First Molecular Evidence of Babesia vogeli, Babesia vulpes, and Theileria ovis in Dogs from Kyrgyzstan.

Tick-borne parasitic diseases cause mild to severe infections among vertebrate hosts, including dogs. Species in the genus Babesia are important tick-borne pathogens and have worldwide distributions. Although there are data on the prevalence and distribution of Babesia species among dogs around the world, there is no information available in Kyrgyzstan, according to a literature review. In this study, 337 dogs were screened by nested PCR for the presence of the 18S small subunit ribosomal RNA (18S SSU rRNA) gene of piroplasm species. Overall prevalence was 6.23% (21/337) for Babesia/Theileria spp. DNA sequencing of positively tested samples revealed that eighteen samples were infected with Babesia vogeli (B. vogeli) (5.34%), two samples with B. vulpes (0.59%), and one sample with Theileria ovis (T. ovis) (0.29%). The phylogenetic analyses and nucleotide sequences in contrast with those present in GenBank revealed that two nucleotide substitutions (594th and 627th) were found between B. vogeli isolates, including ours, indicating that the mutation is relatively rare. The sequences of other pathogens obtained in this study confirmed 100% nucleotide identity with B. vulpes and T. ovis sequences in GenBank. To the best of our knowledge, B. vogeli, B. vulpes, and T. ovis were detected for the first time in dogs from Kyrgyzstan, and it is thought that results will contribute to the understanding of the epidemiology of canine tick-borne pathogens in the country.

Probiotics and prebiotics alleviate behavioral deficits, inflammatory response, and gut dysbiosis in prenatal VPA-induced rodent model of autism.

Autism spectrum disorders are neuropsychiatric conditions characterized by social interaction and communication disorders and repetitive stereotypical behaviors. These disorders are also accompanied by an inflammatory status. Bidirectional communication between microbiome, gut, and brain has been discovered as a major mechanism influencing core symptoms and biomarkers of autism. Therefore, the modulation of the gut microbiota in autism has recently attracted interest. In this study, probiotic- and prebiotic-mediated modulation of the gut microbiota was compared in terms of different symptoms and findings in an experimental autism model. Valproic acid (VPA) (500 mg/kg) was administered to Wistar rats (on prenatal day 12.5) to induce autistic-like behaviors. Based on the supply of probiotics and prebiotics, animals were grouped as control (saline), autistic-like (prenatal VPA), probiotic (prenatal VPA + 22.5 × 109 cfu/day probiotic), prebiotic (prenatal VPA + 100 mg/day prebiotic), and combined treatment (prenatal VPA + 22.5 × 109 cfu/day probiotic + 100 mg/day prebiotic). After the treatment process, behavioral tests (social behaviors, anxiety, stereotypical behavior, sensorimotor gating, and behavioral despair) and biochemical analyses (serum and brain tissue) were conducted, and the quantities of some phyla and genera were determined in stool samples. Significant positive effects of probiotic and combined treatments were observed on the sociability, social interaction, and anxiety parameters. In addition, all three treatments had positive effects on stereotypical behavior. However, the treatments did not affect sensorimotor gating deficits and behavioral despair. Further, probiotic treatment reversed the VPA-induced increase and decrease in serum IL-6 and IL-10 levels, respectively. Combined treatment also significantly increased the IL-10 levels. Prenatal VPA exposure decreased 5-hydroxytryptamine (5-HT) levels in the prefrontal cortex of the brain; however, combined treatment reversed this decrease. Prenatal VPA exposure also caused a decrease in Bacteroidetes/Firmicutes ratio in the gut microbiota, while the probiotic treatment significantly increased this ratio. These findings indicate that probiotic- and prebiotic-mediated microbial modulation may represent a new therapeutic approach to alleviate autistic-like symptoms.

The detection and phylogenetic analysis of Anaplasma phagocytophilum-like 1, A. ovis and A. capra in sheep: A. capra divides into two genogroups.

