Molecular Survey of Parasitic Contamination of Frozen Berries.

Berries represent healthy dietary options and contain bioactive compounds associated with a decreased risk of diseases. Despite representing healthy food choices, these products can be contaminated by pathogenic microorganisms, including parasites. Among foodborne parasites, Giardia duodenalis, Cryptosporidium parvum, Cyclospora cayetanensis, Toxoplasma gondii, and Echinococcus multilocularis are of significant public health importance and have been recently detected in fresh berries in Europe, including Italy. Berries can be purchased fresh or frozen, and it is worrying that even frozen berries could represent a risk for the consumer. In fact, several parasites can resist freezing temperatures and have been responsible for outbreaks of infection. The aim of this study was to investigate the presence of G. duodenalis, C. parvum, C. cayetanensis, T. gondii, and E. multilocularis in frozen berries with simplex and multiplex real-time PCR protocols. A total of 108 packages of mixed frozen berries were bought from supermarkets located in a south-eastern region of Italy. The samples were tested using two simplex real-time PCR protocols targeting C. parvum and G. duodenalis, respectively, and a multiplex real-time PCR targeting C. cayetanensis, T. gondii, and E. multilocularis. None of the investigated parasites were detected in the frozen berry samples tested. This research topic is still unexplored and of great current interest. These results represent a first attempt to investigate parasitic contamination of frozen berries sold on the Italian market, but further large-scale surveys are required.

Seroprevalence and risk factors of Toxoplasma gondii infection in goats from South Punjab Province, Pakistan.

Layyah District in South Punjab Province of Pakistan offers the most intensive caprine economy in the country; its Indus riverine and desert environment makes the area peculiar and worthy of specific investigations. This study aimed to investigate the prevalence of anti-Toxoplasma gondii (T. gondii) IgG-antibody in goats in serum samples and the potential risk factors. The prevalence of T. gondii infection was estimated using a two-stage sample design. All caprine farms in the study area were stratified by size, and from these 110 were randomly selected. Twelve goats (>1-year-old) were selected from each farm and a total of 1320 serum samples were collected and tested by ELISA. A questionnaire on the conditions and management practices of each farm was administered to 110 farmers. Four hundred and sixteen out of 1320 sera samples (31.5%) were found positive and 89% of the flock had at least one seropositive goat. The proportion of seropositive goats tested within each flock ranged from 8.3% to 83.3%. with several factors contributing to this heterogeneity. Goat age played a significant role in the presence of cats. Significant interactions were related to goat farms having floor of dirt and kitten presence. Moreover, age class, abortion history and water source supply were modulated by owner education levels. This is the first study to determine the prevalence of anti-T. gondii antibodies in goats sera in Layyah district and the largest carried out so far in Pakistan. The remarkable presence of T. gondii among goats in areas where goat farming plays a significant economic role may pose a production threat to the small-stock industry, as well as to public health and food safety. Therefore, investigations to identify high-risk goat populations are highly recommended in order to facilitate the implementation of local control strategies.

Dermanyssosis in the Urban Context: When the One Health Paradigm Is Put into Practice.

Poultry red mites (Dermanyssus gallinae) are primarily ectoparasites of laying hens but also parasitize synanthropic birds in urban contexts. This mite can occasionally attack mammals, including humans, and cause mild to severe dermatitis. Attacks by zoonotic Mesostigmata mites are currently an increasing but still neglected problem of urban life. The authors present two cases of dermanyssosis involving two health workers at a hospital, linked to air conditioning outdoor units colonized by pigeons. Videos that describe the environmental contamination by D. gallinae and show where the infestation originated are presented. In addition, the authors update the literature of all urban cases, which, to date, reports over 240 clinical cases, mostly in private homes but also in public buildings. Dermatitis due to these mites is often unrecognized and, therefore, misdiagnosed. This report describes how the two cases herein reported were rapidly resolved thanks to the close cooperation between veterinary parasitologists and allergologists. It is crucial to raise awareness of the problem among general practitioners and specialists. In addition, the authors suggest a reconsideration of urban architectural choices that increase the public health risk posed by dermanyssosis and other diseases related to synanthropic birds.

When there is no communication between urban planners and public health operators: urban Dermanyssus gallinae infestations in humans.

