Frontiers in Zoology

Over the past couple of decades, measuring immunological parameters has become widespread in studies of ecology and evolution. A combination of different immunological indices is useful for quantifying different parts of the immune system and comprehensively assessing immune function. Running multiple immune assays usually requires samples to be repeatedly thawed and re-frozen. There is some evidence that repeated freezing and thawing can affect assay results, but this has never been comprehensively studied in some common ecological immunology assays. We tested the effect of multiple (1, 2, 3, 4, 5, 10) freeze-thaw cycles on the results of four commonly used immunological assays: haemolysis-haemagglutination titres, haptoglobin concentration, bacterial killing capacity and total immunoglobulins (IgY). We tested five different bird species from four different bird orders (Passeriformes, Columbiformes, Charadriiformes and Galliformes), and we included both captive and free-living individuals. In addition, we tested for haptoglobin concentrations and the haemolysis-haemagglutination assay if re-analysing samples 1 year apart led to different results. For the haemolysis-haemagglutination assay we also tested two different sources of rabbit blood, and we compared untreated microtitre plates with plates that were “blocked” to prevent nonspecific interactions between the plate and assay reagents. Repeated freezing and thawing of plasma had no effect on lysis titres, haptoglobin concentrations, bacterial killing capacity, or total immunoglobulin levels. Agglutination titres were unaffected by up to five cycles but were lower after ten freeze-thaw cycles. For the haemolysis-haemagglutination assay and haptoglobin concentrations, re-analysing samples 1 year apart yielded highly correlated data. For the haemolysis-haemagglutination assay, the source of rabbit blood did not influence the results, and the untreated vs. blocked plates differed slightly overall, but at the individual level assay results were highly correlated. Using different rabbit blood sources or different types of microtitre plates yielded highly correlated data. Our data suggest that repeated freeze-thaw cycles do not impair assay results to the point of influencing ecological or evolutionary conclusions. Plasma samples can be safely stored in one tube and thawed repeatedly for different assays. Nevertheless, we recommend consistent treatment of samples in terms of freeze-thaw cycles or other laboratory treatments to minimize the potential for introducing a systematic bias.

Hybridization can have complex effects on evolutionary dynamics in ants because of the combination of haplodiploid sex-determination and eusociality. While hybrid non-reproductive workers have been found in a range of species, examples of gene-flow via hybrid queens and males are rare. We studied hybridization in East African army ants (Dorylus subgenus Anomma) using morphology, mitochondrial DNA sequences, and nuclear microsatellites. While the mitochondrial phylogeny had a strong geographic signal, different species were not recovered as monophyletic. At our main study site at Kakamega Forest, a mitochondrial haplotype was shared between a "Dorylus molestus-like" and a "Dorylus wilverthi-like" form. This pattern is best explained by introgression following hybridization between D. molestus and D. wilverthi. Microsatellite data from workers showed that the two morphological forms correspond to two distinct genetic clusters, with a significant proportion of individuals being classified as hybrids. We conclude that hybridization and gene-flow between the two army ant species D. molestus and D. wilverthi has occurred, and that mating between the two forms continues to regularly produce hybrid workers. Hybridization is particularly surprising in army ants because workers have control over which males are allowed to mate with a young virgin queen inside the colony.

How host manipulation by parasites evolves is fascinating but challenging evolutionary question remains. Many parasites share the capacity to manipulate host behavior increasing their transmission success. However, little is known about the learning and memory impact of parasites on their host. Wolbachia are widespread endosymbionts and infect most insect species. These bacteria are maternally transmitted and mainly alter the reproduction of hosts with weak virulence. We tested the impact of parasites (Wolbachia) on their host learning and memory capacities. To address this question we trained individuals to one direction with positive reinforcement. We compared performances between individual Wolbachia-free, Wolbachia naturally and Wolbachia artificially infected individuals. We report that in the host parasite interaction (Armadillidium vulgare/Wolbachia) naturally infected individuals Wolbachia or transinfected adult with Wolbachia are less likely to learn and memorize the correct direction with social reinforcement compared to Wolbachia-free individuals. Our results imply that Wolbachia impact in the central nervous system of their host altering the memory formation and maintenance. We conclude that host manipulation can affect cognitive processes decreasing host adaptation capacities.

