Question

What information would you need to compare tolerance among sheep? How would you determine if one sheep was more tolerant than another? Draw a graph that illustrates the tolerance of several individual sheep.

Answer 1

Hosts may mitigate the impact of parasites by two broad strategies: resistance, which limits parasite burden, and tolerance, which limits the fitness or health cost of increasing parasite burden. The degree and causes of variation in both resistance and tolerance are expected to influence host–parasite evolutionary and epidemiological dynamics and inform disease management, yet very little empirical work has addressed tolerance in wild vertebrates. Here, we applied random regression models to longitudinal data from an unmanaged population of Soay sheep to estimate individual tolerance, defined as the rate of decline in body weight with increasing burden of highly prevalent gastrointestinal nematode parasites. On average, individuals lost weight as parasite burden increased, but whereas some lost weight slowly as burden increased (exhibiting high tolerance), other individuals lost weight significantly more rapidly (exhibiting low tolerance). We then investigated associations between tolerance and fitness using selection gradients that accounted for selection on correlated traits, including body weight. We found evidence for positive phenotypic selection on tolerance: on average, individuals who lost weight more slowly with increasing parasite burden had higher lifetime breeding success. This variation did not have an additive genetic basis. These results reveal that selection on tolerance operates under natural conditions. They also support theoretical predictions for the erosion of additive genetic variance of traits under strong directional selection and fixation of genes conferring tolerance.

Animals can defend themselves against parasites through either resistance (reducing parasite numbers, for example, by killing them) or tolerance (maintaining health as infections levels increase, for example, by repairing damage). Resistance has been well-studied in wild animals, but tolerance has been less so. We analysed data on body weight collected over 25 years on a natural population of Soay sheep, infected with parasitic gut worms. As parasite burden increased, sheep lost weight. Crucially, there was variation among individuals: some lost weight rapidly with increasing infections , whereas others lost weight slowly . The least tolerant individuals lost 4.5 kg of body weight across the range of parasite burdens that we saw, whereas the most tolerant lost only around 0.36 kg. However, variation in tolerance did not have a heritable genetic basis, so that although tolerance varied between individuals, this was not due to genetic differences. Further analysis revealed that there was natural selection on tolerance. Individuals who were more tolerant of infection produced more offspring over the course of their lives. This study shows that natural selection can act upon resistance and tolerance simultaneously in nature, a result that has implications for both human health and livestock management.

Determining the role of tolerance in host defence against nematodes under natural conditions has important implications for fundamental and applied science. First, it advances our understanding of parasite-mediated selection on hosts, for some of the most prevalent and abundant parasite taxa on Earth . Second, quantifying variation in tolerance may inform management of livestock to enhance productivity during nematode infection –. Third, the quantitative approach to studying variation in nematode tolerance applied here, in a natural animal population, may also prove useful in future studies of how variation in human health with increasing gastrointestinal nematode burdens is generated and/or maintained.

Gaining insight into how tolerance varies and affects host fitness under natural conditions (e.g., limited food, natural infection rates, diverse host and parasite genetics) requires individual-based study of a wild population. Despite this similarity, disease phase curves also have an explicit temporal component over the course of a microparasite infection, whereas we consider tolerance to be the health changes in an individual host across macroparasite infections of varying intensity. The statistical apparatus for dealing with tolerance in this way is very well-developed , whereas disease-phase curves, while currently a very useful conceptual tool , have not yet, to our knowledge, been statistically characterized.