At the international level, it is necessary to apply urban health strategies that can integrate concrete actions to protect and promote health in urban and architectural planning. In cities, the "urban fauna" mostly consists of synanthropic birds (sparrows, starlings, swallows, martins, jackdaws, crows, hawks, gulls, pigeons) that have adapted to a continuous relationship with humans. These animals enrich the ecological network of biodiversity but also pose health problems. The most successful avian colonizers are pigeons (Columba livia), which proliferate due to the abundance of food available to them and the absence of predators. Pigeons may harbor several organisms that are pathogenic for humans, and among these the role of Dermanyssus gallinae should not be underestimated. In the absence of their preferred pigeon host, these mites will move from the nest to windowsills and window frames from which they attack humans. The Authors show that modern architectural design features in towns can favor the establishment and proliferation of pigeons, contributing to the public health risk for dermanyssosis or other diseases related to these birds. They describe an outbreak of dermanyssosis due to incorrect or unsuitable structural interventions, and highlight the need of re-thinking urban architectural choices in order to safeguard public health.

What's in a child's ear? A case of otomyiasis by Sarcophaga argyrostoma (Diptera, Sarcophagidae).

A clinical report of otomyiasis in a 1-year-old girl is reported. A III instar larva of Sarcophaga sp. was microscopically identified and Sarcophaga (Liopygia) argyrostoma (Diptera, Sarcophagidae) was suspected. A molecular method targeting a fragment of the cox1 gene was used to confirm the identity of the specimen. Although myiases are not frequent manifestations in otolaryngology, they should arouse the attention of doctors, social workers and parents dealing with disabled people, the elderly and children. This contribution also highlights the need of combining microscopy and molecular tools to achieve a correct and reliable identification of the specimen/s.

Who Is Dermanyssus gallinae? Genetic Structure of Populations and Critical Synthesis of the Current Knowledge.

Despite the economic and animal welfare importance of the Poultry Red Mite Dermanyssus gallinae, its genetic structure has been studied in a scattered way so far. The prophylaxis and control of such a globally distributed ectoparasite can be significantly improved by understanding its genetic population structure (composition in species and intraspecific variants). The present study aims to establish a rigorous framework for characterizing the neutral genetic structure of D. gallinae based on a literature review combined with an integrative analysis of the data available in GenBank on population-level nucleotide sequence diversity supplemented by a new dataset. The integrative analysis was conducted on sequence data extracted from GenBank coupled with new sequences of two fragments of the mitochondrial gene encoding Cytochrome Oxidase I (CO1) as well as of an intron of the nuclear gene encoding Tropomyosin (Tpm) from several PRM populations sampled from European poultry farms. Emphasis was placed on using the mitochondrial gene encoding CO1 on which the main universal region of DNA barcoding in animals is located. The species D. gallinae sensu lato is a species complex, encompassing at least two cryptic species, i.e., not distinguishable by morphological characters: D. gallinae sensu stricto and D. gallinae L1. Only D. gallinae s.s. has been recorded among the populations sampled in poultry farms worldwide. Current knowledge suggests they are structured in three mitochondrial groups (haplogroups A, B, and C). Haplogroup A is cosmopolitan, and the other two present slightly contrasted distributions (B rather in the northern part of Europe, C most frequently found in the southern part). Recent data indicate that a dynamic geographic expansion of haplogroup C is underway in Europe. Our results also show that NUMT (nuclear mitochondrial DNA) pseudogenes have generated artifactual groups (haplogroups E and F). It is important to exclude these artifact groups from future analyses to avoid confusion. We provide an operational framework that will promote consistency in the analysis of subsequent results using the CO1 fragment and recommendations for future analyses.

Detection of Zoonotic Cryptosporidium ubiquitum in Alpine Wild Ruminants.

Cryptosporidium is a widespread apicomplexan protozoan of major zoonotic importance, characterized by a wide host range, and with relevant economic implications and potential negative effects on livestock and wildlife population dynamics. Considering the recent strong demographic increase of alpine ungulates, in this study, carried out in the Italian Northwestern Alps, we investigated the occurrence of Cryptosporidium spp. in these species and their potential involvement in environmental contamination with Cryptosporidium spp. oocysts. The immune-enzymatic approach revealed a Cryptosporidium prevalence of 1.7% (5/293), 0.5% (1/196) and 3.4% (4/119) in alpine chamois (Rupicapra rupicapra), red deer (Cervus elaphus) and roe deer (Capreolus capreolus), respectively. Positive samples were subjected to polymerase chain reaction (PCR) amplification for the COWP and gp60 genes. The presence of Cryptosporidium was confirmed in one chamois and four roe deer. Sequences obtained clustered within Cryptosporidium ubiquitum, currently recognized as an emerging zoonotic species. This finding represents the first detection of zoonotic C. ubiquitum in chamois and in the Alpine environment. Despite the low observed prevalences, environmental contamination by oocysts could play a role as a potential source of infections for humans and livestock.

