Archetype analysis and the PHATE algorithm as methods to describe and visualize pregnant women's levels of physical activity knowledge.

The knowledge of physical activity (PA) recommended for pregnant women and practical application of it has positive impact on the outcome. Nevertheless, it is estimated that in high-income countries over 40% of pregnant women are insufficiently physically active. One of the reasons is insufficient knowledge pregnant women have about allowed effort during pregnancy and both recommended and not recommended physical activities. Description of knowledge about physical activity the women have and distinguishing patterns of their knowledge is becoming an increasingly important issue. A common approach to handle survey data that reflect knowledge involves clustering methods or Principal Component Analysis (PCA). Nevertheless, new procedures of data analysis are still being sought. Using survey data collected by the Institute of Mother and Child Archetypal analysis has been applied to detect levels of knowledge reflected by answers given in a questionnaire and to derive patterns of knowledge contained in the data. Next, PHATE (Potential of Heat-diffusion for Affinity-based Trajectory Embedding) algorithm has been used to visualize the results and to get a deeper insight into the data structure. The results were compared with picture derived from PCA. Three archetypes representing three patterns of knowledge have been distinguished and described. The presentation of complex data in a low dimension was obtained with help of PHATE. The formations revealed by PHATE have been successfully described in terms of knowledge levels reflected by the survey. Finally, comparison of PHATE with PCA has been shown. Archetype analysis combined with PHATE provides novel opportunities in examining nonlinear structure of survey data and allows for visualization that captures complex relations in the data. PHATE has made it possible to distinguish sets of objects that have common features but were captured neither by Archetypal analysis nor PCA. Moreover, for our data, PHATE provides an image of data structure which is more detailed than interpretation of PCA.

Long-term pulmonary outcomes of young adults born prematurely: a Polish prospective cohort study PREMATURITAS 20.

BACKGROUND: The long-term consequences of prematurity are often not sufficiently recognized. To address this gap, a prospective cohort study, which is a continuation of the multicenter Polish study PREMATURITAS, was conducted, utilizing unique clinical data from 20 years ago. OBJECTIVE: The main goal was to evaluate lung function, detect any structural abnormalities using lung ultrasound, and assess psychological well-being in young adults born between 24 and 34 weeks of gestational age (GA). Additionally, the study aimed to investigate potential associations between perinatal risk factors and abnormalities observed in pulmonary function tests (PFTs) during adulthood. METHODS: The young survivors underwent a comprehensive set of PFTs, a lung ultrasound, along with the quality of life assessment. Information regarding the neonatal period and respiratory complications was obtained from the baseline data collected in the PREMATURITAS study. RESULTS: A total of 52 young adults, with a mean age of 21.6 years, underwent PFTs. They were divided into two groups based on GA: 24-28 weeks (n = 12) and 29-34 weeks (n = 40). The subgroup born more prematurely had significantly higher lung clearance index (LCI), compared to the other subgroup (p = 0.013). LCI ≥ 6.99 was more frequently observed in the more premature group (50% vs. 12.5%, p = 0.005), those who did not receive prenatal steroids (p = 0.020), with a diagnosis of Respiratory Distress Syndrome (p = 0.034), those who received surfactant (p = 0.026), and mechanically ventilated ≥ 7 days (p = 0.005). Additionally, elevated LCI was associated with the diagnosis of asthma (p = 0.010). CONCLUSIONS: The findings suggest pulmonary effects due to prematurity persist into adulthood and their insult on small airway function. Regular follow-up evaluations of young survivors born preterm should include assessments of PFTs. Specifically, the use of LCI can provide valuable insights into long-term pulmonary impairment.

Trends in caesarean section rates in Europe from 2015 to 2019 using Robson's Ten Group Classification System: A Euro-Peristat study.

