Pig semen-the Vector of virus Transmission (4)

Pig semen-the Vector of virus Transmission (4)

Porcine semen as a vector for transmission of viral pathogens-Part 4

Dominiek Maesa,*, Ann Van Sooma, Ruth Appeltanta, Ioannis Arsenakisa, Hans Nauwynckb

A University of Ghent, Mellerbeck, Belgium, College of Veterinary Medicine, Animal Health and Obstetrics, Reproduction

A Department of Reproduction, Obstetrics and Herd Health, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium

B Belgium, Mellerbeck, University of Ghent, College of Veterinary Medicine, Virology Laboratory, Immunology and Parasitology, virus discipline

B Department of Virology, Immunology and Parasitology, Laboratory of Virology, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium

Keyword Keywords:

Pig semen, pig, artificial insemination, virus, review / examination

Semen, Pig, Artificial insemination, Virus, Review

Follow up the above.

6. Precautionary measures

Preventive measures

Artificial insemination centers need to use economical, effective and biosafety methods to produce an appropriate amount of semen with controllable quality. The best way to prevent the spread of disease through semen is to buy boars in good health, implement strict biosafety measures, and monitor boars' health.

Artificial insemination centers need to produce the appropriate quantity of quality-controlled semen ina 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 new boars

Health status of incoming replacement boars

Artificial insemination centers usually replace 60% of boars every year, but new boars pose a great risk of disease, so it is very important to isolate, domesticate, fully observe and test new boars. First of all, we should know the health status of the boar herd. Its health status should be equal to, but preferably higher than that of boars in artificial insemination centers. Through the communication between veterinarians, we can learn about the clinical diagnosis and laboratory tests of new boars, vaccination programs and veterinary measures of the source herd, which can be evaluated. Secondly, new boars should be isolated for 30 to 60 days. The quarantine area is preferably located off the court. Pig herd isolation must abide by the principle of "all in and all out". During isolation, the health status of pigs should be assessed, blood samples should be taken to detect viremia or serum antibodies, and diseases in boars in artificial insemination centres should be vaccinated. Boars infected with dangerous pathogens are not allowed to enter artificial insemination centres. Different artificial insemination centers have different isolation periods, testing schemes and vaccination programs, mainly due to different factors, such as the frequency of introduction of boars and the health status of the original herd animals [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. Precautionary measures

Preventive measures

6.1. Biosafety measures

Biosecurity measures

The most important biosafety measures for artificial insemination centres involve the following: (1) the location and ventilation of artificial insemination centres should be as far away from pigs as possible. An air filtration system is used in dense areas of pigs to ensure that artificial insemination centres avoid the introduction of pathogens through the air [89]. Ensure that the ventilation does not drain the air from the boar station to the laboratory. If the laboratory is close to the pigsty, positive pressure ventilation should be used to minimize air infiltration from the pigsty to the laboratory. (2) there should be strict regulations for employees and visitors who enter the artificial insemination center. Employees need to be quarantined for 1 to 3 days. Contact with pigs is not allowed during quarantine. In addition, it is recommended to implement personnel entry and exit shower measures for all visitors and employees. (3) semen collection and delivery-the place where semen delivery personnel take semen should be located outside the artificial insemination center. The semen collection point should be a separate room, or at least a separate room from the laboratory and boar station. Separate the semen collection points separately, which can avoid the cross-contamination of semen coolers, boots, clothing and so on. The semen placement site of each sow house should be located outside the office. Any items (for example, materials, equipment) sent into an artificial insemination center should not have come into contact with pigs before. Items should be put in a designated room for disinfection before being sent to the artificial insemination center. (4) deratization and insects-rodents and insects can carry and transmit a variety of swine viruses, so appropriate precautions should be taken. (5) Water quality-the water source of boars should be analyzed regularly. If the quality is not up to standard, the source of water should be changed or the water should be treated properly. (6) carry out strict cleaning and disinfection procedures.

