Wild Mammalian Populations Research Articles

Use of lure sticks for non‐invasive genetic sampling of European wildcat populations: lessons learnt and hints for future insights

Abstract Non‐invasive genetic sampling is an increasingly common approach in wildlife research. It allows the gathering of first‐hand data on wild mammalian populations without capturing or handling individuals. For this reason, it has proved to be particularly useful when applied to elusive species living at low population densities and/or hard to identify in the field. The European wildcat represents an interesting case study in this respect. Several papers have been produced in the last decades, in which non‐invasive genetic sampling has been applied. Nevertheless, evidence from different case studies presents a complex scenario, where the efficiency of the method can vary considerably. This paper aimed to analyse possible interpretations of such differences and to identify potential drivers and barriers. 20 papers on the subject have been reviewed and compared, although differences in several details reported in the examined papers limited an in‐depth comparison. The review showed that the overlap of the study period with the reproductive season does not affect the final results of lure stick hair sampling research on the European wildcat. Moreover, valerian lure sticks generally provided positive results in the Continental ecoregion, whereas, in the Mediterranean and Atlantic regions, outcomes were absent or very scarce. Most of the other working hypotheses remain still plausible, despite not yet being definitely provable. Setting up future wildcat monitoring schemes based on effective non‐invasive genetic sampling in different biogeographical regions of Europe is certainly a scope to be pursued. Some suggestions are provided in this respect (e.g. the set of parameters needed to allow further comparisons; the need to test other types of attractants, to make the application of the method possible where the use of valerian was proven to be inefficient or scarcely efficient, in order to allow a better comparison of future results).

Do Wildlife Crossings Mitigate the Roadkill Mortality of Tropical Mammals? A Case Study from Costa Rica

Although Central America is one of the most biologically diverse regions in the world, booming road construction is driving roadkill mortality that threatens to alter the demography, genetic diversity, and viability of wild mammalian populations. Costa Rica has the highest road density in Central America, but the effectiveness of wildlife crossings in mitigating roadkill mortality has not been assessed with controlled experimental studies. Hacienda Baru National Wildlife Refuge is located along a critical biological corridor bisected by one of the busiest highways in the country. The first wildlife underpasses in Costa Rica were built here in 2010 as part of the expansion of Route 34 and subsequently documented by camera-trap and roadkill surveys. Using a control-impact design, we demonstrate a considerable reduction in wildlife mortality in the presence of the crossing structures compared with controls, with the underpasses and overpasses eventually used by 21 mammalian species. Some species made use of the structures right away, while others took over a year to learn to use them. This is the first controlled study in Central America to document the effectiveness of wildlife crossings and provides evidence that well-designed mitigation structures can contribute to wildlife-friendly roadways.

Age of first infection across a range of parasite taxa in a wild mammalian population.

Newborn mammals have an immature immune system that cannot sufficiently protect them against infectious diseases. However, variation in the effectiveness of maternal immunity against different parasites may couple with temporal trends in parasite exposure to influence disparities in the timing of infection risk. Determining the relationship between age and infection risk is critical in identifying the portion of a host population that contributes to parasite dynamics, as well as the parasites that regulate host recruitment. However, there are no data directly identifying timing of first infection among parasites in wildlife. Here, we took advantage of a longitudinal dataset, tracking infection status by viruses, bacteria, protists and gastro-intestinal worms in a herd of African buffalo (Syncerus caffer) to ask: how does age of first infection differ among parasite taxa? We found distinct differences in the age of first infection among parasites that aligned with the mode of transmission and parasite taxonomy. Specifically, we found that tick-borne and environmentally transmitted protists were acquired earlier than directly transmitted bacteria and viruses. These results emphasize the importance of understanding infection risk in juveniles, especially in host species where juveniles are purported to sustain parasite persistence and/or where mortality rates of juveniles influence population dynamics.

