Development of a new PCR protocol for the detection of species and genotypes (strains) of Echinococcus in formalin-fixed, paraffin-embedded tissues

The aim of our study was to establish a new PCR protocol for the detection and discrimination of Echinococcus granulosus complex on one hand and Echinococcus multilocularis in formalin-fixed, paraffin-embedded tissues (FFPTs) on the other. The target sequences for all PCRs are located on a 471bp segment of the mitochondrial ND1 gene, the fragment sizes of the amplification products are 295bp (for the sheep strain of E. granulosus), 204bp (for the pig strain of E. granulosus) and 252bp (for E. multilocularis), respectively. In total, 80 FFPTs from patients with histologically confirmed echinococcosis (76 with E. granulosus and four with E. multilocularis) operated on in Austrian hospitals between 1978 and 2005 were examined. In 68 (85%) samples, we were able to detect specific DNA fragments with our newly established PCR protocols. Thirty-eight (47.5%) of 80 clinical samples were identified as the G1 strain, 26 (32.5%) as the G5, 6 or 7 strains and four (5%) as E. multilocularis. The specificity of all three PCRs was 100%; for the discrimination between G6 and G7 strains, sequencing of an additional 234bp PCR fragment was necessary and showed that three out of 26 G5, 6 or 7 PCR-positive patients were infected with E. granulosus genotype G6 (the camel strain).     

Echinococcus granulosus genotypes circulating in alpacas (Lama pacos) and pigs (Sus scrofa) from an endemic region in Peru

The identification of the genotypes of Echinococcus granulosus present in livestock and wild animals within regions endemic for cystic echinococcosis (CE) is epidemiologically important. Individual strains display different biological characteristics that contribute to outbreaks of CE and that must be taken into account in the design of intervention programs. In this study, samples of hydatid cysts due to E. granulosus were collected from alpacas (4) in Puno and pigs (8) in Ayacucho in Peru, an endemic region for CE. Polymerase chain reaction amplification and DNA sequencing of specific regions of the mitochondrial cytochrome C oxidase subunit 1 and NADH dehydrogenase subunit 1 genes confirmed the presence of a strain common to sheep, the G1 genotype, in alpacas. Two different strains of E. granulosus were identified in pigs: the G1 and the G7 genotypes. This is the first report of the G1 genotype of E. granulosus in alpacas in endemic regions of CE in Peru.     

Echinococcus granulosus strain differentiation in Iran based on sequence heterogeneity in the mitochondrial 12S rRNA gene

Parasite strain characterization is essential for the establishment of a prevention and control strategy in any endemic area. The aim of this study was to characterize different Echinococcus granulosus isolates from Iran by using DNA sequences of the mitochondrial 12S rRNA gene. Thirty livers and lungs of cattle, sheep and goats naturally infected with E. granulosus were collected from abattoirs in northern and western Iran between June and October 2007. These samples yielded 18 fertile cysts which we used for the genetic work. We designed and tested two new primer pairs which specifically amplify portions of the mitochondrial 12S rRNA gene of the two strains (G1 and G6) of E. granulosus known to occur in Iran. One primer pair amplified a fragment of 259 base pairs (bp) from only the G1 strain. The second pair amplified a fragment of 676 bp from the G6 strain. The G1 genotype was identified in all fertile cyst samples, in agreement with previous studies in Iran. Ten of our samples and a single reference sample of the G6 strain were sequenced and compared with the G1 and G6 sequences deposited in GenBank.     

