Microtus Arvalis Research Articles

Genomes of Microtus rodents highlight the importance of olfactory and immune systems in their fast radiation.

The characterization of genes and biological functions underlying functional diversification and the formation of species is a major goal of evolutionary biology. In this study, we investigated the fast radiation of Microtus voles, one of the most speciose group of mammals, which shows strong genetic divergence despite few readily observable morphological differences. We produced an annotated reference genome for the common vole, Microtus arvalis, and resequenced the genomes of 10 different species and evolutionary lineages spanning the Microtus speciation continuum. Our full genome sequences illustrate the recent and fast diversification of this group, and we identified genes in highly divergent genomic windows that have likely particular roles in their radiation. We found three biological functions enriched for highly divergent genes in most Microtus species and lineages: olfaction, immunity and metabolism. In particular, olfaction-related genes (mostly olfactory receptors and vomeronasal receptors) are fast evolving in all Microtus species indicating the exceptional importance of the olfactory system in the evolution of these rodents. Of note is e.g. the shared signature among vole species on Olfr1019 which has been associated with fear responses against predator odours in rodents. Our analyses provide a genome-wide basis for the further characterization of the ecological factors and processes of natural and sexual selection that have contributed to the fast radiation of Microtus voles.

Can repellent crops reduce the abundance of the common vole (Microtus arvalis) as a way to reduce crop damage?

The common vole is one of the most damaging rodents in agriculture. A number of methods are used to suppress its numbers and limit damage. One option is growing crops that are not suitable for voles; this might limit their numbers and distribution. Through long-term monitoring of common voles throughout the Czech Republic, their abundance in eleven crop types (annual, biennial and perennial crops) was evaluated through active burrow counts. Reference crops were selected from perennials and biennials and annual crops were selected as potential repellents. The perennial crops were clover with alfalfa, which serve as the primary habitat for voles. These crops are where voles are found in their highest densities, and from which they spread to the surrounding crops during periods of high abundance. The biennial winter rape was selected because it provides favourable conditions for voles to overwinter and multiply rapidly in the spring. Compared to perennial crops and winter rape, significantly lower numbers of voles were found in onions, poppy seeds and maize. Additionally, fewer voles were also found in mustard compared to perennial crops. Onion and poppy were the only crops to show a significantly lower abundance of voles in comparison to wheat. The annual crops tested are unattractive to voles and due to the cultivation practices used, they do not even have enough weeds as food. As profitable crops they can be cultivated over sufficiently large areas to potentially prevent the spread of voles to surrounding crops, especially during outbreak years.

Fur microbiome as a putative source of symbiotic bacteria in sucking lice

Symbiosis between insects and bacteria has been established countless times. While it is well known that the symbionts originated from a variety of different bacterial taxa, it is usually difficult to determine their environmental source and a route of their acquisition by the host. In this study, we address this question using a model of Neisseriaceae symbionts in rodent lice. These bacteria established their symbiosis independently with different louse taxa (Polyplax, Hoplopleura, Neohaematopinus), most likely from the same environmental source. We first applied amplicon analysis to screen for candidate source bacterium in the louse environment. Since lice are permanent ectoparasites, often specific to the particular host, we screened various microbiomes associated with three rodent species (Microtus arvalis, Clethrionomys glareolus, and Apodemus flavicollis). The analyzed samples included fur, skin, spleen, and other ectoparasites sampled from these rodents. The fur microbiome data revealed a Neisseriaceae bacterium, closely related to the known louse symbionts. The draft genomes of the environmental Neisseriaceae, assembled from all three rodent hosts, converged to a remarkably small size of approximately 1.4 Mbp, being even smaller than the genomes of the related symbionts. Our results suggest that the rodent fur microbiome can serve as a source for independent establishment of bacterial symbiosis in associated louse species. We further propose a hypothetical scenario of the genome evolution during the transition of a free-living bacterium to the member of the rodent fur-associated microbiome and subsequently to the facultative and obligate louse symbionts.