Free-ranging livestock influence species richness, occupancy, and daily behaviour of wild mammalian species in Baluran National Park, Indonesia

In some developing countries, human activities in protected areas threaten wildlife populations and their habitats. This study was conducted to understand the influences of free-range livestock on the wild mammalian population in Baluran National Park, Indonesia. There were 3852 and 1156 cows and goats, respectively, and livestock occupied an area of approximately 55.96km2. The species richness and probability of occupancy in areas with and without livestock were assessed, and the spatial co-occurrence and overlap of daily activity patterns among mammalian species and livestock were measured. A total of 39 camera traps were installed from August 2015 to January 2016, collecting 23,021 independent photographs. In areas with livestock, the number of mammal species (eight) was lower than in areas without livestock (11 species). The most affected species were the large herbivores Bubalus bubalis and Bos javanicus, and a large carnivore Panthera pardus, all of which were absent in areas with livestock. Regardless of the feeding guild, the probability of occupancy of almost all species declined in areas with livestock, except the medium herbivores Paradoxurus hermaphroditus and Hystrix javanica that showed a higher probability of occupancy. The species whose probability of occupancy declined were the carnivores Cuon alpinus and Prionailurus bengalensis; the herbivores Rusa timorensis, Tracypithecus auratus and Muntiacus muntjak; and the omnivore Sus scrofa. In the presence of livestock, R. timorensis and S. scrofa changed their activities from diurnal to nocturnal. Livestock affected most wild mammals in several ways, including by reducing the species richness, lessening the probability of occupancy and changing the daily activity patterns of many animals. This research recommends a significant reduction in the size of the range area for domestic livestock.

Natural epigenetic variation in the female great roundleaf bat (Hipposideros armiger) populations

Epigenetic modifications are considered to have an important role in evolution. DNA methylation is one of the best studied epigenetic mechanisms and methylation variability is crucial for promoting phenotypic diversification of organisms in response to environmental variation. A critical first step in the assessment of the potential role of epigenetic variation in evolution is the identification of DNA methylation polymorphisms and their relationship with genetic variations in natural populations. However, empirical data is scant in animals, and particularly so in wild mammals. Bats are considered as bioindicators because of their sensitivity to environmental perturbations and they may present an opportunity to explore epigenetic variance in wild mammalian populations. Our study is the first to explore these questions in the female great roundleaf bat (Hipposideros armiger) populations using the methylation-sensitive amplified polymorphism (MSAP) technique. We obtained 868 MSAP sites using 18 primer combinations and found (1) a low genomic methylation level (21.3 % on average), but extensive DNA methylation polymorphism (90.2 %) at 5'-CCGG-3' sites; (2) epigenetic variation that is structured into distinct between- (29.8 %) and within- (71.2 %) population components, as does genetic variation; and (3) a significant correlation between epigenetic and genetic variations (P < 0.05). These results may also apply to other wild mammalian populations. The possible causes for the correlation between epigenetic and genetic variations are discussed.

Interplay between plasma oxidative status, cortisol and coping styles in wild alpine marmots, Marmota marmota

Variation in how individuals cope behaviourally and physiologically with stressors is widespread and can have a significant impact on life-history traits and fitness. Individual coping styles are characterised by differential behavioural and adrenocortical reactivity to various challenges. As stress hormones can affect the production of reactive chemical species and the antioxidant status, individuals with different coping styles may differ also in oxidative status. Field studies on wild mammalian populations are few in number and none so far has simultaneously tested the relationship between coping style, adrenocortical reactivity and oxidative status in the same individuals. We measured individual variation in coping styles along a proactive-reactive continuum together with variation in baseline and stress-induced plasma oxidative damage, plasma non-enzymatic antioxidant capacity and cortisol in wild alpine marmots, Marmota marmota. Confirmatory path analysis revealed that different coping styles are accompanied by different baseline and stress-induced plasma oxidative statuses. Our findings also highlight the potential role of cortisol as a mediator of such differences.

Age-specific breeding success in a wild mammalian population: selection, constraint, restraint and senescence

The Selection, Constraint, Restraint and Senescence Hypotheses predict how breeding success should vary with age. The Selection Hypothesis predicts between-individual variation arising from quality differences; the other hypotheses predict within-individual variation due to differing skills or physiological condition (Constraint), residual reproductive lifespan (Restraint), or somatic and reproductive investment (Senescence). Studies tend to focus on either the initial increase in breeding success or later decrease; however, both require consideration when unravelling the underlying evolutionary processes. Additionally, few studies present genetic fitness measures and rarely for both sexes. We therefore test these four hypotheses, which are not mutually exclusive, in a high-density population of European badgers Meles meles. Using an 18-year data set (including 22 microsatellite loci), we show an initial improvement in breeding success with age, followed by a later and steeper rate of reproductive senescence in male than in female badgers. Breeding success was skewed within age-classes, indicating the influence of factors other than age-class. This was partly attributable to selective appearance and disappearance of badgers (Selection Hypothesis). Individuals with a late age of last breeding showed a concave-down relationship between breeding success and experience (Constraint Hypothesis). There was no evidence of abrupt terminal effects; rather, individuals showed a concave-down relationship between breeding success and residual reproductive lifespan (Restraint Hypothesis), with an interaction with age of first breeding only in female badgers. Our results demonstrate the importance of investigating a comprehensive suite of factors in age-specific breeding success analyses, in both sexes, to fully understand evolutionary and population dynamics.

Gestation length variation in a wild ungulate

Summary 1. In species inhabiting a seasonal environment, timing of birth may be highly correlated with fitness. However, we know little about the relative effects of conception date and gestation length in determining birth date in wild mammalian populations. 2. Here, we tested for variation in gestation length using long-term data from a wild ungulate population (red deer, with observations running from 1971 to 2007). High March temperatures during gestation were associated with shorter average gestation length (a decrease of 0·77 days for every degree warmer), but we found no significant correlations with female age, recent reproductive history or offspring sex. 3. We also tested whether early or late conceiving females adjusted gestation length to give birth closer to an optimum time. Simply fitting conception date as a fixed effect in a model of gestation length is not statistically sound as errors in determining conception date could drive an observed negative covariance; consequently we also approached this problem using a novel technique of bivariate models. We found that late conception was indeed correlated with shorter gestation length, but this was not repeatable within females, suggesting either that the observed association may have been a statistical artefact or that gestation length adjustment may only be apparent on the rare occasions that females conceive outside an optimum time window. 4. Although there were no consistent differences between females in gestation length, there were differences between males in the gestation lengths of females with whom they mated. This possibly reflects a local environment effect, grouping females who were primarily in the same area during conception and gestation. 5. We conclude that observed variation in birth date is predominantly due to systematic variation in conception date as opposed to gestation length. Additionally, we highlight the broad range of reproductive tactics utilized by a number of closely related species, and consequently the difficulties inherent in cross-species comparisons of gestation length.

Social and genetic interactions drive fitness variation in a free-living dolphin population

The evolutionary forces that drive fitness variation in species are of considerable interest. Despite this, the relative importance and interactions of genetic and social factors involved in the evolution of fitness traits in wild mammalian populations are largely unknown. To date, a few studies have demonstrated that fitness might be influenced by either social factors or genes in natural populations, but none have explored how the combined effect of social and genetic parameters might interact to influence fitness. Drawing from a long-term study of wild bottlenose dolphins in the eastern gulf of Shark Bay, Western Australia, we present a unique approach to understanding these interactions. Our study shows that female calving success depends on both genetic inheritance and social bonds. Moreover, we demonstrate that interactions between social and genetic factors also influence female fitness. Therefore, our study represents a major methodological advance, and provides critical insights into the interplay of genetic and social parameters of fitness.

MHC screening for marsupial conservation: extremely low levels of class II diversity indicate population vulnerability for an endangered Australian marsupial

Variation at genetic regions of functional significance is a new focus in conservation genetics. One of the prime candidates for such studies is the major histocompatibility complex (MHC). This gene region, with its critical role in immune response in vertebrates, is thought to have a significant impact on population fitness. The screening of diversity at MHC regions in non-model organisms presents a number of challenges that need to be overcome before its widespread incorporation into conservation programs. Here we present an approach and data for the screening of diversity at a class II MHC locus in wild and captive populations of the western barred bandicoot (Perameles bougainville). Primers developed to amplify the DAB gene in the tammar wallaby were modified to amplify the same gene in western barred bandicoots. Alleles were identified by cloning and sequencing of PCR products. Screening of all populations was carried out via high resolution melt analysis (HRM) and revealed just two sequence copies for this gene present in every individual from all populations. Alignment of translated amino acid sequences show that these variants contain many conserved elements indicative of functional MHC class II genes and suggest that they represent separate duplicated loci. Such a low level of MHC diversity is surprising for wild mammalian populations and exposes this species to increased vulnerability to novel disease pathogens.

Tracking mtDNA Heteroplasmy through Multiple Generations in the North Atlantic Right Whale (Eubalaena glacialis)

Mitochondrial heteroplasmy has been identified in a variety of species and can result from either paternal leakage, whereby sperm mitochondria enter the ova during fertilization, or more commonly by the "survival" and proliferation of mutant variants within an organism. From an evolutionary perspective, this process represents the generation of new mitochondrial diversity within a species. Although this has been documented in some mammalian species, it has been reported from relatively few wild mammalian populations and in no wild nonhuman population has the transfer and segregation of mitochondrial heteroplasmy been tracked through multiple generations. We report on the first case of the identification and tracking of mitochondrial control region heteroplasmy through 3 generations in the North Atlantic right whale, Eubalaena glacialis. We also identify the full segregation to the mutant variant within a single generation and thus the development of a new haplotype (haplotype G) in a maternal lineage of this endangered species. Witnessed here is the generation of mitochondrial diversity in a genetically depauperate species.

