We diagnose pathogens by quantitative detection of signature nucleic acid sequences with real-time polymerase chain reaction (PCR) methodology. Our approach to microbiological diagnostics has been featured in eBIOForum Magazine (PDF).
Our lab is based on the application of patented real-time PCR methods developed for research with intracellular Chlamydia bacteria.
Our real-time PCR delivers reliable results with rapid turnaround and results available within 24 hours after sample receipt.
Our approach to FIP diagnosis has been featured in “Veterinary Medicine.” Please see the Feline Infectious Peritonitis assay (PDF) for details.
The Molecular Diagnostics Laboratory has now started a comprehensive case-control study to determine the predictive value of our FIP mRNA PCR, alone and in combination with other molecular methods and clinical-pathological symptoms typically associated with FIP. Please see a one-page abstract of the study here: Auburn University FIP Study (PDF).
Create your own CUSTOM PANEL at a discount – we now offer $10 multiple PCR discounts for every PCR after the first one submitted. The canine blood parasite panel for detection of Hepatozoon spp., Babesia spp., Anaplasma spp., Ehrlichia spp., and Borrelia spp. is the most popular combination requested by our clients. Please see our updated Molecular Diagnostics Submission Form (PDF).
All our real-time PCR assays are designed to detect single copies of nucleic acid targets in a large sample volume (equivalent to 0.1 mL of blood); and to recognize the relevant pathogens by use of fluorescent probes, differentiating them from closely related, but in the particular animal species less pathogenically relevant microbial organisms. Because of these strict quality standards, we typically offer the most sensitive and specific assays for these pathogens. Published investigations that use our assays continually confirm this and create NEW SCIENTIFIC INFORMATION that improves understanding and treatment of infectious diseases. Recent manuscripts from our laboratory were published in highly respected journals. These MANUSCRIPTS describe:
- a new understanding of canine Hepatozoon spp. infection (Li et al., 2008; Allen et al., 2008);
- improved diagnosis and treatment monitoring of Babesia spp. infection in dogs (Wang et al., 2010a);
- unambiguous discrimination of FIV-vaccinated from FIV-infected cats with the assay for feline immunodeficiency virus, and identification of the subtype of the detected FIV strain (Wang et al., 2010b);
- our Salmonella spp. PCR helped to investigate Salmonella enterica infections in leatherback turtle breeding grounds in the Caribbean (Dutton et al., 2013);
- our Chlamydia spp., E. coli O157:H7, and pathogenic Salmonella spp., and Listeria monocytogenes PCRs have been approved by the Food & Drug Administration (FDA) for monitoring of transgenically-produced biotechnology pharmaceutics;
- our pathogenic Leptospira spp. PCR has been proven the most sensitive detection method for leptospira infection in dogs (Xu et al., 2014);
- with custom-developed real-time PCRs we have helped to analyze the impact of the Gulf oil spill on fish health (Crowe et al., 2014);
- and recently we contributed with multiple assays, most prominently for heartworm, to new knowledge about blood-borne parasites in dogs in Nicaragua (Wei at al. Parasites & Vectors 7_126, 2014).
The Molecular Diagnostics Laboratory uses state-of-the-art technology to diagnose pathogens by quantitative detection of signature nucleic acid sequences. The lab is based on the application of patent-pending real-time polymerase chain reaction (PCR) technology developed in Dr. Bernhard Kaltenboeck’s laboratory. Assay costs are $ 80 each, and diagnostic results are available within 24 hours of sample receipt.
All assays are in-house developed for highest sensitivity (detection of single target molecules) and specificity (fluorescent probe detection). Such PCR diagnostics translate immediately into effectively targeted therapy rather than serving mainly as retrospective confirmation.
