Re-emergence of human leishmaniasis in northern Italy, 2004 to 2022: a retrospective analysis.

BackgroundHuman leishmaniasis is a protozoan disease transmitted by sand flies and endemic in the Mediterranean region. In Italy, leishmaniasis is present in the south and the western coastal regions, with an epidemic peak detected in northern Italy in the early 1970s.AimTo examine temporal trends, and demographic, clinical, geographical and environmental features of human leishmaniasis cases recorded by the local health unit (LHU) of Bologna, northern Italy.MethodsIn this retrospective observational study, we analysed human leishmaniasis cases recorded from 2004 to 2022 within the Bologna LHU. We also conducted serological investigations for canine leishmaniasis in owned dogs living near the place of infection of human cases.ResultsIn total, 173 cases of human leishmaniasis were detected, and 154 cases were considered autochthonous. An increase of human cases was observed since 2004, with incidence peaks above 2 cases/100,000 inhabitants in 2013, 2018 and 2022; epidemic peaks were preceded by dry summers. Most cases lived in the plain and hilly areas less than 400 m above sea level and many resided in isolated housing, in city outskirts, and/or near uncultivated areas, watercourses and railway sections. The incidence of canine leishmaniasis did not increase in the study period.ConclusionAn epidemic of human leishmaniasis with fluctuating annual numbers of cases, probably related to environmental and climatic factors, was identified in the Bologna LHU. Understanding the risk factors and the environmental characteristics related to places of infection is crucial to evaluate the public health implications of leishmaniasis.

Eleven Years of Health Monitoring in Wild Boars (Sus scrofa) in the Emilia-Romagna Region (Italy).

In recent years, the growth of wild ungulates has increased the focus on their health monitoring. In particular, the health status of wild boars is relevant for the economic impact on the pig industry. The Emilia-Romagna region activated a wildlife monitoring plan to better evaluate the health status of the wild boar population. Between 2011 and 2021, samples of found dead and hunted wild boar have been examined for trichinellosis, tuberculosis, brucellosis, african swine fever, classical swine fever, Aujeszky's disease, swine vesicular disease, and swine influenza A. Trichinella britovi was identified in 0.001% of the examined wild boars; neither M. bovis nor M. tuberculosis were found in M. tuberculosis complex positive samples; 2.3% were positive for Brucella suis; 29.4% of the sera were positive for Aujeszky's disease virus; and 0.9% of the samples were positive for swine influenza A virus. With an uncertain population estimate, the number of animals tested, the number of positives, and the sampling method do not allow us to make many inferences but suggest the need to implement and strengthen the existing surveillance activity, as it seems to be the only viable alternative for safeguarding animal and human health.

Salmonella Infection in Pigs: Disease, Prevalence, and a Link between Swine and Human Health.

Salmonella is one of the most spread foodborne pathogens worldwide, and Salmonella infections in humans still represent a global health burden. The main source of Salmonella infections in humans is represented by contaminated animal-derived foodstuffs, with pork products being one of the most important players. Salmonella infection in swine is critical not only because it is one of the main causes of economic losses in the pork industry, but also because pigs can be infected by several Salmonella serovars, potentially contaminating the pig meat production chain and thus posing a significant threat to public health globally. As of now, in Europe and in the United States, swine-related Salmonella serovars, e.g., Salmonella Typhimurium and its monophasic variant Salmonella enterica subsp. enterica 1,4,[5],12:i:-, are also frequently associated with human salmonellosis cases. Moreover, multiple outbreaks have been reported in the last few decades which were triggered by the consumption of Salmonella-contaminated pig meat. Throughout the years, changes and evolution across the pork industry may have acted as triggers for new issues and obstacles hindering Salmonella control along the food chain. Gathered evidence reinforces the importance of coordinating control measures and harmonizing monitoring programs for the efficient control of Salmonella in swine. This is necessary in order to manage outbreaks of clinical disease in pigs and also to protect pork consumers by controlling Salmonella subclinical carriage and shedding. This review provides an update on Salmonella infection in pigs, with insights on Salmonella ecology, focusing mainly on Salmonella Choleraesuis, S. Typhimurium, and S. 1,4,[5],12:i:-, and their correlation to human salmonellosis cases. An update on surveillance methods for epidemiological purposes of Salmonella infection in pigs and humans, in a "One Health" approach, will also be reported.

