Occurrence of Blastocystis-subtypes in patients from Italy revealed association of ST3 with a healthy gut microbiota

An epidemiological survey on Blastocystis was carried out enrolling a total of 2524 subjects referred to the Umberto I Academic Hospital in Rome, for the routine parasitological exams, during 2017–2018. The studied population included a sample of immunocompromised individuals (N = 130) followed at the same hospital. DNA sequencing of the small subunit rRNA gene (SSU rDNA) locus was performed on samples positive to the coproparasitological analysis to molecular characterize the Blastocystis-subtypes. Microscopical analysis detected Blastocystis in 192/2524 (7.6%) of the enrolled subjects. It was the organism most frequently identified in the analysed faecal samples diagnosed in single infection (5.6%) or in co-infection with other enteric protozoa (2%). Furthermore, it was found mainly in immunocompromised patients (22.3%) compared to immunocompetent ones (6.8%). As expected, ST3 was the most occurring subtype identified in 40% of the subjects, followed by ST1 (29%), ST2 (16%), ST4 (12%), and ST7 (3%). Next-generation sequencing (NGS) of the 16S rDNA was performed on a sub-sample of Blastocystis-ST3-carriers, homogenous by age and gender, as well as on Blastocystis-free subjects, to profile and compare their gut bacterial composition. A higher bacterial diversity was found in ST3-Blastocystis-carriers, which exhibited a high abundance of Prevotella, Methanobrevibacter and Ruminococcus while, a high percentage of Bacteroides was found in Blastocystis-free subjects. This study evidenced the presence of Blastocystis in 7.6% of faecal samples in Italy and a high circulation of the protist among immunocompromised patients (22.3%). Molecular characterization of positive samples evidenced the occurrence of five different subtypes, including zoonotic ST such as the ST7, highlighting the risk of transmission from animals. Study of the gut microbiota composition confirms previous evidences according to which, the colonisation by Blastocystis would be linked with an eubiotic gut characterized by potentially beneficial species such as Prevotella and Ruminococcus, rather than with a dysbiotic state, with a high abundance of Enterobacteriaceae, and corroborated the role of the protist as “an old friend” of the human gut

Taeniid cestodes in a wolf pack living in a highly anthropic hilly agro-ecosystem

The Italian wolf population in human-modified landscapes has increased greatly in the last few decades. Anthropisation increases the risk of transmission of many zoonotic infections and in this context, control of taeniid cestode species needs to be addressed from a One Health perspective. Predator-prey interactions are at the root of taeniid cestode transmission, and the wolf plays a key role in the maintenance and transmission of taeniids. To date, all available data on the taeniids of wolves in Italy refer to populations living in a wild habitat. Between 2018 and 2019, we investigated taeniids in a wolf pack living in a highly anthropic hilly agro-ecosystem. Thirty-eight faecal samples were collected and analysed, 4 of which were also genetically characterised for individual wolves and belonged to three different animals. Samples collected were analysed microscopically and by molecular analysis in order to identify the taeniid species. Taeniid eggs were detected in 34.2% (13/38) of samples. Within samples positive to taeniid eggs only Echinococcus granulosus s.s. and Taenia hydatigena were identified in 26.3% and 10.5% of the samples, respectively. On microscopic examination, Capillaria spp., Ancylostomatidae and Toxocara canis eggs, Crenosoma vulpis larvae, and coccidian oocysts were also found. The combination of low biodiversity of taeniid species with a high occurrence of E. granulosus s.s. recorded in this study could be the consequence of a deeper link occurring between wolves and livestock in human-modified landscapes than in wild settings

Usual or unusual presentations of Dirofilaria repens in two sibling dogs: a case report