OBJECTIVE: To assess changes in caesarean section (CS) rates in Europe from 2015 to 2019 and utilise the Robson Ten Group Classification System (TGCS) to evaluate the contribution of different obstetric populations to overall CS rates and trends. DESIGN: Observational study utilising routine birth registry data. SETTING: A total of 28 European countries. POPULATION: Births at ≥22 weeks of gestation in 2015 and 2019. METHODS: Using a federated model, individual-level data from routine sources in each country were formatted to a common data model and transformed into anonymised, aggregated data. MAIN OUTCOME MEASURES: By country: overall CS rate. For TGCS groups (by country): CS rate, relative size, relative and absolute contribution to overall CS rate. RESULTS: Among the 28 European countries, both the CS rates (2015, 16.0%-55.9%; 2019, 16.0%-52.2%) and the trends varied (from -3.7% to +4.7%, with decreased rates in nine countries, maintained rates in seven countries (≤ ± 0.2) and with increasing rates in 12 countries). Using the TGCS (for 17 countries), in most countries labour induction increased (groups 2a and 4a), whereas multiple pregnancies (group 8) decreased. In countries with decreasing overall CS rates, CS tended to decrease across all TGCS groups, whereas in countries with increasing rates, CS tended to increase in most groups. In countries with the greatest increase in CS rates (>1%), the absolute contributions of groups 1 (nulliparous term cephalic singletons, spontaneous labour), 2a and 4a (induction of labour), 2b and 4b (prelabour CS) and 10 (preterm cephalic singletons) to the overall CS rate tended to increase. CONCLUSIONS: The TGCS shows varying CS trends and rates among countries of Europe. Comparisons between European countries, particularly those with differing trends, could provide insight into strategies to reduce CS without clinical indication.

Occurrence of Dirofilaria repens in wild carnivores in Poland.

Dirofilaria repens is an expanding vector-borne zoonotic parasite of canines and other carnivores. Sub-clinically infected dogs constitute the most important reservoir of the parasite and the source of infection for its mosquito vectors. However, occurrence of D. repens infection in wild animals may contribute to the transmission of the parasite to humans and may explain the endemicity of filariae in newly invaded regions. The aim of the current study was to determine the occurrence of D. repens in 511 blood and spleen samples from seven species of wild carnivores (wolves, red foxes, Eurasian badgers, raccoons, raccoon dogs, stone martens, and pine martens) from different regions of Poland by means of a PCR protocol targeting the 12S rDNA gene. Dirofilaria repens-positive hosts were identified in seven of fourteen voivodeships in four of the seven regions of Poland: Masovia, Lesser Poland, Pomerania and Warmia-Masuria. The highest prevalence was found in Masovia region (8%), coinciding with the highest previously recorded prevalence in dogs in Central Poland. The DNA of Dirofilaria was detected in 16 samples of three species (total prevalence 3.13%). A low and similar percentage of positive samples (1.9%, 4.2% and 4.8%) was recorded among badgers, red foxes, and wolves, respectively. Dirofilaria repens-positive hosts were identified in seven of fourteen voivodships. Based on detection in different voivodeships, D. repens-positive animals were recorded in four out of the seven regions of Poland: in Masovia, Lesser Poland, Pomerania, and Warmia-Masuria. The highest prevalence of filariae was found in Masovia region (8%), reflecting the highest previously recorded prevalence in dogs (12-50%) in Central Poland. In summary, we conducted the first comprehensive study on the epidemiology of D. repens in seven species of wild hosts in all seven regions of Poland and identified the first case of D. repens infection in Eurasian badgers in Poland and the second in Europe.

Haplotypes of Dirofilaria repens from Poland and selected countries of Central, North-Eastern Europe and the Middle East: An evaluation on the relation between the genetic diversity and the geographic distribution of the fast-spreading parasite.

Subcutaneous dirofilariosis is a fast-spreading infection of dogs, and occasionally of other carnivores and humans. Several factors contribute to its spread, including climate change, which facilitates development and survival of Dirofilaria repens in the mosquito vector. Movement/relocation of infected definitive hosts (dogs) from endemic regions to non-endemic regions is another possible cause of local emergence and the presence of a wide variety of wild reservoirs of the parasite may also contribute to its spread. The main aim of this study was to evaluate the genetic diversity of D. repens from different regions of Europe and to evaluate the spread of identified haplotypes and their geographic origin. A total of 95 D. repens isolates were obtained from Central and Eastern Europe (Poland, Belarus, Ukraine, Austria, Romania), NE Europe (Lithuania, Latvia, Estonia), Italy and Israel. All but two positive samples were obtained from the blood of dogs while one positive sample was obtained from an adult worm from a human case from the Lublin area in SE Poland and one sample was obtained from Anopheles plumbeus mosquito from Austria. Genetic diversity in D. repens isolates was evaluated by PCR amplification and sequencing of three genetic markers, including two mitochondrial genes (mtDNA): the cytochrome c oxidase subunit I (COI) and dehydrogenase subunit I (NADH). Additionally, the genomic marker, internal transcribed spacer 1 (ITS-1) was amplified and sequenced. Haplotypes were differentiated based on sequence alignments by identifying Single Nucleotide Polymorphism (SNPs) using DnaSP and Mega X. PopArt was used to construct a haplotype network including all identified haplotypes. Both mtDNA sequences (COI and NADH) were combined together for phylogenetic and network analyses. Altogether 18 haplotypes (DR1-DR18) were identified in combined mtDNA markers among 95 analysed samples. Haplotype DR1 was the most common encompassing 66 isolates: 42 isolates from Poland (41 from dogs and one from a human), 13 from Lithuania, 4 from Latvia, 2 from Ukraine and 5 from Romania. All other haplotypes grouped around haplotype DR1 separated by 1-5 SNPs, forming a star-like shape. Haplotype DR2 was the second most common haplotype, formed by six isolates from Romania. Interestingly, haplotype DR3 was represented only by four isolates from Israel. The remaining 15 haplotypes were represented by 1-4 isolates of different origins. Our study showed that only minor genetic diversity was found in D. repens since all isolates appear to have clustered in or branched out from haplotype DR1 with 1-5 SNP differences. The genetic diversity appears to be governed by geographic origin since isolates from neighbouring populations (countries) appear to share unique haplotypes while other populations that are geographically distant from individual haplotypes.

Cytokine gene polymorphism and parasite susceptibility in free-living rodents: Importance of non-coding variants.

Associations between genetic variants and susceptibility to infections have long been studied in free-living hosts so as to infer the contemporary evolutionary forces that shape the genetic polymorphisms of immunity genes. Despite extensive studies of proteins interacting with pathogen-derived ligands, such as MHC (major histocompatilbility complex) or TLR (Toll-like receptors), little is known about the efferent arm of the immune system. Cytokines are signalling molecules that trigger and modulate the immune response, acting as a crucial link between innate and adaptive immunity. In the present study we investigated how genetic variation in cytokines in bank voles Myodes glareolus affects their susceptibility to infection by parasites (nematodes: Aspiculuris tianjensis, Heligmosomum mixtum, Heligmosomoides glareoli) and microparasites (Cryptosporidium sp, Babesia microti, Bartonella sp.). We focused on three cytokines: tumour necrosis factor (TNF), lymphotoxin alpha (LTα), and interferon beta (IFNß1). Overall, we identified four single nucleotide polymorphisms (SNPs) associated with susceptibility to nematodes: two located in LTα and two in IFNß1. One of those variants was synonymous, another located in an intron. Each SNP associated with parasite load was located in or next to a codon under selection, three codons displayed signatures of positive selection, and one of purifying selection. Our results indicate that cytokines are prone to parasite-driven selection and that non-coding variants, although commonly disregarded in studies of the genetic background of host-parasite co-evolution, may play a role in susceptibility to infections in wild systems.

Breeding in the pandemic: short-term lockdown restrictions in a European capital city did not alter the life-history traits of two urban adapters.

Humans are transforming natural habitats into managed urban green areas and impervious surfaces at an unprecedented pace. Yet the effects of human presence per se on animal life-history traits are rarely tested. This is particularly true in cities, where human presence is often indissociable from urbanisation itself. The onset of the SARS-CoV-2 outbreak, along with the resulting lockdown restrictions, offered a unique, "natural experiment" to investigate wildlife responses to a sudden reduction in human activity. We analysed four years of avian breeding data collected in a European capital city to test whether lockdown measures altered nestbox occupancy and life-history traits in terms of egg laying date, incubation duration and clutch size in two urban adapters: great tits (Parus major) and blue tits (Cyanistes caeruleus). Lockdown measures, which modulated human presence, did not influence any of the life-history traits investigated. In contrast, the interaction between year and tree cover, a distinct ecological attribute of the urban space, was positively associated with clutch size, a key avian life-history and reproductive trait. This highlights the importance of inter-year variation and habitat quality over human activity on urban wildlife reproduction. We discuss our results in the light of other urban wildlife studies carried out during the pandemic, inviting the scientific community to carefully interpret all lockdown-associated shifts in biological traits. Supplementary Information: The online version contains supplementary material available at 10.1007/s11252-022-01309-5.

Primary Congenital Hypothyroidism in Children Below 3 Years Old - Etiology and Treatment With Overtreatment and Undertreatment Risks, a 5-Year Single Centre Experience.

