Study on the helminth fauna of red foxes and dogs in Liguria (north-west Italy): epidemiological and diagnostic aspects

An epidemiological survey on the helminths of 165 foxes and 450 rural dogs from N-W Italy (Imperia and Savona districts) was conducted between 2010 and 2012. Foxes’ cardiorespiratory system, gastrointestinal tract, liver, urinary apparatus, muscle tissue and rectal faecal samples were examined. For each dog faecal and blood samples were collected: feacal samples were examined by centrifugal floatation and by Baermann technique, blood samples were Knott test, serological examination for antigens of Dirofilaria immitis, histochemical staining and PCR. Dogs’ serum samples were also tested with two newly developed ELISA tests for the detection of circulating A. vasorum antigens and specific antibodies. Several species of Trichuridae nematodes found during the study were examined by scanning electron microscopy (SEM) (applied on eggs) and were subjected to biomolecular techniques to characterize parts of the ribosomal and mitochondrial DNA (for adults and eggs), in order to investigate morphological and genetic aspects of this complex group of nematodes. A wide variety of parasites were found in foxes at necropsy: Angiostrongylus vasorum (78%), Eucoleus aerophilus (42%), Eucoleus boehmi (1 of 2 foxes), Crenosoma vulpis (16%) and Filaroides sp. (5%) in the cardiopulmonary tract; Spirocerca lupi (23.5%), Physaloptera spp. (2.5%), Uncinaria stenocephala (70%), Toxascaris leonina (27%), Toxocara canis (25%), Molineus legerae (24%), Trichuris vulpis (21%), Aonchoteca putorii (9%), Pterygodermatites affinis (5%), cestodes of the genus Mesocestoides (82%), of the genus Taenia (5.5%), of the family Dilepididae (27%) and one trematode species (Plagiorchidae) (0.6%) in the gastrointestinal tract and Pearsonema plica (57%) in the urinary bladder. Echinococcus multilocularis, Trichinella spp. and filarial infections were not found. In dogs’ coprology larvae of A. vasorum (0.6%) and C. vulpis (0.3%) and eggs of E. aerophilus (9.6%), E. boehmi (1.6%), T. canis (20%), T. vulpis (18%), Ancylostomatidae (12%), Capillaria spp. (1.8%), Physaloptera spp. (0.2%), Hymenolepis diminuta (1.1%) and Dipylidium caninum (0.2%) were observed. D. immitis (1.7%), D. repens (1%) and Acanthocheilonema reconditum (7%) were found in dogs, in a region historically free from filariosis.In a subset of dogs (n=347) which were contemporaneously tested with the Baermann method and with the ELISA tests for A. vasorum, a higher prevalence was found with serology (0.3% with the Baermann method, 0.6% with the ELISA tests). No cross reactions were observed with the serological tests in those dogs known to be positive for other parasitic infections; serology proved to be a valid alternative diagnostic technique for A. vasorum. SEM allowed distinguishing differences in the external structure of the eggs of several Trichuridae nematodes (E. boehmi, E. aerophilus, P. plica, Calodium hepaticum and T. vulpis). A part of the small subunit rRNA (18S rRNA) gene and of the mitochondrial cytochrome oxidase subunit I (mt COI) gene was amplified from specimens of T. vulpis, E. aerophilus, E. boehmi, P. plica, A. putorii, C. hepaticum (found in the liver of a fox from Cuneo and of a vole from Switzerland) and Calodium splenaecum (in the spleen of a Crocidura sp. from Switzerland), yielding the first 18S rRNA gene sequences of all the capillariid nematodes and the first mt COI sequences of E. boehmi, P. plica, C. hepaticum, A. putorii and T. vulpis. The 18S rRNA gene is highly conserved among the different species, while interspecies divergence was much higher at the mtCOI gene locus, making it suitable for resolving closely related species and for the future development of Trichuridae species-specific primers