Identification and phylogenetic analysis of Taenia spp. parasites found in wildlife in the Emilia-Romagna region, northern Italy (2017-2022).

The recent expansion of the habitat of several wildlife species, comprising anthropized areas, is a relevant risk factor for many zoonotic diseases and should be considered in national and regional sanitary monitoring systems. We evaluated adult intestinal Taenia spp. parasites isolated from wild carnivores and cystic larval forms isolated from wild mammals analysed at the Istituto Zooprofilattico Sperimentale della Lombardia ed Emilia-Romagna (IZSLER) as part of the regional wildlife sanitary surveillance plan. Then, we assessed parasite species through molecular analysis (multiplex PCR followed by ribosomal 12S subunit gene sequencing) in order to update the epidemiological situation on Taeniids infection in the Emilia-Romagna wildlife, reporting the prevalence of each isolated species. The most commonly isolated species was Taenia serialis, which we detected in both wolves and foxes as definitive hosts and in roe deer as intermediate host. More attention on the distribution of Taeniids in wildlife should be paid, considering their potential zoonotic role: several Taenia spp. (Taenia solium, Taenia multiceps, Taenia serialis, Taenia brauni, Taenia glomerulatus) are known for causing coenurosis in humans, with possible severe or fatal outcomes.

Microbiological and Chemical Analysis of Food Collected Under Official Control in the Emilia-Romagna Region of Northern Italy, 2014-2019.

This study analyzed data from 6 years (2014-2019) of official controls in the Emilia-Romagna region (northern Italy) to investigate the frequencies of human pathogens and chemical hazards in foods during production and distribution. Campylobacter spp. was the most prevalent pathogen, isolated in 4.4% of the 1,078 food samples examined, followed by Salmonella spp. (2.8%), Shiga toxin-producing Escherichia coli (STEC) (1.9%), and Listeria monocytogenes (0.9%). Salmonella serotyping showed that the isolates belonged to the serotypes most commonly isolated from humans in Emilia-Romagna. These serotypes were as follows: S. Infantis (34.8%), mostly isolated from chicken, monophasic S. Typhimurium (1,4, [5],12:i:-) (12.6%), S. Bredeney (8.9%), and S. Derby (8.6%). No Clostridium botulinum, Yersinia spp., and Shigella spp. were isolated. No positivity was detected for hepatitis A virus, while 5.1% of samples taken in the production phase of the food chain were found to be contaminated with norovirus. The chemical analyses identified environmental contaminants within legal limits (heavy metals, 0.6% positive overall; mycotoxins, 0.4% positive overall), analytes subjected to monitoring (perfluoro-alkyl substances (PFASs), 6.2% positive overall; inorganic arsenic, no positives overall) and process contaminants and additives within legal limits (acrylamide, 9.6% positive overall; permitted or nonpermitted additives, 0.9% positive overall). Only one sample showed dioxins and polychlorinated biphenyls (PCBs) at levels higher than the legal limits. The monitoring by competent authorities (CA) of food contamination can generate useful data that can be used as a basis for estimating the exposure to different food contaminants over time and for evaluating the effects of control measures on the contamination of food.

Antimicrobial Resistance and Virulence Factors Assessment in Escherichia coli Isolated from Swine in Italy from 2017 to 2021

Prudent antibiotic use in pigs is critical to ensuring animal health and preventing the development of critical resistance. We evaluated the antimicrobial resistance (AMR) pattern in commensal and enterotoxigenic Escherichia coli (ETEC) isolates obtained in 2017−2021 from pigs suffering from enteric disorders. Overall, the selected 826 E. coli isolates showed the highest level of resistance to ampicillin (95.9%), tetracycline (89.7%), cefazolin (79.3%), and trimethoprim/sulfamethoxazole (74.8%). The resistance rates of the isolates to ampicillin increased (p < 0.05), reaching 99.2% of resistant strains in 2021. Regarding isolates harboring virulence genes, ETEC F18+ were significantly more resistant to florfenicol, gentamicin, kanamycin, and trimethoprim/sulfamethoxazole than ETEC F4+ strains. E. coli lacking virulence factor genes were more resistant to amoxicillin with clavulanic acid and cefazolin, but less resistant to gentamicin (p < 0.01) than isolates harboring virulence factors. Throughout the study period, a significant number of ETEC F18+ isolates developed resistance to florfenicol, gentamicin, and kanamycin. Finally, ETEC 18+ significantly (p < 0.05) increased resistance to all the tested antibiotics. In conclusion, AMR varied for E. coli over time and showed high levels for molecules widely administered in the swine industry, emphasizing the need for continuous surveillance. The observed differences in AMR between commensal and ETEC isolates may lead to the hypothesis that plasmids carrying virulence genes are also responsible for AMR in E. coli, suggesting more research on genetic variation between pathogenic and nonpathogenic E. coli.

