Until now a few studies have been carried out on the gut lymphoid system in fish despite its protective role in the host. Here, we have evaluated the effects of Candida albicans (Ca) and lipopolysaccaridae (LPS) on the pyloric and terminal segments of gut in the rainbow trout Oncorhynchus mykiss. In particular, data show that both Ca and LPS are able to cause apoptosis of intestinal lymphoid cells as detected by the TUNEL procedure. These findings suggest a further modality of gut response in fish to environment antigens.

The knowledge of gametogenesis is of paramount importance to develop a reliable technology for Atlantic bluefin tuna (Thunnus thynnus L.) (ABFT) rearing in captivity. The aims of this study were: a) to evaluate the capacity of male ABFT, confined in captivity before puberty, to finalize spermatogenesis; b) to compare germ cell proliferation between wild and captive ABFT. Testis samples were taken from: a) 13 juvenile ABFT reared in the North Adriatic Sea (Croatia); b) 30 adult ABFT reared in the central and western Mediterranean (Spain, Malta and Italy); c) 20 adult wild ABFT captured by tuna traps in Italy and Morocco. Samples were fixed in 10% formalin, dehydrated in ethanol and embedded in paraffin. Proliferating germ cells were identified through the immunohistochemical detection of proliferating cell nuclear antigen (PCNA). The first spiniform ray of the first dorsal spine was taken from the juvenile fish in order to estimate the age through the count of annual discontinuities. Juvenile ABFT captured before puberty were able to finalize spermatogenesis starting from 3 years of age. Germ cell proliferation was delayed in captive-reared ABFT specimen compared to wild individuals. These results seem to indicate that testis maturation can be anticipated in ABFT caught before puberty, but spermatogenesis is somewhat damaged in adult fish reared in captivity compared to wild individuals.

In all Equestrian sports, the welfare of the horse must be paramount and must never be subordinated to competitive or commercial influences. It is necessary to prohibit any training methods which are abusive or cause fear or for which the horse has not been properly trained. The purpose of this study was to examine the effects of prolonged and intensive physical training on the immune response in trotter horses in order to recommend at the legislator the adoption of guidelines for the welfare of these animals. In fact, during their athletic life and some of them undergo lethal lung infections, therefore it is likely that modifications of physiologic cellular parameters could account for the increased susceptibility to microbial disease. Noteworthy, chronic stress has been shown to be immunosuppressive, whereas acute stress seems to lead to immunoenhancing effects. In particular, we have studied some immune parameters as blood cells distribution, hemoglobin, hematocrit, erythrocyte sedimentation rate, phagocytosis activity, macrophage Migration Inhibiting Factor (MIF) and finally the levels of (1-3)-ß-D-Glucan, as indicator of clearance. Taken together, these findings indicate a condition of reduced immune response in seven trotters after race, to identify possible biomarkers of stress dependent on physical exercise.

The European anchovy, Engraulis encrasicolus, is a multiple-spawning small pelagic fish with a comparatively long reproductive season. From April to October 2009, ovary samples were collected from individuals of the southwestern Adriatic Sea in order to examine ovarian histological changes and assess batch fecundity monthly variations throughout the whole reproductive season. To assess monthly variations of the relative batch fecundity, the correlation between batch fecundity (F) - i.e. the number of oocytes released at each spawning act - and ovary-free body mass (W*) was tested by four regression models; the power equation () was found to be the most suitable to describe correlations. The reproductive season of the anchovy of the central-southern Adriatic population lasts from May to September; in this period, all the oocyte development stages were observed, including hydrated oocytes and postovulatory follicles. In April, most fish had only unyolked oocytes; in October, an extensive atresia of yolked follicles was observed. The slope of all the on monthly regressions did not differ significantly from 1, which shows that relative batch fecundity is constant all over the anchovy size range, throughout the spawning season. In the central-southern Adriatic anchovy population, batch fecundity increased from May to July and then gradually decreased until September. Differences in batch fecundity of the anchovy from different areas of the eastern Atlantic and Mediterranean could possibly be due to both environmental parameters and genetic differences among the different populations.

Contractions of ovarian tunica albuginea, the teleostean cystovary wall layer containing smooth muscle fibres, facilitate oocytes and fluids movements within the ovary, oocytes ovulation and spawning. Fish isotocin, the homologue hormone of mammalian oxytocin, plays a significant role in ovulation, oviduct contraction and spawning. In the present study, ovarian wall spontaneous contraction, as well as isotocin in vitro effect on tunica albuginea contractility, was analysed in female seabream in different reproductive conditions: vitellogenesis, regressing (post-spawning) and extensive atresia. Tunica albuginea spontaneous contractility was recorded using ovary wall strips mounted in an organ bath containing modified Ringer's solution. The strips were then exposed to cumulative doses of isotocin (6, 30, 60 μg/ml). Female seabream in regressing condition exhibited the highest level of tunica albuginea spontaneous contraction amplitude compared with the other two groups. Only fish in vitellogenesis state showed a significant increase in contraction amplitude after isotocin administration at the dose of 30 μg/ml. The same group exhibited also a significant isotocin dose-dependent decrease in the contractile frequency. These results confirm the involvement of isotocin in stimulating tunica albuginea contractile activity during the oestrogen-regulated phase of vitellogenesis, whereas the absence of significant effects of isotocin on ovarian contractility in fish at the regressing state might be ascribed to the occurrence of a contractile activity autonomously regulated by the internal pacemaker system. The absence of exposed isotocin receptors could explain the lack of effects of the isotocin administration in seabream showed extensive atresia of the follicular cells.

