Pro and contra IBR-eradication

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1 Veterinary Microbiology 113 (2006) Pro and contra IBR-eradication Mathias Ackermann *, Monika Engels Institute of Virology, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse 266a, CH-8057 Zurich, Switzerland Abstract Bovine herpesvirus type 1 (BoHV-1) is the causative agent of respiratory and genital tract infections such as infectious rhinotracheitis (IBR), infectious pustular vulvovaginitis (IPV, balanoposthitis (IBP), and abortion. Despite of a pronounced immune response, the virus is never eliminated from an infected host but establishes life-long latency and may be reactivated at intervals. Europe has a long history of fighting against BoHV-1 infections, yet, only a small number of countries has achieved IBR-eradication. Therefore, it seemed appropriate to review the reasoning pro and contra such a task. Clearly, the goal can indeed be achieved as has been demonstrated by a number of European countries. However, detection and stamping out of seemingly healthy virus carriers is inevitable in the process. Unfortunately, the use of vaccines is only of temporary and limited value. Therefore, there are numerous considerations to be put forward against such plans, including the high costs, the great risks, and the unsatisfactory quality of tools. If either control or eradication of IBR is nonetheless a goal, then better vaccines are needed as well as better companion tests. Moreover, better tools for the characterization of viral isolates are required. Collaborative actions to gather viral strains from as many countries as possible for inclusion into a newly created clustering library would be most advantageous. # 2005 Elsevier B.V. All rights reserved. Keywords: Bovine herpesvirus 1; IBR/IPV; Eradication; Review 1. Introduction The bovine herpesvirus type 1 (BoHV-1) belongs to the subfamily Alphaherpesvirinae and is an important pathogen of cattle (Wyler et al., 1989). It is the causative agent of respiratory and genital tract infections such as infectious rhinotracheitis (IBR), infectious pustular vulvovaginitis (IPV, balanopothitis (IBP), and abortion. Due to breeding synchronization * Corresponding author. Tel.: ; fax: address: @vetvir.unizh.ch (M. Ackermann). in cattle, BoHV-1 may also cause abortion storms. Rarely, the virus has been associated with central nervous disorders as well as with death of newborn calves (Metzler et al., 1986; Wyler et al., 1989). Upon primary infection, BoHV-1 replicates in the periphery, i.e. the mucous membranes of either the respiratory or the genital tract. From there, it will gain access to local sensory neurons for establishment of latency in the corresponding ganglia (Ackermann et al., 1982; Ackermann and Wyler, 1984). Furthermore, the virus may spread locally to the deeper respiratory tract, causing IBR, or systemically, which may ultimately lead to abortion as a consequence of infection of the /$ see front matter # 2005 Elsevier B.V. All rights reserved. doi: /j.vetmic

2 294 M. Ackermann, M. Engels / Veterinary Microbiology 113 (2006) fetus. Immunity against BoHV-1, either natural or induced following vaccination, has been reported to protect from clinical disease as well as from the negative consequences attributed to systemic spread of the virus (Wyler et al., 1989; Babiuk et al., 1996; Bosch et al., 1996; Castrucci et al., 2002a,b; Dispas et al., 2003). However, despite of a pronounced immune response, the virus is not eliminated from the infected host upon recovery but establishes life-long latency in the sensory ganglia, from where it may be reactivated at intervals (Wyler et al., 1989; Van Drunen Littel-van den Hurk et al., 1997; Bosch et al., 1998; Castrucci et al., 2002a,b). Therefore, eradication of IBR will always include the destruction of a great number of healthy, seropositive animals, which are considered to represent the virus reservoir because they are persistently infected with BoHV-1. Europe has a long history of fighting against BoHV-1 infections (Ackermann et al., 1990a,b). However, only a small number of countries has achieved the goal of IBR-eradication (Table 1). Therefore, it seems appropriate to review the reasoning pro and contra IBR-eradication. 2. Prevalence and IBR-eradication IBR occurs on all continents, although there are differences in prevalence and incidence. One report from Algeria indicated a seroprevalence of about 20% (Achour and Moussa, 1996). In the USA, Canada, Australia and New Zealand the seroprevalence of BoHV-1 infection is variable but may be very high (E. Breidenbach, Federal Vet. Office, pers. comm.). In most of these countries vaccination is widely used on an individual basis, which may help to decrease the economical losses due to clinical disease but also contributes to the high seroprevalence (e.g. Van Drunen Littel-van den Hurk et al., 1997). Cerqueira et al. (2000) reported a seroprevalence of 56% in the Bahia state, Brazil. In other countries of the Southern Americas the situation may be similar, but also more severe due to the simultaneous presence of BoHV-5 (Metzler et al., 1986; Esteves et al., 2003). According to OIE, 1461 cases (23 outbreaks) were registered in Russia for the year Almost no data are available from the