The illegal rearing and slaughtering of pigs in the wild on the Mediterranean island of Sardinia favor an increase in the biomass of Trichinella britovi in wild boars (Sus scrofa) but do not affect the serological prevalence of infection.

BACKGROUND: Worms of the nematode genus Trichinella are zoonotic pathogens with a worldwide distribution. The first report of Trichinella on the Mediterranean island of Sardinia was for Trichinella britovi, one of the four species of this genus circulating in Europe, which was identified in 2005 following an outbreak of trichinellosis in humans due to the consumption of pork from pigs reared in the wild. Since then, T. britovi larvae have been repeatedly isolated from free-ranging pigs, foxes (Vulpes vulpes) and wild boars (Sus scrofa) sampled in the central-eastern region of the island (Orgosolo municipality), but have never been isolated from samples from other areas of the island. The aim of this study was to investigate the parasitological and serological prevalence of T. britovi infection in wild boars in Sardinia over space [eight wild boar hunting management units (HMUs)] and time (seven wild boar hunting seasons). METHODS: Muscle and serum samples of boars killed in the 2014-2015 to 2020-2121 hunting seasons were collected from eight HMUs of central and south-western Sardinia. Trichinella sp. larvae were detected by artificial digestion of predilection muscles. A total of 4111 serum samples of wild boar were collected from the investigated HMUs and tested by enzyme-linked immunosorbent assay as a screening test and by western blot as a confirmatory test using excretory/secretory antigens. RESULTS: Trichinella britovi muscle larvae were detected in six (0.03%) of the 17,786 wild boars tested. All of the Trichinella sp.-positive wild boars had been hunted in Orgosolo municipality (central-eastern area of the island), except for one, hunted in a neighboring municipality. An overall serological prevalence of 3.8% (95% confidence interval, 3.3-4.5) was detected by western blot. No statistical differences were detected between the HMUs where T. britovi larvae were detected in wild boars, foxes, and free-ranging pigs and those where wild boars, foxes and free-ranging pigs tested negative. CONCLUSIONS: The serological prevalence did not vary between the wild boar populations in which the larval load was detectable by artificial digestion (Orgosolo municipality) and those in which the larval load was below the detection limit. Furthermore, the serological prevalence of anti-Trichinella immunoglobulin G in the wild boar populations remained constant during the study period, which covered seven wild boar hunting seasons. As the transmission events (i.e., the serological prevalence) are stable, the high biomass of the parasite in Orgosolo municipality can only have arisen as a consequence of factors independent of its natural cycle, i.e., the presence of a high number of free-ranging pigs, and the concomitant presence of African swine fever, due to illegal pig slaughtering in the field. This epidemiological situation suggests that the natural cycle of T. britovi may be influenced by inappropriate pig husbandry and slaughtering practices.

An Unexpected Case of Opisthorchis felineus Infection Revealed during Liver Transplantation.

A man with hepatitis B infection was admitted to Pisa University Hospital for hepatological evaluation, which revealed multiple cystic lesions and suggested a cirrhotic evolution. Treatment with Entecavir 0.5 mg/day was started, resulting in rapid viral load suppression and alanine aminotransferase normalization. After 10 years, imaging documented a single nodule of hepatocellular carcinoma (HCC), and a robot-assisted nodule resection was performed. One year later, HCC recurrence prompted orthotopic liver transplantation, during which the patient died because of the sudden rupture of the donor's organ and rapid multiorgan deterioration before retransplantation. During post-mortem liver examination, adult worms were evidenced within large biliary ducts, suggesting infection with Opisthorchis or Clonorchis spp. flukes. Sequencing of the ITS2 locus, following PCR amplification of DNA extracted from liver tissue, revealed 100% identity with the reference sequence of O. felineus. Infection of the patient with O. felineus was confirmed by the presence of specific IgG detected by ELISA in the patient's sera. Two major alkaline phosphatase serum levels peaks observed during the first two years of antiviral therapy support the hypothesis that O. felineus infection worsened liver function. This case report highlights the importance of a very careful screening of parasitic infections in solid organ transplantation candidates.

