IDEXX RealPCR™ Tests

Cats and dogs frequently present with diarrhea, but identifying causative agents often requires numerous tests with varying reliability. The IDEXX Canine and Feline Diarrhea RealPCR™ Panels allow you to screen for and accurately identify multiple infectious causes of diarrhea from only a single fecal sample.

Knowing precisely which intestinal pathogen is affecting your patient gives you the information you need to hone treatment recommendations and improve efficacy. And because it’s RealPCR technology, you can find out more quickly and with greater accuracy exactly what’s contributing to your patients’ diarrhea.

When to use IDEXX Canine and Feline Diarrhea RealPCR™ Panels:

• To identify the pathogen(s) that may be causing or contributing to diarrhea.
• For timely diagnosis and determining appropriate therapies/treatments.
• As a surveillance tool for dog or cat populations in shelters, breeding facilities, etc.*
• To identify and minimize human exposure to zoonotic pathogens.

Canine and Feline Diarrhea Panels and Tests

Specimen requirements:
5 g fresh feces (1 g minimum) in a sterile container; keep refrigerated; collect sample prior to antibiotic administration.

When ordering a panel that includes the Salmonella spp. RealPCR™ Test, also submit 3–5 g fresh feces in fecal culture transport media for reflex Salmonella spp. culture if indicated.

Indications include:

• Presence of intracranial disease manifestations
  • Depression, change in behavior
  • Seizures
  • Ataxia, circling, vestibular disease
  • Cortical blindness
  • Paresis, paralysis
• Finding abnormalities on cerebral spinal fluid (CSF) analysis, including elevated cell count and/or elevated protein
• Presence of unexplained neck pain

Specimen requirements:
0.5 mL (0.1 mL minimum) CSF (preferred) and/or 2 mL EDTA whole blood (LTT); keep refrigerated; collect sample prior to antibiotic administration.

When ordering a panel that includes a Lyme Quant C6® Test, FeLV antigen or FIV antibody by ELISA, also submit 1 mL serum.

Canine Respiratory Disease

Multiple infectious agents can be a cause of respiratory disease in dogs. Prompt identification and treatment of these agents allows you to better manage these cases, may lead to less severe clinical signs and allows for appropriate isolation of infected dogs.

The IDEXX Canine Respiratory Disease (CRD) RealPCR™ Panel provides rapid, sensitive and specific identification for eight infectious agents—all at once.

From 4,700 samples submitted and tested with the CRD RealPCR Panel during a 12-month period, results were:¹

• Positive for any one of the eight infectious agents 53.1% of the time
• Positive for coinfections with two or more infectious agents 43.5% of the time

Prevalence of each component in the CRD RealPCR Panel

Feline Upper Respiratory Disease

Cats presenting with signs of feline upper respiratory disease (URD)—sneezing, oculonasal discharge, conjunctivitis with or without oral ulceration—are probably common in your practice. But until now, there have been no quick, accurate and affordable tests that could determine the causative agents.

Running the quick and easy IDEXX Feline Upper Respiratory Disease (URD) RealPCR™ Panel dramatically increases your chance of accurately identifying the origins of feline URD.

From 7,500 samples submitted and tested with the Feline URD RealPCR Panel during a 12-month period, results were:¹

• Positive for any one of the six infectious agents 71.0% of the time
• Positive for coinfections with two or more infectious agents 42.4% of the time

Prevalence of each component in the Feline URD RealPCR Panel

The results from running this single-panel, real-time PCR test enable you to quickly take the next steps:

• Choose the appropriate treatment
• Quarantine animals as needed
• Improve patient outcome

Canine and Feline Respiratory Panels, Tests and Profiles

Specimen requirements:

Deep pharyngeal swab (with visible organic material on swab; please rub firmly) and a conjunctival swab (wipe eye clean; swab inside of eyelid) in the same tube; collect sample prior to antibiotic administration. Please submit dry, plastic-stemmed swabs, without transport media, in a RTT or an empty, sterile tube; keep refrigerated. When acquiring a pharyngeal swab, precautions should be taken to prevent patient from biting and/or ingesting part of swab.

For feline patients:

• When ordering a feline URD profile that includes a CBC, chemistry or T4, also submit 2 mL serum, 1 mL EDTA whole blood (LTT) and two blood smears.
• When ordering a feline URD profile that includes a urinalysis, also submit 5 mL urine in a sterile container.

Systemic mycoses are associated with high morbidity and mortality especially if undiagnosed and untreated. Because of the length of time, difficulty and zoonotic potential associated with culture, diagnosis has until now relied on serology and/or microscopic identification of the fungus on tissue biopsies or cytologic samples. The Systemic Mycoses RealPCR™ Panel provides a sensitive diagnostic tool to detect these infectious fungal pathogens.