Helminthologic survey of the wolf (Canis lupus) in Estonia, with an emphasis on Echinococcus granulosus

Carcasses of 26 wolves were collected during the 2000/2001 and 2003/2004 hunting seasons and examined for helminths. Thirteen helminth species were recorded: one trematode (Alaria alata), seven cestodes (Diphyllobothrium latum, Mesocestoides lineatus, Taenia hydatigena, Taenia multiceps, Taenia ovis, Taenia pisiformis, and Echinococcus granulosus), and five nematode species (Uncinaria stenocephala, Toxascaris leonina, Toxocara canis, Trichinella nativa, and Trichinella britovi). The most common species were A. alata and U. stenocephala. Mature Echinococcus granulosus was found and described for the first time in Estonia, and its identity verified using PCR-RFLP analysis. Sequencing a fragment of the mitochondrial DNA NADH dehydrogenase 1 (mtND1) gene showed that the E. granulosus strain from Estonia was identical to strain G10, recently characterized in reindeer and moose in Finland.     

Molecular characterization of Echinococcus granulosus from Peru by sequencing of the mitochondrial cytochrome C oxidase subunit 1 gene

Echinococcus granulosus, the etiologic agent of cystic echinococcosis (CE) in humans and other animal species, is distributed worldwide. Ten intra-specific variants, or genotypes (G1-G10), have been defined based on genetic diversity. To determine the genotypes present in endemic areas of Peru, samples were collected from cattle (44), sheep (41) and humans (14) from Junín, Puno Huancavelica, Cusco, Arequipa and Ayacucho. DNA was extracted from protoscolex and/or germinal layers derived from 99 E. granulosus isolates and used as templates to amplify the mitochondrial cytochrome C oxidase subunit 1 gene. The resulting polymerase chain reaction products were sequenced and further examined by sequence analysis. All isolates, independent of the host, exhibited the G1 genotype. Phylogenetic analysis showed that three isolates from Ayacucho shared the same cluster with microvariant G1(4). The G1 genotype is considered the most widespread and infectious form of E. granulosus worldwide and our results confirm that the same patterns apply to this country. Therefore, these findings should be taken into consideration in developing prevention strategies and control programs for CE in Peru.     

Echinococcus granulosus sensu lato genotypes infecting humans – review of current knowledge

Genetic variability in the species group Echinococcus granulosus sensu lato is well recognised as affecting intermediate host susceptibility and other biological features of the parasites. Molecular methods have allowed discrimination of different genotypes (G1–10 and the ‘lion strain’), some of which are now considered separate species. An accumulation of genotypic analyses undertaken on parasite isolates from human cases of cystic echinococcosis provides the basis upon which an assessment is made here of the relative contribution of the different genotypes to human disease. The allocation of samples to G-numbers becomes increasingly difficult, because much more variability than previously recognised exists in the genotypic clusters G1–3 (=E. granulosus sensu stricto) and G6–10 (Echinococcus canadensis). To accommodate the heterogeneous criteria used for genotyping in the literature, we restrict ourselves to differentiate between E. granulosus sensu stricto (G1–3), Echinococcus equinus (G4), Echinococcus ortleppi (G5) and E. canadensis (G6–7, G8, G10). The genotype G1 is responsible for the great majority of human cystic echinococcosis worldwide (88.44%), has the most cosmopolitan distribution and is often associated with transmission via sheep as intermediate hosts. The closely related genotypes G6 and G7 cause a significant number of human infections (11.07%). The genotype G6 was found to be responsible for 7.34% of infections worldwide. This strain is known from Africa and Asia, where it is transmitted mainly by camels (and goats), and South America, where it appears to be mainly transmitted by goats. The G7 genotype has been responsible for 3.73% of human cases of cystic echinococcosis in eastern European countries, where the parasite is transmitted by pigs. Some of the samples (11) could not be identified with a single specific genotype belonging to E. canadensis (G6/10). Rare cases of human cystic echinococcosis have been identified as having been caused by the G5, G8 and G10 genotypes. No cases of human infection with G4 have been described. Biological differences between the species and genotypes have potential to affect the transmission dynamics of the parasite, requiring modification of methods used in disease control initiatives. Recent investigations have revealed that the protective vaccine antigen (EG95), developed for the G1 genotype, is immunologically different in the G6 genotype. Further research will be required to determine whether the current EG95 vaccine would be effective against the G6 or G7 genotypes, or whether it will be necessary, and possible, to develop genotype-specific vaccines.     