Molecular Typing of Leishmania spp. Causing Tegumentary Leishmaniasis in Northeastern Italy, 2014-2020.

Tegumentary leishmaniasis (TL) is endemic but neglected in southern Europe. Therefore, this study aimed to analyze the Leishmania strains causing TL cases in northeastern Italy, where an upsurge of TL cases has been observed in the last decade. Sections from 109 formalin-fixed and paraffin-embedded (FFPE) biopsies of skin and mucosal tissues were collected from TL cases in the selected area. Two DNA targets were amplified and sequenced: the ribosomal internal transcribed spacer 1 (ITS1) and the heat-shock protein 70 gene (hsp70). An in silico analysis was also performed on 149 genomes belonging to the Leishmania donovani complex. A total of 88 out of 109 (80.7%) samples from 83 TL cases were successfully typed by ITS1 and/or hsp70. ITS1 analysis identified L. infantum in 67 cases (91.8%), while L. major (n = 4, 5.5%) and L. tropica (n = 2, 2.7%) were detected in the remaining cases that were categorized as imported. Further, the hsp70 typing of 75 autochthonous cases showed the presence of eight distinct sequence variants belonging to the Leishmania donovani complex, with high genetic variability when compared to known L. infantum populations. In conclusion, our findings show that peculiar L. infantum variants are emerging in the novel focus on TL in northeastern Italy.

Detection and Molecular Characterization of Canine Distemper Virus in Wildlife from Northern Italy.

Canine distemper virus (CDV) is a fatal, highly contagious disease found in wild and domestic carnivores. Several outbreaks have occurred in wildlife in Italy in recent years. This study aims to detect CDV in wildlife following the increasing mortality of foxes (Vulpes vulpes) in the Emilia-Romagna region (northern Italy) observed in 2021. Sixty-seven foxes and one badger (Meles meles) were subjected to necropsy followed by histological examination and were analyzed with molecular techniques to detect the presence of CDV. Of the tested animals, 16% (nine foxes and one badger) were positive for CDV. Phylogenetic analysis showed two different lineages based on complete H gene sequences. The Europe/South America-1 lineage was detected in one fox from Modena, which resembled the CDV variant associated with a previous outbreak in northern Italy in 2018, while the European Wildlife lineage was detected in animals from the Rimini province. Amino acid analysis highlighted a Y549H mutation in all sequences collected, which is commonly associated with increased virulence.

Tegumentary Leishmaniasis in Northeastern Italy from 2017 to 2020: A Neglected Public Health Issue.

Tegumentary leishmaniasis (TL) includes cutaneous (CL) and mucosal (ML) leishmaniasis; despite being endemic in southern Europe, it is often underdiagnosed and underreported. This study aimed to retrospectively examine data collected from patients with TL in a selected area of northeastern Italy (Emilia-Romagna region, RER). A network of 10 diagnostic units within RER was established, and TL cases diagnosed in RER from 2017 to 2020 were evaluated. A total of 135 TL cases were collected (62% male, 38% female); patients ranged from 1 to 84 years, with a median age of 57. Among these cases, 113 (84%) were notified to the public health authorities. The average annual incidence of TL was 0.76 cases per 100,000 inhabitants. Infections were acquired within the RER in 84% of cases; the 113 autochthonous cases were distributed in the foothills areas of the region. We provide evidence of a defined spatial distribution of TL cases in a selected area of northeastern Italy, as well as a relevant number of ML cases. Our observations suggest the need to raise awareness about TL among clinicians and pathologists, promote the molecular confirmation of cases by reference laboratories, and encourage the establishment of surveillance networks for this neglected disease.

Wildlife Hosts of Leishmania infantum in a Re-Emerging Focus of Human Leishmaniasis, in Emilia-Romagna, Northeast Italy.

