Molecular characterization of cattle and sheep isolates of Echinococcus granulosus from Elazig province in Turkey and expression analysis of the non-coding RNAs, egr-miR-7, egr-miR-71 and egr-miR-96.

Cystic Echinococcosis (CE) is a common zoonotic disease seen in human and animals worldwide, caused by the larval form of Echinococcus granulosus. In this study, E. granulosus s.l. species and haplotypes were determined in hydatid cysts isolated from cattle and sheep, and the expression levels of egr-miR-7, egr-miR-71 and egr-miR-96 miRNAs were compared in different cyst structures. A total of 82 (cattle, n = 41; sheep, n = 41) hydatid cyst isolates (germinal membranes and/or protoscoleces) were collected from a slaughterhouse in Elazig province of Turkey. After mt-CO1 gene sequences were made, 81 out of 82 hydatid cyst isolates were determined as E. granulosus s.s. (G1 and G3), while an isolate of cattle origin was determined as Echinococcus canadensis (G6/7). A total of 26 nucleotide polymorphisms and 29 haplotype groups were identified in the samples. miRNA expressions in germinal membranes of sterile cysts and germinal membrane and protoscoleces of fertile cysts were investigated by qRT-PCR and Real Time PCR analyses. It was determined that miRNAs were expressed at high levels in 79.31% of the 29 haplotype groups and at low levels in the remaining 10.34%. In 10 fertile samples of sheep origin, egr-miR-7, egr-miR-71 and egr-miR-96 miRNAs were found to be 44, 168, and 351-fold higher in expression, respectively, in the germinal membrane compared to the protoscoleces. Especially egr-miR-96 may have the potential to be used as biomarkers in the diagnosis of active CE.

Identification of antigen B (AgB) Gene polymorphism in cattle and sheep isolates of Echinococcus granulosus and investigation of effects on serological diagnosis.

Cystic Echinococcosis (CE) is a worldwide common helminth disease caused by the larval form of Echinococcus granulosus. The aim of this study is to determine the genetic differences between distinct isolates of E. granulosus obtained from cattle and sheep and determine the polymorphism of the AgB1 gene by DNA sequence analysis, as well as investigating its relationship with serological response using ELISA and Western Blot tests. For this aim, germinal membranes of hydatid cysts of 30 cattle and 30 sheep from the provinces of Elazig and Erzincan in Turkey and serum samples of these animals were collected. Following isolation of the total genomic DNA, the 12S rRNA gene of all isolates was amplified by PCR for genetic characterization, and the mt-CO1 gene region was examined by DNA sequence analysis. The gDNAs were then amplified by PCR using AgB1-specific primers, and genetic variation was investigated by DNA sequence analysis. At the final stage, all serum samples were analyzed by ELISA and Western Blot tests using a partially purified hydatid cyst fluid antigen. As a result, 114 (95%) of the 120 isolates were determined to be E. granulosus sensu stricto by using 12S rRNA-PCR. Subsequently, the DNA sequence analysis of the remaining 6 samples of the mt-CO1 gene revealed that all samples were E. granulosus sensu stricto. According to the DNA sequence analysis that followed, nucleotide changes in the AgB1 gene were observed in 13 (10.8%) of 120 samples. With this study, 9 (69.2%) out of 13 hydatid cysts in which polymorphism was detected by DNA sequence analysis in their AgB1 gene were found to be positive with ELISA, and 6 (46.1%) were found positive by WB. While 80 (74.7%) of 107 non-polymorphic samples in the AgB1 gene were found to be positive with ELISA, and 75 (70.9%) were positive with WB. As a result, variation in different ratios was determined in the AgB1 gene of E. granulosus sensu stricto, and it was determined that this had a partial effect on serological response.

Molecular characterization and detection of variants of Taenia multiceps in sheep in Turkey.

