Highly Transferable pAQU-Related Plasmids Encoding Multidrug Resistance Are Widespread in the Human and Fish Pathogen Photobacterium damselae subsp. damselae in Aquaculture Areas in the Black Sea.

The marine bacterium Photobacterium damselae subsp. damselae is a pathogen that causes disease in diverse marine animals, and is also a serious opportunistic human pathogen that can cause fatal infections. Strains of this pathogen isolated from diseased European sea bass in aquaculture facilities in the Turkish coast of the Black Sea were found to exhibit reduced sensitivity to multiple antimicrobials. Selected representative strains were subjected to complete genome sequencing and plasmid characterization. It was found that multidrug resistant (MDR) isolates harboured large conjugative plasmids sharing part of their sequence backbone with pAQU-group plasmids, hitherto reported exclusively in China and Japan. Four new pAQU-group versions of plasmids were identified in the present study, containing distinct combinations of the resistance determinants tetB, floR, sul2, qnrVC, dfrA and strAB. Conjugative transfer of pPHDD2-OG2, a representative plasmid of 170,998 bp, occurred at high frequencies (2.2 × 10-2 transconjugants per donor cell), to E. coli and to pathogenic P. damselae subsp. damselae and subsp. piscicida strains. Upon transfer, pPHDD2-OG2 conferred reduced susceptibility to a number of antimicrobials to the recipient strains. Comparative genomics analysis of host strains suggested that these MDR plasmids of the pAQU-group were acquired by different genetic lineages of Pdd. This study provides evidence that P. damselae subsp. damselae isolated from diseased fish constitute a reservoir for conjugative MDR pAQU-group plasmids in the Mediterranean basin, and have the potential to spread to diverse bacterial species.

Natural calcium phosphates from fish bones and their potential biomedical applications.

The treatment and recovery of bio-wastes have raised considerable attention both from the environmental and economic point of view. Every year, a remarkable amount of fish processing by-products are generated and dumped as waste from all over the world. Fish bones can serve as a raw material for the production of high value-added compounds that can be used in various sectors including agrochemical, biomedical, food and pharmaceutical industries. The calcination of fish bones results in a single phase (hydroxyapatite) or bi-phasic (hydroxyapatite-tricalcium phosphate) bioceramics depending on the processing conditions as well as the content of the fish bones. This review summarizes the literature on the production of hydroxyapatite from fish bones and discusses their potential applications in biomedical field. The effect of processing conditions on the properties of final products including Ca/P ratio, crystal structure, particle shape, particle size and biological properties are presented in the light of X-ray diffraction, scanning electron microscopy, transmission electron microscopy, thermogravimetric-differential thermal analysis, bioactivity and biocompatibility investigations.

Photobacterium damselae subsp. damselae, an Emerging Fish Pathogen in the Black Sea: Evidence of a Multiclonal Origin.

UNLABELLED: Photobacterium damselae subsp. damselae is considered to be an emerging pathogen of marine fish of importance in aquaculture, with a notable increase in its geographical distribution during the last several years. In this study, we carried out for the first time to our knowledge a genetic and pathobiological characterization of 14 strains isolated from sea bass (Dicentrarchus labrax) reared in the Southeastern Black Sea, where high mortalities were observed at two aquaculture farms during the summer and autumn of 2011. Heterogeneity was evidenced among strains in phenotypical traits, such as sucrose fermentation, motility, and hemolysis. Although 11 of 14 isolates were hemolytic, we found that all of the isolates lacked the pPHDD1 virulence plasmid that encodes the phospholipase-D damselysin (Dly) and the pore-forming toxin PhlyP, two hemolysins previously reported to constitute major virulence factors for turbot. Subsequent PCR and sequencing analyses demonstrated that the 11 hemolytic isolates harbored a complete hlyAch gene, a chromosome I-borne gene that encodes HlyAch hemolysin, whereas the three nonhemolytic isolates contained hlyAch pseudogenes caused by insertion sequence elements. Virulence challenges with two representative strains revealed that, albeit less virulent than the pPHDD1-harboring strain RM-71, the plasmidless hlyAch-positive and hlyAch-negative Black Sea isolates were pathogenic for sea bass. A phylogenetic analysis based on the toxR gene sequence uncovered a greater diversity in the isolates, indicating that the presence of this pathogen in the Black Sea was not caused by the introduction and spread of a single virulent clone but by the proliferation of different clones. IMPORTANCE: The geographical distribution of marine bacterial pathogens is undergoing a worldwide increase. In particular, bacteria of the group vibrios are increasingly being isolated as the causative agents of disease in novel species of cultivated fish in areas where they had not been previously reported. Here we characterize for the first time to our knowledge a collection of isolates of the fish and human pathogen Photobacterium damselae subsp. damselae from diseased sea bass reared in the Black Sea. We uncovered great genetic diversity in the Black Sea isolates of this pathogen, suggesting a multiclonal origin. We also demonstrate for the first time that these isolates bear pathogenic potential for sea bass cultures by virulence challenges.

Seasonal dynamics of Trichodina spp. on whiting (Merlangius merlangus)in relation to organic pollution on the eastern Black Sea coast of Turkey.

It was determined that there is a relationship between prevalence and mean densities of Trichodina spp. on Merlangius merlangus and organic pollution, measured as levels of nitrite, nitrate and phosphate in the surrounding environment. Monthly, two left anterior gill arcs of 60 M. merlangus (unless otherwise stated) captured with hook and line were fixed in 10% formalin. The number of Trichodina spp. was determined by counting all of the cells with a grid slide. The two right anterior gill arcs were used to prepare dry smears to picture the morphology of the trichodinids and to determine species composition. High levels of prevalence and densities of the protozoan were observed during the late fall, winter and early spring months. Then, prevalence decreased to lower levels but never below 60%, an indication of the important role of M. merlangus for Trichodina spp. in the studied region. A multivariate analysis revealed that the magnitude of prevalence was related to the level of all three parameters: nitrite, nitrate and phosphate (r2=0.59). However, a much stronger relationship between prevalence and nitrate, phosphate, oxygen and temperature (r2=0.89) was detected. Consequently, the seasonal parasite prevalence and density were affected by organic pollution. The timing for a high prevalence and mean densities of this parasite also suggests that primary production may be responsible for the observed seasonal variation in prevalence and mean densities. By using the model describing the relationship between the water quality parameters and the prevalence data of Trichodina spp. recalculated values and observed field data corresponded closely.