Worldwide neonatal screening for congenital hypothyroidism (CH) is a gold standard of active surveillance in newborns. Prompt diagnosis, subsequent timely treatment implementation, and proper dosage of levothyroxine (L-T4) are crucial for normal growth and development, especially of the central nervous system. However, overtreatment may have a potential negative impact on further neurodevelopment. We retrospectively analysed data of 99 newborns with CH diagnosis, referred to the Endocrinology Outpatient Clinic of the Institute of Mother and Child in Warsaw, Poland from the CH screening program from 2017 to 2021. We evaluated the diagnostic process and treatment up to the age of 3 years. We compared groups of children from the first and the second screening groups (FSG, SSG) in the neonatal screening with an evaluation of ultrasound examination (thyroid dysgenesis vs. gland in situ, GIS). The overtreatment and undertreatment risks were assessed and an analysis of the new TSH thresholds was performed. Treatment was implemented at a median of 9 days of life (3 - 27); 8 days (3 - 17) in FSG and 19 (6 - 27) in SSG. The dose of L-T4 differed between FSG and SSG at all three analysed time points (start of the therapy, 12 months, and 3 years) with significantly higher doses in FSG. The same was observed for the patients with thyroid dysgenesis vs. GIS. Screening TSH level was ≥ 28mIU/l in 91.7% of patients with thyroid dysgenesis in comparison to 74.0% of patients with GIS (p= 0.038). The optimally treated group (fT4 in the upper half of the reference range, according to the guidelines) was up to 58.0% of the children during the follow-up. The risk for overtreatment was present in 1/5 of the study group after 12 months and 1/4 after 3 years of L-T4 therapy. Analysis of new TSH thresholds showed an increased prevalence of mild hypothyroidism, GIS, and either euthyroid state or overtreatment while treating with lower L-T4 doses in comparison to the rest of the cohort. The study confirmed the general efficacy of the CH diagnostic pathway and the timely implemented L-T4 therapy. The suspected overtreatment after the first 12 months of L-T4 therapy requires consideration of the earlier diagnosis re-evaluation.

Tick-Borne Pathogens, Babesia spp. and Borrelia burgdorferi s.l., in Sled and Companion Dogs from Central and North-Eastern Europe.

Ticks are important vectors of numerous pathogens of medical and veterinary significance. The aim of the current study was to determine the prevalence of Babesia spp. and Borrelia burgdorferi s.l. in sled and pet dogs from Central and North-Eastern Europe. Neither Babesia spp. nor Borrelia burgdorferi s.l. infections were detected in sled dogs from seven countries (Poland, Lithuania, Latvia, Estonia, Belarus, Russia and Finland). The DNA of Babesia spp. was detected in 100% of symptomatic and 5.4% of asymptomatic pet dogs from Poland. Similarly, the DNA of Babesia spp. was identified in 82% of symptomatic and 3.8% of asymptomatic pet dogs from Ukraine. The DNA of Borrelia burgdorferi s.l. was detected in 4.4% of pet dogs. Molecular typing confirmed the presence of Babesia canis and Borrelia burgdorferi sensu stricto (s.s.) in selected samples. Four dogs were co-infected by B. canis and Borrelia burgdorferi s.l. Tick-borne pathogens constitute a serious health threat to pet dogs in Central and South-Eastern Europe, but were not observed among sled dogs from the same region of Europe nor in the Baltic countries.

Update on prevalence of Babesia canis and Rickettsia spp. in adult and juvenile Dermacentor reticulatus ticks in the area of Poland (2016-2018).

Ornate dog tick, Dermacentor reticulatus is an important vector of Babesia canis, and Rickettsia spp. and other pathogens of veterinary and public health interest. The current study is the first to investigate the long-term changes in prevalence of these pathogens in expanding tick populations in Central Europe. Molecular techniques (PCR, sequencing) were applied for the detection of pathogen DNA in adult (n = 2497) and juvenile ticks (1096 larvae and 410 nymphs). DNA of Rickettsia spp. was identified in 35% of adults and 12.6% of juvenile ticks. DNA of B. canis was detected in 3% of adult ticks and only in ticks from the Eastern region (regional prevalence 6%). As previously, no B. canis-positive ticks were found in Western Poland, including ticks from Wroclaw area (n = 298). DNA of B. canis was identified in 0.33% of juvenile ticks (in 3 pools of larvae and 2 nymphs) from the Eastern region. In the current study we confirmed high occurrence of R. raoultii in adults ticks from all four zones and relatively high prevalence of B. canis in the Eastern population of D. reticulatus, corresponding well with high incidence of canine babesiosis in this area of Poland. Finally, we confirmed R. raoultii and B. canis infection in all life stages of D. reticulatus ticks.

