Luca Bano

Scopus Publications

Effectiveness of a Bivalent Recombinant Vaccine on the Production of Neutralizing Antibodies Against BoNT/C, BoNT/D, BoNT/CD e BoNT/DC in Bovines
Ilenia Drigo, Luca Zandonà, Elena Tonon, Katia Capello, and Luca Bano
MDPI AG
Background/Objectives. Bovine botulism, although relatively rare, presents significant economic losses due to high mortality rates and restrictions on livestock product trade. Vaccination remains the most effective strategy for preventing botulism-related mortality. This study evaluated the efficacy of a bivalent recombinant vaccine targeting the C-terminal portion of the heavy chain (Hc) of botulinum neurotoxin serotype C (BoNT/C) (Hc BoNT/C) and botulinum neurotoxin serotype D (BoNT/D) (Hc BoNT/D) in inducing neutralizing antibodies against these toxins and their mosaic variants BoNT/CD and BoNT/DC in cattle. This comparison aims to improve the design of an optimal recombinant vaccine for preventing bovine botulism caused by the most common serotypes. Methods. Twenty, four-month-old Holstein Friesian calves were randomly assigned to two groups of ten animals: vaccinated group and control group. Sera were collected at various time points to assess antibody titers using ELISA and neutralizing antibody titers using a mouse protection assay. Neutralizing antibody titers were compared to those obtained with a commercially available toxoid vaccine. Results. The recombinant vaccine elicited significant increases in anti-HcBoNT/C and anti-HcBoNT/D IgG antibody levels in vaccinated animals compared to controls animals with no adverse effects. Specifically, post-vaccination, the calves showed no local reactions (swelling, warmth) or behavioral changes suggestive of systemic illness. Neutralizing antibody titers against BoNT/C and BoNT/D were significantly higher in the recombinant vaccine group compared to the toxoid vaccine group. However, the recombinant vaccine showed lower neutralizing activity against BoNT/DC compared to the toxoid vaccine. Conclusions. The bivalent recombinant vaccine demonstrated promising immunogenicity in cattle, inducing high neutralizing antibody titers against BoNT/C and BoNT/D. While effective against these toxins, the lower efficacy against BoNT/DC highlights the need for further research to optimize the vaccine formulation, potentially by incorporating a BoNT/DC Hc component, to provide broader protection against bovine botulism.

Neurotoxic phospholipase A<inf>2</inf>: A proposed cause of equine grass sickness and other animal dysautonomias (multiple system neuropathies)
Bruce McGorum, R. Scott Pirie, Luca Bano, Tracey Davey, John Harris, and Cesare Montecucco
Wiley

Antimicrobial susceptibility of Clostridium botulinum group III field strains isolated in Europe from animal outbreaks
Benedetta Cordioli, Alessia Rizzardi, Angela Guolo, Tiziana Ferro, Cosetta Bacchin, Manuel Garbuio, Fabrizio Anniballi, Paola De Santis, Miriam Koene, Caroline Le Maréchal,et al.
Elsevier BV

Rhodococcus equi infection in a dairy goat from a herd in northeastern Italy

A possible cause and therapy for equine grass sickness
Cesare Montecucco and Luca Bano
Elsevier BV

No viable bacterial communities reside in the urinary bladder of cats with feline idiopathic cystitis
Andrea Balboni, Giovanni Franzo, Luca Bano, Lorenza Urbani, Sofia Segatore, Alessia Rizzardi, Benedetta Cordioli, Matteo Cornaggia, Alessia Terrusi, Kateryna Vasylyeva,et al.
Elsevier BV

