Serological evidence of human exposure to foodborne zoonotic parasites in Romanian patients and possible association with food habits and hygiene.

Humans may become accidental dead-end hosts for a series of zoonotic foodborne parasites, of which Toxoplasma gondii, Echinococcus spp., Toxocara spp., and Trichinella spiralis are of major public health interest, due to their potential pathological implications. The aims of the study were to evaluate the exposure to these pathogens in north-western Romania, and to investigate their potential association to risk factors. From June 2022 to January 2024, 554 patients admitted to a tertiary hospital in north-western Romania were screened for the presence of IgG antibodies against T. gondii, Echinococcus spp., Toxocara canis, and T. spiralis by ELISA, and potential risks were assessed using a questionnaire. Overall, 225 samples (40.6%) were positive for at least one pathogen. The highest seroprevalence for IgG was found for T. gondii (33.9%), followed by Echinococcus spp. (9.1%), T. spiralis (2.9%), and T. canis (1.1%). For T. spiralis, raw meat consumption was associated with positivity. For T. gondii, increased age, rural environment, contact with cats, consumption of unwashed fruits/vegetables and drinking water from unverified sources were significantly associated to seropositivity. The present study provides new insights into the epidemiological status of zoonotic foodborne parasite in Romania, underlining the need to increase awareness on the importance of water, sanitation and food habits in relation with this neglected pathology.

Prevalence and Genotyping of Water- and Food-Borne Parasitic Protozoans (Giardia duodenalis and Cryptosporidium spp.) in Hospitalized Patients from Northwestern Romania.

Giardia duodenalis and Cryptosporidium spp. are important zoonotic protozoan pathogens that infect the gastro-intestinal tract of numerous vertebrates, including humans, and both parasites are responsible for water- or food-borne outbreaks of disease worldwide. Although, globally, both parasites are highly prevalent, particularly in developing countries, epidemiological data from Romania are scarce, and genotyping has rarely been performed. The aims of the present study were to investigate the occurrence and genetic diversity of G. duodenalis and Cryptosporidium spp. in patients hospitalized in Northwestern Romania in relation to clinical and paraclinical presentation and to identify the relative frequency of non-specific symptoms and potential risk factors. Between June 2022 and January 2024, 426 fecal samples were screened for gastro-intestinal parasites by rapid tests and microscopical examination, further confirmed by PCR and sequencing. Giardia duodenalis was detected and characterized in 12 samples (2.82%), while Cryptosporidium parvum was confirmed in four samples (0.94%). A majority of positive patients were symptomatic and reported nausea and vomiting with a significantly higher frequency compared to negative ones. This study provides new insights into the epidemiological status and clinical implications of gastro-intestinal parasite species and genospecies in Romania that are necessary for an in-depth understanding of the potential zoonotic transmission and improvement of patient care.

Toxoplasma gondii in small mammals in Romania: the influence of host, season and sampling location.

BACKGROUND: Toxoplasma gondii is a protozoan parasite that infects a large spectrum of warm-blooded animals, including humans. Small rodents and insectivores play an important role in the epidemiology of T. gondii and may serve as a source of infection for both, domestic and wild definitive felid hosts. Factors influencing the occurrence of T. gondii in wild small mammals are unknown, despite the fact that many intermediate host species are identified. We have used small mammals (Rodentia and Lipotyphla) captured over two years in various habitats, both in urbanised and in natural landscapes. We assessed the importance of land-use, season and host ecology on T. gondii infection. RESULTS: We examined 471 individuals belonging to 20 small mammal species, collected at 63 locations spread over wide altitude, habitat and land-use ranges from Romania. Heart tissue samples were individually analysed by PCR targeting the 529 bp repetitive DNA fragment of T. gondii. The overall prevalence of infection was 7.3%, with nine species of rodents and two species of shrews being found to carry T. gondii DNA. Five species showed high frequency of infection, with the highest prevalence found in Myodes glareolus (35.5%), followed by Spermophilus citellus (33.3%), Sorex minutus (23.1%), S. araneus (21.7%) and Micromys minutus (11.1%). Adults seemed more often infected than young, however when controlling for season, the difference was not significant, as in spring both adults and young showed higher infection rates, but more adults were sampled. Contrary to our expectations, urban/rural areas (with their implicit high density of domestic feline presence) had no effect on infection prevalence. In addition, neither habitat, nor land-use at sampling sites was important as only geographical location and host species were contributing factors to the infection risk. CONCLUSIONS: High prevalence of T. gondii infection showed a highly localised, patchy occurrence, with long living and higher mobility host species being the most common carriers, especially during autumn.

