Evaluation of the Possible Transmission of BSE and Scrapie to Gilthead Sea Bream (Sparus aurata)

In transmissible spongiform encephalopathies (TSEs), a group of fatal neurodegenerative disorders affecting many species, the key event in disease pathogenesis is the accumulation of an abnormal conformational isoform (PrPSc) of the host-encoded cellular prion protein (PrPC). While the precise mechanism of the PrPC to PrPSc conversion is not understood, it is clear that host PrPC expression is a prerequisite for effective infectious prion propagation. Although there have been many studies on TSEs in mammalian species, little is known about TSE pathogenesis in fish. Here we show that while gilthead sea bream (Sparus aurata) orally challenged with brain homogenates prepared either from a BSE infected cow or from scrapie infected sheep developed no clinical prion disease, the brains of TSE-fed fish sampled two years after challenge did show signs of neurodegeneration and accumulation of deposits that reacted positively with antibodies raised against sea bream PrP. The control groups, fed with brains from uninfected animals, showed no such signs. Remarkably, the deposits developed much more rapidly and extensively in fish inoculated with BSE-infected material than in the ones challenged with the scrapie-infected brain homogenate, with numerous deposits being proteinase K-resistant. These plaque-like aggregates exhibited congophilia and birefringence in polarized light, consistent with an amyloid-like component. The neurodegeneration and abnormal deposition in the brains of fish challenged with prion, especially BSE, raises concerns about the potential risk to public health. As fish aquaculture is an economically important industry providing high protein nutrition for humans and other mammalian species, the prospect of farmed fish being contaminated with infectious mammalian PrPSc, or of a prion disease developing in farmed fish is alarming and requires further evaluation

Pathological study of seabass (Dicentrarchus labrax) vibriosis

In this study, juvenile seabass fish were experimentally infected by immersion with two bacterial strains of V. anguillarum, (Van1 and Van731) in three different experiments. Van1, a naturally occurring strain, harbours a 67kb pJM1-like plasmid, which we named pKTP. pKTP encodes the iron-sequestering system of this strain. Using the heat shock method we have also created the plasmidless-derivative strain of Van1 designated as Van731. Both strains identified by PCR detection of the amiB gene. The successful curing of the plasmid was confirmed with angR and angH gene-specific PCR. The ability of the strain to produce histamine was tested by ΗPLC. The first experimental infection aimed to study the Van1 kinetics. The presence of the bacteria in fish tissues was confirmed with 16rRNA FISH using the Vav3 and the EUB 338 probes. Histological and FISH analyses showed that skin was the route of V.anguillarum invasion. The bacteria were found in small aggregates in dermis, as early as 20 hours post infection. At the same time point small numbers of bacteria were seen in the spleen and peritoneum. 72 hours post infection the bacteria were found in all the fish organs examined. The second experimental infection was carried out in order to assess the mortality of fish infected with the strains Van1 and Van 731. The mortality was significantly lower in fish infected with the Van 731 strain indicating that the presence of thepKTP plasmid is particularly important for Van1 pathogenicity. The third experimental infection aimed to investigate the effect of the iron-binding bacterial system on the V.anguillarum ability to induce histopathological lesions. The skin, the spleen and the heart were the organ mostly affected. Less severe lesions were seen in the liver, the head and body kidney, the gills, the posterior intestine and the swim bladder, whereas the lesions were scarce and mild in the remaining organs. The lesions that were consistently found in the organs of fish infected with both strains included oedema, vessel dilatation, congestion, haemorrhage, necrosis and mild inflammatory cell infiltration. The qualitative and quantitative analysis of V.anguillarum-induced lesions showed that the genetically modified strain Van731 caused significantly less pathology in fish. Taken together the results of this study suggest that iron binding system is important but not essential for the pathogenesis of vibriosis of the seabass. Other pathogenetic factors, although less important than the iron-binding system, contribute in the pathogenesis of vibriosis.Πραγματοποιήθηκαν τρεις πειραματικές μολύνσεις ιχθυδίων λαβρακιού, με εμβάπτιση, και χρησιμοποιήθηκαν δύο βακτηριδιακά στελέχη V. anguillarum, το Van1 και το Van731. Το Van1, φυσικό στέλεχος διαπιστώσαμε ότι διέθετε πλασμίδιο μεγέθους 67Kb, που ονομάσαμε pΚΤΡ. Το pΚΤΡ είναι πλασμίδιο όμοιο με το pJM1 και υπεύθυνο για την παραγωγή σιδηροφόρου μορίου από το στέλεχος. To Van731 είναι γενετικά τροποποιημένο στέλεχος που δημιουργήσαμε από το Van1 με θερμική καταπόνηση. To Van731 δεν διαθέτει το pΚΤΡ πλασμίδιο. Τα στελέχη ταυτοποιήθηκαν με PCR, με τη χρήση του γονιδίου amiB. Η επιτυχής αφαίρεση του πλασμιδίου επιβεβαιώθηκε με PCR με τη χρήση των γονιδίων angR και angH. Η παραγωγή ισταμίνης από τα στελέχη επιβεβαιώθηκε με HPLC. Η πρώτη πειραματική μόλυνση πραγματοποιήθηκε για τη μελέτη της κινητικής του Van1. Ο έλεγχος της παρουσίας του βακτηριδίου στους ιστούς επιβεβαιώθηκε με FISH. Για τον υβριδισμό χρησιμοποιήθηκαν οι ιχνηθέτες Vav3 και Eub338. Από την πρώτη πειραματική μόλυνση προκύπτει, ότι πύλη εισόδου του βακτηριδίου στον οργανισμό του ψαριού αποτελεί το δέρμα. Το βακτηρίδιο εντοπίστηκε σε μικρά αθροίσματα στο χόριο του δέρματος στις 20 ώρες μετά τη μόλυνση και σχεδόν ταυτόχρονα στο σπλήνα και στο περιτόναιο. Στις 72 ώρες μετά τη μόλυνση το βακτηρίδιο διαπιστώθηκε στο σύνολο των οργάνων που εξετάστηκαν. Η δεύτερη πειραματική μόλυνση πραγματοποιήθηκε για να ελεγχθεί η προκαλούμενη από τα στελέχη Van1 και Van731 θνησιμότητα. Η μέση θνησιμότητα υπολογίστηκε στο 87,5% και 2,5% αντίστοιχα, αποδεικνύοντας ότι οφείλεται σε σημαντικό βαθμό στο πλασμίδιο που το βακτηρίδιο διαθέτει. Η τρίτη πειραματική μόλυνση πραγματοποιήθηκε για να περιγραφούν οι ιστοπαθολογικές αλλοιώσεις της Δονακίωσης από V. anguillarum στο λαβράκι και να διερευνηθεί σε επίπεδο αλλοιώσεων ο ρόλος του συστήματος δέσμευσης σιδήρου. Σημαντικότερες αλλοιώσεις από το Van1 προκλήθηκαν στο δέρμα, το σπλήνα και την καρδιά, ηπιότερες στο ήπαρ, τον κεφαλικό και σωματικό νεφρό, τα βράγχια, το οπίσθιο έντερο και τη νηκτική κύστη, ενώ ελάχιστες ήταν οι αλλοιώσεις στα υπόλοιπα όργανα. Κυκλοφορικές διαταραχές όπως οιδήματα, διάταση και συμφόρηση αγγείων, και αιμορραγίες, νεκρώσεις και ήπια φλεγμονώδης αντίδραση χαρακτήρισαν την ιστοπαθολογική εικόνα της νόσου στα διάφορα όργανα. Οι αλλοιώσεις ήταν σημαντικά ηπιότερες στα ιχθύδια που μολύνθηκαν με το Van731. Το σύστημα δέσμευσης σιδήρου αποδεικνύεται σημαντικό για την εκδήλωση της παθογένειας της νόσου. Εντούτοις στην παθογένεια εμπλέκονται και άλλοι παθογενετικοί μηχανισμοί του βακτηριδίου