The “best practices for farming” successfully contributed to decrease the antibiotic resistance gene abundances within dairy farms
Barbara Salerno, Matteo Cornaggia, Raffaella Sabatino, Andrea Di Cesare, Claudio Mantovani, Lisa Barco, Benedetta Cordioli, Luca Bano, and Carmen Losasso
Frontiers Media SA
IntroductionFarms are significant hotspots for the dissemination of antibiotic-resistant bacteria and genes (ARGs) into the environment and directly to humans. The prevalence of ARGs on farms underscores the need for effective strategies to reduce their spread. This study aimed to evaluate the impact of a guideline on “best practices for farming” aimed at reducing the dissemination of antibiotic resistance.MethodsA guideline focused on prudent antibiotic use, selective therapy, and hygienic and immune-prophylactic practices was developed and provided to the owners of 10 selected dairy farms and their veterinarians. Fecal samples were collected from lactating cows, dry cows, and calves both before and after the implementation of the guideline. ARGs (blaTEM, ermB, sul2, and tetA) were initially screened by end-point PCR, followed by quantification using digital droplet PCR. ARG abundance was expressed in relative terms by dividing the copy number of ARGs by the copy number of the 16S rRNA gene.ResultsThe ARG abundances were higher in lactating cows compared to other categories. Despite similar levels of antibiotic administration (based on veterinary prescription data from the sampled farms) in both sampling campaigns, the total abundance of selected ARGs, particularly blaTEM and tetA, significantly decreased after the adoption of the farming guidelines.DiscussionThis study highlights the positive impact of prudent antibiotic use and the implementation of farming best practices in reducing the abundance of ARGs. The lactating cow category emerged as a crucial point of intervention for reducing the spread of antibiotic resistance. These findings contribute to ongoing efforts to address antibiotic resistance in farm environments and strengthen the evidence supporting the adoption of good farming practices.

Assessment of the antimicrobial resistance level in two dairy herds with different antimicrobial consumption

Botulism in Cattle: A Case Report of an Outbreak in Sardinia (Italy)
Luigia Pinna, Annamaria Coccollone, Marcella Maxia, Luca Bano, Concetta Scalfaro, Daniela Mandas, and Manuele Liciardi
MDPI AG
Clostridium botulinum is the main causative agent of botulism in humans and animals. The ingestion of the botulinum neurotoxin, usually types C and D, has been shown to produce disease (neurological symptoms) in most botulism cases in cattle. We report an outbreak in Southern Sardinia that involved a livestock farm with 120 animals, 39 of which died. The aim of this report is to describe the course of this outbreak and the progression of symptoms up to the death of some animals; we also describe the therapeutic approach applied in this case and the analytical techniques used to diagnose the disease. Finally, we emphasize the importance of promptly proceeding with the sampling of several matrixes when a suspicion of botulism arises.

Association between ability to form biofilm and virulence factors of poultry extra-intestinal Campylobacter jejuni and Campylobacter coli
Andrea Laconi, Roberta Tolosi, Ilenia Drigo, Luca Bano, and Alessandra Piccirillo
Elsevier BV

Calves as Main Reservoir of Antibiotic Resistance Genes in Dairy Farms
Barbara Salerno, Matteo Cornaggia, Raffaella Sabatino, Andrea Di Cesare, Maddalena Furlan, Lisa Barco, Massimiliano Orsini, Benedetta Cordioli, Claudio Mantovani, Luca Bano,et al.
Frontiers Media SA
A side effect of antibiotic usage is the emergence and dissemination of antibiotic resistance genes (ARGs) within microbial communities. The spread of ARGs among pathogens has emerged as a public health concern. While the distribution of ARGs is documented on a global level, their routes of transmission have not been clarified yet; for example, it is not clear whether and to what extent the emergence of ARGs originates in farms, following the selective pressure exerted by antibiotic usage in animal husbandry, and if they can spread into the environment. Here we address this cutting edge issue by combining data regarding antimicrobial usage and quantitative data from selected ARGs (blaTEM, blaCTXM, ermB, vanA, qnrS, tetA, sul2, and mcr-1) encoding for resistance to penicillins, macrolides-lincosamides-streptogramins, glycopeptides, quinolones, tetracyclines, sulfonamides, and colistin at the farm level. Results suggest that dairy farms could be considered a hotspot of ARGs, comprising those classified as the highest risk for human health and that a correlation existed between the usage of penicillins and blaTEM abundances, meaning that, although the antibiotic administration is not exclusive, it remains a certain cause of the ARGs' selection and spread in farms. Furthermore, this study identified the role of calves as the main source of ARGs spread in dairy farms, claiming the need for targeted actions in this productive category to decrease the load of ARGs along the production chain.

