Paola Pepe

Comparison of individual and pooled faecal samples in sheep for the assessment of gastrointestinal strongyle infection intensity and anthelmintic drug efficacy using McMaster and Mini-FLOTAC.

A field study was conducted to validate pooled faecal samples in sheep for the assessment of gastrointestinal (GI) strongyle infection intensity (faecal egg count - FEC) and anthelmintic drug efficacy (FEC reduction - FECR). Ten sheep farms located in the Campania region of southern Italy were selected for the study. In each farm, individual faecal samples from 20 adult sheep (when possible) were collected, before (D0) and after (D14) an anthelmintic treatment with albendazole. For each farm and at each time point (D0 and D14) the faecal samples were examined individually and as pools. Specifically, three different pool sizes (5, 10 and 20 individual sheep samples) and three different analytic sensitivities (namely 10 using Mini-FLOTAC; 15 and 50 using the two variants of McMaster - McM15 and McM50) were compared for FEC and FECR using individual and pooled faecal samples. GI strongyle intensity (eggs per gram of faeces - EPG) of pooled samples correlated positively with mean EPG of individual samples, with very high correlation coefficients (ranging from 0.94 to 0.99) across the 3 different pool sizes and analytic sensitivities. Mini-FLOTAC was more sensitive compared to the two variants of McMaster (McM15 and McM50) for the diagnosis of GI strongyles in sheep (100% vs 88.5% vs 75.9%) and resulted in significant higher FEC compared to both McM15 and McM50, with a mean difference in egg counts of approximately 90 EPG (p<0.001). The drug efficacy results showed that FECR was higher than 98% at most farms independently of the pool size and analytic sensitivity. With the exception of two farms, FECR was 100% when calculated for individual animals and across the different pool size and analytic sensitivities. In conclusion, the present study highlighted that pooling ovine faecal samples is a rapid procedure that holds promise as a valid strategy for assessing GI strongyles FEC and FECR in sheep.

Mini-FLOTAC, a new tool for copromicroscopic diagnosis of common intestinal nematodes in dogs

BackgroundThe most common intestinal nematodes of dogs are Toxocara canis, hookworm and Trichuris vulpis. The present study was aimed to validate a new copromicroscopic technique, the Mini-FLOTAC and to compare its diagnostic efficiency and sensitivity with four other copromicroscopic techniques: direct smear, tube flotation, Wisconsin method and the FLOTAC dual technique.FindingsTwo experiments were performed. In the first, faecal positive samples collected from 59 stray asymptomatic dogs, of which 21 were naturally infected with ancylostomidae, 13 naturally infected with T. canis and 25 naturally infected with T. vulpis were used to validate the Mini-FLOTAC technique. The second experiment was performed on faecal samples randomly selected from 38 stray asymptomatic dogs to compare the diagnostic efficiency and sensitivity of the different techniques. Samples were fixed with 5% formalin; sodium chloride and zinc sulphate were used for flotation solutions because they performed best for detecting and quantifying intestinal nematode eggs in dogs. Mini-FLOTAC and FLOTAC were the most efficient and sensitive techniques and they gave higher EPG and higher numbers of positive samples in both the experiments, for all three parasites.ConclusionsAs Mini-FLOTAC does not require centrifugation it is a very promising technique for counting helminth eggs in dog faeces.

Detection of Dirofilaria immitis in mid-western arid Argentina

Dirofilariosis, caused by Dirofilaria immitis and D. repens, is (re-) emerging worldwide. Dogs are the main reservoirs, while human infection has recently become an important focus of interest and attention. In Argentina, canine D. immitis infection has been described in eastern and northern subtropical and temperate humid regions, but never reported in mid-western arid regions so far. In this research note we report for the first time the occurrence of autochthonous human and canine D. immitis infection in the region.

