Phylogenetic analysis of a novel Hepatozoon species (Hepatozoon sp. SK3) and an additional yet unknown Hepatozoon species (Hepatozoon sp. BV2) besides H. erhardovae in small rodents from Central Europe.

Hepatozoon spp. are tick-borne apicomplexan parasites of terrestrial vertebrates that occur worldwide. Tissue samples from small rodents and their parasitizing fleas were sampled for molecular detection and phylogenetic analysis of Hepatozoon-specific 18S rRNA gene region. After alignment and tree inference the Hepatozoon-sequences retrieved from a yellow-necked mouse (Apodemus flavicollis) placed into a strongly supported single clade demonstrating the presence of a novel species, designated Hepatozoon sp. SK3. The mode of transmission of Hepatozoon sp. SK3 is yet unknown. It is important to note that this isolate may be identical with the previously morphologically described Hepatozoon sylvatici infecting Apodemus spp.; however, no sequences are available for comparison. Furthermore, the previously reported variants Hepatozoon sp. BV1/SK1 and BV2/SK2 were detected in bank voles (Clethrionomys glareolus). It has been suggested that these variants should be identified as Hepatozoon erhardovae leading to the assumption that BV1 and BV2 are paralogous 18S rRNA gene loci of this species. Evidence has also been presented that fleas are vectors of H. erhardovae. In this study, we show with high significance that only the Hepatozoon sp. BV1 variant, but not BV2, infects the studied flea species Ctenophthalmus agyrtes, Ctenophthalmus assimilis, and Megabothris turbidus (p < 0.001). This finding suggests that Hepatozoon sp. BV2 represents an additional species besides H. erhardovae (= Hepatozoon sp. BV1), for which alternative arthropod vectors or non-vectorial modes of transmission remain to be identified. Future studies using alternative molecular markers or genome sequencing are required to demonstrate that BV1/SK1 and BV2/SK2 are different Hepatozoon species.

Lead content in soil, plants, rodents, and amphibians in the vicinity of a heating plant's ash waste.

This study supplements previous research focused on environmental condition in the vicinity of waste ash material. The main aim of our study was the comparative analysis of lead levels in soil, plant, and animal organisms in the area of the tailings pond and surroundings, using x-ray. Findings confirm that the level of Pb in the top layer of soil is in the range of 20-135 ppm. Lead content in Calamagrostis plant tissues was confirmed only at the tailings pond area, with the highest lead concentrations measured in above-ground components; stems with blooms followed by roots and ground floor sheats. The livers, kidneys, and hearts of Apodemus flavicollis were examined, with findings of higher values in the tailings pond area than in the reference site, and average values of 14.5 ppm for livers, 16.0 ppm for kidneys, and 16.6 ppm for hearts. No significant differences were discovered based on sex and body length/body weight of A. flavicollis individuals. Values for Bombina variegata liver tissue reached an average of 12.3 ppm for individuals caught in a water reservoir without ash sediments, versus 15.7 ppm in those trapped by the edge of then tailings pond area. Females had lower concentrations of lead than males, but with no statistically significant differences found. Despite lower lead levels in soil and ash than expected, concentrations in mammalian and amphibian organs suggest a possible transition of this element into the food chain, and therefore further research in this area is recommended.

Population structure of Apodemus flavicollis and comparison to Apodemus sylvaticus in northern Poland based on RAD-seq.

BACKGROUND: Mice of the genus Apodemus are one the most common mammals in the Palaearctic region. Despite their broad range and long history of ecological observations, there are no whole-genome data available for Apodemus, hindering our ability to further exploit the genus in evolutionary and ecological genomics context. RESULTS: Here we present results from the double-digest restriction site-associated DNA sequencing (ddRAD-seq) on 72 individuals of A. flavicollis and 10 A. sylvaticus from four populations, sampled across 500 km distance in northern Poland. Our data present clear genetic divergence of the two species, with average p-distance, based on 21377 common loci, of 1.51% and a mutation rate of 0.0011 - 0.0019 substitutions per site per million years. We provide a catalogue of 117 highly divergent loci that enable genetic differentiation of the two species in Poland and to a large degree of 20 unrelated samples from several European countries and Tunisia. We also show evidence of admixture between the three A. flavicollis populations but demonstrate that they have negligible average population structure, with largest pairwise FST<0.086. CONCLUSION: Our study demonstrates the feasibility of ddRAD-seq in Apodemus and provides the first insights into the population genomics of the species.

