S36.4: Parental care and offspring immunity

1 Dipartimento di Biologia, Università degli Studi di Milano, via Celoria 26, I-20133 Milano, Italy, fax 39 2 2362726, e-mail n.saino@imiucc.csi.unimi.it; ²Laboratoire d'Ecologie, CNRS URA 258, Université Pierre et Marie Curie, Bat. A, 7ème étage, 7 quai St. Bernard, Case 237, F-75252 Paris Cedex 05, France

In its widest commonly accepted definition, parental care is "any form of parental behaviour that appears likely to increase the fitness of a parent's offspring" (Clutton-Brock 1991). Parental care in birds can therefore start even before fertilisation, through choice of the best time for breeding, defence of a territory that will provide adequate food for the offspring and building of a nest, and continue through laying of large, high quality eggs, and provisioning of young with food and defence against predators up to offspring independence (e. g. Clutton-Brock 1991; Martin 1987, 1995; Mertens 1987).

Parental care in birds has been the subject of intensive study, mainly focusing on the relationships between offspring survival or recruitment and quality of the nesting site, hatching date and extent of parental expenditure (Lack 1950; Martin 1987; van Noordwijk et al. 1995). Perhaps not surprisingly, these studies have shown that parents performing the largest effort in providing parental care produce offspring of relatively high quality that attain large body size and body mass, and fledge or reach independence in better condition.

Experimental work has also shown, with some exceptions, that parents that expend much energy and time on parental care have lower survival prospects and/or future breeding success (e. g. Partridge 1992). Thus, parental care is actually a form of investment where parents trade their current reproductive output against other fitness components such as the probability of surviving to the next reproductive event and ability to allocate further effort in producing progeny (Roff 1992; Stearns 1992).

How does parental care translate into offspring fitness? What mechanisms mediate the effect of intense parental care on offspring survival and probability of recruitment to the breeding population? Choosing nesting sites safe from predators obviously will reduce mortality. Producing large eggs will offer hatchlings large amounts of reserves thus enhancing their survival in the first critical days of life. Timing breeding events so that offspring will enjoy the maximum availability of food when their nutritional demand peaks reduces the chances of offspring starvation. While these considerations make good intuitive sense, they might not tell us the whole story about the pathways of the effects of parental care on important components of offspring fitness.

Most papers in this symposium, as well as many recent studies in evolutionary ecology, have emphasised the evolutionary implications of parasitism (e.g. Price 1980, Loye & Zuk 1991; Clayton & Moore 1997; Møller 1997; Merino & Møller 1999; Møller & Saino 1999a, 1999b; Zuk 1999). Parasites are ubiquitous organisms that, by definition, exploit their hosts for obtaining limiting resources and therefore detract from host fitness. Given their widespread occurrence and diversity, and the threat they impose on hosts, selection must have been strong in favour of the evolution of diverse host defences particularly against virulent parasites. Birds are known to be affected by a wide array of parasites ranging from viruses and bacteria to unicellular parasites and metazoans (Loye & Zuk 1991). Some of these can have profound negative effects on individual fitness. Efficient parental care should therefore allow offspring to avoid infection or, once infection has occurred, to recognise and destroy parasites or control the effects of chronic infection. One way to avoid offspring infection is nesting in parasite-free sites or avoid re-using old nests that might harbour parasites from previous years or previous breeding attempts during the same breeding season, or choose parasite-free food. This of course might require extra parental effort and the benefits have to be traded against the costs of searching for nesting sites and building new nests or longer foraging time. The immune system has evolved to ensure discrimination between self and non-self, thus allowing recognition of potential pathogens, tolerance towards self or non-pathogenic agents, and adaptive response to antigens along two main pathways: cellular and humoral immunity (Roitt et al. 1989; Kuby 1991; Tizard 1991). The relevance of a strong immune response to parasites after infection to an individual's reproduction and survival has been demonstrated in a large number of studies on humans and other vertebrates under controlled laboratory conditions, but evidence is now also accumulating from studies on birds in the field (Roitt et al. 1996; Saino et al. 1997a; Christe et al. 1998).

