The use of muscle biomarkers for assessing physiological effects of heavy metal pollution in the greater white-toothed shrew (Crocidura russula).

The greater white-toothed shrew Crocidura russula has been used as a sentinel species for estimating environmental risks to human populations. Previous studies in mining areas have focused on the liver of shrews as the primary target of physiological and metabolic changes due to heavy metal pollution. However, populations persist even when detoxification by the liver seems to be compromised and damage is observed. These pollutant-adapted individuals inhabiting contaminated sites may exhibit altered biochemical parameters that confer increased tolerance in various tissues other than the liver. The skeletal muscle tissue of C. russula might be an alternative tissue that allows the survival of organisms inhabiting historically polluted sites due to the detoxification of redistributed metals. Organisms from two heavy metal mine populations and one population derived from an unpolluted site were used to determine the detoxification activities, antioxidant capacity, and oxidative damage, as well as cellular energy allocation parameters and acetylcholinesterase activity (a biomarker of neurotoxicity). Muscle biomarkers differ between shrews from polluted sites and shrews from the unpolluted location, with the mine animals showing: (1) a decreased energy consumption concomitant with increased energy reserves and total available energy; (2) reduced cholinergic activity, suggesting an impairment of neurotransmission at the neuromuscular junction; (3) an overall decrease in detoxification capacity and enzymatic antioxidant response and a higher level of lipid damage. Also, some of these markers differed between females and males. These changes may have resulted from a decreased detoxifying capacity of the liver and could potentially bring about significant ecological effects for this highly active species. Heavy metal pollution induced physiological changes in Crocidura russula showing that skeletal muscle may serve as a backup sink organ allowing rapid species adaptation and evolution.

Oxidative damage in the Vesper mouse (Calomys laucha) exposed to a simulated oil spill-a multi-organ study.

Small wild mammals have been used to measure the damage caused by exposure to oil-contaminated soil, including deer mice. However, the study of toxic effects of crude oil using oxidative damage biomarkers in the wild rodent Calomys laucha (Vesper mouse) is absent. This investigation aimed to evaluate the effects of acute exposure to contaminated soil with different concentrations of crude oil (0, 1, 2, 4 and 8% w/w), simulating an accidental spill, using oxidative stress biomarkers in the liver, kidneys, lungs, testes, paw muscle, and lymphocytes of C. laucha. Animals exposed to the contaminated soil showed increases in lipid peroxidation and protein carbonylation at the highest exposure concentrations in most organ homogenates analyzed and also in blood cells, but responses to total antioxidant capacity were tissue-dependent. These results showed that acute exposure to oil-contaminated soil caused oxidative damage in C. laucha and indicate these small mammals may be susceptible to suffer the impacts of such contamination in its occurrence region, threatening the species' survival.

Variation and Selection in the Putative Sperm-Binding Region of ZP3 in Muroid Rodents: A Comparison between Cricetids and Murines.

In mammals, the zona pellucida glycoprotein 3 (ZP3) is considered a primary sperm receptor of the oocyte and is hypothesized to be involved in reproductive isolation. We investigated patterns of diversity and selection in the putative sperm-binding region (pSBR) of mouse ZP3 across Cricetidae and Murinae, two hyperdiverse taxonomic groups within muroid rodents. In murines, the pSBR is fairly conserved, in particular the serine-rich stretch containing the glycosylation sites proposed as essential for sperm binding. In contrast, cricetid amino acid sequences of the pSBR were much more variable and the serine-rich motif, typical of murines, was generally substantially modified. Overall, our results suggest a general lack of species specificity of the pSBR across the two muroid families. We document statistical evidence of positive selection acting on exons 6 and 7 of ZP3 and identified several amino acid sites that are likely targets of selection, with most positively selected sites falling within or adjacent to the pSBR.

Multimarker approach to assess the exposure of the wild rodent Calomys laucha to a simulated crude oil spill.

A mysterious oil spill occurred in the ocean near Brazil in 2019, which affected coastal areas in northeastern Brazil. When oil pollution occurs in coastal zones, organisms such as small mammals can suffer deleterious effects to their health. This study aimed to evaluate the effects of exposure to contaminated sandy soil with different crude oil concentrations in males of the species Calomys laucha. The exposure to crude oil resulted in multiple health issues for the subjects in the very first days of exposure. Furthermore, the exposure resulted in mutagenic damage to bone marrow blood cells and behavioral and morphological alterations, which were almost always in a dose-dependent form. The present study demonstrates the sensibility of the biomarkers used and highlights that small wild mammals such as C. laucha are useful for predicting environmental damage caused by the exposure to crude oil.