In the last decade, an upsurge of human leishmaniasis has been reported in the Emilia-Romagna region, Northeast Italy. Epidemiologic data have raised doubts about the role of dogs as the main reservoirs for Leishmania infantum. In the present study, a total of 1,077 wild animals were screened for L. infantum DNA in earlobe and spleen samples from 2019 to 2022. The lymph nodes were tested only in 23 animals already positive in the earlobe and/or spleen. A total of 71 (6.6%) animals resulted positive in at least one of the sampled tissues, including 3/18 (16.7%) wolves, 6/39 (15.4%) European hares, 38/309 (12.3%) roe deer, 1/11 (9.1%) red deer, 8/146 (4.9%) wild boars, 13/319 (4.1%) red foxes, 1/54 (1.9%) porcupine, and 1/59 (1.7%) European badger. Most of the infected animals (62/71) tested positive only in the earlobe tissue, only four animals (two roe deer and two wild boars) tested positive only in the spleen, and five animals (three roe deer and two red foxes) resulted positive for both tissues. L. infantum DNA was detected in the lymph nodes of 6/23 animals. L. infantum detection occurred in all seasons associated with low real-time PCR Ct values. Further research is needed in order to clarify the role of wildlife in the re-emerging focus of leishmaniasis in Northeast Italy.

Diffuse Large B-Cell Lymphoma (DLBCL) in an Aged Raccoon (Procyon lotor).

A 15-years-old, captive, female raccoon (Procyon lotor) was necropsied after a one-week history of apathy and self-isolation. Gross changes consisted of the severe enlargement of the mesenteric lymph node; hepatosplenomegaly with multifocal to coalescing, white tan nodules in the spleen and liver,; and pale kidneys. Histologically, neoplastic CD79α-positive lymphocytes effaced the mesenteric lymph node and multifocally infiltrated the spleen, liver, and kidneys, and focally infiltrated the heart. Based on pathological and immunohistochemical findings, as well as the canine-adapted World Health Organization (WHO) diagnostic criteria, a diagnosis of diffuse large B-cell lymphoma (DLBCL) was made.

Sand Flies and Pathogens in the Lowlands of Emilia-Romagna (Northern Italy).

Cases of sand fly-borne diseases in the Emilia-Romagna region, such as meningitis caused by Toscana virus and human leishmaniasis, are reported annually through dedicated surveillance systems. Sand flies are abundant in the hilly part of the region, while the lowland is unsuitable habitat for sand flies, which are found in lower numbers in this environment with respect to the hilly areas. In this study, we retrieved sand flies collected during entomological surveillance of the West Nile virus (from 2018 to 2021) to assess their abundance and screen them for the presence of pathogens. Over the four-year period, we collected 3022 sand flies, more than half in 2021. The most abundant sand fly species was Phlebotomus (Ph.) perfiliewi, followed by Ph. perniciosus; while more rarely sampled species were Ph. papatasi, Ph. mascittii and Sergentomyia minuta. Sand flies were collected from the end of May to the end of September. The pattern of distribution of the species is characterized by an abundant number of Ph. perfiliewi in the eastern part of the region, which then falls to almost none in the western part of the region, while Ph. perniciosus seems more uniformly distributed throughout. We tested more than 1500 female sand flies in 54 pools to detect phleboviruses and Leishmania species using different PCR protocols. Toscana virus and Leishmania infantum, both human pathogens, were detected in 5 pools and 7 pools, respectively. We also detected Fermo virus, a phlebovirus uncharacterized in terms of relevance to public health, in 4 pools. We recorded different sand fly abundance in different seasons in Emilia-Romagna. During the season more favorable for sand flies, we also detected pathogens transmitted by these insects. This finding implies a health risk linked to sand fly-borne pathogens in the surveyed area in lowland, despite being considered a less suitable habitat for sand flies with respect to the hilly areas.

SARS-CoV-2 and animals, a long story that doesn't have to end now: What we need to learn from the emergence of the Omicron variant.