Far East. In some countries, the prevalence of IBR may be comparable to other continents. A few Far Eastern countries did never report IBR cases, i.e. the Philippines, Singapore, Sri Lanka, and Vietnam. However, no official eradication programs exist outside of Europe. As far as data are available Table 1 gives an overview of the IBR situation in different European countries, including Switzerland. The first IBR outbreaks in Europe were observed in the 1970s (Edwards, 1988; Metzler et al., 1985). Since that time most European countries reacted with a variety of control programs, the extent of which depended on economical considerations and interests (Franken, 1997; Straub, 1999). Since the EU allows IBR-free member states to request import conditions for cattle, semen and embryos (Noordegraaf et al., 2000), more efforts were undertaken in the late 1990s to achieve eradication of BoHV-1 in the EU (Bätza, 2003; Beer et al., 2003; Trapp et al., 2003). It may be concluded from the data in Table 1 that, if BoHV-1 eradication is the goal, culling of seropositive animals without vaccination has been the most successful method. However, this can only be considered if the seroprevalence of BoHV-1 is relatively low. A decision by governmental agencies to engage against IBR was always of utmost importance for a success (Ackermann et al., 1989, 1990a). To accomplish IBR-eradication, it is advisable, in a first round, to create an IBR-free breeding stock. In many cases, this can be achieved by gradually removing all seropositive cattle from a conventional breeding lot and replacing them with seronegative progeny. ELISAs for analysing individual serum and milk samples or bulk-milk or even dried blood are excellent tools for that purpose (Rosskopf et al., 1994; Spirig et al., 1987, 1989). Clearing feedlots and free ranging foster calve facilities from BoHV-1 is more demanding (Ackermann et al., 1990c). Housing positive and negative groups under different roofs or separation by plastic curtains have been shown to greatly enhance the feasibility of IBR-eradication in feed lot facilities (Fig. 1A). Although IBR is a respiratory illness, the agent is not easily transmitted by aerosol. In contrast, salivation onto feed and movement of contaminated feed from positive groups to negative groups has been found to be a major source of BoHV-1 transmission in fattening units (Fig. 1B). During and after an eradication campaign, annual surveillance by serological means is the most

3 M. Ackermann, M. Engels / Veterinary Microbiology 113 (2006) Table 1 Status of IBR-eradication in selected European countries Country Prevalence before campaign IBR-free a Notifiable Vaccination Stamping out Surveillance Austria 0.58% (1990) Yes Yes Prohibited Yes b Yes Denmark Low (1984) Yes c Yes Prohibited Modified Yes Finland 14 cases Yes d Yes Prohibited Modified Yes Norway Low Yes e Yes Yes Sweden Low Yes f Yes Prohibited Modified Yes Switzerland % (1983) Yes g Yes Prohibited Yes Yes Belgium 62 65% No No Marker vaccine No h No France Variable No No Yes No i No Germany Variable No Yes Yes Modified j Yes Greece No information available No Yes k Hungary 13 79% l No Yes Yes Ireland No information available No No Yes m Italy 62 85% No n Yes Planned Yes Lithunia 17% No Yes Yes Luxembourg No information available No Yes Yes Yes Poland 20 38% No Yes Prohibited Modified Portugal No information available No o Yes Scotland 12% No Spain High No p Yes Yes The Netherlands 40% in dairy cattle No Marker vaccine Modified United Kingdom 2.1% (1964); 10.2% (1986) No Yes Yes q a OIE, b Eradication program compulsory since c : outbreaks in 83 herds. d No cases since e No cases since f No cases since g 29 cases since h Three step eradication program propagated (vaccination, elimination of ge positive animals, stop vaccination). i No general concept; program based on IBR-free herds certification. j Two campaign modes: (1) culling seropositive animals, no vaccination where low prevalence; (2) vaccination with marker vaccine, successive elimination of ge positive animals where high prevalence. k Since l Depending on herd size. m Restricted control measures since n District Bolzano: IBR-free, yet 2 outbreaks in o 102 cases reported in p 827 outbreaks, 4985 cases in q Movement control inside the country (in context with AI centers). Data gained either from the OIE (Handistatus II page) or from literature (Msolla et al., 1981; Edwards, 1988; Eloit, 1997; Franken, 1997; Bosch et al., 1998; Nylin et al., 1998; Köfer et al., 1999; Straub, 1999; Tekes et al., 1999; Thiry et al., 1999; Boelaert et al., 2000; Mars et al., 2000a; Paisley et al., 2001; Castrucci et al., 2002a,b; Limbourg et al., 2002; Bätza, 2003; Trapp et al., 2003; Truyen et al., 2003). important prerequisite for early recognition of newly imported cases and for maintaining the IBR-free status. Since wild ranging animals as well as sheep and goats can be excluded as BoHV-1 reservoirs, movement of seropositive cattle and trade with BoHV-1-positive semen used in artificial insemination are to be considered as the most important ways to reintroduce the virus into IBR-free facilities (Kupferschmied et al., 1986; Six et al., 2001). Vaccination with ge marker vaccines combined with eradication of ge positive animals may appear as the second best variant. However, it would be better to apply this as the strategy of choice only in catteries or regions with high seroprevalence. Notably, not a single country that included vaccination in its eradication program has yet succeeded to eradicate

