Charles River Laboratories

Pharmaceutical manufacturers share a common goal of formulating a safe and effective product. To achieve this, a robust contamination control strategy is a necessity. The importance of this holistic approach is evident by the recent regulatory emphasis in Annex 1 and PDA’s Technical Report 88, “Microbial Data Deviation Investigations in the Pharmaceutical Industry.” However, despite manufacturers’ best efforts, microbial contamination events are all too frequent and demand a thorough investigation to identify root cause.

Poor root cause investigations are commonly cited in FDA Warning Letters. An example from a recent letter states:

For all OOS results found by the retrospective review to have an inconclusive or no root cause identified in the laboratory, include a thorough review of production (e.g., batch manufacturing records, adequacy of the manufacturing steps, suitability of equipment/facilities, variability of raw materials, process capability, deviation history, complaint history, batch failure history). Provide a summary of potential manufacturing root causes for each investigation, and any manufacturing operation improvements. A comprehensive review and remediation plan for your OOS result investigation systems. The CAPA should include but not be limited to addressing the following:

• Quality unit oversight of laboratory investigations

• Identification of adverse laboratory control trends

• Resolution of causes of laboratory variation

• Initiation of thorough investigations of potential manufacturing causes whenever a laboratory cause cannot be conclusively identified

• Adequate scoping of each investigation and its CAPA

• Revised OOS investigation procedures with these and other remediations

Given the scope of what’s expected for a contamination investigation, it is worth investing in prevention and an environmental monitoring program.

  ​When contamination investigations are required, it is important for microbiologists to use all available tools to quickly find the source of the contamination 

When contamination investigations are required, it is important for microbiologists to use all available tools to quickly find the source. Technical Report 88 includes carefully consideredquestions to assist in the investigation. One of the key starting points is a secure microbial identification to the species level. This can help direct the investigation by reviewing the environmental monitoring data to see if the organism has been recovered. If so, strain typing is a valuable tool in identifying and confirming the source.

Strain typing is a molecular method that compares DNA sequences of conserved housekeeping genes to look for nucleotide differences. If two organisms are the same strain, it can indicate that they come from the same source. Thus, strain typing can aid in investigations, and this is supported by regulatory and industry guidance. “Any isolates should be identified at least to species and, if available, strain-typed to aid the follow-up investigations with isolates recovered from the manufacturing or material bioburden…If the same species of microorganism was recovered from EM or supporting activities during batch manufacturing or testing, these colonies should be identified to the strain-type level” (TR 88). Another PDA publication, “Microbial Identification: The Keys to a Successful Program”also states that “having strain typing as part of your identification scheme is very valuable” (pg. 77) and “strain typing provides the ability of tracking the contaminant to its root cause” (pg. 146). USP <1113> also explains the utility and benefit of strain typing.

If the microbial contaminant is the same as a laboratory QC strain, it’s important to first eliminate laboratory error. This can also be done through strain typing. For example, Staphylococcus aureus can be typed through the spaA gene and Pseudomonas aeruginosa can be typed through the ascA, guaA, mutL, and nuoD genes. The genes are sequenced, and then phylogenetic trees are generated and interpreted. In the example below, the laboratory QC strain Pseudomonas aeruginosa ATCC# 9027 was different from the product contaminant and thus the contamination was not due to lab error.