Carmen Maresca

Background: Brucella canis is a zoonotic pathogen associated with reproductive disorders in dogs and represents an emerging public health concern. Dogs are the only known source of infection for humans, and transmission is often associated with close contact, particularly in occupational settings. Reports of canine and human infections in Europe are increasing, underscoring the need for integrated surveillance to assess the risk of introduction and spread. Objectives: This study aimed to investigate the possible circulation of B. canis in different subgroups of dogs from Central Italy, representing diverse risk contexts (stray, breeding, blood donor, refugee-associated, and previously outbreak-linked dogs), and to generate sentinel data to inform further risk-based surveillance and zoonotic risk assessment. Methods: A comprehensive serological, molecular, and bacteriological survey was conducted on 128 dogs sampled in the Umbria region, covering animals from different backgrounds and risk contexts. Blood samples were tested using bacterial culture, real-time PCR, serum agglutination test, complement fixation test, and/or indirect immunofluorescence antibody test. Results: All tested dogs were negative for B. canis. The upper 95% confidence limit for prevalence was 3.5%, suggesting that widespread circulation is unlikely, although a low/moderate prevalence in specific groups cannot be excluded. Conclusions: Although no cases of B. canis were detected, the results provide sentinel information and highlight the need for continued risk-based surveillance, particularly in low-prevalence areas to prevent introduction of the infection and to enable early detection in case of occurrence. As dogs are the only known source of human infection, veterinary monitoring plays a pivotal role in mitigating zoonotic risks and supporting One Health strategies for evidence-based control.

Background: Brucella canis is a zoonotic pathogen associated with reproductive disorders in dogs and represents an emerging public health concern. Dogs are the only known source of infection for humans, and transmission is often associated with close contact, particularly in occupational settings. Reports of canine and human infections in Europe are increasing, underscoring the need for integrated surveillance to assess the risk of introduction and spread. Objectives: This study aimed to estimate the prevalence of B. canis in dogs from Central Italy, a region potentially at risk due to several factors. Methods: A comprehensive serological, molecular, and bacteriological survey was conducted on 128 dogs of various origins, including strays, blood donors, breeding dogs, dogs accompanying refugees, and dogs previously linked to a B. canis outbreak. Blood samples were tested using bacterial culture, real-time PCR, serum agglutination test, complement fixation test, and indirect immunofluorescence antibody test. Results: All tested dogs were negative for B. canis, with an estimated maximum prevalence of 3.5% (95% confidence interval), suggesting a low likelihood of infection within the sampled population. Conclusions: Although no positive cases were detected, maintaining an active and continuous surveillance system is essential to prevent B. canis introduction, particularly through international dog movements and other high-risk scenarios. As dogs represent the only known source of human infection, veterinary surveillance plays a pivotal role in mitigating zoonotic risks. This study highlights the importance of regional surveillance, particularly in low-prevalence areas, as part of broader efforts to support evidence-based control strategies within a One Health framework.

Alternaria toxins (ATs) are a group of toxins produced by Alternaria fungi that frequently contaminate tomatoes and tomato products. Recently, the European Food Safety Authority evaluated ATs for their genotoxic and carcinogenic properties. Alternaria infestation is often controlled using ad hoc treatment strategies (fungicides). In this study, two analytical methods were developed, validated and applied for the determination of five ATs and 195 pesticides in tomato products collected from the Italian market. Two distinct groups, organic (n = 20) and conventional (n = 20) Italian tomato sauces, were characterized in depth. Tenuazonic acid, alternariol and alternariol monomethyl ether were found up to 517, 27 and 7.1 µg/kg, respectively, while pesticides were detected between 0.0026 and 0.0421 mg/kg in conventional products, and, interestingly, up to 0.0130 mg/kg in organic products. No correlation emerged between the detected levels of ATs and pesticides and the type of tomato cultivation, but the probability of pesticide contamination in conventional products was eight times higher than in organics. Some considerations about exposure assessment and risk characterization for ATs were also proposed in the overall population and in more sensitive and/or exposed subgroups, underlining the need for new focused toxicological and monitoring studies to establish reliable reference values. Moreover, these data highlight that fungicide treatments may not protect tomatoes from ATs contamination, although it may remove fungi infestation. As organic product consumption is increasing, it is important to lay down dedicated regulations for maximum permitted levels to ensure the food safety of these products that are often perceived by consumers as a healthier and environmentally friendlier choice.

