Bacterial sterility testing of tissues for transplantation

The National Bacteriology Laboratory provides a centralised bacteriological service to the entire organisation.  As part of its remit, the NBL screens all tissues retrieved by NBS Tissue Banks, which comprise of 40% of those transplanted in the United Kingdom. The laboratory also provides scientific and technical advice and recommends acceptable decontamination methods.

The tissues screened are:

  • Bone (surgical and deceased donors)
  • Tendon
  • Amnion
  • Heart Valve
  • Skin
  • Menisci / Osteochondral

The rejection organisms are specific ‘pathogenic’ organisms such as: Pseudomonas aeruoginosa, Pyogenic streptococci, Clostridium spp., Staphylococcus aureus, fungi and non-viable organisms. However some tissues can still be approved for clinical use provided these samples undergo terminal sterilisation such as irradiation.The screening strategy for cadaveric tissue from deceased donor is based on bioburden determination and detection of specific rejection organisms. A high bioburden, presence of specific organisms or detection of any organisms after exposure to decontaminating agents result in the rejection of tissue. Bone-chip samples from all surgical bones (live donors) are also screened for any contamination.

Tissue samples are received in the NBL in inoculated broth bottles and/or culture plates. Aerobic and anaerobic cultures are performed on all tissues. Broth cultures (Thioglycollate and Tryptic Soy) are incubated at 35°C for 7 days from pre-decontaminated samples and 14 days for post-decontaminated samples. Blood agar plates are incubated at 35°C for 2 days and Sabouraud agar plates for 7 days.

A bar-code labelling system exists for all samples and a dedicated LIMS, called ‘Hematos’ allows the culture results to be entered directly into the computer, thereby obviating transcriptional errors.

The subculture of broths is performed under laminar airfow conditions using a fully automated subculture machine called ‘InocuLab’. Bacterial identification is undertaken when required using automated system such as ‘Phoenix’ which is based on biochemical reactions. Molecular tools such as PFGE and 16S sequencing are also under investigation.

Back to top


Environmental monitoring and identification of action level organisms

The National Bacteriology Laboratory (NBL) has devised and implemented an agreed national environmental monitoring system for both controlled and uncontrolled rooms within the NBS to comply with the current edition of:

  • The 'Rules and Guidance for Pharmaceutical Manufacturers and Distributors'. MHRA
  • The 'Guidelines for the Blood Transfusion Services in the United Kingdom'
  • European Directive 98/79/EC on in-vitro Diagnostic Devices
  • 'A Code of Practice for Tissue Banks for providing tissues of human origins for therapeutic purposes'. Department of health
  • 'Standards for Hematopoietic Progenitor Cell Collection, Processing & Transplantation'. JACIE
  • 'FACT-NETCORD standards'

National environment monitoring forms part of the quality management system for cleanroom and processing facilities, ensuring that products are processed to the highest possible standards and that microbial contamination does not present an unacceptable risk to product quality and safety.Processing areas are defined as either controlled or uncontrolled on the basis of how their air quality is maintained with regards to contamination with particulates and viable micro-organisms. Clean room facilities within the NBS are controlled rooms whereas, for example, processing laboratories are defined as uncontrolled rooms.

The laboratory provides an environmental monitoring service involving the devising of national environmental monitoring programmes, the provision of training and advice on the suitable use of media.

As part of national system, the role of the NBL is to provide an identification and reference laboratory service. The national system is in place for Tissue services at Edgware and Liverpool, Processing at Colindale, Brentwood, Tooting and Manchester, LCBB, Reagents laboratories and the SCI laboratories.

Action level organisms isolated from controlled rooms and from repeat non-compliant monitoring in uncontrolled rooms are identified to either genus or species level depending on the clinical importance of the isolate.

Identification of monitoring sites has been defined by the NBL with support from information and data derived from Hazard Analysis and Critical Control Point (HACCP) studies. These studies use a risk analysis based approach to identify critical points in a process where there is a risk of a microbiological hazard compromising the safety of the final product.

Back to top

Monitoring techniques

A - Air Sampling

Settle Plates

A settle plate is an agar plate that is exposed to the air for a specified time in a specified location. Airborne particles such as dust will fall on to its surface and any attached bacteria will grow on incubation. This test gives an indication of the levels of airborne contamination.

Air Sampler

Air sampling is used to check the microbiological quality of air in controlled rooms. A specially designed air sampling machine samples a predetermined volume of air and impacts it onto an agar surface which can be incubated and counted for colonies.

B - Surface Sampling

Contact Plates

A contact plate is an agar plate with its agar surface raised above the edge of the plate (pictured in donor arm monitoring). The agar surface is pressed against surfaces such as benches and floors to monitor surface contamination. Many contain disinfectant neutralizers to inactivate any residual disinfectant from post disinfection sampling.

Glove Prints

Five finger glove prints are taken on to agar plates from each hand of the operator following aseptic processing in controlled rooms. Glove prints are a means of assessing the potential transfer of bacterial contamination to sterile products as a result of handling by personnel during and/or after aspetic processing. Results are recorded as colony forming units (CFU) per plate. If the CFU has reached or exceeded the action level for the given monitoring site, an investigation and corrective action is instigated including cleaning, isolate identification and repeat monitoring.

Monitoring results are entered onto local database allowing trend analysis and result review.

Back to top

Also see