Bacteriology (Molecular Characterisation of Bacterial Pathogens

In this study, Enterococcus faecalis was isolated from100 samples of apparently healthy and diseased cultured Tilapia fish (Oreochromis niloticus) collected from 4 different fish farms at different localities in Kafr El-Sheikh governorate with an incidence of (20%) and it, also isolated from 12 samples (8 water samples and 4 ration samples) at the same infected farms with incidence of 50% from water samples, but the ration samples give a negative result for Enterococcus faecalis isolation. The antimicrobial susceptibility test has revealed that isolates were sensitive to vancomycin, amoxicillin, ampicillin, norfloxacin, gentamicin and ciprofloxacin, while resistant to cefotaxime, tetracycline, streptomycin and erythromycin. Molecular detection of some virulence genes from 8 Enterococcus faecalis isolates revealed that 3 isolates produced esp gene, 7 isolates produced asa1gene and all isolates produced EF3314 gene, while the cylA was not detected in any of Entero-coccus faecalis isolates. Also PCR results of some resistance genes showed that 8 isolates and 7 isolates were found to produce blaz and tetk genes respectively. Experimental infection test with some isolated Enterococcus faecalis revealed that the isolates caused mortality in fish with characteristic disease signs.

Background: Standard protocols for Pulsed-field gel electrophoresis (PFGE) were adopted and being used in a global scale for surveillance of many bacterial food-borne diseases. Matched PFGE bands are considered according to values of retention factors (rFs) regardless of co-migration of different DNA fragments (having equal or very close molecular weight). Molecular epidemiology is turning toward whole genome sequencing (WGS) which its results are compared using different DNA sequence alignment methods. Although, WGS results can be digested In-silico, PFGE and WGS data are being compared separately. Methodology: To link results of both methods, we describe a new image analysis algorithm that enables identification of how many DNA fragments co-migrate during PFGE. We built a database that compares results of the previously mentioned algorithm to In-silico obtained digestion models (from WGS). Reliability of the image analysis algorithm was also assessed In-silico using novel computer simulation approach. From WGS, 1,816 digestion models (DMs) were obtained as recommended by PulseNet international. Simulation codes were designed to predict PFGE profiles when DMs are separated at 5% PFGE resolution in addition to expected co-migration levels. Results: PFGE simulation has shown that about 35% of DNA fragments co-migrate at 5% PFGE resolution. Similar result was obtained when wet-lab PFGE profiles were analyzed using image analysis algorithm mentioned earlier. In terms of number of PFGE typable DNA fragments, 45,517 were typable (representing 46.54% out of 97,801). Previously mentioned typable fragments (in terms of typable sizes) comprised 91.24% of the sum of nucleotides of all chromosomes tested (7.24 billion bp). However, significant variations were shown within and between different digestion protocols. When image analysis results were compared to DMs, results returned by geltowgs.uofk.edu database revealed reasonable relatedness (Dice coefficient of variation was 0.44) to the most related DM. Conclusions: Identification of co-migration levels will reveal the third dimension of PFGE profiles. This will provide a better way for evaluating isolate relationships. Linking old PFGE results to WGS by means of simulation demonstrated here will provide a chance to link hundreds of thousands of PFGE epidemiological data accumulated during the last 24 years to the new WGS era. Evaluation of population dynamics of pathogenic bacteria will be dee per through place and time. Selection of restriction enzymes for PFGE typing will have a powerful In-silico evaluation tool.

Sheep and lambs are threatened by a number of infectious diseases, among which enterotoxaemia that caused by Clostridium perfringens and other aerobic bacteria. A total of 54 faecal swabs (10 diarrhoeic, 44 apparently healthy) were collected from sheep of different ages in different localities in Alexandria governorate and examined bacteriologically. The results revealed that Clostridium perfringens, Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa could be isolated in a percentage of 44.5, 31.5, 11.1 and 9.3% respectively. Pathogenicity tests were carried out for aerobic bacteria and toxigenic typing of C. perfringens by multiplex PCR was performed which revealed that 25% of isolated C. perfringens were type A. An in vitro sensitivity test of C. perfringens type A was carried out against different antimicrobials and garlic oil, the results showed that it was sensitive to Ciprofloxacin (5µg), Amoxicillin/Clavulaniacid(10µg), Erythromycin(15 µg), Flumequine (30µg), Gentamycin (10µg), Chlormphenicol (30µg), Vancomycin (µg); Nitrofurantoin (300µg), intermediately sensitive to tetracycline (30µg), Enerofloxacin (5µg), and resistant to Amoxicillin (10µg), Neomycin (30µg) Streptomycin (10µg), Trimethoprim/ sulfamethoxazole (25µg) and garlic oil. Proper hygienic measures in the sheep farms and feeding animals with balanced ration are recommended to avoid the infection of sheep with such enteropathogens.

Bacteria play a major role in the biogeochemical cycling of toxic heavy metals. Heavy metal resistant bacteria can
be used as bioremediation agents. The aim of this present investigation was to determine the resistance property of
bacteria isolated from industrial waste dumped soil samples, to heavy metals (Pb2+ and Cd2+) and some antibiotics.
Of 29 isolates, only two bacterial strains were found to be highly resistant to lead and cadmium. Further, these two
bacterial strains were checked for resistance against heavy metals by culturing them in basal medium in which
varying concentrations of heavy metals were incorporated. HML8 and HMC1 isolates showed resistance against
Pb2+ and Cd2+ at all concentrations tested. The morphological and biochemical tests confirmed that the isolates
belonged to Bacillus sp. Both the isolates showed similar pattern of growth at different temperature ranging from
25⁰C to 42⁰C. HMC1 showed better growth in the presence of high NaCl concentration but the same was unable to
grow at high pH. On the other hand HML8 showed good growth in the basal medium at high pH. High salt
concentration affected the growth of HML8 in the basal medium. Both the isolates also exhibited high tolerance to
antibiotics such as Amoxicillin and Penicillin G. The results of the present study clearly suggest that both the strains
can be studied and explored further for biotechnological applications especially for lead and cadmium
detoxification.

Present work is a prospective study on UTI infected diabetic patients from Bangalore region, molecular character and antimicrobial susceptibility of Pseudomonas sp. infections. Among seventy seven samples, 10 % were identified as Pseudomonas sp. are isolated on C-TAB agar media and confirmed by morphological and biochemical tests. On testing antimicrobial activity by well diffusion methods, all organisms showed highest resistance to Ampicillin (77%) followed by amplified fragment length polymorphism. Dendrogram analysis of ten variables of Pseudomonas sp. by using different primer (E+CTT)(M+AAT) and (E+AAG) (M+GGC) gives total diversity among 10 accession showed within 5.4 units of genetic distance.
Key words: Diabetic, infection, antimicrobial, polymorphism, accession.