Occurrence of juvenile Dermacentor reticulatus ticks in three regions in Poland: the final evidence of the conquest.

BACKGROUND: Two populations of Dermacentor reticulatus ticks (Western and Eastern) in Poland are among the most dynamic tick populations in Central Europe. Expansion and settlement of ticks in new localizations depend on the presence of suitable hosts, for both adult and juvenile ticks. METHODS: The current study was planned to complement our previous studies on questing adult ticks and was focused on a collection of juvenile D. reticulatus ticks from rodents from three regions in Poland, defined by the presence/absence of adult ticks (regions of the Western and Eastern tick population and the gap area between them) to confirm the existence of stable populations. Rodent trapping was conducted in open habitats (fallow lands, wasteland and submerged meadows) in 2016-2018 in June, July and/or August to encompass seasonal peaks of larvae and nymph activity. RESULTS: Altogether, three tick species were collected, 2866 D. reticulatus, 2141 Ixodes ricinus and 427 Haemaphysalis concinna. Dermacentor reticulatus was the most common (72.3%) and abundant (mean 17.94 ± 2.62 ticks/rodent) tick species on rodents from the Eastern region; in the Western region infestation of rodents was only 6.8%. Ixodes ricinus was found in all three regions and was the only tick species collected from rodents from the gap area. Haemaphysalis concinna was noted only in the Western region. The highest infestation of juvenile D. reticulatus was recorded on voles (Myodes and Microtus spp.), infestation of I. ricinus was the highest on Apodemus mice, and the majority of H. concinna ticks were collected from root voles Alexandromys oeconomus. CONCLUSIONS: Our study confirmed a stable population of D. reticulatus in Eastern and Central Poland and a lower prevalence and mean abundance of this tick species among rodents from the Western region. A lack of juvenile D. reticulatus on rodents in Niewiadów confirmed the existence of the gap area, free of D. reticulatus ticks.

Pathogens detected in the tick Haemaphysalis concinna in Western Poland: known and unknown threats.

In recent years, a new focus of the relict tick Haemaphysalis concinna was discovered in Western Poland, near Wolsztyn, Greater Poland voivodeship. This species may play an important role in the circulation of pathogens of medical and veterinary importance. In the present study we tested 880 juvenile ticks collected from rodents, including 427 H. concinna, 443 Ixodes ricinus and 10 Dermacentor reticulatus for three of the most common pathogens vectored by ticks in Poland: Rickettsia and Babesia spp. and Borrelia burgdorferi s.l. Additionally, molecular techniques were applied for accurate identification of tick host species (the voles Microtus and Alexandromys). Our study found differences in the range and prevalence of vectored pathogens between the three tick species. DNA of all three pathogens was found in I. ricinus. In juvenile H. concinna, DNA of Babesia microti, Borrelia afzelii and Rickettsia sp. was identified. Moreover, DNA of a new unnamed Babesia species related to B. crassa, was found in two H. concinna nymphs. This genotype of Babesia was previously identified in H. concinna in the Far East and then in Central Europe. DNA of Rickettsia raoulti and B. afzelii was detected in D. reticulatus nymphs. Among rodent hosts, Alexandromys oeconomus seems to be host of the highest significance for juvenile tick stages and was the only host species with B. afzelii detected in blood samples. Using phylogenetic methods, we confirmed a clear division between rodents from the genera Microtus and Alexandromys. Moreover, we found that A. oeconomus trapped in Western Poland clustered with a Central European A. oeconomus allopatric phylogroup.

Long-term study of Borrelia and Babesia prevalence and co-infection in Ixodes ricinus and Dermacentor recticulatus ticks removed from humans in Poland, 2016-2019.

