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In recent years, due to the large Romanian community present in Italy, the retail of foods coming from Eastern Europe has increased. The most common type of violation detected in these foods consists of incorrect labeling and species-replacement frauds. In this paper, the compliance of labels of 43 ethnic processed food coming from Eastern Europe and commercialized in Italy was evaluated by means of PCR and Sanger sequencing. Our data revealed 33% of non-compliant labels in samples containing swine, ruminants, and avian ingredients. These results demonstrate that PCR can be easily used for the identification of species in highly processed products, proving to be a rapid, effective, and economic method. On the other hand, samples reporting fish as ingredients highlighted the ineffectiveness of the applied sequencing protocol, due to the low informative property of targeted fragments or to the lack of consensus sequences in the case of uncommon species.

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Simple and rapid molecular detection technologies for authenticating animal species are urgently needed for food safety and authenticity. This study established a new direct-fast quantitative polymerase chain reaction (qPCR) detection technology for beef to achieve rapid and on-site nucleic acid detection in food. This technology can complete nucleic acid extraction in 4 min using a new type of food nucleic acid-releasing agent, followed by direct amplification of the DNA sample by fast qPCR in 25 min. The results indicated that direct-fast qPCR can specifically identify beef and can also identify 0.00001% of beef components in artificially simulated meat mixtures, with a detection precision variation coefficient of <4%. This method can be used to effectively identify beef in different food samples. As a simple, fast, and accurate molecular detection technology for beef, this method may provide a new tool for the on-site detection of beef components in food.

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Following a request from the European Commission, EFSA was asked to deliver a scientific opinion on the safety of sepiolite as a technological feed additive for all animal species. In 2022, the Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) delivered an Opinion on the safety and efficacy of the same additive. The Panel concluded that sepiolite used as a feed additive is safe for the consumers and the environment, and efficacious as a thickener-suspending agent, binder and anticaking agent in feed for all animal species under the proposed conditions of use. The additive was not considered an eye or skin irritant. However, it was considered a respiratory irritant, a respiratory and dermal sensitiser; owing to the dusting potential and its silica content, the additive was considered a risk by inhalation. Regarding the target species, in the previous Opinion, the Panel concluded on the safety of the additive for dairy ruminants. However, no conclusion could be drawn for all other species/categories. Based on the tolerance studies in chickens for fattening, weaned piglets and trout evaluated in the current assessment, and the one in dairy cows previously assessed, the Panel concluded that the inclusion of sepiolite at the maximum recommended level of 20,000 mg/kg complete feed is safe for all animal species.

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Different strains of Escherichia coli that exhibit genetic characteristics linked to diarrhea pose a major threat to both human and animal health. The purpose of this study was to determine the prevalence of pathogenic Escherichia coli (E. coli), the genetic linkages and routes of transmission between E. coli isolates from different animal species. The efficiency of disinfectants such as hydrogen peroxide (H2O2), Virkon®S, TH4+, nano zinc oxide (ZnO NPs), and H2O2-based zinc oxide nanoparticles (H2O2/ZnO NPs) against isolated strains of E. coli was evaluated. Using 100 fecal samples from different diarrheal species (cow n = 30, sheep n = 40, and broiler chicken n = 30) for E. coli isolation and identification using the entero-bacterial repetitive intergenic consensus (ERIC-PCR) fingerprinting technique. The E. coli properties isolated from several diarrheal species were examined for their pathogenicity in vitro. Scanning electron microscopy (SEM), high-resolution transmission electron microscopy (HR-TEM), Fourier-transform infrared spectrum (FT-IR), X-ray diffraction (XRD), zeta potential, and particle size distribution were used for the synthesis and characterization of ZnO NPs and H2O2/ZnO NPs. The broth macro-dilution method was used to assess the effectiveness of disinfectants and disinfectant-based nanoparticles against E. coli strains. Regarding the results, the hemolytic activity and Congo red binding assays of pathogenic E. coli isolates were 55.3 and 44.7%, respectively. Eleven virulent E. coli isolates were typed into five ERIC-types (A1, A2, B1, B2, and B3) using the ERIC-PCR method. These types clustered into two main clusters (A and B) with 75% similarity. In conclusion, there was 90% similarity between the sheep samples' ERIC types A1 and A2. On the other hand, 89% of the ERIC types B1, B2, and B3 of cows and poultry samples were comparable. The H2O2/ZnO NPs composite exhibits potential antibacterial action against E. coli isolates at 0.04 mg/ml after 120 min of exposure.

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Many different animal species are susceptible to SARS-CoV-2, including a few Canidae (domestic dog and raccoon dog). So far, only experimental evidence is available concerning SARS-CoV-2 infections in red foxes (Vulpes vulpes). This is the first report of SARS-CoV-2 RNA detection in a sample from a red fox. The RT-qPCR-positive fox was zoo-kept together with another fox and two bears in the Swiss Canton of Zurich. Combined material from a conjunctival and nasal swab collected for canine distemper virus diagnostics tested positive for SARS-CoV-2 RNA with Ct values of 36.9 (E gene assay) and 35.7 (RdRp gene assay). The sample was analysed for SARS-CoV-2 within a research project testing residual routine diagnostic samples from different animal species submitted between spring 2020 and December 2022 to improve knowledge on SARS-CoV-2 infections within different animal species and investigate their potential role in a One Health context. Within this project, 246 samples from 153 different animals from Swiss zoos and other wild animal species all tested SARS-CoV-2 RT-qPCR and/or serologically negative so far, except for the reported fox. The source of SARS-CoV-2 in the fox is unknown. The fox disappeared within the naturally structured enclosure, and the cadaver was not found. No further control measures were undertaken.

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Survival crises stalk many animals, especially endangered and rare animals. Accurate species identification plays a pivotal role in animal resource conservation. In this study, we developed an animal species identification method called Analysis of whole-GEnome (AGE), which identifies species by finding species-specific sequences through bioinformatics analysis of the whole genome and subsequently recognizing these sequences using experimental technologies. To clearly demonstrate the AGE method, Cervus nippon, a well-known endangered species, and a closely related species, Cervus elaphus, were set as model species, without and with published genomes, respectively. By analyzing the whole genomes of C. nippon and C. elaphus, which were obtained through next-generation sequencing and online databases, we built specific sequence databases containing 7,670,140 and 570,981 sequences, respectively. Then, the species specificities of the sequences were confirmed experimentally using Sanger sequencing and the CRISPR-Cas12a system. Moreover, for 11 fresh animal samples and 35 commercially available products, our results were in complete agreement with those of other authoritative identification methods, demonstrating AGE's precision and potential application. Notably, AGE found a mixture in the 35 commercially available products and successfully identified it. This study broadens the horizons of species identification using the whole genome and sheds light on the potential of AGE for conserving animal resources.

