Phylogeny of Shrew- and Mole-Borne Hantaviruses in Poland and Ukraine.

Earlier, we demonstrated the co-circulation of genetically distinct non-rodent-borne hantaviruses, including Boginia virus (BOGV) in the Eurasian water shrew (Neomys fodiens), Seewis virus (SWSV) in the Eurasian common shrew (Sorex araneus) and Nova virus (NVAV) in the European mole (Talpa europaea), in central Poland. To further investigate the phylogeny of hantaviruses harbored by soricid and talpid reservoir hosts, we analyzed RNAlater®-preserved lung tissues from 320 shrews and 26 moles, both captured during 1990-2017 across Poland, and 10 European moles from Ukraine for hantavirus RNA through RT-PCR and DNA sequencing. SWSV and Altai virus (ALTV) were detected in Sorex araneus and Sorex minutus in Boginia and the Bialowieza Forest, respectively, and NVAV was detected in Talpa europaea in Huta Dlutowska, Poland, and in Lviv, Ukraine. Phylogenetic analyses using maximum-likelihood and Bayesian methods showed geography-specific lineages of SWSV in Poland and elsewhere in Eurasia and of NVAV in Poland and Ukraine. The ATLV strain in Sorex minutus from the Bialowieza Forest on the Polish-Belarusian border was distantly related to the ATLV strain previously reported in Sorex minutus from Chmiel in southeastern Poland. Overall, the gene phylogenies found support long-standing host-specific adaptation.

Long-term changes in winter abundance of the barbastelle Barbastella barbastellus in Poland and the climate change - Are current monitoring schemes still reliable for cryophilic bat species?

Warmer winters may lead to changes in the hibernation behaviour of bats, such as the barbastelle Barbastella barbastellus, which prefers to hibernate at low temperatures. The species is also known for its large annual fluctuations in the number of wintering individuals, so inference about population trends should be based on long-term data. Prior to 2005, analyses indicated stable or even increasing barbastelle population in Poland. We analysed the results of 13 winter bat counts (2005-2017) of the species from 15 of the largest hibernacula, and additional site of 47 small bunkers, in Poland. The total number of wintering individuals remained stable during the study period, because the barbastelle is not a long-distance migrant, this likely reflects the national population trend. On the basis of mean winter air temperatures we divided the country into four thermal regions. Analyses of barbastelle abundance in hibernacula in the four regions revealed a 4.8% annual mean increase in numbers in the coldest region, where mean winter temperatures were below -2°C, annual mean declines of 3.3% and 3.1% in two warmer regions of western Poland, but no trend in the region of intermediate mean winter temperatures of between -1°C and -2°C. Overall, there was a significant, but weak, negative correlation between the abundance of hibernating individuals and the mean winter temperature. On the other hand, the number of individuals hibernating in small bunkers increased, even though the site was located in one of the warm regions. The results indicate a warming climate will likely reduce the use of large, well-insulated winter roosts by species that prefer colder conditions-and that this is already happening. For forest-dwelling bats, such as the barbastelle, for which monitoring schemes are primarily based on winter surveys of large hibernacula, estimations of population trends may consequently become less reliable.

Ecotype Variation in Trace Element Content of Hard Tissues in the European Roe Deer (Capreolus capreolus).

Animals living in anthropogenic habitats bear a multitude of costs, which are directly or indirectly associated with human activities. Among others, an elevated exposure to environmental pollution can have negative consequences for wildlife populations. We examined the differences in the concentrations of trace elements between the field and forest ecotype of the European roe deer (Capreolus capreolus). Naturally, roe deer inhabited various types of woodlands (forest ecotype), but within the last century, they adapted to life in a human-transformed agricultural areas (field ecotype), which could be associated with an increased exposure to pollution. In this study, we measured concentrations of seven trace metals (barium, copper, iron, lead, manganese, strontium, zinc) and fluoride in skull bones and permanent teeth of more than 230 roe deer from 8 study plots in East-Central Europe. We found that field roe deer had higher concentrations of four trace metals (copper, iron, lead, strontium) and fluoride compared with forest roe deer. These differences were consistent with variations in the general level of environmental contamination within the study plots, as assessed with trace element content in wild plants. Our study indicates that bone and teeth of the European roe deer can be used as a valid indicator of environmental pollution. Also, we expect that elevated exposure of field roe deer to environmental pollution can have negative consequences for wild populations of this species, as well as for the consumers of venison.

A Novel Hantavirus of the European Mole, Bruges Virus, Is Involved in Frequent Nova Virus Coinfections.

Hantaviruses are zoonotic viruses with a complex evolutionary history of virus-host coevolution and cross-species transmission. Although hantaviruses have a broad reservoir host range, virus-host relationships were previously thought to be strict, with a single virus species infecting a single host species. Here, we describe Bruges virus, a novel hantavirus harbored by the European mole (Talpa europaea), which is the well-known host of Nova virus. Phylogenetic analyses of all three genomic segments showed tree topology inconsistencies, suggesting that Bruges virus has emerged from cross-species transmission and ancient reassortment events. A high number of coinfections with Bruges and Nova viruses was detected, but no evidence was found for reassortment between these two hantaviruses. These findings highlight the complexity of hantavirus evolution and the importance of further investigation of hantavirus-reservoir relationships.

Age-Related Patterns in Trace Element Content Vary Between Bone and Teeth of the European Roe Deer (Capreolus capreolus).