Fitness is strongly related to locomotor performance, which can determine success in foraging, mating, and other critical activities. Locomotor performance on different substrates is likely to require different abilities, so we expect alignment between species’ locomotor performance and the habitats they use in nature. In addition, we expect behaviour to enhance performance, such that animals will use substrates on which they perform well. We examined the associations between habitat selection and performance in three species of Oedura geckos, including two specialists, (one arboreal, and one saxicolous), and one generalist species, which used both rocks and trees. First, we described their microhabitat use in nature (tree and rock type) for these species, examined the surface roughnesses they encountered, and selected materials with comparable surface microtopographies (roughness measured as peak-to-valley heights) to use as substrates in lab experiments quantifying behavioural substrate preferences and clinging performance. The three Oedura species occupied different ecological niches and used different microhabitats in nature, and the two specialist species used a narrower range of surface roughnesses compared to the generalist. In the lab, Oedura geckos preferred substrates (coarse sandpaper) with roughness characteristics similar to substrates they use in nature. Further, all three species exhibited greater clinging performance on preferred (coarse sandpaper) substrates, although the generalist used fine substrates in nature and had good performance capabilities on fine substrates as well. We found a relationship between habitat use and performance, such that geckos selected microhabitats on which their performance was high. In addition, our findings highlight the extensive variation in surface roughnesses that occur in nature, both among and within microhabitats.

Inter-specific comparisons of metazoan developmental mechanisms have provided a wealth of data concerning the evolution of body form and the generation of morphological novelty. Conversely, studies of intra-specific variation in developmental programs are far fewer. Variation in the rate of development may be an advantage to the many marine invertebrates that posses a biphasic life cycle, where fitness commonly requires the recruitment of planktonically dispersing larvae to patchily distributed benthic environments. We have characterised differences in the rate of development between individuals originating from a synchronised fertilisation event in the tropical abalone Haliotis asinina, a broadcast spawning lecithotrophic vetigastropod. We observed significant differences in the time taken to complete early developmental events (time taken to complete third cleavage and to hatch from the vitelline envelope), mid-larval events (variation in larval shell development) and late larval events (the acquisition of competence to respond to a metamorphosis inducing cue). We also provide estimates of the variation in maternally provided energy reserves that suggest maternal provisioning is unlikely to explain the majority of the variation in developmental rate we report here. Significant differences in the rates of development exist both within and between cohorts of synchronously fertilised H. asinina gametes. These differences can be detected shortly after fertilisation and generate larvae of increasingly divergent development states. We discuss the significance of our results within an ecological context, the adaptive significance of mechanisms that might maintain this variation, and potential sources of this variation.

Non-native species are often introduced in cities, where they take advantage of microclimatic conditions, resources provided by humans, and competitor/predator release to establish and proliferate. However, native communities in the surrounding rural or natural areas usually halt their spread through biotic resistance, mainly via top-down regulative processes (predation pressure). Here, we show an unusual commensal interaction between exotic and native bird species that favours the spread of the former from urban to rural habitats. We show how Monk parakeets Myiopsitta monachus, an invasive species often introduced in cities worldwide, associated for breeding with a much larger, native species (the white stork Ciconia ciconia) to reduce predation risk in central Spain, thus allowing their colonization of rural areas. Parakeets selected stork nests close to conspecifics and where breeding raptors were less abundant. Parakeets always flushed when raptors approached their nests when breeding alone, but stayed at their nests when breeding in association with storks. Moreover, when storks abandoned a nest, parakeets abandoned it in the following year, suggesting that storks actually confer protection against predators. Our results show how a protective-nesting association between invasive and native species can counteract biotic resistance to allow the spread of an invasive species across non-urban habitats, where they may become crop pests. Monk parakeet populations are now growing exponentially in several cities in several Mediterranean countries, where they coexist with white storks. Therefore, management plans should consider this risk of spread into rural areas and favour native predators as potential biological controllers.

One of the most prominent life-history trade-offs involves the cost of reproduction. Oxidative stress has been proposed to be involved in this trade-off and has been associated with reduced life span. There is currently an unclear relationship between oxidative cost and the reproduction-longevity trade-off. The current study, using a non-lethal and minimally invasive (only a single blood sample and no euthanasia) method, investigated whether an oxidative cost (oxidative stress) to reproduction would be apparent in two long-lived eusocial mole-rats, the naked mole-rat (NMR), Heterocephalus glaber, and the Damaraland mole-rat (DMR), Fukomys damarensis, where breeding colony members live longer than non-breeder conspecifics. We measured the direct redox balance in plasma by measuring the oxidative stress index (OSI) based on the ratio of total oxidant status and total antioxidant activity in breeders and non-breeders of both sexes, in the two species. NMR had significantly higher OSI between breeders and non-breeders of each sex, whereas DMR showed no significant differences except for total antioxidant capacity (TAC). The mode of reproductive suppression and the degree of reproductive investment in NMR may explain to some degree the redox balance difference between breeders and non-breeders. DMR show minimal physiological changes between breeders and non-breeders except for the mode of reproduction, which may explain some variations in TAC and TOS values, but similar OSI between breeders and non-breeders.