This study describes two different manifestations of Dirofilaria repens infection in sibling dogs with microfilaremia. Dog 1, asymptomatic, harbored a gravid female of D. repens on the parietal layer of tunica vaginalis of one testicle and showed a marked circulating eosinophilia (3.3·103/μL). Both testicles were normal in shape and size without any gross lesions. Dog 2 had a pyotraumatic dermatitis. The cases were confirmed by PCR and sequencing. The sequences obtained showed 100% identity with those of D. repens isolated from human scrotum in Croatia. The treatment with moxidectin 2.5% and imidacloprid 10%/kg was effective in eliminating microfilariae after just one application, as demonstrated by negative modified Knott’s tests and PCR analyses of blood samples. This status was maintained during the post-treatment observation period. The classical localization of D. repens in dogs is in subcutaneous tissues, within nodules or free; however, it can also occur with some frequency in testicles, as described in humans. The infection can be associated with circulating eosinophilia or pyotraumatic dermatitis, as reported in this study. Thus, in endemic areas, it is advisable to carefully inspect the removed testicles at neutering since parasite localization can take place without any macroscopic changes. Moreover, in the case of circulating eosinophilia or pyotraumatic dermatitis, investigations should include modified Knott’s test and PCR to ensure that D. repens is not the cause of these alterations. Rapid and sensitive tests for the early detection of infected animals would help to prevent or limit the spread of this zoonosis

Molecular subtyping of blastocystis sp. Isolated from farmed animals in southern Italy

Blastocystis is a common intestinal protist distributed worldwide, infecting humans and a wide range of domestic and wild animals. It exhibits an extensive genetic diversity and, so far, 25 distinct small subunit ribosomal RNA (SSU rRNA) lineages termed subtypes (STs)) have been characterized; among them, 12 have thus far been reported in humans. The aims of the present study were to detect and genetically characterize Blastocystis sp. in synantropic animals to improve our current knowledge on the distribution and zoonotic transmission of Blastocystis STs in Italy. Samples were collected from N = 193 farmed animals and submitted to DNA extraction and PCR amplification of the SSU rRNA. Blastocystis was detected in 60 samples (31.08%) and successfully subtyped. Phylogenetic analysis evidenced that the isolates from fallow deer, goats, and pigs (N = 9) clustered within the ST5; those from pheasants (N = 2) in the ST6; those from chickens (N = 8) in the ST7; those from sheep (N = 6) in the ST10; and those from water buffaloes (N = 9) in the ST14 clade. The comparison between the present isolates from animals and those previously detected in humans in Italy suggested the animal-to-human spillover for ST6 and ST7. The present study represents the widest Blastocystis survey performed thus far in farmed animals in Italy. Further epidemiological studies using molecular approaches are required to determine the occurrence and distribution of Blastocystis STs in other potential animal reservoirs in Italy and to define the pathways of zoonotic transmission

Molecular Identification of New Cases of Human Dirofilariosis (Dirofilaria repens) in Italy

1) Dirofilariosis is a vector‐borne parasitic disease mainly in domestic and wild carnivores caused by Dirofilaria (Noctiella) repens, which is endemic in many countries of the Old World, and D. immitis, which has a worldwide distribution. In recent years, an increase in the number of human cases has been reported, suggesting that dirofilariosis is an emergent zoonosis. Here, we describe further cases (N = 8), observed in Central Italy during the years 2018–2019. 2) Molecular diagnosis was performed on: i) live worms extracted from ocular conjunctiva, cheek, and calf muscle; ii) histological sections of surgically removed nodules from parenchymal lung, coccyx, and breast. 3) Sequence analysis (650‐bp) of the mitochondrial cytochrome oxidase subunit I gene (mtDNA cox1) showed a match of 100% with the sequences of D. repens previously deposited in GenBank. ELISA test to detect IgG against filarial antigens was performed on four patients’ sera and resulted positive in two patients who showed ocular and subcutaneous dirofilariosis, respectively. Microfilariae have been never detected in the peripheral blood of the patients. 4) The occurrence of N = 8 new cases of human D. repens‐infections observed in a two‐year period suggests an increased circulation of the parasite in Italy. Therefore, dirofilariosis should be included in differential diagnosis in patients presenting subcutaneous and/or pulmonary nodules. Molecular diagnosis of the etiological agents is fundamental. Specific serological diagnosis needs to be improved in future research work