be a significant concern, because the modified live canarypox vector lacks the ability to infect mammalian cells. In contrast, modified live virus EVA and VEE vaccines and live anthrax spore vaccines should not be used in pregnant mares. Protection of mares against the potential abortigenic effects of EVA infection is, therefore, best accomplished by completing the primary immunization series before the mare enters the broodmare band and by administering subsequent boosters during the open period before rebreeding [2].

The practice of booster vaccinating mares against multiple diseases to maximize colostral transfer of antibodies to the foal and the fact that mares in broodmare bands are generally middle aged or older results in the typical broodmare receiving multiple doses of many vaccine antigens and adjuvants during her lifetime. In addition to stimulating high levels of antibodies against a range of antigens, this practice may also predispose these mares to a higher rate of local and systemic adverse reactions, an issue that not only warrants further investigation but also may force horse owners and veterinarians to carefully consider strategies for revaccination. Simultaneous administration of multiple vaccines to mares late in gestation is a common practice that may increase the risk of an adverse reaction. In addition, the possibility that "competition" between multiple antigens will compromise the response to some or all of the administered antigens should be considered. When administration of multiple vaccines late in gestation is indicated, it is good practice to administer no more than four antigens at one time and to allow an interval of 2 - 4 wks between administration of different vaccines.

It is widely assumed that pregnant mares are fully capable of mounting appropriate cellular and humoral immune responses to vaccines; however, this issue has received little research attention. Although mares that have been primed before breeding seem to mount appropriate anamnestic responses to vaccines, we have generated preliminary data suggesting that the humoral response to primary vaccination with several inactivated vaccines may be down-regulated during gestation, resulting in failure of vaccinated mares to passively transfer specific antibodies to the foal through colostrum. We are currently researching this issue because, if proven, it has obvious ramifications to the immunoprotection of both the foal and the mare.

3. Considerations for Use of Vaccines in Foals and Weanlings

Challenge studies to support licensing of vaccines by the United States Department of Agriculture (USDA) generally require horses to be antibody negative; therefore, foals are not typically used to avoid the potential confounding effects of maternal antibodies. In addition, studies in foals to document the potential inhibitory effects of maternal antibodies are not required for licensing. It is not, therefore, surprising that few vaccines carry label directions for primary immunization of foals. Of those that do, the range of recommended minimum ages is wide and includes the following:3 months for several inactivated rabies vaccines, 6 months for inactivated influenza and EHV-1 vaccines from one company [g] 9 months for a modified live intranasal strangles vaccine [h] and 11 months for a modified live intranasal influenza vaccine [i]. Considering that foals are at risk for acquiring many of the diseases against which we vaccinate horses and the fact that the period before weaning is regarded by many horse owners and veterinarians as a convenient time to vaccinate, it became common practice during the 1970s and 1980s to start primary vaccination of foals at 3 - 4 months of age. The primary immunization series for multiple antigens could, therefore, be completed before weaning. AAEP vaccination guidelines published during the 1980s and early 1990s supported this practice, despite a paucity of published data documenting efficacy [3].

While foals are considered to be immunocompetent at birth and are capable of mounting both humoral and cellular immune responses to a range of antigens, continued maturation of the immune system after birth is believed to be necessary to achieve optimal immune function. Few studies have, however, been done to determine the age at which various elements of the innate and adaptive immune system become fully competent and optimal responses to vaccines can be achieved. Maternal antibodies, and perhaps other important immune effectors such as lymphocytes, that are concentrated in colostrum and are passively transferred to the foal play a crucial role in defense against pathogens encountered during the first few months of life when endogenous immune function continues to mature. Although several immunoglobulin isotypes are present in colostrum, the sub-isotypes of IgG are absorbed into the systemic circulation of the foal in much higher concentration than either IgM or IgA [4]. Specific IgA is, however, secreted continuously in milk and may provide passive protection against pathogens such as S. equi and enteric organisms by helping coat the pharyngeal and intestinal mucosa, thereby neutralizing pathogens [4,5]. When foals ingest adequate amounts of high-quality colostrum during the first 12 - 24 h after birth, titers of specific antibody in the serum of the foal are typically very similar to the serum titer in the mare at the time of foaling. While the rate of decay of specific sub-isotypes of IgG varies to some extent, the overall half-life of decline of maternal IgG antibodies is typically between 25 and 40 days [6-12]. Thus, the magnitude of the post-nursing antibody titer and the sensitivity of the assay used to detect passively transferred antibodies will determine persistence of these antibodies at measurable levels in the serum of foals. Use of enzyme-linked immunoabsorbent assays (ELISAs) and other sensitive assays has made it possible to detect persistence of maternal antibodies for 6 months or longer in some cases [10].

In addition to passively protecting the foal against pathogens encountered during the first few months of life, maternal antibodies have been shown to exert a profound inhibitory effect on the immune responses of foals to antigens, including those contained in vaccines. Several studies reported by groups in Holland, Ireland, and the United States during the 1990s brought this issue into focus by showing that foals <6 months of age consistently failed to mount serologic responses to inactivated influenza vaccines [8-10,13-16]. Of greater concern was the finding that a high proportion of foals vaccinated under the cover of maternal antibody not only failed to seroconvert in response to the recommended primary series of two or three doses of influenza vaccine, but also many failed to respond to multiple additional doses administered during the next year; this suggests the induction of a potentially detrimental "immunotolerance-like" phenomenon [13,14,17]. Subsequent studies by our group confirmed an apparent lack of response of foals to multiple doses of inactivated influenza vaccines when the hemagglutination inhibition (HI) test was used to detect serologic responses. When the same samples were retested using sensitive isotype-specific ELISA tests, it was found that 6-month-old foals did mount a response that included all IgG sub-isotypes but was less vigorous for the more important virus-neutralizing IgGa and IgGb sub-isotypes than for the less-effective IgG(T) sub-isotype [10]. Although there seemed to be some differences in responses to different vaccines containing different adjuvants, this "misdirection" of isotype responses in favor of IgGT, likely influenced by maternal antibodies, was consistently observed [10]. Subsequent studies in which titers of total, rather than antigen-specific, IgG sub-isotypes were determined documented that the age-related increase in concentrations of IgGb lagged significantly behind increases in concentrations of other isotypes and remained below adult levels beyond 6 months of age [18].

Maternal-antibody interference has now been documented to be a significant issue for several antigens, including tetanus, EEE, WEE, EHV-1, and EHV-4, contained in vaccines administered to foals [1,10,11,19-21]. Even low levels of antibody, below those detectable by many routine serologic tests and below those thought to be protective, can completely block the serologic response to some vaccines, resulting in a potentially prolonged period of susceptibility before the foal is capable of responding appropriately to vaccines [1]. These findings also indicate that it is not typically feasible to test samples from foals serologically to predict whether they will respond to particular vaccines. We now recommend that primary immunization with most vaccines containing inactivated antigens should be delayed until foals are 6 months of age or older and, with the exception of the rabies vaccine, three doses of vaccine should be included in the primary series rather than the two doses routinely recommended by vaccine manufacturers. Typically, the third dose stimulates a serologic response of greater magnitude and durability than the two doses alone and may also set a higher "set-point" for the response to subsequent booster doses [1,10,22]. In contrast to the results cited above, recent studies in our laboratory have shown that maternal antibodies do not seem to exert a marked inhibitory effect on the response of foals to the inactivated West Nile virus vaccine [j] thus permitting antibody-positive foals as young as 3 months of age to be immunized successfully.