Dermanyssus gallinae in non-avian hosts: A case report in a dog and review of the literature.

Non-avian attacks of the worldwide distributed mite Dermanyssus gallinae are occasionally reported. However, it is widely accepted that their occurrence is underestimated. The present study aims to describe the first Italian case of dermanyssosis in a dog, to molecularly characterize the mites collected from the patient and the animal enclosure, where poultry and dog were confined, and to review the current literature on the non-avian attacks by D. gallinae. The dog was successfully treated with an oral sarolaner-based product, followed by a spot-on formulation of imidacloprid and moxidectin. The infestation source was likely attributable to poultry and confirmed by molecular identification of D. gallinae sensu strictu. Ten articles on non-avian D. gallinae attacks in domestic animals and wildlife were retrieved, pointing out the need for more awareness amongst practitioners. The therapeutic effect of available antiparasitic drugs, currently used off-label, should also be better explored in non-avian hosts.

Contamination of fresh produce sold on the Italian market with Cyclospora cayetanensis and Echinococcus multilocularis.

To investigate the presence of Cyclospora cayetanensis, Toxoplasma gondii and Echinococcus spp. in fresh produce sold in Italy, 324 locally produced 'ready-to-eat' (RTE) mixed-salad packages belonging to three brands and 324 berries packages (blueberries and blackberries imported from Peru and Mexico, respectively, and raspberries grown in Italy) were purchased at retail. Nine individual packages from each of the six types of fresh produce were collected monthly for one year, and with the same produce pooled, this resulted in a total of 72 pools for the whole year. Using microscopy (FLOTAC), a Cyclospora-like oocyst was detected in a blueberry sample and a taeniid egg was detected in a RTE-salad sample. Molecular tools confirmed these to be C. cayetanensis and Echinococcus multilocularis, respectively. Toxoplasma gondii was not detected in any of the samples. This study shows for the first time in Europe that imported berries on the Italian market may be contaminated with C. cayetanensis and RTE salads grown in Italy with E. multilocularis. The results indicate a new epidemiological scenario and highlight that current management of fresh produce, locally produced or imported, does not ensure products are free from parasite contamination.

Cutaneous myiasis in cats and dogs: Cases, predisposing conditions and risk factors.

Two cases of cutaneous myiasis diagnosed in 2018 in Emilia-Romagna region (northern Italy) were reported. The first one, described in a domestic cat Felis silvestris catus L. (Carnivora: Felidae) and caused by Calliphora vicina Robineau-Desvoidy (Diptera: Calliphoridae), was the first one of this type ever reported in Italy in cats. The second one was described in a domestic dog Canis lupus familiaris L. (Carnivora: Canidae) and caused by Lucilia sericata (Meigen) (Diptera: Calliphoridae) and was unusual because it occurred in absence of lesions. An extensive literature search on cutaneous myiasis in these two domestic animal species was performed in order to draw attention to predisposing conditions and risk factors.

Diversity of Eimeria Species in Wild Chamois Rupicapra spp.: A Statistical Approach in Morphological Taxonomy.

Wildlife is frequently infected by intestinal protozoa, which may threaten their fitness and health. A diverse community of Eimeria species is known to occur in the digestive tract of mountain-dwelling ungulates, including chamois (genus Rupicapra). However, available data on Eimeria diversity in these taxa is at times inconsistent and mostly dated. In the present study, we aimed to revisit the occurrence of Eimeria spp. in the Alpine subspecies of the Northern chamois (Rupicapra rupicapra rupicapra) and the Apennine subspecies of the Southern chamois (Rupicapra pyrenaica ornata) in Italy, using an integrated approach based on a hierarchical cluster analysis (HCPC) applied to oocyst morphology and morphometry. A total of 352 fecal samples were collected from R. r. rupicapra (n = 262) and R. p. ornata (n = 90). Overall, 85.3% (300/352) of the animals tested microscopically positive to Eimeria spp. Based on morphological analysis, we identified all the eimerian species described in chamois. Through the HCPC method, five clusters were generated, corresponding to E. suppereri, E. yakimoffmatschoulskyi, E. riedmuelleri (two different clusters), and E. rupicaprae morphotypes. The well-defined clusters within E. riedmuelleri support the existence of two distinct morphological groups, possibly referable to different taxonomic units. This study suggests that combining a morphometrical approach with a powerful statistical method may be helpful to disentangle uncertainties in the morphology of Eimeria oocysts and to address taxonomic studies of eimeriid protozoa at a specific host taxon level.

Habronematidosis in Equids: Current Status, Advances, Future Challenges.