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A wide range of animal species show variable susceptibility to SARS-CoV-2; however, host factors associated with varied susceptibility remain to be defined. Here, we examined whether susceptibility to SARS-CoV-2 and virus tropism in different animal species are dependent on the expression and distribution of the virus receptor angiotensin-converting enzyme 2 (ACE2) and the host cell factor transmembrane serine protease 2 (TMPRSS2). We cataloged the upper and lower respiratory tract of multiple animal species and humans in a tissue-specific manner and quantitatively evaluated the distribution and abundance of ACE2 and TMPRSS2 mRNA in situ. Our results show that: (i) ACE2 and TMPRSS2 mRNA are abundant in the conduction portion of the respiratory tract, (ii) ACE2 mRNA occurs at a lower abundance compared to TMPRSS2 mRNA, (iii) co-expression of ACE2-TMPRSS2 mRNAs is highest in those species with the highest susceptibility to SARS-CoV-2 infection (i.e., cats, Syrian hamsters, and white-tailed deer), and (iv) expression of ACE2 and TMPRSS2 mRNA was not altered following SARS-CoV-2 infection. Our results demonstrate that while specific regions of the respiratory tract are enriched in ACE2 and TMPRSS2 mRNAs in different animal species, this is only a partial determinant of susceptibility to SARS-CoV-2 infection.IMPORTANCESARS-CoV-2 infects a wide array of domestic and wild animals, raising concerns regarding its evolutionary dynamics in animals and potential for spillback transmission of emerging variants to humans. Hence, SARS-CoV-2 infection in animals has significant public health relevance. Host factors determining animal susceptibility to SARS-CoV-2 are vastly unknown, and their characterization is critical to further understand susceptibility and viral dynamics in animal populations and anticipate potential spillback transmission. Here, we quantitatively assessed the distribution and abundance of the two most important host factors, angiotensin-converting enzyme 2 and transmembrane serine protease 2, in the respiratory tract of various animal species and humans. Our results demonstrate that while specific regions of the respiratory tract are enriched in these two host factors, they are only partial determinants of susceptibility. Detailed analysis of additional host factors is critical for our understanding of the underlying mechanisms governing viral susceptibility and reservoir hosts.

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Following a request from the European Commission, EFSA was asked to deliver a scientific opinion on the assessment of the application for the renewal of the authorisation of folic acid as a nutritional feed additive. The additive is authorised for use in feed and water for drinking for all animal species. The applicant provided evidence that the additive currently in the market complies with the existing conditions of authorisation and the production process has not been modified. The FEEDAP Panel considers that there is no evidence to revise the conclusions reached in the previous assessment for the safety for the terrestrial species, consumers and for the environment. The use of folic acid in aquatic animal species to cover their nutritional needs is considered safe. However, the Panel is not in a position to set a maximum safe level for all fish and crustacean species. Considering the narrow margin between the requirement and the tolerated levels seen in some aquatic animal species, the FEEDAP Panel considers that supplementation should not exceed the requirements of the different aquatic animal species. The additive is neither a skin irritant nor a dermal sensitiser. The exposure through inhalation is likely. Due to the lack of data, the FEEDAP Panel is not in the position to conclude on the potential of folic acid to be harmful to the respiratory system and irritant to eyes. The Panel retains that the previously made conclusion on the efficacy remains valid.

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Following a request from the European Commission, EFSA was asked to deliver a scientific opinion on the safety and efficacy of Levilactobacillus brevis (formerly Lactobacillus brevis) DSM 23231 as a technological feed additive for all animal species. The Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) concluded that evidence has been provided that the additive currently on the market complies with the existing terms of authorisation. The Panel also concluded that L. brevis DSM 23231 remains safe for all animal species, consumers and the environment under the authorised conditions of use. The additive should be considered a respiratory sensitiser. Based on the studies submitted regarding user safety, the preparation of the additive tested was shown not to be a skin or eye irritant. The Panel was not in the position to conclude on the skin sensitisation potential of the additive. There is no need for assessing the efficacy of the additive in the context of the renewal of the authorisation.

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Laboratory testing methods to confirm the identity of meat products and eliminate food fraud regularly rely on PCR amplification of extracted DNA, with most published assays detecting mitochondrial sequences, providing sensitive presence/absence results. By targeting single-copy nuclear targets instead, relative quantification measurements are achievable, providing additional information on the proportions of meat species detected. In this Methods paper, new assays for horse, donkey, duck, kangaroo, camel, water buffalo and crocodile have been developed to expand the range of species that can be quantified, and a previously published reference assay targeting the myostatin gene has been modified to include marsupials and reptiles. The accuracy of this ratio measurement approach was demonstrated using dPCR with mixtures of meat DNA down to 0.1%. However, the limit of detection (LOD) of this approach is not just determined by the assay targets, but by the samples themselves, with food or feed ingredients and processing impacting the DNA yield and integrity. In routine testing settings, the myostatin assay can provide multiple quality control roles, including monitoring the yield and purity of extracted DNA, identifying the presence of additional meats not detected by the suite of species-specific assays and potentially estimating a sample-specific LOD based on measured copy numbers of the myostatin target. In addition to the myostatin positive control assay, a synthetic DNA reference material (RM) has been designed, containing PCR targets for beef, pork, sheep, chicken, goat, kangaroo, horse, water buffalo and myostatin, to be used as a positive template control. The availability of standardised measurement methods and associated RMs significantly improves the reliability, comparability and transparency of laboratory testing, leading to greater confidence in results.