Canine prostatic serum esterase (CPSE) is considered a useful tool to identify prostate disorders in dogs, with increasing interest in ultrasound (US)‐based sonoelastography to non‐invasively detect prostate disorders. Since no report is available about a possible correlation between these diagnostic tools, we aimed to investigate a possible correlation between strain elastography (SE) and 2D‐shear wave elastography (SWE) and CPSE. Twenty‐one dogs were included and, on each animal, CPSE was evaluated followed by a complete US examination and SE and 2D‐SWE application. Healthy dogs were identified based on the CPSE results. All the dogs included were characterized by normal CPSE values (<52.3 ng/mL) and normal US prostate appearance. The prostate was characterized by intermediate stiffness with SE (pattern III – 84.7% for the left lobe and 79.27% for the right lobe) and softer than the abdominal wall (SR 0.6 for the left lobe and 0.56 for the right lobe), with low values for both m/s and kilopascals (kPa) for 2D‐SWE, pointing that the healthy tissue is not hard. 2D‐SWE results were, respectively, 13.51 ± 5.55 kPa and 2.31 ± 0.42 m/s for the left lobe and 18.05 ± 6.47 kPa and 2.39 ± 0.43 m/s for the right lobe. The significant difference between the right and left measurements expressed with kPa, not evidenced with m/s, can be considered indicative of m/s as the most reliable measurement to be considered regarding the prostate parenchyma. Even though no linear correlation was detected between CPSE and elastography values, these preliminary results evidence that the healthy prostates were characterized by a similar elastographic pattern, thus pointing that these techniques can be potentially useful to be applied in case of prostatic disorders to improve the accuracy of the final diagnosis in a non‐invasive way.

Among the fungi responsible for deep mycosis, the genus Aspergillus plays a predominant role both in human and veterinary medicine. From a “One Health” perspective, infections by Aspergillus spp. often represent a public health problem linked to specific occupational categories that could have a greater risk of inhaling spores and developing any respiratory disease. This preliminary investigation allowed to acquire information about the spread of Aspergillus spp. in avian livestock of the Umbria region (Central Italy), their sensitivity to antifungals, and the presence of mycotoxins in the considered farms. Environmental, feed, animal, and human samples were collected for mycological investigations; chemical analyses were also performed in feed samples. Moreover, prevalence estimated of the fungal isolates were provided for each individual farm sampled. Direct fungal identification was possible in 298 out of the 559 total samples; 162 of the samples were positive for Aspergillus spp. Mycotoxins were detected in 5 out of the 21 feed samples collected. All the aspergilli tested for antifungal susceptibility were resistant to fluconazole. The results obtained show how much the genus Aspergillus is widespread in the investigated farms; therefore, the poultry livestock represents a favorable environment for the maintenance and spread of fungal spores and their potential transmission to animals and humans.

Poultry is the most likely source of livestock-associated Extended Spectrum Beta-Lactamase (ESBL) and plasmid-mediated AmpC (pAmpC)-producing E. coli (EC) for humans. We tested the hypothesis that farming methods have an impact on the load of ESBL/pAmpC-EC in the gut of broilers at slaughter. Isolates (n = 156) of antibiotic-free (AF), organic (O), and conventional (C) animals were characterized for antibiotic susceptibility and antibiotic resistance genes. Thirteen isolates were whole-genome sequenced. The average loads of ESBL/pAmpC-EC in cecal contents were 4.17 Log CFU/g for AF; 2.85 Log CFU/g for O; and 3.88 Log CFU/g for C type (p < 0.001). ESBL/pAmpC-EC isolates showed resistance to antibiotic classes historically used in poultry, including penicillins, tetracyclines, quinolones, and sulfonamides. Isolates from O and AF farms harbored a lower proportion of resistance to antibiotics than isolates from C farms. Among the determinants for ESBL/pAmpC, CTX-M-1 prevailed (42.7%), followed by TEM-type (29%) and SHV (19.8%). Avian pathogenic E. coli (APEC), belonging to ST117 and ST349, were identified in the collection. These data confirm the possible role of a broiler as an ESBL/AmpC EC and APEC reservoir for humans. Overall, our study suggests that antibiotic-free and organic production may contribute to a reduced exposure to ESBL/AmpC EC for the consumer.