The results of studies investigating maternal antibody interference with responses to vaccines should be interpreted with caution, because only humoral responses are typically assessed and infectious challenge is not performed to confirm that the lack of serologic response equates to lack of protection. Lack of a serologic response may correlate well with lack of protection for some diseases, but not for others. In contrast, the presence of a serologic response may not correlate with protection, which is frequently the case for respiratory-tract pathogens. With the exception of the intranasally administered strangles and influenza vaccines [h,i] the modified live virus EHV-1 vaccine [e] the modified live virus EVA vaccine [k] and the canarypox vectored WNV vaccine [f] most commercially available vaccines are inactivated, adjuvanted, and administered by IM injection. As such, they are much more likely to induce a systemic serologic response than either a cell-mediated immune response or local mucosal immunity.

When formulating a vaccination protocol for foals, it is important to determine which one of the following is the most important goal or whether both are equally important:

• To protect the foal and weanling against specific high-risk infectious diseases that affect this age group and have the potential to cause significant disease, either directly or by predisposing the animal to other secondary infections
• To initiate primary immunization to protect against disease later in life

Assessing risk takes into account both the incidence of disease (i.e., the likelihood that the foal will become infected) as well as the risk of serious sequelae or death if the horse does become infected. If the disease affects the foal early in life, such as with rotavirus infection, there is usually insufficient time to induce a protective immune response by actively immunizing the foal. Under these circumstances, the approach should be to maximize the degree of protection passively transferred from the mare through colostrum. Other diseases, such as rabies, affect horses of all ages, but the risk of acquiring infection is generally low.

Diseases of moderate to high risk to young foals but low risk to adults include rotavirus infection (on certain breeding farms in certain years) and, in geographic areas such as Kentucky and some other eastern states, type B botulism. For these diseases, the best approach is to:

1. Booster vaccinate the mare before foaling to maximize uniformity of passive transfer.
2. Ensure good passive transfer of maternal antibodies.
3. Introduce management practices to reduce exposure to the infectious agent.
4. Vaccinate the foal if risk continues beyond first few months of life.

Diseases of moderate to high risk for weanlings and older horses but lower risk to young foals born to vaccinated mares include EHV-4, EHV-1, strangles, influenza, tetanus, and, in the eastern and southeastern United States, EEE. For these diseases, the best approach is to:

1. Vaccinate the mare before foaling to maximize uniformity of passive transfer.
2. Ensure good passive transfer of maternal antibodies.
3. Start foal vaccination after the risk of maternal antibody interference is no longer present in most foals.
4. Introduce management practices to reduce exposure to the infectious agent while primary vaccination is being completed.
5. Use three or more doses of vaccine in the primary series to improve the chances that foals that did not respond to earlier doses will respond when given additional doses later.

Diseases of low risk to foals in most circumstances include rabies, PHF, WEE, and EVA. For these diseases, the best approach is to:

1. Vaccinate the mare before foaling if the disease is a significant risk to adult horses and a vaccine shown to be safe for use in pregnant mares is available. If the available vaccines are not considered safe for use in pregnant mares, administer boosters before breeding.
2. Ensure good passive transfer of maternal antibodies.
3. Start foal vaccination after the risk of maternal antibody interference is no longer present in any foal (typically 9 months to 1 yr of age).

4. Considerations for Specific Diseases

WNV Infection

In the 5 yrs since the first cases of WNV infection in North America were diagnosed in dead birds at the Bronx zoo, this mosquito-borne viral disease has spread in all directions and has become endemic in almost all mainland states and provinces in the United States and Canada. As of March 2005, the disease has been confirmed in >22,000 horses, ~35% of which have died or been euthanized. Although the risk of infection and death seem to increase with increasing age, the disease has been confirmed in foals as young as 3 wks of age. Although cases have been seen virtually year round in the southeastern states, the risk of acquiring infection is highest during those months in which mosquito activity peaks (typically July, August, September, and October in most states).

Two fully licensed vaccines (West Nile Innovator [j] and Recombitek [f]) are available for use in horses. The West Nile Innovator [j] vaccine is inactivated and contains a metabolizable oil adjuvant. This vaccine is available as either a monovalent (single component) or as a multivalent vaccine containing other encephalitis virus antigens (EEE and WEE). The Merial Recombitek [f] vaccine is a modified live canarypox-vectored vaccine. Both vaccines have met USDA requirements for safety in tests involving >600 horses. In challenge models, both have been proven to significantly reduce the magnitude of viremia (amount of virus circulating in the blood) in experimentally infected vaccinated horses compared with non-vaccinated control horses for as long as 12 months after primary vaccination with two doses of vaccine [23,24]. There is clear evidence that vaccination reduces the risk of infection and death after natural challenge in the field setting, although clinical disease may not be fully prevented [25,26].

Directions for use of both vaccines include administration of two doses of vaccine 3 - 6 wks apart (consult the specific label). Optimal protection cannot be expected until 2 wks after administration of the second dose, although experimental studies with the Recombitek [f] vaccine have documented significant protection as early as 26 days after administration of the first dose [27]. The vaccine manufacturers recommend revaccination of previously vaccinated horses on an annual basis or more frequently when local conditions are conducive to a prolonged period of potential exposure to infected mosquito vectors. Annual revaccination is best completed in the spring (late February through early April) before the onset of the insect vector season. In areas in which the mosquito season is prolonged, revaccination twice annually, one time in the spring and again in the late summer or early fall (late July through early September), may be necessary to maximize protection.

Neither of the licensed vaccines carry label recommendations for administration to pregnant mares; therefore, it is recommended that mares be vaccinated before breeding whenever possible. It is, however, well recognized that pregnant mares are at risk of acquiring infection from infected mosquitoes. Consequently, it has become accepted practice by many veterinarians to administer vaccines to pregnant mares on the reasonable assumption that the risk of adverse consequences of WNV infection far exceeds the reported adverse effects of use of vaccines in pregnant mares. Thousands of doses of WNV Innovator [j] vaccine have been administered safely in pregnant mares. In addition, a recently published study failed to document vaccine-associated adverse effects in a large population of pregnant mares [28]. Although the Recombitek [f] vaccine is a live vectored vaccine, the canarypox vector is incapable of replication in mammals and does not induce a viremia that could infect a fetus. In addition, a canarypox-vectored influenza vaccine available in Europe has been licensed for use during pregnancy; thus, the vectored WNV vaccine is unlikely to induce adverse effects in pregnant mares. As with other vaccines, it is sound practice to avoid administering WNV vaccines to mares during the first 60 days of gestation unless conditions of imminent risk prevail.

Booster vaccination of previously primed pregnant mares (4 - 6 wks before foaling using either WNV Innovator [j] or Recombitek [f] vaccine) seems to induce a strong anamnestic serologic response that provides their foals with passive colostral protection lasting 3 - 4 mo [l]. In contrast, preliminary data suggest that a significant proportion of naïve pregnant mares failed to seroconvert when the primary series of WNV Innovator [j] vaccines were administered during the second half of gestation, perhaps reflecting pregnancy-associated down-regulation of Th2 responses [l]. If subsequently proven, this preliminary observation adds further justification to the recommendation that the primary series is best completed before breeding.