Biometric analysis of the cranial features of common voles

Although cranial bones reliably identify the features of mammals, these traits can be affected by sex, population size and seasonality. This paper biometrically analyses the cranial features of the common vole. The research sample consisted of adult (n = 134), subadult (n = 57), of western Slovakia. Overall, software based on the highlighted marks measured 29 cranial features. Significant sexual dimorphism was observed for three features in adult individuals (width of the maxillary tooth row – right side “WMTRR”, maximum height of the mandibula excluding coronoid process “AMdm” and coronoid height of the mandibula “AMd”) and four features in subadults (length of the tooth row in the maxilla “LOSD”, “WMTRR”, length of the molar “LoM” and alveolar length of the lower molars “ALLM”). Results showed a significant positive relationship between body weight and cranial size in 18 of the 29 features. They also suggested a weak but significant positive effect of common vole abundance on cranium size (Chitty effect) likewise in 6 of the 29 features. In several cases, this effect affects, in addition to the abundance, the onset of sexual maturity of individuals.

Structure of the hybrid zone between allied species of the common vole, Microtus arvalis and M. obscurus: Influence of genetic factors and landscape-geographic conditions

Patterns of introgression of several genetic markers across the hybrid zone between allied species of the common vole Microtus arvalis s. str. и M. obscurus were studied in four its sections: in northwest Nizhny Novgorod region, east Vladimir and southwest of Nizhny Novgorod regions, south Lipetsk region, and northwest Voronezh region. Analysis of the clinal variability for three molecular-genetic markers (cytb, tp53, SMCY11) and for karyotypes showed a structural similarity between the “Vladimir – Nizhny Novgorod”, “Nizhny Novgorod”, and “Voronezh” sections. The maximal width was shown for the cytb cline, the minimal width – for the SMCY11 cline; the tp53 cline and chromosomal cline occupy intermediate position for this parameter. Furthermore, in these transects the centre of the cline for the cytb is shifted southeastward (into the distribution range of M. obscurus) from the centres of three other clines. The revealed asymmetric introgression of mitochondrial genome from M. arvalis to M. obscurus may be explained by the fact that the hybrid zone was formed as a result of invasion of M. obscurus into the range of M. arvalis. The “Lipetsk” transect differs from three above-mentioned transects by very narrow clines with nearly coinciding centres. Such characteristics of the “Lipetsk” transect are obviously caused by localization of the hybrid zone in this section along river Voronezh. The obtained results led us to suppose that the structure of the studied hybrid zone is determined mainly by coinciding (or non-coinciding) of its centre with local physical barriers.

Analyzing the spatiotemporal habitat colonization dynamics of the common vole (Microtus arvalis Pallas) in Castilla y León, northwest Spain, using a hotspot-based approach†.

The common vole is an agricultural pest and population outbreaks have caused significant crop damage in the agricultural areas of the west-central part of the Iberian Peninsula (Spain) during the last few decades. Thus, monitoring is imperative to gain a comprehensive insight on its spatiotemporal habitat colonization dynamics. This work was performed on a long-term database with the primary objective of establishing an operational framework for understanding the spatial and temporal distribution of the common vole, all of it based on the Getis-Ord statistics. The temporal evolution of the vole abundance index (VAI) was consistent for the three studied habitats, that is, crops, reservoirs and paths. Furthermore, the majority of common vole abundance peaks coincide with summer periods, especially in annual herbaceous crops. The spatial distribution of vole abundance exhibited a non-random pattern, characterized by spatial clustering. Particularly, the areas with higher significance of this clustering were located at the so-called 'Tierra de Campos' county, covering northern Valladolid, southern Palencia and north-eastern Zamora provinces. Periods of major incidence and concentration were 2013-2014, 2016-2017 and 2019. Common vole temporal occupancy patterns demonstrate that colonization occurred simultaneously in alfalfa reservoirs and dispersion paths, both preceding settlement in annual herbaceous crop plots. The geographic information system (GIS)-hotspots-based methodology proposed here can be valuable for stakeholders involved in integrated management of the common vole, serving as a detector of pest-prone areas in both space and time. These hotspots are useful for predicting future surveillance areas that accurately reflect pest colonization patterns. It was found that common vole abundance along dispersal paths acts as a source of dissemination, preceding the risk of colonization in annual herbaceous crop plots. © 2024 The Author(s). Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Genome-wide relaxation of selection and the evolution of the island syndrome in Orkney voles.

