A modified culture medium for improved isolation of marine vibrios

Marine Vibrio members are of great interest for both ecological and biotechnological research, which often relies on their isolation. Whereas many efforts have been made for the detection of food-borne pathogenic species, much less is known about the performances of standard culture media toward environmental vibrios. We show that the isolation/enumeration of marine vibrios using thiosulfate-citrate-bile salts-sucrose agar (TCBS) as selective medium may be hampered by the variable adaptability of different taxa to the medium, which may result even in isolation failure and/or in substantial total count underestimation. We propose a modified TCBS as isolation medium, adjusted for marine vibrios requirements, which greatly improved their recovery in dilution plate counts, compared with the standard medium. The modified medium offers substantial advantages over TCBS, providing more accurate and likely estimations of the actual presence of vibrios. Modified TCBS allowed the recovery of otherwise undetected vibrios, some of which producing biotechnologically valuable enzymes, thus expanding the isolation power toward potentially new enzyme-producers Vibrio taxa. Moreover, we report a newly designed Vibrio-specific PCR primers pair, targeting a unique rpoD sequence, useful for rapid confirmation of isolates as Vibrio members and subsequent genetic analyses

Effect of chronic heat stress on gastrointestinal histology and expression of feed intake-regulatory hormones in broiler chickens

Heat stress (HS) dramatically impairs the growth performance of broiler chickens, mainly as a consequence of reduced feed intake due to the loss of appetite. This study was aimed at evaluating the alterations induced by chronic HS conditions on the morphological and morphometric features of the gastrointestinal (GI) tract and on the expression of some enteroendocrine cells (EECs) involved in the regulation of feed intake in chickens. Three hundred male chickens (Ross 308) were divided into two experimental groups and raised either in thermoneutral environment for the whole fattening period (0–41 days) (TNT group) or subjected to chronic HS conditions (30 °C for 24 h/day) from 35 to 41 days (HS group). Samples of proventriculus, duodenum, jejunum and cecum were collected from 24 broilers (12/group). Haematoxylin-eosin was used for the morphometric evaluations, while immunohistochemistry was applied for the evaluation of EECs expressing ghrelin (GHR), cholecystokinin (CCK), neuropeptide Y (NPY), glucagon-like peptide-1 (GLP-1), and serotonin (5-HT). In the proventriculus, HS reduced total wall thickness and mucous layer height (P ≤ 0.01) as well as mean diameter, circumference, and area of the compound tubular glands (P ≤ 0.001) with respect to TNT. The small intestine of HS birds was characterised by decreased villous height and total thickness (duodenum, P ≤ 0.01; jejunum, P ≤ 0.001), whereas crypt depth and width were reduced only in the jejunum (P ≤ 0.01). HS had negligible effects on the morphological aspects of the cecum. In the proventriculus, an increase in GHR and NPY EECs was observed in response to HS (P ≤ 0.001). Similarly, the small intestine villi of the HS group showed greater GLP-1 (P ≤ 0.05), 5-HT (P ≤ 0.001) and CCK (P ≤ 0.01) EECs. Moreover, the expression of 5-HT EECs was higher in the duodenal (P ≤ 0.01) and jejunal (P ≤ 0.01) crypts of HS birds, whereas GLP-1 and CCK EECs increased only in jejunal crypts (P ≤ 0.05). Finally, 5-HT EEC expression was increased in the cecum of HS group (P ≤ 0.01). In conclusion, these outcomes demonstrate that chronic HS induces morphometric alterations not only in the small intestine but also in a key organ such as the proventriculus. Furthermore, HS conditions affect the presence and distribution of EECs, suggesting that some GI peptides and biogenic amine may be implicated in the regulation of appetite and voluntary feed intake in heat-stressed broiler chickens

Environment and behavior: Neurochemical effects of different diets in the calf brain

open7noCalves reared for the production of white veal are subjected to stressful events due to the type of liquid diet they receive. Stress responses are mediated by three main stress-responsive cerebral regions: the prefrontal cortex, the paraventricular nucleus of the hypothalamus, and the nucleus of the solitary tract of the brainstem. In the present study, we have investigated the effects of different diets on these brain regions of ruminants using immunohistochemical methods. In this study, 15 calves were used and kept in group housing systems of five calves each. They were fed with three different diets: a control diet, a milk diet, and a weaned diet. Brain sections were immunostained to evaluate the distribution of neuronal nitric oxide synthase and myelin oligodendrocyte glycoprotein immunoreactivity in the prefrontal cortex; the expression of oxytocin in the paraventricular nucleus; and the presence of c-Fos in the A2 group of the nucleus of the solitary tract. The main results obtained indicate that in weaned diet group the oxytocin activity is lower than in control diet and milk diet groups. In addition, weaning appears to stimulate myelination in the prefrontal cortex. In summary, this study supports the importance of maintaining a nutritional lifestyle similar to that occurring in natural conditions.openPeli A.; Grandis A.; Tassinari M.; Bergamini P.F.; Tagliavia C.; Roccaro M.; Bombardi C.Peli A.; Grandis A.; Tassinari M.; Bergamini P.F.; Tagliavia C.; Roccaro M.; Bombardi C