Cardiorespiratory and gastrointestinal parasites of dogs in north-west Italy

The present study was conducted to evaluate the prevalence of gastrointestinal and cardio-respiratory parasites in dogs in Liguria (northwest Italy) which is a region that has scarcely been studied. A total of 450 dogs (260 males and 190 females) were enrolled in the study from 2009 and 2013. All dogs lived in a rural or semi-rural environment in the provinces of Imperia (n = 352) or Savona (n = 98), Liguria. Coprological examinations showed that 197 dogs (43.8 %, CI 38.7-48.9 %) were infected by parasites. Specifi cally, 3.3 % of the dogs presented only cardio-respiratory species, 32.4 % only intestinal species, and 8.0 % presented a mixed infection. The most frequent intestinal parasites were Toxocara canis (20 %), Trichuris vulpis (17.8 %), Ancylostomatidae (12 %), Coccidia (2.7 %), Aonchotheca putorii (1.8 %) and Toxascaris leonina (1.8 %). The cardio-respiratory species found were Eucoleus aerophilus (9.6 %), Eucoleus boehmi (1.6 %), Angiostrongylus vasorum (0.7 %), and Crenosoma vulpis (0.2 %). A total of 116 dogs (25.8 %) were parasitized by a single species, multiple infections were observed in 81 dogs (18.0 %) up to a six-order infection. This preliminary study highlighted that the prevalence of intestinal parasites in investigated area is high. Cardio respiratory parasites were detected in an area that has not been investigated before. The creation of a more extensive sampling programme of the area, on a provincial basis in order to build a more detailed map of prevalences for different species of dog parasites throughout Liguria, a more extensive sampling programme of the area needs to be created - ideally for each province

Occurrence and viability of Anisakis spp. larvae in ready to eat products made of herring (Clupea harengus) sold in Italy

The study aimed to assess the occurrence and viability of Anisakis spp. larvae in ready to eat products made of herring, a common host of Anisakis spp. and the third most commercialized fish species in the EU. A total of 135 products consisting of 50 smoked whole specimens and 85 filleted products (25 smoked, 30 marinated, 30 canned) were sampled from 2016 to 2018. Viscera and muscle of whole herrings were visually inspected and separately digested. Filleted products were also visually inspected and digested. Larvae viability was assessed, then they were counted and microscopically identified to genus level. A subsample was molecularly identified. At least one Anisakis spp. larva was found in 56 products (41.5%), for a total of 1715 larvae collected (0-172 larvae/product). Most of the larvae (91%) were found in the viscera of 49 of the 50 whole herrings (98%). A highly significant difference was observed between the positivity rate and larval density at muscle level, as 149 larvae were found in the muscle of 31 whole herrings (positivity rate 62%, 0.022 larval density/g), while only 7 larvae were found in the 85 filleted products (positivity rate 7%, 0.001 larval density/g). Larvae were molecularly identified as A. simplex. The study showed that obviously contaminated products are commercialized. All the larvae were dead, proving a negligible risk of developing anisakiasis. However, their allergenic potential is debated and the significant difference between muscle infection levels in whole and filleted herrings may result in a different risk of exposure to antigens

DETERMINATION OF OCHRATOXIN A IN FARMED FISH BY ENZYMATIC DIGESTION (ED) COUPLED TO HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY WITH A FLUORESCENCE DETECTOR (HPLC-FLD)

Several studies have demonstrated that fish feeds contain significant concentrations of chemical contaminants, many of which can bioaccumulate and bioconcentrate in fish tissues (1).  The serious concern regarding the use of fish meal and fish oil in the the aquaculture industry has led to extensive search of alternative raw materials for aquafeeds. The most obvious alternatives are oils and proteins of plants origin. The use of these alternative feed ingredients can introduce contaminants that were previously not associated with fish farming such as mycotoxins (2). Ochratoxin A (OTA) is a mycotoxin produced as a secondary metabolite by various Aspergillus and Penicillium species with nephrotoxic, carcinogenic, immunotoxic and teratogenic potential (3). OTA has been found in several food commodities, including cereals and can also be present in food of animal origin as a result of carryover from contaminated feed (3). The aim of the present study was to determine OTA concentrations in muscle, kidney and liver of 10 seabream and 10 seabass of farmed origin collected on the market. Analysis will be performed by using an enzymatic digestion (ED) method coupled to high-performance liquid chromatography with a fluorescence detector (HPLC-FLD). Fish tissues were digested for 1 hour at 37°C with a 1% pancreatin solution in a phosphate buffer and then cleaned up with ethylacetate. After being evaporated to dryness and re-dissolved, the sample was processed using HPLC-FLD. The method was validated for: specificity, recovery, trueness, selectivity, linearity, limit of detection (LOD) and limit of quntification (LOQ), repeatability and reproducibility. Recoveries of analytical method were higher than 85 % for all the matrices. Intra and inter-day repeatability expressed as relative standard deviation were less than 9%. The LOD and LOQ for liver and muscles samples were 0.001 and 0.002 μg/kg, respectively. The LOD and LOQ for kidney samples were 0.01 and 0.02 μg/kg, respectively. The highest concentrations of OTA were found in kidney of the 20 fishes analyzed (rang

Occurrence of microplastics in bivalves: can a systematic literature review support the risk assessment?