Over the past few decades, among equine parasitoses caused by gastrointestinal nematodes, habronematidosis has been discontinuously studied worldwide. Habronematidosis is a parasitic disease distributed all over the world. It is caused by Habronema microstoma, Habronema muscae, and Draschia megastoma (Spirurida, Habronematidae), and it is maintained in the environment by muscid flies which act as intermediate hosts. At larval and adult stages these species live in the stomach of domestic and wild equids. However, the larvae can also be found on the skin, causing lesions known as "summer sores", and occasionally on other body areas, such as ocular and genital mucosa (muco-cutaneous habronematidosis) and lung, liver, brain parenchyma. Depending on the parasite's developmental stage and localization site, clinical signs vary from mild to severe. Habronematidosis is responsible for significant economic losses, mostly when sport horses are affected, because their performances are impaired and the infection can be unaesthetic. We used three on-line databases for searching the articles on habronematidosis according to the selected inclusion criteria; a total of 250 contributions, published between 1911 and 2020 were analyzed. This review summarizes the key features of pathogenesis, epizootiology, diagnosis, treatment, and control of habronematidosis, and highlights the current knownledge about its geographical distribution and spread. Anthelmintic drugs are the most widely-used tools against habronematidosis; given the known risk of anthelmintic resistance in some nematodes affecting horses, this aspect should also be explored for habronematidosis. Dedicated research is essential to fill gaps of knowledge and increase the understanding of habronematidosis to maximize equine health, reduce economic losses and sanitary impact associated with this parasitic infection.

Urogenital myiasis caused by Lucilia sericata (Diptera: Calliphoridae) in a domestic rabbit in Italy.

The report describes a case of urogenital myiasis in a domestic rabbit Oryctolagus cuniculus L. (Lagomorpha: Leporidae) caused by Lucilia sericata (Meigen; Diptera: Calliphoridae) in region Emilia-Romagna (Northern Italy). The case, occurring in June 2018, is the first one involving L. sericata as an agent of myiasis in a domestic rabbit in Italy. Species identification was based on morphological investigations of males through identification keys. The rabbit developed the urogenital myiasis as a consequence of chronic enteritis causing an accumulation of faeces in the perianal and perineal region.

Asia and Europe: So Distant So Close? The Case of Lipoptena fortisetosa in Italy.

In Europe, 5 Lipoptena species have been recorded, including Lipoptena fortisetosa. This species, native to Asian countries, was described as a parasite of sika deer and its appearance in Europe dates back to more than 50 years ago. Lipoptena fortisetosa has been recently reported in Italy, sharing its hosts with Lipoptena cervi. A morpho-molecular approach was developed to determine the phylogenetic interrelationship of Italian and Asian CO1 haplotypes sequenced from Lipoptena fly individuals collected in Italy, and their DNA sequences were compared with conspecifics available in GenBank; morphological key-characters (terminalia) of L. fortisetosa were compared with the original description. Two haplotypes were recorded from Italy and assigned to L. cervi and L. fortisetosa, respectively. The latter was part of the monophyletic clade L. fortisetosa, along with 2 Central European and 2 Korean haplotypes (100% identical to one of the Korean haplotypes); moreover, Italian L. fortisetosa female terminalia were consistent with the original description of Asian individuals. Pending more in-depth investigations, this study provides a first answer to the hypothesis of the recent colonization of Italy by L. fortisetosa from Asia as we did not detect any obvious and stable morphological and molecular differences in specimens from the 2 geographical areas. The presence of the sika deer in Europe was retraced and the possible route traveled by the parasite from Asia and the eco-biological factors that may have enhanced its settlement are discussed.

Efficiency of the Q3 lab-on-chip Real Time-PCR platform for detecting protozoan pathogens in bivalve mollusks.

The zoonotic protozoan parasites Toxoplasma gondii, Cryptosporidium parvum and Giardia duodenalis have been recorded worldwide in economically important edible shellfish, and are thus likely to represent a significant public health risk. Therefore, an innovative, user-friendly diagnostic tool is required in order to improve food safety control. The Q3 system is a miniaturized platform whose efficiency and applicability were investigated and compared with results obtained using standard Real-Time PCR. Tanks of saltwater containing acclimated Mytilus galloprovincialis, Ruditapes philippinarum and Ostrea edulis specimens were spiked with purified Cryptosporidium, Giardia and Toxoplasma cysts/oocysts at different concentrations (i.e., 103, 104 and 105). We then collected 30 specimens for each shellfish species from each group at 24 h and 72 h post-contamination. After DNA extraction, we tested all samples by Real-Time-PCR and Q3, and evaluated the sensitivity, specificity, predictive values, repeatability and concordance between the two systems. Concordance between Real-Time-PCR and Q3 was very good (p < 0.01), especially for Toxoplasma in M. galloprovincialis at both 24 h and 72 h after contamination, and in O. edulis at 72 h. The ability of Q3 to detect all the investigated pathogens was similar to that of Real-Time-PCR, and Q3 was efficient in detecting Toxoplasma in both M. galloprovincialis and O. edulis. This is the first study concerning the use of lab-on-chip technology in a food matrix, and in edible marine mollusks in particular.