Molecular evidence for the presence of a G7 genotype of Echinococcus granulosus in Slovakia

Variability in Echinococcus granulosus is very important epidemiologically since strain characteristics may influence local patterns of transmission of hydatid disease. To classify the genotype presented in pig protoscoleces of the Slovak territory, a DNA-based approach has been used. Nucleotide sequences for a 471 bp region of the mitochondrial NADH dehydrogenase 1 (ND1) gene revealed a substantial affinity of isolates examined to the G7 genotype. Only a 0.9-3.4% sequence variation was recorded for E. granulosus samples compared with the reference G7 variant. To distinguish between G7 and G9 genotypes not differing in ND1 sequences, isolates were additionally examined by PCR-RFLP analysis of the nuclear ITS1 region. The resulting two-banded pattern is characteristic for the G7 strain. The data presented thus provides the first explicit evidence of the G7 genotype in the Slovak region.     

G6PD通过细胞周期蛋白D1/D2调控人黑色素瘤细胞周期的进程

葡萄糖-6-磷酸脱氢酶(G6PD)在人皮肤黑色素瘤A375细胞中处于高表达与高活性状态, 但G6PD在黑色素瘤发生发展过程中的作用及其具体机制尚不明确.本文在前期运用 siRNA方法构建G6PD敲减的黑色素瘤A375稳转细胞(A375-G6PDΔ)基础上,构建表达载体pBabe-puro-G6PDWT在A375-G6PDΔ细胞中过表达野生型的G6PD基因,从而构建G6PD表达恢复的稳转细胞(A375-G6PDΔ-G6PDWT).3株细胞A375-WT、A375-G6PDΔ和 A375-G6PDΔ-G6PDWT经G6PD酶活性测定、MTT测定、克隆形成实验、流式细胞仪分析细胞周期和Western 印迹检测.结果显示,A375-G6PDΔ-G6PDWT细胞的G6PD蛋白表达量 (0.847 ± 0.080)及其活性(0.394 ± 0.029)分别是A375-G6PDΔ的3.28倍(P＜0.01) 和7.34倍(P＜0.01),分别是A375-WT细胞的91-57%和2.12倍(P＜0.05).与A375-WT细 胞相比,A375-G6PDΔ细胞G0/G1期细胞数增加,S期细胞数减少,增殖指数PI降低了25-70%（P＜0.05）,细胞周期蛋白D1/D2、细胞周期蛋白E表达分别下降37.4%、54.3% (P＜0.01)和17.3%;而A375-G6PDΔ-G6PDWT细胞呈现G1/S期阻滞解除,细胞周期蛋白D1/D2蛋白分别恢复到A375-WT细胞的89.5%和87.6%,细胞周期蛋白E表达未见 恢复,呈现生长增殖和克隆形成率的恢复并接近于A375-WT细胞. 结果提示,G6PD通 过细胞周期蛋白D1/D2调控人皮肤黑色素瘤A375细胞G1期向S期转换的进程,这为黑色 素瘤发病机制的研究提供了新的思路.     

Intraspecific variability among NADH dehydrogenase subunit 1 sequences of Taenia hydatigena.

This paper describes intraspecific variability of the partial sequences of the mitochondrial ND1 gene among isolates of Taenia hydatigena from pigs in Poland, Ukraine and Wales. The differences between studied isolates ranged from 0.4 to 5.5%, which exceeds the variability within the same fragment between the different genetic variants of Echinococcus multilocularis and is comparable with the variability between the most closely related strains (G5/G6/G7) of E. granulosus. The biggest difference (5.5%) was found between the geographically most distant Ukrainian and Welsh samples of T. hydatigena while the samples collected from the neighbouring locations in Poland, were most similar to each other.     