Preliminary data suggest that, unlike most other inactivated vaccines administered by IM injection, a three-dose series of WNV Innovator [j] vaccine is capable of inducing seroconversion in a high proportion of 3-month-old foals born to WNV antibody-positive mares [l]. Because the canarypox vector system used in Recombitek [f]. accomplishes transfection of cells and expression of the major E and M peptide antigens of WNV on the surface of antigen-presenting cells in association with MHC class I and class II antigens, it would be reasonable to assume that maternal antibodies would minimally affect cellular and humoral responses to this vaccine. Preliminary data suggest that Recombitek [f] is capable of inducing seroconversion in 3-month-old foals from WNV antibody-positive mares [l].

Pending availability of further data, current recommendations for vaccination of mares and foals against WNV are as follows:

1. Complete primary vaccination of mares before breeding whenever possible.
2. Revaccinate mares once or twice annually, depending on local conditions affecting duration of period of seasonal risk. Time administration of one booster dose to occur 4 - 6 wks before foaling.
3. Start primary vaccination of foals from seropositive vaccinated mares at 3 - 4 months of age using a three-dose primary series.
4. Primary vaccination of foals from non-vaccinated, non-exposed mares should commence at 3 - 4 months of age or younger (as early as 1 month of age) depending on month of birth and seasonal level of activity of mosquito vectors in the area. The three-dose primary vaccination protocol outlined above is recommended, pending acquisition of further data. If field challenge is expected before completion of the primary series, use of the Recombitek [f] vaccine may be advantageous, because this vaccine has been documented to induce protection within 26 days of administration of one dose of vaccine [27].

Tetanus

All horses are at risk for acquisition of tetanus; therefore, routine vaccination using one of the many licensed adjuvanted toxoid vaccines is recommended and has been a central component of immunization programs for horses for many years. Typically, a primary series comprising two doses of toxoid administered at 3- to 6-wk intervals is followed by boosters administered once annually. Tetanus toxoid is a potent antigen that rapidly induces strong serologic responses and protection as soon as 8 days after administration of the first dose [29]. A study conducted in Germany >40 yrs ago showed that the protection induced by three doses of toxoid lasted at least 8 yr and possibly for the lifetime of the horse [30]. As a result, tetanus is now rarely encountered as a clinical entity in horses. Tetanus toxoid is considered to be safe for administration to pregnant mares; therefore, annual revaccination of mares during the last 4 - 6 wks of gestation is recommended to maximize passive transfer of maternal antibodies.

Protection against tetanus seems to be mediated entirely by circulating antibodies, and these antibodies are transferred well through colostrum [31,32]. Thus, foals born to vaccinated mares seem to be at low risk of developing tetanus. Limited and conflicting information is available in the literature regarding the potential for maternal antibody interference with vaccination. Tetanus toxoid is considered to be a potent antigen, and it is widely accepted that concurrent administration of tetanus toxoid and antitoxin at different sites does not interfere with the response to tetanus toxoid [33]. Consequently, it was assumed that maternal antibodies were unlikely to interfere with the response of foals to tetanus toxoid, and it became common practice to initiate primary vaccination of foals against tetanus at 3 months of age. Jansen and Knoetze [34] documented that maternal antibodies inhibited the response of 10- to 18-wk-old foals to a single dose of tetanus toxoid in a water-in-oil emulsion, and Liu [35] recommended primary vaccination of foals against tetanus starting at 4 - 6 months of age. Recent studies employing highly sensitive ELISA tests to measure sub-isotypes of IgG antibodies directed against tetanus toxoid indicate that the serologic response of most 3-month-old foals from antibody-positive vaccinated mares was completely inhibited [10]. In addition, these studies showed that the inhibitory effect of maternal antibodies was still evident, albeit at a reduced level, when primary vaccination was delayed until foals were 6 months of age. Furthermore, levels of anti-tetanus antibodies induced by vaccination were substantially higher when a third dose of vaccine was added to the primary series, regardless of the age at which the primary series was initiated [10,22].

The following protocol is appropriate for tetanus vaccination:

1. Booster vaccinate the mare 4 - 8 wks before foaling.
2. Begin foal vaccination at 6 months of age.
3. Administer three doses of tetanus toxoid in the primary series (two doses administered 4 - 6 wks apart, followed by a third dose 8 - 12 wks later).
4. Booster vaccinate at 12-month intervals thereafter.

WEE, EEE, and VEE

The arthropod-borne diseases EEE and WEE are endemic in many areas of North America; therefore, vaccination of horses in these areas is strongly recommended. Routine vaccination of horses against VEE is not recommended at this time, because the disease has not been active in Mexico or North America for many years. All licensed encephalomyelitis vaccines are inactivated, adjuvated bivalent products containing WEE and EEE antigens [m,n,o] or trivalent containing WEE, EEE, and VEE [p]. After completion of a primary series comprised of two doses of vaccine administered 3 - 6 wks apart, revaccination of horses at intervals of 12 months or less is typically recommended, depending on the duration of the vector season and the specific virus that is the primary threat (WEE or EEE). Annual revaccination in the spring before the onset of the insect vector season seems to be effective in many areas, whereas revaccination at 4- to 6-month intervals may be necessary to achieve optimal protection in the Gulf states and Florida where the risk of arthropod transmission of EEE exists year round. Inactivated bivalent encephalomyelitis vaccines are considered to be safe for use in pregnant mares; therefore, booster vaccination 4 - 8 wks before foaling is routinely recommended.

Although correlates for protection against WEE and EEE are not well established, it is assumed that circulating antibodies are important based on the knowledge that infection is acquired by injection (mosquito bites) and current inactivated vaccines seem to be protective. Neutralizing antibodies to WEE and EEE are transferred passively to foals through colostrum and are detectable in the serum of many foals from vaccinated mares for at least 3 mo [6,11,36]. The half life of decline of colostrally derived antibodies against EEE and WEE have been estimated to be 33 and 20 days, respectively [6,11]. When foals from antibody-positive mares were vaccinated soon after birth with an inactivated EEE vaccine, revaccinated at monthly intervals through 6 months of age, and revaccinated again 13 - 19 months later, they failed to mount a detectable IgG response against EEE [17]. While the authors concluded that this failure to seroconvert indicated that vaccination soon after birth may have induced tolerance to EEE, the ability of these animals to survive virulent challenge and to mount an IgG response within 7 days of challenge suggests that true immunotolerance was not induced. Studies in older foals showed that although the serologic response to vaccination may be blocked by maternal antibodies, these foals were not rendered tolerant to doses of vaccine administered later in life [6]. Studies in our laboratory comparing the responses of 3-month-old foals with those of 6-month-old foals from antibody-positive mares showed that 3-month-old foals consistently failed to mount a serologic response to two doses of inactivated bivalent WEE/EEE vaccine; the majority had not responded even after administration of three doses [1]. Most 6-month-old foals failed to seroconvert after administration of two doses of vaccine; however, the majority responded after administration of a third dose [1].

Because EEE is a highly fatal disease and a year round risk in the Gulf states including Florida, foals in these areas are considered to be at risk for infection and death during their first year of life [6,17,37]. Therefore, vaccination of foals against EEE is recommended in these areas starting at 3 months of age [6]. The efficacy of vaccination programs to prevent EEE have, however, been brought into question by the finding of clinical EEE in vaccinated horses, particularly those <2 yrs of age [37]. This finding suggests that vaccine failure may result from interference by colostrally derived antibodies [6,11,17,36,37]. WEE has a lower mortality than EEE and occurs in midwestern and western states where the risk of infection is highest during the summer and early fall. In most areas, prevalence of WEE infection is low, so the risk of foals acquiring infection during their first year of life is generally low. For this reason, vaccination against WEE can be delayed until the spring of the yearling year for foals born in the late spring and summer months in most areas of the western United States, except in regions where a high prevalence exists.