"Parasite turnover zone" at secondary contact: A new pattern in host-parasite population genetics.

We describe here a new pattern of population genetic structure in a host-parasite system that can arise after secondary contact of previously isolated populations. Due to different generation times, and therefore different tempos of molecular evolution, the host and parasite populations reach different degrees of genetic differentiation during their separation (e.g., in refugia). Consequently, upon secondary contact, the host populations are able to re-establish a single panmictic population across the area of contact, while the parasite populations stop their dispersal at the secondary contact zone and create a narrow hybrid zone. From the host's perspective, the parasite's hybrid zone functions on a microevolutionary scale as a "parasite turnover zone": while the hosts are passing from area A to area B, their parasites turn genetically from the area A genotypes to the area B genotypes. We demonstrate this novel pattern with a model composed of Apodemus mice and Polyplax lice by comparing maternally inherited markers (complete mitochondrial genomes, and complete genomes of the vertically transmitted symbiont Legionella polyplacis) with single nucleotide polymorphisms derived from louse genomic data. We discuss the circumstances that may lead to this pattern and possible reasons why it has been overlooked in studies of host-parasite population genetics.

Chronic exposure to lead and cadmium pollution results in genomic instability in a model biomonitor species (Apodemus flavicollis Melchior, 1834).

Polymetal dust is a common industrial pollutant. While the use of remediation filters and equipment in lead smelters has reduced pollutant emission, surrounding areas remain contaminated due to the long-term transfer of heavy metals along the food chain. Here we assess the mutagenic potential of the lead-zinc smelter near Plovdiv (Bulgaria) situated in an area that has been contaminated with heavy metals for 60 years. We aimed to evaluate the genomic response of the yellow-necked mouse (A. flavicollis), a biomonitor species, in three sampling sites along the pollution gradient. Mice from Strandzha Natural Park were used as a negative control. The bioaccumulation rate of two non-essential heavy metals, lead (Pb) and cadmium (Cd), in liver tissues was determined by atomic absorption spectroscopy. Genetic alterations attributable to chronic exposure to trace levels of heavy metals were assessed in different blood cell populations using two independent methods: a micronucleus test was applied to evaluate the clastogenic and aneugenic alterations in erythrocytes, while a comet assay was used to assess DNA instability, as evidenced by single- and double-stranded breaks and alkali-labile sites, in leucocytes. We observed elevated levels of Pb and Cd in livers derived from mice from the impacted area: the mean Pb concentration (21.38 ± 8.77 µg/g) was two-fold higher than the lowest-observed-adverse-effect levels (LOAELs), while the mean Cd concentration (13.95 ± 9.79 µg/g) was extremely close to these levels. The mean levels of Pb and Cd in livers derived from mice from the impacted area were 31-fold and 63-fold higher, respectively, than the levels measured in mice from the control area. The mean frequency of micronuclei was significantly higher (four-fold) than that observed in the control animals. Furthermore, parameters measured by the comet assay, % tail DNA, tail length and tail moment, were significantly higher in the impact area, indicating the degree of genetic instability caused by exposure to heavy metals. In conclusion, this study shows that despite the reported reduction in lead and cadmium emissions in Bulgaria in recent years, A. flavicollis individuals inhabiting areas subject to long-term contamination exhibit significant signs of DNA damage.

Low-pass single-chromosome sequencing of human small supernumerary marker chromosomes (sSMCs) and Apodemus B chromosomes.