The Atlantic bluefin tuna Thunnus thynnus (ABFT) is intensely fished in the Mediterranean Sea to supply a prosperous capture-based mariculture industry. Liver apoptotic structures and tumor necrosis factor (TNF) gene expression were determined in: wild ABFT caught in the eastern Atlantic; juvenile ABFT reared in the central Adriatic Sea; juvenile ABFT reared in the northern Adriatic Sea; adult ABFT reared in the western Mediterranean. The highest density of liver apoptotic structures was found in the juveniles from the northern Adriatic. Two partial TNF cDNAs (TNF1 and TNF2) were cloned and sequenced. TNF1 gene expression was higher in juveniles than in adults. The highest expression of TNF2 was found in the juveniles from the northern Adriatic. These findings might be related to the juvenile exposure to environmental pollutants.

Ivermectin (IVM) is a macrocyclic disaccharide anthelmintic agent with broad-spectrum antiparasitic action. It is used for controlling internal and external parasites in food-producing animals (bovine, swine and equine species), including donkey. This drug is included in Council Regulation (EEC) 2377/9, annex I, with Maximum Residue Limit (MRL) ranging from 15 to 100 μg kg−1 depending on the species. The pharmacokinetic characteristics of IVM mainly depend on the animal species, formulation and route of administration. Due to its high lipophilicity it accumulates in milk of food producing animals. Detailed IVM pharmacokinetic studies were carried out for horse, pig, cattle, sheep, and goat, but there is lack of information for asinine species. Consequently, the absence of “label milk withdrawal time” and MRLs in donkey’s milk indicates that IVM should not be used in lactating animals. On the contrary, information about IVM residues in milk would be of paramount importance in view of the fact that donkey’s milk is a suggested substitute for infants suffering from hypersensitivity to cow’s, sheep’s and goat’s milk protein or from multiple hypersensitivity. European legislation does not explicitly state that an MRL should be determined for every target species; however it’s interpretation led to MRLs being adopted for each species and each foodstuff. Although IVM is approved for use in all lactating animal species the possibility of an illegal use should be considered. In this paper, the problems arising from IVM administration in lactating donkey are analysed. A particular attention is paid for the existing legislation and some proposals are suggested for regulatory framework formulation.

Melanomacrophage centres (MMCs), located in different organs of non-mammalian vertebrates, play a role in the destruction, detoxification or recycling of endogenous and exogenous materials. Cytochrome P450 monoxygenase 1A (CYP1A) is involved in xenobiotics biotransformation, and its liver expression is considered as a biomarker for detecting exposure to environmental pollutants. Atlantic bluefin tuna (ABFT), Thunnus thynnus L., liver samples were collected from: wild animals caught in the eastern Atlantic; juveniles reared in the central Adriatic; juveniles reared in the northern Adriatic; adults reared in the western Mediterranean. The samples were processed for basic histology, histochemistry and for CYP1A immunodetection. An unexpected high density of MMCs, containing ferric iron and lipofuscin-ceroids, was detected in the juveniles sampled in the northern Adriatic Sea. These individuals showed also a strong anti-CYP1A immunopositivity in hepatocytes and in the epithelium of bile ducts. This study supports the utility of MMCs as biomarkers of fish 'health status' and gives concern for a potential contaminant accumulation in ABFT.

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Angiogenesis is an important pathway of neoplastic growth and progression. Our several published studies suggest that angiogenesis represents an interspecies pathway of tumour growth due to its roles in several spontaneous canine tumour model. Stromal microenvironment, macrophages and mast cells play a main roles in tumour angiogenesis. Furthermore several recent studies strongly suggest that mast cells density and mast cell activation are correlated with microvascular density. In this context mast cells may stimulates angiogenesis by secretion of their angiogenetic factors such as Vascular Endothelial Growth Factor (VEGF), Tryptase and Chynase. VEGF represents the classical pro-angiogenic factor while Tryptase and Chynase are proteases identified as new pro-angiogenic factor.. Interstingly canine mast cell tumour model (CMCTM) represents a unique model to study the role of mast cells in angiogenetic pathway. CMCTM are classified in well, intermediate and poorly differenziated tumours with and different angiogenetic activity and metastatic capacity. We aim to study by mean of immunohistochemistry and ELISA methods primary mast cell tumour and mast cell tumour lines for the expression of angiogenic factors and mast cell activation. In this context will be possible to evaluate the inhibition of pro-angiogenic factors and mast cell degranulation as a new anti-angiogentic and anti tumour therapy.