Survey of co-infection by Salmonella and oxyurids in tortoises

BackgroundSalmonella spp. and oxyurids are among the most prevalent bacterial and parasitic agents in reptiles. These organisms are routinely isolated in healthy tortoises, although heavy infections may cause significant pathology. Tortoises are considered a common source of reptile-associated salmonellosis, an important zoonosis reported worldwide. A survey of the prevalence of Salmonella spp. and oxyurids in 53 tortoises was conducted in southern Italy and a possible correlation between the two pathogens was therefore investigated.ResultsSalmonella spp. and oxyurids were detected with a prevalence of 49.1 and 81.1%, respectively. A significant positive correlation between Salmonella spp. and oxyurids was demonstrated. However, confounding factors related to husbandry could have been involved in determining this correlation.ConclusionsOur results suggest that caution should be exercised in translocation, husbandry, and human contact with tortoises and other exotic pets. Further studies on the epidemiology, molecular characterization and pathogenesis of Salmonella and oxyurids are needed to assess the actual impact of these organisms, as single or associated infections, on tortoises and on other exotic pets.

Angiostrongylus vasorum: epidemiological, clinical and histopathological insights

BackgroundCanine angiostrongylosis is a nematode infection in domestic dogs and wild carnivores. The present report focuses on epidemiological, clinical and histopathological findings in a case of fatal disseminated angiostrongylosis in a dog living in southern Italy and provides data on the extent of the spread of Angiostrongylus vasorum in the same area.Case presentationA 4-year-old female English Setter from the Campania region of southern Italy was referred with a 2-week history of cough and severe respiratory distress that did not respond to antimicrobial therapy. Based on clinical, radiological, echographical and cytological findings (including the presence of larvae), a suspect diagnosis of lungworm infection was performed. After few days the dog died due to progressive clinical aggravation. Complete postmortem examination was conducted within 24 hours from death and samples from lungs, heart, liver, kidney, spleen, stomach and small intestine were fixed in 10% buffered formalin. Grossly, several hemorrhagic foci were observed mostly in the lungs, liver, kidney. Microscopically, the lungs contained numerous, multifocal to coalescing granulomas composed of epitheliod macrophages, multinucleated giant cells and some neutrophils, frequently associated with parasite eggs and larvae. The lungs contained many firm nodules, many adult nematodes approximately 1.5 to 2 cm in length were observed in cut sections and identified as A. vasorum. A subsequent parasitological survey performed with FLOTAC on stray dogs living in the same area showed the presence of A. vasorum larvae in 17 of 1639 stray dogs examined (1.04%).ConclusionThis survey provides new data on distribution of A. vasorum and underlines that canine angiostrongylosis should be considered as differential diagnosis in dogs.

Dicrocoelium dendriticum induces autophagic vacuoles accumulation in human hepatocarcinoma cells.

The relationship between Dicrocoelium dendriticum and cancer has been poorly investigated so far, but a large amount of findings suggest that other trematodes can favour cancer in both animals and humans. In this study, the effects of D. dendriticum on cell proliferation, cell death mechanisms and oxidative stress induction were evaluated in hepatocellular carcinoma (HCC) cell lines (HepG2 and HuH7). Results showed that short time exposure to low concentrations of somatic antigens from D. dendriticum caused slight proliferation in both HepG2 and HuH7 cells while high concentrations and long exposure time to extracts from D. dendriticum caused a significant growth inhibition. This effect was, however, not paralleled by apoptosis but it occurred with an about 40% increase of the formation of autophagic vacuoles. In the same experimental conditions, a strong oxidative stress was recorded with an about 100% increase of the intracellular O(2-). These data suggest the occurrence of an escape anti-apoptotic mechanism in HCC cells. In conclusion, these results suggest a role for D. dendriticum in the chronic oxidative stress and in the regulation of transformation processes in HCC warranting additional investigations in this specific area of research.