In this study, the presence, prevalence, and genotypes of Anaplasma phagocytophilum, A. ovis, and A. capra in sheep were investigated based on 16 S SSU rRNA, groEL, and gtlA gene-specific polymerase chain reaction (PCR), respectively. The sequences of the genes were used for detection of the phylogenetic position of the species. Additionally, a restriction fragment length polymorphism (RFLP) were carried out for discrimination of A. phagocytophilum and related variants (A. phagocytophilum-like 1 and 2). The prevalence of Anaplasma spp. was found as 25.8% (101/391), while it was found that A. ovis, A. phagocytophilum-like 1, and A. capra are circulating in the sheep herds in Kyrgyzstan, according to the PCRs, RFLP and the partial DNA sequencing results. The positivity rates of A. phagocytophilum-like 1, A. ovis, and A. capra genotype-1 were 6.9, 22.5, and 5.3%, respectively. A total of 32 (8.2%) sheep were found to be mix infected. Moreover, phylogenetic analyses and sequence comparison with those available in the GenBank showed that A. capra formed two distinct genetic groups (A. capra genotype-1 and A. capra genotype-2). Considering the zoonotic potential of these species, it may be necessary to make changes in the interpretation of anaplasmosis cases in animals and there is a need for further studies to determine the pathogenicity of the species/genotypes circulating in animals.

First Molecular Detection of Dirofilaria immitis and D. repens in Dogs from Kyrgyzstan.

BACKGROUND: Dirofilaria immitis and Dirofilaria repens are the causative agents of cardiopulmonary and subcutaneous dirofilariosis, respectively. This neglected disease mainly seen in dogs, cats and wild carnivores is re-emerging recent years. No study was conducted on dirofilariosis in dogs in Kyrgyzstan. PURPOSE: The goal of this study was to investigate Dirofilaria species using PCR and sequencing in dogs from Kyrgyzstan. METHOD: Dirofilaria spp. infection in dogs was screened via convential PCR and sequencing in 337 dogs from Kyrgyzstan. RESULT: The overall prevalence of Dirofilaria spp. was 0.59% (2/337): DNA of D. immitis was detected in one sample and DNA of D. repens in second positive sample. In second sample, parallel co-infection of D. repens with Wolbachia was also found. While D. immitis sequence showed 98.70-100% similarity with previously reported sequences of D. immitis from dog blood, D. repens shared 100% identity with other sequences of D. repens. CONCLUSION: These results provided first evidence for Dirofilaria spp. in Kyrgyzstan and emphasized the veterinary and medical importance.

Presence of Toxocara spp. and Other Zoonotic parasites Ova in Children's Playground in Karaman, Turkey

Objective: Human toxocariosis (HT) is a widespread and neglected parasitic disease around the world and it is caused by Toxocara canis and Toxocara cati, a common nematode found in dogs and cats. Childiren are caught to HT after ingestion of embriyonated Toxocara spp. eggs via contaminated materials such as soil, hair and etc. The aim of this study is to investigate Toxocara spp. and other zoonotic parasites in children's playgrounds in Karaman province of Turkey. Methods: In total, 103 samples (68 sand soil, 26 soil and 9 stool) from 20 randomly selected children's playgrounds in May 2018 in Karaman province, were investigated. Samples were examined by flotation in saturated NaCl solution and parasite ova were diagnosed under the light microscope morphologically. Results: Of the 20 screened playgorunds, 11 [55%, confidence interval (CI=33.6-75.2)]and 27 analyzed sample (26.2%, CI=18.4-35.2) were positive one or more parasite species. While Toxocara spp. eggs were the most common species in total (19.4%, CI=12.6-27.8), taeniid (Taenia spp., Echinococcus spp.) eggs and Ancylostoma spp. eggs were found in seven (6.8%, CI=2.97-12.7) and one (0.97%, CI=0.05-4.21) samples respectively. Also, one soil sample was found to be contaminated with both Toxocara and taeniid eggs. Conclusion: These results demonstrate that children's playgrounds in Karaman may be a source for HT and other zoonotic infections. We advise to be fenced children's playgrounds in order to prevent pet animal's accessibility.

Evaluation of Cystic Echinococcosis Cases in Terms of Sociodemographic, Clinical and Hospitalization Features in Karaman Province, Turkey.

