Standard Operating Procedures Bacteriology

Antimicrobial Resistance Surveillance and Research Network- 2019

This Standard Operating Procedure (SOP) manual is intended to assist as a reference material for routine laboratory activities of ICMR’s ‘Antimicrobial Resistance Surveillance and Research Network’ (AMRSN)

1. Introduction

This is a comprehensive standard operative procedure manual for all types of specimens received in a clinical bacteriology diagnostic laboratory serving a tertiary care hospital. The manual has been compiled by referring to international protocols customized to the needs and the infrastructure already available in India or infrastructure that can be achieved by upgradation. Both conventional and automated procedural alternatives are included. It is intended that all participating laboratories, including Nodal Centers and the Regional Center laboratories, will strictly adhere to the procedures. The manual has been organized to place each part of the procedure together, including sample collection, specimen processing, supplies, quality control (QC), and step-by-step testing procedure. This will allow the user to see an overview of the entire procedure together. Guidelines for specimen collection and transport can be separately made available to the collection points and those for processing are to be made available in the processing laboratories. All the laboratories must isolate, identify to species level and carry out susceptibility tests of significant bacterial isolates as per guidelines provided. For isolates, which are difficult to identify, Regional Center laboratories can send the isolates to Nodal Centers for further characterization.

2. Role of the laboratory

Microbiologists play a critical role in gathering data both for clinical and public health decision making. Efficient and accurate microbiologic diagnosis of bacterial infections guides the choice of antibiotics and other treatment options for the patient. Similarly, microbiological surveillance is critical to guide appropriate antibiotic therapy through the identification of local resistance profiles. Thus, the role of the microbiology laboratory is essential to preventing morbidity and mortality from bacterial infections.

3. Biosafety

Laboratorians working with infectious agents are at risk of laboratory-acquired infections as a result of accidents or unrecognized incidents. The degree of hazard depends upon the virulence and dose of the biological agent, route of exposure, host resistance, proper biosafety training and experience with biohazards. Laboratory-acquired infections occur when microorganisms are inadvertently ingested,
inhaled, or introduced into tissues.

While laboratory-acquired infections are not as extensively reported, deadly infections with any of the organisms are possible if appropriate biosafety procedures are not strictly followed in a properly equipped laboratory. Biosafety Level 2 (BSL-2) practices are required for work involving these agents as they present a potential hazard to personnel and the environment.

4. Disinfection

Organisms may have different susceptibilities to various disinfectants. As a surface disinfectant, 70% isopropyl alcohol is generally effective. However, 70% alcohol is not the disinfectant of choice for decontaminating spills. It should be noted that 100% alcohol is not as effective a disinfectant as 70% alcohol. Phenolic disinfectants, although expensive, are effective against many organisms. Phenolic compounds (e.g. Chloroxylenol) are active against vegetative bacteria and lipid-containing viruses and Mycobacteria. They are not active against spores and their activity against nonlipid viruses is variable. Always read disinfectant labels for manufacturers’ recommendations for dilution and for exposure times for efficacy.

An effective general disinfectant is a 1:100 (1% or 10,000 ppm) dilution of household bleach (sodium hypochlorite) in water; at this dilution, bleach can be used for wiping surfaces of benches, hoods, and other equipment. A 1:100 dilution of bleach (10,000 ppm of available chlorine) should be used to clean up spills of cultured or concentrated infectious material where heavy contamination has occurred; however, it is more corrosive, will pit stainless steel, and should not be used routinely. If bleach is used, wipe down the area with 70% alcohol to inactivate the bleach. If bleach is used as a disinfectant, the diluted solutions should be made weekly from a concentrated stock solution.

Refrigerators and freezers

The temperature of laboratory refrigerators and freezers should be monitored daily to ensure that they are functioning properly. They should also be regularly inspected for the presence of broken vials or tubes containing infectious agents. When removing and discarding broken material, laboratorians should wear gloves and PPE. If the broken material is suspected of being infectious, disinfectant should be applied to the affected area and kept in place for at least 15 minutes before removal of the broken material. Refrigerators and freezers should be regularly cleaned with a disinfectant and defrosted to prevent possible contamination or temperature failure.


Standard Operating Procedures 2019