Synanthropic Birds and Parasites

SUMMARY This paper describes the parasitologic findings for 60 synanthropic bird carcasses recovered in the Campania region of southern Italy. Birds consisted of 20 yellow-legged gulls (Larus michahellis), 15 rock pigeons (Columba livia), 15 common kestrels (Falco tinnunculus), and 10 carrion crows (Corvus corone). Each carcass was examined to detect the presence of ectoparasites and then necropsied to detect helminths. Ectoparasites occurred in 100% of the birds examined. In particular, chewing lice were recovered with a prevalence of 100%, whereas Pseudolynchia canariensis (Hippoboscidae) were found only in pigeons with a prevalence of 80%. Regarding endoparasites, a total of seven helminth species were identified: three nematodes (Ascaridia columbae, Capillaria columbae, Physaloptera alata), one cestoda (Raillietina tetragona), one trematoda (Cardiocephalus longicollis), and two acanthocephalans (Centrorhynchus globocaudatus and Centrorhynchus buteonis). The findings of the present study add data to the parasitologic scenario of synanthropic birds. This is important because parasitic infection can lead to serious health problems when combined with other factors and may affect flying performance and predatory effectiveness. RESUMEN Aves sinantrópicas y parásitos. Este trabajo describe los hallazgos parasitológicos de 60 cadáveres de aves sinantrópicas recuperadas en la región de Campania, en el sur de Italia. La aves incluyeron 20 gaviotas patiamarillas (Larus michahellis), 15 palomas domésticas (Columba livia), 15 cernícalos comunes (Falco tinnunculus), y 10 cornejas comunes (Corvus corone). Cada canal se examinó para detectar la presencia de ectoparásitos y luego se realizó la necropsia para detectar helmintos. Los ectoparásitos se presentaron en 100% de las aves examinadas. En particular, se recolectaron piojos masticadores con una prevalencia de 100%, mientras que la Pseudolynchia canariensis (Hippoboscidae) se encontró sólo en las palomas con una prevalencia del 80%. En cuanto a los endoparásitos, se identificaron un total de siete especies de helmintos: tres nematodos (Ascaridia columbae, Capillaria columbae, Physaloptera alata), un cestodo (Raillietina tetragona), un trematodo (Cardiocephalus longicollis) y dos acantocéfalos (Centrorhynchus globocaudatus y Centrorhynchus buteonis). Los resultados del presente estudio contribuyen con datos en el escenario parasitológico de las aves sinantrópicas. Esto es importante ya que la infección parasitaria puede conducir a serios problemas de salud cuando se combina con otros factores y puede afectar al rendimiento de vuelo y la eficacia depredadora.

Ostertagia ostertagi antibodies in bulk tank milk from dairy cattle in Italy: A nation-wide survey

The objective of this study was to investigate the exposure to Ostertagia ostertagi in dairy cattle herds across Italy through measurement of antibody concentration in the bulk tank milk (BTM) and to evaluate the possible effect of regions, seasons and pasture management practices on the level of exposure to the parasite. The O. ostertagi antibody levels in milk were determined using a commercial ELISA kit (SVANOVIR® O. ostertagi-Ab) and expressed as optical density ratio (ODR). From March 2015 to September 2017, BTM samples were collected in 1209 dairy cattle farms located in 15 regions of Northern, Central and Southern Italy. As reported in previous studies, BTM samples were classified in three different categories according to the level of exposure to O. ostertagi as follows: (i) positive with ODR values >0.60; (ii) negative with ODR values <0.30 and (iii) grey zone with ODR values between 0.30 and 0.60. Finally, differences in ODR values between regions, seasons and herd management practices were studied using the one-way analysis of variance. The overall mean ODR revealed a value of 0.49 in Italy; in particular, dairy cattle farms located in Southern Italy showed the highest O. ostertagi BTM ODR values than the other regions (p < 0.05). The ODR values ranged from 0.45 in autumn to 0.53 in spring and the seasonal difference was significant (p < 0.05). Furthermore, a positive association was found between the ODR values and the access to pasture; herds with access to pasture showed significantly higher titres (p < 0.0001) of O. ostertagi antibodies in milk (mean ODR = 0.63) than in those in total-confinement housing (mean ODR = 0.42). This study, the first of its kind in Italy, will provide a quantitative assessment exposure to O. ostertagi of Italian dairy herds and represents a significant step forward in evidence-based medicine for dairy veterinarians, advisors and farmers.

