1. Investigación

Visceral leishmaniosis is the one of the most important protozoal zoonoses in Europe, and it is caused by Leishmania infantum, an intracellular protozoan parasite. The disease is endemic in dogs in the Mediterranean area. The main goal of this work is to correlate the levels of several cytokines linked to immune response against L. infantum infection in two canine breeds. Thirty-one Boxer and twenty-eight Ibizan Hound dogs living in the Valencian Community (East coast of Spain) were analyzed for the presence of anti-Leishmania antibodies in serum by IFAT test. Cytokines IFN-γ, TNF-α, IL-2, IL-6, IL-8, and IL-18 were determined by ELISA commercial tests. The levels of IFN-γ, IL-2, and IL-18 in our study, cytokines linked to a cellular immune response, were higher (p < 0.05) in the Ibizan Hound breed; IL-6 levels were higher, although not significant, and only levels of IL-8 were higher in Boxer than in Ibizan Hound. No expression of TNF-α was found. These results corroborate that Ibizan Hound can develop a protective response against canine leishmaniosis, while Boxer is a susceptible breed. The study of immunological aspects in the different canine breeds may represent a useful tool in the prediction of the disease.

Precision nutrition in broilers requires tools capable of identifying amino acid imbalances individually or in groups, as well as knowledge on how more digestible proteins can be designed for innovative feeding programs adjusted to animals’ dynamic requirements. This work proposes two potential tools, combining traditional nutrition with biotechnological, metabolomic, computational and protein engineering knowledge, which can contribute to improving the precise amino acid nutrition of broilers in the future: (i) the use of serum uric nitrogen content as a rapid biomarker of amino acid imbalances, and (ii) the design and modeling of de novo proteins that are fully digestible and fit exactly to the animal’s requirements. Each application is illustrated with a case study. Case study 1 demonstrates that serum uric nitrogen can be a useful rapid indicator of individual or group amino acid deficiencies or imbalances when reducing dietary protein and adjusting the valine and arginine to lysine ratios in broilers. Case study 2 describes a stepwise approach to design an ideal protein, resulting in a potential amino acid sequence and structure prototype that is ideally adjusted to the requirements of the targeted animal, and is theoretically completely digestible. Both tools can open up new opportunities to form an integrated framework for precise amino acid nutrition in broilers, helping us to achieve more efficient, resilient, and sustainable production. This information can help to determine the exact ratio of amino acids that will improve the efficiency of the use of nitrogen by poultry.

The ideal protein concept refers to dietary protein with an amino acid profile that exactly meets an animal’s requirement. Low-quality protein levels in the diet have negative implications for productive and reproductive traits, and a protein oversupply is energetically costly and leads to an excessive N excretion, with potentially negative environmental impact. Urea Nitrogen (UN), which corresponds to the amount of nitrogen in the form of urea circulating in the bloodstream, is a metabolite that has been widely used to detect amino acid imbalances and deficiencies and protein requirements. This review aims to critically analyse how UN can contribute to accurately implementing the ideal protein concept in monogastric animals, particularly in pig, poultry, and rabbit nutrition (14,000 animals from 76 published trials). About 59, 37, and 4% of trials have been conducted in pigs, poultry, and rabbits, respectively. UN level was negatively correlated to main performance traits (Pearson Correlation Coefficient [PCC] of 􀀀0.98 and 􀀀0.76, for average daily gain and feed conversion ratio, respectively), and lower UN level was related to higher milk yield and concentration. High level of UN was positively correlated to N excretion (PCC = 0.99) and negatively correlated to protein retention (PCC = 􀀀0.99). Therefore, UN in blood seems to be a proper indicator of amino acid imbalance in monogastric animals. Great variability in the use of UN was observed in the literature, including uses as determination medium (blood, plasma, or serum), units, and feeding system used (ad libitum or restricted), among others. A standardization of the methods in each of the species, with the aim to harmonize comparison among works, is suggested. After review, UN measurement in plasma and, whenever possible, the utilization of the same nutritional methodology (ad libitum conditions or restriction with blood sampling after refeeding at standardised time) are recommended. More studies are necessary to know the potential of UN and other bioindicators for amino acid deficiencies evaluation to get closer to the ideal protein concept.