Island populations often experience different ecological and demographic conditions than their counterparts on the continent, resulting in divergent evolutionary forces affecting their genomes. Random genetic drift and selection both may leave their imprints on island populations, although the relative impact depends strongly on the specific conditions. Here we address their contributions to the island syndrome in a rodent with an unusually clear history of isolation. Common voles (Microtus arvalis) were introduced by humans on the Orkney archipelago north of Scotland >5000 years ago and rapidly evolved to exceptionally large size. Our analyses show that the genomes of Orkney voles were dominated by genetic drift, with extremely low diversity, variable Tajima's D, and very high divergence from continental conspecifics. Increased d N/d S ratios over a wide range of genes in Orkney voles indicated genome-wide relaxation of purifying selection. We found evidence of hard sweeps on key genes of the lipid metabolism pathway only in continental voles. The marked increase of body size in Orkney-a typical phenomenon of the island syndrome-may thus be associated to the relaxation of positive selection on genes related to this pathway. On the other hand, a hard sweep on immune genes of Orkney voles likely reflects the divergent ecological conditions and possibly the history of human introduction. The long-term isolated Orkney voles show that adaptive changes may still impact the evolutionary trajectories of such populations despite the pervasive consequences of genetic drift at the genome level.

Impact of environmental factors on small mammals and their fleas

Abstract Small mammals in the Horšianska dolina-Valley were researched first in 1984–86 and later in 2016–19. This paper seeks to identify and assess the species composition and dynamics of fleas (Siphonaptera) found in the valley on small mammals of Rodentia and Eulipotyphla and to compare findings from these two periods separated by thirty years. The results indicated deviations in small mammal and flea community similarity as well as in the small mammal indicator species for each habitat and flea indicator species for the small mammal species. A total 12 different species of small mammals were captured, and 12 flea species were collected. Eight flea species were detected in 1984–86 and ten species were found in 2016–19. In both periods A. penicilliger, C. assimilis, C. solutus, D. dasycnema, N. fasciatus were captured. The differences were that in the earlier period, the species uncovered were C. congener, P. bidentata, and P. soricis, while in the later period they were C. agyrtes, C. bisoctodentatus, M. turbidus, and R. integella. In the relationship between fleas and small mammals, C. solutus was the specific species for Apodemus flavicollis, while A. penicilliger and C. agyrtes were for Clethrionomys glareolus, and C. assimilis was for Microtus arvalis.

Effects of an Anti-Fertility Product on Reproductive Structures of Common Vole Males and Residues of Compounds.

Some rodent species cause significant damage to agriculture and forestry, and some can transmit pathogens to humans and livestock. The common vole (Microtus arvalis) is widespread in Europe, and its population outbreaks have resulted in massive crop loss. Bait-based fertility control could contribute to rodent pest management. Bait containing 4-vinylcyclohexene diepoxide (VCD) and triptolide (TP), registered as ContraPest®, was delivered to male common voles for 14 or 28 consecutive days. The effects on reproductive structures and residues in the liver and testes were assessed. There was no effect on testis weight, sperm viability, sperm motility and oxidative stress in sperm cells. Results regarding the mitochondrial membrane potential of sperm, DNA fragmentation and progressively motile sperm cells were inconclusive. However, there was an increase in morphological sperm defects in voles treated for 14/28 days and fewer normal sperm cells in voles treated for 28 days. There were no TP residues in the testes, few and low TP residues and no VCD residues in liver tissues, making considerable secondary exposure to non-target species unlikely. Treatments with VCD + TP seemed to have minor effects on the reproductive organs of males. Further studies should evaluate the effect of VCD + TP on females and on the reproductive success of common voles and other pest rodent species.

WILDbase: towards a common database to improve wildlife disease surveillance in Europe.

BackgroundTo be better prepared for emerging wildlife-borne zoonoses, we need to strengthen wildlife disease surveillance.AimThe aim of this study was to create a topical overview of zoonotic pathogens in wildlife species to identify knowledge gaps and opportunities for improvement of wildlife disease surveillance.MethodsWe created a database, which is based on a systematic literature review in Embase focused on zoonotic pathogens in 10 common urban wildlife mammals in Europe, namely brown rats, house mice, wood mice, common voles, red squirrels, European rabbits, European hedgehogs, European moles, stone martens and red foxes. In total, we retrieved 6,305 unique articles of which 882 were included.ResultsIn total, 186 zoonotic pathogen species were described, including 90 bacteria, 42 helminths, 19 protozoa, 22 viruses and 15 fungi. Most of these pathogens were only studied in one single animal species. Even considering that some pathogens are relatively species-specific, many European countries have no (accessible) data on zoonotic pathogens in these relevant animal species. We used the Netherlands as an example to show how this database can be used by other countries to identify wildlife disease surveillance gaps on a national level. Only 4% of all potential host-pathogen combinations have been studied in the Netherlands.ConclusionsThis database comprises a comprehensive overview that can guide future research on wildlife-borne zoonotic diseases both on a European and national scale. Sharing and expanding this database provides a solid starting point for future European-wide collaborations to improve wildlife disease surveillance.

