Identifying Borrelia burgdorferi as the causative agent of Lyme disease in 1981 was a watershed moment in understanding the major impact that tick-borne zoonoses can have on public health worldwide, particularly in Europe and the USA. The medical importance of tick-borne diseases has long since been acknowledged, yet little is known regarding the occurrence of emerging tick-borne pathogens such as Borrelia spp., Anaplasma phagocytophilum, Rickettsia spp., Bartonella spp., “Candidatus Neoehrlichia mikurensis”, and tick-borne encephalitis virus in questing ticks in Romania, a gateway into Europe. The objective of our study was to identify the infection and co-infection rates of different Borrelia genospecies along with other tick-borne pathogens in questing ticks collected from three geographically distinct areas in eastern Romania. We collected 557 questing adult and nymph ticks of three different species (534 Ixodes ricinus, 19 Haemaphysalis punctata, and 4 Dermacentor reticulatus) from three areas in Romania. We analyzed ticks individually for the presence of eight different Borrelia genospecies with high-throughput real-time PCR. Ticks with Borrelia were then tested for possible co-infections with A. phagocytophilum, Rickettsia spp., Bartonella spp., “Candidatus Neoehrlichia mikurensis”, and tick-borne encephalitis virus. Borrelia spp. was detected in I. ricinus ticks from all sampling areas, with global prevalence rates of 25.8%. All eight Borrelia genospecies were detected in I. ricinus ticks: Borrelia garinii (14.8%), B. afzelii (8.8%), B. valaisiana (5.1%), B. lusitaniae (4.9%), B. miyamotoi (0.9%), B. burgdorferi s.s (0.4%), and B. bissettii (0.2%). Regarding pathogen co-infection 64.5% of infected I. ricinus were positive for more than one pathogen. Associations between different Borrelia genospecies were detected in 9.7% of ticks, and 6.9% of I. ricinus ticks tested positive for co-infection of Borrelia spp. with other tick-borne pathogens. The most common association was between B. garinii and B. afzelii (4.3%), followed by B. garinii and B. lusitaniae (3.0%). The most frequent dual co-infections were between Borrelia spp. and Rickettsia spp., (1.3%), and between Borrelia spp. and “Candidatus Neoehrlichia mikurensis” (1.3%). The diversity of tick-borne pathogens detected in this study and the frequency of co-infections should influence all infection risk evaluations following a tick bite.

In industrialized countries, Hepatitis E is a recognized zoonosis, with wild boar and swine representing the main reservoirs for zoonotic genotype HEV-3 in Europe. Data related to HEV infection in wild boar population in Romania are restricted to serological surveys. Therefore, our main goal was to determine the HEV prevalence in wild boar population and to characterize HEV strains circulating in Romania. Using TaqMan real-time RT-PCR assay, we analyzed the presence of RNA HEV in 45 liver samples and five spleen samples collected from 50 wild boars. Samples were collected during the 2013-2015 hunting seasons. Nine samples of 50 were tested positive for HEV RNA, resulting an overall prevalence of 18%. Phylogenetic analysis revealed that the isolates clustered in different HEV-3 monophyletic groups, depending on the sampling county. This is the first study signalling, based on molecular analysis, the presence of HEV in wild boar population from Romania. Also, in this study, we report the detection of HEV in splenic tissue from wild boar.

A serosurvey for Tahyna virus (TAHV), a mosquito-borne California encephalitis orthobunyavirus (Peribunyaviridae) endemic to Europe, was performed to estimate the activity of TAHV on a broad geographic scale. Sera from wild boar (Sus scrofa), roe deer (Capreolus capreolus) and red deer (Cervus elaphus) were collected from Austria, Hungary and Romania. Samples were tested for neutralizing antibodies against TAHV using a virus microneutralization assay. The results demonstrate that TAHV transmission to mammals is widespread in Europe, particularly in the wild boar population where the mean rate of seroconversion is 15.2%.

The prevalence of the strains of extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae is continuously increasing at the global level. The appearance of ESBL enzymes represents a danger for the efficacy of treatments with beta-lactam antibiotics (Măciucă I., 2015). The aim of the study resided in assessing the prevalence of ESBL-positive strains of E. coli and K. pneumoniae in pets that were treated with antibiotics (Enrofloxacin, Ciprofloxacin, Cefadroxil) for various bacterial infectious diseases. In February 2015, 29 faeces samples were collected at the rectal level from dogs and cats. The samples were collected with the help of sterile buffers. For the screening of the strains of (ESBL)-producing Enterobacteriaceae, the Oxoid Brilliance chromogenic ESBL Agar medium was used, a specific medium for the isolation of (ESBL)-producing Enterobacteriaceae because it contains cefpodoxim, a second-generation cephalosporin to which all the extended-spectrum beta-lactamase (ESBL)-producing strains are resistant. The phenotypic confirmation of the isolated ESBL strains was achieved by using the combined disc method (Clinical and Laboratory Standards Institute, 2014). The taxonomic classification of the strains that were isolated was achieved by testing some minimal biochemical characteristics with the help of the MIU, TSI, EMBA, TBX tests. The E. coli ATCC 25922 and K.pneumoniae ATCC 700603 strains were used as a reference for quality control for the antibiotic sensitivity test.The results have been interpreted according to the CLSI 2014 standard. As a result of sample processing, we noticed a prevalence of 62.06% in the individuals who were carriers of E. coli and K. Pneumoniae ESBL-positive strains.

Hepatitis E virus (HEV) is the causal agent of the hepatitis E transmitted primarily via the faecal-oral route. HEV belongs to the family Hepeviridae, with HEV strains isolated from human and swine classified into the Orthohepevirus genus Orthohepevirus A species. The disease is considered as an emerging zoonosis with worldwide distribution based on recent advances showing that HEV strains circulating in domestic and wild pigs are genetically related to strains identified in autochthonous human cases.The aim of the present study was to determine the seroprevalence of HEV in wild boar from Galaţi and Buzău counties, as a preliminary stage of the evaluation of the HEV distribution in wild boar population completed by molecular identification and characterization Serum samples were collected from 68 wild boars during the hunting season, between December 2014 and February 2015, in 33 hunting funds from two Romanian Counties: Galați and Buzău. For serological analysis all samples were tested using a commercially available HEV antibody assay: ID Screen® Hepatitis E Indirect Multispecies ELISA kit (IDVet Diagnostics, France).HEV antibodies have been detected in 7 out of 68 serums, representing an overall prevalence of 10.29%. Seropositive animals were identified in both counties, respectively 3 out of 30 (10%) for Buzău County and 4 out of 38 (10.52%) in Galați County. The prevalence rates determined in this study are comparable to those determined in other European Countries.Our results provided premises to investigate the hepatitis E virus presence in Romanian wild boar as well as in other wild animals, which are considered as potential HEV reservoirs.