Is ITS-2 rDNA suitable marker for genetic characterization of Sarcoptes mites from different wild animals in different geographic areas?

The present study examined the relationship among individual Sarcoptes scabiei mites from 13 wild mammalian populations belonging to nine species in four European countries using the second internal transcribed spacer (ITS-2) of nuclear ribosomal DNA (rDNA) as genetic marker. The ITS-2 plus primer flanking 5.8S and 28S rDNA (ITS-2+) was amplified from individual mites by polymerase chain reaction (PCR) and the amplicons were sequenced directly. A total of 148 ITS-2+ sequences of 404 bp in length were obtained and 67 variable sites were identified (16.59%). UPGMA analyses did not show any geographical or host-specific clustering, and a similar outcome was obtained using population pairwise Fst statistics. These results demonstrated that ITS-2 rDNA does not appear to be suitable for examining genetic diversity among mite populations.

Aryl hydrocarbon receptor (AHR) and AHR nuclear translocator (ARNT) expression in Baikal seal ( Pusa sibirica) and association with 2,3,7,8-TCDD toxic equivalents and CYP1 expression levels

Most toxic effects of 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) and related planar halogenated aromatic hydrocarbons (PHAHs) are mediated by ligand-activated aryl hydrocarbon receptor (AHR) signaling pathway. To understand the regulation mechanism of AHR and AHR nuclear translocator (ARNT) expression in wild Baikal seal ( Pusa sibirica) population contaminated by PHAHs, the present study investigated hepatic mRNA expression levels of AHR and its heterodimer, ARNT genes, in association with biological index (age, gender and body weight), PHAH accumulation and expression levels of cytochrome P450 (CYP) 1A and 1B. While there was no gender difference, the AHR mRNA expression levels were increased with ages ( p = 0.014) and body weights ( p = 0.015), indicating that AHR expression might be affected by these biological factors. The AHR mRNA expression levels exhibited significant positive correlations with total TEQs and most of individual congener TEQs derived from polychorinated dibenzo- p-dioxins, dibenzofurans and non- ortho coplanar polychorinated biphenyls (PCBs), indicating the transcriptional up-regulation of AHR expression by these congeners. On the other hand, there was no significant correlation between individual TEQs from mono -ortho coplanar PCBs and AHR expression. These results imply the structure-related transcriptional activity of AHR among PHAHs congeners. AHR mRNA levels showed positive correlations with both CYP1A protein ( p = 0.039) and CYP1A1 mRNA expression levels ( p = 0.046). In contrast to AHR expression, neither the total nor individual congener TEQs influenced ARNT at the transcriptional level. ARNT mRNA showed significant negative correlations with CYP1A/1B protein ( p = 0.027 and p = 0.006) and CYP1A1 mRNA expression levels ( p = 0.039), implying the existence of different transcriptional regulation between AHR and ARNT genes and negative regulation by CYP1A/1B-mediated signaling pathways. The present findings may render significant insight on the basic mechanisms underlying regulation of AHR and ARNT expressions associated with biological factors and PHAH exposure in wild mammalian populations.

Induction of hepatic P450-dependent monooxygenase in feral mice from a PCDD/PCDF-contaminated area

Increased PCDD/PCDF levels were detectable in liver of feral mice (Microtus arvalis) living in PCDD/PCDF contaminated areas (up to 27800 ng TE/kg soil) of a former cable pyrolysis plant, in comparison with animals from a control area. The pattern of PCDD/PCDF congeners showed marked differences between soil and liver. Hepatic 7-ethoxyresorufin O-deethylase (EROD; a functional parameter for PCDD/PCDF-inducible cytochrome P450IA1) was increased in feral mice in relation to the degree of soil contamination with PCDD/PCDF. The data suggest that hepatic EROD activity is a useful bioindicator of the exposure of wild mammalian populations with PCDD/PCDF.

Heterogeneity of amino acid sequence in hippopotamus cytochrome c.

