Diseases of Swine (8th edition)/Chapter 64

Although natural selection has been genetically improving disease resistance in pigs for millennia, pathological organisms are also evolving to survive in an ever changing environment and host organism. Humans confound the situation by constantly changing the environment and managerial practices used to raise pigs. Alterations in animal density, vaccinations, and antibiotics have significantly modified this ecosystem and have made the relationship between hosts' genetics and susceptibility to acquired diseases extremely complex.

A few general theories from quantitative genetics form the basis of these genetic mechanisms (Falconer 1981). First, an animal's phenotype is the result of its genotype and the environment in which it developed. Animal breeders usually partition phenotypic variation into its causal components and describe it with the following equation:


 * phenotypic variation (P) = genetic variation (G) + environmental variation (E) + interaction between G and E

Second, genetic variation is partitioned into the components of additive variation, dominance variation, and epistatic variation. An important concept to remember is that selection, whether natural or imposed by humans, utilizes only additive genetic variation. Heritability is the proportion of phenotypic variation that is due to additive genetic variation and is defined as


 * heritability (h2) = additive genetic variation/ phenotypic variation

An analogous definition of heritability, and the one most used by producers, is the response to selection relative to the superiority of the selected parents. If a population of pigs has an average tenth-rib fat measurement of 2.5 cm (h$2$ = 0.40) and the average of the selected parents is 2.0 cm, then we would expect the progeny to have an average tenth-rib fat measurement of 2.3 cm; that is, 0.40(2.0 - 2.5) + 2.5 = 2.3. Even though the selection differential of the parents was 0.5 cm, only 40% of that difference is expected to be passed on, because the heritability of the trait is 0.40. Heritabilities less than 0.20 are considered low, those between 0.20 and 0.40 moderate, and those above 0.40 high.

A trait with low heritability should not be assumed to be unaffected by genetics. In fact, any trait that differs between breeds or responds to heterosis has a significant genetic component. Heterosis is the difference in performance of crossbred animals from the average performance of purebred animals and is attributed to genes with dominant and epistatic action. Traits most affected by heterosis are those pertaining to fitness (longevity, reproduction rate, etc.). It is quite common to observe that F1 females will have litters approximately 10% larger than the average litter size of the two breeds used to produce the crossbred gilt. This larger litter size is because of the heterosis (or hybrid vigor) present in crossbred animals.

Producers would like to be able to breed pigs resistant or resilient to invasion of all pathogens (general disease resistance). However, the etiologies of diseases can be quite different and the host has different mechanisms which it uses to prevent pathogens from causing disease. The two basic lines of a pig's defense are antibody-mediated and cell-mediated immunity. The defense mechanism utilized depends on the pathogen. It also appears that these mechanisms are controlled by different genes located throughout the genome. The situation is further complicated by clear evidence that resistance to certain pathogens can be largely controlled by nonimmunological factors. Identification of the genetic mechanisms that affect susceptibility should be easier for diseases with simple etiologies than for diseases with complex etiologies. When resistance/susceptibility is clearly defined and controlled by a single gene, scientists can use the recently developed swine genetic map (Rohrer et al. 1996) to identify the chromosomal region of the genome which possesses the gene. Genetic improvement of disease resistance in pigs requires identifying heritable phenotypes (measurements) of resistance which are well defined, accurately measured, and highly correlated with incidence of disease (marker traits). If we focus selection on resistance to a single pathogen, we may produce animals that are only resistant to a specific disease. Selection of animals less susceptible to many pathogens requires measurements of marker traits for antibody-mediated immunity (AMI) and for cell-mediated immunity (CMI) and that the phenotypes be incorporated into a selection index (general disease resistance).

To date, very few studies of swine have examined the genetics of disease resistance. Therefore, this review begins by summarizing studies of disease resistance relevant to swine in two livestock species, chickens and cattle, and one model species, the mouse. We then discuss studies conducted in swine that attempt to determine differences between breeds and those calculating heritabilities associated with disease resistance. We conclude with identification of genes possibly involved in protecting the pig from infection and a discussion of the potential of gene-mapping technologies to identify genes or markers that segregate with genes which affect disease resistance.