Chuột chũi Damaraland (Fukomys damarensis) là một loài động vật có vú sống dưới lòng đất, có tính xã hội cao, trong đó sự phân bổ sinh sản cực đoan với một con cái (nữ hoàng) chiếm hữu việc sinh sản trong mỗi thuộc địa. Các con cái không sinh sản trong sự hiện diện của nữ hoàng bị ức chế sinh lý đến mức không rụng trứng. Sự chặn này được cho là do sự gián đoạn trong việc tiết hormone giải phóng gonadotropin (GnRH) từ vùng dưới đồi. Để hiểu rõ hơn về các cơ chế sinh lý cơ bản của việc ức chế sinh sản ở những con cái có thứ bậc thấp, chúng tôi đã nghiên cứu sự biểu hiện của các thụ thể hormone steroid và enzyme chuyển đổi androgen aromatase trong các vùng não trước liên quan đến kiểm soát hành vi sinh sản ở các con cái sinh sản và không sinh sản từ các thuộc địa nguyên vẹn. Thêm vào đó, chúng tôi đã đưa vào phân tích các con cái đã thoát khỏi sự ức chế xã hội khi được tách ra khỏi sự hiện diện của nữ hoàng. Chúng tôi đã phát hiện ra sự biểu hiện của thụ thể androgen, thụ thể estrogen α và aromatase ở một số vùng não trước của chuột chũi Damaraland cái. Phân bố của chúng tương ứng với các phát hiện trước đó ở các loài động vật có vú khác. Việc định lượng tín hiệu lai cho thấy nữ hoàng có sự biểu hiện thụ thể androgen tăng lên so với các con cái không sinh sản và thực hiện sự không sinh sản trong hầu hết các vùng não được nghiên cứu, bao gồm vùng trước hành não giữa (MPOA), hạch chính của hạch nằm ở rìa đường cùng (BSTp), nhân trung gian bên dưới vùng dưới đồi (VMH), nhân vòng cung (ARC) và hạch amygdala giữa (MeA). Hơn nữa, các con cái sinh sản có sự biểu hiện thụ thể estrogen α tăng lên trong nhân màng ngoài tâm thất trước (AVPV) và trong MeA, trong khi đó sự biểu hiện aromatase trong AVPV giảm đáng kể so với các con cái không sinh sản. Sự vắng mặt của sự ức chế xã hội có liên quan đến sự biểu hiện thụ thể androgen tăng lên trong ARC, sự biểu hiện thụ thể estrogen α tăng lên trong MeA và BSTp và sự biểu hiện aromatase giảm trong AVPV. Nghiên cứu này cho thấy rằng sự ức chế xã hội và tình trạng sinh sản ảnh hưởng khác nhau đến hiện tượng nội tiết thần kinh của chuột chũi Damaraland cái. Mô hình biểu hiện khác nhau của thụ thể estrogen α và aromatase trong AVPV giữa các con cái sinh sản và không sinh sản hỗ trợ cho quan điểm rằng vùng này đóng vai trò quan trọng trong việc trung gian việc ức chế sinh lý ở các con cái có thứ bậc thấp.

Multidirectional interactions in social networks can have a profound effect on mate choice behavior; e.g., Poecilia mexicana males show weaker expression of mating preferences when being observed by a rival. This may be an adaptation to reduce sperm competition risk, which arises because commonly preferred female phenotypes will receive attention also from surrounding males, and/or because other males can copy the focal male's mate choice. Do P. mexicana males indeed respond to perceived sperm competition risk? We gave males a choice between two females and repeated the tests under one of the following conditions: (1) an empty transparent cylinder was presented (control); (2) another ("audience") male inside the cylinder observed the focal male throughout the 2nd part, or (3) the audience male was presented only before the tests, but could not eavesdrop during the actual choice tests (non-specific sperm competition risk treatments); (4) the focal male could see a rival male interact sexually with the previously preferred, or (5) with the non-preferred female before the 2nd part of the tests (specific sperm competition risk treatments). The strength of individual male preferences declined slightly also during the control treatment (1). However, this decrease was more than two-fold stronger in audience treatment (2), i.e., with non-specific sperm competition risk including the possibility for visual eavesdropping by the audience male. No audience effect was found in treatments (3) and (5), but a weak effect was also observed when the focal male had seen the previously preferred female sexually interact with a rival male (treatment 4; specific sperm competition risk). When comparing the two 'non-specific sperm competition risk' treatments, a very strong effect was found only when the audience male could actually observe the focal male during mate choice [treatment (2)]. This suggests that focal males indeed attempt to conceal their mating preferences so as to prevent surrounding males from copying their mate choice. When there is no potential for eavesdropping [treatment (3)], non-specific specific sperm competition risk seems to play a minor or no role. Our results also show that P. mexicana males tend to share their mating effort more equally among females when the resource value of their previously preferred mate decreases after mating with a rival male (perceived specific sperm competition risk), but this effect is comparatively weak.