Why swine semen becomes a vehicle for virus transmission (Part Four)
Swine semen - a vehicle for virus transmission (Part Four)
Porcine semen as a vector for transmission of viral pathogens-Part 4
Dominiek Maesa,*, Ann Van Sooma, Ruth Appeltanta, Ioannis Arsenakisa, Hans Nauwynckb
a Belgium, Merelbeke, Ghent University, Faculty of Veterinary Medicine, Herd Health and Reproduction, Reproduction Section
a Department of Reproduction, Obstetrics and Herd Health, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium
b Belgium, Merelbeke, Ghent University, Faculty of Veterinary Medicine, Laboratory of Virology, Immunology and Parasitology, Virology Section
b Department of Virology, Immunology and Parasitology, Laboratory of Virology, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium
Keywords:
swine semen, pigs, artificial insemination, viruses, review
Semen, Pig, Artificial insemination, Virus, Review
Continued from above...
6. Preventive measures
Preventive measures
Artificial insemination centers need to adopt economical, effective and biosecure methods to produce an appropriate amount of semen of controllable quality. The best way to prevent disease transmission through semen is to purchase boars of excellent health status, implement strict biosecurity measures, and monitor boar health.
Artificial insemination centers need to produce the appropriate quantity of quality-controlled semen in a cost-effective, efficient, and biosecure process. The best way to prevent disease transmission via semen is to purchase boars with a preferred health status, to maintain very strict biosecurity measures, and to monitor the health of the animals.
6.1. Health status of newly introduced boars
Health status of incoming replacement boars
Artificial insemination centers typically replace 60% of their boars each year, but newly introduced boars bring great disease risk, so quarantine, acclimatization, adequate observation and testing of newly introduced boars are crucial. First, the health status of the herd of origin of the boars should be established. Its health status should be equal to, but preferably higher than, that of the boars at the artificial insemination center. Through communication between veterinarians, information on the clinical diagnosis and laboratory testing of the newly introduced boars, vaccination programs and veterinary measures of the herd of origin can be obtained, and the boars can be evaluated on this basis. Second, newly introduced boars should be quarantined separately for 30 to 60 days. The quarantine area is best located off-site. Herd quarantine must follow the "all-in, all-out" principle. During quarantine, herd health status should be assessed, blood samples should be collected to test for viremia or serum antibodies, and at the same time, vaccination against diseases present in the boars of the artificial insemination center should be carried outtreatment. Boars infected with dangerous pathogens must not enter the artificial insemination center. Quarantine periods, testing protocols and vaccination protocols differ among artificial insemination centers, mainly determined by different factors such as the frequency of boar introduction and the health status of the animals in the herd of origin [88].
Because new boar entry poses a great risk for disease, and because typically 60% of the boars in AI centers are replaced on a yearly basis, it is of paramount importance for incoming boars to implement proper isolation and acclimation procedures, with adequate time for observation and testing. First, the health status of the source herd should be known. The health status should be similar but preferably higher than the health status of the AI center. It could be evaluated through vet-to-vet communications of the diagnostics and laboratory examinations, vaccination programs, and veterinary involvement of the source herd. Second, replacement boars should be housed for 30 to 60 days in a separate isolation facility. Such a quarantine unit should be located preferably off-site. Isolation populations must always be flowed all-in/all-out. During this quarantine period, the health status should be evaluated, blood sampling could be performed to detect possible viremia or the presence of serum antibodies, and vaccinations can be applied against diseases present in resident boars of the AI center. Animals that are infected with pathogens of concern should not enter the AI center. The isolation period, the testing protocols, and the vaccination schemes vary between AI centers and depend on different factors such as the frequency of boar introduction and the health status of the animals of the origin herds [88].