SELECTION EXPERIMENTS IN CHICKENS AND MICE
Selection upon marker traits for CMI and AMI has produced birds with significantly different measurements for the traits, but in all studies to date the resistance of these birds to diseases has been tested only with the virus causing Marek's disease. Divergent selection for levels of IgG at 10 weeks of age (Tamaki 1981) and antibody titers in response to vaccinations with sheep red blood cells (Pinard et al. 1992) were conducted to determine the heritability of AMI response. The heritability of CMI was evaluated in chickens by selecting on response to natural infections with Leucocytozoon caulleryi (Okada et al. 1985) and degree of splenomegaly in graft-versus-host reactions (Okada and Mikami 1974). Selection in each line of birds resulted in chickens with divergent phenotypes for the trait selected upon. Okada and Yamamoto (1987) measured responses to vaccination with sheep red blood cells, bovine serum albumin, and lipopolysaccharides in animals from both populations selected for CMI and the AMI line of Tamaki (1981) and found that birds selected for high immune response generally produced more antibodies to the antigen challenge than birds from the low-response selection lines. In addition, birds in highresponse lines had a higher spleen-to-live-weight index but were usually more susceptible to a challenge from Marek's disease. Contrarily, Pinard et al. (1992) determined that birds selected for greater antibody response to sheep red blood cells were more resistant to Marek's disease than those selected for low response. Okada and Yamamoto (1987) were also able to show significant differences in susceptibility to Marek's disease in animals from similar genetic backgrounds but with different major histocompatibility complex (MHC) genotypes. This indicates that at least one of the genes responsible for resistance to Marek's disease may be either a member of the MHC or located near the MHC.

Biozzi et al. (1982) summarized the results of an elaborate selection experiment in mice for high and low antibody production in response to erythrocyte antigens from either sheep or pigeons and in response to two different species of Salmonella (S. typhimurium and S. oranienburg). A line was also vaccinated with bovine serum albumin and rabbit gamma globulin. Selection was quite effective and heritabilities for all marker traits were about 0.20. Once the response to selection had reached a plateau, each selected line was rechallenged to determine correlated responses to the selection imposed. Biozzi et al. (1982) found that the high-response line had greater antibody titers than the low line following vaccination with numerous antigens different from those used in the selection program. However, macrophage antigen catabolism was much lower in the high-response line. Thus, these animals were more susceptible to diseases that require macrophage-dependent protection (CMI). Because of the greater processivity of the macrophages in the low-response line, these animals possessed a greater innate resistance to pathogens that require CMI (five pathogens were studied, including S. typhimurium and Brucella abortus). Innate immunity to parasitic infections and rabies virus, which requires AMI protection, was similar in both lines. Acquired immunity was greater in the high-response line for parasitic infections and rabies, while the low line had superior acquired immunity to bacterial infections. In all cases studied, selection for greater antibody production (AMI) resulted in reduced CMI responses, yielding animals more susceptible to particular pathogens. These results clearly indicate that selection did not result in animals resistant to all types of pathogens, that genetic mechanisms which regulate AMI are not identical to those affecting CMI, and the two different genetic mechanisms may be negatively correlated. Biozzi et al. (1982) hypothesized that the negative genetic correlation between AMI and CMI responses allows animal populations to maintain maximum resistance to all pathogens it may encounter to permit survival of the species. These results indicate that selection for AMI should be quite successful in pigs, but that selection for animals with general disease resistance to most pathogens will be quite difficult or even impossible.

