PROCEDURE FOR BACTERIAL CONTAMINATION & PATHOGEN TESTING OF DRAINS

1. OBJECTIVE:

To lay down a procedure for Procedure for bacterial contamination & pathogen testing of drains.

• SCOPE:

This procedure is applicable for procedure for bacterial contamination & pathogen testing of drains.

• RESPONSIBILITY:

Microbiologist

• ACCOUNTABILITY:

QC Manager.

•  PROCEDURE:
  • Collect the sample as on weekly basis.
  • Take sterile 250 ml of sachatte bottle & sterile pipette (25 ml) and bulb. Take sample by using sterile pipette 25 ml about 50 ml.
  • After received the sample note down of detail of drains of sample in Drain Sample Inward in Format.
  • Analysis the sample as respect parameter:
  • Microbial contamination
  • Total bacterial contamination
  • Total Fungal Contamination
  • Pathogen
  • Total Bacterial Contamination:
  • Take 1 ml of sample by sterile pipette / micro pipette in duplicate sterilized Petri dishes 90 mm. Pour 15 – 20 ml of SCDA, mix and incubate at 30°C to 35°C for 5 days.
  • Total Fungal Contamination: Take 1 ml of sample by sterile pipette / micro pipette in duplicate sterilized Petri dishes 90 mm. Pour 15 – 20 ml of SDA, mix and incubate at 20°C to 25°C for 5 days.
  • Pathogen: E. coli:
  • Take 1 ml from sample solution and incubate with 100 ml soybean casein digest broth at 30 – 35⁰C for 18 -24 hours.
  • After incubation shake the broth and transfer 1 ml to 100ml of MacConkey Broth, incubate at 42 – 44⁰C for 24-48 hours.
  • After completion of incubation, subculture on a plate of MacConkey agar and incubate at 30 – 35⁰C for 18 -72 hours.
  • Acceptance Criteria:
  • If Growth of pink, non-mucoid colonies indicates the presence of Escherichia coli.
  • Maintain the record.
  • Pathogen: Salmonella:
  • Take 10 ml of sample and incubate with 100ml of soyabean casein digest broth at 30 – 35⁰C for 18 -24 hours
  • After incubation shake the broth and transfer 0.1 ml to 10 ml of Rappaport Vassiliadis Salmonella Enrichment broth medium and incubate at 30 – 35⁰C for 24 -48 hours.
  • After completion of incubation, subculture on a plate of Wilson and Blair’s agar and incubate at 30 – 35⁰C for 24 -48 hours.
  • If Growth of Green colonies with black center develop and in 48 hours the colonies become uniformity black. Colonies surrounded by a dark zone and metallic sheen indicates the presence of Salmonella.
  • If sub cultured on plates of Xylose Lysine Deoxycholate agar and incubate at 30 – 35⁰C for 24 -48 hours. Well developed, red colonies with or without black center indicates possibility of Salmonella.
  • Acceptance Criteria:
  • If Growth of red colonies with or without black center indicates the presence of Salmonella.
  • Maintain the record.
  • Pathogen: Pseudomonas aeruginosa:
  • Take 1ml of sample and incubate with soyabean casein digest broth at 30 – 35⁰C for 18 -24 hours.
  • After completion of incubation, subculture on a plate of Cetrimide agar and incubate at 30 – 35⁰C for 18-72 hours.
  • Acceptance Criteria:
  • If Growth of greenish colony indicates the presence of Pseudomonas aeruginosa.
  • Maintain the record.
  • Pathogen: Staphylococcus aureus:
  • Take 1 ml of sample and incubate with soyabean casein digest broth at 30 – 35⁰C for 18 -24 hours.
  • After completion of incubation, subculture on a plate of Mannitol salt agar and incubate at 30 – 35⁰C for 18 -72 hours.
  • Acceptance Criteria:
  • If Growth of yellow or white colony with yellow zone indicates the presence of staphylococcus aureus.
  • Maintain the record.
  • Acceptance Criteria for Microbial Contamination & Pathogen:
  • Microbial Contamination: To be record.
  • Pathogens: Should be absent.
  • Frequencies for sampling and analysis: Every Saturday (If Saturday is OFF, sample and analysis of drain point on next working day).

• TRAINING:

Trainer   : Manager – Quality Control

Trainees : Staff of the microbiology departments

• DISTRIBUTION:

Master Copy                  :           QA Department

Controlled Copy            :           Microbiology Department

Display Copy                 :           Microbiology Department (If Required)

• REFERENCES:

In-House

1. ABBREVIATIONS:

Abbreviations	Extended Form
SOP	Standard Operating Procedure
NA	Not Applicable
MB	Microbiology
Sr. No.	Serial Number
QA	Quality Assurance
DEPT.	Department
IPA	Isopropyl Alcohol
%	Percentage