Involvement of individual subsites and secondary substrate binding sites in multiple attack on amylose by barley alpha-amylase

Barley alpha-amylase 1 (AMY1) hydrolyzed amylose with a degree of multiple attack (DMA) of 1.9; that is, on average, 2.9 glycoside bonds are cleaved per productive enzyme-substrate encounter. Six AMY1 mutants, spanning the substrate binding cleft from subsites -6 to +4, and a fusion protein, AMY1-SBD, of AMY1 and the starch binding domain (SBD) of Aspergillus niger glucoamylase were also analyzed. DMA of the subsite -6 mutant Y105A and AMY1-SBD increased to 3.3 and 3.0, respectively. M53E, M298S, and T212W at subsites -2, +1/+2, and +4, respectively, and the double mutant Y105A/T212W had decreased DMA of 1.0-1.4. C95A (subsite -5) had a DMA similar to that of wild type. Maltoheptaose (G7) was always the major initial oligosaccharide product. Wild-type and the subsite mutants released G6 at 27-40%, G8 at 60-70%, G9 at 39-48%, and G10 at 33-44% of the G7 rate, whereas AMY1-SBD more efficiently produced G8, G9, and G10 at rates similar to, 66%, and 60% of G7, respectively. In contrast, the shorter products appeared with large individual differences: G1, 0-15%; G2, 8-43%; G3, 0-22%; and G4, 0-11% of the G7 rate. G5 was always a minor product. Multiple attack thus involves both longer translocation of substrate in the binding cleft upon the initial cleavage to produce G6-G10, essentially independent of subsite mutations, and short-distance moves resulting in individually very different rates of release of G1-G4. Accordingly, the degree of multiple attack as well as the profile of products can be manipulated by structural changes in the active site or by introduction of extra substrate binding sites.     

Genetic characterization of Echinococcus granulosus in camels, cattle and sheep from the south-east of Iran indicates the presence of the G3 genotype

Echinococcus granulosus, the aetiologic agent of cystic echinococcosis (CE), is one of the most important zoonotic helminthes worldwide. Isolates of the parasite show considerable genetic variation in different intermediate hosts. Several genotypes and species are described in different eco-epidemiological settings. This study investigated E. granulosus genotypes existing in livestock and humans from the province of Kerman, located in south-eastern Iran, using sequencing data of cox1 and nad1 mitochondrial genes. Fifty-eight E. granulosus isolates, including 35 from sheep, 11 from cattle, 9 from camels and 3 from goats, were collected from slaughterhouses throughout Kerman. One human isolate was obtained from a surgical case of CE. Mitochondrial cox1 and nad1 regions were amplified using polymerase chain reaction (PCR) and 38 isolates were sequenced. Genotypes G1 (73.7%), G3 (13.2%) and G6 (13.1%) were identified from the isolates. G1 was the most common genotype from sheep (86.7%), cattle (80%), camels (44.4%) and goats (100%). Sheep, cattle and camels were also found to be infected with the G3 genotype (buffalo strain). The human isolate was identified as the G6 genotype. Results showed that the G3 genotype occurred in different animal hosts in addition to G1 and G6 genotypes.     

Genetic Characterization of Human-Derived Hydatid Cysts of Echinococcus granulosus Sensu Lato in Heilongjiang Province and the First Report of G7 Genotype of E. canadensis in Humans in China