Genome Sequence of Campylobacter Strain 19-13652, Isolated from Breeding Pheasants
Arianna Peruzzo, Carmen Losasso, Adriana Di Castri, Ilenia Drigo, Luca Bano, and Massimiliano Orsini
American Society for Microbiology
We report the whole-genome sequence of a Campylobacter strain that was isolated from breeding pheasants presenting “bulgy eyes” in Italy. Traditional molecular typing methods did not return any reliable result. Whole-genome sequencing and sequence comparison with known genomes did not meet the criteria for assignment to an existing species.

Severe gastroenteropathy associated with Clostridium perfringens isolation in starving juvenile sturgeons
Ginevra Brocca, Samuele Zamparo, Tobia Pretto, Alessandro Calore, Andrea Marsella, Romy Lucon Xiccato, Matteo Cornaggia, Luana Cortinovis, Luca Bano, Anna Toffan,et al.
Wiley

Extensive genome exploration of clostridium botulinum group III field strains
Silvia Fillo, Francesco Giordani, Elena Tonon, Ilenia Drigo, Anna Anselmo, Antonella Fortunato, Florigio Lista, and Luca Bano
MDPI AG
In animals, botulism is commonly sustained by botulinum neurotoxin C, D or their mosaic variants, which are produced by anaerobic bacteria included in Clostridium botulinum group III. In this study, a WGS has been applied to a large collection of C. botulinum group III field strains in order to expand the knowledge on these BoNT-producing Clostridia and to evaluate the potentiality of this method for epidemiological investigations. Sixty field strains were submitted to WGS, and the results were analyzed with respect to epidemiological information and compared to published sequences. The strains were isolated from biological or environmental samples collected in animal botulism outbreaks which occurred in Italy from 2007 to 2016. The new sequenced strains belonged to subspecific groups, some of which were already defined, while others were newly characterized, peculiar to Italian strains and contained genomic features not yet observed. This included, in particular, two new flicC types (VI and VII) and new plasmids which widen the known plasmidome of the species. The extensive genome exploration shown in this study improves the C. botulinum and related species classification scheme, enriching it with new strains of rare genotypes and permitting the highest grade of discrimination among strains for forensic and epidemiological applications.

Genomic analysis of extra-intestinal Campylobacter jejuni and Campylobacter coli isolated from commercial chickens
Andrea Laconi, Ilenia Drigo, Nicola Palmieri, Lisa Carraro, Elena Tonon, Raffaela Franch, Luca Bano, and Alessandra Piccirillo
Elsevier BV
Campylobacter jejuni and Campylobacter coli have commonly been considered harmless commensal inhabitants of the chicken gut; however, these Campylobacter spp. are known to be able to multiply in the gut and invade other tissues, negatively affecting host health and performance. In this study, fourteen Campylobacter spp. were isolated from chickens showing foci of necrosis on the liver surface resembling lesions observed in cases of avian vibrionic hepatitis/spotty liver disease. The whole genome sequences of the fourteen isolates were analysed and their virulomes compared to those of Campylobacter reference sequences, aiming to investigate the possible association between virulence genes and the observed pathological lesions. Nine C. jejuni and five C. coli were studied. These Campylobacter shared twelve virulence factors with other isolates originated from chicken livers and hosted a higher number of virulence-associated genes in comparison to the reference genomes, including genes encoding for factors involved in adherence to and invasion of the intestinal epithelial cells. Our findings seem to point out that these twelve common virulence-associated genes, together with the presence of a high number of virulence factors involved in adherence, invasion and motility, might be responsible for the extra-intestinal spread of our isolates and the colonization of parenchymatous tissues, possibly causing the pathological lesions observed.

Closing clostridium botulinum group III genomes using long-read sequencing
Cedric Woudstra, Tommi Mäklin, Yagmur Derman, Luca Bano, Hanna Skarin, Christelle Mazuet, Antti Honkela, and Miia Lindström
American Society for Microbiology
Clostridium botulinum group III is the anaerobic Gram-positive bacteria producing the deadly neurotoxin responsible for animal botulism. Here, we used long-read sequencing to produce four complete genomes from Clostridium botulinum group III neurotoxin types C, D, C/D, and D/C. The protocol to obtain high-molecular-weight DNA from C. botulinum group III is described.

Business intelligence tools to optimize the appropriateness of the diagnostic process for clinical and epidemiologic purposes in a multicenter veterinary pathology service
Nicola Pozzato, Laura D’Este, Laura Gagliazzo, Marta Vascellari, Monia Cocchi, Fabrizio Agnoletti, Luca Bano, Antonio Barberio, Debora Dellamaria, Federica Gobbo,et al.
SAGE Publications
Laboratory tests provide essential support to the veterinary practitioner, and their use has grown exponentially. This growth is the result of several factors, such as the eradication of historical diseases, the occurrence of multifactorial diseases, and the obligation to control endemic and epidemic diseases. However, the introduction of novel techniques is counterbalanced by economic constraints, and the establishment of evidence- and consensus-based guidelines is essential to support the pathologist. Therefore, we developed standardized protocols, categorized by species, type of production, age, and syndrome at the Istituto Zooprofilattico Sperimentale delle Venezie (IZSVe), a multicenter institution for animal health and food safety. We have 72 protocols in use for livestock, poultry, and pets, categorized as, for example, “bovine enteric calf”, “rabbit respiratory”, “broiler articular”. Each protocol consists of a panel of tests, divided into ‘mandatory’ and ‘ancillary’, to be selected by the pathologist in order to reach the final diagnosis. After autopsy, the case is categorized into a specific syndrome, subsequently referred to as a syndrome-specific panel of analyses. The activity of the laboratories is monitored through a web-based dynamic reporting system developed using a business intelligence product (QlikView) connected to the laboratory information management system (IZILAB). On a daily basis, reports become available at general, laboratory, and case levels, and are updated as needed. The reporting system highlights epidemiologic variations in the field and allows verification of compliance with the protocols within the organization. The diagnostic protocols are revised annually to increase system efficiency and to address stakeholder requests.

Clostridioides difficile in calves in central italy: Prevalence, molecular typing, antimicrobial susceptibility and association with antibiotic administration
Francesca Blasi, Carmela Lovito, Elisa Albini, Luca Bano, Gastone Dalmonte, Ilenia Drigo, Carmen Maresca, Francesca Romana Massacci, Serenella Orsini, Sara Primavilla,et al.
MDPI AG
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.

Detection of Active BoNT/C and D by EndoPep-MS Using MALDI Biotyper Instrument and Comparison with the Mouse Test Bioassay
Ilenia Drigo, Elena Tonon, Simone Pascoletti, Fabrizio Anniballi, Suzanne R. Kalb, and Luca Bano
MDPI AG
Botulinum neurotoxins (BoNTs) are among the most poisonous known biological substances, and therefore the availability of reliable, easy-to use tools for BoNT detection are important goals for food safety and human and animal health. The reference method for toxin detection and identification is the mouse bioassay (MBA). An EndoPep-MS method for BoNT differentiation has been developed based on mass spectrometry. We have validated and implemented the EndoPep-MS method on a Bruker MALDI Biotyper for the detection of BoNT/C and D serotypes. The method was extensively validated using experimentally and naturally contaminated samples comparing the results with those obtained with the MBA. Overall, the limit of detection (LoD) for both C and D toxins were less than or equal to two mouse lethal dose 50 (mLD50) per 500 µL for all tested matrices with the exception of feces spiked with BoNT/C which showed signals not-related to specific peptide fragments. Diagnostic sensitivity, specificity and positive predictive value were 100% (95% CI: 87.66–100%), 96.08% (95% CI: 86.54–99.52%), and 93.33% (95% CI: 78.25–98.20%), respectively, and accuracy was 97.47% (95% CI: 91.15–99.69%). In conclusion, the tests carried out showed that the EndoPep-MS method, initially developed using more powerful mass spectrometers, can be applied to the Bruker MALDI Biotyper instrument with excellent results including for detection of the proteolytic activity of BoNT/C, BoNT/D, BoNT/CD, and BoNT/DC toxins.

