Phylogenetic Characterization of Orthohantavirus dobravaense (Dobrava Virus).

We report complete coding sequences of Orthohantavirus dobravaense (Dobrava virus) Igneada strains and phylogenetic characterization of all available complete coding sequences. Our analyses suggested separation of host-dependent lineages, followed by geographic clustering. Surveillance of orthohantaviruses using complete genomes would be useful for assessing public health threats from Dobrava virus.

Borrelia miyamotoi in wild rodents from four different regions of Turkey.

Borrelia miyamotoi is a tick-borne zoonotic agent that causes hard tick-borne relapsing fever, an emerging disease in humans. Some small mammalian and bird species are reported to be reservoirs of B. miyamotoi. This study aims to examine Borrelia species present in rodents captured from rural areas of Turkey. Blood samples of rodents were initially screened with Borrelia 16S rRNA qPCR. The Borrelia flaB gene was subsequently amplified by conventional PCR, after which all positive samples were sequenced. Borrelia miyamotoi was observed in nine out of 536 blood samples (1.7%) collected from wild rodents. Phylogenetic analysis showed that all positive samples belonged to the European genotype clade of B. miyamotoi. PCR positivity was 5.3%, 3.7%, and 1.8% in Apodemus uralensis, Apodemus flavicollis, and Myodes glareolus, respectively. Borrelia burgdorferi sensu lato that causes Lyme borreliosis in humans could not be detected in the rodents. In this study, presence of B. miyamotoi DNA is reported for the first time in rodents in Turkey.

Partial External Biliary Diversion for Severe Diarrhea After Liver Transplant in Patients with Progressive Familial Intrahepatic Cholestasis Type 1.

Progressive familial intrahepatic cholestasis is a heterogeneous group of autosomal recessive disorders, and liver transplant is the only curative treatment. A biliary diversion operation for disruption of enterohepatic circulation in patients with progressive familial intrahepatic cholestasis type 1 without cirrhosis is another option. We present a pediatric patient with progressive familial intrahepatic cholestasis type 1 who underwent liver transplant due to end-stage liver disease. After transplant, diarrhea and growth retardation complications resolved after partial external biliary diversion surgery.

First molecular detection and identification of Leishmania species in small wild rodents from Turkey.

Leishmaniasis is a parasitic disease infecting animals and humans. Two clinical forms (Visceral and cutaneous leishmaniasis) and four species are reported to be present in Turkey. Several studies have investigated canine and human leishmaniasis in Turkey but no study was performed to screen the infection among wild rodents, so far. The present study aims to investigate the role of small wild rodents as reservoir animals for Leishmania spp. in different regions of Turkey. Formalin-preserved tissue samples (spleen, liver, lung) of 712 rodents from 30 provinces were screened for the presence of Leishmania spp. DNA. Before DNA extraction, tissues were dried, rehydrated, and homogenated. Leishmania screening in rodent tissues and species determination was performed with a combination of real-time kDNA and ITS1 polymerase chain reaction protocols. Eight (1.12%) out of 712 animals were found to be positive for Leishmania spp. DNA and species typing revealed five L. infantum, two L. tropica and one L. major among positives. Leishmania major and L. infantum DNA were detected in Apodemus spp. from Zonguldak province located in the Western Black Sea Region, while L. tropica DNA was found in Meriones sp. and Gerbillus dasyurus from Adana and Hatay provinces located in Eastern Mediterranean Region of Turkey. The present study is first to report natural infection of L. infantum, L. major and L. tropica in small wild rodents in Turkey, suggesting their possible roles as reservoirs. Further studies are needed for planning epidemiological studies and also for developing rodent control measures in risky endemic areas to break the transmission cycle.

Altitudinal Effects on Innate Immune Response of a Subterranean Rodent.