Population effects of heavy metal pollution in wild Algerian mice (Mus spretus).

Heavy metal mining is one of the largest sources of environmental pollution. The analysis of different types of biomarkers in sentinel species living in contaminated areas provides a measure of the degree of the ecological impact of pollution and is thus a valuable tool for human and environmental risk assessments. In previous studies we found that specimens from two populations of the Algerian mice (Mus spretus) living in two abandoned heavy metal mines (Aljustrel and Preguiça, Portugal) had higher body burdens of heavy metals, which led to alterations in enzymatic activities and in haematological, histological and genotoxic parameters, than mice from a nearby reference population. We have now analysed individuals from the same sites at the biometric and genetic levels to get a broader portrayal of the impact of heavy metal pollution on biodiversity, from molecules to populations. Size and shape variations of the mouse mandible were searched by implementing the geometric morphometric method. Population genetic differentiation and diversity parameters (φST estimates; nucleotide and haplotype diversities) were studied using the mitochondrial cytochrome b gene (Cytb) and the control region (CR). The morphometric analyses revealed that animals from the three sites differed significantly in the shape of the mandible, but mandibular shape varied in a more resembling way within individuals of both mine sites, which is highly suggestive for an effect of environmental quality on normal development pathways in Algerian mice. Also, antisymmetry in mandible size and shape was detected in all populations, making these traits not reliable indicators of developmental instability. Overall little genetic differentiation was found among the three populations, although pairwise φST comparisons revealed that the Aljustrel and the Preguiça populations were each differentiated from the other two populations in Cytb and in CR, respectively. Genetic diversity parameters revealed higher genetic diversity for Cytb in the population from Aljustrel, while in the population from Preguiça diversity of the two markers changed in opposite directions, higher genetic diversity in CR and lower in Cytb, compared to the reference population. Demographic changes and increased mutation rates may explain these findings. We show that developmental patterns and genetic composition of wild populations of a small mammal can be affected by chronic heavy metal exposure within a relatively short time. Anthropogenic stress may thus influence the evolutionary path of natural populations, with largely unpredictable ecological costs.

Phylogeny and adaptation shape the teeth of insular mice.

By accompanying human travels since prehistorical times, the house mouse dispersed widely throughout the world, and colonized many islands. The origin of the travellers determined the phylogenetic source of the insular mice, which encountered diverse ecological and environmental conditions on the various islands. Insular mice are thus an exceptional model to disentangle the relative role of phylogeny, ecology and climate in evolution. Molar shape is known to vary according to phylogeny and to respond to adaptation. Using for the first time a three-dimensional geometric morphometric approach, compared with a classical two-dimensional quantification, the relative effects of size variation, phylogeny, climate and ecology were investigated on molar shape diversity across a variety of islands. Phylogeny emerged as the factor of prime importance in shaping the molar. Changes in competition level, mostly driven by the presence or absence of the wood mouse on the different islands, appeared as the second most important effect. Climate and size differences accounted for slight shape variation. This evidences a balanced role of random differentiation related to history of colonization, and of adaptation possibly related to resource exploitation.

R2d2 Drives Selfish Sweeps in the House Mouse.

A selective sweep is the result of strong positive selection driving newly occurring or standing genetic variants to fixation, and can dramatically alter the pattern and distribution of allelic diversity in a population. Population-level sequencing data have enabled discoveries of selective sweeps associated with genes involved in recent adaptations in many species. In contrast, much debate but little evidence addresses whether "selfish" genes are capable of fixation-thereby leaving signatures identical to classical selective sweeps-despite being neutral or deleterious to organismal fitness. We previously described R2d2, a large copy-number variant that causes nonrandom segregation of mouse Chromosome 2 in females due to meiotic drive. Here we show population-genetic data consistent with a selfish sweep driven by alleles of R2d2 with high copy number (R2d2(HC)) in natural populations. We replicate this finding in multiple closed breeding populations from six outbred backgrounds segregating for R2d2 alleles. We find that R2d2(HC) rapidly increases in frequency, and in most cases becomes fixed in significantly fewer generations than can be explained by genetic drift. R2d2(HC) is also associated with significantly reduced litter sizes in heterozygous mothers, making it a true selfish allele. Our data provide direct evidence of populations actively undergoing selfish sweeps, and demonstrate that meiotic drive can rapidly alter the genomic landscape in favor of mutations with neutral or even negative effects on overall Darwinian fitness. Further study will reveal the incidence of selfish sweeps, and will elucidate the relative contributions of selfish genes, adaptation and genetic drift to evolution.