Antigen discovery by bioinformatics analysis and peptide microarray for the diagnosis of cystic echinococcosis.

BACKGROUND: Cystic echinococcosis (CE), caused by Echinococcus granulosus sensu lato, is a neglected zoonosis. Its diagnosis relies on imaging, supported by serology, while only imaging is useful for staging and follow-up. Since diagnostic tools and expertise are not widely available, new accurate and easily implementable assays for the diagnosis and follow-up of CE are highly needed. METHODOLOGY/PRINCIPAL FINDINGS: We aimed to identify new E. granulosus antigens through a bioinformatics selection applied to the parasite genome, followed by peptide microarray screening and validation in ELISA, using independent panels of sera from patients with hepatic CE and clinically relevant controls. From 950 proteins selected in silico, 2,379 peptides were evaluated by microarray for IgG reactivity and eight candidates selected for validation. Reactivity to one peptide was significantly higher in the CE group (p = 0.044), but had suboptimal diagnostic accuracy. CONCLUSIONS/SIGNIFICANCE: Here we performed bioinformatics analysis and peptide microarray for antigen discovery, useful for the diagnosis of CE. Eight candidates were selected and validated. Reactivity to one peptide associated to CE but had suboptimal diagnostic accuracy. Importantly, the database developed in this study may be used to identify other antigenic candidates for CE diagnosis and follow-up.

Serological testing for Trichinella infection in animals and man: Current status and opportunities for advancements.

Serological tests are widely used for the detection of Trichinella spp. infections in animals and humans. Despite some limitations, (such as low sensitivity in the early period after infection) ELISA and western blot testing have demonstrated good performance when excretory/secretory products from muscle larvae are used as antigens in agreement with the International Commission on Trichinellosis. Over recent decades, considerable progress has been made in the characterization of Trichinella-derived molecules in the hope of improving diagnosis, mainly during the early days post infection. Despite these efforts, validated tests using characterized antigens for early diagnosis are still not available. However, combining currently available sero-diagnostic tools with clinical and epidemiological data provides valuable information on Trichinella infections in humans and animals as shown in this review.

Animal welfare and zoonosis risk: anti-Trichinella antibodies in breeding pigs farmed under controlled housing conditions.

BACKGROUND: Domesticated pigs are the main source of Trichinella sp. infections for humans, particularly when reared in backyards or free-ranging. In temperate areas of southern Europe, most pigs are farmed under controlled housing conditions, but sows and sometimes fattening pigs have access to outdoors to improve animal welfare. The aim of the present study was to investigate whether outdoor access of breeding pigs farmed under controlled housing conditions can represent a risk for Trichinella sp. transmission when the farm is located in an agricultural area interspersed with wooded areas and badlands, where Trichinella spp. could be present in wildlife. METHODS: Serum samples were collected from 63 breeding sows and one boar before and after their access to an open fenced area for 2 months and from 84 pigs that never had outdoor access. Samples were screened for anti-Trichinella antibodies by ELISA, and positive sera were confirmed using Western blot (Wb) excretory/secretory antigens. To detect Trichinella sp. larvae, muscle tissues from serologically positive and negative pigs were tested by artificial digestion. RESULTS: Thirteen (20.6%) sows and one boar tested positive with both ELISA and Wb. No larvae were detected in muscle samples of serologically positive and serologically negative pigs. Positive serum samples were then tested by Wb using crude worm extract as antigens. The Wb banding pattern displayed was that characteristic of encapsulated species (Trichinella spiralis or Trichinella britovi). CONCLUSIONS: The detection of anti-Trichinella antibodies without larvae in the pig muscles, supported by epidemiological data, suggests that pigs may have been exposed to T. britovi. This study stresses the importance of instigating monitoring systems at farm level to prevent Trichinella sp. transmission and to investigate, through a landscape parasitological study, the suitability of a site before the planting of a high containment level pig farm in which the sows can have outside access to improve their welfare during pregnancy.