Consider testing dogs or cats with:

• Nonspecific clinical signs, such as fever, lethargy, anorexia, weight loss, lymphadenopathy
• Respiratory signs, including dyspnea, coughing, nasal mass or discharge, sneezing, epistaxis
• Ocular signs, including uveitis, chorioretinitis, keratitis, panophthalmitis, acute blindness, exophthalmos
• Neurological signs, including depression, seizures, ataxia, behavioral changes
• GI signs, including large and/or small bowel diarrhea with protein-losing enteropathy
• Musculoskeletal signs, including lameness, bone pain

Canine and Feline Systemic Mycoses Panel and Tests

Specimen requirements:
Ideal sample depends on clinical manifestation (see below). Keep all samples refrigerated.

• For systemic, nonspecific manifestations: 2 mL EDTA whole blood (LTT).
• For respiratory manifestations:
  • Deep pharyngeal swab (with visible organic material on swab; please rub firmly) and/or nasal swab. Please submit dry, plastic-stemmed swabs, without transport media, in a RTT or in an empty, sterile tube. Multiple swabs can be submitted together in the same tube. When acquiring a pharyngeal swab, precautions should be taken to prevent patient from biting and/or ingesting part of swab.
  • Tracheal wash or bronchoalveolar lavage fluid: 0.5 mL (0.1 mL minimum).

• Aspirate* or biopsy* of nasal mass.
• For ocular manifestations: 0.5 mL (0.1 mL minimum) uveal fluid, ocular tissue aspirate* or biopsy.*
• For neurological manifestations: 0.5 mL (0.1 mL minimum) CSF.
• For GI manifestations: 5 g (1 g minimum) fresh feces in a sterile container.
• For skin lesions: Tissue aspirate*, 0.5 mL (0.1 mL minimum) fluid, if available, or biopsy.*
• For bone lesions: Bone aspirate* or biopsy.*

IDEXX Reference Laboratories has introduced several new panels for vector-borne disease testing that combine both serology and real-time polymerase chain reaction (PCR). These next-generation IDEXX Vector-borne Disease Panels take advantage of the benefits of both methodologies, making it easy and cost-effective for veterinarians to make complete and accurate diagnosis.

Combining serology and PCR increases diagnostic sensitivity and reduces the chance for false negatives. The IDEXX RealPCR™ component of the vector-borne disease panels detects pathogenic nucleic acid (DNA), identifying infectious agents of clinical disease as well as coinfections, and the serology portion of the panel may reveal exposure to multiple pathogens and provides an indication of the duration of infection. For Lyme disease, the IDEXX C₆ technology is the preferred diagnostic method of Borrelia burgdorferi infection.

When to use Vector-borne Disease Panels:

• A sick patient with clinical signs and/or laboratory abnormalities consistent with a vector-borne disease

  Examples:

  • An animal presenting with clinical signs of lethargy, anorexia, lameness and fever
  • A sick patient with clinical laboratory findings that include anemia, thrombocytopenia, leukocyte abnormalities and/or hyperglobulinemia

• Patients with suspected subclinical infections as based upon history, physical examination, serological results and clinical laboratory findings.

  Example: Clinically healthy animals with a history of tick exposure, physical examination findings of organomegaly or lymphadenomegaly, and laboratory abnormalities such as thrombocytopenia or anemia

• Patients previously diagnosed with a vector-borne disease and are in need of monitoring therapy

  • The Lyme Quant C₆® Test is run as part of several IDEXX Vector-borne Disease Panels and can indicate waning antibody levels
  • For other pathogens, a negative PCR result indicates a reduction in pathogen load

Note: Positive serologic results alone in a clinically healthy animal with normal laboratory results is NOT an indication for performing an IDEXX Vector-borne Disease Panel.

Canine and Feline Vector-borne Disease Panels and Tests

Specimen requirements:
2 mL EDTA whole blood (LTT); keep refrigerated; collect sample prior to antibiotic administration.

When ordering a panel that includes a Lab 4Dx Test, Lyme Quant C6 Test or FeLV antigen and FIV antibody by ELISA, also submit 1 mL serum.

Anemia Panel

Specimen requirements:
1 mL serum, 2 mL EDTA whole blood (LTT); keep refrigerated; collect sample prior to antibiotic administration.

Blood Donor Panel

Specimen requirements:
1 mL serum, 2 mL EDTA whole blood (LTT); keep refrigerated; collect sample prior to antibiotic administration.

Clinical signs of canine distemper virus (CDV)—vomiting, diarrhea, respiratory and even acute and progressive neurologic signs—may be difficult to distinguish from other infectious and noninfectious diseases in young dogs with unknown vaccine histories.

The IDEXX Canine Distemper Virus (CDV) RealPCR™ Test can definitively diagnosis distemper infection to:

• Better guide appropriate therapy
• Support isolation of puppy/dog to prevent spread of infection
• Prepare the pet owner for possible development of neurologic consequences
• Help owners and clinicians make informed decisions about therapy versus euthanasia

Canine Distemper Virus (CDV) Test

Note:

For patients with respiratory manifestations who have been vaccinated with a modified-live vaccine in the last four weeks, see the Canine Distemper Virus (CDV) Quant RealPCR™ Test (test code 3265).