The emergence of Clostridioides difficile as the main agent of antibiotic-associated diarrhoea has raised concerns about its potential zoonotic role in different animal species. The use of antimicrobials is a major risk factor for C. difficile infection. Here, we provide data on C. difficile infection in dairy and beef calves in Umbria, a region in central Italy. This cross-sectional study focuses on prevalence, risk factors, ribotypes, toxinotypes and antimicrobial resistance profiles of circulating ribotypes. A prevalence of 19.8% (CI95%, 12–27.6%) positive farms was estimated, and the prescription of penicillins on the farms was associated with C. difficile detection (OR = 5.58). Eleven different ribotypes were found, including the ST11 sublineages RT-126 and -078, which are also commonly reported in humans. Thirteen isolates out of 17 showed resistance to at least one of clindamycin, moxifloxacin, linezolid and vancomycin. Among them, multiple-drug resistance was observed in two isolates, belonging to RT-126. Furthermore, RT-126 isolates were positive for tetracycline resistance determinants, confirming that tetracycline resistance is widespread among ST11 isolates from cattle. The administration of penicillins increased the risk of C. difficile in calves: this, together with the recovery of multi-resistant strains, strongly suggests the need for minimising antibiotic misuse on cattle farms.

Clostridium (C.) perfringens is the causative agent of several diseases and enteric infections in animals and humans. The pathogenicity of the bacterium is largely mediated by the production of a wide range of toxins. Individual C. perfringens strains produce only subsets of this toxin repertoire, which permits the classification in seven toxinotypes (A–G). In addition, a variety of minor toxins further characterizes the single strains. The aim of this work was to evaluate, using Polymerase Chain Reaction (PCR) assays, the diversity of 632 C. perfringens strains isolated in Italy over 15 years. The genotyped strains were analyzed to determine the presence of major and minor toxins (cpe, consensus, and atypical cpb2), their geographical origins, and the source of isolation (animal species or food). Our study shows that toxinotype A had the greatest representation (93%) and correlated mainly with consensus cpb2 in a variety of animal species, as well as with atypical cpb2 in the five food samples. Type D, associated with cpe and atypical cpb2 minor toxins, was identified in 3% of the cases, and type F was identified in 2.5%. Seven type C isolates (1.1%) were detected in cattle, whereas the only type B atypical cpb2 isolated in Italy was detected in a goat, and one type E cpe+atypical cpb2 was detected in a sheep. Type G was not detected.

The overuse of antibiotics in livestock contributes to the antibiotic resistance pandemic. The assessment of the actual antibiotic consumption is crucial in limiting the expansion of the problem effectively. The aim of this study was to provide the first qualitative and quantitative analysis of antimicrobial usage using data from paper-based registers on dairy and beef farms located in the Umbria region, Italy. Antimicrobial therapies of a one-year period were collected from 101 farms with at least 50 cattle each. Defined daily doses (DDD_vet) and defined course doses (DCD_vet) were calculated per administration route and antimicrobial class. The total courses administered were fewer in beef (330.7 × 10⁻³ DCD_vet/year) than in dairy farms (1034.1 × 10⁻³ DCD_vet/year). The use of the highest priority critically important antimicrobials (HPCIAs) was higher (p = 0.0033) in dairy than in beef herds. In terms of DDD_vet, the parenteral fluoroquinolone administration ranked second and fourth on dairy and beef farms, respectively; the consumption of beta-lactams was ten times higher on dairy than on beef farms. Our results confirm that intensive dairy management practices are associated with increased antibiotic consumption and highlight the necessity to strengthen the existing stewardship programs by involving all stakeholders in effective antimicrobial resistance reduction plans.