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The aim of this study was to investigate the changes in human and animal milk oligosaccharides over lactation. In total, 89, 97, 115, and 71 oligosaccharides were identified in human, bovine, goat, and camel milk. The number of common oligosaccharides between camel and human milk was the highest (16 and 17 in transitional and mature milk). With respect to the absolute concentration of eight oligosaccharides (2'-FL, 3-FL, α3'-GL, LNT, LNnT, 3'-SL, 6'-SL, and DSL), 2'-FL, 3'-FL, LNT, and LNnT were much higher in human than three animal species. 3'-SL had a similar concentration in bovine colostrum (322.2 µg/mL) and human colostrum (321.0 µg/mL), followed by goat colostrum (105.1 µg/mL); however, it had the highest concentration in camel mature milk (304.5 µg/mL). The ratio of 6'-SL and 3'-SL (1.77) in goat colostrum was similar to that in human colostrum (1.68), followed by bovine colostrum (0.13). In terms of changes of eight oligosaccharides over lactation, they all decreased with the increase of lactation in bovine and goat milk; however, α3'-GL, 2'-FL, and 3-FL increased in camel species, and LNT increased first and then decreased over lactation in human milk. This study provides a better understanding of the variation of milk oligosaccharides related to lactation and species.

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GeneFields®-Hair is a simple analysis kit that uses nucleic acid chromatography and polymerase chain reaction (PCR) to identify animal species of hair-like food contaminants. In this study, we evaluated GeneFields®-Hair as a simple and rapid method for identifying animal species from hair-like materials collected in forensic science, such as at crime scenes. The use of this kit with other human biological materials (whole blood, head dandruff, nails, saliva, oral mucosa, sebum, and urine) was also investigated. Animal body hair samples were pretreated by grinding in a buffer solution, centrifuged, and the supernatant was used for PCR. Nucleic acid chromatography of the PCR products allowed the identification of the animal species by the presence or absence of coloration on the decision line. For human biological materials, nucleic acid chromatography was performed after the appropriate pretreatment like body hair material. The determination of some animal species was difficult, even if they had a dedicated DNA Strip determination line. Furthermore, animals from the same family but different genera were sometimes detected on the same determination line. All the human biological samples were correctly identified. Smartphone photographs of the coloration of the judgment line were processed using the ImageJ software for quantitative determination.

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Typically much smaller in number than their mainland counterparts, island populations are ideal systems to investigate genetic threats to small populations. The Svalbard reindeer (Rangifer tarandus platyrhynchus) is an endemic subspecies that colonized the Svalbard archipelago ca. 6,000-8,000 years ago and now shows numerous physiological and morphological adaptations to its arctic habitat. Here, we report a de-novo chromosome-level assembly for Svalbard reindeer and analyze 133 reindeer genomes spanning Svalbard and most of the species' Holarctic range, to examine the genomic consequences of long-term isolation and small population size in this insular subspecies. Empirical data, demographic reconstructions, and forward simulations show that long-term isolation and high inbreeding levels may have facilitated the reduction of highly deleterious-and to a lesser extent, moderately deleterious-variation. Our study indicates that long-term reduced genetic diversity did not preclude local adaptation to the High Arctic, suggesting that even severely bottlenecked populations can retain evolutionary potential.