European wild rabbit (Oryctolagus cuniculus) has been defined as a keystone species in the Mediterranean ecosystem. Rabbits have been classed as “endangered” by the IUCN within their native range. In this sense, animal nutrition may play a fundamental and limiting role in the conservation of wild species. The overarching goal of ecological nutrition is to unravel the extensive web of nutritional links that direct animals in their interactions with their ecological environments. The main aim of this work was to evaluate the effect of different feed intake, geographic location, animal sex, and reproductive stage on glucose, non-esterified fatty acids (NEFA), and plasmatic urea nitrogen (PUN), albumin, glutamate, and total protein metabolites. Additionally, we examined the potential of these metabolites as biomarkers. Full stomach contents and blood samples were collected from European wild rabbits (n = 89) for the analysis of the metabolites described above. Our work shows that the levels of these metabolites are affected by the sex of the animals, as well as by their reproductive stage (glucose, NEFA and albumin). There were signs of better optimisation of resources by females than by other groups of animals. These data may be interesting in the study of nutritional components that could be affecting physiological state of this species.

European wild rabbit (Oryctolagus cuniculus) populations have drastically reduced, and recently, rabbits have been classed as “endangered” by the IUCN. This animal plays an important ecological role in Mediterranean ecosystems and its introduction could significantly contribute to ecological restoration. Rabbits have high nutrient requirements that apparently cannot be covered in all ecosystems, and there are clues that nutrition can limit their abundance and density. On the other hand, some studies reflect the effects of food availability on the metabolomic status of other animal species, but there are no specific studies on this keystone species. The main aim of this work is to find biomarkers to assess the previous levels of ingestion of European rabbits (Oryctolagus cuniculus). To address this gap, gastric content and blood samples were collected from European rabbits (n = 99) in a Mediterranean area for the analysis of glucose, non-esterified fatty acids (NEFA), plasmatic urea nitrogen (PUN), albumin, glutamate and total protein metabolites. Depending on their previous feed intake (gastric content and the ratio between the gastric content and the weight of the animal), the animals were divided into two groups (lower and normal previous feed intake). Our work shows that the metabolomic profiles of the animals were affected. Levels of glucose (+82%; p = 0.0003), NEFA (􀀀61%; p = 0.0040) and PUN (+139%; p < 0.001) were different in the animals with lower previous feed intake than the animals with normal previous feed intake. This work summarises that metabolic phenotype can be interesting when seeking to discover the limiting nutrients and food availability in diets that could affect the ecological fitness and conservation of European wild rabbits. It is important to mention that in this work, only the effects on six different metabolites have been analysed and more studies are necessary to complement the knowledge of possible metabolites that indicate the level of ingestion in this species and others. These (and new) biomarkers could be used as a tool to provide information about individual or population characteristics that other physiological parameters cannot detect, improving the conservation physiology field.

Background: The Ibizan Hound is a canine breed native to the Mediterranean region, where leishmaniosis is an endemic zoonosis. Several studies indicate low prevalence of this disease in these dogs but the underlying molecular mechanism remains unknown. Methods: In this study, qualitative immunological and genomic profiles of this breed have been analyzed. Results: Our analysis shows relevant differences between the cytokine serum profile of Ibizan Hound dogs and previously published data from other canine strains. Additionally, several genetic risk variants related to the immune response, regulation of the immune system, and genes encoding cytokines and their receptors have been studied. The most relevant genes that presented such fixed polymorphisms were IFNG and IL6R. Other variants with frequencies ≥ 0.7 were found in the genes ARHGAP18, DAPK1, GNAI2, MITF, IL12RB1, LTBP1, SCL28A3, SCL35D2, PTPN22, CIITA, THEMIS, and CD180. Epigenetic regulatory genes such as HEY2 and L3MBTL3 showed also intronic polymorphisms. Conclusions: Our analysis and results indicate that the regulation of immune responses is different in Ibizan Hounds compared to other breeds. Future studies are needed to elucidate whether these differences are related to the low prevalence of L. infantum infection in the Ibizan Hound.