The Role of Wild Boars in the Circulation of Tick-Borne Pathogens: The First Evidence of Rickettsia monacensis Presence.

Most wild mammals can serve as hosts both for tick-borne pathogens (TBPs) and for the ticks themselves. Among these, wild boars, due to their large body size, habitat and life span, show high exposure to ticks and TBPs. These species are now one of the widest-ranging mammals in the world, as well as the most widespread suid. Despite the fact that certain local populations have been decimated by African swine fever (ASF), wild boars are still considered overabundant in most parts of the world, including Europe. Altogether, their long-life expectancy, large home ranges including migration, feeding and social behaviors, wide distribution, overabundance and increased chances of interactions with livestock or humans make them suitable sentinel species for general health threats, such as antimicrobial-resistant microorganisms, pollution and ASF geographical distribution, as well as for the distribution and abundance of hard ticks and also for certain TBPs, such as Anaplasma phagocytophilum. The aim of this study was to evaluate the presence of rickettsial agents in wild boars from two counties in Romania. Among 203 blood samples of wild boars (Sus scrofa ssp. attila) collected during 3 (2019-2022) hunting seasons (September-February), 15 were found positive for tick-borne pathogen DNA. Six wild boars were positive for A. phagocytophilum DNA presence and nine for Rickettsia spp. The identified rickettsial species were R. monacensis (six) and R. helvetica (three). No animal was positive either for Borrelia spp., Ehrlichia spp. or Babesia spp. To the best of our knowledge, this is the first report of R. monacensis in European wild boars, thus adding the third species from the SFG Rickettsia, in the epidemiology of which this wild species may have a role as a reservoir host.

Milk Pathogens in Correlation with Inflammatory, Oxidative and Nitrosative Stress Markers in Goat Subclinical Mastitis.

Goat mastitis is still frequently diagnosed in dairy farms, with serious consequences on milk quality and composition. The aim of this study was to establish correlations between milk microorganisms and biochemical parameters in goats with no signs of clinical mastitis. Thus, 76 milk samples were collected from a dairy goat farm, Carpathian breed, followed by microbiological, molecular (16S rRNA sequencing) and somatic cells analysis, determination of lactate dehydrogenase (LDH), ß-glucuronidase, catalase (CAT), glutathione peroxidase (GPx) activity, total antioxidant capacity (TAC), nitric oxide (NO) and lipid peroxides (LPO) using spectrophotometry and the ELISA method for 8-hydroxy-deoxyguanosine (8-OHdG) as the oxidative DNA damage indicator. Samples positive for bacterial growth showed a significant (p < 0.05) increase in the number of somatic cells, LDH and ß-glucuronidase activity, as well as higher levels of CAT, GPx, NO, LPO and 8-OHdG compared with pathogen-free milk whereas TAC was lower in milk from an infected udder. These findings suggest that subclinical mastitis is associated with increased enzymatic activity and induction of oxidative stress. Nevertheless, changes in biochemical parameters tended to vary depending on the pathogen, the most notable mean values being observed overall in milk positive for Staphylococcus aureus.

Serological Survey of Mosquito-Borne Arboviruses in Wild Birds from Important Migratory Hotspots in Romania.

