Detection of infectious agents from environmental air dust

Inventors

Henderson, Kenneth S. • Coiro, John M. • Bilecki, Brian M. • Schupsky, Thomas P.

Assignees

Charles River Laboratories International Inc

Abstract

Embodiments of the present disclosure are directed to systems and methods for collection and analysis of environmental air dust (EAD) within an individually ventilated cage rack (IVR) environment for detecting pathogens. The method includes collection of an EAD sample by a collection media, isolation of a plurality of nucleic acids (e.g., RNA and/or DNA) representative of one or more infectious agents from the EAD sample, optional reverse transcription of RNA to cDNA if the isolated nucleic acids contain RNA, amplification of the cDNA and/or DNA (e.g., by polymerase chain reaction (PCR)), and assay interpretation. Optionally, the EAD sample may be analyzed with one or more other sample types (e.g., fecal pellets, oral swabs, body swabs, tissue, etc.) to improve detection of low-copy organisms.

Core Innovation

The invention describes a method for detecting at least one pathogen using environmental air dust captured from airflow within an exhaust plenum of an individually ventilated cage rack (IVR). A test sample comprises environmental air dust captured by a collection media, where a surface of the collection media is oriented at an acute angle with respect to a direction of airflow within the exhaust plenum during capture. Nucleic acids are isolated from the test sample, amplified, and analyzed to identify the presence or absence of at least one pathogen.

The capture geometry can be defined by an acute angle oriented relative to airflow, including an angle between 15° and 25°. Nucleic-acid isolation includes isolating a plurality of nucleic acids and, in refinements, is performed using magnetic isolation, column-based nucleic-acid isolation, organic extraction methods, or alkaline lysis. The workflow includes amplifying at least one of the plurality of nucleic acids using loop-mediated isothermal amplification or polymerase chain reaction (PCR), followed by analysis of amplified nucleic acids.

In an additional configuration, the invention describes detecting one or more pathogens attached to environmental air dust by receiving a test sample comprising environmental air dust captured from airflow passing through an enclosure by air filtering collection media within an exhaust plenum. The enclosure includes a chamber containing an animal cage in fluid communication with the airflow, and the environmental air dust is released by agitation of soiled bedding by a test animal positioned within the animal cage. As in the IVR approach, nucleic acids are isolated, amplified, and analyzed to identify the presence or absence of a pathogen using a collection-media surface oriented at an acute angle relative to the direction of airflow within the exhaust plenum during capture.

Claims Coverage

The independent claims cover two main detection methods. Across these claims, inventive features include acute-angle oriented collection media in an exhaust plenum for environmental air dust capture, nucleic-acid isolation and amplification from the dust, and pathogen presence/absence analysis; the second independent claim adds enclosure-based dust release by agitation of soiled bedding by a test animal.

Collectively, the independent claims define pathogen detection from environmental air dust captured in an exhaust plenum using collection media with an acute-angle orientation, followed by nucleic-acid isolation, amplification, and analysis to identify pathogen presence or absence. The second independent claim further requires an enclosure in which dust is released by agitation of soiled bedding by an animal in fluid communication with airflow to the exhaust plenum.

Stated Advantages

Documented Applications