Done and Dusted
Microbial Solutions
Ken Henderson, PhD

Done and Dusted

AALAS showcases the latest findings on PCR of EAD, and there are many

Monitoring the health of research mice requires a lot of effort. For years, soiled bedding sentinels did a lot of the heavy lifting by detecting circulating pathogens that could pose a threat to the colony.

Will they soon be out of a job? The PCR-based surveillance of entire individually ventilated racks referred to as exhaust air dust (EAD) testing, which Charles River helped pioneer, continues to expand. More laboratory animal facilities are using it as a component of routine health monitoring and some, if not many, have replaced soiled bedding sentinels with PCR of EAD or at least are using fewer sentinel mice for a smaller list of agents.

This is obviously good news from an animal welfare perspective. Anything we can do to reduce our use of research animals is a positive development for the field. But the interesting thing from my perspective is that PCR of EAD really came about because of advances in nucleic acid detection technology and our collective desire to find better ways of tracking certain infectious agents—adenovirus for example—that for whatever reason did not transmit successfully through soiled bedding. Cage Exhaust SwabOnce we were able to demonstrate that you could effectively use exhaust air filters to swab for pathogens, and we had a high-throughput method liked PCR to test many different agents at one time, EAD was able to advance. And advance it has.

This week, at the American Association for Laboratory Animal Science's annual meeting in Phoenix, one of the largest gatherings of laboratory animal caretakers, veterinarians and facility managers in the world, we will get a chance to hear some of the latest findings on PCR of EAD and address some of the questions and concerns laboratories that are using, or are considering using it, have. It's a good forum to wrestle with these issues and share experiences. The large number of poster presentations, abstract platform sessions and group discussions shows how much the topic has grown in the two years since we first reported on the PCR of EAD at an AALAS conference. Last year there were seven papers, this year 15 posters and platforms are being presented during the five-day meeting on alternative health monitoring technologies, 13 of which deal in some way with PCR of EAD. Here is a sampling of some of the studies being highlighted:

  • Cornell University scientist Erin Daugherity will be presenting data that shows PCR of EAD is sensitive enough to detect pathogens not readily transferred via dirty bedding. In this case, the agent in question was Pneumocystis murina, a fungus causing chronic pneumonia in immune-compromised mice. The pathogen is hard to detect in dirty bedding but monthly EAD samples from vertical and horizontal plenums, which pipe air in and out from the individually ventilated caging (IVC) system, detected Pneumocystis within a month and was later confirmed by testing pooled lung tissue from 10 mice housed in the suspected cages. In contrast, the sentinel bedding system missed the Pneumocystis entirely.
  • A separate study from Johns Hopkins scientist Anna Goodroe evaluated PCR of EAD as an adjunct to the sentinel screening of Pneumocystis carinii in laboratory rats. This study came about after the lab had a hard time detecting and eradicating Pneumocystis with only sentinel screening. The JH team found that using rack exhaust plenum PCR in combination with serology worked better in identifying and eliminating the agent.
  • Another study Charles River conducted with Allentown Inc., a manufacturer of cage racks, evaluated a method that simplifies the process of EAD testing on IVC racks that do not use cage level filtration. An in-line filter manifold was inserted into the exhaust air stream of the plenum, thus making it easier to replace the filter. We found that PCR of EAD placed in the manifold detected agents not found in sentinel mice by PCR or using traditional methods.
  • There are also multiple papers being presented on alternative PCR testing systems more suited to IVCs that block the majority of exhaust air dust from exiting the case and therefore make plenum testing impossible. One study being presented by Philip Gerwin from Memorial Sloan-Kettering Cancer Center, and which Charles River was a collaborator on, expands on some of his earlier work. Gerwin studied the reliability of PCR testing of cage filter tops as a method for detecting five different types of pinworms and fur mites, and compared the results to PCR samples collected directly from the animals and traditional testing methods. The study found that the filter top PCR testing of cage filters was a reliable alternative for detection.
  • A PCR of EAD study conducted by the Salk Institute of Biologic Studies in La Jolla found some inconsistencies with PCR of EAD testing. The study focused on mouse norovirus, which is one of the most prevalent viral pathogens in rodents. The virus is transmitted by fecal-oral route and is shed consistently, so it is effectively transmitted to soiled bedding sentinels. The group found the soiled bedding sentinels detected the norovirus up to four weeks in both low and high prevalent colonies, while the PCR of EAD was positive at two weeks but alternated between positive and negative thereafter. Not too distant from these findings our lab and others have found that MNV can usually be detected by EAD PCR, but lower copies available for testing does challenge the system if samples are not correctly collected on accommodating rack systems. This discrepancy does underline that methods of testing and assays as well as sample collection method and even rack type may influence the success of an EAD testing program.

And some of those questions will, I'm sure, be raised during a panel discussion that I'm chairing with animal scientists who are using PCR of EAD and other alternative health monitoring technologies to talk about what's working and what's missing. The group includes Tina Grove of Princeton University, Julie Watson of Johns Hopkins, Sylvia Banks from the University of Oregon, Mark Bates from Virginia Commonwealth University and Erin Daugherity.

So stay tuned!

If you have would like to read more about PCR of EAD, check out this recent Eureka post and this recent article in R&D Magazine.

(Photos credit: Matt Newby, Charles River Labs)

How to cite:

Henderson, Ken. Done and Dusted. Eureka blog. Nov 2, 2015. Available: