New insights on Avian orthoreovirus and Chicken astrovirus co-infection in an Italian broiler flock: preliminary biomolecular and pathological results

Summary Common pathogens of intensive poultry farms, either parasitic or bacterial, such as Coccidia or Salmonella, are well known and strictly controlled by veterinary management. This case study reports an unusual case of runting stunting syndrome (RSS) observed on a Sicilian poultry farm of broiler chickens during 2019. The investigation was carried out on five chickens which present delayed in body weight and growth performance. Animals showed also difficulty in deambulation and diarrhea. At necropsy, intestinal lesions were detected in three of the five clinical cases. Gut samples were collected and analyzed to identify potential pathogens responsible for the RSS. Presence of viruses was detected by using quantitative reverse transcription PCR (RT‑qPCR), while selected tissues were fixed and embedded in paraffin wax according to routine procedures. All histological sections were stained with hematoxylin‑eosin. RT‑qPCR successfully detected both Chicken astrovirus (CAstV) and Avian orthoreovirus (ARV). Histology evidenced severe specific lesions on the intestinal mucosa in liver and kidneys. Chicken astrovirus and Avian orthoreovirus RNA was also detected in cecal tonsils, kidney and liver, thus implying their possible primary role in inducing the disease. Further studies are needed to evaluate the role of other possible factors (low biosecurity measures, e.g.) and, most of all, the consequences in terms of economic losses and animal health impairment

Does TLS Exist in Canine Mammary Gland Tumours? Preliminary Results in Simple Carcinomas

Abstract: Neoplastic progression is influenced by the expression of tumour antigens that activate an anti-tumour immune response. Human medical studies show that this body defence is carried out in secondary lymphoid organs (SLOs) but also directly in the tumour through organized cellular aggregates that are called tertiary lymphoid structures (TLSs). However, their occurrence has different meanings in different tumour types. For example, the presence of TLSs in breast cancer is associated with the most aggressive subtypes. This paper aimed to study TLSs in canine mammary simple carcinomas. A morphological assessment of the inflammatory infiltrate was performed on H&E sections of fifty cases. Immunohistochemistry was then carried out to typify the inflammatory cells in the tumour microenvironment. Results showed that, sometimes, inflammatory infiltrates were organized in follicles close to high-grade carcinomas, simulating a lymphoid organization, as in breast cancer. Therefore, we can assume that even in canine mammary tumours, TLSs exist and they are entities to consider due to their presence in the most aggressive histotypes or tumours with a high degree of malignancy

Cardiac filariosis in migratory Mute swans (Cygnus olor) in Sicily

Sarconema eurycerca is a common parasitic disease of North America swans and geese. The infection has been correlated with severe heart lesions, often resulting in cardiac failure and death of the animals. Heartworms infections have been previously reported in European swans, and specifically in the United Kingdom and Nederland. Both the countries are characterized by a cold temperate weather, similar to the one that can be found in swan wintering areas of U.S.A. and Canada. The first record of cardiac filariasis associated with Sarconema eurycerca infection in four swans in Italy. Twelve mute swans were examined during avian influenza surveillance activities on migratory birds. Birds were collected in the year 2006, in wintering areas of Eastern Sicily (Italy). Four of the twelve swans showed necrotic-haemorrhagic myocarditis with intra-lesional nematodes. Morphological characteristics identified the parasite as a filarial nematode. Birds lungs samples were used for parasites DNA extraction. The latter was used as template for polymerase chain reaction (PCR) amplification and sequencing of part of the 12S rDNA gene. Comparison of genomic DNA extracted from a reference S. eurycerca isolate confirmed parasite identity and provided the first sequence resources for this species of value to future diagnostic and epidemiological studies

Molecular Characterization of Antimicrobial Resistance and Virulence Genes of Bacterial Pathogens from Bovine and Caprine Mastitis in Northern Lebanon

