Microbiological quality of marketed fresh and frozen seafood caught off the Adriatic coast of Croatia

Fresh and frozen seafood products (fish, shellfish, crustaceans, molluscs) in wide use in Croatia and typical for the Mediterranean diet, were examined for presence of microbiological contamination through winter and summer seasons. Total bacterial counts of aerobic mesophilic bacteria (AB), aerobic psychrophilic bacteria (AP), Salmonella spp., Enterobacteriaceae, Escherichia coli, Staphylococcus aureus, sulphite-reducing clostridia (SRC), Listeria monocytogenes, Vibrio cholerae and V. parahaemolyticus tests were performed. The microbiological quality of individual samples varied widely between the animal species and within winter/summer seasons regarding total counts of aerobic mesophilic and psychrophilic bacteria. The poorest quality was for (both summer and winter) fish samples, where 66.6 % of fresh and frozen fish were found unacceptable by Croatian standards. The overall prevalence of V. parahaemolyticus was 5 %. Its recovery rate was higher in fresh/frozen shellfish in both seasons than in other specimens or other storage/season conditions. Fresh crustaceans sampled in winter demonstrated significantly higher aerobic mesophilic counts than frozen ones. Unacceptable Enterobacteriaceae levels were obtained in 40 % of the summer fresh fish samples. The results of this survey constitute an indicator of bacteriological contamination of a variety of seafood. The findings could serve as a basis for future testing of seafood, and possibly as a template for developing a regional/Mediterranean testing scheme on the microbial contamination of seafood in order to establish data with comparative epidemiological and statistical values

Identification and functionality of bacterial strains in lyophilized probiotic preparations on the Croatian market

Istraživački cilj je uspostaviti pouzdan metodološki pristup za robusnu analizu probiotičkih pripravaka. Prioritetno je definirana ispravnost taksonomske nomenklature i usklađenost koncentracije probiotičkih stanica s preporučenom dnevnom dozom. Mikrobiološka analiza i združena primjena fenotipskih i genotipskih metoda isključila je prisutnost neprobiotičkih bakterija. Probiotički pripravci su sigurni za ljudsku primjenu jer su prema metaboličkim, fiziološkim i fermentacijskim profilima potvrđeni kao vrste iz rodova Lactobacillus ili Bifidobacterium te je ustanovljena niska pojavnost fenotipske rezistencije na antibiotike. Analizom SDS-PAGE proteinskih profila temeljem dendrograma generiranih pomoću Gel Compare II, sojevi su klasificirani u Lactobacillus ili Bifidobacterium rod. Razlikovanje sojeva utemeljeno je na analizi polimorfizma slučajno umnoženih fragmenata DNA, čija je biotipizacija provedena umnažanjem V9 regije 16S rRNA Bifidobacterium ili 16S/23S intergenske rRNA „spacer“ regije Lactobacillus vrste. Sojevi su identificirani sekvencioniranjem 16S rRNA gena. Potencijal kolonizacije i jačanja funkcije intestinalne barijere određeni su karakterizacijom adaptacije na ekstremne uvjete mikrookoliša i gastrointestinalnog trakta, adhezije na mucin i proteinske ligande intestinalnih niša. Pojedini sojevi iskazuju aktivnost hidrolaze žučnih soli, asimilaciju kolesterola ili prebiotičkih supstrata i kompetitivne ekskluzije enteropatogena. Razvijeni integrirani pristup obuhvaća primjenu molekularne metodologije i in vitro eksperimentalnih modela za validaciju probiotičkih bakterija.The research goal was to define a reliable methodological approach for robust analysis of probiotic preparations. The correctness of the taxonomic nomenclature and the compliance of the concentration of probiotic cells with the recommended daily dose was defined as a priority. Microbiological analysis of probiotic products and the combined application of phenotypic and genotypic methods ruled out the presence of contaminating non-probiotic bacteria. The products are safe for use due to the low incidence of phenotypic resistance to antibiotics of strains which are according to metabolic, physiological characteristics and carbohydrate fermentation profiles grouped to Lactobacillus and Bifidobacterium species. SDS-PAGE of soluble cell or surface proteins and computer-aided comparison of the electrophoretic protein patterns generated by Gel Compare II classified bacterial strains into Lactobacillus spp. or Bifidobacterium spp. RAPD-PCR was used to generate unique and identifying DNA profiles of Lactobacillus and Bifidobacterium strains, whose biotyping was performed by amplification of V9 region of 16S rRNA Bifidobacterium or 16S/23S intergenic rRNA "spacer" region of Lactobacillus species. Probiotic strains were identified by 16S rRNA sequencing. Colonization potential and enhancement of intestinal barrier function were determined based on the adaptation to the extreme microenvironment conditions and gastrointestinal tract and adhesion to the mucin and protein ligands of intestinal niches. Strains show bile salt hydrolase activity and cholesterol or prebiotics assimilation and competitive exclusion of enteropathogens. The developed integrated approach includes the application of molecular methodology and in vitro experimental models for the validation of probiotic bacteria