Leishmaniosis, one of themost important zoonoses in Europe, is caused by Leishmania infantum, an intracellular protozoan parasite. This disease is endemic in theMediterranean area, where the main reservoir is the dog. Several studies indicate a possible susceptibility to L. infantum infection with clinical signs in some canine breeds. One of them is the boxer breed, which shows a high prevalence of disease. In this study, immunological and genomic characterization of serum samples from boxer dogs living in theMediterranean area were evaluated to analyze the immune response and the possible genetic explanation for this susceptibility. Serum levels of cytokines IFN- , IL-2, IL-6, IL-8, and IL-18 were determined by ELISA commercial tests, while the genotyping study was performed using the CanineHD DNA Analysis BeadChip. The results show relevant differences in the serum levels of cytokines compared to published data on other canine breeds, as well as sequence changes that could explain the high susceptibility of the boxer breed to the disease. Concretely, polymorphic variants in the CIITA, HSF2BP, LTBP1,MITF, NOXA1, PKIB, RAB38, RASEF, TLE1, and TLR4 genes were found, which could explain the susceptibility of this breed to L. infantum infection.

Toxoplasma gondii is an obligate intracellular parasite that causes toxoplasmosis, with approximately one third of the population around the world seropositive. The consumption of contaminated food is the main source of infection. These include meat products with T. gondii tissue cysts, and dairy products with tachyzoites. Recently, contamination has been detected in fresh products with oocysts and marine products. Despite the great health problems that are caused by T. gondii, currently there are no standardized methods for its detection in the food industry. In this review, we analyze the current detection methods, the prevalence of T. gondii in different food products, and the control measures. The main detection methods are bioassays, cell culture, molecular and microscopic techniques, and serological methods, but some of these do not have applicability in the food industry. As a result, emerging techniques are being developed that are aimed at the detection of multiple parasites simultaneously that would make their application more efficient in the industry. Since the prevalence of this parasite is high in many products (meat and milk, marine products, and vegetables), it is necessary to standardize detection methods, as well as implement control measures.

Honeybee populations have locally and temporally declined in the last few years because of both biotic and abiotic factors. Among the latter, one of the most important reasons is infection by the microsporidia Nosema ceranae, which is the etiological agent of type C nosemosis. This species was first described in Asian honeybees (Apis cerana). Nowadays, domestic honeybees (Apis mellifera) worldwide are also becoming infected due to globalization. Type C nosemosis can be asymptomatic or can cause important damage to bees, such as changes in temporal polyethism, energy and oxidative stress, immunity loss, and decreased average life expectancy. It causes drastic reductions in workers, numbers of broods, and honey production, finally leading to colony loss. Common treatment is based on fumagillin, an antibiotic with side effects and relatively poor efficiency, which is banned in the European Union. Natural products, probiotics, food supplements, nutraceuticals, and other veterinary drugs are currently under study and might represent alternative treatments. Prophylaxis and management of affected colonies are essential to control the disease. While N. ceranae is one potential cause of bee losses in a colony, other factors must also be considered, especially synergies between microsporidia and the use of insecticides.

Different university degrees focus on students acquiring theoretical and practical knowledge, aiming to develop their professional activity in the future. However, the usual study plans often forget other skills that will be very useful for the correct performance of their professional activity. In the case of veterinarians, these can range from dialogue with farmers to the unification of knowledge, so that they can provide a simple and effective solution to the different questions that may arise throughout their work activity. On the other hand, the perception of the world and the ways of acquiring knowledge have been changing over the years. Currently, our students require new ways of being presented with the information and knowledge that they should acquire, using, in most cases, new technologies. The present study was carried out with two cases. First, we used gamification through role-play as an alternative methodology to generate a method to unify the knowledge acquired in the subject and, mainly, to acquire skills such as the transfer of this acquired knowledge to other classes and situations. The second case aims to verify if the use of new technologies, specifically the use of interactive videos, can improve the acceptance of students and their training. A total of 2 h of videos were recorded, and 31 min and 42 s of that footage were ultimately used. A special edition and some specific illustrations and designs were made for this work, taking care of the format–background relationship. The results obtained show that these alternative-learning methodologies could be applied to many subjects, so that students, in a playful and relaxed way, are able to unify all the knowledge they are acquiring in their training as veterinarians, preparing them to face the exercise of their future professional activity with greater ease and safety. Finally, we provide the degree of acceptance of these new learning methodologies by students.