4 296 M. Ackermann, M. Engels / Veterinary Microbiology 113 (2006) Fig. 1. Essentials for IBR-eradication in feedlots (Ackermann et al., 1990c). (A) Bird s view of a feedlot stable with boxes for groups of animals to both sides of the maintenance corridor. In most instances, animals of the same age will be grouped together. In the case presented, the groups containing the oldest animals are to the right and the youngest to the left. Groups of animals indicated in red contain at least one BoHV-1 seropositive member but the entire group is considered to be infected. Upon slaughtering of the animals in the most mature group, the remaining groups will be moved from the left to the box next right. If possible one box will be kept empty between positive (red) and negative (green) groups. A plastic curtain hanging from the ceiling helps efficiently to prevent transmission of BoHV-1 from positive to negative groups. (B) The same stable is shown from the front view, with the maintenance corridor in the centre. In the presented case, the air had been shown to form a circular flow, with no aeration of the animals in the groups of the opposite side of the maintenance corridor. Feed was deposited into cribs next to the maintenance corridor. It was noted that BoHV-1 was efficiently transmitted upon transferring contaminated feed from older, positive animals to younger seronegative animals. IBR. Strategies on a voluntary basis leading to certificates for IBR-free herds may be beneficial. However, they are costly for individual herd owners and a country will not reach an IBR-free status within an appropriate time. 3. The virus strikes back Austria, Denmark, Finland, Norway, Sweden, and Switzerland successfully eradicated BoHV-1 infection (Ackermann et al., 1989, 1990b; Nylin et al., 1998;

5 M. Ackermann, M. Engels / Veterinary Microbiology 113 (2006) Köfer et al., 1999; Noordegraaf et al., 2000; Paisley et al., 2001). However, 29 new Swiss cases were recorded throughout the last decade. The new cases were detected in the course of the official annual serosurveillance. The selected number of screened farms and animals was based on a confidence level of 99% for a herd prevalence of less than 0.1%, which was considered appropriate to maintain the IBR-free status (M. Reist, Federal Vet. Office, pers. comm.). In some cases the origin of the newly introduced virus could be determined. For example, one outbreak in Switzerland was tracked back to the import of latently infected animals. A second case involved trade of seropositive animals with unknown history, while a third case was associated with an outbreak of anaplasmosis (Hofmann-Lehmann et al., 2004). Previously, import of BoHV-1-positive semen was shown to be the cause of IBR-outbreaks (Kupferschmied et al., 1986). However, most officially IBR-free countries have their histories of unexplained new cases (Table 1). In any case, IBR-free countries are endangered as long as IBR-eradication is not a common goal within countries that participate in mutual cattle trade and production of semen for artificial insemination. In view of this, it seems reasonable to trace newly introduced cases of IBR back to their origins in order to minimize further damage. If latently infected animals from affected premises are available, virus shedding can be provoked by the application of dexamethasone (Ackermann et al., 1982; Ackermann and Wyler, 1984; Spirig et al., 1989). Consequently, the virus may be isolated and characterized. Due to its high G + C content, sequencing of the BoHV-1 genome is extremely demanding (Meyer et al., 1997). Therefore, systematic sequencing of individual isolates will be difficult to perform. Our laboratory has developed a clustering system, which is based on the patterns from four different restriction enzymes established on a panel of about 100 different BoHV-1 strains (Fig. 2) (Wyss, 2001). These enzymes are HpaI with 8 sites, HindIII with 12 sites, SfiI with 49 sites, and PstI with 95 restriction sites predicted by the published sequence of BoHV-1. By clustering of bands within a defined band group after digestion with HindIII, HpaI, SfiI and PstI, numeric codes were generated for each virus. To Fig. 2. The clustering principle. Band groups (clusters) were defined after digestion of BoHV-1 strains with the enzymes HindIII, HpaI, SfiI and PstI. Numeric codes, based on the number of bands within each cluster, were attributed to each virus strain. The migration of individual bands within the clusters served for refined analysis. The HindIII pattern of 3 BoHV-1 strains are shown. This pattern is divided into three clusters, cluster 1 containing 9 bands, cluster 2 three bands, and cluster 3 one band, resulting in the code 931 for the viruses presented. Migration differences of bands in cluster 1 as bands E and F, e.g. either running as two separate bands or as double-band, are marked by a Roman number. The migration property of bands in cluster 2 is in accordance with the BoHV-1 subtyping by Metzler et al., 1985.