Based on current knowledge, the following vaccination protocol for encephalomyelitis is an appropriate compromise:

1. Booster vaccinate the mare 4 - 6 weeks before foaling.
2. Begin foal vaccination at 6 months of age or during the spring of the yearling year using three doses in the primary series; booster vaccinate annually thereafter.
3. In the southeastern United States where the risk of EEE infection is high year round, start vaccinating foals at 3 - 4 months of age; use three or more doses in the primary series.

Equine Herpes Virus Type-1 and Equine Herpes Virus Type-4 Infection

EHV-1 and EHV-4 infect the host through the respiratory tract after inhalation or ingestion of infective aerosols or direct contact with infected horses or products of EHV-1 abortion. Viremia frequently occurs after infection with EHV-1, which can lead to myeloencephalopathy, abortion, or birth of infected non-viable foals. However, manifestations of infection with EHV-4 (rhinopneumonitis) are generally confined to the respiratory tract, because EHV-4 does not typically infect endothelial cells or produce a cell-associated viremia [38]. Horses frequently become infected with EHV-4 and EHV-1 during the first year of life and develop clinical signs of respiratory disease; however, they fail to clear the virus, resulting in a chronic carrier state for both viruses in the majority of the horse population [39,40]. Infection may recrudesce later in life as a clinically apparent or unapparent infection that results in an increase in serum neutralizing (SN) antibody titer. Consequently, many horses have detectable levels of SN antibody to both EHV-1 and EHV-4 in their serum [39,41].

Correlates for protection against EHV-1 and EHV-4 infection have been extensively investigated but are not yet clearly defined. Infection with EHV-1 induces a strong humoral response, but protection from reinfection is short lived and is not achieved until the horse has experienced multiple infections with homotypic virus [38]. No clear relationship exists between protection from EHV-1 infection and concentrations of circulating antibody induced by vaccination or infection, but the duration and amount of virus shedding from the nasopharynx is reduced in animals with high levels of circulating neutralizing antibody [38]. As with other herpes viruses, mucosal immunity and cell-mediated responses likely play a role as important as circulating neutralizing antibodies in protection against EHV-1 infection [42]. Evidence for the role of cell mediated immunity comes from studies showing that the presence of major histocompatibility complex (MHC) class 1 restricted cytotoxic T-lymphocyte precursors in peripheral blood are correlated with protection [38]. Because EHV-4 replication is largely confined to epithelial cells of the upper respiratory tract, it is likely that mucosal immunity is important in protection [38]. While circulating antibodies alone do not prevent EHV-4 infection, high levels of vaccine-induced circulating virus neutralizing antibody markedly reduce virus shedding and clinical signs after challenge infection [38].

The principal indication for use of EHV vaccines is prevention of EHV-1-induced abortion in pregnant mares. Consistent vaccination seems to reduce the frequency and severity of herpes virus-induced disease. Field experience suggests that while the incidence of sporadic EHV-1 induced abortions in individual mares has not changed, the incidence of abortion storms caused by EHV-1 has declined significantly since the introduction and widespread use of EHV-1 vaccines in the United States; however, convincing evidence is lacking [39,43]. Outbreaks of abortion and associated perinatal foal death do, however, continue to occur on occasion in herds of vaccinated mares. Of the vaccines currently licensed for use in pregnant mares in the United States, only inactivated monovalent EHV-1 vaccines [a,b] containing abortogenic strains of EHV-1 carry a label claim for preventing abortion, whereas at least one bivalent EHV-1/4 vaccine is licensed for prevention of abortion in Europe [q]. One of the vaccines available in North America [a] incorporates both the 1p and 1b subtypes of EHV-1 to reflect the documented increase in the proportion of EHV-1 abortions caused by the 1b subtype that occurred during the 1980s compared with earlier years [44]. Pregnant mares should be vaccinated during the fifth, seventh, and ninth months of gestation. Many veterinarians also recommend a dose during the third month of gestation, although documentation of added benefit of this approach is lacking. Similarly, vaccination of mares with an inactivated EHV-1/EHV-4 vaccine at the time of breeding and again 4 - 6 wks before foaling is commonly practiced to enhance concentrations of colostral immunoglobulin for transfer to the foal; however, there are no published reports to document the effectiveness of this approach in raising titers of specific antibody in mares that have been vaccinated against EHV-1 three times during the previous 5 months. Vaccination of barren mares and stallions with either a bivalent EHV-1/4 vaccine or a monovalent EHV-1 vaccine before the start of the breeding season and thereafter at 6-month intervals is recommended with the goal of increasing herd immunity in an attempt to reduce viral shedding and challenge to pregnant mares on breeding farms [39].

A modified live virus EHV-1 vaccine [e] has been used as an aid to prevention of EHV-1 abortion by some practitioners for many years, but this vaccine is not currently labeled for this use. However, several recent developments have created a renewed interest in the potential for use of modified live virus (MLV) vaccines for protecting horses against manifestations of EHV-1 and EHV-4 infection. Sequencing of the EHV-1 genome has made it possible to document the nature of the mutation encoding for attenuation, mediated through truncation of the gp2 glycoprotein, of the KyA strain [45]. Similar studies may soon yield information regarding the mutation underlying attenuation of the RAC-H strain from which Rhinomune [e] was derived. In addition, studies in Europe have documented the efficacy of an intranasally administered temperature-sensitive live EHV-1 vaccine in preventing abortion, neurologic disease, and respiratory disease after experimental challenge with virulent EHV-1 [42,46]. This renewed interest in modified live vaccines as well ongoing investigation of recombinant vaccines expressing the major EHV glycoprotein antigens will likely lead to improved approaches to immunoprophylaxis in the near future.

Because currently available inactivated vaccines do not block infection with EHVs, the most we can hope for when using inactivated vaccines is reduction of severity of clinical signs and attenuation of virus shedding to help protect herd mates. Challenge studies in weanlings aged 5 - 8 months have clearly shown the efficacy of an inactivated whole virus EHV-1/4 vaccine in significantly reducing clinical manifestations and virus shedding induced by virulent EHV-1 challenge administered 2 wks after completion of the two-dose primary series [47]. Efficacy was clearly correlated with vaccine-induced antibody levels at the time of challenge in this study [47]. Studies investigating the serologic response of 3-month-old and 5-month-old foals to this same vaccine documented failure of a high proportion of foals to seroconvert, suggesting that this vaccine is likely to be less effective in protecting younger foals than in protecting the foals aged 5 - 8 months cited in the above challenge study [21].

