Importance of an Efficient Quality Assurance System in Microbiology Testing

In the nutraceuticals, pharmaceuticals, and cosmetics industries, quality means conformance to standards set by both the manufacturer and the industry. These standards must not only reflect a strong emphasis on the desired features of a product but also on its safety and efficacy.

To maintain quality, quality control (QC) procedures are employed. These procedures are in place to identify any action that does not meet the specified standards for quality. As a tool, QC is said to be corrective in nature. Its goal is to find and eliminate where problems in quality originate from. An example of QC in a typical micro laboratory is the regular monitoring of reagents and test samples.

Microbiology Instruments Appropriate for Enzyme Testing

Enzymes are intricate, stable, globular proteins that serve as catalysts in chemical reactions. These reagents reduce the activation energy in order to increase the rate of the reaction at the same temperature. Their stability prevents them from being consumed like other reagents in the reaction. While they can accelerate chemical reactions by more than 1,000 times, the effect of enzymes can either be enhanced by promoters, or reduced by inhibitors. Most industrial enzymes are obtained from microorganisms like fungi and bacteria, though they can also be obtained from animal and plant sources, unless they are activated.

Nutraceutical Rapid Microbiology Detection Systems

Consumers want quality assurance and the FDA is backing them with their cGMP guidelines. Many manufacturing companies are choosing to build in-house programs or signing up with contract labs to beat their existing microbiology quality assurance practices. As a manufacturer, you should always be prepared to test your raw materials and products for unacceptable levels of certain microorganisms.

The Pharmaceutical Industry and Its Dependence on Microbiology

Put simply, microbiology entails the study of microorganisms that one can't see with the naked eye. These are from bacteria, fungi, protozoa and such similar organisms. There is an intricate association between microbes to specific diseases. This has kept scientists at the front on research concerning them for centuries. The role of microbiology on advances within the healthcare industry, and especially the pharmaceutical industry, has led to great discoveries ranging from medicines and vaccines to devices.

Microbiology Quality Assurance and Rapid Microbiological Methods

When it comes to microbiology quality assurance, this is the process whereby the quality of laboratory reports can be guaranteed. Quality control is the part of quality assurance that is mainly concerned with controlling errors when tests are being performed as well as in the verification of the test results. Materials, equipment and procedures have to be controlled adequately. Culture material has to undergo testing for sterility as well as performance. Every laboratory should have Standard Operating Procedures (SOPs). Quality assurances should be incorporated at every stage of a microbiological procedure, which can be done by ensuring that the laboratory is well lit, dust free and air-conditioned. 

Understanding Microbiological Quality Control and Assurance Solutions

In the ever-changing pharmaceutical industry, competition is rife when it comes to the speedy development of products. Microbiological testing is essential to ensure safe products are made without any environmental hazards and at a reduced cost. This has led to an increased use of rapid detection kits that cut costs and time in detection of pathogens in any given sample. 

Regulations and Requirements for Pharmaceutical Microbiology Labs

In microbiology, quality assurance stands for the measures put in place to ensure quality control, as well as quality improvement. Quality control stands for the constant observation of work practices, paraphernalia and chemical substances so as to spot and correct defects. Once there are proper quality control measures in place, then quality improvement comes into focus as a way of making existing systems more efficient.

Rapid Microbiological Testing of E. coli with the BioLumix Vial

Escherichia coli are Gram negative rod-shaped organisms found naturally in the lower intestines of warm blooded organisms. Most serotypes of this organism are relatively harmless, making up a small percentage of bacterial colonization in the gut. These serotypes prevent the establishment of pathogenic strains.

E. coli is one of the most common bacteria found in the gut of animals. This includes humans. There are other animal species that contain this organism in the gut including reptiles and fish. E. coli colonizes the gut and can cause infection in the urinary tract and brain stem (meningitis) as well as intestinal diseases referred to as gastroenteritis. There are five classes of E. coli that produce disease. The most serious disease is the Enterohemorrhagic (EHEC) class. These organisms can cause diarrhea distinct from some others (including Shigella) in that there is copious bloody discharge and no fever. The life threatening situation is its toxic effects on the kidneys (hemolytic uremia).