Angiogenesis is an important pathway of neoplastic growth and progression. Our several published studies suggest that angiogenesis represents an interspecies pathway of tumour growth due to its roles in several spontaneous canine tumour model. Stromal microenvironment, macrophages and mast cells play a main roles in tumour angiogenesis. Furthermore several recent studies strongly suggest that mast cells density and mast cell activation are correlated with microvascular density. In this context mast cells may stimulates angiogenesis by secretion of their angiogenetic factors such as Vascular Endothelial Growth Factor (VEGF), Tryptase and Chynase. VEGF represents the classical pro-angiogenic factor while Tryptase and Chynase are proteases that presents new pro-angiogenic factor. We aim to study the role of mast cells in neoplastic growth and progression in canine mast cell tumour model. Canine mast cell tumour model represents a unique model with well, intermediate and poorly differenziated tumour and differently angiogenetic activity. We aim to study with immunohistochemistry and ELISA methods primary mast cell tumour and mast cell tumour lines for the expression of angiogenic factors and mast cell activation. In this context will be possible to hypothesize the inhibition of pro-angiogenic factors and mast cell degranulation as a new anti-angiogentic and anti tumour therapy.

Fish liver is constituted of hepatocyte cords pervaded by a network of sinusoids. In fish liver, macrophages tend to give rise to melano-macrophage centres (MMCs). The aim of this study was to: a) characterize histochemically Atlantic bluefin tuna (Thynnus thynnus L.) (ABFT) MMCs; b) evaluate the use of MMCs as indicator of health status. Liver samples were taken from: a) wild adult males captured by traditional traps in Sardinia and Morocco; b) captive adult males experimentally reared in sea cages in Spain; c) captive juvenile males commercially reared in Croatia. The samples were fixed in 10% formalin, dehydrated in ethanol and embedded in paraffin wax. Sections were stained with: haematoxylin-eosin, α-naphtyl acetate esterase (ANAE) for macrophages; Mallory's method for lipofuscin/ ceroid, Perl’s stain for hemosiderin; antibodies against vitellogenin (VTG) and cytocrome 450P1A (CYP1A) mono-ossigenase. MMCs showed lysosomial activity and contained lipofuscins/ ceroids and hemosiderin. MMCs density was higher in the Croatian group in comparison to the other two fish groups. Moreover, individuals with hepatocytes immunopositive to VTG and CYP1A were found only in the Croatian group, thus indicating the exposure of these fish to environmental pollutants. This study indicates a role of MMCs as metabolic dumps and confirmed their utility as biomarker of fish health state

To evaluate the effect of decreasing dietary protein on growth performance, carcass traits, and intestinal mucosal morphometry, 180 female Hubbard strain broiler chickens were divided into 3 groups and fed 3 isoenergetic diets ad libitum from 14 d of age until slaughter age (49 d). The treatments varied according to 3 protein levels: high-protein diet (HiP, 22.5% CP, DM basis), medium-protein diet (MedP, 20.5% CP), and low-protein diet (LowP, 18.5%). Diets were obtained by replacing wheat middlings with soybean meal and were formulated to meet or exceed broiler amino acid requirements of the NRC. Morphometric indices of duodenum, jejunum, and ileum were measured at the end of the feeding period and included villus height, crypt depth, villus-to-crypt ratio, and apparent villus surface area. The dietary protein level had a significant effect on final BW of birds, whereas ADG, ADFI, and feed efficiency remained unaffected by dietary treatment. The muscle (breast and drumstick) yields were significantly higher in birds fed the HiP diet compared with those of the MedP and LowP diets. Meat quality traits were not affected by the protein level. The villus surface area of all intestinal segments did not change among groups. Instead, reducing the dietary protein level to 20.5% resulted in a higher villus height and villus height to crypt depth ratio in the duodenum and ileum. On the basis of our findings, even if the high-protein diet promoted meat yield, a medium-protein diet could positively support broiler growth performance, as confirmed by favorable morphometric features of the intestine.

Dog mast cell tumor (MCT) is common in dog. The etiology of canine MCTs is unknown, but it is probably multi-factorial. Its incidence is higher than it has found in human. There are demonstrated several common biological and clinical characteristics in both species. Cutaneous mast cells are located in the dermis and hypodermis. The objective of this study is to detect of MC on Toluidine Blue stained slides. There were examined 74 dogs of difference breeds and aged, from Tirana city. Six of them demonstrated the skin canine tumors. Skin samples were obtained from these animals. Macroscopic examination of the tumor revealed nodular ulcerated lesion with areas of necrosis and hemorrhage, accompanied with normal superjacent epidermis and annexes. Serial sections obtained from biopsy specimens were processed with toluidine blue staining pH 4.5, specific for MC identification. This study suggests that Toluidine blue, pH 4.5 stain may give a good information about skin tumors in dog, histologically with benign behavior.