Short communication: Detection of undeclared presence of bovine milk in buffalo yogurt.

The authenticity of buffalo (Bubalus bubalis) dairy products is a focal issue, considering the increasing demand for buffalo milk products. Therefore, the aim of this study was to investigate the undeclared presence of bovine (Bos taurus) milk in buffalo yogurt, to understand which risk factors might make the product vulnerable to fraud. Real-time PCR assay showed the undeclared presence of bovine DNA in addition to buffalo DNA in 18 of 72 samples. Given the widespread lack of data on the presence of undeclared milk species in buffalo dairy products, the study provides a significant insight into the incidence of fraud in the buffalo dairy field. The data from this study could help improve the analysis of food safety risks along the buffalo milk supply chain and in the dairy processing industry, perceived as being highly vulnerable to food fraud, and prioritize target areas for food policy making to steer and enforce European food fraud regulations.

Electrochemical Preparation of Synergistic Nanoantimicrobials.

The rapid spreading of resistance among common bacterial pathogens towards the misused antibiotics/disinfectant agents has drawn much attention worldwide to bacterial infections. In light of this, the present work aimed at the realization of core-shell nanoparticles possessing remarkable antimicrobial properties thanks to the synergistic action of the metal core and the disinfectant shell. Copper nanoparticles stabilized by benzalkonium chloride were prepared, characterized, and implemented in poly-vinyl-methyl ketone to obtain nanoantimicrobial composite coatings. Bioactivity tests are reported, proving the excellent disinfectant properties of the proposed nanomaterials, as compared to one of the well-known and strongest silver-based nanoantimicrobials. Applications are also briefly described.

Occurrence of Aichi virus in retail shellfish in Italy.

AiV-1 is considered an emerging human enteric pathogens and foodborne transmission has been documented as an important source of exposure for humans, chiefly in relation to non-safe, risky food habits. We surveyed the presence of AiV-1 in retail shellfish, including oysters and mussles, identifying the virus in 3/170 (1.8%) of the analysed samples. The AiV-1 positive samples were of different geographic origin. Upon sequence analysis of a portion of the 3CD junction region, two AiV strains identified from harvesting areas in Northern Italy were characterised as genotype B and displayed 99-100% identity at the nucleotide level to other AiV-1 strains detected in sewages in Central Italy in 2012, suggesting that such strains are stably circulating in Italian ecosystems. Interestingly, a strain identified from mussles harvested in Southern Italy could not be characterised firmly, as inferred in the Bayesian analysis and by sequence comparison, indicating that different AiV strains are also circulating in Italy. Viral contamination in retail shellfish challenges the microbiological guidelines for food control and requires the development and optimization of additional diagnostic and prevention strategies.

Occurrence of potentially pathogenic arcobacters in shellfish.

Considering that several recent cases of human gastroenteritis have been associated with species from the Arcobacter genus, and that few data are currently available about the occurrence of this genus in Italian shellfish, the aim of the present study was to evaluate the occurrence of Arcobacter spp. and the presence of virulence-associated genes. The approach consisted of cultural and biomolecular (multiplex-PCR and 16S-RFLP) methods identifying isolates, followed by PCR assays aimed at the cadF, ciaB, cjl349, irgA, hecA putative virulence genes. Arcobacter spp. was detected in 16/70 (22.8%) shellfish samples. Specifically, Arcobacter spp. was highlighted in 10/42 (23.8%) mussel and in 6/28 (21.4%) clam samples. Subsequently, biomolecular assays revealed Arcobacter butzleri in 12/16 (75%) and Arcobacter cryaerophilus 1B in 4/16 (25%) isolates. PCRs aimed at the five putative virulence genes demonstrated widespread distribution of these genes among Arcobacter isolates and some differences from the results published by other authors. Our research provides more information regarding the health risks associated with the consumption of raw bivalve molluscs and underlines the need to implement an adequate control plan by performing intensive and continuous monitoring in order to guarantee human health.

Occurrence of mislabeling in meat products using DNA-based assay.

Considering that the authentication of food contents is one of the most important issues for the food quality sector, and given the increasing demand for transparency in the meat industry followed the horsemeat scandal in Europe, this study investigates processed-meat products from Italian markets and supermarkets using the mitochondrial cytochrome b gene qualitative PCR identification system in order to verify any species substitution or mislabeling. The results revealed a high substitution rate among the meat products, highlighting a mislabeling rate of 57 %, and consequently, considerable discordance with the indications on the labels, which raises significant food-safety and consumer-protection concerns.

Occurrence of Prototheca spp. in cow milk samples.

Protothecosis is a potential zoonotic disease associated with bovine mastitis which can be transmitted to humans through contaminated milk. Considering the increasing prevalence of bovine mastitis due to Prototheca species, individual cow milk samples were analyzed using microbiological examination and biomolecular assay. Aspects related to health requirements for milk production, clinical and histological bovine mastitis were also described. The results showed 24/257 (9.3%) culture-positive samples and 42/257 (16.3%) PCR-positive samples. Moreover in 5 cows with somatic cell count over 106/mL presented histological features of mastitis. This study reveals that the presence of Prototheca species in dairy herds was related to the hygienic conditions of the milking equipment, showing an emerging public health issue.

Evaluation of hepatitis E virus RNA persistence in experimentally contaminated cured pork liver sausages.

Hepatitis E is a disease sustained by RNA viruses, which have four different genotypes, all of which are responsible for acute forms of hepatitis. Genotypes 1 and 2 infect only humans, causing epidemics mainly transmitted by contaminated water, while geno-types 3 and 4 are zoonotic, and the infection is linked to the consumption of raw or undercooked meat or meat products. Hepatitis E virus (HEV) genotypes 3 and 4 have been detected in domestic Suidae, considered the asymptomatic reservoir of HEV, and in wild animals such as wild boar and deer. Despite scientific studies that have highlighted the presence of HEV in cured meat products, such as pork liver sausages, the viral persistence in the different production steps of curing has not been evaluated. Therefore, this study aimed to evaluate the persistence of HEV genotype 3 during the different curing and storage times of experimentally contaminated pork liver sausages using biomolecular methods. The sausages tested positive at all curing and storage times. This study confirms the potential risk attributed to pork liver sausages in HEV transmission. However, to guarantee an efficient risk assessment, future studies will be performed to correlate the presence of HEV RNA with infectious viral particles.

The EU Interreg Project "ADRINET": Assessment of Well-Known and Emerging Pollutants in Seafood and Their Potential Effects for Food Safety.

Anthropogenic activities lead to the spread of chemicals and biological materials, including plastic waste, toxic metals, and pharmaceuticals, of which the impact on the Mediterranean Sea is of high concern. In this context, the EU Interreg Italy-Albania-Montenegro Project "ADRINET (Adriatic Network for Marine Ecosystem) _244" (2018-2020) arises. It aims to carry out biomonitoring campaigns in the main commercial interest of fish and cephalopod species, such as Sparus aurata, Dicentrarchus labrax, Sepia spp., and Loligo spp. sampled in three different subregions of the Mediterranean Sea. The presence of the main environmental contaminants, such as cadmium, microplastics, and antibiotics was investigated in these seafood samples. Contamination by cadmium and antibiotics in the seafood investigated in our study was negligible. However, a high value of microplastics was detected in the stomach and gut of Sparus aurata and Dicentrarchus labrax. Overall, even though the presence of microplastics needs to be investigated by further studies, the results confirmed that the environmental conditions of the three bays investigated by the ADRINET project partners (Italy, Albania, Montenegro) are positive and not affected by intensive anthropogenic activity.

ERIC-PCR genotyping of paenibacillus larvae in southern Italian honey and brood combs.

Given the considerable economic loss to beekeepers worldwide and the possible public health implications related to the presence of antibiotics in honey, an American Foulbrood (AFB) monitoring/prevention program for Paenibacillus larvae is regarded as essential. This study investigates the occurrence and distribution of P. larvae genotypes in honey and brood combs from Apulia (Italy). Genotyping of P. larvae isolates using ERIC-PCR generated a total of four different ERIC banding patterns (ERIC-A, ERIC-B, ERIC-C, ERIC-D), including fragments ranging from 200 to 3000 bp. Considering that the genotype has an influence on P. larvae infections and multi-genotype infections of colonies or apiaries may increase the complexity of P. larvae infections by influencing the type and speed of the development of clinical symptoms, the findings of the present study could be helpful for training veterinarians, bee inspector's extension staff, and beekeepers, thus improving the detection of AFB infections in the field.

Occurrence and characterization of Arcobacter spp. from ready-to-eat vegetables produced in Southern Italy.