DISEASE RESISTANCE IN RUMINANTS
Mastitis has been the chief focus of studies of genetic mechanisms of disease resistance in cattle. The etiology of mastitis is quite complex since many pathogens can actually cause the disease and numerous environmental factors can influence its progression, resulting in low heritability. Estimates of heritability for direct measures of mastitis (incidence of subclinical and clinical mastitis) range from 0.06 to 0.12 (Shook 1989). Because incidence of mastitis is a subjective measurement that cannot be standardized across producers, the dairy industry uses the marker trait of somatic cell count (SCC) in milk samples to indicate a cow's susceptibility to mastitis. However, SCC can also be influenced by many factors and its heritability is similar to that of mastitis, but the genetic correlation between the two traits is high (0.5-0.9). Most dairy cows in the United States have SCC measurements, because it is one of the measurements taken by the Dairy Herd Improvement Association. Because SCC is an objective measurement and recorded on a large number of cows, estimated breeding values for sires can be computed with relatively high accuracies to permit producers to select for a reduction in SCC and thus decrease the incidence of mastitis in their herds. In the United States, this is the only type of selection for disease resistance practiced by the dairy industry, and probably by any livestock industry. In Norway, producers also have access to the estimated breeding value (EBV) for incidence of ketosis (Shook 1989). The estimates of heritability for incidence of ketosis (about 0.1) indicate that a selection program should be effective if incidences of ketosis were recorded for a large number of cows and included in a national sire summary (Shook 1989).

Evidence of genetic variability for susceptibility to infestations of specific parasites such as protozoa, nematodes, and arthropods was summarized in an excellent review by Wakelin (1978), and most estimates of heritability for these traits are in the moderate to high range (0.2-0.5). More recently, scientists have detected evidence of major genes affecting tick resistance in cattle (Frisch 1994) and resistance to Haemonchus contortus in sheep (Albers et al. 1987). An interesting note is that both major genes identified appear to be dominant, indicating that only one copy of the resistant allele is necessary to confer resistance. The superior resistance of Bos indicus cattle to external parasites is well documented (Hutt 1958). In addition to their innate resistance to tick infestations, it has also been reported that Bos indicus cattle are resistant to tick-borne diseases (Hutt 1958). This resistance is probably not due to an immunological factor. Rather, it is most likely that animals with low tick burdens have a much lower probability of being infected.

Burton et al. (1989) studied the responsiveness of dairy calves to injections with human erythrocytes and ovalbumin to determine whether animals that produced more antibodies had a greater level of general immune resistance. While they found a low correlation between general resistance and antibody production in response to erythrocytes and ovalbumin, they did observe that animals with higher titers to erythrocyte antigens had a lower incidence of diarrhea. No correlations were detected for incidence of pneumonia. Burton et al. (1989) were able to calculate a heritability of 0.48 for antibody production in response to ovalbumin and 0.31 for human erythrocytes, which would indicate that selection for these marker traits would be quite successful even though the level of general disease resistance the animals would possess is questionable.

Because of the MHC role in immune function, it has been studied by numerous laboratories for its impact on general disease resistance. Part of the genetic basis of resistance to mastitis can be attributed to genes within the bovine MHC or other closely linked genes (Lundén et al. 1990; Weigel et al. 1990; Andersson-Eklund and Danell 1993; Mallard et al. 1995). Lewin and Bernoco (1986) were also able to show an association between MHC haplotypes and progression of bovine leukemia virus (BLV). Even though the effect was not seen for initial infection, there was evidence of resistance to BLV-dependent B-cell proliferation and lymphocytosis (Lewin and Bernoco 1986). Other diseases in which susceptibility may be controlled by the MHC in cattle are squamous cell eye carcinoma, trypanosomiasis, and the tick Boophilus microplus (Tizard 1987), but a single MHC type that appears to convey the most resistance to all pathogens has yet to be found.

RESISTANCE TO SPECIFIC DISEASES IN SWINE
In spite of the apparent success of early attempts to select for disease resistance in swine, these results have not been followed up. German veterinarians in the 1940s (Hutt 1958) developed a line of pigs that were resistant to erysipelas. In this study, pigs were selected based on their response to inoculations with the causative pathogen (Erysipelothrix rhusiopathiae), and after a few generations of selection a line completely resistant to chronic and severe acute infections was produced (Hutt 1958). Cameron et al. (1942) reported the identification of pigs resistant to Brucella suis. They were able to show that the observed resistance was genetic, as they conducted studies on three subsequent generations of pigs. They proposed that the resistance allele was recessive, but their results indicate that additional genes were also affecting resistance or that a phenomenon known as incomplete penetrance was present (Cameron et al. 1942). Incomplete penetrance is when only a portion of animals with a given genotype express the expected phenotype. This phenomenon has been observed for the porcine stress syndrome locus, where some animals that are homozygous for the stress-susceptible allele do not react to halothane anesthesia, but the reasons behind incomplete penetrance have yet to be identified. Hutt (1958) also presents evidence for genetic disease resistance in swine to hog cholera, dysentery, and atrophic rhinitis. Reports of additional attempts at selection for disease resistance do not appear in number until the 1970s.