The Soay sheep (Ovis aries) population of St. Kilda has been a model system in which to explore heterogeneity in a wide array of quantitative traits . The sheep harbour gastrointestinal nematodes, and several causes of heterogeneity in nematode resistance, including host genetics sex, age and body weight  have been identified. However, the degree of variation in host tolerance of gastrointestinal nematodes and any associations of tolerance with fitness are unknown. Here, we study tolerance in terms of changes in body weight with increasing parasite burden. Because body weight is the single biggest predictor of annual fitness through positive effects on survival and reproductive success in this population, it is an appropriate proxy (sensu ) for host fitness in our analyses. We expected that any weight-associated tolerance variation predictive of fitness of sheep would also be relevant to parasite fitness, as assumed by theory due to the persistent shedding of parasite transmission stages by tolerant individuals that survive despite high parasite burdens. With these motivations, we used longitudinal sampling of known individuals, a population pedigree, and a novel statistical workflow (see below) to quantify (i) the average association between body weight and parasite burden in the population; (ii) between-individual variation in tolerance, quantified as the slope of body weight on parasite burden; (iii) the additive genetic basis of tolerance; and (iv) the strength and direction of selection on tolerance. Our results reveal that individuals vary in their tolerance of nematode infection and that tolerance is under positive phenotypic selection through lifetime breeding success (LBS).

Mean relative LBS Estimated selection gradient -0.5 - SE 4 Tolerance WT WT-Age 1 3 Tolerance quartile

Positive phenotypic selection for increased tolerance in Soay sheep.

(A) Mean relLBS is higher in individuals that were more tolerant of infections. The plot was generated from individual estimates of tolerance slopes and relLBS from the model shown in Table 2. Individuals in the four tolerance quartiles are predicted to lose varying amounts of weight between infection levels of 0 and 2,000 strongyle eggs/gram of faeces.B) Estimated selection gradients calculated from the bivariate model of body weight (WT) and relLBS, which is shown in full in Table 2. Selection gradients were calculated for each of 1,000 posterior estimates of the individual VCV matrix as described in the text for individual variation in body weight; in the slope of body weight on FEC—that is, tolerance; and in the slope of body weight on age. Points show mean estimated selection gradient of each trait on LBS±95% CI.

Mean body weight (kg) 10.0 - - о 2000 400 800 1200 1600 Strongyle FEC (eggs/gram)

Mean, population-level tolerance of Soay sheep to gastrointestinal nematodes.

Body weight (kg) Number of individuals Number of individuals Less tolerant More tolerant 0 500 1000 2000 18 20 22 24 26 28 -4.2 -3.2 -2.4 -1.6 -0.8 Strongyle FEC (eggs/gram) Intercept of body weight (kg) Change in body weight (kg)

Significant individual-level variation in tolerance of gastrointestinal nematodes by the Soay sheep

As a result  data were collected from 1988 to 2012 and consist of complete demographic data (on annual survival and reproductive success) plus faecal egg counts (FECs) of highly prevalent gastrointestinal strongyle nematodes as a measure of parasite burden and body weight from 4,934 captures of 2,438 individuals of known age and sex born between 1980 and 2012. Around 50% of the study population are captured and sampled each August, though not necessarily the same 50%. Many of our individuals were captured across many years (up to 12), whereas some were captured only once, for instance as lambs. Data from once-captured individuals are essential because they enhance estimation of the model intercept and the statistical power for our random regression analyses . A comprehensive genetic pedigree has been constructed using data on 315 highly informative SNPs, allowing us to determine the genetic basis of body weight and of tolerance to infection. Breeding success in females is evaluated by behavioural observations of lambs and ewes and confirmed using genetic markers, whereas breeding success in males is evaluated using genetic markers (see Materials and Methods for further details on all aspects of data collection).

Longitudinal multivariate data are required to address questions pertaining to individual variation in tolerance and its fitness consequences, but determining the most appropriate statistical framework for such analyses has proved challenging . Random regression models are mixed-effects models that include one or more random slope terms alongside standard random intercept terms. These random slope terms allow estimation of the between-individual variance in a linear function: for example, tolerance may be defined as the slope of individual health or fitness on parasite burden. These models have recently been advocated as a means of quantifying and exploring individual variation in tolerance . Combining this approach with widely used pedigree-based “animal models” allows further separation of individual variation in tolerance slopes into additive genetic and environmental components allowing us to estimate genetic variance for tolerance. Multivariate versions of these models can estimate the covariance between a measured trait and an index of fitness and thus the strength and direction of selection , allowing selection on tolerance to be estimated as the covariance between the slope of health (estimated as body weight; on parasite burden and lifetime fitness. Finally, the results of these analyses allow calculation of selection gradients a measure of the strength of natural selection on a trait that is broadly used in evolutionary biology that quantifies the relative strength of selection on each trait in question. Here, we utilise this workflow to determine the extent of phenotypic and genetic variance in nematode tolerance and whether it is under natural selection in a wild mammal population.