Given that the number of foodborne illness outbreaks linked to the consumption of ready-to-eat vegetables has been widely documented and considering that data on the occurrence of Arcobacter spp. in such foodstuffs are lacking, the aim of the present study was to evaluate the presence of Arcobacter spp. and the occurrence of virulence factors as well as to genotype Arcobacter spp. in ready-to-eat (RTE) vegetable samples, using cultural and biomolecular assays. Arcobacter spp. was detected in 16/110 (14.5%) samples, with A. butzleri being detected in 15/16 and A. cryaerophilus in 1/16 isolates. PCRs aimed at the nine putative virulence genes demonstrated widespread distribution of such genes among A. butzleri and A. cryaerophilus isolates. In addition, multilocus sequence type (MLST) analysis revealed a low genetic diversity within the arcobacters isolates. The results underline the need to develop an appropriate surveillance system based on biomolecular characterization for an integrated microbiological risk assessment of ready-toeat vegetables, and consequently of composite foods.

Evaluation of the persistence of SARS-CoV-2 (ATCC® VR-1986HK™) on two different food contact materials: flow pack polyethylene and polystyrene food trays.

Even though SARS-CoV-2's primary transmission pathway is person-to-person, the role played by surfaces and food contact materials in carrying viral RNA should be further explored. For this purpose, the study aimed to investigate the persistence of SARS-CoV-2 using the strain ATCC® VR-1986HK™ on flow pack polyethylene (FPP) and polystyrene food trays (PFT). Samples of FPP and PFT were contaminated with heat-inactivated SARS-CoV-2 and were incubated at a temperature of 24 ± 1 °C and at controlled relative humidity (RH 65%). The experimental design included analyses at the time 0, 3, 6, 12, 24, 36, 48 and after every 24 h until the viral RNA was no longer detectable. The results showed a significant decrease (P < 0.001) in viral copy numbers on PFT within 3 h (24% reduction) and, at 72 h, the viral RNA had fallen below the limit of detection. Regarding the FPP, it was necessary to wait 24 h for a significant decrease (P = 0.015) in the viral load (14% reduction), while the detection threshold was reached at 96 h. These findings showed that the viral RNA persists longer on flow pack polyethylene samples than on polystyrene food trays, thus highlighting the importance of material characteristics in the persistence of SARS-CoV-2.

Determination of antibiotic residues in honey in relation to different potential sources and relevance for food inspection.

Honey contaminations could derive from intensive agriculture and industrial activities, but also from beekeeper treatments. In EU no MRLs for antibiotics in honey are set, only a minimum required performance limit for chloramphenicol of 0.3 µg kg-1 is recommended. Screening tests are available, characterised by their rapidity and simple use. Due to their high rate of false positives and the need to meet zero tolerance levels for antibiotics, their presence in samples was investigated using a liquid chromatography High Resolution Mass Spectrometry (LC-HRMS) multiclass antibiotic residue method, comparing the results with those of previous screening tests. The confirmatory method showed good sensitivity: CCα and CCß ranging from 0.03 to 4.80 ng g-1 and from 0.12 to 5.56 ng g-1, respectively. Ninety-eight honey samples from different geographical areas, analysed by two screening tests, showed a high percentage of false positives. This is fundamental to guarantee honey safety, especially, for organic production.

Occurrence of Listeria monocytogenes in ready-to-eat foods from supermarkets in Southern Italy.

The study provides data on the prevalence of Listeria monocytogenes in ready-to-eat (RTE) foods from supermarkets in Southern Italy. The pathogen was detected in 105/1045 (10%) RTE food samples. In particular, it was highlighted in 4/392 (1%) pastries, 23/112 (20.5%) vacuum-packaged sliced salami samples, 2/108 (1.9%) cream cheese samples, 31/115 (27%) mayonnaise based deli salads and 45/132 (34.1%) smoked salmon samples. The mozzarella samples were L. monocytogenes negative. Given the considerable public health implications, the study confirms that surveillance of listeriosis in Europe should be improved and coordinated between European Union Member States in order to better estimate the burden of disease and to prevent foodborne outbreaks, assessing the human health risk arising from RTE foods.

Evaluation of the Occurrence of False Aneurysms During Halal Slaughtering and Consequences on the Animal's State of Consciousness.