Cystic echinococcosis (CE) caused by the larval stage of Echinococcus granulosus sensu lato (s.l.) is one of the most important zoonotic parasitic diseases worldwide and 10 genotypes (G1–G10) have been reported. In China, almost all the epidemiological and genotyping studies of E. granulosus s.l. are from the west and northwest pasturing areas. However, in Heilongjiang Province of northeastern China, no molecular information is available on E. granulosus s.l. To understand and to speculate on possible transmission patterns of E. granulosus s.l., we molecularly identified and genotyped 10 hydatid cysts from hepatic CE patients in Heilongjiang Province based on mitochondrial cytochrome c oxidase subunit I (cox1), cytochrome b (cytb) and NADH dehydrogenase subunit 1 (nad1) genes. Two genotypes were identified, G1 genotype (n = 6) and G7 genotype (n = 4). All the six G1 genotype isolates were identical to each other at the cox1 locus; three and two different sequences were obtained at the cytb and nad1 loci, respectively, with two cytb gene sequences not being described previously. G7 genotype isolates were identical to each other at the cox1, cytb and nad1 loci; however, the cytb gene sequence was not described previously. This is the first report of G7 genotype in humans in China. Three new cytb gene sequences from G1 and G7 genotypes might reflect endemic genetic characterizations. Pigs might be the main intermediate hosts of G7 genotype in our investigated area by homology analysis. The results will aid in making more effective control strategies for the prevention of transmission of E. granulosus s.l.     

Description of Echinococcus granulosus genotypes in human hydatidosis in a region of southern Chile

INTRODUCTION: Echinococcus granulosus species has a wide variety in both geography and hosts; indeed, 10 genotypes have been reported in studies on material of animal origin. The aim of this study was to genotype E. granulosus obtained from human hydatid cysts. MATERIALS AND METHODS: The hydatid fluid and sand was collected from patients who underwent surgery for hepatic and pulmonary hydatidosis at Hospital Regional in Temuco, Chile, between 2004 and 2005. Two PCR systems were used: PCR Eg 9 and PCR Eg 16. The RsaI enzyme was used for RFLP. The genotype was confirmed using the sequence of one fragment of 366 bp from a mitochondrial gene (cox1). RESULTS: The DNA of protoscolices from 24 samples was analyzed, 4 of them from pulmonary cysts and 20 from hepatic cysts. The 366 bp fragment was amplified in 20 out of 24 samples (83.3%). Enzymatic digestion revealed the presence of 3 possible genotypes: in 20 out of 21 samples (95,2%), a restriction was observed corresponding to the G1 or G7 genotypes; in the remaining sample genotype G4 or G7 was observed. Sequencing confirmed the presence of G1 genotype for 19 samples and G6 genotype for the remaining sample (G4 or G7 according to PCR-RFLP). CONCLUSION: The PCR-RFLP technique enabled three possible genotypes present (G1 or G7, G4 or G7) to be established. Sequencing allowed us to decisively identify the G1 and G6 genotypes in our study group. Previous studies agree with the identification of the G1 genotype in our country. We consider it significant that the G6 genotype is present in Chile for its epidemiological implications.     

Using mitochondrial and nuclear markers to evaluate the degree of genetic cohesion among Echinococcus populations

Based on the distinctiveness of their mitochondrial haplotypes and other biological features, several recent publications have proposed that some Echinococcus granulosus strains should be regarded as separate species. However, the genetic cohesion of these species has not been extensively evaluated using nuclear markers. We assess the degree of polymorphism of the partial mitochondrial cox1 (366 bp), the nuclear mdh (214 bp) and EgAgB4 (281-283 bp) genes of E. granulosus sensu lato isolates collected from areas where different strains occur sympatrically. Five distinct mitochondrial haplotypes were determined by direct sequencing (G1, G2, G5, G6 and G7). The mdh genotypes were first screened by SSCP: three alleles were identified (Md1-Md3), which were further confirmed by nucleotide sequencing. For EgAgB4, which was analysed by direct sequencing the PCR products, two groups of sequences were found: EgAgB4-1 and EgAgB4-2. No haplotype-specific mdh or EgAgB4 sequences occur. Nevertheless, alleles Md1 and Md2 and type 1 sequences of EgAgB4 showed a higher frequency within the group of haplotypes G1-G2, while allele Md3 and EgAgB4-2 are most frequent in the G5-G7 cluster. By AMOVA it is shown that 79% of the total genetic variability is found among haplotype groups. These findings are compatible with two not mutually exclusive evolutionary hypotheses: (a) that haplotypes share an ancestral polymorphism, or (b) that the reproductive isolation between parasites with distinct haplotypes is not complete, leading to gene introgression. The biologic and epidemiologic consequences of our findings are discussed.     