Habitat type modulates sharp body mass oscillations in cyclic common vole populations

Cyclic rodent populations exhibit pronounced changes in body mass associated with the population cycle phase, long-known as Chitty effect. Although Chitty effect is a common epiphenomenon in both America and Europe, there is still incomplete evidence about the generality of these patterns across the entire range of most species. Moreover, despite decades of research, the underlying factors driving Chitty effect remains poorly understood. Here, we examined the influence of intrinsic and extrinsic factors that may underlie observed patterns in vole size variation in the Iberian common vole Microtus arvalis asturianus. We weighed and measured 2816 adult voles that were captured during 6 trapping periods. Vole numbers and body mass showed strong period- and phase-related variation both in females and males, demonstrating marked Chitty effect in the studied population. Body mass of adult males correlated with body length, evidencing that heavier males are also structurally larger. Statistical models showed that probability of occurrence of large-sized vole (> 37 g) was significantly more likely in reproductive males, during increase and peak phases, and it was modulated by habitat, with crop fields and field margins between crops showing an increased likelihood. We suggest an effect of the habitat on vole body mass mediated by predation.

Echinococcus multilocularis infection affects risk-taking behaviour in Microtus arvalis: adaptive manipulation?

Manipulation of host behaviour by parasites to enhance transmission to the next host is a fascinating phenomenon that has interested scientists since the 1970s. It has been proposed that infection with the cestode Echinococcus multilocularis produces an impairment of the antipredatory behaviour in the rodent intermediate host common vole, Microtus arvalis, which may facilitate transmission of the tapeworm to the canid final host. In this study, we observed the behaviour of infected common voles at 12 weeks post-infection, when protoscoleces production and maturation commonly occurs, in order to assess behavioural changes compared to uninfected controls, that might ease predation in the wild. Infected and uninfected voles were monitored for 24 h to observe their spontaneous activity. In addition, the next day, both infected and uninfected voles were subjected to 4 different behavioural tests: open field test, barrier test, platform test and air-puff test in a running wheel. No significant difference between uninfected and infected voles emerged during the behavioural tests. However, observation of spontaneous activity revealed that infected voles increased their feeding frequency and spent significantly more time above bedding even when not eating, compared to the uninfected controls. In the wild, these behavioural changes increase the animals exposure to predators, raising their chance of becoming prey. These findings are the first direct evidence consistent with behavioural manipulation by E. multilocularis on common voles.

Odour preferred males led to a higher offspring number in the common vole

Very recently, an interesting phenomenon was described in the common vole; vole parents with similar locomotor ability produced significantly larger litters. Positive assortative mating is a tendency to prefer individuals with similar phenotypes. We tested whether this also applies to smell similarity. Odour preference was tested in a T-maze, where each female was presented with two male odours, i.e. shavings together with feces and urine from home boxes. After female preference was established, the female was either paired with a preferred male (chosen) or paired with a non-preferred male (opposite choice). For analysis of the relationship to odour preference, genotyping of major histocompatibility complex (MHC) Class II DRB was done using amplicon sequencing. In the set of 45 individuals from two populations, we recovered 38 nucleotide haplotypes (alleles). Similarity of alleles in parent pairs according to the indexes of Sørensen–Dice (S–D) and Jaccard were calculated. Values of these indexes in parental pairs with preferred males were significantly higher (more similar) than in not preferred. The number of offspring in parental pairs with preferred males were significantly higher than in not preferred males. However, there is no correlation between the mentioned indexes and the number of offspring. The relationship between the success of reproduction and alleles is not clear-cut, this may be influenced by the measure of similarity we used, or by something that we could not detect.