Taenia multiceps is a cestode (family Taeniidae) that in its adult stage lives in the small intestine of dogs and other canids. The metacestode, known as Coenurus cerebralis, is usually found in the central nervous system including brain and spinal card in sheep and other ruminants. The presence of cysts typically leads to neurological symptoms that in the majority of cases result in the death of the animal. Coenurosis could cause high losses in sheep farms because the disease commonly affects young animals. A total of 20 C. cerebralis isolates collected from naturally infected sheep in Mardin province of Turkey were characterized through the polymerase chain reaction and sequencing of a fragment of cytochrome c oxidase subunit 1 (CO1) gene. The results showed that the CO1 gene sequences were highly conserved in C. cerebralis isolates. Phylogenetic analysis based on partial CO1 gene sequences revealed that C. cerebralis isolates were composed of three different variants.

Epidemiological survey and molecular characterization of Echinococcus granulosus in cattle in an endemic area of eastern Turkey.

Cystic echinococcosis (CE) is a zoonotic disease affecting mainly various species of livestock and humans. A survey of cystic echinococcosis in cattle was conducted from December 2008 to April 2009 in an endemic area of eastern Turkey. A total of 1758 cattle were examined and hydatid cysts were found in 33.9% of the cattle (595/1758). Most of the cattle (75.8%) had hydatid cysts only in the lungs, 10.9% only in the liver, 12.4% in the both liver and lungs, 0.7% in the spleen and 0.2% in the heart. 220 of these cysts were examined by PCR of 12S rRNA gene and sequencing of mt-CO1 gene. 147 of 220 cattle isolates showed the same band pattern with 12S rRNA analyses and were identified as G1-G3 complex (Echinococcus granulosus sensu stricto) and also 28 of these were confirmed by mitochondrial CO1 sequencing as G1 genotype. The other 73 samples that did not amplified with 12S rRNA gene specific primers were analyzed for mt-CO1 gene and only 7 samples yielded 446bp product and after the CO1 sequencing these were identified as G3 genotype. This study confirms the predominance of the sheep strain (G1 genotype) in Turkey and this is the most comprehensive genetic survey of cattle CE in Turkey.

Molecular genetic characterization of different isolates of Echinococcus granulosus in east and southeast regions of Turkey.

We used PCR-RFLP analysis of ribosomal ITS1 fragment using four different restriction enzymes and DNA sequencing of mitochondrial CO1 gene to investigate the genetic characteristics of isolates of Echinococcus granulosus obtained from different hosts (179 sheep, 19 cattle, 7 goat, 1 camel, 1 dog and 1 human) and regions (Elazig, Malatya, Erzurum, Van, Diyarbakir and Sanliurfa) of Turkey. The report represents the most comprehensive genotypic investigation of E. granulosus isolates undertaken in Turkey, with Turkish samples of cattle, goat, camel and dog origin being characterized for the first time. We show that the predominant genotype involved in E. granulosus transmission in Turkey is the common sheep strain (G1 genotype) infecting humans, cattle, sheep, goats, camels as well as the dog definitive host. Nevertheless, we urge that coordinators of local control programs in Turkey should take into consideration the potential occurrence and risk of additional strains of E. granulosus infecting humans and animal hosts, and plan accordingly.

Seroprevalence of hypodermosis in cattle in some provinces of Turkey.

The aim of this study was to determine the seroprevalence of hypodermosis in cattle in the east and southeast of Turkey. For this purpose, a total of 634 sera samples of cattle were collected from Malatya, Elazig and Diyarbakir provinces of east and southeast of Turkey from November 2005 to February 2006. The sera were analyzed using a Hypodermin C antigen by means of indirect ELISA. One hundred and forty eight (23.3%) out of 634 cattle were seropositive for hypoderma antibodies. The highest percentage of seropositivity were detected at Elazig province (26.3%) followed by Malatya (22.3%) and Diyarbakir provinces (22.1%). The seropositivity rate was higher in female (31%) than male (14.1%). When the mean is considered by animal breed, the highest seropositivity was detected at local breed (27.7%) following crossbreed (26.8%) and purebreed (19.7%). There was a positive relation between age and seropositivity. Seropositivity rate was 15.9% in 2 and under ages while these rates were 38.1% and 30.4% in 3-4 ages and 5 and up ages, respectively.