6. Preventive measures
Preventive measures
6.1. Biosecurity measures
Biosecurity measures
The most important biosecurity measures for artificial insemination centers involve the following [3,88]: (1) Location and ventilation equipment of the artificial insemination center -- it should be as far away from pig herds as possible. Air filtration systems should be used in areas with dense pig populations to ensure that the artificial insemination center is protected from airborne pathogen introduction [89]. Ensure that ventilation equipment does not exhaust air from the boar station toward the laboratory. If the laboratory is adjacent to the pig barn, positive-pressure ventilation should be used to minimize air infiltration from the barn into the laboratory. (2) Strict rules should apply to employees and visitors entering the artificial insemination center. Employees need to be quarantined for 1 to 3 days. During the quarantine period, contact with pigs is not allowed. In addition, personnel shower-in/shower-out measures are recommended for all visitors and employees. (3) Semen collection point and delivery -- the location where semen delivery personnel collect semen should be outside the artificial insemination center. The semen collection point should be a separate room, or at least a room separate from the laboratory and the boar station. Keeping the semen collection point separate can minimize cross-contamination of semen coolers, boots, clothing, etc. The semen drop-off point for each sow barn should be located outside the office. Any items brought into the artificial insemination center (e.g., supplies, equipment) should not have come into contact with pigs beforehand. Items should be placed in a designated room for disinfection before being brought into the artificial insemination center. (4) Rodent and insect control - rodents and insects can carry and transmit manyswineviruses, so appropriate preventive measures should be taken. (5) Water quality - the water source for boars should be analyzed regularly. If the quality is substandard, the water source should be replaced or the water appropriately treated. (6) Implement strict cleaning and disinfection procedures.
The most important biosecurity measures for AI centers relate to the following items [3,88]: (1) location of the AI center and ventilation -- the AI center should be located as far from pigs as feasible. Filter systems for incoming air could be used in pig-dense areas to safeguard AI centers against entry of airborne pathogens [89]. The facilities should be oriented so that exhaust air from the boars does not exhaust toward the laboratory. If the laboratory is connected to the animal facility, there should be positive pressure ventilation to minimize air infiltration from the animals into the laboratory. (2) Employees and visitors -- strict regulations should be in place for persons entering AI centers. A downtime of 1 to 3 days may be required for persons entering the AI center. During this downtime period, persons are not allowed to have contact with pigs. In addition, it is recommended to have a shower-in/shower-out policy for all visitors and employees. (3) Semen pickup point and deliveries -- the semen pickup point for the courier delivery personnel should be located at the outside of the AI center. This pickup point should be in a separate room, or as a minimum, a separate room from both the laboratory and the stud population. By having a separate pickup point, cross-contamination of semen coolers, boots, clothing, and so forth is minimized. Drop-off points at each of the sow units should be located outside of the office. Any deliveries (e.g., supplies, equipment) on the AI center should not have been exposed to pigs previously. They should be delivered directly to a designated receiving chamber and properly disinfected before entering the AI center. (4) Rodent and insect control - rodents and insects are capable of carrying and transmitting several pig viruses, and therefore, proper prevention measures should be in place. (5) Water quality - water sources for boars should be analyzed regularly. If the quality is insufficient, the water source should be changed or the water should be treated properly. (6) Implementation of strict cleaning and disinfection procedures.
6. Preventive measures
Preventive measures
6.3 Boar health monitoring and vaccination
Animal health monitoring and vaccination
The most common approach is to monitor the health status and clinical presentation of boars. If clinical problems occur, semen must not be collected until the animal has recovered. Automating the measurement of boar feed intake and water consumption makes it easier to detect sick boars at an early stage. Targeted monitoring of boar health status and performance several times a day is still a very basic method and cannot independently monitor viral infection and/or the presence of virus in semen. Therefore, it is recommended to establish standard protocols for immunization, deworming and sampling for specific diseases, and to review them regularly. Vaccination of boars can very effectively eliminate viruses or reduce virus shedding, and reduce the risk of virus transmission through artificial insemination. First, artificial insemination centers need to comply with the country's animal health regulations, for example, mandatory vaccination against ADV, because it is part of official virus eradication programs (e.g., Eastern Europe), while in other countries that have officially eradicated ADV, ADV vaccination may be prohibited (e.g., Western Europe, North America). The combination of highly effective recombinant marker vaccines and various precise enzyme-linked immunosorbent assays (ELISA) has made it realistically possible to eradicate ADV over large areas of the world (e.g., many European countries and North America) [48]. The European Pharmacopoeia requires that after boars are vaccinated with injectable ADV vaccine, their semen must not be used [90]. For other diseases not required by laws and regulations, artificial insemination centers can decide for themselves which vaccine to use. Vaccination against PPV and PCV2 may help reduce virus shedding after infection [91]. In the case of PRRSV, within 50 days after boars are vaccinated with modified live vaccine, the shedding of field virus in the body can be reduced or eliminated [92,93]. It cannot be ignored that modified live vaccines provide only partial immune protection, resulting in shedding of vaccine virus in semen for up to 39 days [17,92]. In contrast, inactivated vaccines cannot significantly reduce the shedding of field virus in semen [92].