Accuracy of an experimental whole-blood test for detecting reactivation of echinococcal cysts.

BACKGROUND: Cystic echinococcosis (CE) is a complex disease for which clear understanding of clinical manifestations is needed to avoid misdiagnosis, inappropriate treatment, and severe complications. We evaluated the accuracy of a whole-blood stimulation test based on Interleukin (IL)-4 detection in response to Antigen B (AgB) of Echinococcus granulosus sensu lato to discriminate cyst viability and detect cyst reactivation in patients with hepatic CE. METHODOLOGY/PRINCIPAL FINDINGS: Thirty patients with CE3b cysts and 37 patients with spontaneously-inactivated CE4-CE5 cysts were recruited (T0). After enrollment, 5 patients with CE3b cysts received albendazole, resulting in cyst solidification (CE4) in 4/5. Within a two-year follow-up, the whole-blood test was repeated at two time-points, in ≥14 (T1) and in ≥4 (T2) patients per group. IL-4 and a panel of other soluble factors were measured in the stimulated plasma. Baseline IL-4 levels were significantly higher in patients with CE3b compared to those with CE4 cysts (p = 0.006). Test accuracy for CE3b diagnosis had a sensitivity of 33-60% and a specificity of 76-95%, depending on the cut-off applied. Overall, IL-4 levels did not change significantly over time in either group; however, patients within the CE3b group showed a significant decrease of IL-1ra, IL-6, IL-8, G-CSF, IFN-γ, IP-10, MCP-1, MIP-1α, FGF at T1 compared to T0 (p≤0.042). CONCLUSIONS/SIGNIFICANCE: Whole-blood IL-4-response to AgB is significantly higher in patients with active compared to inactive CE but apparently not modulated over time after treatment. On the contrary, the levels of IL-1ra, IL-6, IL-8, G-CSF, IFN-γ, IP-10, MCP-1, MIP-1α, FGF significantly decreased in active CE during follow-up. Additional studies are needed to understand whether these findings might have a clinical significance for patients' follow-up.

Collaborative Studies for the Detection of Taenia spp. Infections in Humans within CYSTINET, the European Network on Taeniosis/Cysticercosis.

Laboratory tools for diagnosing taeniosis/cysticercosis in non-endemic countries are available; however, there is little data on their performance. To provide information on the sensitivity, specificity, and reproducibility of these tools, inter-laboratory studies were organized within the EU COST-Action CYSTINET (TD1302). Two serological and one coprological Ring Trials (RTs) were organized to test a panel of human-derived sera and stool samples using assays routinely conducted by the participating laboratories to detect Taenia spp. infections. Four Western blots (WBs) and five ELISAs were used by nine laboratories for cysticercosis diagnosis. In the first serological RT, the overall sensitivity was 67.6% (95% CI, 59.1-75.4), whereas specificity was 97% (95% CI, 89.8-99.6). WBs recorded the best accuracy. A second serological RT was organized, to assess the three tests most frequently used during the first RT. Two out of six laboratories performed all the three tests. The overall sensitivity and specificity were 52.8% (95% CI, 42.8-62.7) and 98.1% (95% CI, 93.2-99.7), respectively. Laboratory performance strongly affected test results. Twelve laboratories participated in the coprological RT using conventional microscopy and six laboratories used molecular assays. Traditional diagnosis by microscopy yielded better results than molecular diagnosis. This may have been influenced by the lack of standardization of molecular tests across participating laboratories.

Surveillance of foodborne parasitic diseases in Europe in a One Health approach.