[Seroprevalence of Toxoplasma gondii in dogs in the province of Kocaeli]. / Kocaeli yöresi köpeklerinde Toxoplasma gondii seroprevalansi

The aim of this study was to investigate the seroprevalence of Toxoplasma gondii in pet dogs and stray dogs in Kocaeli province using the Sabin-Feldman Dye Test. A total of 116 dogs were examined and 81 (69.8%) were found to be positive. Seropositivity was found to be 59.6% in male dogs and 78.1% in female dogs (p<0.05). Seropositivity was 62.5% in pet dogs and 71.7% in stray dogs (p>0.05). The seropositivity was found to be 68.2%, 75.6% and 62.5% in the 0-1, 1-3, and over 3 age groups, respectively. As a result, the high seropositivity of dog toxoplasmosis in Kocaeli province has been found to be important.

[Molecular genetic characterization of genus Echinococcus.]. / Echinococcus cinsinin moleküler genetik karakterizasyonu.

Morphological, biological and epidemiological features of the adult and metacestode stage of the genus Echinococcus have been the most frequently used criteria for the characterization of species. But strain characterization is more complex and is based on the morphological, biological, biochemical, epidemiological and molecular features. Although morphological and biological studies have provided extremely useful information for strain characterization, that these features may be variable must be taken into consideration. They may be influenced by host and environmental factors and may not reflect distinctness at a genetic level. Although, conventional techniques may be affected by the environment or host, molecular techniques allow a direct characterization of the genome of the parasite and they are not affected by environmental factors. In this review, various data concerning molecular typing methods of the genus Echinococcus have been summarized.

[Seropositivity of hydatidosis in cattle with eosinophilia.]. / Eozinofilik sigirlarda hidatidosis seropozitifligi.

The aim of this study was to investigate the seropositivity of hydatidosis in a group of cattle with and without eosinophilia. Of the 597 cattle, 79 (13.2%) were found to have eosinophilia with absolute eosinophilic counts. Out of 79 cattle, 62 (78.4%) and 48 (60.7%) were found to be positive for hydatidosis by ELISA and IFAT, respectively. Out of 79 cattle without eosinophilia, 38 (48.1%) were found to be seropositive by ELISA and 34 (43.1%) by IFAT. Our data indicated that seropositivity of hydatidosis in cattle with eosinophilia was higher than the group without eosinophilia and it will be useful to investigate livestock with eosinophilia for hydatidosis.

[Determination of Hypoderma bovis and Przhevalskiana silenus somatic antigens using SDS-PAGE.]. / Hypoderma bovis ve Przhevalskiana silenus somatik antijenlerinin SDS-PAGE ile tanimlanmasi

The aim of this study was to examine larval antigens obtained from the third instar larvae of Hypoderma bovis and Przhevalskiana silenus using the sodium dodecyl-sulphate polyacrylamide gel electrophoresis (SDS-PAGE) method. SDS-PAGE separation was performed using 12% separation gel and 5% stacking gel. At the end of the SDS-PAGE examination, 11 and 19 different polypeptide bands were detected between 6-66 kDa and 6-100 kDa in the third instar larvae antigen of H. bovis and P. silenus, respectively.

[Survey of Neospora caninum in cattle in some provinces in the Eastern Anatolian region.]. / Dogu Anadolu bölgesinin bazi illerinde bulunan sigirlarda Neospora caninum'un arastirilmasi

This study was carried out in order to investigate the presence of Neospora caninum in cattle in the Elazig, Malatya, Mus and Bingöl provinces from January 2003- May 2004. Blood samples were collected from 513 cattle and 32 aborting cows that were of various breeds and ages in all provinces. Sera were obtained from these animals and antibodies against N. caninum were investigated using the enzyme-linked immunosorbent assay (ELISA). A commercially available competitive ELISA (cELISA) kit (VMRD, Inc., Veterinary Medical Research and Development, Pullman, WA, USA. Product Code 5NO5.20) was used to detect the N. caninum antibodies in the sera. Out of 513 cattle in the region, 36 (7.01 %) were found to be seropositive by cELISA. Seropositivity rates obtained by cELISA were 15.00% in Elazig, 4.00% in Malatya, 4.86% in Mus and 4.69% in Bingöl. One of the 32 aborting cows (3.12 %) was found to be seropositive against N. caninum.