Immunohistochemical Distribution of Serotonin Transporter (SERT) in the Optic Lobe of the Honeybee, Apis mellifera

Visual information is processed in the optic lobes, which consist of three retinotopic neuropils. These are the lamina, the medulla and the lobula. Biogenic amines play a crucial role in the control of insect responsiveness, and serotonin is clearly related to aggressiveness in invertebrates. Previous studies suggest that serotonin modulates aggression-related behaviours, possibly via alterations in optic lobe activity. The aim of this investigation was to immunohistochemically localize the distribution of serotonin transporter (SERT) in the optic lobe of moderate, docile and aggressive worker honeybees. SERT-immunoreactive fibres showed a wide distribution in the lamina, medulla and lobula; interestingly, the highest percentage of SERT immunoreactivity was observed across all the visual neuropils of the docile group. Although future research is needed to determine the relationship between the distribution of serotonin fibres in the honeybee brain and aggressive behaviours, our immunohistochemical study provides an anatomical basis supporting the role of serotonin in aggressive behaviour in the honeybee

The comparison of latero-medial versus dorso-palmar/plantar drilling for cartilage removal in the proximal interphalangeal joint

The aims of the present study were to compare the percentages of articular cartilage removed using a lateral drilling approach of the proximal interphalangeal joint (PIPJ) and a dorsal drilling approach, and to assess the usefulness of digital fluoroscopy when performing a lateral drilling approach. Sixty cadaveric PIPJs were drilled using a surgical drill bit to remove the articular cartilage. The limbs were divided into three groups containing 10 forelimbs and 10 hindlimbs each. One group received the dorsal drilling approach, the second one received the lateral drilling approach and the last one received the lateral drilling approach under digital fluoroscopy guidance. The percentage of articular cartilage removed from each articular surface was assessed using Adobe Photoshop® software. The percentages of removed cartilage turned out to be significantly higher with lateral approach, especially under fluoroscopic guidance, both in the forelimbs (p = 0.00712) and hindlimbs (p = 0.00962). In conclusion, the lateral drilling approach seems to be a minimally invasive technique with which to perform PIPJ arthrodesis, even more efficient than the previously reported dorsal approach

Neural Crest-Derived Chondrocytes Isolation for Tissue Engineering in Regenerative Medicine

Chondrocyte transplantation has been successfully tested and proposed as a clinical procedure aiming to repair articular cartilage defects. However, the isolation of chondrocytes and the optimization of the enzymatic digestion process, as well as their successful in vitro expansion, remain the main challenges in cartilage tissue engineering. In order to address these issues, we investigated the performance of recombinant collagenases in tissue dissociation assays with the aim of isolating chondrocytes from bovine nasal cartilage in order to establish the optimal enzyme blend to ensure the best outcomes of the overall procedure. We show, for the first time, that collagenase H activity alone is required for effective cartilage digestion, resulting in an improvement in the yield of viable cells. The extracted chondrocytes proved able to grow and activate differentiation/dedifferentiation programs, as assessed by morphological and gene expression analyses

Cellular Distribution of Canonical and Putative Cannabinoid Receptors in Canine Cervical Dorsal Root Ganglia

open10noGrowing evidence indicates cannabinoid receptors as potential therapeutic targets for chronic pain. Consequently, there is an increasing interest in developing cannabinoid receptor agonists for treating human and veterinary pain. To better understand the actions of a drug, it is of paramount importance to know the cellular distribution of its specific receptor(s). The distribution of canonical and putative cannabinoid receptors in the peripheral and central nervous system of dogs is still in its infancy. In order to help fill this anatomical gap, the present ex vivo study has been designed to identify the cellular sites of cannabinoid and cannabinoid-related receptors in canine spinal ganglia. In particular, the cellular distribution of the cannabinoid receptors type 1 and 2 (CB1 and CB2) and putative cannabinoid receptors G protein-coupled receptor 55 (GPR55), nuclear peroxisome proliferator-activated receptor alpha (PPARα), and transient receptor potential vanilloid type 1 (TRPV1) have been immunohistochemically investigated in the C6-C8 cervical ganglia of dogs. About 50% of the neuronal population displayed weak to moderate CB1 receptor and TRPV1 immunoreactivity, while all of them were CB2-positive and nearly 40% also expressed GPR55 immunolabeling. Schwann cells, blood vessel smooth muscle cells, and pericyte-like cells all expressed CB2 receptor immunoreactivity, endothelial cell being also PPARα-positive. All the satellite glial cells (SGCs) displayed bright GPR55 receptor immunoreactivity. In half of the study dogs, SGCs were also PPARα-positive, and limited to older dogs displayed TRPV1 immunoreactivity. The present study may represent a morphological substrate to consider in order to develop therapeutic strategies against chronic pain.openChiocchetti R, Galiazzo G, Tagliavia C, Stanzani A, Giancola F, Menchetti M, Militerno G, Bernardini C, Forni M, Mandrioli LChiocchetti R, Galiazzo G, Tagliavia C, Stanzani A, Giancola F, Menchetti M, Militerno G, Bernardini C, Forni M, Mandrioli