Supernumerary chromosomes sporadically arise in many eukaryotic species as a result of genomic rearrangements. If present in a substantial part of species population, those are called B chromosomes, or Bs. This is the case for 70 mammalian species, most of which are rodents. In humans, the most common types of extra chromosomes, sSMCs (small supernumerary marker chromosomes), are diagnosed in approximately 1 of 2000 postnatal cases. Due to low frequency in population, human sSMCs are not considered B chromosomes. Genetic content of both B-chromosomes and sSMCs in most cases remains understudied. Here, we apply microdissection of single chromosomes with subsequent low-pass sequencing on Ion Torrent PGM and Illumina MiSeq to identify unique and repetitive DNA sequences present in a single human sSMC and several B chromosomes in mice Apodemus flavicollis and Apodemus peninsulae. The pipeline for sequencing data analysis was made available in Galaxy interface as an addition to previously published command-line version. Human sSMC was attributed to the proximal part of chromosome 15 long arm, and breakpoints leading to its formation were located into satellite DNA arrays. Genetic content of Apodemus B chromosomes was species-specific, and minor alterations were observed in both species. Common features of Bs in these Apodemus species were satellite DNA and ERV enrichment, as well as the presence of the vaccinia-related kinase gene Vrk1. Understanding of the non-essential genome elements content provides important insights into genome evolution in general.

Parasitic nematodes of the genus Syphacia Seurat, 1916 infecting Muridae in the British Isles, and the peculiar case of Syphacia frederici.

Syphacia stroma (von Linstow, 1884) Morgan, 1932 and Syphacia frederici Roman, 1945 are oxyurid nematodes that parasitize two murid rodents, Apodemus sylvaticus and Apodemus flavicollis, on the European mainland. Only S. stroma has been recorded previously in Apodemus spp. from the British Isles. Despite the paucity of earlier reports, we identified S. frederici in four disparate British sites, two in Nottinghamshire, one each in Berkshire and Anglesey, Wales. Identification was based on their site in the host (caecum and not small intestine), on key morphological criteria that differentiate this species from S. stroma (in particular the tail of female worms) and by sequencing two genetic loci (cytochrome C oxidase 1 gene and a section of ribosomal DNA). Sequences derived from both genetic loci of putative British S. frederici isolates formed a tight clade with sequences from continental worms known to be S. frederici, clearly distinguishing these isolates from S. stroma which formed a tight clade of its own, distinct from clades representative of Syphacia obvelata from Mus and S. muris from Rattus. The data in this paper therefore constitute the first record of S. frederici from British wood mice, and confirm the status of this species as distinct from both S. obvelata and S. stroma.

Population dynamics of wild rodents induce stochastic fadeouts of a zoonotic pathogen.

Stochastic processes play an important role in the infectious disease dynamics of wildlife, especially in species subject to large population oscillations. Here, we study the case of a free ranging population of yellow-necked mice (Apodemus flavicollis) in northern Italy, where circulation of Dobrava-Belgrade hantavirus (DOBV) has been detected intermittently since 2001, until an outbreak emerged in 2010. We analysed the transmission dynamics of the recent outbreak using a computational model that accounts for seasonal changes of the host population and territorial behaviour. Model parameters were informed by capture-mark-recapture data collected over 14 years and longitudinal seroprevalence data from 2010 to 2013. The intermittent observation of DOBV before 2010 can be interpreted as repeated stochastic fadeouts after multiple introductions of infectious rodents migrating from neighbouring areas. We estimated that only 20% of introductions in a naïve host population results in sustained transmission after 2 years, despite an effective reproduction number well above the epidemic threshold (mean 4·5, 95% credible intervals, CI: 0·65-15·8). Following the 2010 outbreak, DOBV has become endemic in the study area, but we predict a constant probability of about 4·7% per year that infection dies out, following large population drops in winter. In the absence of stochastic fadeout, viral prevalence is predicted to continue its growth to an oscillating equilibrium around a value of 24% (95% CI: 3-57). We presented an example of invasion dynamics of a zoonotic virus where stochastic fadeout have played a major role and may induce future extinction of the endemic infection.

Bartonella, Rodents, Fleas and Ticks: a Molecular Field Study on Host-Vector-Pathogen Associations in Saxony, Eastern Germany.