In the context of climate change, globalization, and enhanced human traveling, arboviruses continue to represent a threat to public health. West Nile and Usutu viruses are mosquito-borne viruses belonging to the Flaviviridae family (Flavivirus genus) and members of the Japanese encephalitis virus serocomplex. Included in the Togaviridae family (Alphavirus genus), the Sindbis virus is also vectored by mosquitoes. In the present study, we aimed to analyze the presence of antibodies concerning the abovementioned viruses in migratory and resident birds in the South-Eastern region of Romania, as avian hosts represent the main reservoir for human infection. Blood samples were collected from wild birds between May 2018 and October 2019 in nine locations from three counties. All the samples were serologically tested by ELISA and a serum neutralization test. Overall, a seroprevalence of 8.72% was registered for the West Nile virus, 2.71% for the Usutu virus, and 0% for the Sindbis virus. To our best knowledge, this is the first large-scale comprehensive study to assess the West Nile virus seropositivity in wild birds and the first serological confirmation of the Usutu virus in wild birds in Romania. Moreover, this is the only follow-up study reviewing the current seroprevalence of the Sindbis virus in Romania since 1975.

Efficacy of Artemisia annua against Coccidiosis in Broiler Chickens: A Field Trial.

(1) Background: Various studies on artemisinin and its derivatives have shown that Artemisia annua may be of therapeutic interest for different diseases, including chicken coccidiosis. This study aimed to evaluate the effects of Artemisia annua on farm-reared broiler chickens by analyzing both the anticoccidial efficacy and its effect on the intestinal microbiota of poultry. (2) Methods: The experiment was performed within three houses on a broiler chicken farm located in Romania. House 1 was the experimental group and received a diet with an addition of A. annua. Houses 2 and 4 were the control groups and received anticoccidials. The prophylactic efficacy of A. annua against coccidiosis was evaluated by recording the weight gain, feed conversion rate, number of oocysts per gram of feces, lesion score, and mortality rate. (3) Results: The chickens fed with A. annua showed a decreasing trend in the number of oocysts per gram of faeces, and their lesion score was 80% lower than in the control group. The weight gains of the chickens treated with A. annua was lower, whilst the feed conversion rate was better than in controls. (4) Conclusions: Artemisia annua showed promising results in the prophylaxis of coccidiosis. Overall, the broiler chickens that received A. annua presented promising zootechnical performances and medical data related to coccidiosis and gut microbiota.

Seroprevalence Rates against West Nile, Usutu, and Tick-Borne Encephalitis Viruses in Blood-Donors from North-Western Romania.

INTRODUCTION: West Nile virus (WNV), Usutu virus (USUV), and the tick-borne encephalitis virus (TBEV) are all arboviruses belonging to Flaviviridae family. All are characterized by vectorial transmission and sometimes associated with neuroinvasive infections. The circulation of these viruses is considered endemic in parts of Europe, with human cases reported in many countries. Among hosts, the viruses are vectored by hematophagous arthropods, such as mosquitoes (WNV, USUV) and ticks (TBEV). Considering the currently outdated knowledge regarding the epidemiology of these viruses in Romania, the aim of our study was to assess the seroprevalence rates of WNV, USUV, and TBEV among healthy blood donors in north-western Romania. METHODS: Human blood samples from healthy donors were collected between November 2019 and February 2020 in six counties from the north-western region of Romania. The samples were serologically tested by ELISA and serum neutralization test. RESULTS: Overall, we obtained a seroprevalence of 3.17% for WNV, 0.08% for TBEV, and 0% for USUV. CONCLUSION: Despite the low seroprevalence of WNV, USUV, and TBEV in our study, we highlight the need for continuous nationwide vector and disease surveillance and implementation of control measures. Further research is required for an optimal overview of the epidemiological status of the Romanian population regarding these flaviviruses together with countrywide awareness campaigns.

