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Et donc nous avons. Females suspected of being pregnant were transferred to individual cages to establish a breeding colony of voles. Each cage contained a thick layer of standard sawdust c. To prevent the development of ectoparasites i. During handling, all voles from the breeding colony were inspected for ectoparasites in order to ensure vector-free conditions in the cages and animal house.
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No ectoparasites were noted at any time after initial caging, neither on the dams nor on the pups. Non-pregnant females were then released at their original trap lines. Pups were kept together with their dams for one month. In the third week of life we weighed the pups and collected blood samples from the tail tip of each individual. Then pups and dams were released at the trap lines at which the dams had been caught originally. Two thin blood smears were prepared from drops of blood taken from the heart autopsies or tail tip alive processing of wild-caught voles and pups.
Molecular techniques were used for the detection of Babesia in adult voles males and females , embryos and pups. Embryos were isolated from the uterus and individually processed autopsies following two washes in sterile water, to minimize contamination with maternal blood. We autopsied embryos from 20 litters 16 obtained in and 4 litters from from females that succumbed under anesthesia Fig.
Organs mainly hearts and lungs together, and brains, livers, spleens and kidneys, if distinguishable were isolated from embryos with sterile dissecting instruments. The remaining 13 litters were in earlier stages of pregnancy 1—2 trimester and were too small diameter of the embryo together with amniotic sac less than 1 cm to enable the isolation of specific internal organs. The scheme of the study.
Detection and genotyping of B. The primers and thermal profile used in this study have been described previously [ 24 ]. Negative controls were performed in the absence of template DNA. In the second step nested reaction , primers GR and GF were used.
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As positive controls we used the genomic DNA of B. PCR products were subjected to electrophoresis on a 1. Selected PCR products from voles trapped in and , all pregnant females and dams, and from at least two pups per litter were sequenced by a private company Genomed S. Consensus sequences were compared with sequences deposited in the GenBank database using BioEdit tool [ 29 ]. The statistical approach adopted has been documented comprehensively in our earlier publications [ 30 — 33 ]. Prevalence percentage of animals infected was analysed by maximum likelihood techniques based on log-linear analysis of contingency tables.
Beginning with the most complex model, involving all possible main effects and interactions, those combinations not contributing significantly to explanation of variation in the data were eliminated stepwise, beginning with the highest-level interaction. Statistical analysis was carried out using SPSS v. Abundance of B. When samples were only positive by PCR, an intensity of 0. The number of wild-caught voles by year of study, host species, age and sex is provided in Table 1. In total, voles of three species were trapped and sampled: common voles, M. Prevalence of B.
Overall, prevalence of B. In prevalence of B. Among field voles, prevalence of B. Data on the abundance of B. Abundance was calculated on the basis of microscopical observation of blood smears for wild-caught M. The mean abundance of B. Mean abundance of B. There were no significant differences in mean abundance of B. Altogether female voles were trapped, among which 44 were pregnant thus providing 27 litters embryos and pups from Babesia -positive females and 17 litters from Babesia -negative mothers for analysis of vertical transmission Fig.
The overall prevalence of B. Highest prevalence was in pregnant female M. There were significant differences in the prevalence of B. All pregnant M. Of the 44 pregnant females, 11 were kept in captivity until pup delivery, and these provided 10 litters from Babesia -positive females host species and litter size provided in Table 5 and 1 litter 6 pups from a Babesia -negative M.
Reliable analysis of the prevalence of infections in embryos was possible for embryos from another 20 litters [14 litters from Babesia -positive females Fig. These embryos were of an appropriate size to enable autopsy and isolation of organs heart with lungs, for all samples.
Among Babesia -positive pregnant females, 11 were M. Babesia -positive tissues heart and lungs in embryos were found in Among the three host species, no B. Among two Babesia -positive M. In summary, B. Babesia microti DNA was detected in all M. In one litter from a Babesia -positive M.
Among 9 litters from M. There was no significant difference between male and female pups born from infected dams: Interestingly, a higher percentage of Babesia -positive offspring was obtained in the first year of the study when we focused on embryos, in comparison to , when the focus was on pups Tables 4 and 5.
Two litters 6 pups of M. The other litter was delivered by the only one Babesia -negative M. All the other pups delivered by 9 Babesia -positive dams 40 Babesia -positive and 10 Babesia -negative pups; Table 5 survived until the end of the experiment. The mean litter size for all 11 dams was 5. The effect of Babesia infection in the dam on the litter size could not be reliably analyzed as there was only one litter from a Babesia -negative dam with 6 pups and the mean litter size for Babesia -positive dams was 5.
The mean body mass of M. Male pups of M. The mean weight was almost identical for Babesia -positive pups and Babesia -negative pups The abundance of B. Alignment of the sequences revealed that two main B. Lower similarity for several sequences was the result of some non-specific background amplification of DNA. The IRU2 Munich-like genotype was dominant among positive embryos Among the B. The B. The IRU2 genotype of B. The final step of our study on vertical transmission was to determine the B. In 3 cases the genotypes of B. In 4 cases the B. Thus, the dominant B. Interestingly, for two females infected with the B.
In one case the genotype of B. In one case the B. In four other litters, where the B. In this study we reported a high prevalence of B. We also reported a complex circulation of two main rodent B. Although the present study focused primarily on the occurrence of vertical transmission of B. The first study on B. Overall prevalence of B. However, the markedly lower prevalence of B. Building on this first study, where B. A similar pattern was observed in abundance of B. Interestingly, both parameters were lowest in the third species, M.
This species is rarely reported from our study sites in the Mazury Lake District [ 36 ] and as our data show, this is a species of much lower significance as a reservoir host of B. Interestingly, over the long-term, there were a few years when B. Our finding that M. Surveys in the UK, where M. High prevalence of B. The overall prevalence in the community of voles in the current study was similar to prevalence in Omsk region, Russia We found intriguing the generally low infestation of I.
Therefore we tested the hypothesis that high prevalence of B. Quite clearly our observations, whether based on pregnant females-embryos or dams-pups, support our hypothesis, both revealing a high rate of vertical transmission in M. This rate of Babesia -positive offspring derived from Babesia -positive female voles may be compared with an overall prevalence of B. However, to enable a more meaningful comparison, estimation of Babesia -positive offspring should include also Babesia -negative offspring of Babesia -negative females.
To support the latter hypothesis on the negative impact of congenitally acquired Babesia infection , we compared selected parameters between Babesia -positive and Babesia -negative litters litter size and pups i. I would expect Ethereum to rally leading up to the event on the eighth of February. But I would be cautious of a pullback around or just after the launch. The growing demand from both the retail and institutional crowds can be attributed to the growing activity and hype surrounding the decentralized finance sector. The trading volume on decentralized exchanges has also been increasing steadily.
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