Bartonellae cause zoonotic diseases and are transmitted by arthropods. Rodents are reservoirs for most Bartonella spp. As the knowledge about Bartonella in rodents and their parasitizing ectoparasites is scarce in Germany, this study's objectives were to investigate Bartonella spp. in small mammals and in their ectoparasites. A total of 79 small mammals (seven species) were captured and their ectoparasites collected at seven sites around Leipzig, Saxony, Germany, in 2010 and 2011. Altogether, 79 spleen samples, 135 fleas (five species) and 365 ticks (three species) were investigated for Bartonella spp. by PCR targeting the ITS 16S-23S rRNA region. In total, 52 (65.8 %) small mammals, 73 (54.1 %) fleas and 51 (16.3 %) ticks were positive for Bartonella spp. Most small mammals were positive for uncultured Bartonella sp. (n = 29) followed by Bartonella grahamii (n = 12), Bartonella taylorii (n = 8) and Bartonella sp. N40 (n = 3). Likewise, most fleas were positive for uncultured Bartonella sp. (n = 45) followed by B. grahamii (n = 14), B. taylorii (n = 8), B. sp. N40 (n = 5) and Bartonella elizabethae (n = 2). Most ticks were positive for B. sp. (n = 19) followed by B. grahamii (n = 10), Bartonella chomelii (n = 3), B. taylorii (n = 2) and B. sp. N40 (n = 1). This study's results suggest that rodents and fleas may be reservoirs and vectors, respectively. Zoonotic B. grahamii and B. elizabethae were found in rodents and their fleas. Therefore, humans may contract Bartonella infection by contact to wild rodents. Ticks seem of minor importance in transmitting Bartonella spp. found in fleas and rodents. However, ticks might be vectors of B. chomelii.

Advantages of masting in European beech: timing of granivore satiation and benefits of seed caching support the predator dispersal hypothesis.

The predator satiation and predator dispersal hypotheses provide alternative explanations for masting. Both assume satiation of seed-eating vertebrates. They differ in whether satiation occurs before or after seed removal and caching by granivores (predator satiation and predator dispersal, respectively). This difference is largely unrecognized, but it is demographically important because cached seeds are dispersed and often have a microsite advantage over nondispersed seeds. We conducted rodent exclosure experiments in two mast and two nonmast years to test predictions of the predator dispersal hypothesis in our study system of yellow-necked mice (Apodemus flavicollis) and European beech (Fagus sylvatica). Specifically, we tested whether the fraction of seeds removed from the forest floor is similar during mast and nonmast years (i.e., lack of satiation before seed caching), whether masting decreases the removal of cached seeds (i.e., satiation after seed storage), and whether seed caching increases the probability of seedling emergence. We found that masting did not result in satiation at the seed removal stage. However, masting decreased the removal of cached seeds, and seed caching dramatically increased the probability of seedling emergence relative to noncached seeds. European beech thus benefits from masting through the satiation of scatterhoarders that occurs only after seeds are removed and cached. Although these findings do not exclude other evolutionary advantages of beech masting, they indicate that fitness benefits of masting extend beyond the most commonly considered advantages of predator satiation and increased pollination efficiency.

Recent increase in prevalence of antibodies to Dobrava-Belgrade virus (DOBV) in yellow-necked mice in Northern Italy.

Dobrava-Belgrade virus (DOBV) is the most pathogenic hantavirus in Europe with a case-fatality rate of up to 12%. To detect changes in risk for humans, the prevalence of antibodies to DOBV has been monitored in a population of Apodemus flavicollis in the province of Trento (northern Italy) since 2000, and a sudden increase was observed in 2010. In the 13-year period of this study, 2077 animals were live-trapped and mean hantavirus seroprevalence was 2·7% (s.e. = 0·3%), ranging from 0% (in 2000, 2002 and 2003) to 12·5% (in 2012). Climatic (temperature and precipitation) and host (rodent population density, rodent weight and sex, and larval tick burden) variables were analysed using Generalized Linear Models and multi-model inference to select the best model. Climatic changes (mean annual precipitation and maximum temperature) and individual body mass had a positive effect on hantavirus seroprevalence. Other possible drivers affecting the observed pattern need to be studied further.

Seroprevalence of Tick-Borne Encephalitis (TBE) Virus Antibodies in Wild Rodents from Two Natural TBE Foci in Bavaria, Germany.

