Microbiological analysis of seawater and black mussels (Mytilus galloprovincialis Lamarck, 1819) of Sinop province and molecular characteriziation of isolated Escherichia coli strains

Fecal coliform and fecal streptococcus contamination at coastal waters is one of the common problems of public health all over the world. In this study, the physico-chemical and microbiological analysis of seawater and mussel (Mytilus galloprovincialis) samples collected from 15 different stations in Sinop between May-2011 and October-2011 were investigated. The characterization of a total of 41 strains of Escherichia coli including 40 native isolates and 1 reference strain also were carried out according to phenotypic characterization, antibiogram, whole cell and extracellular protein profiles, plasmid DNA profiles and randomly amplified polymorphic DNA (RAPD) method.According to microbiological analyses, it was shown that the numbers of fecal coliform and fecal streptococcus for the seawater and mussel samples were over recommended values in some stations. Further, the microbiological analyses of seawater examples collected from different stations showed that the difference between total coliform, fecal coliform and fecal streptococcus numbers (p<0.05) was significant for each station, while the difference was not significant for total aerobe bacteria numbers (p>0.05). In addition, the difference between whole counting results for mussel samples collected from different stations was not significant (p<0.05), too.According to morphological and physiological properties, it was determined that the native isolates belonged to E. coli species. The antibiotic susceptibility of all strains against 7 standard drugs was tested using disc diffusion method. In accordance with the result of antibiogram, the antibiotic resistivities of the isolates were as bacitracine (100%), novabiosin (100%), amphicilline (12.5%), tetrasiklin (7.5%), seftazidim (5%) and imipenem (2.5%), respectively. On the other hand, it was found that whole strains were susceptible to polimiksin B (100%).The numerical analysis of whole cell protein profiles revealed that E. coli strains were separated to 2 major groups at similarity levels of 72%, 78% and 87% or above. Moreover, the numerical analysis of extracellular protein profiles of strains were generated 2 major groups at similarity levels of 75%, 78% or above. Eventually, it was pointed out that extracellular protein profiles provided better distinction than whole cell protein profiles.Plasmid DNA analyses results confirmed that 22 strains consisted of plasmids molecular weight changing between 24.500-1.618 bp. The plasmid molecular weighing 14.700 bp was found as common 21 of these strains.A numerical analysis of the DNA band profiles obtained by using 2 different primers of 20 E. coli strains was carried out by using UPGMA method. A dendrogram generated by using RBM-6 primer revealed that 13 strains produced bands divided into 2 major clusters at similarity levels of 12% or above. However, a dendrogram generated by using COL-I primer displayed that 14 strains produced bands scattered into 2 major clusters at similarity levels of %26 and over.Consequently, in the study, the microbiological analysis results of the samples of seawater and mussel collected from some stations indicated that microbial pollution degree had reached to serious size. It was also determined that while the morphological and physiological properties, the numerical analysis of whole cell and extracellular protein profiles produced by SDS-PAGE technique at the discrimination of isolated E. coli strains was enough at the species level, RAPD-PCR technique can be provide fast and more useful differentiation at the strain level.Keywords: Fecal coliform, E. coli, Antibiogram, SDS-PAGE, Protein Profiles, Plasmid, RAPD-PCR