Microplastics (MPs) are a global environmental issue, particularly affecting the aquatic ecosystem. Due to their small size (<5 mm), MPs can be absorbed or ingested by aquatic organisms, and transferred through food webs. Toxic effects due to the ingestion of MPs, alone or contaminated with additives or pollutants, have been hypothesized. Human exposure is inevitable, also following accumulation in the food chain. Seafood, especially bivalves, being filter feeders and consumed as whole, are an important potential pathway. The scientific interest in the topic is rising, and several narrative reviews on MPs in food, including seafood, have been published since the publication of a statement on the presence of MPs and nanoplastics in food by EFSA in 2016 (1), highlighting a scarcity of data on MPs occurrence. The aim of this review was to systematically revise scientific papers (SPs) to assess the occurrence of MPs in different categories of bivalves (mussels, clams, oysters and scallop) worldwide. A double-step filtration was used, applying increasingly stricter quality criteria. Data on MPs abundance were first discussed focusing on all the investigated species and geographical areas. Then, a subset of SPs selected in the second filtering step was used to calculate the weighted MPs mean abundance and the human exposure per serving size. In the first filtering process 87 SPs, published between 2014 and 2020 in 30 different scientific journals, were retained. Overall, 67 species, 6 genera and 1 family of bivalves were analysed. Mussels were the most analysed (61 SPs), followed by clams (55 SPs), oysters (31 SPs), and scallops (7 SPs). Mytilus edulis and M. galloprovincialis were the most investigated species, followed by P. viridis, Mytilus sp., R. philippinarum and C. gigas. All these are commercial species, globally farmed and distributed. Marine FAO areas 61 and 27 were most investigated. Overall, MPs mean abundance was variably reported, as well as the use of different methods and procedural controls. Therefore, in this study, the weighted MPs mean abundance was calculated only including data from a subset of SPs (n=32; 37%). The overall weighted MPs mean abundance including data from all FAO areas was 1.19 MPs/g ww. The highest value was observed in FAO area 61 (2.33 MPs/g ww), while values <1 MPs/g ww were observed in FAO areas 27 and 57. Among bivalve categories, the highest weighted MPs mean abundance (overall FAO areas) was observed for scallops (1.99 MPs/g ww), followed by mussels (1.71 MPs/g ww), clams (0.84 MPs/g ww) and oysters (0.65 MPs/g ww). Thus, the consumption of standard portions of each bivalve category determines the ingestion of a different number of MPs depending on the FAO area; the highest value (⁓645 MPs) would be ingested with a portion of mussels from FAO area 61. Our findings confirmed the existence of quality issues and the lack of analytical standardization. A disparity among investigated species and geographical areas was observed, and only three studies addressed processed products. These aspects affect the outcome of systematic reviews to support risk assessment; future studies should explore the issue of MPs adopting an interdisciplinary perspective, integrating different technical and scientific competences to collect evidences for risk assessment and management

A Comparison of Marketing Techniques Among Military Recruiters

The U.S. Department of Defense spent $11 billion in enlistment and retention bonuses from 2006 to 2010, which had only a marginally positive effect on the enlistment rate for the Army. The case study addressed this business problem of recruiting by exploring marketing strategies successful recruiting professionals used to motivate individuals to join the military. The purpose of this study was to determine effective recruiting strategies. Therefore, it incorporated the conceptual framework of emergent strategy theory, which postulated the best strategies are neither completely planned nor completely random, but are rather an adaptation to changing dynamics and circumstances. The population consisted of 2 former recruiters, 1 from the Army and another from the Marine Corps, as well as 38 college students located in Wilkes-Barre, Pennsylvania. Data for the study morphed from face-to-face interviews and 3 focus groups comprised of 10 to 15 students each, for the purpose of addressing the research question. Data analysis occurred through a process of coding and theming. The 9 themes identified included tell the story, advertising strategies, and fit for duty. A lesson learned from these themes was that the key for successful recruiting strategies lies in aligning with the wants and needs of individuals in the target demographic. If senior leaders in the Department of Defense followed the recommendations provided, each of the branches of the military service could potentially achieve higher recruiting rates at a lower cost. The study could result in social change whereby eligible recruits could view the Army and Marine Corps as professions of arms in which individuals can live out their ideals of patriotism but also have a good quality of life due to the benefits of military service