Game animals, such as the roe deer (Capreolus capreolus), have long been used as bioindicators of environmental contamination. Most ecotoxicological research on ungulates has focused on trace element content in soft tissues and antlers. Also, only fragmentary information exists about whether and how trace element concentrations vary with the age of wild-living animals and whether these age-related patterns are similar for different types of tissues. The purpose of this study was to measure concentrations of seven trace metals (barium, copper, iron, lead, manganese, strontium, zinc) and fluoride in bone and teeth of roe deer and to determine whether significant variation is evident with individual age. For this purpose, we collected permanent molars and fragments of mandible bone from more than 130 female roe deer in Central Poland. We found that concentrations of four trace elements (barium, manganese, zinc, and fluoride) in teeth of deer showed positive linear relationships with individual age. No such trends were recorded for trace element content in bone. We suggest that these striking differences in age-related patterns of trace element bioaccumulation between bone and permanent teeth of roe deer might be explained by higher turnover rate and constant remodelling of bone tissue. The results suggest that analysis of permanent teeth may be useful for assessing throughout-life intoxication by environmental pollution in the roe deer and possibly in other mammal species. Our study reinforces the need to carefully account for age-related variation in ecotoxicological research on wild-living animals.

Isolation and partial characterization of a highly divergent lineage of hantavirus from the European mole (Talpa europaea).

Genetically distinct hantaviruses have been identified in five species of fossorial moles (order Eulipotyphla, family Talpidae) from Eurasia and North America. Here, we report the isolation and partial characterization of a highly divergent hantavirus, named Nova virus (NVAV), from lung tissue of a European mole (Talpa europaea), captured in central Poland in August 2013. Typical hantavirus-like particles, measuring 80-120 nm in diameter, were found in NVAV-infected Vero E6 cells by transmission electron microscopy. Whole-genome sequences of the isolate, designated NVAV strain Te34, were identical to that amplified from the original lung tissue, and phylogenetic analysis of the full-length L, M and S segments, using maximum-likelihood and Bayesian methods, showed that NVAV was most closely related to hantaviruses harbored by insectivorous bats, consistent with an ancient evolutionary origin. Infant Swiss Webster mice, inoculated with NVAV by the intraperitoneal route, developed weight loss and hyperactivity, beginning at 16 days, followed by hind-limb paralysis and death. High NVAV RNA copies were detected in lung, liver, kidney, spleen and brain by quantitative real-time RT-PCR. Neuropathological examination showed astrocytic and microglial activation and neuronal loss. The first mole-borne hantavirus isolate will facilitate long-overdue studies on its infectivity and pathogenic potential in humans.

Whole-Genome Sequence of a Novel Hantavirus Isolated from the European Mole (Talpa europaea).

The complete genome sequence of Nova virus, a novel hantavirus isolated from a European mole (Talpa europaea) captured in central Poland, was determined. The availability of this sequence will facilitate the search for other mole-borne hantaviruses and will accelerate the acquisition of new knowledge about their phylogeography and evolutionary origin.

Co-circulation of soricid- and talpid-borne hantaviruses in Poland.

Previously, we reported the discovery of a genetically distinct hantavirus, designated Boginia virus (BOGV), in the Eurasian water shrew (Neomys fodiens), as well as the detection of Seewis virus (SWSV) in the Eurasian common shrew (Sorex araneus), in central Poland. In this expanded study of 133 shrews and 69 moles captured during 2010-2013 in central and southeastern Poland, we demonstrate the co-circulation of BOGV in the Eurasian water shrew and SWSV in the Eurasian common shrew, Eurasian pygmy shrew (Sorex minutus) and Mediterranean water shrew (Neomys anomalus). In addition, we found high prevalence of Nova virus (NVAV) infection in the European mole (Talpa europaea), with evidence of NVAV RNA in heart, lung, liver, kidney, spleen and intestine. The nucleotide and amino acid sequence variation of the L segment among the SWSV strains was 0-18.8% and 0-5.4%, respectively. And for the 38 NVAV strains from European moles captured in Huta Dlutowska, the L-segment genetic similarity ranged from 94.1%-100% at the nucleotide level and 96.3%-100% at the amino acid level. Phylogenetic analyses showed geographic-specific lineages of SWSV and NVAV in Poland, not unlike that of rodent-borne hantaviruses, suggesting long-standing host-specific adaptation. The co-circulation and distribution of BOGV, SWSV and NVAV in Poland parallels findings of multiple hantavirus species co-existing in their respective rodent reservoir species elsewhere in Europe. Also, the detection of SWSV in three syntopic shrew species resembles spill over events observed among some rodent-borne hantaviruses.

Boginia virus, a newfound hantavirus harbored by the Eurasian water shrew (Neomys fodiens) in Poland.

BACKGROUND: Guided by decades-old reports of hantaviral antigens in the Eurasian common shrew (Sorex araneus) and the Eurasian water shrew (Neomys fodiens) in European Russia, we employed RT-PCR to analyze lung tissues of soricine shrews, captured in Boginia, Huta Dlutowska and Kurowice in central Poland during September 2010, 2011 and 2012. FINDINGS: In addition to Seewis virus (SWSV), which had been previously found in Eurasian common shrews elsewhere in Europe, a genetically distinct hantavirus, designated Boginia virus (BOGV), was detected in Eurasian water shrews captured in each of the three villages. Phylogenetic analysis, using maximum likelihood and Bayesian methods, showed that BOGV formed a separate lineage distantly related to SWSV. CONCLUSIONS: Although the pathogenic potential of BOGV and other recently identified shrew-borne hantaviruses is still unknown, clinicians should be vigilant for unusual febrile diseases and clinical syndromes occurring among individuals reporting exposures to shrews.