The negative association between strongyle nematode burden and body weight is likely to arise from parasite-induced anorexia and parasite- and immune-mediated damage to the intestinal wall that causes diarrhoea and/or decreased absorption of protein . Thus, Soay sheep that lost weight slowly with increasing strongyle burden (the more tolerant individuals) may be able to maintain feeding and/or digestive efficiency in the face of increasingly heavy infections, and/or to repair damage to the gut wall. Our models control for variation in body weight due to skeletal size associated with age and sex. This means that our estimate of weight loss associated with heavier strongyle infections is likely to be due to a loss of body condition, reflected in nutritional state or fatness. In any case, the observed variation in the slope of body weight on FEC was substantial, with the most tolerant individuals losing approximately 18 g of body weight per 100 eggs per gram of faeces, and the least tolerant losing 226 g per 100 eggs per gram, a 13-fold difference. Although this is an observational study, we did account statistically for temporal  and individual differences (e.g., in behaviour and heft/spatial allegiance)  affecting strongyle exposure risk in this population. This was accomplished by fitting random effects of individual identity and year in our models, and by collecting all samples at the same time of the year. In addition, we accounted for age in all models, which is the key driver of between-individual variation in parasite infracommunity (the species composition of a host individual's parasite fauna) in this population . There is also evidence in this population that coinfection with prevalent Eimeria protozoan parasites does not affect the association between body weight and strongyle FEC . Finally, as is discussed in more detail below, all the evidence collected thus far suggests that the relatively intolerant individuals identified by this analysis are not merely paying a cost of resistance: analysis has revealed that body weight is either not significantly associated or is positively associated with antibody responses, including those specific to Teladorsagia circumcincta suggesting that nematode-resistant sheep do not pay a cost in terms of reduced body weight. We are therefore able to report that the interindividual variation in tolerance reported here is unlikely to be attributable to variation in exposure or to costs of resistance.

everal empirical studies have shown that variation in tolerance has a genetic basis: Host strains differ in their slopes of fitness or health on infection intensity . We found that variation in tolerance (the slope of body weight on FEC) does not appear to be due to additive genetic effects. Indeed, epidemiological feedbacks and positive frequency dependence, all else being equal, are expected to purge genetic variation for tolerance.As the molecular and cellular mechanisms of tolerance in animals are elucidated—and we expect that they will be, given the recent surge in interest —we will gain greater insight into the causes of this variation. A major challenge for the future will be to determine the contributions of variation in the parasite infracommunity and parasite as well as host genetics to variation in defence strategies.

Variation in the rate at which individuals lose weight with increasing strongyle FEC appears to have important selective consequences in this population. Tolerance was under positive selection in the population, with more tolerant individuals having higher LBS. Previous work on the population shows comparable positive selection for higher body weight  and greater strongyle resistance . Together, these selection analyses reveal that in this population, greater weight, resistance, and tolerance are all independently associated with greater LBS. These results clearly demonstrate that tolerance plays a major role in defence against parasite infection in wild vertebrates, varies considerably between individuals, and that this variation is under relatively strong selection through LBS.

(i) show that host body weight declines with increasing infection intensity;(ii) reveal between-individual variation in the decline in weight with infection intensity, and therefore among-individual variation in tolerance slopes; (iii) demonstrate that among-individual variation in tolerance does not have an additive genetic (heritable) basis; and (iv) reveal that individual tolerance is associated with LBS and, having accounted for selection on other correlated traits, is under relatively strong positive selection. Thus, tolerance varies between individuals and natural selection can act upon it in the wild.