BACKGROUND: We aimed to investigate the cystic echinococcosis (CE) epidemiology in Karaman Province from 2010 to 2017 using data from the provincial state hospital. METHODS: Overall, 482 cases were determined from Karaman State Hospital, Karaman Province, Turkey from 2010 to 2017. Records were investigated in terms of year, gender, age, cyst location, rural and urban households and duration of hospital stay. RESULTS: The CE incidence was 22.40 per 100 000 people after final diagnosis with focused abdominal ultrasound. Totally, 482 people including 166 (34.4%) male and 316 (65.6%) female were with CE. The mean age of patients was 50.3±17.48 yr. More CE cases were reported in the age group of 51-60 yr than in the other age groups. CE was mostly seen in liver (470, 97.5%). Moreover, 28.4% of patients were resided in rural areas. The observed frequencies of demographic features of female gender, infected liver, residence in provincial centers, referral to general surgery policlinic and no hospitalization were significantly higher than expected frequencies (P<0.05). Sixty (12.4%) patients with CE had been hospitalized. There was a statistically significant positive relationship between age of hospitalized patients and duration of hospitalization (P<0.05). Of the patients 73.2% were admitted to general surgery. CONCLUSION: CE is of great importance to public health considerations in the Karaman Province and we advocate the implementation of eradication programs to decrease the CE cases number.

Haemoparasitic agents associated with ovine babesiosis: A possible negative interaction between Babesia ovis and Theileria ovis.

Babesiosis, theileriosis, and anaplasmosis are the most common tick-borne diseases in sheep. The majority of anaplasmosis and theileriosis are subclinical; however, babesiosis causes severe infections in small ruminants. Although there are many reports of co-infections with the agents of these diseases, their clinical severity compared with either of the infections alone is unknown. Within the host, interactions between co-infecting species may cause variations in clinical presentation and response to therapy. The aim of this study was to determine the tick-borne agents in sheep located at sites where fatal disease outbreaks caused by babesiosis have commonly been reported. Two hundred and nine sheep with clinical signs suggestive of ovine babesiosis were included in the study. The initial diagnosis of haemoparasites was based on clinical symptoms and microscopy and was confirmed using PCR assays. The blood samples were examined for the presence of Babesia ovis (B. ovis), Anaplasma ovis (A. ovis), A. phagocytophilum, and Theileria ovis (T. ovis). The results showed 86.12% of the animals were infected with one or more pathogens. B. ovis was the dominant pathogen. Overall, the infection rate of B. ovis, A. ovis, T. ovis, and A. phagocytophilum was 70.81%, 56.94%, 21.05%, and 2.39%, respectively. The infection rate of B. ovis alone (31.11%) was higher than A. ovis (9.44%) or T. ovis (1.67%) alone. Co-infections were found at a higher percentage (57.78%) than single infections (42.22%). A. ovis was detected in the blood of a high percentage (98.07%) of co-infected animals. Coexistence of B. ovis and A. ovis (34.45%) was more common than other combinations of species. There was a noticeably low level of co-occurrence between B. ovis and T. ovis (1.11%). During the study, 11 sick animals did not survive despite treatment. Seven were infected with B. ovis alone, three had a dual infection with B. ovis and A. ovis, and one had B. ovis, A. ovis, and T. ovis.

Molecular detection and characterization of Hepatozoon spp. in dogs from the central part of Turkey.

Canine hepatozoonosis is a tick-borne protozoal disease caused by Hepatozoon spp. Two species of Hepatozoon are currently known to infect dogs as Hepatozoon canis and H. americanum. Although H. canis generally causes a chronic infection with relatively mild clinical alterations compared to H. americanum, infection by H. canis can be life-threatening. The disease is widespread in USA, Africa, Europe, South America, and Asia. To determine the frequency of infection with Hepatozoon spp. in stray dogs from Central Anatolia Region of Turkey, a total of 221 blood samples collected over a three-year period were evaluated by using genus specific Polymerase Chain Reaction (PCR) designed to amplify a fragment of 666bp located in 18 S rRNA gene of Hepatozoon spp. Eight (3.61%) blood samples were positive for Hepatozoon spp. For the classification of species, all positive PCR products were purified with a PCR purification kit and sequenced. Sequencing results of eight representative amplicons indicated that 6 were 98-99% identical to the sequence of H. canis and the other 2 sequences were 95-97% identical to the sequence of Hepatozoon spp. So it was named Hepatozoon sp. MF. A phylogenetic tree was constructed from the sequences of the tick-borne agents identified previously and in this study using the neighbor-joining method. The nucleotide sequences were compared to the H. canis sequences reported in Turkey using the nucleotide Basic Local Alignment Search Tool (BLAST) program. The results of this study are significant in terms of the presence of a novel canine Hepatozoon genotype.