Echinococcus multilocularis: developmental stage-specific expression of Antigen B 8-kDa-subunits

Antigen B (AgB) initially found in hydatid cyst fluid of Echinococcus granulosus is a polymeric lipoprotein of 160 kDa, and is an aggregate of several different but homologous small proteins with approximately 8 kDa. Four genes encoding these 8-kDa-subunits have been identified from E. granulosus. In this study we isolated five genes encoding 8-kDa-subunits of AgB from Echinococcus multilocularis. Sequence comparison of isolated cDNA clones demonstrated that one of these five clones was completely identical to EmAgB8/1 which had been isolated previously by our group, and three of them were 94.5, 90.8, and 91.9% homologous to E. granulosus antigen B 8-kDa subunit genes, EgAgB8/2, EgAgB8/3, and EgAgB8/4, respectively. The remaining clone shared 51-58% homology with the nucleotide sequences of AgB genes. Gene-specific RT-PCR and Western blot analyses revealed that these genes were expressed in a developmentally regulated manner in E. multilocularis vesicles, protoscoleces, and immature adult worms. Possible functions of different expression manners are also discussed.     

Functional analysis of the terminase large subunit, G2P, of Bacillus subtilis bacteriophage SPP1

The terminase of bacteriophage SPP1, constituted by a large (G2P) and a small (G1P) subunit, is essential for the initiation of DNA packaging. A hexa-histidine G2P (H6-G2P), which is functional in vivo, possesses endonuclease, ATPase, and double-stranded DNA binding activities. H6-G2P introduces a cut with preference at the 5'-RCGG downward arrowCW-3' sequence. Distamycin A, which is a minor groove binder that mimics the architectural structure generated by G1P at pac, enhances the specific cut at both bona fide 5'-CTATTGCGG downward arrowC-3' sequences within pacC of SPP1 and SF6 phages. H6-G2P hydrolyzes rATP or dATP to the corresponding rADP or dADP and P(i). H6-G2P interacts with two discrete G1P domains (I and II). Full-length G1P and G1PDeltaN62 (lacking domain I) stimulate 3.5- and 1.9-fold, respectively, the ATPase activity of H6-G2P. The results presented suggest that a DNA structure, artificially promoted by distamycin A or facilitated by the assembly of G1P at pacL and/or pacR, stimulates H6-G2P cleavage at both target sites within pacC. In the presence of two G1P decamers per H6-G2P monomer, the H6-G2P endonuclease is repressed, and the ATPase activity stimulated. Based on these results, we propose a model that can account for the role of terminase in headful packaging.     

Genotypic characterization of Iranian camel (Camelus dromedarius) isolates of Echinoccocus granulosus

In the present study, Echinoccocus granulosus isolates collected from camels (Camelus dromedarius) in eastern Iran were characterized based on the nucleotide sequences of mitochondrial CO1 and NDI genes. The molecular results for camel isolates demonstrated that at least 2 different genotypes are present, i.e., a buffalo genotype (G3) and the camel genotype (G6). Although the sequences of the Iranian camel genotype (G6) are completely homologous to the reference sequence of G6 (M84666) of E. granulosus , a nucleotide mutation (C to T at position 168) was detected in the CO1 sequences of the Iranian G3 isolates (HM626405) when compared with the reference G3 genotype (M84663). The findings of the present study represent the demonstration of the buffalo strain in camels. As previously reported, humans can be infected by this genotype; accordingly, the epidemiological importance of this genotype in the camel population should be considered in further studies.     