Efficacy of λ-cyhalothrin, amitraz, and phoxim against the poultry red mite Dermanyssus gallinae De Geer, 1778 (Mesostigmata: Dermanyssidae): an eight-year survey.

Dermanyssus gallinae (De Geer, 1778) is a major problem for the poultry industry worldwide, as it negatively affects virtually all kinds of rearing systems. Therefore, the control of infestation has become a routine process, and its economic cost is constantly increasing. Until now, most of the control strategies have relied on the use of synthetic chemical drugs, but their efficacy is often questioned by the emergence and diffusion of resistant mite populations. With this in mind, the efficacy of λ-cyhalothrin, amitraz, and phoxim has been verified by testing them against 86 mite populations collected from the same number of poultry farms in Italy from 2008 to 2015. Assays were performed according to the filter paper method using the recommended, half, quarter, double and quadruple doses. The results showed that phoxim and amitraz were the most effective acaricides (median efficacies 80.35% and 80.83%, respectively), but amitraz exhibited a sharp fall in its efficacy during 2011 and 2012, while phoxim maintained its high effectiveness up to 2015, when it dropped. The overall median efficacy of λ-cyhalothrin was 58.33%. The data also highlighted the importance of the use of the right concentration, as an increase in dosage was not always useful against resistant populations, while its reduction also diminished efficacy, simultaneously increasing the risk for the development of resistance.

Molecular epidemiology of Blastocystis sp. in dogs housed in Italian rescue shelters.

Blastocystis is a ubiquitous protozoan with a wide range of hosts. In humans, its presence has been associated with gastrointestinal disorders, although its role as a pathogen still needs to be elucidated. Until now, 17 Blastocystis subtypes (STs) have been identified, with ST1-ST4 the most commonly found in humans. Among domestic animals, the same STs reported in humans have been detected in dogs. An epidemiological survey on dog kennels was carried out to evaluate the prevalence of Blastocystis and the STs involved. Overall, 99 faecal samples were collected from the rescue shelters. Blastocystis detection was performed through conventional barcoding PCR targeting the 1800-bp SSU-rDNA, followed by sequencing and phylogenetic analysis. Blastocystis DNA was found in 21 faecal samples (21.2%), and all samples were successfully sequenced and identified as ST3 in a unique monophyletic group. The presence of Blastocystis was reported for the first time in dogs from Italy, with the identification of ST3, the subtype most commonly found in humans.

Dermanysuss gallinae attacks humans. Mind the gap!

Dermanyssus gallinae is a haematophagous ectoparasite primarily known as a pest of domestic and wild birds. It occasionally feeds on a range of mammals, and, more importantly, is of growing concern in human medicine. This review highlights mite attacks on people working with poultry, and updates the increasing incidence of dermanyssosis in urban environments in Europe. Although several cases of dermanyssosis have been documented, there are a number of reasons why diagnosis of D. gallinae infestations in humans is likely to be underestimated. Firstly, medical specialists are not well aware of D. gallinae infestations in humans. There is also a lack of collaboration with specialists from other disciplines. The problem is compounded by misdiagnoses and by the lack of diagnostic tools. We review the literature on human dermanyssosis cases in Europe, and also provide information on the epidemiology, clinical, histo-pathological and immunological aspects of dermanyssosis. We stress the need for improved recognition of this challenging infestation in humans, and provide straightforward recommendations for health practitioners, starting with collection of the correct anamnestic information and including appropriate management methods for case recognition and resolution. Finally, we indicate the most urgent areas to be addressed by future research. RESEARCH HIGHLIGHTS Dermanyssus gallinae is of growing concern in human medicine. Most physicians are not well aware of dermanyssosis in humans. Bio-epidemiological and clinical aspects of this ectoparasitosis are highlighted. Practical key actions for diagnosis and correct management of infestation in humans are provided.

Correction to: A formulation of neem and hypericum oily extract for the treatment of the wound myiasis by Wohlfahrtia magnifica in domestic animals.

The given names and family names of the authors of the originally published version of this article was inadvertently interchanged. Correct presentation of the author names are presented here.