In 2012, WHO/FAO ranked 24 foodborne parasites (FBP) using multicriteria decision analysis (MCDA) to provide risk assessors with a basis for prioritising control of highly ranked FBP on the global level. One conclusion was that ranking may differ substantially per region. In Europe, the same methodology was used to rank FBP of relevance for Europe. Of the 24 FBP, the top-five prioritised FBP were identified for Europe as Echinococcus multilocularis, Toxoplasma gondii, Trichinella spiralis, E. granulosus, and Cryptosporidium spp., all of which are zoonotic. The objective of the present study was to provide an overview of surveillance and reporting systems in Europe for these top five prioritised FBP in the human and animal populations, to identify gaps, and give recommendations for improvement. Information on the surveillance systems was collected from 35 European countries and analysed according to the five different regions. For most FBP, human surveillance is passive in most countries and regions in Europe and notification differs between countries and regions. Adequate surveillance programmes for these FBP are lacking, except for T. spiralis, which is notifiable in 34 countries with active surveillance in susceptible animals under EU directive. Although human and animal surveillance data are available for the five prioritised FBP, we identified a lack of consistency in surveillance and reporting requirements between national experts and European bodies. Recommendations for improved surveillance systems are discussed.

HERBIVORES AS ACCIDENTAL HOSTS FOR TRICHINELLA: SEARCH FOR EVIDENCE OF TRICHINELLA INFECTION AND EXPOSURE IN FREE-RANGING MOOSE (ALCES ALCES) IN A HIGHLY ENDEMIC SETTING.

Herbivores can be accidental hosts for the zoonotic nematode parasites Trichinella spp., which are endemic at high prevalence in wildlife in northeastern Europe. Using direct and indirect detection methods for Trichinella spp., we investigated samples from 463 wild moose (Alces alces) harvested by hunters in Estonia in 2015. A total of 460 moose were tested directly by artificial digestion of diaphragm muscle, 463 moose were tested indirectly by enzyme-linked immunosorbent assay (ELISA), and 34 moose also by western blot. Positive-control reference sera were from other host species. Nematode larvae were found in six muscle samples; five of which were pooled samples. None of the larvae were identified as Trichinella spp., based on their morphology and molecular analyses. Twelve moose (2.6%) were positive by ELISA, but none were positive by the western blot test. Trichinella spp. infection was not detected, but ELISA results may suggest Trichinella spp. exposure in a small proportion of moose in Estonia.

Glutathione-S-transferase omega 1 and nurse cell formation during experimental Trichinella infection.

The glutathione-S-transferases omega (GSTO) are multifunctional enzymes involved in cellular defense. During the nurse cell (NC) formation in Trichinella spiralis infection, the structural and regulatory genes of the skeletal muscle cell are downregulated and a new phenotype is acquired which advances parasite growth and survival. Previous studies showed that the GSTO1 is overexpressed in the NC during T. spiralis infection. To clarify the role of GSTO1 during NC formation, we evaluated the production of this enzyme by immunohistochemistry (IHC) in the diaphragms of mice experimentally infected with T. spiralis at 15, 28 and 60 days post infection (dpi); phosphorylation of Akt (p-Akt) and JNK1 (p-JNK1) were also evaluated. Furthermore, we evaluated the in vitro effects of T. spiralis excretory/secretory (ES) products from muscle larvae on specific functions (viability, proliferative response, apoptosis) in two cell lines (HeLa and U937), as well as its ability to induce GSTO1, p-AkT, p-ERK1/2 and p-JNK1. Results showed that GSTO1 was elevated in NC present in the diaphragms of T. spiralis experimentally infected mice at 15 dpi and progressively increased up to 60 dpi. The activation pattern of Akt in NC was similar to that of GSTO1, whereas JNK1 was never phosphorylated. ES induced a dose-dependent proliferative response in U937 cells, at 24 h and 48 h of treatment, but not in HeLa cells. However, after 72 h following treatment, significant cell death was observed in both cell lines at all doses. The apoptotic index (a.i.) was significantly higher than in untreated cells in both cell lines but only at the highest concentration of ES tested. Furthermore, Western Blots revealed that cells treated with ES for 24, 48 and 72 h, exhibited time-dependent overexpression of GSTO1, whereas p-Akt appeared only after 24 h of treatment. The p-ERK-1/2 peaked at 24 h then declined at 48 h and 72 h after treatment; however, it remained significantly higher than in untreated cells. No changes were observed in p-JNK1 at 24 and 48 h after treatment but a sharp increase in p-JNK1 was observed at 72 h. Also in HeLa cells, ES induced a small but significant increase in GSTO1 expression after 24 and 48 h of treatment where p-JNK1 was present only after 72 h of treatment. In conclusion, T. spiralis ES can reproduce in vitro the modifications observed inside the NC during experimental infection in mice.

