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We describe group B Streptococcus linked to disease in farmed pigs and wild porcupines in Italy. Occurrence in pigs was attributed to transmission from nonpasteurized bovine milk whey. Antimicrobial-resistance profiles in isolates from porcupines suggest no common source of infection. Our findings expand the known host range for group B Streptococcus disease.

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.

In recent years human diseases due to mosquito-borne viruses were increasingly reported in Emilia-Romagna region (Italy), from the chikungunya virus in 2007 to the West Nile virus (WNV) in 2008. An extensive entomological survey was performed in 2009 to establish the presence and distribution of mosquito arboviruses in this region, with particular reference to flaviviruses. From May 6 to October 31, a total of 190,516 mosquitoes were sampled in georeferenced stations, grouped in 1,789 pools according date of collection, location, and species, and analyzed by reverse transcription polymerase chain reaction (RT-PCR) to detect the presence of RNA belong to Flavivirus genus. WNV was detected in 27 mosquito pools, producing sequences similar to those of birds and human strains obtained in 2008 outbreak, pointed out the probable virus overwintering. Isolation of WNV was achieved from one of these pools. Moreover 56 pools of mosquitoes tested positive for Usutu virus (USUV). Most PCR positive pools consisted of Culex pipiens, which also was the most analyzed mosquito species (81.4% of specimens); interestingly, USUV RNA was also found in two Aedes albopictus mosquito pools. Simultaneous circulation of WNV and USUV in the survey area was highlighted by occurrence of 8 mosquito WNV- and USUV-positive pools and by the overlaying of the viruses \"hot spots\", obtained by kernel density estimation (KDE) analysis. Land use of sampled stations pointed out a higher proportion of WNV-positive Cx. pipiens pool in rural environments respect the provenience of total sampled pool, while the USUV-positive pools were uniformly captured in the different environments. Obtained data highlighting the possible role of Cx. pipiens mosquito as the main vector for WNV and USUV in Northern Italy, and the possible involvement of Ae. albopictus mosquito in USUV cycle. The described mosquito-based surveillance could constitute the foundation for a public health alert system targeting mosquito borne arboviruses.

Background Tegumentary leishmaniasis (TL) caused by Leishmania infantum is an overlooked yet re-emerging disease endemic in Mediterranean Europe. Currently, no standardized molecular surveillance of circulating Leishmania strains is performed in European endemic areas, despite the potential public health implications of parasite biodiversity. This study aims to characterize parasite population haplogroups causing TL in two active endemic areas in southern Europe, i.e. Bologna (northern Italy) and Fuenlabrada (central Spain). Methods In this retrospective study, we typed 87  L. infantum  samples from TL cases in the areas of Bologna and Fuenlabrada; these areas hosted the main European foci of human TL occurring in the last 15 years. Two Leishmania genomic typing targets were used: the heat shock protein 70 (Hsp70 ) and the cysteine peptidase b (Cpb ). Simpson’s index was used to calculate the discriminatory power of the used typing methods. Results Typing results depicted the presence of a heterogeneous parasite population circulating in Bologna with two main haplogroups, i.e. Hsp70 (A)_ Cpb (F) ( n  = 7, 30.4%) and Hsp70 (G)_ Cpb (E/F) ( n  = 7, 30.4%), differing from the reference L. infantum strain JPCM5 haplogroup and partially overlapping with L. donovani lineages. Among the samples from Fuenlabrada, n  = 19 samples were typed by both targets, revealing a homogeneous population expressing Hsp70 (A) and Cpb (E), matching the JPCM5 reference strain haplogroup. Overall, the Cpb typing method exhibited higher discrimination power as compared to the Hsp70 method (Simpson’s index of diversity, P -value < 0.05). Conclusions Our findings show differences among L. infantum populations causing TL in two southern European epidemiological foci of human leishmaniasis and support the recent discovery of L. infantum/L.donovani hybrid strains circulating in northern Italy. These results underscore the critical need to identify the circulating Leishmania strains in endemic areas and assess their potential public health implications in active foci. Graphical Abstract

West Nile (WNV) and Usutu (USUV) viruses are mosquito-borne flaviviruses. Thanks to their importance as zoonotic diseases, a regional plan for surveillance of Arboviruses was implemented in Emilia-Romagna in 2009. The province of Ferrara belongs to the Emilia-Romagna region, and it is an endemic territory for these viruses, with favorable ecological conditions for abundance of mosquitoes and wild birds. From 2015 to 2019, we collected 1842 dead-found birds at a wildlife rehabilitation center, which were analysed by three different PCRs for the detection of WNV and USUV genomes. August was characterized by the highest infection rate for both viruses. Columbiformes scored the highest USUV prevalence (8%), while Galliformes and Strigiformes reported the highest prevalence for WNV (13%). Among Passeriformes (the most populated Order), Turdus merula was the most abundant species and scored the highest prevalence for both viruses. To optimize passive surveillance plans, monitoring should be focused on the summer and towards the avian species more prone to infection by both viruses.