The only reported study calculating the heritability of contracting a swine disease was conducted by Kennedy and Moxley (1980). They estimated the heritability for incidence of atrophic rhinitis to be 0.12. Incidence of infection appeared to be more heritable than severity of infection, using severity scores ranging from 0 to 5 (h$2$ = 0.03). Other groups have estimated heritabilities for immune response (measured as antibody titers) to vaccination for atrophic rhinitis. These estimates of heritability were generally low (h$2$ = 0.12-0.15) (Rothschild et al. 1984; Meeker et al. 1987b) unless the measurements were recorded after maternal immunity was exhausted (h$2$ = 0.5) (Meeker et al. 1987b). Both studies indicated that maternal or litter effects substantially affected antibody titers when serum samples were collected early in life and maternal antibodies were present in the offspring. Breed effects detected for immune response to Bordetella bronchiseptica by both Rothschild et al. (1984) and Meeker et al. (1987a) were similar, with Landrace and Yorkshire breeds reportedly superior to Duroc. Contrary to expectation, neither study detected any evidence for heterosis. Hutt (1958) had previously noted that Canadian producers had reduced the incidence of rhinitis when they deliberately selected against "dish-faced" pigs. However, a systematic effort to pursue this observation is lacking.

Meeker et al. (1987a, b) also evaluated the immune response to vaccination for pseudorabies. They estimated the heritability for antibody production to be 0.18 but were unable to detect any breed differences or evidence of heterosis. The correlation between response to pseudorabies versus atrophic rhinitis vaccines was extremely low (Meeker et al. 1987a). More important, correlations with immune response and production traits were low but indicated that individuals with high antibody titers grew slower (Meeker et al. 1987a). Either the pigs producing more antibodies were more susceptible to other diseases or they were slower growing pigs.

Swine also have genetically based resistance to the nematodes Trichinella spiralis, Ascaris suum, and Strongyloides ransomi. Wakelin (1978) presented heritabilities for resistance to A. suum and S. ransomi similar to those found for other species (moderate heritabilities). Madden et al. (1990, 1993) demonstrated that some swine were able to eliminate encysted T. spiralis muscle larvae and that this ability was associated with the pig's MHC genotype. They documented that the elimination of the encysted larvae was an immunological response mediated or facilitated via eosinophils (Madden et al. 1993) but were unable to detect any differences in expression of class I or II MHC molecules. Whether the observed association of the ability to eliminate encysted larvae with MHC haplotype was due to MHC genes or genes closely linked to the MHC still needs to be determined.

Hutt (1958) also reported a study that identified a major gene for susceptibility to preweaning diarrhea, but follow-up studies were not conducted to further characterize these results. Susceptibility to two types of Escherichia coli obtained from young diarrheic pigs has been shown to be controlled by a single gene for each pathogen. In both cases susceptibility appears to be the dominant trait. Gibbons et al. (1977) determined the inheritance of resistance to E. coli F4+ (K88) strains of bacteria. Bertschinger et al. (1993) studied the susceptibility to F18 (F107) fimbriated E. coli. Both pathogens cause diarrhea when the bacteria are able to colonize on the epithelial lining of the small intestine. Colonization is only permitted when receptors for that particular strain are present on the intestinal walls. These strains of E. coli do not appear to be pathogenic in pigs that do not permit colonization. Edfors-Lilja et al. (1986) were able to demonstrate that pigs which had receptors for E. coli F4+ (K88) had a lower rate of preweaning growth, indicating that their performance was suffering due to their susceptibility to the pathogen. However, the pigs displayed compensatory gain during postweaning growth periods. Since resistance/susceptibility to both pathogens was clearly defined and controlled by single genes, scientists have identified the regions of the genome which possess both genes. Two groups have studied the genetic location of the F4 (K88) receptor and have mapped it to chromosome 13 (Guérin et al. 1993; Edfors-Lilja et al. 1995). Recently, Vögeli et al. (1996) mapped the receptor for F18 fimbriated E. coli to a region on chromosome 6. Efforts are currently under way to identify the genes for the receptors for both strains of E. coli.