Molecular identification of Theileria and Babesia in ticks collected from sheep and goats in the Black Sea region of Turkey.

A molecular survey was undertaken in the Black Sea region of Turkey to determine the presence of Theileria and Babesia species of medical and veterinary importance. The ticks were removed from sheep and goats, pooled according to species and locations, and analyzed by PCR-based reverse line blot (RLB) and sequencing. A total of 2241 ixodid ticks belonging to 5 genus and 12 species were collected and divided into 310 pools. Infection rates were calculated as the maximum likelihood estimation (MLE) with 95% confidence intervals (CI). Of the 310 pools tested, 46 (14.83%) were found to be infected with Theileria or Babesia species, and the overall MLE of the infection rate was calculated as 2.27% (CI 1.67-2.99). The MLE of the infection rates were calculated as 0.691% (CI 0.171-1.78) in Haemaphysalis parva, 1.47% (CI 0.081-6.37) in Rhipicephalus sanguineus, 1.84% (CI 0.101-7.87) in Ixodes ricinus, 2.86% (CI 1.68-4.48) in Rhipicephalus turanicus, 5.57% (CI 0.941-16.3) in Hyalomma marginatum, and 6.2% (CI 4.02-9.02) in Rhipicephalus bursa. Pathogens identified in ticks included Theileria ovis, Babesia ovis, Babesia bigemina, and Babesia microti. Most tick pools were infected with a single pathogen. However, five pools displayed mixed infections with T. ovis and B. ovis. This study provides the first molecular evidence for the presence of B. microti in ticks in Turkey.

Molecular identification of Theileria and Babesia in sheep and goats in the Black Sea Region in Turkey.

This study was carried out to investigate presence and distribution of Theileria and Babesia species via microscopic examination and reverse line blotting (RLB) techniques in sheep and goats in the Black Sea region of Turkey. For this purpose, 1,128 blood samples (869 sheep and 259 goats) were collected by active surveillance from sheep and goats in different provinces of various cities in the region in the years 2010 and 2011. Smears were prepared from the blood samples, stained with Giemsa, and examined under the light microscope for Theileria and Babesia piroplasms. The genomic DNAs were extracted from blood samples. The length of 360-430-bp fragment in the variable V4 region of 18S SSU rRNA gene of Theileria and Babesia species was amplified using the gDNAs. The polymerase chain reaction products were hybridized to the membrane-connected species-specific probes. A total of 38 animals (3.37%) including 34 sheep (3.91%) and 4 goats (1.54%) were found to be positive for Theileria spp. piroplasms in microscopic examination of smears while Babesia spp. piroplasm could not detected. Infection rates were 34.64% in sheep, 10.04% in goats, and totally 28.99% for Theileria ovis while 0.58% in sheep and totally 0.44% for Babesia ovis. However, Theileria sp. OT3 was detected in 2.65% of sheep and 2.04% of all animals; besides Theileria sp., MK had 0.58% prevalence in sheep and 0.77% in goats, with a total 0.62% with RLB. Although T. ovis and Theileria sp. MK were determined in both sheep and goats, B. ovis and Theileria sp. OT3 were observed only in the sheep. These results provide the first detailed molecular data for sheep and goat theileriosis and babesiosis in the region.

[Malaria epidemiology in Bitlis from 1998 to 2008]. / Bitlis Ili'nde 1998-2008 Yillari Arasinda Sitma Epidemiyolojisi.

OBJECTIVE: Malaria is still an important public health problem both in Turkey and the world. In this investigation, the records of patients with malaria that had been detected by the Health Directorship of Bitlis between 1998 and 2008 were examined. METHODS: The retrospective study was performed on data from the Provincial Health Directory. During this 11-year period, a total of 86,951 blood samples were taken by active and pasive surveillance. Thin and thick blood smears stained with Giemsa were examined by immersion objective under microscope. RESULTS: A total of 659 (0.75%) malaria cases were detected. Of these cases, 368 (55.84%) were male and 291 (44.16%) female. It was also observed that the positive cases were found mostly between 1998-2000 and showed an increase between May-September and an important decrease from 2001. Out of the 659 cases of malaria, 599 (90.9%) cases were indigenous, 60 (9.1%) cases were imported and in all cases the determinant was Plasmodium vivax. CONCLUSION: It is hoped that, with this study, the data will contribute to the epidemiology of malaria and its prevention in Bitlis.