Development, maintenance and functioning of the immune system, however, require energy. Recent estimates of the costs of the different components of the immune system have led to a remarkable 20-25% of basal metabolic rate for mice (V. Apanius, pers. comm.). If this estimate also applies to birds, a large proportion of the energy passed on by parents to offspring in altricial species has to be allocated to immune function. Since the amount of total energy intake that can be allocated to immunity is likely to be limited by the competing demands for other metabolic or anabolic activities during offspring ontogeny, it follows that more intense parental care, for example in terms of food provisioning, will lead to a larger absolute share of energy available to offspring immunity.

In addition, functioning of the immune system also requires specific materials, molecules that serve as promoters or regulators of an efficient immune response (Chandra & Newberne 1977; Willis & Baker 1981; Tsiagbe et al. 1987). For example, particular kinds of cellular immune responses in birds have been shown to depend on the availability in the diet of particular amino acids. Young birds subjected to amino acids deficient diets had reduced macrophage function which, in turn, can have far reaching effects on other aspects of immunity since macrophages are also involved in antigen presentation, production of cytokines, and other fundamental roles (Wan et al. 1989). Other components of the diet such as carotenoids that are not synthesised by birds and can be acquired only through ingestion might play a crucial role in regulating and promoting both cellular and humoral immunity (see Bendich 1989; Chew 1993 and references therein; Jyonouchi et al. 1994, 1995). Hence, it is not only quantity that matters, but also quality of the food provided by parents.

Adult birds are themselves faced with the problem of fending off parasites. This implies that parents have to deal not only with immunity of their offspring, but also with their own immunocompetence. A very general consequence of sexual reproduction is that the interests of parents partly differ from that of their offspring. Parents have to decide which share of limiting resources available should be allocated in order to promote their offspring immunity without severely detracting from their own immunity. Moreover, parents have to decide which of their offspring deserve the largest share of the resources critical to their immunity, ultimately basing their decision on the reproductive value of each offspring.

Basically, the study of the evolution of parental strategies in relation to offspring immunity can therefore be set in the framework of general theory on evolution of life histories and optimality. However, the focus of the attention should also be directed to a particular determinant of offspring fitness, defence against parasites and immunity, which has been mostly neglected by ornithologists and evolutionary biologists so far. In another paper in this symposium (Saino & Møller 1999a) we point out some of the possible reasons for the lack of attention to the consequences of host-parasite interactions on the coevolution of behavioural and physiological traits in populations of parasites and the hosts they exploit, and host immune system.

This being acknowledged, a few recent studies have started investigating this promising field in evolutionary biology and, among other things, shedding light on how parents can promote fitness of their offspring by enhancing their ability to produce adaptive immune responses. In the next section we will briefly review the results of these studies. In the following section we will shortly deal with possible future expansions of this research program.

A few recent studies have addressed the question of the extent to which parental care, essentially in terms of quantity and quality of food provisioned and breeding date, affects specific components of offspring immunity.

Two correlative studies on the Barn Swallow Hirundo rustica (Saino et al. 1997b; Saino & Møller, submitted) consistently showed that nestlings in large broods have lower levels of T-lymphocyte cell-mediated immunity, as indicated by swelling response to a subcutaneous injection of a mitogenic lectin, phytohaemagglutinin (PHA; see Goto et al. 1978; McCorkle et al. 1980; Lochmiller et al. 1993 for methods), than nestlings in small broods. Parents of large broods fed offspring at relatively high rates but feeding rate to each nestling decreased with brood size thus suggesting that more food per unit time was provided to each nestling in small broods (Saino et al. 1997b; Saino and Møller, unpublished results).

In a companion manipulative study (Saino et al. 1997b), nestlings of experimentally enlarged broods had lower per capita feeding rates and, correspondingly, smaller body mass to body size ratio, and a lower level of T-lymphocyte cellular response to PHA than their siblings in broods whose size was reduced. Hence, there seems to be a causal link between parental effort per offspring and immunocompetence. Interestingly, in the same cross-fostering experiment, a statistically significant effect of parentage on T-lymphocyte mediated immunity of offspring raised in different environments was observed. This result is compatible with the idea that this particular kind of immunity has a component of additive genetic variance. Alternatively, female Barn Swallows could affect immunocompetence of their offspring through the quality of the eggs they laid. Female birds are known to deliver immunoglobulins, which will represent the first line of defence for their offspring before maturation of their immune system, to the eggs they lay. Moreover, cellular immunity has been shown to depend on the quality of food received by young birds, and particularly on the content of particular amino acids in the diet. Eggs containing relatively large amounts of critical nutrients may produce offspring capable of producing large immune response some days after hatching, perhaps through a precocial and better maturation of specific immune functions.