Structure of the Hybrid Zone between Allied Species of the Common Vole, Microtus arvalis and M. obscurus: Influence of Genetic Factors and Landscape-Geographical Conditions

Structure of the Hybrid Zone between Allied Species of the Common Vole, Microtus arvalis and M. obscurus: Influence of Genetic Factors and Landscape-Geographical Conditions

Second-breeding events of the common kestrel (Falco tinnunculus) in the Czech Republic

Abstract The laying of two clutches in a single breeding season can increase the fitness of the parent birds and stabilise a population. Our study analysed the rate of second-breeding attempts in the common kestrel (Falco tinnunculus) population in the Czech Republic. In the research, a total of 8,049 common kestrel nests were checked between 1979 and 2019. Of the 1,197 breeding events recorded during the study period in Moravia, 1,184 cases were first clutches and 13 cases (1.1%) were second breeding attempts. In Bohemia, a total of 6,852 breeding events were observed during the same study period, of which 6,811 were first clutches, 32 cases were late breeding attempts and 9 cases (0.1%) were second clutches. Second breeding events occurred only rarely, mainly during population outbreaks of prey species. During the mass outbreak of the common vole (Microtus arvalis) population in the Czech Republic in 2019, 5 (5.2%) of the 97 studied common kestrel pairs nested for the second time in Moravia; in Bohemia, second clutches were recorded in 3 (8.1%) of the 37 studied pairs. The second breeding attempts were recorded at an altitude of 272 ± 123 m (mean ± SD, range 179–560 m, n = 22) and commenced between 24 June and 25 July (29 June on average, n = 5). The range of the time gap between the first and the second clutch was 54 to 92 days (70.8 ± 16.3 days, n = 5). The second clutches consisted of an average of 4.1 ± 0.9 eggs (range 3–6, n = 12), with an average of 3.7 ± 0.9 fledglings in each successful breeding pair (range 2–5, n = 11) and 2.6 ± 1.9 fledglings for each initiated breeding attempt (range 0–5, n = 16).

Comparison of baiting strategies in common vole management.

Worldwide, pest rodents can cause extensive damage to agriculture, forestry, food storage, and infrastructure and pose a risk to public health and livestock due to the spread of zoonotic pathogens. In Europe, the most common pest rodent species is the common vole (Microtus arvalis). Management during periodic outbreaks largely relies on rodenticidal bait with zinc phosphide. Efficient baiting with rodenticides or possibly anti-fertility products in the future require baiting methods that allow a sufficient proportion of the population to consume an effective dose of bait. We used a bait with the quantitative marker ethyl-iophenoxic acid (Et-IPA) to evaluate baiting strategies in enclosure experiments. This wheat-based bait with Et-IPA was placed in bait boxes or directly into the tunnel system entrances in different seasons and common vole abundances. Voles were live-trapped, individually marked and blood samples were collected to relate Et-IPA blood residues to bait uptake. The percentage of animals consuming bait was not heavily affected by the baiting strategy but voles had higher Et-IPA blood residues if tunnel baiting was used in autumn and if bait boxes were used in winter. Non-reproductive as well as lighter animals tended to have higher Et-IPA blood residues than reproductive individuals, whereas sex had no effect. Population density had a negative effect on the probability of residues present as well as on Et-IPA blood concentration. The results of this study might help to improve baiting techniques to manage overabundant rodent pest species regardless of the compounds to be delivered. © 2024 Julius Kühn-Institut. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Exploring the influence of density-dependence and weather on the spatial and temporal variation in common vole (Microtus arvalis) abundance in Castilla y León, NW Spain.

The common vole has invaded the agroecosystems of northwestern Spain, where outbreaks cause important crop damage and management costs. Little is yet known about the factors causing or modulating vole fluctuations. Here, we used 11 years of vole abundance monitoring data in 40 sites to study density-dependence and weather influence on vole dynamics. Our objective was to identify the population dynamics structure and determine whether there is direct or delayed density-dependence. An evaluation of climatic variables followed, to determine whether they influenced vole population peaks. First- and second-order outbreak dynamics were detected at 7 and 33 study sites, respectively, together with second-order variability in periodicity (2-3 to 4-5-year cycles). Vole population growth was explained by previous year abundance (mainly numbers in summer and spring) at 21 of the sites (52.5%), by weather variables at 11 sites (27.5%; precipitation or temperature in six and five sites, respectively), and by a combination of previous abundance and weather variables in eight sites (20%). We detected variability in vole spatiotemporal abundance dynamics, which differs in cyclicity and period. We also found regional variation in the relative importance of previous abundances and weather as factors modulating vole fluctuations. Most vole populations were cyclical, with variable periodicity across the region. Our study is a first step towards the development of predictive modeling, by disclosing relevant factors that might trigger vole outbreaks. It improves decision-making processes within integrated management dealing with mitigation of the agricultural impacts caused by voles. © 2023 Society of Chemical Industry.

