Zoonotic Threats: The (Re)emergence of Cercarial Dermatitis, Its Dynamics, and Impact in Europe.

Cercarial dermatitis (CD), or "Swimmer's itch" as it is also known, is a waterborne illness caused by a blood fluke from the family Schistosomatidae. It occurs when cercariae of trematode species that do not have humans as their definitive host accidentally penetrate human skin (in an aquatic environment) and trigger allergic symptoms at the site of contact. It is an emerging zoonosis that occurs through water and is often overlooked during differential diagnosis. Some of the factors contributing to the emergence of diseases like CD are related to global warming, which brings about climate change, water eutrophication, the colonization of ponds by snails susceptible to the parasite, and sunlight exposure in the summer, associated with migratory bird routes. Therefore, with the increase in tourism, especially at fluvial beaches, it is relevant to analyze the current epidemiological scenario of CD in European countries and the potential regions at risk.

Vernonia britteniana Root Phytochemical Studies, In Vitro Cercaricidal Activity on the Larval Stage of Schistosoma mansoni and Antioxidant Activities.

Vernonia britteniana Hiern. (Asteraceae) is a medicinal plant used in traditional Angolan medicine against schistosomiasis. Our study aimed to investigate the phytochemical composition and the cercaricidal and antioxidant activities in vitro of a traditional herbal preparation (Water-Vbr) and a 70% hydroethanolic extract (EtOH70%-Vbr) prepared with this medicinal plant. The activity of the extracts against Schistosoma mansoni cercariae was assessed at different extract concentrations (500, 438, and 125 µg/mL) and at different time intervals, and the phytochemical profiles were obtained by LC-UV-ESI/MS-MS. In addition, the major chemical classes of the identified metabolites were quantified by colorimetry, and the antioxidant potential was assessed using the DPPH and FRAP methods. After 30 min, 100% cercarial mortality was observed at a concentration of 500 µg/mL after exposure, and after 120 min, an LC50 of 438 µg/mL was observed for both extracts. Phenolic acid derivatives (chlorogenic acid, caffeic acid; 3,4-di-O-caffeoylquinic acid; 3,5-di-O-caffeoylquinic acid; and 4,5-di-O-caffeoylquinic acid) and triterpenoids (stigmastane-type steroidal saponins; vernoamyoside D and vernonioside D1; vernoamyoside B; and vernoniamyoside A and C) were identified as the main secondary metabolites. The Water-Vbr extract showed the highest antioxidant activity-DPPH: IC50 = 1.769 ± 0.049 µg/mL; FRAP: mean = 320.80 ± 5.1325 µgAAE/g.

Human Toxocariasis in Portugal-An Overview of a Neglected Zoonosis over the Last Decade (2010-2020).

Toxocariasis is one of the most widespread and important zoonotic parasitic diseases, although neglected. Data regarding human Toxocara infection in Portugal are almost absent. This article gives an overview of the situation of toxocariasis in Portugal over the last decade based on casuistic data. A total of 846 serum samples from individuals suspected of toxocariasis, collected from 2010 to 2020, were analyzed at the Institute of Hygiene and Tropical Medicine. Sera were tested for IgG antibodies to Toxocara canis excreted-secreted larval antigens by enzyme-linked immunosorbent assay and counterimmunoelectrophoresis. Positivity was detected in 18.8% (159/846) [CI 95%: 16.3-21.6], with positives detected throughout continental Portugal. Overall, 59.7% of the positives were diagnosed in younger than 20 years (35.2% aged 0-9 years and 24.5% aged 10-19 years). Eosinophilia was the most frequent feature reported (27.7%). Pediatrics (41.5%) and Infectiology (25.8%) were the specialties with the highest number of positives. An average of 77 samples/year were received, recording a maximum positivity in 2012 (41.5%, n = 27/65) and a minimum in 2020 (6.4%, n = 3/47). These numbers may reflect the effectiveness of current preventive measures, highlighting the need to maintain public awareness to control this helminthozoonosis and promote a higher public health standard.