Second outbreak of Trichinella pseudospiralis in Europe: clinical patterns, epidemiological investigation and identification of the etiological agent based on the western blot patterns of the patients' serum.

Trichinellosis is a zoonotic disease due to the ingestion of raw or undercooked meat from animals infected with the larvae of nematodes belonging to the genus Trichinella. In January-February 2015, an outbreak of trichinellosis occurred in Genoa, Northern Italy. The epidemiological link was traced back to a dinner served at an agritourism farm on 31 December 2014, where a majority of the 52 guests had consumed the 'beef' steak tartare. The source of infection was not traced; however, it was noted that the amount of beef purchased officially for providing at the dinner did not correspond with that served, suggesting that meat of a different origin had been added to the beef to prepare the steak tartare. Clinical and laboratory data of 30 individuals out of the 52 (57.7%), of which four were hospitalized, were consistent with that of the case definition of trichinellosis. Western blot patterns of the sera from patients with confirmed trichinellosis were similar to the diagnostic pattern identified for the reference sera of Trichinella pseudospiralis but different from those of the control sera tested for patients infected with Trichinella spiralis and Trichinella britovi. Identification of T. pseudospiralis as the aetiological agent responsible for the outbreak of trichinellosis using an indirect tool represents an advancement in the epidemiological investigation of this zoonotic disease.

Differences in larval survival and IgG response patterns in long-lasting infections by Trichinella spiralis, Trichinella britovi and Trichinella pseudospiralis in pigs.

BACKGROUND: Domesticated and wild swine play an important role as reservoir hosts of Trichinella spp. and a source of infection for humans. Little is known about the survival of Trichinella larvae in muscles and the duration of anti-Trichinella antibodies in pigs with long-lasting infections. METHODS: Sixty pigs were divided into three groups of 20 animals and infected with 10,000 larvae of Trichinella spiralis, Trichinella britovi or Trichinella pseudospiralis. Four pigs from each group were sacrificed at 2, 6, 12, 18 and 24 months post-infection (p.i.) and the number of larvae per gram (LPG) of muscles was calculated. Serum samples were tested by ELISA and western blot using excretory/secretory (ES) and crude antigens. RESULTS: Trichinella spiralis showed the highest infectivity and immunogenicity in pigs and larvae survived in pig muscles for up to 2 years p.i. In these pigs, the IgG level significantly increased at 30 days p.i. and reached a peak at about 60 days p.i., remaining stable until the end of the experiment. In T. britovi-infected pigs, LPG was about 70 times lower than for T. spiralis at 2 months p.i. and only very few infecting larvae were detected at 6 months p.i., whereas no larvae were detected at 12, 18 and 24 months p.i. At 6 months p.i., degenerated/calcified larvae and cysts were detected in the muscles by trichinoscopy and histology. The IgG pattern showed by T. britovi-infected pigs was similar to that of T. spiralis-infected pigs, although seroconversion occurred some days later. The larval burden of T. pseudospiralis was slightly greater than for T. britovi at 2 months p.i., but no larvae were detected at 6 and 12 months p.i. In T. pseudospiralis-infected pigs, seroconversion occurred slowly, as in T. britovi-infected pigs. The IgG level showed a significant drop at 6 months p.i. and declining to the cut-off value at 12 months p.i. CONCLUSIONS: The longer survival of T. spiralis in pigs in comparison with the other two species highlights its exceptional dissemination potential. These results provide an explanation of the controversial data collected by parasitological and serological tools in the course of epidemiological investigations.

Epidemiology and surveillance of human (neuro)cysticercosis in Europe: is enhanced surveillance required?