Tick-borne encephalitis (TBE) is Eurasia's most important tick-borne viral disease. Rodents play an important role as natural hosts. Longitudinal studies on the dynamics of the seroprevalence rates in wild rodents in natural foci over the year are rare, and the dynamics of the transmission cycle still need to be understood. To better understand the infection dynamics, rodents were captured in a capture-mark-release-recapture-study in two natural foci in Bavaria, Germany, monthly from March 2019 to October 2022. Overall, 651 blood and thoracic lavage samples from 478 different wild rodents (Clethrionomys glareolus and Apodemus flavicollis) were analyzed for antibodies against tick-borne encephalitis virus (TBEV) by indirect immunofluorescence assay (IIFA) and confirmed using a serum neutralization test (SNT). Furthermore, a generalized linear mixed model (GLMM) analysis was performed to investigate ecological and individual factors for the probability of infection in rodents. Clethrionomys glareolus (19.4%) had a higher seroprevalence than A. flavicollis (10.5%). Within Cl. glareolus, more males (40.4%) than females (15.6%) were affected, and more adults (25.4%) than juveniles (9.8%). The probability of infection of rodents rather depends on factors such as species, sex, and age than on the study site of a natural focus, year, and season. The high incidence rates of rodents, particularly male adult bank voles, highlight their critical role in the transmission cycle of TBEV in a natural focus and demonstrate that serologically positive rodents can be reliably detected in a natural focus regardless of season or year. In addition, these data contribute to a better understanding of the TBEV cycle and thus could improve preventive strategies for human infections.

From trees to fleas: masting indirectly affects flea abundance on a rodent host.

Mast seeding causes strong fluctuations in populations of forest animals. Thus, this phenomenon can be used as a natural experiment to examine how variation in host abundance affects parasite loads. We investigated fleas infesting yellow-necked mice in beech forest after 2 mast and 2 non-mast years. We tested 2 mutually exclusive scenarios: (1) as predicted by classical models of density-dependent transmission, an increase in host density will cause an increase in ectoparasite abundance (defined as the number of parasites per host), versus (2) an increase in host density will cause a decline in flea abundance ("dilution," which is thought to occur when parasite population growth is slower than that of the host). In addition, we assessed whether masting alters the relationship between host traits (sex and body mass) and flea abundance. We found a hump-shaped relationship between host and flea abundance. Thus, the most basic predictions are too simple to describe ectoparasite dynamics in this system. In addition, masting modified seasonal dynamics of flea abundance, but did not affect the relationship between host traits and flea abundance (individuals with the highest body mass hosted the most fleas; after controlling for body mass, parasite abundance did not vary between sexes). Our results demonstrate that pulses of tree reproduction can indirectly, through changes in host densities, drive patterns of ectoparasite infestation.

Continuous presence of genetically diverse rustrela virus lineages in yellow-necked field mouse reservoir populations in northeastern Germany.

Rustrela virus (RusV; species Rubivirus strelense, family Matonaviridae) was discovered in different zoo animal species affected by fatal encephalitis. Simultaneous RusV RNA detection in multiple yellow-necked field mice (Apodemus flavicollis) suggested this rodent as a reservoir of RusV. Here, we investigated 1,264 yellow-necked field mice and sympatric other small mammals from different regions in Germany for RusV RNA using an optimized reverse transcription-quantitative polymerase chain reaction (RT-qPCR) protocol and high-throughput sequencing. The investigation resulted in the detection of RusV RNA exclusively in 50 of 396 (12.6 per cent) yellow-necked field mice but absence in other sympatric species. RT-qPCR-determined tissue distribution of RusV RNA revealed the highest viral loads in the central nervous system, with other tissues being only very rarely affected. The histopathological evaluation did not reveal any hints of encephalitis in the brains of infected animals despite the detection of viral RNA in neurons by in situ hybridization (ISH). The positive association between the body mass of yellow-necked field mice and RusV RNA detection suggests a persistent infection. Phylogenetic analysis of partial E1 and full-genome sequences showed a high diversification with at least four RusV lineages (1A-1D) in northeastern Germany. Moreover, phylogenetic and isolation-by-distance analyses indicated evolutionary processes of RusV mostly in local reservoir populations. A comparison of complete genome sequences from all detected RusV lineages demonstrated a high level of amino acid and nucleotide sequence variability within a part of the p150 peptide of the non-structural polyprotein and its coding sequence, respectively. The location of this region within the RusV genome and its genetic properties were comparable to the hypervariable region of the rubella virus. The broad range of detected RusV spillover hosts in combination with its geographical distribution in northeastern Germany requires the assessment of its zoonotic potential and further analysis of encephalitis cases in mammals. Future studies have to prove a putative co-evolution scenario for RusV in the yellow-necked field mouse reservoir.

