The APHI Laboratory
Small Beginnings to Valuable Institute Resource
The APHI laboratory was started approximately 25 years ago to enable the continuation of a collaborative research and outreach project between CSU and CDC that aimed to modify an available enzyme-linked immunosorbent assay (ELISA) and conduct validation trials. Today, the APHI laboratory is a centralized resource for Institute faculty and local, regional and international collaborators. Its mission is to utilize a broad range of techniques to provide the capacity for disease diagnosis and preventive measures through applied research. Although the scope of laboratory activities performed is extremely diverse, the focus is research and training related to the diagnosis and control of infectious diseases important to animal agriculture, trade, and public health in the United States.
Basic Research Linked to Population-level Application
Responding to the charge to link basic research findings to the diagnosis and prevention of animal diseases on a population basis, laboratory personnel explored the application of diagnostic and screening tests for important animal diseases in US livestock and wildlife populations. A five-antigen ELISA was developed; validity of the ELISA was determined using several thousand cattle and cervid serum samples. Investigators examined various alternative antigens for their usefulness for the diagnosis of TB in other species. As a result of this work, a multiple-antigen ELISA was found to be very useful as a screening test for TB in elephants.
In addition to a bovine TB screening test validation, a rapid and mass sample testing system was applied for the surveillance of Chronic Wasting Disease (CWD) in US deer. This application used an existing rapid test for Bovine Spongiform Encephalopathy (BSE) in use in Europe and assessed its suitability as a test for CWD. This assessment led to the application of a reliable surveillance system for CWD in several states in the USA. Work within the APHI lab was instrumental in the support of the Institute’s mission to conduct applied animal health research that results in the development of animal health policy for improved decision-making.
Since its creation, the APHI laboratory continues to innovate as it supports investigators with diagnostic tools for their research activities. Laboratory personnel have utilized a broad range of techniques to determine disease status in animal populations and molecular approaches to study different animal disease agents. The APHI laboratory is unique in that it has provided the capacity within one facility for applied research in bacteriology, virology, molecular biology, assay development and optimization, serology, and training in various techniques.
Since its inception, the APHI laboratory has been a training site for more than 40 undergraduate, graduate, and post-doctoral students as well as visiting scientists from several countries. These individuals were trained in diverse skills relative to veterinary epidemiology and related fields. Furthermore, student researchers and visiting colleagues received help from APHI laboratory staff on their research projects. Laboratory assistance varied; several researchers needed training in laboratory techniques or help with sample processing. Other researchers required much more, including the development of laboratory methods before beginning their projects.
In addition to providing specific contributions to student research projects, APHI laboratory personnel develop and optimize molecular approaches to study agents of animal disease. Their expertise has been shared through training courses and collaborations with visiting research scientists. For example, APHI Laboratory personnel developed and experimentally evaluated a single-tube one-step multiplex reverse transcription – polymerase chain reaction (RT-PCR) for the detection and differentiation of VSV strains. Although RT-PCRs for VSV had been reported for VSV, they were not routinely performed in diagnostic laboratories and the diagnostic sensitivity had not been reported. The PCR developed and optimized by APHI laboratory personnel was rapid, sensitive, and reliable. This test detected VSV RNA in infected insects at a level surpassing that of isolation and correctly identified the VSV virus serotype. This work was continued by a visiting scientist from the National Veterinary Laboratories in Islamabad, Pakistan, as he collaborated with APHI laboratory personnel to learn molecular diagnostic methods. His work improved the sensitivity of the original multiplex RT-PCR for the detection and differentiation of VSV-IND and VSV-NJ in a single assay.
Current Research, Facilities, and Capabilities
Currently, APHI laboratory personnel are actively involved in the research of several important animal diseases – strangles in horses, clostridium infection in cattle, and avian influenza. Additionally, the APHI laboratory is starting work on a new, innovative project – using volatile organic compounds (VOCs) and bacterial nucleic acids to detect TB, MAP, and brucellosis in animals.
The APHI Laboratory is well equipped with basic laboratory equipment – incubators, refrigeration units and freezers, microscopes with cameras, mixers, pH meters, refrigerated centrifuges, balances – in addition to autoclaves, biosafety cabinets, laminar flow hoods, spectrophotometers, a luminometer, fluorometer, Experiontm DNA, RNA, and protein bioanalyzer, chemiluminscent, fluorescent and visible light digital imager for gel and blot analysis/documentation, automated microplate incubators and washers, bead mill homogenizers, recirculating water baths, and thermal cyclers, including one for Real Time PCR.
The facilities and equipment available in the APHI Laboratory enable the fulfillment of the laboratory mission to be a centralized resource with the capacity for disease diagnosis and preventive measures through applied research in bacteriology, virology, and molecular biology. Laboratory personnel continue to utilize a broad range of techniques to determine disease status in animal populations and molecular approaches to study different animal disease agents.
- Development and optimization of methods for antigen and antibody detection, including ELISA, dot blots, western blots, and rapid strip tests.
- Development and characterization of hybridoma cell lines.
- Development, optimization and/or adaptation of molecular methods for pathogen detection from a multitude of sample types.
- Enhancement of utility of existing tests by the adaptation, development and application of optimal methods for processing and testing of both conventional and unconventional sample types for evaluation of disease status.
- Development and adaptation of methods for extraction of DNA and RNA from various samples – feces, whole and processed tissues, swabs.
- Diagnostic test performance evaluation and validation. The goal of test implementation in the field setting requires rigorous quality control and validation of such tests, and sets the APHI laboratory apart from the typical diagnostic laboratory or classic molecular research laboratory.
- Investigation of novel vaccination strategies, including the use of RNA interference.
Contact: Joni Triantis Van Sickle, Laboratory Coordinator