Funkcni usporadani nervovych spojeni zprostredkujicich vyvolane odmitani potravy u krys

A major goal of present neuroscience is to understand the principles of learning and memory. Rapid development of methodological tools of molecular and cellular biology has considerably advanced our ability to study the synaptic and subsynaptic events underlying neural plasticity. The need to find an instance of behavioral learning which can proceed under conditions comparable with those prevailing in isolated tissues called attention of neurobiologists to the CTA paradigm, the critical associative phase of which is not prevented by anaesthesia, hypothermia or functional decortication. During CTA acquisition, gustatory information about novel food (conditioned stimulus - CS) is entered into a temporary file storing gustatory short term memory (GSTM) traces where it waits several hours for the possible onset of visceral symptoms of intoxication (unconditioned stimulus - US). The result of the GSTM-US association is formation of the gustatory long term memory trace (GLTM), corresponding to the CTA engram. Acquisition and retrieval of CTA is implemented by the interaction of several brain structures. The role of three important gustatory and visceral relays in CTA acquisition and retrieval- gustatory neocortex (GC), amygdala (AM) and PBN was studied in lateralization experiments. Reversible unilateral lesions of PBN and AM or of AM and GC were induced by injection of TTX either in the same hemisphere or in the opposite hemispheres during CTA acquisition. It was found that contralateral TTX injections into the above structures caused a complete blockade of CTA acquisition which was not affected by ipsilateral TTX administration. Retrieval of CTA acquired during unilateral blockade of PBN or AM in one hemisphere has been examinated during unilateral TTX blockade of GC alone, AM alone or during combined GC and AM inactivation either in the same or in contralateral hemispheres. A clear CTA retrieval impairment was found only when structures in the opposite hemispheres were blocked. These results indicate a complete lateralization of CTA engram can be formed in each hemisphere separately without participation of the contralateral hemisphere. The contribution of protein kinase C (PKC) to the acquisition of conditioned taste aversion (CTA) was tested by andinjection of three PKC inhibitors - polymyxin B, H7 and staurosporine into the PBN. From the tested drugs only polymyxin B prevented CTA acquisition. Application of H7 and staurosporine into the PBN did not impair CTA learning. The blocking effect of polymyxin B is dose dependent (5 and 10 mM concentration did not disrupt CTA formation) and site specific (application of polymyxin B into the visual cortex did not elicit CTA blockade). The ability of polymyxin B to discrupt CTA learning is not due to irreversible damage of PBN. These results suggest that polymyxin B blocks acquisition of CTA in some nonspecific way not necessarily involving inhibition of PKC. This conclusion is supported by failure of two other more specific PKC inhibitors to affect CTA learning. These results are in agreement with previous findings that only relatively nonspecific inhibitors like 2- deoxy-galactose or colchicine (Bures 1991, Bielavska and Bures 1993) can mimick the disruptive effect of TTX on CTA acquisition when injected into the PBN. The exact role of PKC in CTA acquisition still remains openAvailable from STL Prague, CZ / NTK - National Technical LibrarySIGLECZCzech Republi