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Background: Dirofilaria immitis, also known as heartworm, is one of the most important parasitic nematodes of domestic dogs, causing a potentially serious disease, cardiopulmonary dirofilariosis, which can be lethal. This species seems to be less 'expansive' than its sister species Dirofilaria repens, and it is believed that climate change facilitates the spread of this parasite to new non-endemic regions. Methods: In total, 122 heartworm isolates were analysed from nine endemic countries in Europe (Portugal, Spain, Italy, Greece, Hungary, Romania, Slovakia, and Ukraine) and a single isolate from Bangladesh by amplification and sequencing of two mitochondrial (mt) DNA markers: cytochrome c oxidase subunit 1 (COI) and dehydrogenase subunit 1 (NADH). The main aim of the current study was to determine the genetic diversity of D. immitis and compare it with D. repens haplotype diversity and distribution. DNA was extracted from adult heartworms or microfilariae in blood. Most isolates originated from dogs (Canis lupus familiaris) while 10 isolates originated from wildlife species from Romania, including eight isolates from golden jackals (Canis aureus), one isolate from a Eurasian otter (Lutra lutra) and one isolate from a red fox (Vulpes vulpes). Results: Median spanning network analysis was based on the combined sequence (1721 bp) obtained from two mt markers and successfully delineated nine haplotypes (Di1-Di9). Haplotype Di1 was the dominant haplotype encompassing 91 out of the 122 sequences (75%) from all nine countries and four host species. Haplotype Di2 was the second most common haplotype, formed solely by 13 isolates from Italy. The remaining sequences were assigned to Di3-Di9 haplotypes, differing by 1–4 SNPs from the dominant Di1 haplotype. There was evidence for geographical segregation of haplotypes, with three unique haplotypes associated with Italy and four others associated with certain countries (Di4 and Di7 with Slovakia; Di8 with Greece; Di6 with Hungary). Conclusion: Diversity in D. immitis mt haplotypes was lower by half than in D. repens (9 vs. 18 haplotypes in D. immitis and D. repens, respectively), which may be associated with the slower expansion of heartworm in Central and NE Europe. NADH gene appears to be conserved in Dirofilaria sp. by showing lower genetic diversity than the analysed COI gene. [ABSTRACT FROM AUTHOR]

Introduction and objective. It is easier and non-invasive to obtain faecal samples compared with blood samples. Molecular techniques may enable detection of parasites even in tiny amounts of blood-containing faeces. We aimed to compare the sensitivity of detection of three Babesia species and Hepatozoon canis in blood and faecal samples, including samples derived from naturally infected hosts. Materials and method. Three groups were involved: 1) Nine BALB/c mice infected with Babesia microti sampled during acute (n=3), post-acute (n=3) and chronic phases of infection (n=3); 2) Eight dogs with symptoms of babesiosis; 3) Six red foxes infected with B. vulpes, one fox infected with B. canis, four foxes infected with H. canis. Genomic DNA was extracted from blood and faeces by use of commercial kits and amplified with genus-specific primers in one-step or nested PCR reactions. Selected PCR products were sequenced. Results. No positive results for faecal samples were obtained from H. canis-positive foxes in contrast to Babesia spp. infections. Positive results from PCRs were obtained for all BALB/c mice (100%), five dogs (62.5%) and four of seven foxes (57.1%). Successful sequencing was obtained for six selected murine samples (B. microti), four canine samples (B. canis) and for one fox sample (B. vulpes). The success of B. microti detection in murine faecal samples from acute, post-acute and chronic phases was identical (100%). Conclusions. Detectability of Babesia spp. infections was lower in naturally infected dogs and foxes, compared to experimentally infected mice. Detection of DNA in faecal samples can be useful in the detection of Babesia infection in populations from which blood samples are hard to obtain, but due regard must be given to the possibility that prevalence of infection may be severely underestimated. [ABSTRACT FROM AUTHOR]