The amino acid sequences of chymotryptic and tryptic peptides of Hippopotamus amphibius cytochrome c were determined by a recent modification of the manual Edman sequential degradation procedure. They were ordered by comparison with the structure of the hog protein. The hippopotamus protein differs in three positions: serine, alanine, and glutamine replace alanine, glutamic acid, and lysine in positions 43, 92, and 100, respectively. Since the artiodactyl suborders diverged in the mid-Eocene some 50 million years ago, the fact that representatives of some of them show no differences in their cytochromes c (cow, sheep, and hog), while another exhibits as many as three such differences, verifies that even in relatively closely related lines of descent the rate at which cytochrome c changes in the course of evolution is not constant. Furthermore, 10.6% of the hippopotamus cytochrome c preparation was shown to contain isoleucine instead of valine at position 3, indicating that one of the four animals from which the protein was obtained was heterozygous in the cytochrome c gene. Such heterogeneity is a necessary condition of evolutionary variation and has not been previously observed in the cytochrome c of a wild mammalian population.

Spinal arthritis (spondylosis deformans) in the red fox, Vulpes vulpes, with some methodology of relevance to zooarchaeology

The skeletons of 252 adult foxes killed in suburban London were used to study the development of spondylosis deformans, a condition of the spinal column previously unrecorded in the Red fox. Although the average age of the specimens was only one year nine months, 34·5% of the population were found to be affected with spondylosis. The distribution of osteophytes on the vertebral column is shown, and factors such as age and trauma, which affect the development of the condition, are examined. The significance of such well-developed pathological conditions in a wild mammalian population is discussed.

The Induction of Tolerance of Skin Homografts in Rats with Pooled Cells from Multiple Donors

Summary An investigation has been made of the possibility of conferring tolerance in respect of skin homografts from any donor in a heterogeneous rat population by inoculation of newborn animals with pooled cell suspensions prepared from multiple, randomly selected donors. Subjects for these experiments were Wistar rats, from a large, closed, but noninbred colony, whose heterogeneity was such that the median survival time of skin homografts exchanged between them was about 12 days, although a small proportion of the animals (10%) were genetically tolerant, accepting their grafts for longer than 50 days. Subcutaneous injection of newborn animals with 15,000,000 to 20,000,000 pooled splenic cells from standard panels of 10 donors was almost ineffective in prolonging the lives of test skin homografts, but following intravenous inoculation, the grafts on about 30% of the subjects lived for longer than 50 days—an arbitrary criterion of a high degree of tolerance. Similar dosages of pooled newborn or infant donor splenic cells conferred a high degree of tolerance upon about 60% of the subjects and pooled adult bone marrow cells were even more effective since more than 80% of the animals were highly tolerant. Evidence is presented that if newborn Wistar rats are injected with cells from newborn or adult spleens or from adult bone marrow prepared from the derived inbred Lewis strain, instead of from pooled Wistar donors, then the proportion that became tolerant of Lewis grafts (60–70%) does not differ significantly according to the type of cell inoculated, so that within the dosage limits employed, cells from infant and adult spleen, and from adult bone marrow, must be considered as potentially equally effective in conferring tolerance. It is argued that the different results obtained with the different tissues when pooled cells from heterogeneous donors were employed are expressions of the varying extents to which immunologic interactions occurred on the part of immunologically competent, lymphoid-type cells present in the inoculums. These reactions may have led to the destruction of all cells of some donor genotypes before they had had an opportunity to induce tolerance in respect of some important transplantation antigens of the population. The high degree of efficacy of pooled marrow is attributable to its very low content of immunologically competent cells, whereas that of infant spleen probably depends upon the fact that here the majority of the abundant lymphoid cells are too immature to interact, or may even become tolerant of many of the “foreign” antigens present. Evidence is presented that the “polyvalent” tolerance induced by pooled donor cells is highly colony-specific; although it extends to the majority of members of the colony, and also to members of the derived Lewis inbred strain, it is completely ineffective in respect of skin homografts derived from unrelated chocolate B.N. inbred strain donors. It is suggested on the basis of these findings that the probability of being able to confer polyvalent tolerance in a truly wild mammalian population is extremely remote because of the large number of different histocompatibility genes present that would have to be represented in the pooled, tolerance-conferring inoculums. During the course of this study evidence has been obtained that, contrary to the accepted view, the duration of the tolerance-responsive period in the rat closely resembles that characteristic of the mouse. A preliminary account of “runt disease”—the outcome of the reaction of inoculated adult lymphoid tissue cells against a homologous infant host—as it expresses itself in rats is given.