Spotted Fever Group Rickettsia spp. Diversity in Ticks and the First Report of Rickettsia hoogstraalii in Romania.

Tickborne bacterial pathogens have been described worldwide as risk factors for both animal and human health. Spotted fevers caused by Rickettsiae may cause non-specific symptoms, which make clinical diagnosis difficult. The aim of the current study was to evaluate and review the diversity of SFG Rickettsiae in ticks collected in 41 counties in Romania. A total of 2028 questing and engorged ticks collected in Romania belonging to five species were tested by PCR amplification of Rickettsia spp. gltA and 17-D gene fragments: Ixodes ricinus (n = 1128), Dermacentor marginatus (n = 507), D. reticulatus (n = 165), Rhipicephalus rossicus (n = 128) and Haemaphysalis punctata (n = 100). Five Rickettsia species were identified following DNA sequence analysis: R. helvetica, R. monacensis, R. slovaca, R. raoultii, and R. hoogstraalii. The most common species detected was R. monacensis. Moreover, R. hoogstraalii was detected for the first time in Romania and in R. rossicus ticks. The detection of R. raoultii and R. monacensis in questing larvae of Hae. punctata suggests the possible transovarial transmission of these Rickettsia species in ticks. The detection of R. hoogstraalii for the first time in Romania increases the reported SFG Rickettsia diversity in the country.

Seroprevalence of antibodies against Borrelia burgdorferi sensu lato in healthy blood donors in Romania: an update.

BACKGROUND: The Borrelia burgdorferi sensu lato (s.l.) genogroup is the causative agent responsible for Lyme borreliosis, a common tick-borne infectious disease in some temperate regions of the Northern Hemisphere. In humans, the clinical manifestations of Lyme borreliosis vary from dermatological infection to severe systemic manifestations. In Romania, data on the seroprevalence of Lyme borreliosis and associated risk factors are scarce and outdated, as the only seroprevalence study with a large dataset was published more than 20 years ago. Therefore, the aim of the present study was to evaluate the seroprevalence for Borrelia burgdorferi s.l. in healthy blood donors from six Romanian counties and identify the associated risk factors. METHODS: The study was conducted among 1200 healthy blood donors aged between 18 and 65 years during November 2019 and September 2020 from six counties in the northwestern and central parts of Romania. A two-tiered testing strategy was applied. Positive and equivocal immunoenzymatic test results for IgG and IgM antibodies were further confirmed by Western blot. RESULTS: Serum samples from 20% of the blood donors had positive or equivocal IgG and IgM ELISA index values. In total, 2.3% of the serum samples for IgG and 1.8% for IgM were positive by Western blot. The seroprevalence for both antibodies varied between 1.5% (Satu-Mare) and 6.5% (Bistrița-Nasaud) in the six counties investigated. The highest seroprevalence was observed in men (4.7%), in blood donors performing their professional activities outdoors (4.2%), and in those aged ≥ 56 years (8%). CONCLUSIONS: These findings confirm the presence of specific IgG and IgM antibodies to B. burgdorferi s.l. among healthy blood donors from Romania. Furthermore, potential risk factors, such as gender, age, and behavior, associated with the presence of positive B. burgdorferi s.l. antibodies among healthy blood donors were identified.

Rickettsia spp. in bats of Romania: high prevalence of Rickettsia monacensis in two insectivorous bat species.