Dirofilaria immitis and Angiostrongylus vasorum: The current situation of two major canine heartworms in Portugal.

Cardiopulmonary nematodes are life-threatening pet parasites increasingly reported throughout Europe, with overlapping endemic areas. Dirofilaria immitis is a mosquito-borne whilst Angiostrongylus vasorum is a snail-borne pathogen. Both adult nematodes reside in the pulmonary arteries and right cardiac ventricle of domestic and wild canids, causing a wide spectrum of clinical presentations ranging from cough, dyspnoea and exercise intolerance to severe vascular and pulmonary disease with hearth failure that may lead to death. Information about the prevalence and distribution of cardiopulmonary parasites is essential for the control of animal diseases and, in the case of D. immitis, for the control of potentially associated illnesses in humans. However, in Portugal, heartworm studies are limited to few surveys and case reports, possibly underestimating the relevance of these nematodes. The present work reviews the data on cardiopulmonary dirofilariosis and angiostrongylosis in dogs in Portugal, providing a comprehensive update of the epidemiological situation during the past 20 years.

Human gastric hyperinfection by Anisakis simplex: A severe and unusual presentation and a brief review.

Anisakiasis is an emerging marine food-borne zoonosis resulting from the accidental ingestion of Anisakis larvae, through the consumption of raw or undercooked infected seafood products. The first case of human gastric hyperinfection by Anisakis simplex with an unusual and severe presentation, occurring in a Portuguese woman, is described in this article. Over 140 anisakid larvae were removed by gastroscopy. Massive infection is uncommon in areas where the consumption of raw fish is not part of the traditional diet, as is the case in Portugal. The increased consumption of raw seafood products is considered a health determinant in the rise in cases of anisakiasis. However, clinicians should be aware of the emergence of these infections, not only because of the new dietary habits of the population, but also because of the high prevalence of Anisakis larvae in the different fish species usually consumed by the population, collected on the Portuguese coast.

Molecular characterization of Dirofilaria spp. circulating in Portugal.

BACKGROUND: Dirofilariosis is a potentially zoonotic parasitic disease, mainly transmitted by mosquito vectors in many parts of the world. Data concerning the canine Dirofilaria species currently circulating in Portugal is scarce. Thereby, a large-scale study was conducted to determine the Dirofilaria spp. present in Portugal, based on a molecular approach, and also to optimize a reliable and highly sensitive species-specific polymerase chain reaction (PCR) assay that could be used for the simultaneous detection and differentiation of Dirofilaria immitis, Dirofilaria repens, and other concurrent filarial species in animal reservoirs. METHODS: Blood samples were collected from three districts of Portugal (Coimbra, Santarém and Setúbal) between 2011 and 2013. Samples were tested using rapid immunomigration tests (Witness® Dirofilaria), modified Knott's technique and acid phosphatase histochemical staining. In addition, molecular analysis was performed by amplification of the internal transcribed spacer (ITS) region using two different PCR protocols, specific for molecular screening of canine filarial species. RESULTS: Of the 878 dogs sampled, 8.8% (n = 77) were positive for D. immitis circulating antigen and 13.1% (n = 115) positive for microfilariae by the modified Knott's technique. Of the 134 samples tested by acid phosphatase histochemical staining, 100 (74.6%) were positive for D. immitis. Overall, 13.7% (n = 120) were positive by PCR for D. immitis by ITS2, of which 9.3% (67/720) were also positive by ITS1. ITS2 PCR was the most sensitive and specific method, capable of detecting mixed D. immitis and A. reconditum infections. Heterozygosity, in the form of double peaks, was detected by sequencing of both ITS regions. No D. repens was detected by any of the diagnostic methods. CONCLUSIONS: The present study confirmed D. immitis as the dominant species of the genus Dirofilaria infecting Portuguese dogs, based on sequencing of ITS1 and ITS2 PCR fragments. Additionally, ITS2 PCR was the most adequate method for diagnosis and prevalence estimation.