Immune defense is costly to maintain and deploy, and the optimal investment into immune defense depends on risk of infection. Altitude is a natural environmental factor that is predicted to affect parasite abundance, with lower parasite abundance predicted at higher altitudes due to stronger environmental stressors, which reduce parasite transmission. Using high and low altitude populations of the Turkish blind mole-rat (TBMR) Nannospalax xanthodon, we tested for effects of altitude on constitutive innate immune defense. Field studies were performed with 32 wild animals in 2017 and 2018 from two low- and one high-altitude localities in the Central Taurus Mountains, at respective altitudes of 1010 m, 1115 m, and 2900 m above sea level. We first compared innate standing immune defense as measured by the bacteria-killing ability of blood serum. We then measured corticosterone stress hormone levels, as stressful conditions may affect immune response. Finally, we compared prevalence and intensity of gastrointestinal parasites of field-captured TBMR. We found that the bacteria-killing ability of serum is greater in the mole-rat samples from high altitude. There was no significant difference in stress (corticosterone) levels between altitude categories. Coccidian prevalence and abundance were significantly higher in 2017 than 2018 samples, but there was no significant difference in prevalence, abundance, or intensity between altitudes, or between sexes. Small sample sizes may have reduced power to detect true differences; nevertheless, this study provides support that greater standing innate immunity in high altitude animals may reflect greater investment into constitutive defense.

Characterization of Bartonella taylorii Strains in Small Mammals of the Turkish Thrace.

Rodents play role as a reservoir for some Bartonella species which cause different clinical manifestations in humans. Bartonella spp. existence in rodents of Turkish Thrace has been detected for the first time, and the risky habitat types were evaluated for the infection. Ninety individuals belonging to three small rodent species were screened by PCR, and the overall prevalence of Bartonella infection was 22.2%. The strains were characterized molecularly based on the phylogenetic analyses of two housekeeping genes, rpoB and gltA. They clustered with B. taylorii. The significant effects of habitat types and rodent species on Bartonella infections were observed. It was detected that B. taylorii prevalence was the highest in the swamp forest habitat and A. flavicollis species. The present study demonstrates that A. flavicollis is the reservoir of B. taylorii in the European part of Turkey.

Molecular Survey of Babesia microti (Aconoidasida: Piroplasmida) in Wild Rodents in Turkey.

Babesia microti (Aconoidasida: Piroplasmida) (Franca, 1910) is an important tick-borne zoonotic parasite with rodents serving as reservoir hosts. In the present study, 536 rodents were captured from Burdur, Bartin, Giresun, and Yozgat provinces of Turkey between the years 2010 and 2012, and blood samples were examined for the presence of Babesia spp. using conventional PCR which targeted the 18S rRNA gene. The sequence analysis of PCR amplicons was tested for B. microti as well as for Hepatozoon spp., and Sarcocystis spp. Overall, 5.8% of the rodents were positive for B. microti: 41% in Myodes glareolus, 7.7% in Chionomys roberti, and 2% in Apodemus spp., whereas no Babesia DNA was detected in Mus macedonicus and Microtus spp. Six rodents were positive for Hepatozoon spp. and one rodent was positive for Sarcocystis spp. Overall, 14.9 and 4.5% of rodents captured from Bartin and Giresun provinces, respectively, were PCR positive for B. microti, whereas none of rodents captured in Burdur and Yozgat were positive for Babesia spp. The sequence data of B. microti from rodents revealed that all sequences belonged to the zoonotic genotype. Sequences of B. microti obtained from rodents of the Bartin province were genotypically closer to European isolates, whereas those obtained from rodents of the Giresun province were closer to Russian and Mongolian isolates.

Portal Hypertensive Biliopathy as a Cause of Severe Cholestasis in Children With Congenital Hepatic Fibrosis.

Portal hypertensive biliopathy may occur in patients with noncirrhotic hepatic fibrosis. Portal hypertensive biliopathy treatment should be focused on management of portal hypertension and relief of biliary obstruction. In patients with noncirrhotic portal fibrosis and symptomatic portal hypertensive biliopathy, portal decompression surgery by proximal splenorenal shunt is one successful treatment option.

A novel genetic lineage of Tula orthohantavirus in Altai voles (Microtus obscurus) from Turkey.