- Anaplasmosis (PDF)
- Bartonella henselae (PDF)
- Canine Babesiosis (PDF)
- Canine Distemper Virus (PDF)
- Canine Ehrlichiosis (PDF)
- Canine Hepatozoonosis (PDF)
- Canine Lyme Borrelia (PDF)
- Chlamydia spp (PDF)
- Escherichia coli O157:H7 (PDF)
- Feline Immunodeficiency Virus (PDF)
- Feline Infectious Peritonitis Virus (PDF)
- Heartworm (PDF)
- Hemotrophic Mycoplasmosis (PDF)
- Influenza A Virus (PDF)
- Leptospira spp (PDF)
- Listeria monocytogenes (PDF)
- Pseudomonas aeruginosa (PDF)
- Salmonella spp (PDF)
For detection of extremely low numbers of Chlamydia bacteria, we optimized existing real-time PCR technology (PDF) at several steps from sample collection through analysis of duplex reverse-transcriptase PCR assays.
- Sampling (PDF) was optimized for preservation of target nucleic acids in specimens. This is important for detection of low pathogen concentrations.
- Nucleic acid extraction (PDF) was improved for maximum concentration of rare target nucleic acids such that as few as 7 copies of a nucleic acids per milliliter of original sample (blood, urine, tissue, …) can be reliably detected.
- Chemistry (PDF) and thermal cycling strategy (PDF) of real-time PCR was optimized for robust amplification.
- For quantification of gene transcription, a method for simultaneous amplification of 2 target mRNAs (PDF) in a single real-time PCR was developed. This method provides high accuracy, speed, sensitivity, and ease of detection and quantification of gene activity, and is also the basis of our reverse transcription PCRs for detection of RNA viruses.
- New assays can typically be developed and fully validated within 2 weeks if target nucleic acids are available. We are continuously developing new assays for diagnostic targets of interest. We are also interested in suggestions for assays of diagnostic value and encourage inquiries about such new assays. Please contact us with your questions and/or suggestions.
The combined use of modified nucleic acid extraction and real-time PCR methodology in our diagnostic platform can generate results within 3 hours. Batch nucleic acid extraction followed by real-time PCR on the same day allows communication of results within 24 hours. Finally, the robust assay technology ensures reliable results.
- Identify sampling requirements for each assay.
- Collect Specimen: Sterile EDTA blood is acceptable for blood-borne infectious agents. Do NOT refrigerate or freeze specimens.
- Submit specimens with the Molecular Diagnostics Submission Form (PDF) by Courier or Mail to:
Molecular Diagnostics Laboratory
College of Veterinary Medicine, Department of Pathobiology
252-A Greene Hall
Auburn, AL 36849-5519
Phone: (334) 844-2648
- Specimen collection with our Sample Submission Kit ensures maximum sensitivity of the PCR. The buffer in the collection tubes is designed for optimum preservation of nucleic acids and their recovery in subsequent extraction. No refrigeration or freezing of samples is required. Please reference the submission kit’s Material Safety Data Sheet (PDF) before and during use.
- International Shipping: canine and feline samples may be shipped without US import permits. Please check the international shipping requirements (PDF).
1. What are the basics of your “molecular diagnostics”?
We first extract the nucleic acids from the samples, then use a technique called polymerase chain reaction (PCR) to detect the presence of pathogens. PCR is a Nobel prize-winning molecular biology technique that amplifies certain nucleic acids exponentially, with the help of an enzyme called Taq polymerase. With this technique, we can detect single copies of the target DNA and amplify it to billions of copies that we can easily detect and quantify. For the RNA viruses (such as canine distemper virus, influenza virus and FIP virus), we do RT-PCRs which involves conversion of RNA to DNA followed by PCR.
