Enterococci Research Paper
MATERIALS AND METHODS The Present study was carried out in the Department of Microbiology ,SRM Medical college Hospital and research centre from 2012 to .A total of 128 consecutive isolates of Enterococci were obtained from clinical specimens like urine,pus,blood and other body fluids in the diagnostic section of Central Microbiology Laboratory and processed as per standard recommendation procedures.
Inclusion criteria: All the Enterococcal isolates from clinical samples such as urine,pus,blood,wound swab,catheter tip and other body fluids are included.
Exclusion criteria: All commensal enterococcal isolates from anatomical sites like gastro-intestinal tract,female genital tract and oral cavity are excluded.
PROCESSING
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Positive test is indicated by the development of cherry red colour .
Quality control : Positive - Enterococcus faecalis ATCC 29212 Negative - Streptococcus agalactiae SPECIATION OF ENTEROCOCCI
SUGAR FERMENTATION TESTS: For identification of species, 1% of sugars(Glucose, arabinose,raffinose,mannitol,sorbitol,sucrose,lactose ) in peptone water with Bromothymol blue (0.002%) as the indicator was used. To each tube of Sugars,1-2 drops of 18-24 hour BHI broth culture is added and incubated at 37 C overnight.
Interpretation: Sugar Fermentation is indicated by the change of colour to yellow.
Arginine Dihydrolase test : Moeller's decarboxylase basal broth with 1% Arginine along with an aminoacid free control were inoculated with the test strain.Both the tubes were overlaid with sterile liquid paraffin and incubated at 37 C overnight.The colour of the indicator reverting back to original (purple) indicating arginine hydrolysis was considered as positive provided the control tube remains yellow indicating fermentation.
Quality control :Positive - Enterobacter
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PCR program : Initial denaturation at 94C for 3 minutes followed by 35 cycles of denaturation at 94 C for 1 minute, annealing at 58 C for 1 minute,extension at 72 C for 1 minute.The final extension was at 72 C for 5 minutes.
Analysis of PCR products:
PCR products were subjected to 2% Agarose gel electrophoresis using TAE buffer.5 µl of Ethidium bromide was added.Electrophoresis run at 50V .Gel viewed in UV Transilluminator and documented using gel documentation
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