Further molecular discrimination of Spanish strains of Echinococcus granulosus

We have designed two polymerase chain reaction (PCR) primer sets (PEg9F1-PEg9R1 and PEg16F1-PEg16R1) and two PCR protocols (Eg9-PCR and Eg16-PCR) for discrimination of Echinococcus granulosus genotypes. The oligonucleotide sequences originate from two E. granulosus DNA multiplex-PCR amplification fragments, previously reported, that allows species-specific discrimination between Taenia saginata, Taenia solium, and E. granulosus. The Eg9-PCR, Eg16-PCR, and Eg9-PCR linked restriction fragment length polymorphism (RFLP) analysis was used to characterize 53 E. granulosus isolates from the central region of Spain, highly endemic for echinococcosis. The analysis resulted in: (i) the discrimination of E. granulosus from Echinococcus multilocularis; (ii) the characterisation and discrimination of discrete E. granulosus strains from Spain; and (iii) the identification of two distinct genotypes within E. granulosus Spanish pig isolates. To further characterize the genetic variants in pigs, fragments of the NADH dehydrogenase I (ND1) and the cytochrome c oxidase subunit I (CO1) genes were amplified from parasite DNA and sequenced. The results again revealed the presence of two distinct genotypes: the G1 (sheep-dog strain) and G7 (pig-dog strain) genotypes. This observation could have important consequences for human health in Spain. Furthermore, the Eg9-PCR, Eg16-PCR, and Eg9-PCR-RFLP protocols can be used as additional methods to discriminate various E. granulosus genotypes.     

Canine echinococcosis in Kyrgyzstan: using prevalence data adjusted for measurement error to develop transmission dynamics models

Echinococcosis is a major emerging zoonosis in central Asia. A cross-sectional study of dogs in four villages in rural Kyrgyzstan was undertaken to investigate the epidemiology and transmission of Echinococcus spp. A total of 466 dogs were examined by arecoline purgation for the presence of Echinococcus granulosus and E. multilocularis. In addition, a faecal sample from each dog was examined for taeniid eggs. Any taeniid eggs found were investigated using PCR techniques (multiplex and single target PCR) to improve the diagnostic sensitivity by confirming the presence of Echinococcus spp. and to identify E. granulosus strains. A total of 83 (18%) dogs had either E. granulosus adults in purge material and/or E. granulosus eggs in their faeces as confirmed by PCR. Three genotypes of E. granulosus: G1, G4 and the G6/7 complex were shown to be present in these dogs through subsequent sequence analysis. Purge analysis combined with PCR identified 50 dogs that were infected with adult E. multilocularis and/or had E. multilocularis eggs in their faeces (11%). Bayesian techniques were employed to estimate the true prevalence, the diagnostic sensitivity and specificity of the procedures used and the transmission parameters. The sensitivity of arecoline purgation for the detection of echinococcosis in dogs was rather low, with a value of 38% (credible intervals (CIs) 27-50%) for E. granulosus and 21% (CIs 11-34%) for E. multilocularis. The specificity of arecoline purgation was assumed to be 100%. The sensitivity of coproscopy followed by PCR of the isolated eggs was calculated as 78% (CIs 57-87%) for E. granulosus and 50% (CIs 29-72%) for E. multilocularis with specificity of 93% (CIs 88-96%) and 100% (CIs 97-100%), respectively. The 93% specificity of the coprological-PCR for E. granulosus could suggest coprophagia rather than true infections. After adjusting for the sensitivity of the diagnostic procedures, the estimated true prevalence of infection of E. granulosus was 19% (CIs 15-25%) and the infection pressure in the dog population was estimated to be 0.29 infections per year (CIs 0.014-0.75). Logistic regression analysis failed to identify any significant risk factors for infections for E. granulosus. After adjusting for the sensitivity of the test procedures, the estimated true prevalence for E. multilocularis was 18% (CIs 12-30%). Dogs that were restrained had a significantly lower prevalence of E. multilocularis of 11% (CIs 6-29%) compared with 26% in free-roaming dogs (CIs 17-44%) and independently within these groups hunting dogs were more likely to be infected than non-hunting dogs.