OBJECTIVES: To report on relevant national surveillance systems of (N)CC and taeniasis (the infection with the adult tapeworm) in the European Union/European Economic Area and to assess the magnitude of (N)CC occurrence by retrieving information on cases for the period 2000-2016. METHODS: (N)CC cases were retrieved via national reporting systems, a systematic literature search, contact with clinicians and a search for relevant 'International Statistical Classification of Diseases and Related Health Problems' (ICD)-based data. RESULTS: Mandatory notification systems for (N)CC were found in Hungary, Iceland and Poland. Ten cases were reported in Poland and none in Hungary and Iceland. Through the systematic literature review and information given by clinicians, 263 individual and 721 aggregated (N)CC cases from 19 European countries were identified. ICD-based data were obtained from five countries. From 2000 to 2016, a total of 3489 cases (N)CC cases were coded: 832 in Italy, eight in Latvia, 357 in Portugal, 2116 in Spain and 176 in Sweden. CONCLUSION: Despite being classified as a possible eradicable disease, (N)CC is still diagnosed across Europe, yet its true extent and impact remain unclear.

OBJECTIFS: Rapporter sur les systèmes nationaux de surveillance pertinents de la (neuro)cysticercose (N)CC et de la téniase (infection par le ténia adulte) dans l'Union européenne/l'Espace économique européen, et évaluer l'ampleur de l'occurrence de la (N)CC en reprenant des informations sur les cas durant la période 2000-2016. MÉTHODES: Les cas de (N)CC ont été repris à partir des systèmes nationaux de notification, une recherche systématique de la littérature, des contacts avec des cliniciens et une recherche de données pertinentes basées sur la 'Classification Statistique Internationale des Maladies et Problèmes de Santé Connexes' (ICD). RÉSULTATS: Des systèmes de notification obligatoires pour la (N)CC ont été trouvés en Hongrie, en Islande et en Pologne. Dix cas ont été rapportés en Pologne et aucun en Hongrie et en Islande. Grâce à la revue systématique de la littérature et aux informations fournies par les cliniciens, 263 cas individuels et 721 cas agrégés de (N)CC de 19 pays européens ont été identifiés. Des données ICD ont été obtenues de cinq pays. De 2000 à 2016, un total de 3489 cas de (N)CC ont été codés: 832 en Italie, 8 en Lettonie, 357 au Portugal, 2116 en Espagne et 176 en Suède. CONCLUSION: Bien qu'elle soit classée comme une maladie pouvant être éradiquée, la (N)CC est toujours diagnostiquée à travers l'Europe, mais sa véritable ampleur et son impact restent incertains .

Anisakis Sensitization in the Croatian fish processing workers: Behavioral instead of occupational risk factors?

We undertook the first study systematically evaluating the risk of Anisakis-sensitization in Croatian fish-processing workers and potential genetic susceptibility to anisakiasis. Anti-Anisakis IgE seroprevalence and risk factors for 600 employees of Croatian fish processing facilities and 466 blood donor controls, were assessed by indirect ELISA targeted with: recombinant Ani s 1 and Ani s 7 allergens, an Anisakis crude extract, the commercial ImmunoCAP kit, and questionnaires. Genetic susceptibility to anisakiasis was evaluated by genotypisation of human leukocytes alleles (HLA). Anti-Anisakis seropositive and a fraction of negative subjects were also assessed by ELISA and Western Blot (WB) for IgG seroprevalence to Trichinella spp. Overall, the observed anti-Anisakis seroprevalence inferred by indirect ELISA was significantly higher in fish processing workers (1.8%, 95% CI 0.9-3.3%) compared to the controls (0%, 0-0.8%). Seven out of 11 Ani s 1 and Ani s 7-positives and none of selected 65 negative sera, tested positive on whole-Anisakis extract (ImmunoCAP), whereas Anisakis crude extract ELISA detected 3.9% (2.4-6.0%) seropositives in fish processing workers, three (14%) of which showed IgE reactivity to milk proteins. The highest risk associated with Anisakis-sensitization among workers was fishing in the free time, rather than any of attributes related to the occupational exposure. Although no association was observed between anti-Anisakis seropositivity and wearing gloves or protective goggles, the majority of workers (92%) wore protective gloves, minimizing the risk for Anisakis sensitization via skin contact. Six HLA alleles within DRB1 gene were significantly associated with seropositivity under dominant, allelic or recessive models. All sera confirmed negative for anti-Trichinella spp. IgG. The study exhaustively covered almost all marine fish processing workers in Croatia, reflecting real-time Anisakis sensitization status within the industry, already under the influence of wide array of allergens.