6 298 M. Ackermann, M. Engels / Veterinary Microbiology 113 (2006) analyze a new virus strain, this code can be used to find the most similar strains out of the previously determined panel. In a second step the band lengths within each cluster are compared for a more accurate analysis. In the case of digestion with HindIII the clustering was combined with the original subtyping of BoHV-1 strains according to Metzler et al. (1985). Fig. 2 shows the principle of the system. The clustering codes (fragment numbers of each cluster with each enzyme) were then listed as numeric codes in a table (Wyss, 2001). For the characterization of a given new isolate, BoHV-1 strains are selected with the same or similar numeric codes. Then, the four established restriction enzyme patterns are compared to the new one. Using this strategy, it was possible to identify the origin of a BoHV-1 strain, which had been isolated from imported semen and which had led to a new outbreak in 1983 (Ackermann et al., 1989; Wyss, 2001). The analysis of this strain by the clustering system revealed that the virus was identical to a modified live vaccine strain that is widely used in the USA and Canada. Indeed, the semen originated from Northern America (Kupferschmied et al., 1986). In a second case, originating from a 2002 outbreak, we found HindIII and HpaI patterns that were identical to an ancient Swiss IPV isolate. However, the SfiI and PstI patterns were similar, although not identical, to two ge marker vaccine strains. We hypothesized that the 2002 isolate may be a recombinant between a vaccine and an ancient Swiss IPV strain. We suggest, therefore, to increase the number of entries into the clustering data base in order to create a broadly applicable tool for tracing the origins of novel IBR outbreaks in Europe. 4. Achievements by vaccination A number of conventional modified live (mlv) and inactivated IBR vaccines as well as subunit vaccines and ge-deleted marker vaccines have been used (Van Drunen Littel-van den Hurk et al., 1997; Belknap et al., 1999; Castrucci et al., 2002a; Limbourg et al., 2002)(Table 2). These vaccines reduce the severity of disease and also reduce virus replication and transmission, but they are not able to prevent BoHV-1 infection. Concerning the safety of mlv vaccines, no difference between conventional and marker vaccines exists with regard to virus shedding and transmission upon intranasal vaccination (Kaashoek et al., 1994; Lemaire et al., 2001). Yet, the risk of perpetuation of the marker vaccine virus in a herd is still thought to be minimal (Mars et al., 2000a). Nevertheless, mlv vaccines, when applied intranasally, were able to establish latency with occasional reactivation and shedding. Indeed, a ge-deleted virus was isolated from two cows in a field study (Dispas et al., 2003). A constant concern is the possibility of recombination between ge deleted vaccine virus and wild-type virus, which has been shown to be possible under experimental conditions (Schynts et al., 2003). However, the time interval between two successive infections may be of great influence on the recombination probability (Meurens et al., 2004). Fortunately, recombination events have not yet been detected in the field. A spontaneous BoHV-1 mutant differing from the original vaccine strain has been isolated by Rijsewijk et al. (1999). Finally, contamination of mlv vaccines by other agents has recently been experienced in the Netherlands (Bruschke et al., 2001). Several lots of a marker vaccine were contaminated by Table 2 Properties of current BoHV-1 vaccines Vaccine Protection from disease Prevention of latency Concerns Companion test Inactivated Conventional Yes No Safe No ge-deleted Yes No Safe Available Subunit Yes No Safe Possible Mlv Contamination Conventional Yes No Stability No ge-deleted Yes No Recombination Available

7 M. Ackermann, M. Engels / Veterinary Microbiology 113 (2006) bovine viral diarrhea virus of type I or type II, the latter being a highly pathogenic virus, which led to hundreds of deaths among the vaccinated animals. Although neither conventional nor ge marker vaccines are able to prevent infection with wild-type virus, there is a general agreement that the circulation of wild-type virus is significantly diminished in vaccinated herds (Bosch et al., 1997; Mars et al., 2000b, 2001; Trapp et al., 2003). The mlv vaccines in general were shown to be the most efficient in view of clinical protection and reduction of virus shedding (Bosch et al., 1998). Surprisingly, for vaccination of latently infected animals an inactivated vaccine proved to be more efficient than an mlv vaccine (Bosch et al., 1997). Nevertheless, based on a field study with an inactivated ge marker vaccine, it was concluded that major outbreaks can still occur when BoHV-1 is introduced into the vaccinated herd (Bosch et al., 1998). The main goal of the use of