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Human activities increasingly challenge wild animal populations by disrupting ecological connectivity and population persistence. Yet, human-modified habitats can provide resources, resulting in selection of disturbed areas by generalist species. To investigate spatial and temporal responses of a generalist carnivore to human disturbance, we investigated habitat selection and diel activity patterns in caracals (Caracal caracal). We GPS-collared 25 adults and subadults in urban and wildland-dominated subregions in Cape Town, South Africa. Selection responses for landscape variables were dependent on subregion, animal age class, and diel period. Contrary to expectations, caracals did not become more nocturnal in urban areas. Caracals increased their selection for proximity to urban areas as the proportion of urban area increased. Differences in habitat selection between urban and wildland caracals suggest that individuals of this generalist species exhibit high behavioral flexibility in response to anthropogenic disturbances that emerge as a function of habitat context.

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Following a request from the European Commission, the Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on concentrated liquid l-lysine, l-lysine monohydrochloride and concentrated liquid l-lysine monohydrochloride produced by a genetically modified strain of Escherichia coli (NITE BP-02917) as nutritional and as sensory (flavouring compound) feed additives for all animal species. In 2022, the FEEDAP Panel issued an opinion on the safety and efficacy of these products. In that assessment, the FEEDAP Panel could not exclude the potential presence of recombinant DNA derived from the genetically modified production organism in the products. The applicant provided supplementary data to exclude the presence of recombinant DNA derived from the production organism in the final products. Based on the new data provided, the FEEDAP Panel concluded that no DNA of the production strain E. coli NITE BP-02917 was detected in concentrated liquid l-lysine, l-lysine monohydrochloride and concentrated liquid l-lysine monohydrochloride.

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Continental East Asia has a mild Pleistocene climate and a complex recent geological history. Phylogeographic studies of animals over the last 30 years have produced several distinctive patterns. Glaciation refugia are numerous and are not restricted to any particular regions. Most of them are localized and species-specific, although several large refugia, for example the mountains of SW China, are shared by multiple species and have refugia-within-refugia. Furthermore, postglaciation range expansion events vary greatly in time, scale and direction. Large-scale south-to-north post-LGM expansions are few and mostly occurred in the northern regions. Additionally, several unique geographic features, including the three-step terrain of China and the northern arid belt, have significant impacts on many species histories. Overall, the impacts of Pleistocene glaciations, particularly the LGM, on species history vary drastically from nondetectable to significant. The impacts are the least for species from the southwestern region and are most dominant for species from the north. Geological events play a more significant role in shaping species history than Pleistocene climatic changes. Phylogeographic patterns among animals species are highly consistent with those of plants. Future phylogeographic endeavour in East Asia should be hypothesis-driven and seek processes that underlie common patterns. The wide use of genomic data allow accurate estimates of historical population processes and exploration of older history beyond the Pleistocene.

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Social and academic interest in animal abuse has recently increased thanks to greater awareness of the importance of biodiversity in promoting sustainability. The redefinition of human-animal relationships, in the context of the fight against speciesism and the defense of veganism, has also contributed to this greater attention. Moreover, public awareness of animal rights has strengthened social reactions to violence against animals, though there are still some social sectors that are indifferent to these changes. Thus, better knowledge of the psychological mechanisms underlying reactions to animal abuse could contribute to better informal, social control of such abuse. The main aim of this study is to analyze the relationships between psychopathy, empathy with people, and empathy with nature arising from people's reactions to protected and domestic animal abuse and illegal dumping. Also, as previous studies have shown differences between men and women, both in animal abuse and in personality traits, thus gender is taken into account when analyzing these relationships. A total of 409 people, resident in a highly environmentally protected territory, participated in this study. They were aged between 18 and 82 years old and 49.9% women. Participants were asked about assigned punishments, as well as the probability of intervening personally and/or calling the police, in relation to ten scenarios, based on press releases, describing one of three types of transgression of environmental laws: abuse of protected animals, abuse of domestic animals or illegal dumping. They also responded to Spanish adaptations of the Inventory of Callous Unemotional Traits, the Basic Empathy Scale, the Dispositional Empathy with Nature Scale, and the Social Desirability Scale. Each participant was randomly given ten scenarios corresponding to just one transgression type but all the personality scales. Results show that people's reactions were greater for abuse of domestic than protected animals or for illegal dumping, irrespective of gender. Empathy with nature was related to the reaction against animal abuse more than empathy with people and psychopathy. Results are discussed highlighting the need for future research into similarities and differences between animal abuse and other types of environmental offences, which have many victims but no single being suffering.