In the studies on Barn Swallows, the quality of food received by nestlings in their first days of life clearly affected their ability to mount an immune response to PHA. When artificially provisioned with food rich in proteins, nestlings exhibited a larger immune response than nestlings that were fed only by their parents (Saino et al. 1997b). Hence, both quantity and quality of food were relevant for an efficient T-lymphocyte mediate immune response to an antigenic stimulus.

Cellular immunity of Barn Swallow nestlings was positively correlated with their body mass. Moreover, in years in which ecological conditions during the breeding season could be independently estimated to be adverse, nestlings had smaller immunocompetence than in favourable years. Finally, nestlings of second broods were less immunocompetent than those in first broods (Saino & Møller, submitted). Several studies on birds have shown that offspring hatched early in the breeding season, in first broods and in favourable years have relatively large chances of short-term survival and recruitment to the breeding population (e.g. Sorci et al. 1997 and references therein), and this was also the case also in the Barn Swallow population that we studied. In addition, offspring with larger body mass at the time of fledging usually have relatively large chances of survival (e.g. Lindén et al. 1992; Newton 1992). Hence, the results from the Barn Swallow studies suggest that the effect of rearing conditions on offspring survival and recruitment might be at least partly mediated by immune function. These considerations will serve as a basis to formulate working hypotheses in experiments in which body mass and immunocompetence are manipulated and their independent effect on survival and recruitment investigated.

Work on European Magpies Pica pica also provided evidence in favour of an effect of breeding date on nestling survival mediated by a decline in immunocompetence as breeding season progressed (Sorci et al. 1997). In a study in which first clutches were experimentally induced to fail by breaking the eggs, nestlings from replacement clutches hatched later than those from control clutches and showed weaker cellular response to a subcutaneous challenge with PHA. Although the results of this experiment are clearly consistent with the prediction of a negative effect of hatching date on immune function of offspring, they are also susceptible to alternative interpretations. Females forced to lay a replacement clutch may produce eggs of inferior quality compared to those of their first clutch for example in terms of protein content or concentration of micro-nutrients relevant to immune system maturation, regulation and function (e.g. carotenoids). In addition, laying two clutches may reduce the ability of females to provide parental care to her offspring, thus negatively affecting their nutritional condition and, hence, immunocompetence.

Immunity is a multifaceted defence system which basically implies discrimination by the host between self and non-self. Once a foreign agent for which no tolerance exists is recognised by immunocompetent cells or molecules, a complex series of physiological processes is activated, ultimately leading to a humoral or cell mediated response. An immune response to an antigen requires energy consuming cell proliferation and production of antibodies. On these premises, a straightforward prediction is that an immune response to an antigen will probably negatively affect the ability of the host to produce an efficient immune response to other antigens, since resources available for immune responses are likely to be limiting, and immune function will compete with other metabolic functions and anabolic processes during nestling ontogeny. Such a trade-off between different components of immunity has been studied in Barn Swallow nestlings immunised with an intra-peritoneal injection of a multigenic antigen in the form of a suspension of sheep red blood cells (SRBC) in phosphate buffered saline (PBS) (Saino & Møller, unpublished results). Sheep red blood cells are widely used for immunological tests (Roitt et al. 1996). Parents of broods containing some SRBC-immunised nestlings did not feed nestlings at higher rates than parents of control broods. Consistent with the prediction, SRBC-immunised nestlings showed a smaller cellular mediated response as indicated by the wing web swelling response to subcutaneous inoculation of PHA than their control siblings not immunised with SRBC. Although this study did not directly investigate the relationship between intensity of the response to the two immunogenic stimuli, it clearly suggests that nestlings that had to mount a humoral response to SRBC had reduced cellular mediated immunity, consistent with the existence of a trade-off between the two kinds of immunity.