GENERAL DISEASE RESISTANCE IN SWINE
Two studies have been conducted to determine the heritability of antibody production in response to injections with bovine serum albumin and sheep erythrocytes (Buschmann et al. 1974; Huang 1977). These studies were conducted to determine if antibody production to these antigens would be useful marker traits in selection programs to improve AMI. Heritability estimates were high for both traits (0.4-0.5), although neither study demonstrated whether these marker traits were correlated with improved disease resistance in the animals.

The realization that CMI and AMI were controlled by different genetic mechanisms prompted additional studies to further characterize the genetic mechanisms responsible for disease resistance. Selection of Yorkshire pigs for high and low "general immune response" in an attempt to produce pigs resistant to most pathogenic organisms is currently under way. The general immune response is based on a selection index of measures of AMI and CMI, as well as a measure for innate immunity (Mallard et al. 1992). The phenotypes recorded for AMI were serum IgG concentrations (IGG) and antibody response to hen egg-white lysozyme (HEWL). CMI is recorded as lymphocyte proliferation in response to concanavalin A in vitro (CONA), delayed hypersensitivity to purified protein derivative of tuberculin (DHT), and innate immune response monitored by evaluating uptake and killing of S. typhimurium. However, the heritability of the last trait was too low for effective selection, and it was eliminated from the selection index (Mallard et al. 1993).

No differences were detected for antibody avidity between the lines in the first generation (Appleyard et al. 1992) or in monocyte superoxide anion production or class II MHC antigen expression by the third generation (Groves et al. 1993). Interestingly, there were significant differences between the lines for CMI and AMI response parameters by the third generation (Groves et al. 1993). Heritability estimates for the four remaining traits were calculated after four generations of selection. The range of estimates was from 0.13 for IGG to 0.32 for HEWL, while CMI responses possessed intermediate estimates (0.26 for DHT and 0.19 for CONA) (Mallard et al. 1993). Mallard et al. (1993) also determined that the high-response line of pigs grew faster and reached market weight approximately 10 days earlier than the low-response line, indicating that the high-response line was healthier and more disease resistant. When infected with Mycoplasma hyorhinis, the high-response line also had higher serum antibody titers and lower incidences of peritonitis, pleuritis, and pericarditis, but a higher incidence of arthritis (Mallard et al. 1993).

Edfors-Lilja et al. (1994b) estimated the heritabilities of numerous measures of immune function in Yorkshire pigs. They measured white blood cell counts and determined the percentages of lymphocytes and polymorphonuclear leukocytes. In addition, the level of interferon-a was recorded to determine innate immunity to viral infections. Serum IgG concentration was determined to reflect AMI, and the CMI marker traits recorded were interleukin-2 production and lymphocyte proliferation due to concanavalin A stimulation. Estimates of heritability were moderate to high for most traits except interferon-a production (0.08) and serum IgG concentrations (0.00) (Edfors-Lilja et al. 1994b). Phenotypic correlations between CMI responses were moderate, but all other correlations were low. In a different population of pigs, Edfors-Lilja et al. (1994a) were also able to detect a quantitative trait locus (QTL) for total white blood cell count at chromosome 1 and suggestive evidence of another QTL on chromosome 8 for the same trait. They were unable to detect any loci affecting percentages of polymorphonuclear leukocytes, lymphocytes, or eosinophils in their population.