BACKGROUND: Lyme borreliosis (LB) is the most common vector-borne disease in Europe. Monitoring changes in the prevalence of different Borrelia species in ticks may be an important indicator of risk assessment and of differences in pathogenicity in humans. The objective of our study was to assess the prevalence, co-infection and distribution of Borrelia and Babesia species in ticks removed from humans in a large sample collected during a study period of 4 years. METHODS: The ticks were collected throughout Poland from March to November over 4-year period from 2016 to 2019. All ticks (n = 1953) were morphologically identified in terms of species and developmental stage. Molecular screening for Borrelia and Babesia by amplification of the flagellin gene (flaB) or 18S rRNA marker was performed. Pathogen identity was confirmed by Sanger sequencing or PCR-restriction fragment length polymorphism analysis. RESULTS: The ticks removed from humans in Poland during this study belonged to two species: Ixodes ricinus (97%) and Dermacentor reticulatus (3%). High Borrelia prevalence (25.3%), including B. miyamotoi (8.4%), was confirmed in Ixodes ricinus ticks removed from humans, as was the change in frequency of occurrence of Borrelia species during the 4-year study. Despite Babesia prevalence being relatively low (1.3%), the majority of tested isolates are considered to be pathogenic to humans. Babesia infection was observed more frequently among Borrelia-positive ticks (2.7%) than among ticks uninfected with Borrelia (0.8%). The most frequent dual co-infections were between Borrelia afzelii and Babesia microti. The presence of Borrelia was also confirmed in D. reticulatus (12.7%); however the role of these ticks in spirochete transmission to susceptible hosts is still unclear. CONCLUSIONS: Although the overall risk of developing LB after a tick bite is low in Europe, knowledge of the prevalence and distribution of Borrelia and Babesia species in ticks might be an important indicator of the risk of both these tick-borne diseases.

Monitoring the expansion of Dermacentor reticulatus and occurrence of canine babesiosis in Poland in 2016-2018.

BACKGROUND: The significance of tick-borne diseases has increased considerably in recent years. Because of the unique distribution of the tick species Dermacentor reticulatus in Poland, comprising two expanding populations, Eastern and Western that are separated by a Dermacentor-free zone, it is important to conduct studies on the process of tick expansion and emergence of canine babesiosis. The main aim of the current study was to monitor the expansion of D. reticulatus populations from spring 2016 to autumn 2018 to determine (1) the actual geographical range of this tick species, and (2) and the seasonal/annual shift in range limits and changes in distance between Western and Eastern populations of ticks (the size of the non-endemic area). METHODS: Ticks were collected in spring/autumn during a 3-year study. From each season and year at least three pairs of sites from the Western and Eastern populations were selected. Then the mean distance between paired sites was calculated for each season and year. We collected and analyzed data from veterinary clinics on the number of canine babesiosis cases treated in the clinic during a whole year (2018). RESULTS: Accordingly, further expansion of the two D. reticulatus populations was recorded, mainly along river basins. Marked colonization of the gap zone was observed, with a mean annual shift in the range of 2.5-10 km and a steadily decreasing distance between the two tick populations. The occurrence of babesiosis in different regions revealed low numbers of cases in Western Poland (19 cases/year) and the gap area (only 7 cases/year) and high incidence (up to 250 cases/1000 dogs) and fatality (total 3.65%) in Central and Eastern Poland. Strong associations were found geographically between tick and babesiosis occurrence and temporally in the seasonal patterns of occurrence of ticks and outbreaks of babesiosis. CONCLUSIONS: We documented the shift in range limits and continued process of colonization of the gap zone accompanied by the emergence of canine babesiosis in the Eastern expansion zone. Updated maps of the distribution of ticks and occurrence of babesiosis in different regions of Poland have allowed us to predict of the emergence of pathogens vectored by D. reticulatus. Incidence (per 1000 dogs) of canine babesiosis in veterinary clinics by current range of D. reticulatus.

Emerging risk of Dirofilaria spp. infection in Northeastern Europe: high prevalence of Dirofilaria repens in sled dog kennels from the Baltic countries.

Dirofilariasis is a fast-spreading disease of dogs and humans in Europe. We investigated whether Dirofilaria spp. have spread northwards in Europe, invading the Baltic countries. Altogether, 424 blood samples were collected from eight countries in the period 2017-2019, including 227 samples from sled dogs and 197 samples from other dogs. PCR amplification and sequencing were conducted employing three genetic markers (mitochondrial [mt] 12S rDNA, mt cytochrome c oxidase subunit I [COI] gene and mt dehydrogenase subunit I [NAD1] gene). The SNAP test (IDEXX) for detection of D. immitis infections was also implemented. The DNA of D. repens was detected in 59 of 424 dogs (prevalence 13.9%). D. repens was found in sled dogs from Lithuania, Latvia, Poland and Belarus. Only one dog from Estonia was infected, apparently an imported case. The highest prevalence was recorded in Lithuania (38%). Among pet dogs from the Ukraine, six dogs tested positive (3.8%). Our study has revealed a high prevalence of D. repens infections in Lithuania and Latvia, but no evidence for spread of the heartworm D. immitis. We conclude that sled dog kennels constitute hot spots for D. repens transmission.

The red fox (Vulpes vulpes), a possible reservoir of Babesia vulpes, B. canis and Hepatozoon canis and its association with the tick Dermacentor reticulatus occurrence.