OIE, the world organization for animal health, recently released an update on the state of the art of knowledge regarding SARS-CoV-2 in animals. For farmed animals, ferrets and minks were found to be highly susceptible to the virus and develop symptomatic disease both in natural conditions and in experimental infections. Lagomorphs of the species Oryctolagus cuniculus are indicated as highly susceptible to the virus under experimental conditions, but show no symptoms of the disease and do not transmit the virus between conspecifics, unlike raccoon dogs (Nyctereutes procyonoides), which in addition to being highly susceptible to the virus under experimental conditions, can also transmit the virus between conspecifics. Among felines, the circulation of the virus has reached a level of cases such as sometimes suggests the experimental use of vaccines for human use or treatments with monoclonal antibodies. But even among wild animals, several species (White-tailed deer, Egyptian rousettes, and minks) have now been described as potential natural reservoirs of the virus. This proven circulation of SARS-CoV-2 among animals has not been accompanied by the development of an adequate surveillance system that allows following the evolution of the virus among its natural hosts. This will be all the more relevant as the surveillance system in humans inevitably drops and we move to surveillance by sentinels similar to the human flu virus. The lesson that we can draw from the emergence of Omicron and, more than likely, its animal origin must not be lost, and in this mini-review, we explain why.

Host preference and Leishmania infantum natural infection of the sand fly Phlebotomus perfiliewi in northern Italy.

The host preference of hematophagous insects is important in determining the cycle of pathogens that they potentially transmit; for example, sand flies are competent vectors of Leishmania parasites. In this work, we evaluated the host preference of sand flies collected in the Emilia-Romagna region of Italy in 2018 and 2019 in an area in which Leishmania infantum circulates actively. Out of about 30,000 sampled sand flies, we obtained 252 engorged females, which were processed to identify the sources of blood meals. Sampling data collected confirmed a positive phototropism of Phlebotomus (Ph.) perfiliewi respect to Ph. perniciosus and the enhanced efficiency of light traps in collecting engorged females compared with traps baited with carbon dioxide. We identified blood source in 185 females (183 Ph. perfiliewi, two Ph. pernicious). The most bitten animal was the roe deer (49.5%), followed by humans (29.2%), hare (7.1%) and cow (4.7%). Other animals, including wild boar, horse, donkey, porcupine, chicken and red fox, were less represented (<2%), while the blood of dogs and rodents were not detected. In addition, we singly screened engorged females for Leishmania founding 5 positive specimens, fed on roe deer (4) and man (1), providing evidence of parasite circulation in a sylvatic environment, where presence of dogs was not common. These findings suggest the existence of an uncharacterized Leishmania reservoir in the surveyed area.

Myocardial Injury Complicated by Systolic Dysfunction in a COVID-19-Positive Dog.

A six-year-old Cavalier King Charles spaniel was referred with a two-month history of severe exercise intolerance and syncope. Clinical signs had developed during a local wave of coronavirus disease (COVID-19) two weeks after its family members had manifested symptoms of this viral disease and their positivity to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was confirmed. Cardiologic assessment documented myocardial injury complicated by systolic dysfunction. An extensive diagnostic work-up allowed us to rule out common causes of myocardial compromise, both infective and not. Accordingly, serological and molecular tests aimed at diagnosing SARS-CoV-2 infection were subsequently performed, especially in light of the dog's peculiar history. Results of such tests, interpreted in the light of previous findings and current knowledge from human medicine, supported a presumptive diagnosis of COVID-19-associated myocardial injury, a clinical entity hitherto poorly described in this species.

Co-Circulation of Phleboviruses and Leishmania Parasites in Sand Flies from a Single Site in Italy Monitored between 2017 and 2020.

Sand flies transmit Leishmania infantum, which is responsible for causing leishmaniasis, as well as many phleboviruses, including the human pathogenic Toscana virus. We screened sand flies collected from a single site between 2017 and 2020 for the presence of both phleboviruses and Leishmania. The sand flies were sampled with attractive carbon dioxide traps and CDC light traps between May and October. We collected more than 50,000 sand flies; 2826 were identified at the species level as Phlebotomus perfiliewi (98%) or Phlebotomus perniciosus (2%). A total of 16,789 sand flies were tested in 355 pools, and phleboviruses were found in 61 pools (6 Toscana virus positive pools, 2 Corfou virus positive pools, 42 Fermo virus positive pools, and 7 Ponticelli virus positive pools, and 4 unidentified phlebovirus positive pools). Leishmania was found in 75 pools and both microorganisms were detected in 16 pools. We isolated nine phleboviruses from another 2960 sand flies (five Ponticelli viruses and for Fermo viruses), not tested for Leishmania; the complete genome of a Fermo virus isolate was sequenced. The simultaneous detection in space and time of the Fermo virus and L. infantum is evidence that supports the co-circulation of both microorganisms in the same location and partial overlap of their cycles. A detailed characterization of the epidemiology of these microorganisms will support measures to limit their transmission.