Aquaculture bacterial bank in Greece - methodology and first results

A collection a pathogenic bacterial strains isolated from natural disease outbreaks in Greek fish farms was founded in Greece by the cooperation of three laboratories: a) Lab. of Ichthyology, School of Veterinary Medicine, Aristotle University of Thessaloniki, b) Lab. of Ichthyopathology Faculty of Veterinary Medicine, University of Thessaly and c) Institute of Aquaculture, Hellenic Center for Marine Research Athens. This project is funded by the Hellenic Ministry of Rural Development and Food - Fisheries Department. After isolation the bacteria are characterized biochemically and molecularly (PCR). Their pathogenicity is tested using experimental challenges. Theisolates are then lyophilizised and stocked at -80 oC. The results so far indicate that Listonella anguillarum, Vibrio alginolyticus and Photobacterium damsela subsp piscicida are the main pathogens that occur in Greek mariculture

Beneficial bacteria inhibit cachexia

Muscle wasting, known as cachexia, is a debilitating condition associated with chronic inflammation such as during cancer. Beneficial microbes have been shown to optimize systemic inflammatory tone during good health; however, interactions between microbes and host immunity in the context of cachexia are incompletely understood. Here we use mouse models to test roles for bacteria in muscle wasting syndromes. We find that feeding of a human commensal microbe, Lactobacillus reuteri, to mice is sufficient to lower systemic indices of inflammation and inhibit cachexia. Further, the microbial muscle-building phenomenon extends to normal aging as wild type animals exhibited increased growth hormone levels and up-regulation of transcription factor Forkhead Box N1 [FoxN1] associated with thymus gland retention and longevity. Interestingly, mice with a defective FoxN1 gene (athymic nude) fail to inhibit sarcopenia after L. reuteri therapy, indicating a FoxN1-mediated mechanism. In conclusion, symbiotic bacteria may serve to stimulate FoxN1 and thymic functions that regulate inflammation, offering possible alternatives for cachexia prevention and novel insights into roles for microbiota in mammalian ontogeny and phylogeny.National Institutes of Health (U.S.) (Grant R01CA108854)National Institutes of Health (U.S.) (Grant U01 CA164337