2. What samples do you need and how much?
We use EDTA-whole blood in most cases. We also accept other samples for certain tests and they may be more indicative of the positivity/negativity. For example, we can use cerebrospinal fluid (CSF) for canine distemper; abdominal fluid, feces or tissue/organ biopsies for FIP. Recommended samples are mentioned in the notes on each assay. Please make sure to ship EDTA-blood (purple-top tubes) instead of untreated blood, which will clot and cause problems during nucleic acid extraction and PCR. Blood treated with heparin (green-top tubes) for prevention of clotting should not be used because heparin interferes with nucleic acid extraction and PCR. For undiluted fluid samples, we use 400 microliters for our test, but we can process sample volumes as low as 10 microliters. So it’s OK if the sample submitted is less than 400 microliters, although that may decrease the sensitivity of the test. For swab samples, we don’t have a threshold for how much we can accept. If the sample is too large, we will extract just an aliquot.
3. Should we ship the samples by express shipping? What couriers do you accept?
It’s OK to ship the EDTA-blood samples by regular mail, but any other sample should be shipped by express courier (FedEx, UPS, DHL, etc. are all acceptable). Nucleic acids degrade slowly during shipping, which affects the sensitivity of detection. It is recommended to ship the samples using our Sample Submission Kit, which helps preserve the nucleic acids. (see Specimen Submission).
4. Do you accept samples on weekends?
There are no weekend deliveries of mail or express courier at Auburn University. So please make sure to ship for delivery on Friday, or store the sample over the weekend in your refrigerator and ship on Monday.
5. Do we ship the samples frozen?
No. Please ship the samples at room temperature or refrigerated. Freezing and thawing releases enzymes that rapidly destroy nucleic acids. Since frozen polar packs, added as refrigerant to a shipment, will freeze the sample, avoid the polar packs and ship at room temperature.
6. How much do the tests cost?
The tests are $80/test. If you request two different tests on one sample (for example, a Babesia test and an Ehrlichia test for one blood sample), the second test is $70. We also provide the FIP mRNA Multi-test for $150 or $230, in which we process two or three different samples (usually fluid, biopsy, and blood).
7. How do you bill us?
You can write a check to us when you ship the sample. Otherwise we will send a monthly bill. Checks should be payable to: Department of Pathobiology or Molecular Diagnostics. We cannot accept a credit card as payment. There is no need to open an account with us before sending a sample.
8. What is the turnaround time?
The turnaround time is usually one day. For any sample we receive, we will fax or email the report by the end of the next business day.
Call us at (334) 844-2648
9. What is the Sample Submission Kit? How much is it and how do we get it?
Please check our webpage Sample Submission Kit. We provide the kit with 10 individually labeled tubes with buffer in them to help stabilize the nucleic acids. Enclosed are also 5 HistoBrush swabs for collection of swabs. All specimens must be immersed, and liquid specimens mixed with the buffer. The Sample Submission Kits have a shelf life of at least 1 year.
There are three ways to order the Sample Submission Kit:
- Call us at (334) 844-2648
- Fill in our MD Submission Form and fax to 334-844-2652. Please note attention to Molecular Diagnostics Lab
- Email us at email@example.com.
Please specify how many Submission Kits you would like to order, your physical address, recipient name and contact information.
10. What tests do you provide now?
Currently we provide tests for:
- Bartonella henselae
- Canine / feline Anaplasmosis (Anaplasma phagocytophilum / platys)
- Canine Babesiosis (Babesia gibsoni / canis)
- Canine Distemper Virus
- Canine Ehrlichia spp. (E. canis, E. chaffeensis, E. ewingii)
- Canine Hepatozoonosis (Hepatozoon americanum / canis)
- Chlamydia spp.
- Escherichia coli O157:H7
- Feline Infectious Peritonitis mRNA
- Feline / canine hemotrophic Mycoplasmosis (Mycoplasma haemocanis / haemofelis)
- Heartworm (Dirofilaria immitis / repens)
- Leptospira spp. (pathogenic)
- Listeria monocytogenes
- Lyme Borrelia spp.
- Pan Influenza A Virus
- Pseudomonas aeruginosa
- Salmonella spp.
11. How sensitive are the tests?
We can detect single copies of the pathogen DNA or RNA, which translates to approximately 10 copies in each milliliter of the original sample.