Antimicrobial use and antimicrobial resistance: standpoint and prescribing behaviour of Italian cattle and pig veterinarians
G. Pozza, A. Pinto, S. Crovato, G. Mascarello, L. Bano, M. Dacasto, A. Battisti, B. Bartoli, L. Ravarotto, and S. Marangon
Informa UK Limited
Abstract The scientific community considers the improper use of antimicrobials in farm animals among the causes of the insurgent bacterial resistance contributing to the wider pool of resistance at the animal/human interface with serious public health implications. The present study aims to describe the current perceptions regarding antimicrobial use (AMU) and resistance (AMR) and the prescribing behaviour of two different target groups: cattle and pig veterinary practitioners working in Italy. A semi-structured questionnaire was designed and administered using the Computer-Assisted Web Interviewing method. Among the 789 participants who completed the questionnaire, 53.5% and 12.2% were cattle and pig veterinarians, respectively. Differences between the two groups emerged in their opinions concerning different topics. For example, 69.4% and 85.8% of cattle veterinarians claimed to ‘somewhat or strongly agree’ with the statements ‘prescribing broad-spectrum antibiotics in rearing farms increases AMR’ and ‘the preventive use of antibiotics fosters the development of AMR’, against the observed 59.4% and 69.8% of pig veterinarians (p = .050 and p = .000, respectively). Moreover, 26% of pig veterinarians claimed to ‘somewhat or strongly agree’ with the statement ‘alternative methods currently available could be an efficient alternative to antimicrobial treatment’, against 16.1% of cattle veterinarians (p = .022). At the same time, no differences between the two groups emerged regarding the danger inadequate AMU in animals poses on both animal and human health. From collected data, the need to provide appropriate training strategies with the aim of adopting a behaviour more consistent with the guidelines for prudent AMU in cattle and pig production systems has emerged. HIGHLIGHTS 85.8% of cattle vs. 69.8% of pig veterinarians agreed with the statement ‘the preventive use of antibiotics fosters the development of AMR’. 64.5% of cattle and 69.1% of pig veterinarians declared they suggest/prescribe alternative approaches to the use of antimicrobials. ‘Hygiene-biosecurity-management’ and ‘nutritive principles’ are the alternative strategies most suggested by cattle and pig veterinarians, respectively.

Culture-Dependent and Sequencing Methods Revealed the Absence of a Bacterial Community Residing in the Urine of Healthy Cats
Andrea Balboni, Giovanni Franzo, Luca Bano, Stefano De Arcangeli, Alessia Rizzardi, Lorenza Urbani, Sofia Segatore, Federica Serafini, Francesco Dondi, and Mara Battilani
Frontiers Media SA
A growing number of studies suggest that the lower urinary tract of humans and dogs can harbor a urinary microbiota. Nevertheless, a certain concern has developed that the microbiota reported could be due to unaccounted contamination, especially in low-biomass samples. The aim of this study was to investigate the bacterial community which populates the urine of healthy cats using two approaches: a culture-dependent approach which consisted of the expanded quantitative urine culture (EQUC) techniques capable of identifying live bacteria not growing in standard urine cultures, and a culture-independent approach which consisted of 16S ribosomal RNA next generation sequencing (16S rRNA NGS) capable of identifying bacterial DNA and exploring microbial diversity with high resolution. To avoid confounding factors of possible bacterial contamination, the urine was sampled using ultrasound-guided cystocentesis, and several sample controls and negative controls were analyzed. The urine sampled from the 10 cats included in the study showed no bacterial growth in the EQUC procedure. Although several reads were successfully originated using 16S rRNA NGS, a comparable pattern was observed between urine samples and the negative control, and no taxa were statistically accepted as non-contaminant. Taken together, the results obtained allowed stating that no viable bacteria were present in the urine of healthy cats without lower urinary tract disease and urinary tract infections, and that the bacterial DNA detected was of contaminant origin.