The most general method is to monitor the health of the boars and the presence of clinical signs. In case of clinical problems, no semen should be collected until the animal has recovered. Feed and drinking water intake of the boars may be measured in an automated way, allowing to identify diseased boars more easily and at an early stage. Monitoring the health and performance of the boars, even if applied accurately and more than once a day, remains a very basic approach that is insufficient as a stand-alone measure to detect viral infections and/or the presence of viral pathogens in semen. Therefore, it is recommended to develop standard protocols for vaccinations, deworming, and sampling for specific diseases and to review these on a regular basis. Vaccination of boars can be highly effective in eliminating or decreasing shedding of viruses and decreasing the risk of virus transmission by AI. First of all, AI centers need to adhere to the animal health regulations of the country, e.g., vaccination against ADV may be mandatory because it is part of an official eradication program of the virus (e.g., Eastern Europe), whereas in other countries, vaccination may be forbidden because the country is officially free of ADV (e.g., Western Europe, North America). The combination of highly efficacious marker vaccines and accurate differential ELISAs has made eradication of ADV from large areas of the world practical and feasible (e.g., many European countries, North America) [48]. According to requirements of the European Pharmacopoeia, ADV vaccines for parenteral use must not be transmitted by semen [90]. For other diseases, no legal regulations are in place and the AI center can decide which vaccines to use. Vaccination against PPV and PCV2 may help to reduce shedding of the virus after infection [91]. In the case of PRRSV, the use of an attenuated vaccine shortened or eliminated virus shedding in boars challenged with wild-type virus for 50 days after vaccination [92,93]. Apart from the fact that only partial protection is conferred against infection, vaccination with an attenuated vaccine led to semen shedding of the vaccine virus for up to 39 days [17,92]. In contrast, an inactivated vaccine did not clearly reduce subsequent shedding of wild-type virus in semen [92].
Because possible viral infections (such as PRRSV) need to be detected as early as possible, health monitoring is crucial for artificial insemination. Generally, blood and semen samples need to be tested for the presence of viruses and/or antibodies. Testing procedures vary greatly among artificial insemination centers. Because blood and semen testing sometimes yield different results, and because of uncertain factors such as the optimal testing frequency, sample size, diagnostic methods and interpretation of test results, testing procedures are complex [87].
Health monitoring programs are critically important in AI centers, as possible infection with pathogens (e.g., PRRSV) should be detected as soon as possible. Mostly, both blood and semen samples are tested for the presence of virus and/or antibodies. Monitoring programs can very between AI centers. They can be complex because blood and semen tests can sometimes generate different information, and uncertainties exist regarding optimal testing frequency, numbers of animals to test, diagnostic test to choose, and how to interpret the test results [87].
6. Preventive measures
Preventive measures
6.4 Virus-free semen collectiontechnology
Technologies to obtain virus-free semen
Processing semen and adding antimicrobial agents can dilute semen and reduce bacterial contamination, but these measures cannot eliminate viruses [4,5]. At present, there is no effective antiviral agent that can guarantee virus-free semen. Many proteins, as well as steroid hormones, enzymes and other substances, are present in semen and can to some extent prevent viral infection, but these substances may also damage sperm. Therefore, a delicate balance needs to be maintained [94].