Predation risk modulates diet-induced obesity in male C57BL/6 mice.

OBJECTIVE: In this study, the behavioral and physiological changes induced by experimentally varying the risk of predation in male mice fed a high-fat diet were examined. In particular, the study aimed to assess whether the risk of being predated modulates the body weight gain, providing an ecological context for the obesity resistance observed in many species of small mammals. METHODS: Body weight, food intake, physical activity, and core body temperature of 35 male C57BL/6 mice were monitored for 20 days, while feeding a high-fat diet. A third of the animals were exposed to elevated risk of predation through exposure to the sounds of nocturnal predatory birds, and these were compared to animals exposed to a neutral noise or silence. RESULTS: Male mice exposed to predation risk had significantly lower weight gain than the neutral or silent groups. Reduced food intake and increased physical activity were the main proximal factors explaining this effect. The risk of predation also induced changes in boldness. CONCLUSIONS: This study provides evidence supporting the role of predation risk on body weight gain of small mammals.

Geno- and cyto-toxicity in free-living rodent Mus spretus exposed to simulated onshore oil spill.

This study investigated geno- and cyto-toxic damage in the free-living rodent, Mus spretus after exposure to a simulated spill of crude oil on soil. The results revealed increased mutagenicity and cytotoxicity in bone marrow cells and increased DNA damage in blood cells. Exposure to crude oil increased sperm abnormalities, with lasso-like folds being the most common. These results point to the value of this rodent in serving as a sentinel species for the monitoring and prediction of environmental hazards.

Metallothionein levels in Algerian mice (Mus spretus) exposed to elemental pollution: an ecophysiological approach.

The potential use of metallothioneins (MTs) as biomarkers of trace metal contamination was evaluated for the first time in the Algerian mouse (Mus spretus). Mice were collected seasonally in an abandoned mining area (Aljustrel) and in a reference area, both located in southern Portugal. MT levels were quantified in liver and kidney by differential pulse polarography and hepatic elemental concentrations (Mn, Fe, Cu, Zn, Se) were determined by particle-induced X-ray emission. Hepatic iron and selenium concentrations were elevated in mice from Aljustrel mine when compared to reference animals. MTs levels were averagely higher in mice from Aljustrel than those originated from the reference area. A season-dependent significant effect was found on the hepatic and renal MT concentrations, characterized by higher levels in winter and lower in autumn. In contaminated mice positive relationship between liver elemental contents (Cu in autumn and Fe in winter) and MTs were found. The seasonal variation of MT suggests that probably physiological and environmental factors could influence hepatic and renal MT induction. Results seem to imply that some environmental disturbance occur in the vicinity of the Aljustrel mine. Therefore, for the management purposes MT levels should be followed in liver of M. spretus, especially in winter. Furthermore, other physiological factors that could influence MT expression and turnover in Algerian mouse should also be monitored.

How does the greater white-toothed shrew, Crocidura russula, responds to long-term heavy metal contamination? -- A case study.

Heavy metals accumulation in parallel with the evaluation of physiological and biochemical effects resulting from continued metal exposure were considered here using for the first time the great white-toothed shrew Crocidura russula as an in vivo model. Shrews were originated from an abandoned lead/zinc mining area and from a reference area, both in Alentejo, southern Portugal. Hepatic contents of nickel, copper, zinc, cadmium, mercury and lead were quantified by Inductively Coupled Plasma Mass Spectrometer (ICP-MS). Haematological parameters (white blood cells, red blood cells, haemoglobin and haematocrit) were obtained in a Coulter Counter Analyser and biochemical markers of the redox balance (glutathione S-transferase, glutathione peroxidase, and glutathione reductase) activities were measured spectrophotometrically using a Duo-50 spectrophotometer. Compared with control animals, significantly higher concentration of hepatic cadmium (9.29 vs. 1.18 micorg/g dry weight) and nickel (1.56 vs. 0.343 microg/g dry weight) were detected in the shrews collected in the mining area. However, no significant changes were observed on haematological or enzymatic parameters in animals exposed to metal pollution. The obtained results show that shrews are good bioaccumulators of toxic heavy metals, but very tolerant to their effects, revealing an interesting long-term adaptation to polluted environments. In addition, this study provides reference values for haematological parameters and antioxidant enzymes levels in C. russula, which may be relevant for comparative purposes in further studies.