Curcumin Generates Oxidative Stress and Induces Apoptosis in Adult Schistosoma mansoni Worms.

Inducing apoptosis is an interesting therapeutic approach to develop drugs that act against helminthic parasites. Researchers have investigated how curcumin (CUR), a biologically active compound extracted from rhizomes of Curcuma longa, affects Schistosoma mansoni and several cancer cell lines. This study evaluates how CUR influences the induction of apoptosis and oxidative stress in couples of adult S. mansoni worms. CUR decreased the viability of adult worms and killed them. The tegument of the parasite suffered morphological changes, the mitochondria underwent alterations, and chromatin condensed. Different apoptotic parameters were determined in an attempt to understand how CUR affected adult S. mansoni worms. CUR induced DNA damage and fragmentation and increased the expression of SmCASP3/7 transcripts and the activity of Caspase 3 in female and male worms. However, CUR did not intensify the activity of Caspase 8 in female or male worms. Evaluation of the superoxide anion and different antioxidant enzymes helped to explore the mechanism of parasite death further. The level of superoxide anion and the activity of Superoxide Dismutase (SOD) increased, whereas the activity of Glutathione-S-Transferase (GST), Glutathione reductase (GR), and Glutathione peroxidase (GPX) decreased, which culminated in the oxidation of proteins in adult female and male worms incubated with CUR. In conclusion, CUR generated oxidative stress followed by apoptotic-like-events in both adult female and male S. mansoni worms, ultimately killing them.

Morphological Characteristics of Schistosoma mansoni PZQ-Resistant and -Susceptible Strains Are Different in Presence of Praziquantel.

Schistosomiasis is one of the most common human parasitic diseases whose socioeconomic impact is only surpassed by malaria. Praziquantel (PZQ) is the only drug commercially available for the treatment of all schistosome species causing disease in humans. However, there has been stronger evidences of PZQ-resistance on Schistosoma mansoni and thus it is very important to study the phenotypic characteristics associated with it. The aim of this study was to evaluate morphological alterations in S. mansoni PZQ-resistant adult worms and eggs, by comparing a PZQ- resistant strain obtained under PZQ drug pressure with a PZQ-susceptible strain. For this, scanning electronic microscopy was used to assess tegumental responsiveness of both strains under PZQ exposure, and optical microscopy allowed the monitoring of worms and eggs in the presence of the drug. Those assays showed that PZQ-susceptible worms exposed to the drug had more severe tegumental damages than the resistant one, which had only minor alterations. Moreover, contrary to what occurred in the susceptible strain, resistant worms were viable after PZQ exposure and gradually regaining full motility after removal of the drug. Eggs from resistant strain parasites are considerably smaller than those from susceptible strain. Our results suggest that there might be a difference in the tegument composition of the resistant strain and that worms are less responsive to PZQ. Changes observed in egg morphology might imply alterations in the biology of schistosomes associated to PZQ-resistance, which could impact on transmission and pathology of the disease. Moreover, we propose a hypothetical scenario where there is a different egg tropism of the S. mansoni resistant strain. This study is the first comparing two strains that only differ in their resistance characteristics, which makes it a relevant step in the search for resistance determinants.

Seroprevalence of circulating Angiostrongylus vasorum antigen and parasite-specific antibodies in dogs from Portugal.

Angiostrongylus vasorum is a nematode that lives in the pulmonary arteries and right cardiac ventricle of domestic dogs and wild canids. It is increasingly being reported in several European countries and North America. This parasite induces inflammatory verminous pneumonia, causing severe respiratory disease in dogs. In some instances, coagulopathies, neurological signs and even death may occur. Scant data are available regarding the occurrence of A. vasorum in Portugal. Therefore, sera of 906 shelter dogs from North to South mainland Portugal were collected. ELISAs to detect A. vasorum circulating antigen and specific antibodies against this parasite were performed. A total of six dogs [0.66 %, 95 % confidence intervals (CI) 0.24-1.43] were positive for both A. vasorum antigen and antibody detection, indicating an active infection, and 12 dogs (1.32 %, CI 0.68-2.30) were A. vasorum antibody-positive only. Regions with antigen- and antibody-positive animals overlapped and were distributed over nearly all sampled areas in the country. This is the first large-scale ELISA-based serological survey for A. vasorum in dogs from Portugal. The endemic occurrence of A. vasorum in dogs from different geographical areas of Portugal is therefore confirmed.