Effectiveness of rodenticides based on bromadiolone against the common vole Microtus arvalis Pall. in laboratory conditions

The number of preparations based on the 2nd generation anticoagulant bromadiolone in the catalog of plant protection products against mouse-like rodents in Russia is growing, but information about the effectiveness of this active substance against the common vole Microtus arvalis Pall. is very limited. In a laboratory study of bromadiolone, acute LD50 = 3.3 + 0.8 mg/kg was shown, increased resistance equal to LD50 5.2 mg/kg was detected in individuals selected on the basis of resistance to anticoagulants of the 1st generation against the background of a diet with a high content of vitamin K (green food). Rodents overcame relative stability in the laboratory with a 3-day consumption of bait (0.05 g of bromadiolone/kg). In the field, the effectiveness of treatments may decrease and lead to the selection of resistant individuals. Bromadiolone-based rodenticidal baits are suitable for controlling the number of mouse-like rodent communities with the dominance of the common vole and carrying out treatments at the beginning of population growth.

Genome-wide support for incipient Tula hantavirus species within a single rodent host lineage.

Evolutionary divergence of viruses is most commonly driven by co-divergence with their hosts or through isolation of transmission after host shifts. It remains mostly unknown, however, whether divergent phylogenetic clades within named virus species represent functionally equivalent byproducts of high evolutionary rates or rather incipient virus species. Here, we test these alternatives with genomic data from two widespread phylogenetic clades in Tula orthohantavirus (TULV) within a single evolutionary lineage of their natural rodent host, the common vole Microtus arvalis. We examined voles from forty-two locations in the contact region between clades for TULV infection by reverse transcription (RT)-PCR. Sequencing yielded twenty-three TULV Central North and twenty-one TULV Central South genomes, which differed by 14.9-18.5 per cent at the nucleotide and 2.2-3.7 per cent at the amino acid (AA) level without evidence of recombination or reassortment between clades. Geographic cline analyses demonstrated an abrupt (<1 km wide) transition between the parapatric TULV clades in continuous landscape. This transition was located within the Central mitochondrial lineage of M. arvalis, and genomic single nucleotide polymorphisms showed gradual mixing of host populations across it. Genomic differentiation of hosts was much weaker across the TULV Central North to South transition than across the nearby hybrid zone between two evolutionary lineages in the host. We suggest that these parapatric TULV clades represent functionally distinct, incipient species, which are likely differently affected by genetic polymorphisms in the host. This highlights the potential of natural viral contact zones as systems for investigating the genetic and evolutionary factors enabling or restricting the transmission of RNA viruses.

Strengthening the genomic surveillance of Francisella tularensis by using culture-free whole-genome sequencing from biological samples.

Francisella tularensis is a highly infectious bacterium that causes the zoonotic disease tularemia. The development of genotyping methods, especially those based on whole-genome sequencing (WGS), has recently increased the knowledge on the epidemiology of this disease. However, due to the difficulties associated with the growth and isolation of this fastidious pathogen in culture, the availability of strains and subsequently WGS data is still limited. To surpass these constraints, we aimed to implement a culture-free approach to capture and sequence F. tularensis genomes directly from complex samples. Biological samples obtained from 50 common voles and 13 Iberian hares collected in Spain were confirmed as positive for F. tularensis subsp. holarctica and subjected to a WGS target capture and enrichment protocol, using RNA oligonucleotide baits designed to cover F. tularensis genomic diversity. We obtained full genome sequences of F. tularensis from 13 animals (20.6%), two of which had mixed infections with distinct genotypes, and achieved a higher success rate when compared with culture-dependent WGS (only successful for two animals). The new genomes belonged to different clades commonly identified in Europe (B.49, B.51 and B.262) and subclades. Despite being phylogenetically closely related to other genomes from Spain, the detected clusters were often found in other countries. A comprehensive phylogenetic analysis, integrating 599 F. tularensis subsp. holarctica genomes, showed that most (sub)clades are found in both humans and animals and that closely related strains are found in different, and often geographically distant, countries. Overall, we show that the implemented culture-free WGS methodology yields timely, complete and high-quality genomic data of F. tularensis, being a highly valuable approach to promote and potentiate the genomic surveillance of F. tularensis and ultimately increase the knowledge on the genomics, ecology and epidemiology of this highly infectious pathogen.