The red fox (Vulpes vulpes) is known to be a reservoir host of various vector-borne protozoan parasites. Yet, the impact of these parasitic infections on the health status of the red fox is unknown. In this study, we compared the prevalence of haemoprotozoa in juvenile and adult foxes from four regions in Poland. The abundance of questing adult Dermacentor reticulatus ticks was shown to differ between regions and include a tick-endemic and non-endemic region, as well as two zones in which D. reticulatus recently expanded: an eastern and a western zone. Spleen and kidney enlargement indices were compared between infected, co-infected and non-infected foxes to evaluate pathogenic output of parasitic infections. Blood samples were collected from 383 necropsied red foxes and 25 live-trapped red fox cubs. The weights of spleens, kidneys and bodies were recorded for each necropsied individual. Blood and spleen DNA samples were screened by PCR to detect 18S rRNA and cox1 gene fragments of Babesia spp., and 18S rRNA of Hepatozoon spp. Prevalence of infection and mean organ weight indices were compared in relation to fox age and sex, origin (zone) and infection status. Hepatozoon canis (174/381; 45.7%) and Babesia vulpes (117/381; 30.7%) were the dominant pathogens infecting adult foxes. Babesia canis DNA was detected in 9 (2.4%) individuals. Two fox cubs from Western Poland were infected with B. vulpes (2/14; 14%), while H. canis infection (16/25; 59%) was detected in cubs from all three regions where trapping was carried out. Infection with B. vulpes was associated with significantly higher weights of spleens and kidneys. Spleen weight/ body weight and mean kidney weight/ body weight indices were shown as relevant in evaluating changes occurring during infection with B. vulpes. Babesia and Hepatozoon spp. infections differed in red foxes from D. reticulatus-endemic, non-endemic and newly inhabited areas. The prevalence of B. canis in foxes from four regions of Poland reflects the prevalence recorded in questing adult D. reticulatus collected in these areas. This finding suggests a contribution of red foxes to the establishment of new foci of B. canis-infected D. reticulatus ticks in areas recently invaded by this tick species. Spleen and kidney enlargement was identified in foxes infected with B. vulpes, particularly in individuals co-infected with B. vulpes and H. canis.

Ectoparasites of red foxes (Vulpes vulpes) with a particular focus on ticks in subcutaneous tissues.

In the present study, we identified the ectoparasite communities of red foxes in three regions of Poland that encompassed two endemic regions for the occurrence of Dermacentor reticulatus, as well as a region that is free of this tick species ('gap' area). Our study sites were selected to enable the role of foxes as hosts for juvenile (nest dwelling) and adult (exophilic) D. reticulatus ticks to be determined, and to assess their contribution to the spread of this important vector of Babesia canis. We compared also ectoparasite communities between adult foxes with those of fox cubs. Finally, we carried out a systematic search for subcutaneous ticks determining their prevalence and abundance. In 2016-2018, 366 adult foxes and 25 live-trapped cubs were examined for ectoparasites. Ectoparasites were identified based on morphological features, PCR amplification and sequencing. The total prevalence of ectoparasites was higher in cubs (68%) than in adults (62.8%). In adults, 15 parasite species were recorded, including four tick species, seven flea species, scabies, and one Anopluran species each in the genera Felicola and Lipoptena. In cubs, six ectoparasite species were found, including Ixodes kaiseri, a species not found in adults. Although Ixodes ricinus and D. reticulatus were the dominant tick species on adult foxes, no D. reticulatus ticks were found on cubs. Subcutaneous ticks were common (38%) and abundant in all areas. Molecular analysis of subcutaneous nodules allowed the identification of 17 I. ricinus and five D. reticulatus. In conclusion, red foxes play a minor role as hosts of D. reticulatus.

The Efficiency of Live-Capture Traps for the Study of Red Fox (Vulpes vulpes) Cubs: A Three-Year Study in Poland.