marker vaccines in IBReradication programs is the expectation to protect the animals from disease in combination with the possibility to differentiate vaccinated from infected animals. Although there is no doubt about the advantage of this approach, the following considerations have to be made. Firstly, the detection of infected animals is based on the assumption that all BoHV-1 wild-type strains will express ge and that all these animals will develop antibodies against ge. However, wild-type strains with variant ge have been detected that did not react with certain ge specific monoclonal antibodies (Rijsewijk et al., 2000; Egyed et al., 2000). Further in vitro and in vivo analyses of one of those field isolates, however, revealed only minor differences between the variant and a wild-type virus. Therefore it was concluded that variant virus strains may not be a danger with respect to the ge marker vaccines (Egyed et al., 2000). Secondly, the rapidity of the development of immunity as well as the duration of protection has to be considered. It has been reported that mlv marker as well as conventional vaccines were able to induce protection against disease already 2 3 days after vaccination, although specific antibodies against BoHV-1 were not detected at that time (Kaashoek and Van Oirschot, 1996). Therefore, these vaccines are propagated for use in urgency situations (Trapp et al., 2003). The protection to infection has been claimed to persist for 6 9 months after vaccination with either inactivated or mlv vaccines (Trapp et al., 2003). Thirdly, in experimental studies, ge antibodies were first detected only 2 5 weeks post infection (Beer et al., 2003). Thus, anti-ge antibodies may be missed early after wild-type infection. Lemaire et al. (1999, 2001) even reported on animals, which had been infected in the presence of maternal antibodies and which were later identified as seronegative (thus also ge-negative), latently infected virus carriers. Such incidences may be disadvantageous for successful IBR-eradication campaigns and, even worse, may negatively affect the overall confidence in veterinary vaccines. In conclusion, vaccination with ge marker vaccines combined with the detection of ge positive animals may be considered very early in an eradication program and in countries with a high BoHV-1 prevalence. However, the problems outlined above should not be underestimated, especially in countries where eradication programs enter the final phase. 5. Conclusions A number of reasons speak in favour of IBReradication, including political arguments as well as considerations of a better health system and an improved health status of our cattle. Furthermore, it may be stated that the goal can indeed be achieved as has been demonstrated by a number of countries. The price for such an achievement is, unfortunately, very high. Detection and stamping out of seemingly healthy virus carriers is inevitable in the process. Unfortunately, the use of vaccines is only of temporary and limited value. Therefore, there are at least as many considerations against such plans. The costs of such campaigns have to be weighted against the benefits. Furthermore, the absence of a specific virus from its ecological niche will create a vacuum. Who can tell, which other virus will fill in for the previous one? In any event, if either control or eradication of IBR is nonetheless a goal, there is still an urgent need for improvement of tools. Better vaccines are needed as well as better companion tests. Moreover, better tools for the characterization of viral isolates are required. Since, due to technical limitations, sequencing is not an option, the clustering system mentioned above may be worth to be further developed. Collaborative actions to collect viral strains from as many countries as possible for inclusion into the present clustering library would be most advantageous. Finally,

8 300 M. Ackermann, M. Engels / Veterinary Microbiology 113 (2006) more basic research on the fascinating BoHV-1 may ultimately provide solutions, which will render the need for IBR-eradications obsolete. Acknowledgments Our work has been funded by the Swiss Federal Veterinary Office and the University of Zurich. We thank C. Griot, L. Bruckner and collaborators for support with reactivation experiments in their facility. Furthermore, the personal communications of E. Breidenbach and M. Reist, Federal Vet. Office, are greatly appreciated. Finally, we thank Sandra Wyss for her excellent contributions to the design of the clustering system. References Achour, H.A., Moussa, A., Serological and virological studies on the infectious bovine rhinotracheitis in Algeria. J. Vet. Med. B 43, Ackermann, M., Wyler, R., The DNA of an IPV strain of bovid herpesvirus 1 in sacral ganglia during latency after intravaginal infection. Vet. 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