Orthohantaviruses (family Hantaviridae order Bunyavirales) are emerging pathogens with a significant impact on human health. They are transmitted via aerosolized excreta of rodents which also act as reservoir hosts, constituting a unique route for dispersion. Dobrava-Belgrade and Puumala orthohantaviruses have been previously reported from Anatolia, in rodents, case reports and occasional outbreaks. We have collected rodents at several locations during a surveillance study in eastern Anatolia. The specimens were morphologically-identified and various tissues were screened via a generic orthohantavirus reverse transcription polymerase chain reaction assay. DNA barcoding via mitochondrial cytochrome b sequencing was performed in rodents with detectable orthohantavirus sequences. High throughput sequencing was performed for viral genome characterization. Fifty rodents were collected and identified morphologically as Microtus spp. (96%) and Apodemus spp. (4%). Orthohantavirus sequences were detected in lung and spleen or liver tissues of 4 voles (8%), barcoded as Microtus obscurus. The virus sequences were identified as Tula orthohantavirus (TULV) and near-complete genomic segments of the prototype viral genome, tentatively named as the Tula orthohantavirus-Turkey (TULV-T), could be characterized. Putative open reading frames for viral nucleocapsid and a nonstructural protein on the S segment, glycoproteins G1 and G2 on the M segment and viral replicase on the L segment were identified on the TULV-T. Several minor sequence variants were further characterized. No evidence of recombination could be detected and pairwise comparisons displayed over 95% amino acid sequence identities to various Eurasian TULV strains. Phylogenetic analyses revealed distinct clustering of all genome segments from previously-characterized TULV strains via various approaches and models. Here, TULV-T constituted a novel lineage, forming an intermediate among Asian and European TULV lineages. This report describes the initial documentation of TULV circulation and its potential reservoir in Anatolia. The extent of virus dispersion, alternate hosts or outcomes of human exposure require elucidation.

Dobrava hantavirus variants found in Apodemus flavicollis mice in Kirklareli Province, Turkey.

Hantaviruses infect humans via inhalation of viral particles within secretions of infected rodents or rarely through direct contact with infected rodents. Determining the prevalence of hantavirus infections among rodent populations is of vital importance to obtain information on hantavirus-related cases and to predict possible outbreaks. We hypothesized that DOBV strains circulating in the Thrace Region in Turkey would be related to other Balkan DOBV strains. In this study, hantavirus infections in the rodent population of the Kirklareli-Igneada Region (north-western Turkey, near the Bulgarian border) were investigated. This region is of particular importance, as it is located in the south-eastern margin of the European continent and was used as an entrance point of Asian faunal elements into Europe. DOBV infection was detected in eight of 73 rodents; all were of the Apodemus flavicollis species. Partial sequences of the viral S-, M-, and L-genome segments were recovered and compared with previously reported DOBV sequences. The newly characterized Turkish strains were similar to other DOBV variants. Silent nucleotide mutations were dominant. The hantavirus prevalence in the Igneada region was similar to what has been reported in Greece and Bulgaria. For the first time, the M-segment sequences of DOBV from Turkey were recovered and genetic data of hantaviruses from Thrace region of Turkey were obtained.

[Optimization of ELISA and immunoblot methods for the detection of IgG antibodies against old world hantaviruses in wild rodents]. / Yabani kemiricilerde Eski Dünya hantavirus IgG antikorlarinin saptanmasi için ELISA ve immünoblot yöntemlerinin optimizasyonu.