The Role of Efflux Pumps in Schistosoma mansoni Praziquantel Resistant Phenotype.

BACKGROUND: Schistosomiasis is a neglected disease caused by a trematode of the genus Schistosoma that is second only to malaria in public health significance in Africa, South America, and Asia. Praziquantel (PZQ) is the drug of choice to treat this disease due to its high cure rates and no significant side effects. However, in the last years increasingly cases of tolerance to PZQ have been reported, which has caused growing concerns regarding the emergency of resistance to this drug. METHODOLOGY/PRINCIPAL FINDINGS: Here we describe the selection of a parasitic strain that has a stable resistance phenotype to PZQ. It has been reported that drug resistance in helminths might involve efflux pumps such as members of ATP-binding cassette transport proteins, including P-glycoprotein and multidrug resistance-associated protein families. Here we evaluate the role of efflux pumps in Schistosoma mansoni resistance to PZQ, by comparing the efflux pumps activity in susceptible and resistant strains. The evaluation of the efflux activity was performed by an ethidium bromide accumulation assay in presence and absence of Verapamil. The role of efflux pumps in resistance to PZQ was further investigated comparing the response of susceptible and resistant parasites in the absence and presence of different doses of Verapamil, in an ex vivo assay, and these results were further reinforced through the comparison of the expression levels of SmMDR2 RNA by RT-PCR. CONCLUSIONS/SIGNIFICANCE: This work strongly suggests the involvement of Pgp-like transporters SMDR2 in Praziquantel drug resistance in S. mansoni. Low doses of Verapamil successfully reverted drug resistance. Our results might give an indication that a combination therapy with PZQ and natural or synthetic Pgp modulators can be an effective strategy for the treatment of confirmed cases of resistance to PZQ in S. mansoni.

First molecular identification of mosquito vectors of Dirofilaria immitis in continental Portugal.

BACKGROUND: Canine dirofilariasis due to Dirofilaria immitis is known to be endemic in continental Portugal. However, information about the transmitting mosquito species is still scarce, with only Culex theileri identified to date, albeit with L1-2, through dissection. This study was carried out to investigate the potential vectors of Dirofilaria spp. in continental Portugal. METHODS: Mosquitoes were collected in three distinct seasons (Summer, Autumn and Spring), 2011-2013, in three districts. CDC traps and indoor resting collections were carried out in the vicinity of kennels. Mosquitoes were kept under controlled conditions for 7 days to allow the development of larval stages of Dirofilaria spp.. DNA extraction was performed separately for both head+thorax and abdomen in order to differentiate infective and infected specimens, respectively, in pools, grouped according to the species and collection site (1-40 specimen parts/pool), and examined by PCR using pan-filarial specific primers. Mosquito densities were compared using non-parametric tests. Dirofilaria development units (DDU) were estimated. RESULTS: In total, 9156 female mosquitoes, from 11 different species, were captured. Mosquito densities varied among the 3 districts, according to capture method, and were generally higher in the second year of collections. From 5866 specimens screened by PCR, 23 head+thorax and 41 abdomens pools, corresponding to 54 mosquitoes were found positive for D. immitis DNA. These belonged to 5 species: Culex (Cux) theileri (estimated rate of infection (ERI)=0.71%), Cx. (Cux) pipiens f. pipiens and f. molestus (ERI=0.5%), Anopheles (Ano) maculipennis s.l. (ERI=3.12%), including An. (Ano) atroparvus, Aedes (Och) caspius (ERI=3.73%) and Ae. (Och) detritus s.l. (ERI=4.39%). All but Cx. pipiens, had at least one infective specimen. No D. repens infected specimens were found. Infection rates were: 3.21% in Coimbra, 1.22% in Setúbal and 0.54% in Santarém. DDU were at least 117/year in the study period. CONCLUSIONS: Culex theileri, Cx. pipiens, An. maculipennis s.l. An. atroparvus, Ae.caspius and Ae. detritus s.l. were identified as potential vectors of D. immitis in three districts of Portugal, from Spring to Autumn, in 5 of the 6 collection dates in 2011-2013. Implications for transmission, in the context of climate changes, and need for prophylactic measures, are discussed.