Distribution of Cannabinoid Receptors in Keratinocytes of Healthy Dogs and Dogs With Atopic Dermatitis

It is commonly accepted that some form of skin barrier dysfunction is present in canine atopic dermatitis (AD), one of the most common cutaneous pruritic inflammatory diseases of dogs. The impaired skin barrier function facilitates the penetration of allergens and subsequently stronger sensitization responses. The role of the endocannabinoid system (ECS) in the physiology and pathology of the skin is becoming increasingly established. It has been demonstrated that cannabinoid receptors are expressed in healthy and diseased skin and, based on current knowledge, it could be stated that cannabinoids are important mediators in the skin. The present study has been designed to immunohistochemically investigate the expression of the cannabinoid receptors type 1 (CB1R) and 2 (CB2R) and the cannabinoid-related receptors G protein-coupled receptor 55 (GPR55), transient receptor potential vanilloid 1 (TRPV1) and ankyrin 1 (TRPA1), peroxisome proliferator-activated receptors alpha (PPARα), and serotoninergic receptor 1a (5-HT1aR) in keratinocytes of healthy dogs and of dogs with AD. Samples of skin tissues were collected from 7 healthy controls (CTRL-dogs) and from 8 dogs with AD (AD-dogs). The tissue samples were processed using an immunofluorescence assay with commercially available antibodies, and the immunolabelling of the receptors studied was quantitatively evaluated. The keratinocytes of the CTRL- and the AD-dogs showed immunoreactivity for all the receptors investigated with a significant upregulation of CB2R, TRPA1, and 5-HT1aR in the epidermis of the AD-dogs. The presence of cannabinoid and cannabinoid-related receptors in healthy keratinocytes suggested the possible role of the ECS in canine epidermal homeostasis while their overexpression in the inflamed tissues of the AD-dogs suggested the involvement of the ECS in the pathogenesis of this disease, having a possible role in the related skin inflammation and itching. Based on the present findings, the ECS could be considered a potential therapeutic target for dogs with AD

Characterization of translationally controlled tumour protein from the sea anemone Anemonia viridis and transcriptome wide identification of cnidarian homologues

Gene family encoding translationally controlled tumour protein (TCTP) is defined as highly conserved among organisms; however, there is limited knowledge of non-bilateria. In this study, the first TCTP homologue from anthozoan was characterised in the Mediterranean Sea anemone, Anemonia viridis. The release of the genome sequence of Acropora digitifera, Exaiptasia pallida, Nematostella vectensis and Hydra vulgaris enabled a comprehensive study of the molecular evolution of TCTP family among cnidarians. A comparison among TCTP members from Cnidaria and Bilateria showed conserved intron exon organization, evolutionary conserved TCTP signatures and 3D protein structure. The pattern of mRNA expression profile was also defined in A. viridis. These analyses revealed a constitutive mRNA expression especially in tissues with active proliferation. Additionally, the transcriptional profile of A. viridis TCTP (AvTCTP) after challenges with different abiotic/biotic stresses showed induction by extreme temperatures, heavy metals exposure and immune stimulation. These results suggest the involvement of AvTCTP in the sea anemone defensome taking part in environmental stress and immune responses

Effect of essential oils on the oxyntopeptic cells and somatostatin and ghrelin immunoreactive cells in the european sea bass (Dicentrarchus labrax) gastric mucosa

open9noThis research was founded by “Fondazione CARISBO” project 2017/0312, Bologna, ItalyThe current work was designed to assess the effect of feed supplemented with essential oils (EOs) on the histological features in sea bass’s gastric mucosa. Fish were fed three diets: control diet (CTR), HERBAL MIX® made with natural EOs (N-EOs), or HERBAL MIX® made with artificial EOs obtained by synthesis (S-EOs) during a 117-day feeding trial. Thereafter, the oxyntopeptic cells (OPs) and the ghrelin (GHR) and somatostatin (SOM) enteroendocrine cells (EECs) in the gastric mucosa were evaluated. The Na+K+-ATPase antibody was used to label OPs, while, for the EECs, anti-SOM and anti-GHR antibody were used. The highest density of OP immunoreactive (IR) area was in the CTR group (0.66 mm2 ± 0.1). The OP-IR area was reduced in the N-EO diet group (0.22 mm2 ± 1; CTR vs. N-EOs, p < 0.005), while in the S-EO diet group (0.39 mm2 ± 1) a trend was observed. We observed an increase of the number of SOM-IR cells in the N-EO diet (15.6 ± 4.2) compared to that in the CTR (11.8 ± 3.7) (N-EOs vs. CTR; p < 0.05), but not in the S-EOs diet. These observations will provide a basis to advance current knowledge on the anatomy and digestive physiology of this species in relation to pro-heath feeds.openMazzoni M.; Lattanzio G.; Bonaldo A.; Tagliavia C.; Parma L.; Busti S.; Gatta P.P.; Bernardi N.; Clavenzani P.Mazzoni M.; Lattanzio G.; Bonaldo A.; Tagliavia C.; Parma L.; Busti S.; Gatta P.P.; Bernardi N.; Clavenzani P