This study evaluated the occurrence of false aneurysms and reflexes in bovines, associated with the point along the neck where the cut was performed. The survey was carried out on a total of 1200 male beef cattle, belonging mainly to the Charolais and Limousin breeds, aged between twelve and twenty-four months. In the slaughterhouse, three operators, identified by convention as Operator C4, Operator C2, and Operator C1, performed the Halal slaughtering. Each operator carried out the slaughter of 400 animals and, on the basis of his own professional training, performed the cutting of the vessels at different points along the neck: Operator C4 between the third and fourth cervical vertebrae; Operator C2 at the level of the second cervical vertebra; and Operator C1 at the first cervical vertebra. The occurrence of false aneurysms was assessed on the basis of the different cutting site used by the three operators. Then, the evaluation of consciousness indicators, that is, rhythmic breathing and eye response, closely related to a slow bleeding process and to a delayed loss of brain function, was carried out 90 s post-bleeding. The group of cattle slaughtered by Operator C4 had a prevalence of false aneurysms of 10.25%. Conversely, the other two groups of cattle slaughtered by Operators C2 and C1 showed false aneurysm formation in 7.25% of each case. Further, 37.5% of the animals (18 for Operator C4, 17 for Operator C2, and 16 for Operator C1) with signs of consciousness 90 s after sticking were consequent to the onset of false aneurysms and, more precisely, they were owing to a late second cut of the vessel carried out by the operator, when false aneurysms occurred. The results of the research showed that the cutting point chosen by the operator is a key element in determining the onset of false aneurysms.

Classification of bovine carcasses: New biometric remote sensing tools.

Slaughtering plants approved by the European Union have specific processes to guarantee that beef carcasses or halfcarcasses, of no less than eight months of age, are provided with health mark and then classified and identified according to the EU carcass classification grid. This classification is based on three criteria: i) Category, ii) Conformation (SEUROP) and iii) Fat Cover (FC). At the end of the classification process each carcass is given a code, consisting of two letters and a number: this operation is called Identification. The aim of our study was to evaluate how the European beef carcass quality classification is determined according to the experience of the personnel involved, then comparing the results with those yielded by the Android platform application. West Systems, through its West- Zootech division, has developed an Android platform application (SEUROP APP) that allows SEUROP and FC classification with a smart-phone. The photo taken with the smart-phone will yield the necessary angular parameters to determine the conformation class depending on the animal's muscular mass and based on the convexity of some areas on the half-carcass. It also evaluates the ratio between surface of lean tissue and total carcass surface in order to determine the fat cover and complete the classification. The SEUROP APP was able to obtain objective measurements for as much as 84% of the assessments made during the research and development phase.

Pesticides and Environmental Contaminants in Organic Honeys According to Their Different Productive Areas toward Food Safety Protection.

Monitoring contaminant residues in honey helps to avoid risks to human health, as it is a natural product widely consumed in all population groups, including the most vulnerable, such as children and the elderly. This is important for organic honey production that may be negatively influenced by geographical area pollution. Considering the importance of collecting data on the occurrence of various xenobiotics in different geographical areas, this study aimed to investigate the presence of contaminant residues (persistent organic pollutants (POPs) and pesticides, including glyphosate and metabolites) in organic honey samples from different production areas using different analytical methods, in order to confirm their incidence and possible impact on the food safety traits of organic production. Regarding POPs, traces of benzofluoroanthene and chrysene were detected in honey from intensive orchards and arable lands. Traces of all polychlorobiphenyl (PCB) congeners were detected at different percentages in almost all of the samples, regardless of the origin area. Traces of polybromodiphenylethers (PBDE 28, 33, and 47) were found in different percentages of samples from all of the geographical areas examined. Traces of organochlorines (OCs) and organophosphates (OPs) were identified in honey samples belonging to all of the geographical areas. No glyphosate, glufosinate, and aminomethylphosphonic acid (AMPA) residues were detected.

Consumer attitudes towards animal welfare and their willingness to pay.

The interest of European consumers towards animal welfare can be influenced by several variables, both related to the consumers themselves and to the different countries of the EU. In order to assess animal welfare at farm level, it is essential to develop animal-based measures in accordance with the animals' actual welfare state in terms of their behaviour, health and physiology. The search for valid and reliable indicators is a key objective of several research programs especially for assessing welfare at farm level and the tools may include surveys addressed to farmers. However, there is a need to guarantee financial support for farmers who breed animals in accordance with such welfare conditions, to cover their additional costs. The aim of the study was to investigate the eating habits of Italian consumers regarding meat consumption linked to their knowledge of animal welfare and to their willingness to pay. We investigated consumers' understanding of animal welfare using a questionnaire (based on a list of twenty-three closed-ended questions) designed for collecting data from large numbers of respondents and multivariate statistical analysis. The data in our study showed that the variable with the greatest influence on purchase price was the place of meat purchase. As regards level of education, it appears that people with a high level of education are more concerned about animal welfare and, consequently, are willing to spend a higher price when buying meat. Consumer attention to the animal-welfare issue is on the rise and, in parallel with this growth, there is also a greater willingness to pay, i.e. a surcharge for the products obtained in the respect of animal welfare. This growth is influenced by the awareness and knowledge of the characteristics of animal welfare.