Mastitis is an infectious disease encountered in dairy animals worldwide that is currently a growing concern in Lebanon. This study aimed at investigating the etiology of the main mastitiscausing pathogens in Northern Lebanon, determining their antimicrobial susceptibility profiles, and identifying their antimicrobial resistance (AMR) genes. A total of 101 quarter milk samples were collected from 77 cows and 11 goats presenting symptoms of mastitis on 45 dairy farms. Bacterial identification was carried out through matrix-assisted laser desorption/ionization-time of flight mass spectrometry. Antimicrobial susceptibility was tested by disc diffusion and broth microdilution methods. Molecular characterization included polymerase chain reaction (PCR) screening for genes encoding extended-spectrum beta-lactamases (ESBLs) and plasmid-mediated AmpC among Enterobacterales isolates, and virulence factors among Staphylococcus isolates. Escherichia coli isolates were subjected to phylogenetic typing by a quadruplex PCR method. The most frequently identified species were Streptococcus uberis (19.2%), Streptococcus agalactiae (15.1%), E. coli (12.3%), and Staphylococcus aureus (10.96%). Gram-positive bacteria were resistant to macrolides and etracycline, whereas gram-negative bacteria displayed resistance to ampicillin and tetracycline. Two ESBL genes, blaTEM (83.3%) and blaOXA (16.7%), and one AmpC beta-lactamase gene, blaCMY-II (16.7%), were detected among six E. coli isolates, which mainly belonged to phylogenetic group B1. Among Staphylococcus spp., the mecA gene was present in three isolates. Furthermore, four isolates contained at least one toxin gene, and all S. aureus isolates carried the ica operon. These findings revealed the alarming risk of AMR in the Lebanese dairy chain and the importance of monitoring antimicrobial usage

Effects of Microencapsulated Blends of Organics Acids (OA) and Essential Oils (EO) as a Feed Additive for Broiler Chicken. A Focus on Growth Performance, Gut Morphology and Microbiology

Simple Summary Replacing antibiotics with natural alternative compounds in poultry feeding is being increased in the last few years to challenge the antibiotic resistance problem. Among natural compounds, organic acids and essential oils could be a favorable option. The goal of the trial was testing the dietary supplementation of a blend of organic acids and essential oils in broiler diets in order to evaluate growth performance and gut healthiness. The blend of organic acids and essential oils improved growth performances at the end of the growing period and favorably affected, to a certain extent, gut morphology at different gut districts. Moreover, a selective microbial control against Clostridium perfringens, Enterobacteriaceae, Enterococci and Mesophilic bacteria was found. Additionally, in litter, organic acids and essential oils dietary treatment drove to an overall decrease of Mesophilic bacteria and Enterococci counts. Overall, dietary strategy oriented to a supplementation of a mixture of organic acids and essential oils in broiler diets could offer some favorable perspectives in order to maintain adequate growth performance and gut healthiness either in term of morphology or of microbiology. Nevertheless, improving knowledge on the mechanisms of action of these natural additives together with a potential synergistic action is pivotal to clarify their potential as antibiotic replacers. Abstract The goal of the trial was testing the effects of a blend of organic acids and essential oils dietary supplementation on growth performance and gut healthiness in broiler chickens. In total, 420 male Ross 308 chicks (1-day old) were randomly assigned to two dietary treatments: basal (BD) and organic acids and essential oils (OA&EO) diets (three replicates/treatment; 70 broilers/replicate). BD group received commercial diets whereas OA&EO group basal diets + 5 g/kg of microencapsulated organic acids and essential oils. OA&EO treatment improved the average daily gain (p < 0.01) and feed conversion ratio at 37–47 days compared to BD treatment. OA&EO treatment improved gut morphology mostly at ileum and duodenum levels in terms of villi height, crypt depth, number of villi, mucosa thickness and villi area at 24 and 34 sampling days. A certain selective action against Clostridium perfringens in ileum of OA&EO group was shown at 33 (p = 0.053) and 46 days (p = 0.09) together with lower median values for Enterobacteriaceae, Enterococci, Mesophilic bacteria and Clostridium perfringens at ceca level. Overall, organic acids and essential oils supplementation improved growth performance in the final growth stage and some morphological gut traits and reduced to a certain extent Clostridium perfringens count in ileum

Morphological and Immunohistochemical Examination of Lymphoproliferative Lesions Caused by Marek's Disease Virus in Breeder Chickens