Investigation and response to a large outbreak of leptospirosis in field workers in Lower Saxony, Germany.

Between June and August 2014, 45 cases of leptospirosis were notified among workers on two strawberry farms in North-West Germany. We describe the characteristics of the outbreak and the actions taken to prevent further cases. The activities of the local, federal and national public health and veterinary authorities included collection of case data, laboratory testing of human specimens and of small mammals trapped on the fields, investigation of weather data, as well as information provided to farmers, field workers, physicians and to the authorities in Poland and Romania. Of the 45 identified cases (median age 22, 60% male), 47% were hospitalized. Characteristic symptoms were fever ≥38.5°C, generalized muscle pain and an increase in renal or liver enzymes. Thirteen cases were laboratory confirmed by serological and/or molecular methods. ELISA tests for Leptospira IgG and IgM-antibodies were positive in those samples taken >5 days after hospitalization. The probable causative agent was identified as Leptospira kirschneri serovar Grippotyphosa. Leptospira-specific DNA was found in kidneys of 67% of 64 trapped small mammals and was further identified as Leptospira kirschneri multi locus sequence type 110. During the estimated time period of human infections, the affected region faced warm weather with heavy rainfalls. The results of this investigation are in accordance with the theory of a chain of infection from mice to field workers during warm and humid weather. In 2015, a campaign was initiated to inform physicians, farmers and workers to enhance prevention measures, such as the use of personal protective equipment and early consultation of physicians in case of illness. Since then, no further outbreak occurred.

Identification of Dobrava-Belgrade Virus in Apodemus flavicollis from North-Eastern Italy during Enhanced Mortality.

Hantaviruses include several zoonotic pathogens that cause different syndromes in humans, with mortality rates ranging from 12 to 40%. Most commonly, humans get infected through the inhalation of aerosols or dust particles contaminated with virus-containing rodent excreta. Hantaviruses are specifically associated with the host species, and human cases depend on the presence and the dynamics of reservoir hosts. In this letter, we report the identification of Dobrava-Belgrade virus (DOBV) in the yellow-necked mouse (Apodemus flavicollis) from Italy. The virus was detected in the mountainous area of the province of Udine, bordering Austria and Slovenia, during an event of enhanced mortality in wild mice and voles. Despite serological evidence in rodents and humans that suggested the circulation of hantaviruses in Italy since 2000, this is the first virological confirmation of the infection. Phylogenetic analyses across the whole genome of the two detected viruses confirmed the host-specificity of DOBV sub-species and showed the highest identity with viruses identified in Slovenia and Croatia from both A. flavicollis and humans, with no signs of reassortment. These findings highlight the need for ecologists, veterinarians and medical doctors to come together in a coordinated approach in full compliance with the One Health concept.

Leptospira interrogans Sensu Lato in Wild Small Mammals in Three Moravian Localities of the Czech Republic.

Leptospirosis is a widespread zoonosis, affecting humans, domestic animals and wildlife, with small mammals as a reservoir of this infection. In recent years, this disease has been re-emerging and affects approximately 1 million people all over the world each year. Due to this disease having a significant health impact, it is important to identify the source and method of infection. The risk of Leptospira sp. infection is higher mainly in the cities of developed and industrialised countries. The aim of the study was the detection of antibodies against Leptospira sp. in some wild small mammals captured in the Czech Republic. In total, samples of 855 animals captured in three locations of Moravia during a six-year study (2010-2015) were examined by a microscopic agglutination test, using eight serovars of Leptospira interrogans sensu lato, representing serogroups Grippotyphosa, Icterohaemorrhagiae, Australis, Canicola, Sejroe, Javanica, Pomona and Pyrogenes, as antigens. Antibodies to Leptospira sp. were detected in 6.1% (52/855) of animals, with a prevalence of 6.4% (51/801) and 1.9% (1/54) in rodents and insectivores, respectively. The only statistically significant difference (p ≤ 0.05) was in prevalence between individual species (0-33%), while there were no differences in sex (6.7% in females and 5.1% in males), locality (1.8-8%) and year of trapping (0-8.4%). Only two serovars, L. interrogans serovar Pomona and L. interrogans serovar Grippotyphosa, were detected in 5.5% and 0.5% of animals, respectively. The prevailing serovar of pathogenic L. interrogans s.l. can be identified in a number of infected people in the Czech Republic. The composition of vaccines should be based on the current occurrence of Leptospira serovars in the actual territory. For this reason, the occurrence of Leptospira and its serovars should therefore be regularly monitored.