Species specificity preliminary evaluation of an IL-4-based test for the differential diagnosis of human echinococcosis.

The diagnosis of cystic echinococcosis (CE) is based on imaging, while serology is a complementary test of particular use when imaging is inconclusive. Serology has several limitations. Among them, false-positive results are often obtained in subjects with alveolar echinococcosis (AE), rendering difficult the differential diagnosis. We set up an immune assay based on IL-4-specific production after stimulating whole blood with an antigen B (AgB)-enriched fraction from E granulosus that associates with CE and CE cysts in active stage. We aimed to evaluate potential cross-reactivity of this test using samples from patients with AE. Twelve patients with AE were recruited; IL-4 levels ranged from 0 to 0.07 pg/mL. Based on the previously identified cut-off of 0.39 pg/mL using samples from patients with CE, none of samples from AE patients scored positive. In contrast, almost 80% of samples from AE patients scored positive in serology tests based on different E granulosus-derived antigenic preparations. Our preliminary data show that this experimental whole-blood assay has no cross-reactivity in our cohort of patients with AE, in turn indicating a high specificity of the assay for CE diagnosis. This result supports further work towards the development of improved diagnostic tests for CE.

Retrospective analysis of hospital discharge records for cases of trichinellosis does not allow evaluation of disease burden in Italy.

Human trichinellosis is a disease caused by nematode worms of the genus Trichinella. In Italy, as well as in most other European countries, notification of Trichinella infections in humans is mandatory; however, no information is available on the number of cases occurring annually. The aim of the present study was to retrospectively evaluate the burden of trichinellosis in Italy from 2005 to 2016. Hospital discharge records (HDRs) showing the code for trichinellosis (124) were registered and screened. Results were then compared with yearly reports issued by the Italian National Reference Laboratory for Trichinella (NRLT), with reports from the European Centre for Disease Prevention and Control (ECDC), and with literature data. A total of 102 HDRs revealed that the 124 code was erroneously reported in 72 (70.6%) records. Out of the 30 (29.4%) records with a correct diagnosis of trichinellosis, nine cases were reported by HDRs only, 21 cases were documented by both HDRs and the NRLT, whereas the NRLT documented 100 additional cases. In the studied period, the average yearly incidence was 0.01 cases per 100,000 inhabitants. This study highlights the limitations of using HDRs to obtain a clear picture of the prevalence and incidence of trichinellosis in Italy. These findings demonstrate the need to intensify the surveillance system for trichinellosis through the development of an Italian registry. This would allow the identification of patients with severe infections and pauci-symptomatic patients, and would avoid the need for clinical analyses and unnecessary treatments, reducing the resulting economic burden on the Italian National Health Service.

Delivery of SA35 and SA40 peptides in mice enhances humoral and cellular immune responses and confers protection against Cryptosporidium parvum infection.