Genomic Characterization of Arcobacter butzleri Isolated From Shellfish: Novel Insight Into Antibiotic Resistance and Virulence Determinants.

Arcobacter (A.) butzleri is an emerging pathogenic microorganism, whose taxonomy has been recently suggested to be emended to the Aliarcobacter (Al.) butzleri comb. nov. Despite extensive taxonomic analysis, only few fragmented studies have investigated the occurrence and the prevalence of virulence and antibiotic resistance determinants of this species in strains isolated from shellfish. Herein we report for the first time the whole genome sequencing and genomic characterization of two A. butzleri strains isolated from shellfish, with particular reference to the antibiotic, heavy metals and virulence determinants. This study supported the taxonomic assignment of these strains to the Al. butzleri species, and allowed us to identify antibiotic and metal resistance along with virulence determinants, also additional to those previously reported for the only two A. butzleri strains from different environments genomically characterized. Moreover, both strains showed resistance to ß-lactams, vanocomycin, tetracycline and erythromycin and susceptibility to aminoglycosides and ciprofloxacin. Beside enlarging the availability of genomic data to perform comparative studies aimed at correlating phenotypic differences associated with ecological niche and geographic distribution with the genetic diversity of A. butzleri spp., this study reports the endowment of antibiotic and heavy metal resistance and virulence determinants of these shellfish-isolated strains. This leads to hypothesize a relatively high virulence of A. butzleri isolated from shellfish and prompt the need for a wider genomic analysis and for in vitro and in vivo studies of more strains isolated from this and other ecological niches, to unravel the mechanism of pathogenicity of this species, and the potential risk associated to their consumption.

Detection of a honeybee iflavirus with intermediate characteristics between kakugo virus and deformed wing virus.

Iflavirus RNA was detected in honeybee colonies displaying unduly aggressive behavior and with no evidence of morphological alterations. Sequence analysis of the RNA-dependent RNA polymerase (RdRp) revealed that the iflavirus strain was more similar (> 99% aa) to Deformed Wing Virus (DWV), that has been associated with morphological alterations in bees, rather than to the newly-described Kakugo Virus (KV) (about 95% aa), that has been associated with increased aggressiveness. Therefore, the iflavirus strain detected in the Italian hives genetically resembled DWV but was apparently associated with a KV-like phenotype. RT-PCR detected the iflavirus RNA in the abdomen of the workers, and only in one case was the virus detected in the head. No viral RNA was detected in the drones, a pattern of virus distribution across the honeybee casts that is in apparent conflict with the higher rates of infestation of drones by the mite Varroa distructor. The identification of a virus with apparently intermediate features between DWV and KV open new perspectives on the patho-biological role of iflaviruses in honeybees.

Simultaneous Quantitative Detection of Six Families of Antibiotics in Honey Using A Biochip Multi-Array Technology.

Chemical residues of veterinary drugs such as streptomycin, chloramphenicol, macrolides, sulphonamides, tetracyclines, quinolones and aminoglycosides and other contaminants such as pesticides and heavy metals have been found in honey, leading to concerns for human health. Indeed, there is a growing interest in their presence and persistence in the environment because low levels of antibiotics may favour the proliferation of antibiotic-resistant bacteria. Moreover, antibiotics present in honey may produce residues in foodstuffs, causing adverse effects on humans such as allergic reactions, toxic effects and damage to the central nervous systems. For food and health/safety reasons, antibiotic drugs are not authorized for the treatment of honey bees in the EU, even though these antimicrobial drugs have been approved in many third-party countries. For this reason, contaminated honey products can still be found in European markets. Therefore, there is a need to develop a precise, accurate and sensitive analytical method that may be used to simply and rapidly detect these compounds in honey. The aim of our study was to detect the presence of antibiotics in Apulian honey using the Anti-Microbial array II (AM II) as an innovative screening method to test the health quality of honey and honey products.