Safe and efficient techniques for the live capture of carnivores are limited. In this study, we identified some of the factors that could affect the success of capturing red fox cubs with live capture traps (also known as cage traps). During a three-year period, we analysed 32 captures of 25 fox cubs (1.3 captures/fox). We assessed the impact of the following factors: sex of animals, month of trapping, weather conditions recorded for each trap-night, the willingness of cubs to explore and enter cage traps, the researchers' activity around den complexes before trapping and distances to the nearest village or farm. The overall trap rate (32 captures, including recaptured cubs) and the trap rate for individual cubs (25 captures) was 11.2 cubs/100 trap-nights and 8.7/100 trap-nights, respectively. Animals other than foxes were captured only three times, thus the selectivity of the cage-trapping method was high (32/35 = 91.4%). The probability of capturing one cub per night was 70.2% (32 cubs/ 47 nights). Cubs inhabiting dens in the vicinity of human settlements were less likely to explore and enter traps. Vixens were more likely to relocate their litters if the activity of the staff setting the traps was intense at the trapping site. The success of trapping was higher during poor weather as, for example, during rain or thunderstorms. None of the trapped animals suffered any injuries. Whereas cage trapping can be an effective and safe capture method for juvenile foxes, capture efficiency is affected by the experience of the trappers and a range of other factors including weather and distance to human settlements.

The prognostic role of BRAF and WNT pathways activation in kinase inhibitors-naïve clinical stage III cutaneous melanoma.

The results of local-regional advanced melanoma (stage III) management are still not satisfactory. Particularly, there is no personalized treatment in stage III melanoma patients due to the lack of useful classical pathological markers for prognostication of indolent or aggressive course of the disease. The aim of this study was to explore melanoma genomic landscape by means of the mutational profiling of 50 genes influencing carcinogenesis pathways in the randomly selected 93 kinase inhibitor-naïve (KI-naïve) stage III patients. The genomic alterations were found in 27 out of 50 tested genes and at least one pathogenic variant was detected in 77 out of 93 cases (82.7%). Survival rate was negatively affected by the presence of the somatic mutations in AKT1, ATM, CDH1 and SMARCB1, while the BRAF+ status in KI-naïve stage III population correlated with the longer median overall survival. Genomic alterations in WNT pathway correlated with extranodal adipocyte tissue involvement (P = 0.027) and higher number of metastatic lymph nodes (P = 0.045). In terms of survival, the Cox model confirmed the worse prognosis in patients with mutation in the WNT pathway [hazard ratio (HR) = 2.9, P = 0.017], and better prognosis in cases with mutations in BRAF pathway (HR = 0.5, P = 0.004). WNT/ß-catenin pathway alteration was associated with more advanced/aggressive disease. From this perspective, the concept of blocking the activity of the WNT pathway in selected cases appears promising and complementary to the BRAF inhibition therapeutic option for the future.

Rodents as intermediate hosts of cestode parasites of mammalian carnivores and birds of prey in Poland, with the first data on the life-cycle of Mesocestoides melesi.

BACKGROUND: Rodents constitute an important part of the diet of many carnivore species. This predator-prey food chain is exploited by helminth parasites, such as cestodes, whose larval stages develop in rodents and then mature to the adult stage in predators. The main aim of our study was to use molecular techniques for identification of cestode species recovered from both intermediate and definitive hosts, with a particular focus on the genus Mesocestoides. METHODS: Larval cestodes were obtained during our long-term studies on rodent helminth communities in the Mazury Lake District in the north-east Poland in 2000-2018. Cestode larvae/cysts were collected from body cavities or internal organs (e.g. liver) during autopsies. Adult tapeworms were derived from nine red foxes, three Eurasian badgers and one Eurasian lynx. PCR amplification, sequencing and phylogenetic analyses were conducted employing three genetic markers: 18S rDNA, mitochondrial (mt) 12S rDNA and the mt cytochrome c oxydase subunit 1 (cox1) gene fragment. RESULTS: Altogether 19 Mesocestoides samples were analyzed, including 13 adult tapeworms from definitive hosts and six larval samples from 4 bank voles and 2 yellow-necked mice. Phylogenetic analyses revealed three well-supported trees of similar topology. In each case the Mesocestoides samples formed two separate clades. All isolates from foxes, the lynx isolate and two isolates from rodents grouped with Mesocestoides litteratus. Four isolates from rodents and all three isolates from Eurasian badgers were resolved in a separate clade, most similar to North American M. vogae (syn. M. corti). Examination of fixed, stained adult specimens from Eurasian badgers revealed consistency with the morphology of Mesocestoides melesi. Therefore, this clade is likely to represent M. melesi, a species first described in 1985 from the Eurasian badger Meles meles. Molecular analysis allowed also the identification of Taenia crassiceps, Hydatigera kamiyai and Cladotaenia globifera among larvae derived from rodents. CONCLUSIONS: Molecular and phylogenetic analyses support the recognition of M. melesi as a valid species. Our data represent the first record of the larvae of this species in rodents. This is the first report on the occurrence of H. kamiyai in rodents from Poland.