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Energy investment in reproduction is predicted to trade off against other necessary physiological functions like immunity, but it is unclear to what extent this impacts fitness in long-lived species. Among mammals, female primates, and especially apes, exhibit extensive periods of investment in each offspring. During this time, energy diverted to gestation and lactation is hypothesized to incur short and long-term deficits in maternal immunity and lead to accelerated ageing. We examined the relationship between reproduction and immunity, as measured by faecal parasite counts, in wild female chimpanzees (Pan troglodytes schweinfurthii) of Kibale National Park, Uganda. While we observed higher parasite shedding (counts of eggs, cysts and larvae) in pregnant chimpanzees relative to cycling females, parasites rapidly decreased during early lactation, the most energetically taxing phase of the reproductive cycle. Additionally, while our results indicate that parasite shedding increases with age, females with higher fertility for their age had lower faecal parasite counts. Such findings support the hypothesis that the relatively conservative rate of female reproduction in chimpanzees may be protective against the negative effects of reproductive effort on health. This article is part of the theme issue ‘Evolution of the primate ageing process’.
Wild female chimpanzees were studied in Kibale National Park, Uganda, between July 2015 and December 2017. Kibale is located in southwestern Uganda and comprises 795 km2 of tropical and sub-tropical rainforest [39]. The Kanyawara community of chimpanzees within Kibale was habituated for long-term study in 1987 and numbered 49–55 chimpanzees during the study period. Our sample included four subadult (aged 10–13) females, one of whom experienced a pregnancy, and 17 adult (aged 14 to 56) females from the Kanyawara community. The Ngogo community, habituated for study in 1995, averages 190–200 chimpanzees and provided samples from 60 adult females (aged 14 to 68) during this study. Although Kanyawara and Ngogo are separated by only approximately 10 km and dispersal between them is common [40], the Ngogo chimpanzees have access to significantly higher densities of food resources [38], have a higher energy balance than Kanyawara chimpanzees [41], and have reported the highest survival rates of any known wild chimpanzee community [42]. However, estimated weaning ages are similar [43,44]. Thus, the comparison between Kanyawara and Ngogo allows us to examine whether resource access moderates the effects of reproductive effort on parasite shedding. Reproductive status was categorized into four levels: cycling, pregnant, early lactation and late lactation. Pregnancy was determined by back-calculating 228 days from the birth date of the most recently born offspring [45]. Although chimpanzee infants are often not fully weaned until four to six years of age [43,44,46], nursing intensity and maternal energy costs decline precipitously by about two years postpartum [47,48]. Thus, we defined early lactation as the first two years postpartum with a living infant. Females with infants older than two years, or whose infants had died, were categorized as either cycling or in late lactation, depending on whether they had exhibited maximally tumescent sexual swellings postpartum or remained in lactational amenorrhoea, respectively. The cycling category also included nulliparous females after they exhibited their first maximally tumescent sexual swellings. The long-term reproductive effort is ordinarily quantified by examining parity. For our question, this measure was not ideal because some females experienced many early infant deaths, thus their parity was high but they had invested very little in lactation. To better address long-term reproductive effort, we calculated a measure of cumulative reproductive effort which captured the amount of time that females had spent in pregnancy and the more costly phase of early lactation. This included the sum of all months in which a female was pregnant or had a living infant under 2 years of age. In some cases (n = 20 of 81 females), females were already mothers at first identification at the onset of the long-term study, thus, it was necessary to estimate their early fertility. We used the known relationship between cumulative pregnancy and lactation months and age for those females of known parity (n = 61 females) to estimate the missing years of reproductive effort (electronic supplementary material, figure S1). The positive linear relationship between cumulative pregnancy and lactation months and age represented a yearly gain of 1.79 pregnancy and 4.76 lactation months, respectively, for the average female aged 10 to 38 years. These slopes were multiplied by the number of years the female was not observed and added to known pregnancy and lactation months for the female in question. We defined the total number of pregnancy and lactation months, observed and estimated, as cumulative reproductive effort (CRE). Among the 20 females for which an estimation of CRE was required, three of the oldest females from Ngogo had only one observed birth between them and would have required calculation for the majority, if not all, of their reproductive effort. For this reason, they were removed from analyses. Of the remaining 17 females that required estimation of a component of their CRE, a mean of approximately 7.75 years (range 2–18 years) required estimation, and their average age during the study was 43 years (range: 25–56 years). Faecal samples were preserved in the field in 10% formalin, transported to the University of Wisconsin-Madison, Madison, WI, USA, and stored at room