MAPPING IMMUNE RESPONSE GENES
The human genome initiative and construction of genetic maps for livestock have greatly expanded our knowledge of the mammalian genome and increased the number of mapped and sequenced porcine genes. The final goal of the human genome initiative is to completely sequence the entire genome. As this goal is approached, more genes will be identified, and as their function is determined, our knowledge of how genes interact will grow immensely. Genes responsible for immune function (MHC genes) were some of the first porcine genes mapped and sequenced. Most of the initial studies of genes involved in immune function were conducted in mice and humans, but soon after their discovery, the porcine homologues of these genes were cloned and positioned on the porcine gene map. Recent reviews have been published on the status of porcine immunoglobulins (Butler and Brown 1994), MHC (Lunney 1994), and cytokines (Murtaugh 1994). While all of these genes are known to have crucial roles in combating disease, the only repeatable association between genetic variation within one of these genes and disease resistance was the association between MHC genotype and susceptibility to T. spiralis (Madden et al. 1990, 1993).

New information is also indicating other types of genes possibly involved in disease resistance. A class of genes which have antibacterial properties has recently been identified in swine (Storici and Zanetti 1993a, b; Gudmundsson et al. 1995). When the results from this research are combined with the identification of genes coding for the receptor molecules required for E. coli pathogenicity, research into a new class of disease resistance genes will open. As the functions of additional genes sequenced from the human genome become known, our understanding of the complex relationship between host and pathogen will increase. This knowledge will allow us to identify new marker traits associated with resistance or susceptibility and to produce the genetic markers useful for marker-assisted selection.

OVERVIEW
Selection of pigs for disease resistance to certain pathogens can be quite successful. However, it is quite clear that no one gene, or form of a gene, can confer resistance to all pathogens. Selection for general disease resistance is more complex because the different methods of immune protection are governed by different genes in various locations of the genome. For example, selection for increased CMI response could lead to animals with reduced AMI response and vice versa (Biozzi et al. 1982). Disease resistance may be controlled by balancing selection to ensure that the population has adequate immune response to any kind of pathogen (Biozzi et al. 1982). In light of work by Edfors-Lilja et al. (1994b) and Mallard et al. (1993), it would appear that the forces of balancing selection are not being imposed since selection can be effective for CMI and AMI function with no negative genetic correlations between the two types of immunity detected. Therefore, appropriate selection of marker traits to measure and subsequent weighing of each measure ment in a selection index should be an effective method to improve the general disease resistance of pigs. Obviously, additional research is essential to identify marker traits that are easily measured at low cost, have moderate to high heritabilities, and are highly correlated to an animal 's ability to resist infection by various pathogens.

Response to selection for disease resistance can also be improved. Implementation of a progeny-testing system when a marker trait has a low heritability will improve the accuracy of identifying superior parents and increase the rate of genetic progress. Genetic mapping of the swine genome has already provided markers associated with disease resistance for the two strains of E. coli F18+ and F4+. Once producers are able to identify animals containing the disease-resistant allele(s) at these genomic locations, they can initiate marker-assisted selection to increase the frequency of the resistant alleles and reduce the incidence of disease in their herds. Certainly other loci will be detected and used to genetically improve the resistance of pigs in the relatively near future.

Of course, any selective process must take into account the antagonisms it may encounter. It is quite likely that during the selection of animals resistant to a pathogen, the pathogen will evolve to survive in the improved host. Since bacteria and viruses evolve more rapidly over a given period of time than mammalian organisms do, the genetic response of the pig may be offset by the rapid evolutionary response of the pathogen. A corollary to this is already seen in the reduced efficacy of antibiotics as bacteria gain antibiotic resistance. As selection programs are initiated, they must take into account the probability that some of these organisms will evolve in response to selection for resistance.

Moreover, interactions between the environment and the genetics of the pig no doubt exist. Under experimental conditions, Gray et al. (1994) found that sheep selected for resistance to nematodes had fecal egg counts four times higher than unselected sheep when challenged with extremely high levels of Haemonchus contortus larvae. If significant genetic-by-environment interactions exist, pigs selected for improved disease resistance in one environment may be more susceptible to disease when raised in another. These interactions will significantly increase the complexity of a structured selection program.

In summary, it is likely that pigs which are less susceptible to pathogens can be selected. It is also likely that there will always be diseases that affect pigs. If we endeavor to improve pigs through selection, we must consider that other pathogens or modified pathogens will develop and will need to be addressed. With a multifaceted defense relying on management practices, antibiotics, and swine genetics, we should be able to overcome these new challenges.