Visceral leishmaniasis (VL) caused by Leishmania infantum is an ongoing health problem in southern Europe, where dogs are considered the main reservoirs of the disease. Current data point to a northward spread of VL and canine leishmaniasis (CanL) in Italy, with new foci in northern regions previously regarded as non-endemic. Multilocus microsatellite typing (MLMT) was performed to investigate genetic diversity and population structure of L. infantum on 55 samples from infected humans, dogs and sand flies of the E-R region between 2013 and 2017. E-R samples were compared with 10 L. infantum samples from VL cases in other Italian regions (extra E-R) and with 52 strains within the L. donovani complex. Data displayed significant microsatellite polymorphisms with low allelic heterozygosity. Forty-one unique and eight repeated MLMT profiles were recognized among the L. infantum samples from E-R, and ten unique MLMT profiles were assigned to the extra E-R samples. Bayesian analysis assigned E-R samples to two distinct populations, with further sub-structuring within each of them; all CanL samples belonged to one population, genetically related to Mediterranean MON-1 strains, while all but one VL cases as well as the isolate from the sand fly Phlebotomus perfiliewi fell under the second population. Conversely, VL samples from other Italian regions proved to be genetically similar to strains circulating in dogs. A peculiar epidemiological situation was observed in northeastern Italy, with the co-circulation of two distinct populations of L. infantum; one population mainly detected in dogs and the other population detected in humans and in a sand fly. While the classical cycle of CanL in Italy fits well into the data obtained for the first population, the population found in infected humans exhibits a different cycle, probably not involving a canine reservoir. This study can contribute to a better understanding of the population structure of L. infantum circulating in northeastern Italy, thus providing useful epidemiologic information for public health authorities.

The Trypanosoma theileri group includes several trypanosome species hardly distinguishable due to the lack of discriminating morphological characters. Trypanosomes belonging to this group have been isolated from different bovine, ovine, and cervids in Europe, Africa, Asia, and Americas. The principal vectors of the T. theileri group are considered tabanid flies; however, T. melophagium is transmitted exclusively by sheep keds. In 2016, 128 sand flies out of 2,728 trapped in Valsamoggia municipality, Italy, were individually dissected and an unknown trypanosome strain, named TrPhp1, was isolated from a female of the sand fly Phlebotomus perfiliewi. Sequence analysis placed this trypanosome in the T. theileri group with very high homology to other trypanosomes detected in European cervids. This is the first report of the T. theileri group isolation from a sand fly, and the possible role of this insect group in the trypanosome transmission cycle is discussed. Within the T. theileri group, the phylogenetic analysis distinguished several lineages, which, unfortunately, do not correspond with their host specificity and their taxonomic status remains ambiguous.

Paenibacillus larvae (P. larvae) is responsible for American foulbrood (AFB), the most severe bacterial disease of honeybees. The enumeration of P. larvae spores in substrates taken from hives allows for the identification of the contamination levels of the colonies, mostly in those with atypical symptoms or with asymptomatic infections; in these cases, it is essential for the effective control of American foulbrood (AFB). In this work we described a new quantitative TaqMan® probe-based real-time PCR (qPCR) assay, targeting the 16S rRNA gene of P. larvae, used for the quantification of P. larvae spores in powdered sugar samples collected from hives, in comparison to the culture. A total of 105 colonies were selected, belonging to 10 apiaries with different levels of infection, located in northern Italy. The proportions of positive colonies was 54% (57/105) with the culture and 66% (69/105) with qPCR. A significant difference between the two methods was found with McNemar’s test (p = 0.02). Out of the 51 positive samples by both methods, 45 showed higher infection by qPCR compared to the culture. A close concordance with the clinical–epidemiological status of the hives was observed by both methods, with higher infection levels found by qPCR.

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 1077 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.

Bacterial diseases are widespread in pet birds, posing a severe threat in nestlings and birds with weakened immune systems, often resulting in high mortality during the first days after hatching. This study aimed to describe the pathological findings of a natural bacterial co-infection with Klebsiella pneumoniae and Pseudomonas aeruginosa in nestling European goldfinches (Carduelis carduelis), as a cause of mortality after hatching. Eight nestlings kept in a breeding facility showed an inability to move, anorexia, and respiratory distress, and were found dead between 1 and 4 days of life. Gross pathological findings included diffuse pneumonia with edema and multifocal hemorrhages and occasionally hepatic necrosis. On histopathology, the main findings in all examined birds included severe, subacute bronchopneumonia and severe necrotizing hepatitis. Gram-negative bacilli were observed in parabronchial walls, within pulmonary arteries and surrounding hepatocellular necrotic foci. Lungs, livers and kidneys were sampled for bacteriological examination, resulting in two Gram-negative bacterial isolates. Four housekeeping genes (i.e., 16S rRNA; rpoB; khe; ecfX) were amplified and sequenced for bacterial identification at species level. Although K. pneumoniae and P. aeruginosa are common Gram-negative pathogens and are often co-isolated in human bacterial pneumonia, co-infection with these bacteria has not been documented in nestling goldfinches to date. Pathogen identification is essential for formulating a correct etiological diagnosis and further selecting the most appropriate therapeutic strategy.