Specific antibodies against both EHV-1 and EHV-4 are passed in colostrum [12,19-21,48]. Field studies with modified live EHV-1 vaccines indicate that colostral antibodies exert a profound inhibitory effect on serologic responses to vaccination up to at least 5 months of age [19,49,50]. However, a cytotoxic cellular immune response to both EHV-1 and EHV-4 was induced in a substantial percentage of foals vaccinated with a modified live EHV-1 vaccine in the presence of maternal antibody, although humoral responses were often absent [51]. Recent studies with two different commercially available inactivated bivalent EHV-1/4 vaccines and one inactivated EHV-4/influenza vaccine have shown that the majority of foals from EHV-vaccinated mares do not mount a detectable neutralizing antibody response to vaccines administered at 3 and 4 months of age, even when three doses are administered in the primary series [1,20,21]. An increased proportion of foals responded when vaccinated with a three-dose series starting at 5 - 6 months of age, but a substantial number still failed to seroconvert [1,21]. Some foals with low or undetectable levels of SN antibody at the time of vaccination failed to mount a serologic response, suggesting that low levels of antibody, below the lower limit of detection of the SN test based on EHV-1 antigen, are capable of inhibiting the serologic response to inactivated EHV-1/4 vaccines [21]. The failure of a large proportion of foals <6 months of age to mount serologic responses to inactivated EHV-1/4 vaccines and the influence of antibody titer at the time of vaccination on failure to respond has been confirmed using sensitive gD and gG ELISAs in studies on commercial stud farms in Australia [52]. In parallel studies, these researchers concluded that mares were the source of infection for foals and that intensive use of inactivated EHV-1/4 vaccines on breeding farms in Australia had minimally impacted the infection rate of young foals and weanlings with EHV-1 and EHV-4 [40,53].

The obvious dilemma in designing a vaccination strategy to prevent EHV-1 and EHV-4 infection in foals and weanlings is that if primary immunization is delayed until 6 months of age or older when maternal antibody titers have declined to low levels and are less likely to interfere with vaccination, then the foals are likely to encounter field infection before completion of the three-dose primary series. Thus, it is unreasonable to expect a high degree of efficacy for vaccination programs designed to protect foals and weanlings against EHV infection using available vaccines.

Under circumstances where vaccination of foals against EHV-1 and EHV-4 is elected, the following compromise program may be used:

1. Vaccinate the mare during the fifth, seventh, and ninth months of gestation using an inactivated EHV-1 vaccine that is licensed as an aid to prevention of abortion.
2. Booster vaccinate the mare 4 - 6 wks before foaling with an inactivated bivalent EHV-1/4 vaccine.
3. Start foal vaccination at 4 - 6 months of age using two doses of an inactivated bivalent vaccine or a modified live EHV-1 vaccine administered 3 - 4 wks apart; administer a third dose 8 - 12 wks later.
4. Revaccinate at 4 - 6 month intervals thereafter using either an inactivated bivalent vaccine or a modified live EHV-1 vaccine.

To be completely effective in blocking primary infection and establishment of a lifelong carrier state with EHV-1 and EHV-4, future vaccination strategies should be directed at inducing a strong mucosal immune response in the upper respiratory tract during the first few weeks of life at a time when high levels of maternal antibodies are present. Promising progress toward this goal was reported recently by Patel et al. [54] who documented that intranasal administration of a single dose of temperature-sensitive modified live EHV-1 vaccine to maternal antibody-positive foals aged 1.4 - 3.5 months afforded partial but significant protection against febrile respiratory disease, viremia, and virus shedding after intranasal challenge with virulent EHV-1 performed 8 wks after vaccination.

Influenza

Infection with the orthomyxovirus influenza A-equine-2 is a common cause of rapidly spreading outbreaks of respiratory disease in horses in North America and Europe. Although horses of all ages are susceptible, young performance and show horses seem to be at the greatest risk of acquiring infection, because they have a high likelihood of contact with other horses and are often encountering the virus for the first time. While the high "traffic" in horses from different sources and the high proportion of young susceptible animals on breeding farms would be expected to contribute to a high risk for acquisition of influenza infection, documented outbreaks of influenza in foals residing on breeding farms on which adults are regularly vaccinated against influenza are uncommon [55]. Therefore, there seems to be no reason to vaccinate young foals from vaccinated mares as was recommended in the past [35,55,56].

Immunity after natural infection with influenza persists for >1 yr and is not dependent on high levels of circulating antibody [38,57-59]. In addition to invoking a strong serologic response, natural infection induces large amounts of virus-specific secretory virus neutralizing IgA in nasal secretions and genetically restricted cytotoxic T lymphocytes (CTL) that kill infected cells [58,60]. Memory CTL can be detected in the peripheral blood for at least 6 months after infection [60]. In contrast, immunity after vaccination with conventional parenterally administered inactivated vaccines is short lived and is highly correlated with levels of circulating antibody directed against surface hemagglutinin antigens [58,59,61]. With the possible exception of immune-stimulating complex vaccines, inactivated vaccines administered by IM injection have limited potential to induce CTL responses or nasal secretory IgA responses [58,59,62].

Challenge studies have documented that the cold-adapted modified live influenza A-equine-2 vaccine [i] licensed for intranasal administration induces protection lasting 6 months or longer [63,64]. As with natural infection, protection induced by this vaccine does not depend on stimulation of high levels of circulating antibody [63,64]. Although published data regarding the safety of Flu-Avert I.N. [i] in pregnant mares are not available, the cold-adapted strain of influenza virus contained in the vaccine does not replicate at core body temperatures; thus, viremia and dissemination to the gravid uterus are highly unlikely [64]. Many practitioners, therefore, administer this vaccine at 6- to 12-month intervals as part of their core vaccination program for broodmares and other horses on breeding farms. Because the intranasally administered vaccine does not induce high levels of circulating antibody, potent inactivated injectable influenza vaccines should be used to booster vaccinate pregnant mares before foaling.

Several studies published over the last 15 yrs have documented that antibodies against influenza A-equine-1 and A-equine-2 viruses are passively transferred to foals through colostrum [8-10,14,16,56]. While Liu et al. [56] showed a rapid decline in levels of maternal antibodies, which resulted in many foals being seronegative for influenza antibodies by 4 wks of age, other workers have shown that these antibodies decline more slowly with a half life of ~30 - 38 days and that they are detectable in some foals beyond 6 months of age [8-10,14-16]. These workers also showed that the majority of foals from vaccinated mares did not seroconvert when vaccinated with inactivated adjuvanted whole virus or subunit influenza vaccines administered at 3 months of age, whereas a higher proportion of 6-month-old foals responded when given three doses of influenza vaccine in the primary series [8-10,14-16]. In a study to investigate the finding that many yearling horses had low or undetectable levels of HI antibody despite having received multiple doses of inactivated influenza vaccine during the first year of life, Cullinane et al. [14] found that a substantial number of foals vaccinated at 3 months of age not only failed to respond serologically to the subunit vaccine used in the primary series but also failed to respond to four or more additional doses of either an inactivated subunit or whole virus vaccine administered over the next year. This result suggested that early vaccination in the presence of maternal antibody had induced immunotolerance to influenza vaccines, as determined by lack of serologic responses [14]. Similar results were obtained in our laboratory when the responses of 3-month-old antibody-positive foals to inactivated whole virus influenza vaccines were assessed using the HI test. However, when the same samples were retested using a sensitive ELISA assay that detects sub-isotypes of IgG, evidence of induction of tolerance was not found. Instead, misdirection of the immune response in favor of IgGT rather than the IgGa and IgGb sub-isotypes believed to be important for protection was documented [10]. In the same study, we found that while the response of 6-month-old foals from seropositive mares was superior to that of 3-month-old foals (in terms of both the percent of foals seroconverting and the magnitude of the resulting antibody titers), the response of yearlings was clearly superior to that of 6-month-old foals [1]. While 60% of the yearlings seroconverted to influenza A-equine-2 after two doses of vaccine, all seroconverted after three doses and >50% developed HI titers of >1:1000 [1]. Based on these data, it was concluded that titers will likely persist at a protective level for a much longer duration after a three-dose primary series than after a two-dose series. This conclusion is also supported by results of studies in Europe to the extent that administration of a third dose, 2 - 4 months after the second dose, is now strongly recommended [1,8-10,14,59]. Because the inhibitory effects of maternal antibodies on responses to inactivated influenza vaccines may persist for up to 9 months in foals born to mares with high titers, primary vaccination of foals from vaccinated mares should be dela The MLV intranasal vaccine [i] is licensed for use in horses 11 months of age or older. While this vaccine seems to be safe for use in foals as young as 2 months of age [65], published data regarding the potential for maternal antibody interference are lacking. Preliminary studies suggest that maternal antibodies may interfere with the response of foals aged between 3 and 6 months, whereas foals with maternal antibody vaccinated at 7 months of age were protected against virulent challenge [66,67]. Pending publication of well-controlled studies, it is recommended that if the first dose of Flu-Avert I.N. [i] vaccine is administered before 11 months of age, a second dose should be administered at 11 months of age or older [67]. A live canarypox-vectored recombinant vaccine expressing the hemagglutinin genes of equine influenza viruses [r] is available in Europe and is approved for use in pregnant mares and foals as young as 4 months of age [59]. Although there are no published reports regarding the influence of maternal antibodies on responses to this vaccine, the recommended minimum age for vaccination of foals from immunized mares is 5 months.