Dirofilariasis by Dirofilaria repens: an imported case and a brief review.

Human infections caused by Dirofilaria repens, a cosmopolitan zoonotic parasitosis endemic in Southern and Eastern Europe and Asia still is an underdiagnosed infection due to parasite identification difficulties. Here, we report the first human case of subcutaneous dirofilariasis by D. repens diagnosed in Portugal. This was probably an imported case from India, as judged by epidemiological and clinical data. With this presentation we aim to alert clinicians for the emergence of vector-borne zoonoses associated with global warming and international travel. This case showed that differential diagnosis of D. repens in subcutaneous nodules is needed, in order to avoid further complications.

Prostate adenocarcinoma associated with prostatic infection due to Schistosoma haematobium. Case report and systematic review.

Schistosomiasis affects more than 240 million people worldwide, an infection which may cause urogenital manifestations including, among others, squamous bladder cancer and prostate involvement. We describe the first case of a prostate adenocarcinoma associated with prostatic Schistosoma haematobium infection occurring in Angola. Prostate carcinoma was suspected because of high levels of prostate-specific antigen. This observation prompted us to review the literature on schistosomiaisis with respect to genital pathology and prostate cancer. Described genital manifestations in men include funiculitis, epididymitis, granulomata of the seminal vesicles, testicular masses, and prostate lesions which may cause haematospermia and infertility. In contrast to bladder cancer, only 12 reports including the present case on 17 cases on prostate carcinoma associated with schistosomiasis have been published worldwide. The rarity of reports on prostate carcinoma associated with schistosomiasis is partly due to diagnostic constraints, and its incidence is underestimated. However, in emerging countries, the incidence of prostate cancer appears to increase mainly as a result of urbanization and improved access to health care where schistosomiasis prevalence is decreasing.

Prevalence and seasonal variations of canine dirofilariosis in Portugal.

Dirofilariosis is a severe vector-borne emergent disease that is spreading worldwide and becoming a serious threat to human and veterinary public health. Portugal, a Mediterranean country, has favorable climate conditions for mosquito development and survival. At present, accurate data on the prevalence and epidemiological pattern of dirofilariosis in Portugal is scarce and outdated. To study these trends, a project was developed to assess the current prevalence of Dirofilaria immitis infection in shelter dogs as well as the prevalence of Dirofilaria species present in three coastal regions in central Portugal: Coimbra, Santarém, and Setúbal. Blood samples were collected from 696 shelter dogs during three consecutive years: 2011, 2012, and 2013. A rapid immunomigration technique was performed to detect female D. immitis antigens. Concurrently, to detect and identify circulating microfilariae, a modified Knott's technique and acid phosphatase histochemical staining were also performed. Of the 696 dogs sampled, 105 were positive for D. immitis, with an overall prevalence of 15.1%. Forty of the 105 dogs were antigen negative but were positive for D. immitis microfilariae. Three animals were co-infected with D. immitis and Acanthocheilonema dracunculoides, and there was also one dog infected only with A. dracunculoides, all confirmed by polymerase chain reaction. The highest prevalence of canine heartworm during the 3-y period was in Setúbal (24.8%), followed by Coimbra (13.8%), and Santarém (13.2%), with significant inter-district differences. Our results demonstrate a higher prevalence of dirofilariosis compared with findings of previous studies and show an increasing rate of infection in the southern areas of Portugal attributed, at least in part, to bioclimatic and ecological factors. The present study updates the epidemiological situation and correlates the risk of dirofilariosis transmission within each region. These findings are highly relevant to both human and veterinary public health, contributing to the general awareness of pet owners and veterinarian practitioners and reinforcing the need for effective control measures against vectors and preventive therapy in companion animals.