In altricial birds body mass or size hierarchies among nestlings in a brood may result from the adoption of a brood reduction strategy which allows parents to differentially allocate parental care, mainly in the form of food provisioning, to individual nestlings depending on contingent food abundance (Stenning 1996). Such hierarchies may arise by means of hatching asynchrony, variance in egg size leading to differential size or condition among siblings, early established sexual dimorphism, or parents allocating more resources to offspring with larger reproductive value, including those that, either for their genetically based better resistance or random effects, are the least infested by parasites (e.g. Parsons 1970; Schifferli 1973; Clark & Wilson 1981). Irrespective of the mechanisms generating a hierarchy among siblings, this should produce non negligible variance among siblings in the ability to produce an immune response. Once a hierarchy has been established, it can pay parents in terms of fitness to have one or more offspring that have poor immunological defences against parasites since these nestlings will be preferentially attacked by parasites thus leaving the high ranking offspring relatively unaffected. This so called 'tasty chick' hypothesis (Christe et al. 1998) is particularly interesting because it provides a basis to interpret the occurrence of unevenly distributed virulent parasites among siblings observed in some species. Christe et al. (1998) have provided observational evidence in support of one of the main predictions of the 'tasty chick' hypothesis: House Martin Delichon urbica nestlings with the lowest immunocompetence are more likely to die before fledging. Future studies will possibly allow to test whether parasites differentially infest the least immunocompetent nestlings in a brood. On the other hand, Saino et al. (1998) could find only ambiguous evidence for a negative relationship between parasite infestation and immunocompetence. In an experiment in which the ectoparasite load of a dipteran ectoparasite in Barn Swallow nests was manipulated, parents did not increase their feeding effort. Nestlings in broods inoculated with parasites had larger concentrations of lymphocytes and eosinophils in peripheral blood and faster feather development than nestlings in control broods. Possibly as a consequence of no adjustment of feeding effort by parents to heavily parasitised offspring, lympho-proliferative response and eosinophilia were to the detriment of somatic growth and concentration of proteins in the plasma, indicating that a trade-off might exist between immune response to parasites and other aspects of nestling growth and condition.

To the best of our knowledge, only these few studies have been published, while some more are under way on the relationship between parental care and immunity of offspring in birds. The implications of host-parasite interactions on the evolution of parental care are therefore an almost virgin field of investigation and the studies we have briefly reviewed above constitute only the very first steps in this direction. The integration of immunological approaches in studies on host-parasite interactions will certainly help to clarify the effect of parasites on the evolution of parental strategies and the precise mechanisms mediating the effect of parental care on offspring survival and recruitment.

In this section we will briefly discuss a few questions that future research might be able to address.

(1) What are the specific proximate determinants of immunocompetence of young birds?

Nutritional condition as influenced by parental care and sib competition may obviously play a central role in determining the condition of animals (Chandra & Newberne 1977; Gershwin et al. 1985); however, ascertaining which diet components affect specific aspects of immunity will require carefully conducted experiments. Availability of proteins has been shown to affect immunocompetence in birds (e.g. Lochmiller et al. 1993). However, micro-nutrients such as carotenoids might also play a role in stimulation and regulation of immune function (Bendich 1989; Chew 1993). In addition, maternal effects such as the amount and quality of nutrients in the eggs could also be involved and their detection will require manipulation of female condition before laying.

(2) Do parents trade immunocompetence of offspring against their own ability to mount adaptive immune responses to parasites? If food is limiting immune responses of both offspring and parents, conditions exist for an inter-generational conflict of interest. Under such circumstances parents may have to decide how much of their crucial nutrients they should allocate to their offspring without compromising their own immune functions. Parasites are ubiquitous and can severely affect condition and survival of both parents and offspring. The need for efficient immune defences is therefore expected to exert a powerful selective effect in shaping the evolution of optimal parental strategies (Saino et al. 1999).

(3) Does current immune defence of offspring influence allocation of parental care to individual offspring? Offspring survival is a matter of strong immune responses. When ecological conditions prevent parents from providing adequate care to all their progeny, they are expected to invest differentially in relatively more immunocompetent offspring since they will ensure the largest fitness reward.