Current recommendations for vaccination against influenza using vaccines licensed in North America are as follows:

1. Maintain all horses on breeding farms on a regular program of influenza vaccination using either MLV intranasal vaccine at 6 - 12-month intervals or an inactivated injectable vaccine at 4 - 6-month intervals.
2. For pregnant mares, time administration of one of the boosters to occur 4 - 8 wks before foaling; use an inactivated vaccine.
3. If MLV intranasal vaccine is used in the immunization program, administer the first dose at 11 months of age, administer a second dose 3 - 6 months later, and continue at 6-month intervals thereafter. If the first dose is administered before 11 months of age, revaccinate at 11 months of age or older and continue as described.
4. If inactivated vaccines are used, begin foal vaccination at 6 - 12 months of age using three or more doses in the primary series. Administer the first two doses 3 - 6 wks apart and wait another 8 - 12 wks to administer the third dose; revaccinate at 4- to 6-month intervals thereafter if risk continues.

Vaccination of foals born to seronegative, non-vaccinated mares can commence at 3 months of age or earlier if significant risk of exposure to influenza exists.

Strangles

Infection of horses with Steptococcus equi subsp. equi continues to cause troublesome outbreaks of strangles throughout North America, despite the availability and widespread use of vaccines; this indicates that the efficacy of strangles vaccines is suboptimal [68]. Protection against S. equi infection seems to be mediated by a combination of mucosal IgG and IgA antibodies produced locally in the nasopharynx and opsonic IgG antibodies in serum [69-72]. Licensed strangles vaccines include inactivated, adjuvanted M-protein cell wall extracts [s,t]. These vaccines induce a good opsonophagocytic antibody response in serum but a minimal mucosal response, which likely accounts for the incomplete protection observed when they are used in the field [72,73]. However, data do exist to document that vaccination using injectable M protein vaccines significantly reduces the attack rate and severity of strangles in herds with endemic infection [73-75]. An attenuated live vaccinevac_eq_strangles [h] for intranasal administration was introduced onto the market several years ago and is now in widespread use. Vaccines of this type have been shown to induce a relevant mucosal immune response and partial or complete protection, but they may do so without inducing a strong serologic response [68,76].

Because young horses are typically more susceptible to strangles than older horses and the disease is spread by both direct and indirect contact with symptomatic horses or asymptomatic carriers, strangles is common on breeding farms. Broodmares and foals are, therefore, the primary target of preventive strategies, including vaccination. Despite the incomplete efficacy of strangles vaccines, vaccination is indicated for horses likely to experience a substantial risk of exposure, such as those being introduced to or born onto premises where strangles is endemic. After completion of a two- or three-dose primary series, revaccination of horses at 6-month intervals using either the M-protein or intranasal vaccine is typically recommended. Because injectable M-protein vaccines induce better serologic responses than do MLV intranasal vaccines, administration of this type of vaccine as one of the semi-annual boosters to pregnant mares 4 - 8 wks before foaling is recommended to maximize passive protection of the foal.

Antibodies of the IgG and IgA class recognizing the M-protein of S. equi are passively transferred to the foal through colostrum and are also present in the milk of immune mares [5]. Antibodies of predominantly the IgGb isotype are absorbed from colostrum and redistributed to the nasopharyngeal mucosa [4]. These IgGb antibodies, along with the SeM-specific IgA antibodies that are present in milk and passively coat the pharyngeal mucosa of nursing foals, provide protection to most nursing foals up to the time of weaning [4,5,68,69]. Serologic (ELISA) responses to M-protein vaccines are poor in foals, most likely because of the inhibitory effect of maternal antibodies. The modified live intranasally administered vaccine may be less subject to interference by circulating maternal antibodies; however, this issue has not been investigated, and the manufacturers do not recommend administration of this vaccine to horses <9 months of age. Considering that on farms where strangles is endemic, foals often become infected around the time of weaning at 4 - 8 months of age, it is difficult to protect them if vaccination is delayed until 9 months of age.

The following vaccination protocol seems to be a reasonable compromise on breeding farms on which the risk of strangles infection is high and mares are on a regular vaccination program:

1. Booster vaccinate the mare 4 - 8 wks before foaling using an inactivated M-protein vaccine.
2. Begin foal vaccination using the intranasal live vaccine at 4 - 9 months of age; use the recommended two-dose primary series followed by boosters at 6- to 12-month intervals.
3. Alternatively, begin foal vaccination at 4 - 6 months of age with an intramuscularly administered M-protein vaccine; use at least three doses in the primary series and booster vaccinate at 6-month intervals.

Rabies

Most horses in North America are at risk for exposure to rabies virus, because the disease is endemic in wildlife reservoirs in many regions. Although rabies is rare in horses, the disease is invariably fatal, and every case carries with it substantial public health implications. Therefore, vaccination of horses against rabies is recommended. The rabies vaccines currently licensed for use in horses [u,v,w] are inactivated, tissue culture-derived products that induce strong serologic responses after a single dose [a]. Although correlates for protection against infection with rabies virus in horses are not well defined, it is logical to assume that protection would correlate with titers of circulating antibody, because infection is usually acquired by systemic injection through bites by rabid animals. In humans, post-vaccination antibody titers are used to predict protection and to assess the need for post-exposure vaccination or administration of immune serum. In dogs, however, post-vaccination serologic test results were not found to be completely predictive of resistance to challenge exposure during tests performed with certain inactivated vaccines [31]. Published results of challenge studies assessing efficacy of rabies vaccines licensed for use in horses in North America are not available.

Label directions on inactivated rabies vaccines licensed for use in horses [u,v,w] suggest administration to foals aged 3 months or older using one dose of vaccine in the primary series followed by a second dose at 1 yr of age. Thereafter, annual revaccination is recommended. None of the licensed vaccines carries a specific label approval for use in pregnant mares; therefore, it is recommended that mares be revaccinated before breeding whenever possible. Because rabies antibodies persist in serum for a prolonged period, foals born to mares that are revaccinated while open acquire substantial titers of rabies antibody after ingesting colostrum [l].