Genome sequence of the fish brain bacterium Clostridium tarantellae
Luca Bano, Matthias Kiel, Gabriele Sales, Andrew C. Doxey, Michael J. Mansfield, Haleluya T. Wami, Marco Schiavone, Ornella Rossetto, Marco Pirazzini, Ulrich Dobrindt,et al.
American Society for Microbiology
Eubacterium tarantellae was originally cultivated from the brain of fish affected by twirling movements. Here, we present the draft genome sequence of E. tarantellae DSM 3997, which consists of 3,982,316 bp. Most protein-coding genes in this strain are similar to genes of Clostridium bacteria, supporting the renaming of E. tarantellae as Clostridium tarantellae .

Botulism in wild birds and changes in environmental habitat: A relationship to be considered
Elena Circella, Antonio Camarda, Luca Bano, Giacomo Marzano, Roberto Lombardi, Francesco D’Onghia, and Grazia Greco
MDPI AG
Any human activity, even if aimed at the improvement of a natural area, can potentially affect wildlife, leading to possible short-term or long-term changes due to the human–wildlife interaction. In this study, a botulism outbreak which occurred in waterfowl in a nature reserve after a conservative environmental action is reported. More than 180 different species of wild birds, including seventy waterfowl species, live in the area. The wildlife reserve rangers built an artificial pond equipped with draining canals in the wetland in order to improve the environment of waterfowl species and to facilitate their supply of food. Then, presumably due to tidal rides, gray mullets (Mugil cephalus) arrived from the sea and settled in the pond. The number of fishes gradually increased, and several fishes died with a peak of mortality in the summer of 2017, creating a great amount of decaying organic material and the optimal conditions for Clostridium botulinum growth and toxin production. A botulism outbreak then occurred rapidly and was characterised by flaccid paralysis and sudden mortality of the birds. Seven mallard ducks (Anas platyrhynchos), 4 common teals (Anas crecca), 1 garganey (Anas querquedula), 2 wood sandpipers (Tringa glareola), 1 little egret (Egretta garzetta), 1 little grebe (Tachybaptus ruficollis), and 4 Eurasian coots (Fulica atra) were found dead. Interestingly, the toxin identified as responsible for the disease outbreak was the mosaic of type C and D toxins (C/D type). The prompt removal of the fish carcasses led to a rapid resolution of the outbreak of the disease, highlighting the relevance of a correct management for any action in environmental contexts. The conclusion is that any human activity in wildlife habitats should be carefully considered in order to assess the possible impacts and to quickly identify the possible risks of changes in wildlife population.

Complete genome sequences of three rabbit endogenous lentivirus type K viruses obtained from commercial meat rabbits in Italy
Gianpiero Zamperin, Adelaide Milani, Alice Fusaro, Alessia Schivo, Luca Zandonà, Luca Bano, and Isabella Monne
American Society for Microbiology
Rabbit endogenous lentivirus type K (RELIK) was discovered in the genome of the European rabbit ( Oryctolagus cuniculus ). In our study, we present three complete genome sequences of RELIK viruses generated using a target amplification approach performed on the RNA of commercial rabbits from Italy.

Erratum to “Investigation of Clostridium botulinum group III's mobilome content” [Anaerobe 49 (2018)71–77](S1075996417302329)(10.1016/j.anaerobe.2017.12.009)
Cédric Woudstra, Caroline Le Maréchal, Rozenn Souillard, Fabrizio Anniballi, Bruna Auricchio, Luca Bano, Marie-Hélène Bayon-Auboyer, Miriam Koene, Isabelle Mermoud, Roseane B. Brito,et al.
Elsevier BV

Luca Bano

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Scopus Publications