Survey on the presence of bacterial, fungal and helminthic agents in off-leash dog parks located in urban areas in central-italy

Off-leash dog parks are designated public spaces where dogs can move freely, under their owners’ supervision. These areas, allowing animals to socialize and move freely, are fundamental for dogs’ welfare. However, different pathogens, even zoonotic, may be excreted by the attending animals and contaminate the environment. The aim of the present study was to verify the occurrence of bacterial, fungal and parasitic pathogens in off-leash dog parks located in Florence (central Italy). Between March and May 2019, 83 fecal samples, 43 soil samples and 23 water samples (from fountains and puddles) collected from 26 off-leash fenced areas were examined. Fecal samples scored positive for Yersinia spp. (n = 7), Listeria innocua (n = 4), Toxocara canis eggs (n = 2) and Ancylostoma caninum/Uncinaria stenocephala eggs (n = 1). Keratinophilic geophilic fungi (mostly Microsporum gypseum /A. incurvatum) were recovered from 43 soil samples belonging to 23 out of 26 parks, along with Microsporum canis in a single case. Prototheca spp. was never isolated from water samples, while Trichosporon sp. was cultured in two cases, alone and in association with Geotrichum candidum. These results show that dogs did not act as important carriers for the investigated bacterial and parasitic pathogens, although examined areas may represent a risk for the spreading of some dermatophytoses to both pets and their owners. Periodical examinations to assess the main bacteriological, parasitological and mycological pathogens in different samples collected in off-leash dog parks should be carried out in a One-Health perspective

Larval ascaridoid nematodes in horned and musky octopus (Eledone cirrhosa and E. moschata) and longfin inshore squid (Doryteuthis pealeii): Safety and quality implications for cephalopod products sold as fresh on the Italian market

The aim of this study was to evaluate the occurrence, infection level and distribution of ascaridoid larvae in cephalopod products sold in Italy. Data on the species most commonly commercialized as whole and fresh on the Italian market were collected. After comparing commercial and literature data, Eledone spp., comprising E. cirrhosa and E. moschata (horned octopus and musky octopus, respectively) and Doryteuthis pealeii (longfin inshore squid) were selected, as they had been rarely investigated. Overall, 75 Eledone spp. caught in the Mediterranean Sea (FAO area 37) and 70 D. pealeii from the Northwest Atlantic Ocean (FAO area 21) were examined by visual inspection and artificial digestion (viscera and mantle separately). Parasites were submitted to morphological and molecular analysis. Prevalence (P), mean intensity (MI) and mean abundance (MA) were calculated. In D. pealeii, 2 nematode larvae molecularly identified as Anisakis simplex s.s. were found in the viscera and in the mantle of two specimens (P: 2.9% 95% CI: 0–6.8%; MI: 1; MA: 0.028). In Eledone spp. 9 nematode larvae molecularly attributed to Hysterothylacium spp. were found in the mantle of 5 specimens (P: 6.7% 95% CI: 1–12.3%; MI: 1.8; MA: 0.12). This is the first report of A. simplex s.s. in D. pealeii. Considering the zoonotic and allergenic potential of these larvae and their localization also in the edible part (mantle), a potential public health issue exists

DNA barcoding reveals chaotic labeling and misrepresentation of cod (Xue) products sold on the Chinese market

The increasing rate of seafood frauds, especially in the case of highly priced species, highlights the need of verifying the identity of fish products. This paper describes the application of DNA barcoding to the identification of 52 products commercialized with the Chinese term 鳕 (Xue, Cod) in supermarkets (Nanjing and Shanghai) and in the online market. Considering the lack of harmonization around the definition of Cod, the mislabeling rate was assessed according to three increasingly stringent definitions: Cod meaning Gadiformes species; Cod meaning Gadus spp.; Cod not meaning any specific species, since a qualifier (“Atlantic”, “Pacific” or “Greenland”) should be added in order to refer to Gadus morhua, Gadus macrocephalus or Gadus ogac, respectively. Results highlighted a very high mislabeling rate, which exceeded 60% even with the less stringent definition. Interestingly, only 42.3% of the samples were Gadiformes, while the others were Perciformes, Pleuronectiformes or toxic Tetraodontiformes species. Economic, ecological and health issues arising from the misuse of the term Cod are discussed in the light of the leading role of China in the seafood worldwide industry and of the increased national consumption of marine species