Documentation of rabies in reportedly vaccinated horses, most of which were <2 yr of age, has brought into question the efficacy of label recommendations for primary vaccination of foals against rabies [77]. Recent studies in our laboratory have shown that the serologic response of most 3-month-old foals from antibody-positive mares is completely blocked, even when a two-dose primary vaccination series is used. Although the response to the first dose of vaccine is typically blocked in 6-month-old foals from antibody-positive mares, these foals seem to seroconvert after administration of a second dose administered 4 wks later [l].

Pending publication of results of further studies, the following approach is recommended for the use of inactivated rabies vaccines in mares and foals in rabies-endemic areas:

1. Vaccinate mares before breeding.
2. Begin the primary vaccination series for foals from vaccinated mares at 6 months of age or older using two doses administered ~4 wks apart; for foals from non-vaccinated mares, the primary vaccination series can be started as early as 3 months of age and may comprise only one dose, although a two-dose series will likely induce more durable protection.
3. Booster vaccinate as yearlings and then annually thereafter.

Rotavirus Infection

Equine rotavirus is one of the most important causes of infectious diarrhea in foals during the first few weeks of life and often causes outbreaks involving the majority of the foal crop on individual farms [78-80]. Older foals and adult horses are more resistant to infection. Rotaviruses are classified into seven serogroups (A through E) based on common antigens in each group [80]. Until recently, all equine rotavirus isolates were classified as belonging to serogroup A, which includes 14 serotypes (G1 through G14); of these, five (G3, G5, G10, G13, and G14) representing four genotypes (P1, P7, P12, and P18) have been identified and characterized in horses [81-83]. Most equine rotavirus isolates are of the P12 genotype and G3 serotype (previously referred to as H-2) and include two subtypes (subtype 1 and 2) [84].

An inactivated rotavirus A vaccine [d] containing the G3 (H-2) serotype in a metabolizable oil-in-water emulsion is conditionally licensed for use in many states in the United States. The primary indication for this vaccine is administration to pregnant mares on endemic farms as an aid to preventing diarrhea in their foals caused by infection with rotaviruses of serogroup A. Label recommendations call for a three-dose series of the vaccine to be administered during the eighth, ninth, and tenth months of each pregnancy. This protocol has been shown to induce a significant increase in serum concentrations of neutralizing antibody in vaccinated mares [85] and to significantly increase concentration of antibodies of the IgG, but not IgA, subclass in the colostrum and milk of vaccinated mares [86]. The concentration of the passively derived rotavirus-specific antibody of the IgG subclass in the serum of foals (up to 90 days of age) from vaccinated mares after nursing is significantly higher than that measured in serum of foals born to non-vaccinated mares [85,86]. A field study showed this vaccine to be safe and provided circumstantial evidence of at least partial efficacy. An approximate two-fold higher incidence of rotaviral diarrhea was found in foals from non-vaccinated mares compared with those from vaccinated mares, although this difference did not prove to be statistically significant [85]. Similarly, a controlled field study in Argentina in which an inactivated aluminum hydroxide-adjuvanted vaccine containing the G3P2 (SA11), G3P12 (H2), and G6P1(Lincoln) strains was administered to 100 mares at 60 days and again at 30 days before foaling showed a substantial reduction in the incidence and severity of rotaviral disease in foals from vaccinated mares compared with foals from non-vaccinated mares [87]. Challenge studies involving two inactivated rotavirus vaccines administered in a similar manner to pregnant mares in Japan showed that their foals were not completely protected against infection but had a substantial reduction in severity of clinical signs after challenge [82].

The major correlate for protection against rotaviral infection seems to be mucosal immunity, predominantly mucosal IgA, in the gastrointestinal tract. Studies of the immunoglobulin isotype responses of mares and of antibodies passively transferred to their foals after parenteral vaccination of the mares with inactivated rotavirus vaccines indicate that this approach is unlikely to provide foals with intestinal mucosal protection in the form of IgA [86]. Consequently, it is not surprising that current protocols do not provide complete protection. In addition, because the conditionally licensed vaccine available in the United States contains only the G3 serotype of the A serogroup, it cannot be expected to protect against infection with all field strains.

The recommended program for the use of this vaccine is as follows:

1. Vaccinate the mare during the eighth, ninth, and tenth months of gestation.
2. Repeat the three-dose vaccination series during each subsequent pregnancy.

The potential for serologic responses to primary vaccination with the rotavirus vaccine to be suppressed during pregnancy should be investigated to determine whether completion of the primary series before breeding might be advantageous.

Toxicoinfectious Botulism - "Shaker Foal Syndrome"

Of the eight distinct toxins produced by subtypes of Clostridium botulinum, types B and C are associated with the majority of botulism outbreaks; virtually all cases of toxicoinfectious botulism ("shaker foal syndrome") are caused by type B [88-90]. Shaker foal syndrome is an important problem affecting foals between 2 wks and 8 months of age in Kentucky and the mid-Atlantic states; it also occurs sporadically in other areas [88-90]. A C. botulinum type B toxoid vaccine [c] is available in North America with the primary indication being the prevention of shaker foal syndrome in endemic areas [91]. A similar toxoid is available to protect foals in endemic areas in Australia [92]. Antibodies against toxin type B do not cross protect against other toxin types. Protection seems to be mediated primarily by circulating antibodies, because a program of mare vaccination has been shown to offer protection to foals that achieve good passive transfer of colostral antibodies; this protects the foal for 8 - 12 wks [88,91]. Maternal antibodies do not seem to interfere with the response of foals to primary vaccination against botulism [93]. Therefore, a primary series comprising three doses of vaccine can be initiated at 2 - 3 months of age or earlier.

The recommended program for the use of BotVax B [c] is as follows:

1. Vaccinate the mare during the eighth, ninth, and tenth months of gestation.
2. Vaccinate the mare 4 - 6 wks before foaling in subsequent pregnancies.
3. Vaccinate the foal beginning at 2 - 3 months of age if the risk of infection continues.

Potomac Horse Fever

Recent documentation of the involvement of operculate freshwater snails and aquatic insects such as caddisflies and mayflies in the life cycle of Neoricketsia (formerly Ehrlichia) risticii has permitted formulation of focused control measures directed at minimizing exposure of horses to the habitats occupied by these species during the summer and fall months when disease risk is highest in endemic areas [94]. Risk reduction is best accomplished by denying horses access to river banks, creek beds, and irrigation ditches as well as pastures that have recently been flooded or flood irrigated.