(4) Do parents have reliable clues in order to assess offspring immune condition and level of parasite infestation? In order to perform optimal parental care, parents must be able to assess current level of infection and immunological status of their individual offspring. On the other hand, offspring are selected to manipulate their parents in order to obtain the maximum possible share of care, at least to the extent that reduction of care to nest mates does not negatively affect their own inclusive fitness. A reliable signal is one that cannot be faked by the signaller. Parents are therefore expected to rely only on nestling signals that honestly reveal offspring condition. A new model of honest signalling of condition by nestlings is centred on carotenoids, which are the determinants of the yellow-red colour of the gape of nestlings of some species. Carotenoids have the dual function of natural pigments (Goodwin 1984) and promoters and regulators of immune function. In order to assess current level of infection of offspring, parents may assess gape colour as an indication of the amount of carotenoids that can be stored in peripheral tissues of offspring. Other things being equal, nestlings with brightly red gapes signal that few carotenoids are being used for immunity and, thus, freedom from parasites. This idea stems from the observation that parasitism leads to a reduction in carotenoid based coloration of feathers and exposed skin surfaces in poultry and other birds (e.g. Bletner et al. 1966, Yvore & Mainguy, 1972; Kowalski & Reid, 1970; see also Goodwin, 1984).

(5) Does offspring sex matter for parental investment in offspring immune function? Recent studies have shown that birds can manipulate the sex ratio of their offspring in relation to ecological conditions during breeding or sexual attractiveness of their father (e.g. Ellegren et al. 1997; Komdeur et al. 1997). Generally speaking, male vertebrates appear to be more susceptible to parasites than females possibly owing to sexual differences in profiles of immunosuppressive androgens (Folstad & Karter 1992). Since sexual dimorphism in hormonal profiles and, thus, susceptibility to parasitism may become established early in life, parents may invest differentially in offspring of the two sexes. Such differential investment may depend on parental condition or the differential reproductive value of sons and daughters which, in turn, will depend on contingent ecological conditions and the abundance of parasites. Adjustment of sex ratio may occur both at fertilisation and after hatching.

(6) How far reaching in a bird's life are the consequences of parental care it experienced on the development and functioning of its immune system? This highly speculative question is also relevant for the study of evolution of parental care since the duration of these effects will affect the intensity of selection in favour of parental care oriented towards favouring immunocompetence of offspring.

(7) Does parental care affect natal dispersal ability via an effect on offspring immunity? Dispersal has both advantages and costs. For example, one frequently invoked benefit of breeding far from the natal area is avoidance of deleterious effects of inbreeding on offspring fitness (e.g. Greenwood 1980; Johnson & Gaines 1990). Among several potential costs, one that has received almost no mention in the evolutionary literature is that dispersing individuals might have to face a parasite fauna different from that to which they have been exposed in their natal area, and to which they have acquired resistance. A rather frequent observation in parasitological studies is that prevalence (proportion of infected hosts) and abundance of individual parasite species can greatly vary between populations and even between demes of a single population. Only individuals in prime immunological condition may be able to sustain the cost of being infected by new parasite strains encountered after dispersal. Any of the effects of parental care on offspring immunity which lasts long enough to affect immunocompetence after dispersal may therefore affect an individual's decision about dispersal. Thus, parental care may promote dispersal ability and offspring fitness if dispersal is beneficial for example in terms of outbreeding.

This line of reasoning, however, could also be reversed. Parasites have shorter generation times than their hosts and, at any given moment, are more likely than their hosts to be a step ahead in the coevolutionary arms race. The case could therefore be made that a local strain of parasites is more adapted to exploit the population of hosts were it lives than hosts belonging to any other population or deme (Gandon et al. 1996). Under these circumstances, while paying the costs of dispersal (in terms, for example, of unfamiliarity with a new environment), a dispersing individual may benefit from being exposed to parasites not particularly adapted to exploit it. On the other hand, philopatric individuals may be those with an efficient immune system and benefit from living and breeding in a familiar environment. If anything, current evidence argues in favour of this latter scenario. Indeed, the largest recruitment rates into the natal populations is that of individuals born early in the season (Lack 1950, 1968), and immunocompetence of offspring has been shown to decline as the season progresses (Sorci et al. 1997; Saino & Møller, submitted). These findings suggest that it is offspring with relatively large immunocompetence that return to their natal area. Properly designed experiments in which, for example, eggs or hatchlings are cross-fostered between nests in different demes will clarify if offspring are less susceptible to parasitism once translocated to an area different from their original one than nestlings that are exposed to parasite strains typical of their parental area.

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