temperature until processing. One gram of preserved faecal material was first removed from each sample and resuspended in 10% formalin. Gastrointestinal parasite eggs, cysts and larvae were sedimented using a protocol adapted from Ash & Orihel [49]. Briefly, one gram of faeces was drained of excess formalin and the sediment was washed through two sheets of grade 10 cheesecloth with 0.08% sodium chloride solution into a reservoir. The slurry was resuspended in 0.08% sodium chloride in a 15 ml tube filled to capacity and spun on a centrifuge at 500 rcf for one minute to form a pellet of eggs, cysts and larvae. The sodium chloride was then decanted, and the process repeated twice. After three washes with sodium chloride, the pellet was suspended in 10 ml of 10% formalin and 3 ml of ethyl-acetate, shaken vigorously for 30 s and then centrifuged at 500 rcf for 2 min. The resulting pellet was viewed under 10× magnification using compound light microscopes for the identification of helminth eggs and larvae. All measurements and photographs were taken under 40× objective, and final measurements were recorded in millimeters. One-half slide from each sample was processed at 40× objective for the identification of protozoan cysts and their enumeration by species as many (greater than three views where more than two parasites were visible in the field of view), moderate (at least three views where more than two parasites were present in the field of view), few (at least three views where more than one parasite was present in the field of view), rare (one to five parasites seen on entire slide) or absent (no parasite observed). Four response variables from parasitological analyses were calculated. First, we quantified parasite richness as the number of different parasite eggs, cysts and larval morphotypes identified in each faecal sample (including both helminths and protozoans). Next, we examined prevalence (i.e. presence/absence) and intensity (i.e. total egg count) of nodule worms from the genus Oesophagostomum. Nematodes of this genus are known pathogens of humans and non-human primates and can cause weakness, diarrhoea, abdominal pain, weight loss and internal lesions to the body cavity and organs in chimpanzees [50]. They are also known to occur at high prevalence in Kibale chimpanzees [51–57]. Similarly, we assessed the prevalence of Iodamoeba sp. infection, as measured by the presence or absence of cysts. Iodamoeba are protozoans that are generally considered non-pathogenic symbionts [58]. Oesophagostomum and Iodamoeba were the most prevalent helminths and protozoans, respectively, observed in our study (see Results), making them amenable to quantitative analysis to examine the influence of reproduction on relatively pathogenic versus non-pathogenic parasites. The final dataset for analysis included 78 female chimpanzees (n = 412 observations) aged 10 to 56 years old, including 21 females (n = 150 faecal samples) from Kanyawara and 57 females (n = 262 faecal samples) from Ngogo (electronic supplementary material, figure S2). Six females in the 10- to 15-year-old age class were new immigrants to the communities, which can be stressful for young females [59]. However, the new immigrants did not differ in measures of parasitism from the broader sample based on preliminary analyses, so we did not include immigration status as a fixed effect in our models. A table of descriptive statistics for fixed effects and response variables is provided in the electronic supplementary material, table S3. All analyses were conducted in R [60] with the faecal sample as the unit of analysis, and chimpanzee identity and year and month of sample collection included in all models as random effects. As measures of parasite eggs, cysts and larvae are often overdispersed among hosts, and count data often include many zeros (i.e. no infection observed), several distributions were considered. The following distributions were selected as they represented the best fit to each type of response based on plots of distributions and residuals: Poisson for parasite richness, binomial for both prevalence models and negative binomial for Oesophagostomum intensity. To examine associations between reproduction and parasite shedding, we examined our four parasite response variables in generalized linear or logistic mixed models using the lme4 [61] and glmmTMB [62] packages. All models included reproductive status (prediction 1), CRE (prediction 2), community, age in years (continuous) and season (i.e. wet and dry). Because CRE and age were intrinsically related and led to multicollinearity in our models, they could not be used in the same model in their raw forms. After conducting exploratory analyses which revealed that models with age alone, as assessed by a likelihood-ratio test, produced better fits than those with CRE alone, we transformed the CRE measure by calculating the residuals of a simple linear regression of CRE on age [63]. Using the residual CRE in the analysis allowed us to assess the influence of CRE on parasite shedding, independent from the influence of age. Residual CRE and age were both mean-centred for modelling. We also considered interaction terms between the community and reproductive status and between community and CRE in all models, addressing the prediction that habitat quality might moderate trade-offs between reproduction and parasite shedding (prediction 3). All two-way interactions were explored and eliminated from the model if they were non-significant. Significance of fixed effects from regression models was determined by Wald Chi-square tests. If a fixed effect was a factor with more than two levels, a Tukey test was employed to determine significant differences between pairs of means.