Why Test for the Presence of E. coli?

E. coli is often used as indicator organisms to test the effectiveness of effluent disinfection in a wastewater treatment plant, on animal products as well as in nutraceutical and pharmaceutical products. While these organisms are generally harmless, they do live under the same conditions that human pathogens live. Since we cannot test for every pathogen, we test for easily detectable indicator organisms. The assumption is that if we kill the indicator organisms then we most likely kill the pathogens during effluent disinfection. E. coli has reemerged as an indicator, partly facilitated by the introduction of newer methods that can rapidly identify E. coli.

Current Methodology

The current methodology can take anywhere from 3-7 days and includes multiple broths, agars, transfers and temperatures. Different methodologies are utilized by the various industries. Below are some examples:

Nutraceutical and Dietary supplements: The protocol described in USP <2022> requires absence of the organisms in 10 grams of product. A 1:10 dilution of product is made into TSB or another enrichment medium. This mixture is incubated for 24 to 48 hours at 30°C to 35°C, and then 1.0 mL is transferred to 10 mL of MacConkey broth. This mixture is incubated for 24 to 48 hours at 42°C to 44°C. A loop from the MacConkey broth is transferred to MacConkey agar and the plate is incubated for 18 to 24 hours at 30°C to 35°C. If typical colonies appear, these colonies are then transferred to Levine Eosin Methylene Blue agar and incubated 24 to 48 hours at 30°C to 35°C. If none of the colonies exhibit green metallic sheen under reflected light or if none of the colonies exhibit a blue-black appearance under transmitted light, the sample meets the requirement for the absence of Escherichia coli. Because results can sometimes be misread due to interpretation of plates, identification may be run on the sample(s) adding another 2-3 days of testing.

Pharmaceutical: The protocol described in USP <62> again requires the absence of E. coli in 10 grams of product. A 1:10 dilution is made and incubated for 18 to 24 hours at 30°C to 35°C. From this mixture, 1.0 mL is transferred to 100 mL of MacConkey broth and incubated 24 to 48 hours at 42°C to 44°C. A loop is transferred to MacConkey agar and this plate is incubated 18 to 72 hours at 30°C to 35°C. Growth of typical colonies indicates the presence of E. coli which is confirmed with an identification test.

Food Testing: The food industry follows Bacteriological Analytical Manual (BAM). In most cases, the Most Probable Number (MPN) method is utilized. This is labor intensive, multi-step assay consists of presumptive (in LST tubes), confirmed (in BGLB tubes), and completed phases (in EC tubes). In the assay, serial dilutions of a sample are inoculated into broth media. Analysts score the number of gas positive (fermentation of lactose) tubes, from which the other 2 phases of the assay are performed and then uses the combinations of positive results to consult a statistical table. From this table, the analyst is able to estimate the number of organisms present. The 3-tube MPN test is used for testing most foods. The 5-tube MPN is used for water, shellfish and shellfish harvest water testing, and there is also a 10-tube MPN method that is used to test bottled water or samples that are not expected to be highly contaminated. Positive EC tubes must be transferred to L-EMB agar and if typical colonies are observed, they must be further identified.

The BioLumix E. coli Vial

The E. coli vial, or EC vial, is a membrane vial that is monitor by the fluorescent signal in the instrument. The membrane separates the incubation zone from the reading zone. If E. coli is present, it utilizes MUG (4-Methylumbelliferyl-3-D-Glucuronide) through an enzymatic reaction to create fluorescence. A 1:10 dilution is made in TSB or a similar growth medium and this mixture is incubated for 18-24 hours at 35°C. From the TSB dilution, 0.1 mL is added to an EC vial containing the MUG supplement and then tested in the BioLumix instrument. The EC assay runs for 18 hours in the instrument. An example of E. coli curves is shown in the graph. If detection occurs, a confirmation test is performed. From the time the sample is prepared to the time the confirmation is complete, the EC test takes only 2 days, saving valuable time.

The BioLumix assay is much simpler to perform, requiring less labor and disposables as any to the standard method. It is faster (completed in 30 hours), accurate, automated, and it eliminates any product interference due to the two zone vial.