The most important biosecurity measures for AI centers relate to the following items [3mem88]: (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. Precautionary measures

Preventive measures

6.3 Health surveillance and vaccination of boars

Animal health monitoring and vaccination

The most common method is to monitor the health status and clinical manifestations of boars. If there are clinical problems, semen should not be collected until the animal recovers. Automatic methods are used to measure the food intake and water consumption of boars, making it easier to detect sick boars at an early stage. The health status and performance of boars are monitored many times every day, but this is still a very basic method and cannot independently monitor virus infection and / or the presence of viruses in semen. Therefore, it is recommended that standard programmes for immunization, disinfestation and sampling for specific diseases be developed and reviewed on a regular basis. Boar immunization can be very effective in eliminating the virus or reducing the spread of the virus, and reduce the risk of virus transmission by artificial insemination. First, artificial insemination centres need to comply with the country's animal health regulations, for example, compulsory vaccination of ADV because it is part of an official programme to eliminate the virus (e.g. Eastern Europe), while in other countries where ADV has been officially cleared, ADV immunization may be banned (e.g. Western Europe, North America). The combination of highly efficient recombinant labeled vaccines and a variety of accurate enzyme-linked immunosorbent assay (ELISA) has made it possible to eliminate ADV on a large scale around the world, such as many European countries and North America [48]. European Pharmacopoeia requires that after boars are vaccinated with ADV vaccine, their semen cannot be used [90]. For other diseases that are not required by laws and regulations, the artificial insemination center may decide which vaccine to use. Vaccination with PPV and PCV2 vaccines may help to reduce the spread of virus after infection [91]. As far as PRRSV is concerned, the detoxification ability of wild virus in boars can be reduced or eliminated within 50 days after vaccination with attenuated vaccine. It can not be ignored that the attenuated vaccine can only provide partial immune protection, resulting in the release of the vaccine virus in semen for as long as 39 days. [17,92] . In contrast, inactivated vaccine could not significantly reduce the excretion of wild 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 of the need to detect possible viral infections (such as PRRSV) as soon as possible, health surveillance is essential for artificial insemination. In general, it is necessary to detect the presence of viruses and / or antibodies in blood and semen samples. The testing procedures of different artificial insemination centers vary greatly. Because sometimes blood and semen tests have different results, and there are uncertain factors, such as the best test frequency, sample number, diagnostic methods and interpretation of test results, the detection procedure is very 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. Precautionary measures

Preventive measures

6.4 virus-free semen collection technique

Technologies to obtain virus-free semen

Semen treatment and addition of antimicrobials can dilute semen and reduce bacterial contamination, but these measures do not remove viruses. At present, there is no effective antiviral agent to ensure that semen is virus-free. There are many proteins in semen, as well as steroids, enzymes and other substances that can prevent viral infection to some extent, but these substances can 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].

Pig semen colloid centrifugation has a history of at least 20 years, from the early density gradient centrifugation to the recently developed monolayer centrifugation (SLC) [95]. Compared with density gradient centrifugation, the latter is more convenient. The use of single-layer centrifugation can eliminate pathogens (bacteria and viruses), thereby improving the biosafety of semen and reducing the use of antimicrobials. In the study of Blomqvist et al. [96], more than 99% of PCV2 could be removed from semen, and the initial infection virus titer was reduced to 3.25 to 3.82 TCID50/ 50 μ L.

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/ 50 μ L.

Martinez-Alborcia et al [97] proposed that monolayer centrifugation (SLC) of semen samples with Androcoll-P colloid before semen cryopreservation can improve the sperm cryopreservation survival rate and change the function of cryopreservation sperm. A subsequent study showed that SLC treatment of boar ejaculatory semen with Androcol-P could completely deal with the sperm-rich part and improve the frozen viability of sperm and the fertilization ability of thawed sperm [98]. This technology can remove pathogens and improve the cryopreservation of spermatozoa, which are beneficial to the frozen semen of boars with genetic advantages and make it easier for international trade. 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 will quickly introduce the virus in pig semen into sows, and then cause infection in pigs, so infected semen will bring risk to breeding pigs. The testing of boars before admission, routine monitoring during the artificial insemination center, regular testing before semen delivery, and the implementation of strict biosafety measures can enhance producers' trust in artificial insemination technology. it is believed that it can provide a large amount of semen with safe and good genetic characteristics. At the same time, the prevention of virus contamination of semen is a top priority. Once semen is infected, it is more difficult to remove the virus than to prevent it. However, the development of new semen processing methods, such as SLC, will gradually increase people's confidence in artificial insemination, a safe technique for pig breeding.

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.

Finish

-The end-