BACKGROUND: Spotted fever group rickettsiae represent one of the most diverse groups of vector-borne bacteria, with several human pathogenic species showing an emerging trend worldwide. Most species are vectored by ticks (Ixodidae), with many zoonotic reservoir species among most terrestrial vertebrate groups. While the reservoir competence of many different vertebrate species is well known (e.g. birds, rodents and dogs), studies on insectivorous bats have been rarely performed despite their high species diversity, ubiquitous urban presence and importance in harboring zoonotic disease agents. Romania has a high diversity and ubiquity of bats. Moreover, seven out of eight SFG rickettsiae species with zoonotic potential were previously reported in Romania. Based on this, the aim of this study was to detect Rickettsia species in tissue samples in bats. METHODS: Here we report a large-scale study (322 bats belonging to 20 species) on the presence of Rickettsia spp. in Romanian bat species. Tissue samples from insectivorous bats were tested for the presence of Rickettsia DNA using PCR detection amplifying a 381 bp fragment of the gltA gene. Positive results were sequenced to confirm the results. The obtained results were statistically analyzed by chi-squared independence test. RESULTS: Positive results were obtained in 14.6% of bat samples. Sequence analysis confirmed the presence of R. monacensis in two bat species (Nyctalus noctula and Pipistrellus pipistrellus) in two locations. CONCLUSION: This study provides the first evidence of a possible involvement of these bat species in the epidemiology of Rickettsia spp., highlighting the importance of bats in natural cycles of these vector-borne pathogens.

Wide Distribution and Diversity of Malaria-Related Haemosporidian Parasites (Polychromophilus spp.) in Bats and Their Ectoparasites in Eastern Europe.

Malaria is responsible for major diseases of humans, while associated haemosporidians are important factors in regulating wildlife populations. Polychromophilus, a haemosporidian parasite of bats, is phylogenetically close to human-pathogenic Plasmodium species, and their study may provide further clues for understanding the evolutionary relationships between vertebrates and malarial parasites. Our aim was to investigate the distribution of Polychromophilus spp. in Eastern Europe and test the importance of host ecology and roost site on haemosporidian parasite infection of bats. We sampled bats and their ectoparasites at eight locations in Romania and Bulgaria. DNA was extracted from blood samples and ectoparasites and tested individually for the presence of DNA of Polychromophilus spp. using a nested PCR targeting a 705 bp fragment of cytB. Two species of Polychromophilus were identified: Po. melanipherus in Miniopterus schreibersii and associated ectoparasites and Po. murinus in rhinolophid and vespertilionid bats (6 species) and their ticks and nycteribiid flies. Only cave-dwelling bat species (and their ectoparasites) showed infections, and we found a strong correlation between infections with Polychromophilus parasites and Nycteribiidae prevalence. We report the high genetic diversity of Polychromophilus spp. in Eastern Europe, suggesting that the simultaneous presence of varied host and vector assemblages enhances bat haemosporidian parasite diversity.

Experimental infection with Toxoplasma gondii in broiler chickens (Gallus domesticus): seroconversion, tissue cyst distribution, and prophylaxis.

Toxoplasma gondii is a widespread zoonotic protozoan that infects most species of mammals and birds, including poultry. This study aimed to investigate the course of T. gondii infection and the efficacy of diclazuril and Artemisia annua in preventing infection in experimentally infected chickens. Seventy-five 1-month-old chickens, female and male, were randomly divided into five groups (n = 15 each) as follows: (1) uninfected untreated (negative control, NC); (2) infected with T. gondii genotype II/III isolated from a wild cat (group WC); (3) infected with T. gondii genotype II isolated from a domestic cat (group DC); (4) infected with T. gondii domestic cat strain and treated with the anticoccidial diclazuril (group DC-D); and (5) infected with T. gondii domestic cat strain and treated with the medicinal plant Artemisia annua (group DC-A). Clinical signs, body temperature, mortality rate, weight gain, feed conversion ratio, hematological parameters, and the presence of T. gondii-specific IgY antibodies were recorded in all groups. Five chickens per group were euthanized 28 days post-infection (p.i.) and their brains, hearts, and breast muscle tested for T. gondii by mouse bioassay and polymerase chain reaction (PCR). No clinical signs related to the experimental infection were observed throughout the study period. T. gondii-specific antibodies were detected by day 28 p.i., but not in all infected chickens. Overall, T. gondii DNA was detected (bioassay or tissue digests) in all infected and untreated chickens (10/10), while viable parasite (bioassay) was isolated from 7 out of 10 chickens. The parasite was most frequently identified in the brain (7/10). There were no differences in the T. gondii strains regarding clinical infection and the rate of T. gondii detection in tissues. However, higher antibody titers were obtained in chickens infected with T. gondii WC strain (1:192) comparing with T. gondii DC strain (1:48). A. annua reduced replication of the parasite in 3 out of 5 chickens, while diclazuril did not. In conclusion, broiler chickens were resistant to clinical toxoplasmosis, irrespective of the strain (domestic or wild cat strain). The herb A. annua presented prophylactic efficacy by reduced parasite replication. However, further studies are required aiming at the efficacy of diclazuril and A. annua for the prevention of T. gondii infection in chickens using quantitative analysis methods.