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Glutathione S-transferases (GSTs) are conjugating enzymes involved in drug metabolism, antioxidant defence, and cell signalling. Herein, we investigated hepatic GST conjugation in several mouse and rat strains, including both sexes, with a direct comparison to humans.Using general and isoform-selective substrates, all mouse strains had significantly greater activities than humans for total cytosolic GST, GST-M, GST-T, and microsomal GST activities. Some strains had significantly greater GST-P activities compared to humans. Sex differences between males and females were evident in all strains for total cytosolic GST, GST-M, and GST-P, and sex differences in GST-T and microsomal GST activities within strains were noted.All rats had significantly greater activities than humans for GST-M and GST-T; only some strains were significantly greater than humans for GST-P, total cytosolic GST, and microsomal GST. Sex differences within strains showed significantly greater GST-M and GST-T activities in males compared to females. Select strains showed sex differences for total cytosolic and microsomal GST activities; there were no sex differences in GST-P activities.Significant differences in glutathione conjugation between humans and rodents exist, including sex differences. This highlights the need for careful animal selection in pre-clinical studies where GSTs are the primary metabolic pathway.

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Following a request from the European Commission, the EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on the safety of lignosulphonate, when used as a technological additive (functional group: binders) in feed for all animal species. In two previous assessments, the FEEDAP Panel concluded that the maximum level proposed of 10,000 mg lignosulphonate/kg complete feed is safe for weaned piglets, pigs for fattening, chickens for fattening, laying hens and cattle for fattening, but a margin of safety could not be identified. Consequently, these conclusions could not be extrapolated to other animal species/categories. In the current application, the applicant proposed to reduce the maximum content of lignosulphonate in feed for all animal species to 8,000 mg/kg complete feed. The results of two newly submitted tolerance studies allowed the Panel to conclude that 8,000 mg lignosulphonate/kg complete feed is also safe for dairy cows and salmonids, with a margin of safety of at least 1.25. Considering the results of the studies previously assessed and those of the two new tolerance studies, the FEEDAP Panel concluded that lignosulphonate is safe for all animal species when used at a maximum content of 8,000 mg/kg complete feed.

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Following a request from the European Commission, the Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on the safety of disodium 5'-inosinate (also known as inosine monophosphate, IMP) produced by Corynebacterium stationis KCCM 80235 as a sensory additive, functional group flavouring compounds, for all animal species. In 2022, the FEEDAP Panel issued an opinion on the safety and efficacy of the product. In that assessment, the Panel could not conclude on the safety of the additive for the target species, consumers, users and the environment due to uncertainties on the genetic basis of the streptomycin resistance of the production strain C. stationis KCCM 80235 and on the possible presence of its recombinant DNA in the final product. The applicant provided supplementary data which elucidated the genetic basis of the streptomycin resistance of the production strain and excluded the presence of its DNA in the final product. Therefore, the FEEDAP Panel concluded that IMP produced by C. stationis KCCM 80235 is safe for the target species, consumers, users and the environment. The FEEDAP Panel reiterated its previous concerns on the safety for the target species when using IMP in water for drinking due to hygienic reasons.