Hantaviruses infect humans via inhalation of viral particles in infected rodents' secretions such as saliva, urine and faeces or via direct contact with infected rodents. The rodent species that are known as the carriers of Dobrava (DOBV), Puumala (PUUV), Saaremaa (SAAV), Tula (TULV) and Seoul (SEOV) viruses are found in our country. The presence of specific antibodies against hantaviruses have been demonstrated in rodents collected from Black Sea and Aegean Regions of Turkey in 2004 for the first time. The first hantavirus-related hemorrhagic fever with renal syndrome (HFRS) cases were reported in Black Sea region in 2009. The determination of the hantavirus prevalence in wild life and rodent populations in the field is crucial for the information about hantavirus-related cases and to clarify the state of risk. There is no commercial product optimized for the screening of rodent serum samples in terms of HFRS agents like DOBV and PUUV that are widely seen in Eurasia as well as Turkey. In this study, the antigens belonging to the commercial enzyme-linked immunoassay (ELISA) and immunoblot tests that are produced for the screening of human sera were used for the development of antibody screening tests against hantavirus in rodent sera and were optimized. The most appropriate serum and conjugate dilutions were determined for the optimization of ELISA (Anti-Hantavirus Pool ELISA; Euroimmun, Germany) and immunoblot (Euroline Anti-Hanta Profile 1 strips; Euroimmun, Germany) methods. Optimized ELISA method was used for the screening and optimized immunoblot method was used for the confirmation. A total of 84 wild rodent sera that belonged to Apodemus and Microtus species were evaluated with this procedure and the cut-off value, sensitivity and specificity of optimized ELISA method were determined. For the optimization of ELISA 1/50, 1/100 and 1/200 serum dilutions and 1/10.000, 1/20.000 and 1/40.000 conjugate dilutions were tested. For the optimization of immunoblot, 1/50 and 1/100 serum dilutions and 1/5.000 and 1/10.000 conjugate dilutions were tested. The horseradish peroxidase conjugated goat anti-mouse IgG for ELISA and the alkaline phosphatase conjugated goat anti-mouse IgG for immunoblot were used. We followed the manufacturer's recommendations for the incubation parameters, substrate and the number of washes. 1/50 serum dilution and 1/10.000 conjugate dilution for ELISA and 1/100 serum dilution and 1/5.000 conjugate dilution for immunoblot were determined as optimal concentrations. By using the optimized ELISA, 26.2% (22/84) of rodents were found positive for hantavirus antibodies according the determined cut-off value (OD(450/620): 0.325). By using immunoblot as a confirmatory test, 20 out of 22 ELISA positive samples could be studied because of the insufficient amount of sera and 17 of them was found positive in terms of DOBV antibodies. Of these rodents 11 were Apodemus flavicollis, three were Apodemus agrarius, two were Microtus guentheri and one was Apodemus sylvaticus. When the results of ELISA were compared to immunoblot results, the optimized ELISA's sensitivity and specificity were found as 100% and 95%, respectively. In this study, a method that can be used in the screening of rodent sera was constituted which uses commercial antigens that can be provided easily, gives fast and reliable results. Similar serological methods optimized for different types of rodents are of great importance for the realization of active follow-up and monitoring of the studies in the field.

Dobrava-Belgrade virus in Apodemus flavicollis and A. uralensis mice, Turkey.

In 2009, human Dobrava-Belgrade virus (DOBV) infections were reported on the Black Sea coast of Turkey. Serologic and molecular studies of potential rodent reservoirs demonstrated DOBV infections in Apodemus flavicollis and A. uralensis mice. Phylogenetic analysis of DOBV strains showed their similarity to A. flavicollis mice-borne DOBV in Greece, Slovenia, and Slovakia.

Chromosome differentiation of four 2n = 50 chromosomal forms of Turkish mole rat, Nannospalax nehringi.

Nannospalax is a genus of blind rodents adapted to living in underground. The species have numerous chromosomal forms in Turkey, and their taxonomic position is still unknown. In this study, 15 mole rats of four different 2n = 50 forms were used; C- and G- banding processes were applied; and a comparison was made accordingly. Karyological results showed that the 2n = 50S form is a new form for Turkish blind mole rats. 2n = 50S form is determined from Andirin (Kahramanmaras) and has NF = 70. The 2n = 50W form, on the other hand, differs from the others with NF = 74 form. C-banding results showed that heterochromatin blocks of all 2n = 50 are different, while only the 2n = 50W form has telomeric heterochromatin blocks. G-banding results, however, displayed homologies and differences among the chromosomal forms. After comparison, we determined that Robertsonian fusion is an efficient force on chromosomal evolution in blind mole rats in Turkey, and that telomeric heterochromatin is a distinctive character for the 2n = 50W form. We suggest that the chromosomal changing mechanism should be independent from climatic peculiarities. These results support the theory that ancestral karyotype should have the largest distribution in a chromosomally variable species.