Microorganisms identification process

• Any specified identification techniques of the In-house isolates and their library will help pharmaceutical to follow the proper guidelines.
• Mainly three method of identification process.

1. Phenotypic identification

2. Proteotypic identification

3. Genotypic identification

• The genetypic method is considered to be more accurate than the phenotype which useful for sterility testing.
• Genotypic identification uses the DNA sequences where are phenotype identification involves describes the protein or peptide sequence.
• Accurate classification of un known bacterial isolates is an essential first step in understanding the impact these organisms have on an environmental monitoring programme.
• They are many methods, technologies and strategies utilized to determine the identity of unknown microorganisms, however the selection of these methods is often impacted by more than performance of technology.
• Cost, time and amount is impact the identification process.

1. Proteotypic identification : Bruker Bio Typen  MALDI-TOF spectroscopy.

2. Metabolic Profiling :  Biomerieux -VITEK 2 Compact 

This equipment given the results based up on biochemical tests.

3. DNA sequence analysis:

Related : Environmental Monitoring Isolates and water Isolates

What is role of viable monitoring programme in pharmaceutical manufacturing facility?

• Environmental monitoring programme(Viable monitoring programme) to prevent product contamination before release on the market.
• In any environment where humans operators are present, microbial contamination at some level is inevitable. 
• Zero contamination at all locations during every aseptic processing operation is technically not possible and this is unrealistic.
• The complete absence of growth  on a microbial sample means only that growth was not discovered, it doesn't means that the environment is free of contamination.
• Viable monitoring programme involves  reduce the microbial contamination on pharmaceutical product. 
• If any viable monitoring excursion was observed during filling activity or any critical activity, that batch was not discarded, before release that batch proper investigation is required. 
• These excursion  are key elements of  proper investigation and identification of contamination.
• In the case of an isolated single excursion, establishing a definitive cause probably will not be possible, and only general corrective measures can be considered.
• Corrective actions may include but are not limited,

           1. Revision of the sanitization program.

           2. Selection of antimicrobial agents.

           3. Review of microbiological sampling methods and techniques

           4. Increased surveillance of personnel practices, possibly including written critiques of aseptic                methods and techniques
            5. When higher than typical recovery levels for glove and garment contamination are                             observed,additional training for gowning practices may be indicated.

           

• The viable monitoring programme ensure that an aseptic processing area maintained in an adequate level of control.
• Viable monitoring programme is a qualitative exercise and semi quantitative exercise this not given more accurate results such as aseptic processing, conclusion regarding lot acceptability should not be made on the basis of environmental sampling results alone.
• Environmental monitoring cannot prove or disprove in absolute terms the sterility of a lot of product.
• Environmental monitoring can only assure those responsible for a process that a manufacturing facility consistent, validated state of control. Care should be taken to avoid drawing inappropriate conclusions from monitoring results.
• Environmental microbial monitoring and analysis of data by qualified personnel can assist in ensuring that a suitable state of control is maintained.

What is the importance of maintain In-house isolate and their library in sterile pharmaceuticals?

• Identification of environment isolates very important and mentioned in different guidelines.(USP General chapter 1116, EU GMP, WHO-GMP and PIC/S).
• A lot of microorganisms are found in the air and water of pharmaceutical manufacturing area.
• These organisms some of these may be pathogenic and non pathogenic.
• Pathogenic organisms harmful for the products and humans, these should be addressed  and monitored frequently.
• These environmental Isolates are main source of the product contamination.
• Use full of investigation of source of contamination in pharmaceutical product.
• Every new isolate found  should be investigate and possible source.
• Once In-house isolates are identified, it's important to have a complete information (Library) of the organisms. Likes characteristics, photographs,Type of of identification tests and Identification results, source of possible contamination.
• In-house library should be maintained and updated regularly must be reliable and accurate.
• In-house library could be easily traced and if any previous history out come.
• These In-house isolates presence and using MLT validation, Disinfectant validation and sterility validation and to be prove these organisms also testing the supported to nutrient media.

As per USP

• A successful environmental control program includes an appropriate level of identification of the flora obtained by sampling.
• A knowledge of the flora in controlled environments aids in determining the usual microbial flora anticipated for the facility and in evaluating the effectiveness of the cleaning and sanitization procedures, methods, agents, and recovery methods. 
• The information gathered by an identification program can be useful in the investigation of the source of contamination, especially when recommended detection frequencies are exceeded.
• Identification of isolates from critical and immediately adjacent areas should take precedence over identification of microorganisms from noncritical areas.
•  Identification methods should be verified, and ready-to-use kits should be qualified for their intended purpose.

Chemical Indicators(Autoclave tape) composition

Chemical Indicator is called Process Indicator or Class-1 Indicators or Autoclave tape.

Chemical Composition:

1. Semi-Bleached craft paper
2. Natural Rubber Saturent
3. Natural Rubber adhesive
4. Butylated Urea- Formaldehide resin
5. Acrylate copolymer
6. Lead  carbonate hydroxide
7. Ethyl Alcohol

• Process indicator tapes with lead or without lead available.
• Lead is very poisonous metal, this material is neurotoxic and can cause brain disorders.
• Process indicator tapes are available in the market with different brand names.
• Process indicator tapes which contains lead basically converted in to light colour to dark after processing in sterilization.
• Disposal of these tapes containing lead is very important. These tapes can't be disposed off anywhere because it can cause health risk to living beings.
• These lead containing tapes should be disposed off as hazardous chemical waste by taking precautions.
• But now a days, there is another option available in the market that is class 1 indicator tapes without lead.
• These tapes are very safe to use and doesn't come under the category of hazardous waste.
• Before purchase you can check whether lead free is mentioned on the tape or not, and verify the MSDS also.
• In pharmaceutical companies, class 1 indicator tapes with lead or without lead are commonly used.
• Autoclave tape is also called adhesive tape or process indicator or chemical indicator.
• Autoclave tape Verify the sterilization process.
• Autoclave tape is an adhesive tape used in autoclaving (Heating under high pressure with steam to sterilize) to indicate whether a specific temperature has been reached.
• Autoclave tape works by changing color after exposure to temperature commonly used in sterilization process, typically 121 degree C in a steam autoclave.

Related : Classification of Chemical indicators