Simple Summary The poultry industry is the most intensive and fastest growing among all livestock production systems, and, in the last decades, it has expanded exponentially due to an increasing demand for meat and eggs. Marek’s disease is a highly contagious and rapidly progressive lymphoproliferative disease. It is one of the most dangerous diseases of those affecting the sector because it causes important economic losses. Although widely controlled by vaccination programs, sometimes chickens are not totally protected, and the presence of virulent field strains can allow outbreaks. This case describes the occurrence of Marek’s disease observed in a breeder chicken flock that reported an increase in mortality rate (+0.4–0.6%) after the 32nd week. Histological analysis has highlighted severe lesions on visceral organs of chickens caused by Marek’s disease, especially in the intestinal tract of a hen that had a tumor mass in the distal part of the cloaca. Immunohistochemical staining confirmed the disease-associated tumor. The aim of this study was to underline the importance of vaccine administration related to the maintenance of proper biosecurity practice, especially in the first week of the raising cycle. In addition, monitoring for disease even after vaccination is crucial to minimize economic loss. Abstract Marek’s disease is widely controlled by vaccination programs; however, chickens are not totally protected, especially immediately after the vaccination when a strong challenge could interfere with the effectiveness of vaccination in the absence of proper biosecurity practice. This case report describes the occurrence of Marek’s disease (MD) observed in a breeder chicken flock reared southeast of Sicily. MD outbreak occurred from 32 to 47 weeks with an increase in weekly mortality rate (+0.4–0.6%). Overall, mortality rate related to Marek’s disease was about 6% at the end of the cycle. Carcasses of chickens found during the occurrence of disease underwent necropsy, and tissues were collected to confirm the infection. Gizzard, cecal tonsil, intestine, spleen and tumor mass were collected and analyzed from a carcass of one hen, 32 weeks old and apparently asymptomatic. Multiplex real-time PCR performed on spleen tissues detected the presence of MD virus pathogenic strain. Macroscopic and microscopic evaluation of the rest of the samples confirmed the neoplastic disease. Moreover, the immunophenotype of the tumor cells was identified as CD3 positive by immunohistochemical (IHC) staining. The vaccinated flock had become rapidly infected with the MD virus, which proves that the challenge of the MD virus was too strong in the rearing house at the beginning of the cycle, causing the outbreak

Investigation of an Autochthonous Outbreak of Bovine Besnoitiosis in Northwestern Sicily

Abstract Bovine besnoitiosis is a parasitic disease caused by the protozoan Besnoitia besnoiti, leading to infertility in bulls and abortions in cows. In Italy, it is considered an emerging disease, recently introduced by the importation of animals from Spain and France. In the last decade, many outbreaks have been reported and confirmed in native cattle, mostly in northern and central Italy. This study reports on an autochthonous outbreak of bovine besnoitiosis in two nearby farms located in northwestern Sicily. A 15-month-old Limousine bull born on the farm showed typical clinical signs of the chronic disease phase, such as edema of the scrotum with subsequent hyperkeratosis associated with eschars and thickening of the skin. A histopathological examination revealed the presence of Besnoitia tissue cysts containing bradyzoites in the eyes, tendons, testicles, dermis, and nictitating membrane. A serological investigation using a commercial ELISA kit revealed a high seroprevalence of the antibody anti-B. besnoiti (79.2% for the farms in this study). Clinical disease showed low prevalence (1.5%) despite the high seroprevalence of specific antibodies in the herd, confirming that bovine besnoitiosis is an emergent endemic pathogen in Sicily, but its clinical behavior still remains sporadic