First multilocus sequence typing (MLST) of Giardia duodenalis isolates from humans in Romania.

BACKGROUND: Giardia duodenalis is one of the most prevalent and highly diverse human parasites, encompassing a complex of eight genetically distinct assemblages, each further divided into sub-assemblages. While in recent years, G. duodenalis genotype distribution patterns in humans have been intensely studied, there is still very little information available on the diversity of Giardia genotypes and sub-assemblages infecting people in Romania. In the present study, we investigated the genetic diversity of Giardia duodenalis in asymptomatic patients from Romania. METHODS: Over an 11-month period, human feces from 7805 healthy adults were screened by microscopic analysis for G. duodenalis cysts during their obligatory periodic check-ups. DNA extraction was performed from microscopic-positive fecal samples, followed by multilocus sequence typing of four genetic loci of the ITS region, gdh, tpi and bg genes, followed by DNA sequencing and phylogenetic analysis. Statistical analysis was performed using EpiInfo 2000 software. RESULTS: The prevalence of giardiasis in the present study was 0.42% (33/7805). Twenty-three samples (76.67%) were successfully genotyped at each locus. The bg and tpi genes had the highest typing success rate (100%). The identified assemblages were assemblage A in 27 cases (subtypes A2 and A3), and B in 3 cases. CONCLUSIONS: To our knowledge, the present study is the first report of multilocus sequence typing of G. duodenalis isolated from humans in Romania. The present results may shed light on G. duodenalis infection in humans at a regional and national level, thus increasing awareness against this parasitic infection.

Multiple Tick-Borne Pathogens in Ixodes ricinus Ticks Collected from Humans in Romania.

Ticks are medically important vectors of infectious diseases that are able to transmit pathogens to humans and animals. Tick-borne diseases represent a major health concern, posing an increasing risk to the public health during the last century and affecting millions of people. The aim of the current study was to provide epidemiological data regarding the presence of certain tick-borne pathogens in ticks feeding on humans in Romania. Overall, 522 Ixodes ricinus ticks collected from humans were screened for six pathogens: Borrelia spp., Neoehrlichia mikurensis, Babesia spp., Coxiella spp., Bartonella spp., and Francisella tularensis. Ticks attached to humans were collected between 2013-2015 in Cluj County, Romania. Conventional, nested and quantitative PCR were used to detect specific genetic sequences of each pathogen. For identifying the infectious agents, positive samples were sequenced. The infection prevalence was 21.07% from which 8.18% were mixed infections. The detected agents were Borrelia spp., N. mikurensis and Babesia spp. The present data reveal the endemic occurrence of potentially zoonotic pathogens in Romania. Revealing the current distribution of tick-borne pathogens in ticks collected from humans may provide new insights in understanding the complex ecology of tick-borne diseases and enlightens current knowledge about the infection prevalence at local, regional and national levels.

Molecular Survey of Metastrongyloid Lungworms in Domestic Cats (Felis silvestris catus) from Romania: A Retrospective Study (2008-2011).

: Background: Lungworms are recognized as important agents in the pathology of the respiratory system in domestic cats. While Aelurostrongylus abstrusus is worldwide known and studied, Troglostrongylus brevior has gained the attention of the scientific community only in the last decade. The pathogenicity of this species seems to be higher than A. abstrusus, causing more severe clinical presentations and being potentially fatal, especially in young animals. METHODS: In this study, 371 DNA isolates of faecal samples were tested by multiplex polymerase chain reaction for the presence of A. abstrusus, T. brevior, and Angiostrongylus chabaudi. RESULTS: The results showed that 30.2% and 6.7% of the investigated domestic cats were positive for A. abstrusus and T. brevior respectively, stressing out the importance of these parasites as agents of respiratory conditions in domestic cats from Romania. None of the samples were positive for A. chabaudi. The age, the outdoor access, and the lack of deworming were identified as significant risk factors for infection with A. abstrusus. CONCLUSIONS: This paper represents the first report of T. brevior in domestic cats from Romania. Moreover, it presents the most northern localization in Europe of T. brevior.

Borrelia spp. in small mammals in Romania.

BACKGROUND: Small mammals play an important role in the life-cycle of ticks and are reservoirs for several zoonotic pathogens. The aim of this study was to provide epidemiological data regarding the presence of Borrelia spp. in tissues of small mammals from Romania. METHODS: We examined 401 individuals belonging to 11 small mammal species collected in Romania. Collections cover the largest effort to survey these reservoirs in the country. Tissue samples were analyzed by multiplex qPCR targeting the ospA gene of Borrelia burgdorferi (s.l.) and a part of the flaB gene of B. miyamotoi. Positive samples were further analysed by conventional PCR and sequenced. RESULTS: The overall prevalence of infection with Borrelia spp. in small mammal tissues was 4.9%. The most commonly detected species were B. afzelii, followed by B. garinii/B. bavariensis, B. miyamotoi and B. burgdorferi (s.s.). To our knowledge, we report for the first time the detection of Borrelia spp. in Crocidura leucodon and C. suaveolens, and B. miyamotoi in the liver of Myodes glareolus. CONCLUSIONS: To our knowledge, our study evaluates for the first time the occurrence of Borrelia spp. in small mammals in Romania, contributing to a better knowledge of the distribution of these bacteria. This survey upgrades previous data on the spatial distribution of the pathogens and reveals the importance of animal surveillance regarding Lyme borreliosis and relapsing fever caused by B. miyamotoi.

Toxoplasma gondii in water buffaloes (Bubalus bubalis) from Romania: what is the importance for public health?

The purpose of our study was to evaluate the prevalence of Toxoplasma gondii infection in autochthonous Carpathian buffaloes from northwestern Romania by serology, PCR techniques, and mouse bioassay. Agreement between MAT and ELISA, correlation between indirect and direct detection methods, and risk factors were evaluated. The apparent overall seroprevalence of T. gondii was 8.1% by MAT and 6.6% by ELISA. The agreement between ELISA and MAT was fair. The apparent seroprevalence was significantly higher in adult buffaloes (12.5%) compared to calves (0.0%) and juveniles (1.9%) by MAT. Most of the positive adult buffaloes detected by MAT had antibodies at a low sera dilution and the highest dilution was 1:768 in a juvenile female (30 months). No viable T. gondii was detected by mouse bioassay, as no T. gondii cyst or DNA was found in the brain of mice and they did not seroconvert. However, T. gondii DNA was detected in two buffaloes: in a 30-month-old male buffalo by qPCR on the diaphragm digest and in a 252-month-old female buffalo by RE nPCR on the mesenteric lymph node. Both animals were negative in MAT and ELISA. The total prevalence of T. gondii by direct detection methods was 2.7%. There was no correlation between indirect and direct detection methods. Since no viable T. gondii was detected in buffaloes, the risk of human infection from buffalo meat is minimal. Buffaloes' biological response to a T. gondii infection appears to be very similar to the response of cattle.