[Schistosomiasis and acute appendicitis]. / Schistosomose e apendicite aguda.

Acute appendicitis associated to Schistosoma haematobium and S. mansoni infection has been found in patients submitted to urgent appendectomy at the Hospital Américo Boavida in Luanda. Due to the high prevalence and morbidity caused by schistosomiasis (or bilharziasis) in the country, we suspect that the involvement of Schistosoma infection on appendicular pathology could be very frequent, in particular for those individuals more exposed to the parasite transmission. We report two clinical cases of acute appendicitis whose surgical specimens of the appendix revealed S. haematobium and S. mansoni eggs in histological samples. The reported patients live in endemic areas and have been exposed to schistosome during childhood, which may explain the infection's chronicity. Information of these clinical cases could be relevant, particularly for surgery specialists and clinical pathologists, due to the possibility of finding more patients with concurrent appendicitis and schistosomiasis.

A ocorrência de casos de apendicite aguda causada por Schistosoma haematobium e S. mansoni é uma situação clínica que tem vindo a ser observada em doentes submetidos a apendicectomia na urgência do Hospital Américo Boavida em Luanda. Atendendo à prevalência e morbilidade da schistosomose (ou bilharziose) no país, admitimos que esta infeção parasitária seja um fator causal muito frequente de apendicite, particularmente nas populações mais expostas à transmissão. Descrevemos dois casos de apendicite aguda associada à presença de ovos destas espécies de Schistosoma atingindo a mucosa do apêndice visíveis por histologia, em doentes naturais de zonas endémicas de Angola, com exposição precoce aos parasitas, o que justifica a cronicidade da infeção. Consideramos que a divulgação destes casos será particularmente relevante para as equipas de cirurgia geral e de anatomopatologia, atendendo à necessidade de execução sistemática de exames histológicos em todas as peças operatórias relacionadas.

Unusual panuveitis in a child: toxocariasis associated with ocular myiasis.

Most panuveitis in children are caused by infectious agents. A detailed clinical history and clinical examination are helpful in the diagnosis, but specific techniques are sometimes required to identify the causing specimen. We report the first published case of panuveitis in a child caused by simultaneous ocular infection by Toxocara canis and a fly larva and the innovative use of immunodiffusion technique in the vitreous for the diagnosis.

Detection of Wolbachia in Dirofilaria infected dogs in Portugal.

Wolbachia pipiens, an intracellular endosymbiont bacteria of filarial nematodes, has been implicated in the pathogenesis of filarial diseases, in particular in heavy Dirofilaria spp. infections. Antibiotic therapy (doxycycline) against Wolbachia has been proven to be suitable adjunct therapy, prior to adulticide treatment of canine dirofilariosis. Despite its importance, investigation on the Wolbachia/Dirofilaria complex in Portugal had not been undertaken so far. This study reports the first detection of Wolbachia in Dirofilaria spp. infected dogs in the context of an ongoing epidemiological survey in central-south regions in the country. Wolbachia DNA was detected by PCR in 52.6% (20/38) of canine blood samples positive for Dirofilaria immitis based on parasitological (Knott's and Acid Phosphatase) and serological (Witness(®)Dirofilaria) methods. No Wolbachia DNA could be detected in samples from dogs with occult infections (parasite negative but antigen positive). The lack of Wolbachia detection in some microfilaremic dogs was somewhat unexpected and needs to be elucidated in further studies, as the presence or absence of these bacteria in association with microfilaria is of importance for veterinarians in the management and control of canine dirofilariosis.