BACKGROUND: Cryptosporidium parvum is a major cause of diarrhea in children and ruminants at the earliest stages of life. Maternal antibodies represent the main shield of neonate mammals for most of the infections. Two recombinant antigens (SA35 and SA40), portions of two C. parvum proteins, were tested for their ability to induce immune responses in adult mice and for protection on neonate BALB/c mice born from females immunised by mucosal delivery of both peptides. METHODS: Adult BALB/c mice were intraperitoneally immunised with SA35 and SA40, separately or mixed, and their immune response was characterised. Furthermore, BALB/c pregnant mice were immunised by mucosal delivery with an SA35/40 mix, before and during pregnancy. Soon after birth, their offspring were infected with two doses (1 × 105 and 5 × 103) of C. parvum oocysts and the parasitic burden was determined at 5 and 9 days post-infection. RESULTS: Intraperitoneal immunisation with SA35 and SA40 induced specific IgG and IgG1 in serum, specific IgA in the intestinal mucosa, increase of CD3+/CD4+ and CD30+ cells in splenocytes, which produced IFN-γ. Neonates born from immunised mice and infected with 1 × 105 oocysts showed a significant reduction of oocysts and intestinal forms (23 and 42%, respectively). A reduction of all parasitic forms (96%; P < 0.05) was observed when neonates were infected with 5 × 103 oocysts. CONCLUSIONS: SA35 and SA40 peptides induce specific humoral and cell-mediated immune responses to C. parvum in adult mice. Moreover, mucosal administration of the SA35/40 mix in pregnant mice reduces C. parvum burden in their litters.

Evaluation of a commercial enzyme-linked immunosorbent assay (ELISA) for detecting antibodies against Toxoplasma gondii from naturally and experimentally infected pigs.

BACKGROUND: Toxoplasmosis is one of the most frequent parasitic infections in animals causing reproductive disorders and thus notable economic losses in productivity. Among food animals, pigs along with sheep and goats possess the highest incidence of Toxoplasma gondii cysts in meat, and play a role as a source of human infection. METHODS: The commercial ELISA kit (PrioCHECK® Toxoplasma Ab SR, Prionics Schlieren-Zurich, Switzerland) for the detection of specific antibodies against T. gondii in swine serum was compared with a commercial IFAT (indirect fluorescent antibody test) (Toxo-Spot IF, bioMérieux, France), used as the reference test. RESULTS: The kappa value obtained comparing the results performed on sera by ELISA with the by IFAT was 1. By a receiver operating characteristics curve analysis, the commercial ELISA had a relative sensitivity of 100%, and a relative specificity of 100% respect to IFAT. CONCLUSIONS: The commercial ELISA showed a very good agreement with the commercial IFAT in the detection of serum antibodies to Toxoplasma in pigs. Our study confirmed the usefulness of the commercial ELISA kit for the detection of antibodies against T. gondii in pigs, representing a valuable tool to improve the diagnostic activity for T. gondii in swine populations at the farm level or at the slaughterhouse, contributing to the control of this widespread infection.

Comparative analysis of excretory-secretory antigens of Anisakis simplex, Pseudoterranova decipiens and Contracaecum osculatum regarding their applicability for specific serodiagnosis of human anisakidosis based on IgG-ELISA.

Serodiagnosis of human anisakidosis is presently hampered by the current lack of standardised serological assays that allow sensitive and specific detection of Anisakidae-specific antibodies in human patients. In the present study, we comparatively evaluated the diagnostic value (by IgG-ELISA) of excretory-secretory antigens (ESAgs) of Anisakis simplex, Pseudoterranova decipiens and Contracaecum osculatum, representing the most frequently found genera responsible for human infection. In addition, we tested also a mix of the three ES preparations (Mix-ESAgs) as well as two recombinant allergens of A. simplex, rAni s 1 and rAni s 7. ES antigen from C. osculatum yielded the best diagnostic performance in IgG-ELISA-based serodiagnosis of the Spanish anisakidosis patients investigated in this study (relative serodiagnostic sensitivity 100%; specificity 89%) as compared to A. simplex ES-antigen (93% versus 57%) and P. decipiens (67% versus 93%) or a mix of the three ES antigens (100% versus 44%), respectively. Cross-reactions of C. osculatum ES antigen with serum-antibodies from patients suffering from other helminth infections were rare and were exclusively found with few sera from toxocariasis, ascariasis, and filariasis patients. The two recombinant allergens rAni s 1 and rAni s 7 did not prove sufficiently sensitive and specific in order to justify a further evaluation of these antigens regarding their suitability in IgG-ELISA-based serodiagnosis of human anisakidosis. In conclusion, the C. osculatum-ESAg-ELISA remains as key candidate to be further assessed for the serodiagnosis of symptomatic anisakidosis in different endemic regions.