[Tick (Ixodoidea) and flea (Siphonaptera) species on three red foxes (Vulpes vulpes) in Erzurum province]. / Erzurum Ilinde Üç Kirmizi Tilkide (Vulpes vulpes) Kene (Ixodoidea) ve Pire (Siphonaptera) Türleri.

In this study, three red foxes (Vulpes vulpes) which died after traffic accidents were examined in terms of ectoparasites, in the province of Erzurum in January of 2009. 13 ticks and 74 fleas were collected from the foxes and taken to the laboratory in separate glasses containing 70% alcohol. Ticks were taken directly from 70% alcohol and have been identified under the stereo microscope. Fleas were cleared in 10% KOH solution during 4-13 days and washed in distilled water 3-4 times and were identified under the light microscope. Two tick species Ixodes hexagonus Leach, 1815 (5 males, 7 females) and Haemaphysalis numidiana Neumann, 1905 (1 male) and four flea species Pulex irritans Linne, 1758 (23 males, 37 females), Chaetopsylla globiceps Tacshenberg, 1880 (11 female), Ctenocephalides canis Curtis, 1826 (2 female) and Ctenocephalides felis felis Bouche, 1835 (1 female) were identified. In this study H. numidiana have been identified in the fox for the first time.

Measurement of gross alpha and gross beta activity concentrations in human tooth.

The gross alpha and gross beta activity concentrations were measured in human tooth taken from 3 to 6 age-groups to 40 and over ones. Accumulated teeth samples are investigated in two groups as under and above 18 years. The gross alpha and beta radioactivity of human tooth samples was measured by using a gas-flow proportional counter (PIC-MPC 9604-alpha/beta counter). In tooth samples, for female age-groups, the obtained results show that the mean gross alpha and gross beta activity concentrations varied between 0.534-0.203 and 0.010-0.453 Bq g(-1) and the same concentrations for male age-groups varied between 0.009-1.168 and 0.071-0.204 Bq g(-1), respectively.

Evaluation of a PCR and comparison with RLB for detection and differentiation of Theileria sp. MK and other Theileria and Babesia species of small ruminants.

Theileria sp. MK in sheep and goats were detected first time by polymerase chain reaction (PCR) and detection limit of PCR and reverse line blotting (RLB) were compared. A part of 18S ssu rRNA gene was amplified from blood samples that were taken from sheep and goats naturally infected with Theileria sp. MK by PCR. Detection limit of both PCR and RLB methods was one infected cell in 10(7) sheep erythrocytes. Nine hundred twenty field samples that had been tested previously by RLB were evaluated by the PCR assay. As found by RLB previously, 12 of 920 (1.30%) samples were detected as positive by PCR. Two positive PCR products, one of which was from sheep and the other from goat, were sequenced. These sequences were identical to the reported nucleotide sequence of Theileria sp. MK. It is concluded that the PCR described in this study will be useful for epidemiological studies and for discrimination between Theileria sp. MK and other Theileria species. In addition, PCR has superiority over RLB because of its ease of use and time period required.

[Use of multiplex PCR for the diagnosis of Theileria annulata and Theileria buffeli]. / Theileria annulata ve Theileria buffeli'nin Teshisinde Multiplex PCR'in Kullanilmasi.

The species causing theileriosis in cattle in Turkey are Theileria annulata and T. buffeli. While T. buffeli is low in pathogenicity or non-pathogenic , T. annulata is very pathogenic and causes tropical theileriosis with high morbidity and mortality in cattle. In this study, a multiplex PCR was used for a simultaneous diagnosis of these species. Genes for the merozoite surface antigen (Tams 1) and the major piroplasm surface protein (MPSP) were amplified with PCR for T. annulata and T. buffeli, respectively. It was found that both single and mixed infection with T. annulata and T. buffeli could be diagnosed with multiplex PCR.