Temporal Partitioning between Forest-Dwelling Small Rodents in a Mediterranean Deciduous Woodland.

Temporal partitioning is reported as one of the main strategies adopted by coexisting mammal species to limit interspecific competition and behavioural interference. In the last decades, camera-trapping surveys have provided valuable insights in assessing temporal niche and activity rhythms of medium and large-sized mammalian species. Conversely, this method has been poorly applied to small rodents. In this work we aimed at assessing temporal niche partitioning between two species of forest-dwelling small rodents-Apodemus flavicollis and Clethrionomys glareolus-by means of intensive camera-trapping. Camera traps were placed in areas where previous genetic analyses have confirmed the only presence of A. flavicollis amongst wood mice species, to prevent misinterpretation of records. We collected 124 independent records of A. flavicollis and 67 records of C. glareolus over three years. The former was mostly nocturnal, with activity peaking after midnight, whereas the latter was mostly active at dawn and dusk. Therefore, a limited temporal overlap was observed, confirming the potential for interspecific competition. Intraguild interference competition between A. flavicollis and C. glareolus may play a pivotal role forcing C. glareolus to be more active in daylight hours where, the more strictly nocturnal A. flavicollis is present. Nocturnal activity of C. glareolus was limited and not influenced by moon phases, whereas A. flavicollis was mostly active in the darkest nights, avoiding bright moonlight nights.

B Chromosomes' Sequences in Yellow-Necked Mice Apodemus flavicollis-Exploring the Transcription.

B chromosomes (Bs) are highly polymorphic additional chromosomes in the genomes of many species. Due to the dispensability of Bs and the lack of noticeable phenotypic effects in their carriers, they were considered genetically inert for a long time. Recent studies on Bs in Apodemus flavicollis revealed their genetic composition, potential origin, and spatial organization in the interphase nucleus. Surprisingly, the genetic content of Bs in this species is preserved in all studied samples, even in geographically distinct populations, indicating its biological importance. Using RT-PCR we studied the transcription activity of three genes (Rraga, Haus6, and Cenpe) previously identified on Bs in A. flavicollis. We analysed mRNA isolated from spleen tissues of 34 animals harboring different numbers of Bs (0-3).The products of transcriptional activity of the analysed sequences differ in individuals with and without Bs. We recorded B-genes and/or genes from the standard genome in the presence of Bs, showing sex-dependent higher levels of transcriptional activity. Furthermore, the transcriptional activity of Cenpe varied with the age of the animals differently in the group with and without Bs. With aging, the amount of product was only found to significantly decrease in B carriers. The potential biological significance of all these differences is discussed in the paper.

Divergence in Coding Sequence and Expression of Different Functional Categories of Immune Genes between Two Wild Rodent Species.

Differences in immune function between species could be a result of interspecific divergence in coding sequence and/or expression of immune genes. Here, we investigate how the degree of divergence in coding sequence and expression differs between functional categories of immune genes, and if differences between categories occur independently of other factors (expression level, pleiotropy). To this end, we compared spleen transcriptomes of wild-caught yellow-necked mice and bank voles. Immune genes expressed in the spleen were divided into four categories depending on the function of the encoded protein: pattern recognition receptors (PRR); signal transduction proteins; transcription factors; and cyto- and chemokines and their receptors. Genes encoding PRR and cyto-/chemokines had higher sequence divergence than genes encoding signal transduction proteins and transcription factors, even when controlling for potentially confounding factors. Genes encoding PRR also had higher expression divergence than genes encoding signal transduction proteins and transcription factors. There was a positive correlation between expression divergence and coding sequence divergence, in particular for PRR genes. We propose that this is a result of that divergence in PRR coding sequence leads to divergence in PRR expression through positive feedback of PRR ligand binding on PRR expression. When controlling for sequence divergence, expression divergence of PRR genes did not differ from other categories. Taken together, the results indicate that coding sequence divergence of PRR genes is a major cause of differences in immune function between species.