Pregnant mares and foals are at risk for becoming infected with N. risticii and developing fulminant clinical PHF, which may prove fatal or result in abortion of an infected fetus [95]. Recovery after natural infection with N. risticii induces a strong antibody response and durable protection from reinfection lasting 20 months or longer [96]. However, presence of antibodies does not necessarily correlate with protection, and it is likely that cell-mediated responses play a crucial role. A β-propiolactone inactivated host cell-free N. risticii vaccine protects mice against homologous challenge. Several inactivated PHF vaccines for IM administration [x,y,z,aa,bb] are licensed for use in horses with the label claim that they aid in prevention of equine monocytic ehrlichiosis. In a series of studies in which ponies were challenged intravenously with N. risticii ~4 wks after completion of the two-dose primary vaccination series using a formalin-inactivated, aluminum hydroxide adjuvanted vaccine [cc]. Ristic et al. [97] reported that 78% of experimentally infected ponies were protected against all clinical manifestations of disease except fever and 33% were protected against all signs, including fever. A published non-controlled field study involving the same vaccine documented induction of serologic responses in most vaccinated horses and a substantial reduction in disease prevalence, morbidity, and mortality compared with data collected in a previous year when horses were not vaccinated [95,98]. In contrast to the results of the studies cited above, an epidemiological investigation involving a large number of horses failed to show any clinical or economic benefit from annual vaccination with currently available vaccines in the state of New York [99,100]. Failure of a substantial number of individual horses to mount an immune response to inactivated PHF vaccines, heterogeneity of N. risticii isolates, and much more rapid waning of protective immunity after vaccination than after natural infection likely account for vaccine failure [95,101]. Despite the above considerations, many practitioners and horse owners feel that available vaccines do induce at least partial protection when administered at 4- to 6-month intervals with administration of one booster being timed to precede the anticipated period of peak challenge.

A study of antibody-positive mares and their foals on a farm in Maryland showed that antibodies to N. risticii were passed through the colostrum and persisted in the serum of ~33% of foals for up to 20 wks, whereas 67% of foals were antibody negative by 12 wks of age [102]. On the basis of these findings and the apparent susceptibility to infection of two foals vaccinated earlier than 12 wks of age, the authors recommended that vaccination of foals from antibody-positive mares should begin with a two-dose primary series starting at 3 - 5 months of age followed by administration of one subsequent booster dose within the next few months [98]. However, the efficacy of this recommendation requires further study. Vaccination of foals in endemic areas is further complicated by the distinct seasonal incidence of disease in July, August, and September, a time when the majority of foals are aged between 3 and 6 months.

Vaccination of foals against PHF is only indicated in endemic areas, and the efficacy of vaccination protocols for foals has not been proven. Pending results of further studies, this protocol is generally followed:

1. Booster vaccinate the mare 4 - 8 wks before foaling.
2. Begin foal vaccination at 3 - 5 months of age or older using a three-dose primary series.
3. Booster vaccinate at 4- to 6-month intervals thereafter.

Equine Viral Arteritis

EVA is endemic in most populations of Standardbred horses, but clinical manifestations of infection are rare despite high seroprevalence [2,103]. Seroprevalence of infection in Thoroughbreds and Quarter horses is low, although outbreaks of both the respiratory and venereal forms of infection have been documented in Thoroughbred horses during the last 25 yrs [2,104]. Although horizontal transmission of the EVA virus during respiratory disease is known to be important in the propagation of outbreaks, venereal transmission from asymptomatic carrier stallions that shed virus in semen is the most important means of transmission that maintains the virus in certain populations of horses [2,105]. EVA can be transmitted by natural cover or by artificial insemination using fresh, chilled, or frozen semen. It seems that a substantial number of Warmblood and sport horse stallions imported from Europe are chronic unapparent shedders of this virus in their semen [104]. While all asymptomatically infected stallions are seropositive (i.e., have circulating neutralizing antibody), carrier stallions are often not identified at the time of importation, because testing for EVA antibodies is not required for importation [104]. These stallions have been the source of several localized outbreaks of EVA on breeding farms or boarding facilities in recent years [106]. Although mares do not become latently infected, exposure of pregnant mares to infected horses during outbreaks by direct or indirect contact may lead to embryonic loss, abortion, neonatal death, and further dissemination of the virus [2,106].

Establishment of the carrier state seems to be dependent on the high levels of androgens circulating in intact stallions and can be prevented by vaccinating colts before they are used for breeding or, preferably, before puberty [2]. Transmission to mares can be prevented by vaccination before breeding to a carrier stallion. The modified live virus vaccine [k] available for use in the United States has been shown to induce good protection against infection and the development of clinical signs.

Neutralizing antibodies to EVA are passively transferred through colostrum to foals and decline with a mean half life of ~32 days [7]. In one study, all foals from seropositive mares were found to be negative for antibodies to EVA by 8 months of age, suggesting that this would be an appropriate age to begin primary immunization [7]. Vaccinated horses can be expected to become seropositive for life, which may complicate export to countries that require serologic testing before importation. Maintenance of accurate vaccination records is, therefore, essential, and documentation of seronegative status before vaccination may be necessary to satisfy the importation regulations of some countries.

Although vaccination against EVA is occasionally recommended as an aid to prevention of horizontally transmitted infection during outbreaks, vaccination is typically reserved for horses on breeding farms and other facilities on which the risk of infection is high and for mares being bred naturally or by artificial insemination to carrier stallions, as follows:

1. Vaccinate the mare before breeding using the modified live vaccine.
2. Begin foal vaccination at 8 - 10 months of age with one dose of vaccine followed by annual boosters thereafter.

Development and marketing of a marker vaccine that allows vaccinated horses to be distinguished from inapparently infected carriers would greatly facilitate control, and even eradication, of EVA in horse populations.

Footnotes

1. Pneumabort-K +1b, Fort Dodge Animal Health, Overland Park, KS 66210.
2. Prodigy, Intervet, Millsboro, DE 19966.
3. BotVax B, Neogen, Lansing, MI 48912.
4. Equine Rotavirus Vaccine, Fort Dodge Animal Health, Overland Park, KS 66210.
5. Rhinomune, Pfizer, New York, NY 10017.
6. Recombitek, Merial Ltd., Duluth, GA 30096.
7. Calvenza EIV, EHV, and EIV/EHV, Boehringer Ingelheim GmbH, 55216 Ingelheim, Germany.
8. Pinnacle I.N., Fort Dodge Animal Health, Overland Park, KS 66210.
9. Flu-Avert I.N., Intervet, Millsboro, DE 19966.
10. West Nile Innovator, Fort Dodge Animal Health, Overland Park, KS 66210.
11. Arvac, Fort Dodge Animal Health, Overland Park, KS 66210.
12. Wilson WD, Mihalyi J. Unpublished data, 2005.
13. Encevac with Havlogen, Intervet, Millsboro, DE 19966.
14. Encephaloid Innovator, Fort Dodge Animal Health, Overland Park, KS 66210.
15. Cephalovac EW, Boehringer Ingelheim GmbH, 55216 Ingelheim, Germany.
16. Cephalovac VEW, Boehringer Ingelheim GmbH, 55216 Ingelheim, Germany.
17. Duvaxyn EHV-1/4, Intervet, Millsboro, DE 19966.
18. PROTEQ-FLU, Merial Ltd., Duluth, GA 30096.
19. Strepvax II, Boehringer Ingelheim GmbH, 55216 Ingelheim, Germany.
20. Srepguard with Havlogen, Intervet, Millsboro, DE 19966.
21. Rabvac 3, Fort Dodge Animal Health, Overland Park, KS 66210.
22. RM Imrab 3, Merial Ltd. , Duluth, GA 30096.
23. Rabguard TC, Pfizer, New York, NY 10017.
24. Mystique, Intervet, Millsboro, DE 19966.
25. Potomavac, Merial Ltd., Duluth, GA 30096.
26. PotomacGuard, Fort Dodge Animal Health, Overland Park, KS 66210.
27. PHF-Gard, Pfizer, Inc., New York, NY 10017.
28. Equovum PHF, Boehringer Ingelheim GmbH, 55216 Ingelheim, Germany.
29. PHF-Vax, Schering-Plough Corp., Union, NJ 07083.