Animal Coronaviruses and SARS-COV-2 in Animals, What Do We Actually Know?

Coronaviruses (CoVs) are a well-known group of viruses in veterinary medicine. We currently know four genera of Coronavirus, alfa, beta, gamma, and delta. Wild, farmed, and pet animals are infected with CoVs belonging to all four genera. Seven human respiratory coronaviruses have still been identified, four of which cause upper-respiratory-tract diseases, specifically, the common cold, and the last three that have emerged cause severe acute respiratory syndromes, SARS-CoV-1, MERS-CoV, and SARS-CoV-2. In this review we briefly describe animal coronaviruses and what we actually know about SARS-CoV-2 infection in farm and domestic animals.

Distribution, virulence, genotypic characteristics and antibiotic resistance of Listeria monocytogenes isolated over one-year monitoring from two pig slaughterhouses and processing plants and their fresh hams.

Listeria monocytogenes contamination in raw pork and ready to eat foods is an important food safety concern, also for the increasing detection of antimicrobial-resistant isolates. Data on L. monocytogenes occurrence, persistence, distribution and genetic characterization in two different plants, namely in continuum from slaughtered pigs, environment and unfinished products (fresh hams) were observed by one-year monitoring and were integrated with their antimicrobial resistance patterns. A total of 98 samples out of the overall 1131 (8.7%) were positive for L. monocytogenes, respectively 2.6% and 13.2% in plants A and B: only three serotypes were identified, 1/2c (50%), 1/2b (36.7%) and 1/2a (13.27%), and strains were classified in 35 pulsotypes and 16 clusters by PFGE; a unique P-type was highlighted according to the detection of virulence genes. The contamination flow of L. monocytogenes has a low occurrence in slaughterhouse (Plant A = 1.1%, Plant B: 3.1%; p > 0.05) and increased throughout the processing chain with trimming area as the most contaminated (Plant A: 25%, Plant B: 57%; (p < 0.05)), both in the environment and in unfinished products (80% in hams before trimming in plant B). The dominant role of environmental contamination in post-slaughter processing is confirmed to be a significant cause of meat contamination by L. monocytogenes. Very high levels of resistance were observed for clindamycin (57%) and high resistance levels (>20-50%) to ciprofloxacin, oxacillin, levofloxacin and daptomycin, confirming the L. monocytogenes resistance trend to a wide range of antimicrobial agents. A total of 11 L. monocytogenes isolates were multidrug resistant and 7 out of them were isolated from slaughtered pigs. An interesting significant (p < 0.05) statistical correlation has been found between resistance to some antimicrobial agents and lineage/serotypes. Microbiological sampling of food and environments after sanitization are commonly used as verification procedure for the absence of L. monocytogenes in food plants and to give assurance of food safety, but strains characterization is necessary for industries to target specific control measures, like the enforcement of the hygiene program and of the control of operator activities, at least for permanent strains. The only presence of L. monocytogenes could not be considered as the conclusive assessment of a potential risk for public health, also in terms of emerging and emerged antimicrobial resistances.

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.

Isolation and Molecular Typing of Leishmania infantum from Phlebotomus perfiliewi in a Re-Emerging Focus of Leishmaniasis, Northeastern Italy.

Visceral leishmaniasis (VL) caused by Leishmania (L.) infantum is a public health threat in the Emilia-Romagna region, northeastern Italy, but its epidemiology has not been fully elucidated in this area. The objective of this study was to characterize Leishmania infection in sand flies collected in a re-emerging focus of VL in the Bologna province. During the summer of 2016, 6114 sand flies were collected, identified, and tested for Leishmania detection. Of the identified sand flies, 96.5% were Phlebotomus (P.) perfiliewi and 3.5% were P. perniciosus. Detected parasites were characterized by biomolecular methods (multilocus microsatellite typing and characterization of repetitive region on chromosome 31), and quantified by real-time PCR. The prevalence of Leishmania infection in individually-tested P. perfiliewi sand flies varied from 6% to 10% with an increasing trend during the season. Promastigotes of L. infantum were isolated by dissection in one P. perfiliewi female; the isolated strain (Lein-pw) were closely related to Leishmania parasites from VL cases in northeastern Italy, but differed from strains isolated in dogs from the same area. Our findings strongly support the vector status of P. perfiliewi for human VL in the study area.