BOVINE BESNOITIOSIS: A CASE REPORT IN SICILY

IINTRODUCTION Bovine besnoitiosis is a parasitic disease caused by the tissue cyst forming protozoan Besnoitia besnoiti The European Food Safety Authority ( has reported that bovine besnoitiosis is re emerging in Europe 1 It represents a chronic and debilitating disease that is responsible for severe economic losses such as poor body condition, decreased milk production, infertility in bulls and occasional abortion 2 Autochthonous outbreaks of the disease were reported in Central Italy for the first time in 2009 3 In Sicily, although the disease was observed in a heifer in 2014 4 no epidemiological data is known AIM OF THE WORK This study describes an autochthonous outbreak of Bovine besnoitiosis in North Western Sicily a bull that shows typical clinical symptoms related to bovine besnoitiosis and moreover, the serological investigation of the herd, suspected of being infected by B besnoiti MATERIALS AND METHODS In a 16 month old affected bull, clinical status was evaluated both in acute and chronic stage After slaughtering, samples of skin, conjunctiva, tongue, and tendons were collected for histopathological examination Furthermore, i n the bull and in the Limousine herd of 72 animals, serological investigation using ID Screen® Besnoitia Indirect 2 0 ID vet was performed. RESULT The bull showed typical clinical signs of besnoitiosis such as: high fever, diffuse oedema at the joints of limbs and scrotum, hyperemia of muzzle and eyes, orchitis , thickening of the nasal planum and wheezing. About 1 0 12 days later, skin lesions ( hyperkeratosis, alopecia, dandruff and scab) appeared (Figure 1). At the 58th day after the beginning of the first signs, the animal showed diffuse skin thickening, was emaciated and reluctant to move. Histological examination showed multifocal coalescing thick wall cysts surrounding parasitophorous vacuole containing thousands of banana shaped bradyzoites all the collected samples (Figure 2 3). ELISA confirmed the seropositivity of the bull, moreover, 79,17% of the herd resulted positive (Tab 1) CONCLUSION. Finally, since all the animals were born in the farm or were bought from nearby farms this case confirms the presence of autochthonous B. besnoiti also in Sicily and may contribute to a further epidemiological study in this area. REFERENCES [1] EFSA Journal, 2010, EFSA Journal 8: 1499-514 [2]Gutiérrez Expósito et al., 2017 Int J Parasitol . 47: 737 51 [3]A. Gentile et al., 2012 Vet Parasitol . 184: 108 15 [4]Di Marco et al., 2016 LXX SISVET. 430

Effect of Mycoplasma agalactiae mastitis on milk production and composition in Valle dell Belice dairy sheep

Contagious agalactia (CA), a disease caused by Mycoplasma agalactiae and other pathogenic mycoplasmas, is a well-known multietiological syndrome affecting dairy breeds of sheep and goats in the Mediterranean basin. The aim of this work was to study the effect on milk production and composition of mastitis caused by M. agalactiae in Valle del Belice dairy sheep. All ewes were manually milked twice daily and the milk from both daily milking was analysed for milk composition and somatic cell counts. Moreover the morning milk samples were collected aseptically from each animal for bacteriological analyses. A mixed linear model was utilised to consider milk production and composition between animals infected by CA and healthy animals. After bacteriological investigation using both cultural and molecular methods, 37 ewes were found to be infected by M. agalactiae while 50 uninfected ewes were randomly selected from the same herds to compare milk production and composition between infected and healthy animals. Statistical analyses showed that the infection with M. agalactiae had a significant effect on yield and some milk components. In particular, infected ewes showed lower milk production with lower lactose content and higher somatic cell counts. The implementation of disease control programmes based on rapid laboratory diagnosis and modern control methods is desirable for Mediterranean endemic areas.Highlights Contagious agalactia is caused by M. agalactiae and affects small ruminant dairy farms in the Mediterranean basin. Contagious agalactia is endemic in many countries and has a severe health and economic impact. Effect on milk production and composition of mastitis caused by M. agalactiae

Polybutylene succinate artificial scaffold for peripheral nerve regeneration

Regeneration and recovery of nerve tissues are a great challenge for medicine, and positively affect the quality of life of patients. The development of tissue engineering offers a new approach to the problem with the creation of multifunctional artificial scaffolds that act on various levels in the damaged tissue, providing physical and biochemical support for the growth of nerve cells. In this study, the effects of the use of a tubular scaffold made of polybutylene succinate (PBS), surgically positioned at the level of a sciatic nerve injured in rat, between the proximal stump and the distal one, was investigated. Scaffolds characterization was carried out by scanning electron microscopy and X-ray microcomputed tomography and magnetic resonance imaging, in vivo. The demonstration of the nerve regeneration was based on the evaluation of electroneurography, measuring the weight of gastrocnemius and tibialis anterior muscles, histological examination of regenerated nerves and observing the recovery of the locomotor activity of animals. The PBS tubular scaffold minimized iatrogenic trauma on the nerve, acting as a directional guide for the regenerating fibers by conveying them toward the distal stump. In this context, neurotrophic and neurotropic factors may accumulate and perform their functions, while invasion by macrophages and scar tissue is hampered