Metal bioaccumulation in the greater white-toothed shrew, Crocidura russula, inhabiting an abandoned pyrite mine site.

Hepatic and renal concentrations of iron, magnesium, zinc, lead, copper, manganese, mercury, cadmium, molybdenum, chromium, and nickel were quantified in shrews (Crocidura russula) inhabiting a pyrite mine site in Portugal. Several morphological parameters (body weight, residual index, and relative weights) were also examined to clarify the physiological effects of pollution. Shrews from the mine showed increased bioavailability of Fe, Pb, Hg, Cd, Mo, and Ni in comparison with reference specimens. Adult shrews had the highest Cd levels while Cr and Ni concentrations diminished. Intersexual differences were found for Mo and Ni. As a consequence of metal pollution, the relative hepatic weight was higher in shrews from the mine site when compared with reference specimens. These data indicate that C. russula is a good bioindicator of metal pollution. We also evaluated the toxic effects of Pb, Hg, Cd, and Ni, because several shrews from the polluted site showed high concentrations of these metals. To approximate at the real biological impact of abandoned mines, after this first step it is necessary to associate the bioaccumulation levels and morphological effects with other physiological, ecological and genetical biomarkers.

Chromosomal phylogeny of Robertsonian races of the house mouse on the island of Madeira: testing between alternative mutational processes.

The ancestral karyotype of the house mouse (Mus musculus) consists of 40 acrocentric chromosomes, but numerous races exist within the domesticus subspecies characterized by different metacentric chromosomes formed by the joining at the centromere of two acrocentrics. An exemplary case is present on the island of Madeira where six highly divergent chromosomal races have accumulated different combinations of 20 metacentrics in 500-1000 years. Chromosomal cladistic phylogenies were performed to test the relative performance of Robertsonian (Rb) fusions, Rb fissions and whole-arm reciprocal translocations (WARTs) in resolving relationships between the chromosomal races. The different trees yielded roughly similar topologies, but varied in the number of steps and branch support. The analyses using Rb fusions/fissions as characters resulted in poorly supported trees requiring six to eight homoplasious events. Allowance for WARTs considerably increased nodal support and yielded the most parsimonious trees since homoplasy was reduced to a single event. The WART-based trees required five to nine WARTs and 12 to 16 Rb fusions. These analyses provide support for the role of WARTs in generating the extensive chromosomal diversification observed in house mice. The repeated occurrence of Rb fusions and WARTs highlights the contribution of centromere-related rearrangements to accelerated rates of chromosomal change in the house mouse.

Molecular phylogeny of the speciose vole genus Microtus (Arvicolinae, Rodentia) inferred from mitochondrial DNA sequences.

Voles of the genus Microtus represent one of the most speciose mammalian genera in the Holarctic. We established a molecular phylogeny for Microtus to resolve contentious issues of systematic relationships and evolutionary history in this genus. A total of 81 specimens representing ten Microtus species endemic to Europe as well as eight Eurasian, six Asian and one Holarctic species were sequenced for the entire cytochrome b gene (1140 bp). A further 25 sequences were retrieved from GenBank, providing data on an additional 23, mainly Nearctic, Microtus species. Phylogenetic analysis of these 48 species generated four well-supported monophyletic lineages. The genus Chionomys, snow voles, formed a distinct and well-supported lineage separate from the genus Microtus. The subgenus Microtus formed the strongest supported lineage with two sublineages displaying a close relationship between the arvalis species group (common voles) and the socialis species group (social voles). Monophyly of the Palearctic pitymyid voles, subgenus Terricola, was supported, and this subgenus was also subdivided into two monophyletic species groups. Together, these groupings clarify long-standing taxonomic uncertainties in Microtus. In addition, the "Asian" and the Nearctic lineages reported previously were identified although the latter group was not supported. However, relationships among the main Microtus branches were not resolved, suggesting a rapid and potentially simultaneous radiation of a widespread ancestor early in the history of the genus. This and subsequent radiations discernible in the cytochrome b phylogeny, show the considerable potential of Microtus for analysis of historical and ecological determinants of speciation in small mammals. It is evident that speciation is an ongoing process in the genus and that the molecular data provides a vital insight into current species limits as well as cladogenic events of the past.