Semen processing and addition of antimicrobials may have a possible dilution effect and decrease bacterial contamination, but these measures do not eliminate viruses [4,5]. Effective antiviral agents to render semen virus-free are currently not (yet) used in practice. Many different proteins found in semen along with steroid hormones, enzymes, and other substances may provide some protection against viral infections, but at the same time, these substances could damage sperm. Therefore, a delicate balance of these substances needs to be maintained [94].
Colloidal centrifugation of swine semen has a history of at least 20 years, from early density gradient centrifugation to the more recently developed single-layer centrifugation (SLC) [95]. Compared with density gradient centrifugation, the latter is more convenient. The use of single-layer centrifugationtechnologycan eliminate pathogens (bacteria and viruses), thereby improving the biosafety of semen and reducing the use of antimicrobial agents. In the study by Blomqvist et al. [96], more than 99% of PCV2 could be removed from semen, with the initial infectious virus titer reduced to 3.25 to 3.82 TCID50/50μL.
Colloid centrifugation of boar semen has been reported sporadically for at least 2 decades, beginning with density gradient centrifugation and progressing more recently to single-layer centrifugation (SLC) [95]. The latter method is easier to use and less time-consuming than density gradient centrifugation. The technique can be used to remove pathogens (bacteria and viruses), thus improving biosecurity of semen doses and potentially reducing the use of antimicrobials. In a study by Blomqvist et al. [96], it was possible to remove more than 99% of PCV2 from semen with an initial infectious virus titer of 3.25 to 3.82 TCID50/50μL.
Martinez-Alborcia et al. [97] proposed that before semen freezing, single-layer centrifugation (SLC) of semen samples using Androcoll-P colloid can improve sperm cryosurvival and alter the function of cryopreserved sperm. A subsequent study showed that SLC treatment of single-ejaculate boar semen using Androcol-P can process the entire sperm-rich fraction, improving sperm cryosurvival and the fertilizing ability of sperm after thawing [98]. Thistechnologycan eliminate pathogens and improve sperm cryopreservation, both of which help make frozen semen from genetically superior boars easier to trade internationally. However, SLC needs further research and improvement, for example, to make it faster and more reliable.
Martinez-Alborcia et al. [97] reported that SLC of semen samples using a pig-specific colloid called Androcoll-P before freezing enhances sperm cryosurvival rates and modifies the functionality of the cryosurviving sperm. A subsequent study showed that SLC processing of boar ejaculates using Androcoll-P is able to process the entire sperm-rich fraction of a boar ejaculate, improving both the sperm cryosurvival and the fertilizing ability of thawed spermatozoa [98]. The removal of pathogens coupled with improved cryopreservation are two important advantages of the technique allowing easier international trade of frozen–thawed semen doses from genetically valuable boars. However, further research is necessary to improve SLC, e.g., by making the technique less time-consuming and the results less variable.
7. Conclusion
Conclusions
Artificial insemination can rapidly introduce viruses from swine semen into sow herds, thereby causing infection and disease in the herd, so virus-carrying semen poses a risk tobreeding pigherds. Testing before boars enter the facility, routine monitoring during their time at the artificial insemination center, regular testing before semen delivery, and the implementation of strict biosecurity measures can all enhance producers' trust in artificial inseminationtechnology, believing that it can provide large quantities of safe semen with excellent genetic traits. At the same time, preventing viral contamination of semen is of paramount importance; once semen carries virus, eliminating the virus is more difficult than preventing it. However, the development of new semen processing methods (such as SLC) will gradually give people more confidence in artificial insemination as a safe pig breedingtechnology. ⟦42 End
Viral contamination of porcine semen poses a risk for breeding herds because AI may lead to fast introduction of viruses into sow populations and to loss of disease-free status for these pathogens. Pre-entrance testing of boars, routine health monitoring of boars during their stay in the AI center, regular testing of semen before delivery, and enforcement of strict biosecurity protocols at the AI center provide most producers with a high level of confidence that AI is a relatively safe means to offer unlimited exchange of desirable genetic characteristics. Prevention of viral contamination of semen should be a primary focus because it is easier to prevent contamination than to eliminate viruses once they are present in semen. Nevertheless, research and development of novel semen processing treatments such as SLC may even further instill confidence in the use of AI as a safe reproductive technology for pig production.
Done
-The end-