Innovative tools for the diagnosis of Echinococcus granulosus in definitive hosts

The aim of this study was to develop and validate an innovative protocol for the diagnosis of Echinococcus granulosus and other Taeniidae in dogs. For this purpose, three experiments were performed, using faecal samples from naturally infected dogs. Firstly, the FLOTAC technique was calibrated with five flotation solutions: saturated sodium chloride (specific gravity, s.g. = 1.20), zinc sulphate (s.g. = 1.35), zinc chloride (s.g. = 1.45), Breza (s.g. = 1.30) and modified Breza (s.g. = 1.40). Then, FLOTAC was compared with four techniques of flotation in centrifuge using: zinc sulphate (s.g. = 1.20), Breza (s.g. = 1.30), modified Breza (s.g. = 1.40), and zinc chloride (s.g. = 1.45). Finally, four different protocols of DNA extraction were compared for Taeniidae identification: QIAamp Tissue Kit and QIAamp Stool from eggs; QIAamp Stool and Wizard Magnetic Purification System for Food from faeces. FLOTAC with zinc sulphate was the most efficient method to detect Taeniidae eggs, showing highest mean of eggs per gram (EPG) of faeces. The QIAamp Stool, using eggs concentrated by FLOTAC, was the best method for DNA extraction. The combination of these protocols provided the highest number of positive samples with PCR, i.e., 47/50 (94.0%). The three negative samples showed a low faecal egg count (2 EPG) below the detection limit (4 EPG) of the protocol. From sequencing of the 47 positive samples: 6 samples were identified as E. granulosus sensu stricto (s.s.), 28 as Taenia hydatigena and 6 as T. pisiformis. A co-infection between different genera of Taeniidae was found in 7 samples.

A national survey of Ixodidae ticks on privately owned dogs in Italy

BackgroundThe geographical distribution of ticks on companion animals needs to be monitored to develop and plan effective control measures, as suggested by the European Scientific Counsel on Companion Animal Parasites. The aim of this study was to conduct the first Italian national survey of tick distribution on privately owned dogs.MethodsThe study was performed over 20 months (February 2016 - September 2017) and involved 153 veterinary practices in 64 different provinces covering 17/20 (85%) Italian regions. Participating practitioners were asked to examine five different dogs per month at random and complete a questionnaire for each dog. Differences in tick infestation associated with: sex, age and hair length (long and short); the dog’s habitat (indoor or outdoor/kennel); and the dog’s environment (urban or rural/sylvatic) were evaluated. The attachment site of ticks on the dog was also recorded. Acaricide efficacy was evaluated for the subset of dogs for which complete information on product used, date of sampling and date of last ectoparasiticide treatment was available.ResultsOf the 3026 dogs examined, 1383 (45.7%) were carrying at least one tick. Overall, 2439 tick samples were collected and a total of 14 tick species identified. Rhipicephalus sanguineus group were the most predominant ticks (63.6%), followed by Ixodes ricinus (30.6%) and I. hexagonus (5.6%). Twenty-four dogs had mixed tick infestations. Long-haired dogs had a higher tick infestation risk as did dogs with outdoor and rural/sylvatic lifestyles. Ticks were located on the head (37.4%), the neck (28.8%), the muzzle (15.5%) and the back (15.3%). A higher prevalence of Rhipicephalus was found in the interdigital spaces (10.8%) compared to Ixodes (0.2%). Finally, ectoparasiticide treatments were found significantly protective against tick infestation, especially orally administered formulations.ConclusionsPrivately owned dogs in Italy have a high prevalence (45.7%) of infestation with ixodid ticks and this risk varies by dog phenotype and lifestyle.