Specimen requirements:

Ideal sample depends on clinical manifestation (see below). Keep all samples refrigerated.

• For neurological manifestations: 0.5 mL (0.1 minimum) CSF and/or 2 mL EDTA whole blood (LTT).
• For respiratory manifestations: Deep pharyngeal swab (with visible organic material on swab; please rub firmly) and a conjunctival swab (wipe eye clean, swab inside of eyelid), in the same tube. Please submit dry, plastic-stemmed swabs, without transport media, in a RTT or in an empty, sterile tube.
• For GI manifestations: 2 mL EDTA whole blood (LTT) and 5 g fresh feces (1 g minimum).
• With no distinct clinical manifestations: 2 mL EDTA whole blood (LTT) and conjunctival swab (wipe eye clean, swab inside of eyelid). Please submit dry, plastic-stemmed swabs, without transport media, in a RTT or in an empty, sterile tube.

Many dogs develop clinical signs of distemper within days of adoption from a kennel or shelter, where they have often received a recent vaccination. Because most distemper vaccines consist of attenuated strains, highly sensitive real-time PCR test systems will pick up the replicating vaccine strain in about 20% of dogs over the course of a few weeks after each immunization.1 Until now, interpreting a positive test result in recently vaccinated dogs was challenging.

The CDV Quant RealPCR™ Test provides a quantitative measure of the CDV viral load. The amount of virus during infection is typically exponentially more than would be detected because of recent vaccination. Therefore, this quantitative information allows the test to discriminate vaccine interference from infection with a wild-type strain of distemper virus.

Canine Distemper Virus (CDV) Test

Note:

Use for dogs with respiratory signs consistent with canine distemper virus that have been vaccinated with a modified live vaccine within the previous 4 weeks. For dogs with clinical manifestations requiring other nonrespiratory types or in dogs for which vaccine interference is not a concern, please see Distemper Virus (CDV) RealPCR™ Test—Canine (Test Code 2265).

Specimen requirements:

Deep pharyngeal swab (with visible organic material on swab; please rub firmly) and a conjunctival swab (wipe eye clean, swab inside of eyelid) in the same tube. Please submit dry plastic-stemmed swabs, without transport media, in RTT or empty, sterile tube (keep refrigerated).

The IDEXX Leptospira spp. RealPCR™ Test enables an early and accurate diagnosis of canine leptospirosis. A potentially life-threatening disease, leptospirosis most commonly manifests in dogs as severe acute kidney disease. In addition, urinary shedding of leptospires by infected animals poses a significant zoonotic risk to dog owners.

The IDEXX Leptospira spp. RealPCR Test offers advantages over serological testing:

• Early detection provides the ability to diagnosis acute infection prior to antibody development.
• Detects urinary shedding in sick and healthy animals to aid in diagnosis and minimize zoonotic risk.
• No cross-reactivity and no interference of results by currently available bacterin-based vaccines, as determined by a recent study performed at Cornell University College of Veterinary Medicine (Richard E. Goldstein, DVM, DACVIM, DECVIM-CA, oral communication, June 2009).

Leptospirosis should be considered in every dog presenting with acute renal disease, especially as early diagnosis and treatment improves the prognosis of infected animals. Use the test code below when ordering the IDEXX Leptospira spp. RealPCR Test.

Canine Leptospira spp. Test

Specimen requirements:
2 mL EDTA whole blood (LTT) and 2 mL urine in a sterile container; keep refrigerated; collect sample prior to antibiotic administration.

Anemia Panel with FeLV/FIV by ELISA

Specimen requirements:
1 mL serum, 2 mL EDTA whole blood (LTT); keep refrigerated; collect sample prior to antibiotic administration.

Blood Donor Panel

Specimen requirements:
1 mL serum, 2 mL EDTA whole blood (LTT); keep refrigerated; collect sample prior to antibiotic administration.

Feline Coronavirus (FCoV) Test

Specimen requirements:
Ideal sample depends on clinical manifestation. 2 mL (0.1 mL mimimum) pleural or abdominal effusion (preferred specimen), CSF, EDTA whole blood (LTT), or 5 g fresh feces (1 g minimum) in a sterile container; keep refrigerated.

Feline hemotropic mycoplasma (FHM) is a common cause of severe hemolytic anemia in cats.

In a recent nationwide study:¹

• 12.7% of healthy blood donor pet cats were infected with feline hemotropic mycoplasma species
• A higher rate of FHM infection was found in outdoor cats and cats exposed to fleas

Until now, diagnosing feline hemotropic mycoplasmosis (formerly hemobartonellosis or feline infectious anemia) has been a challenge. The pathogenicity of FHM species differs:

• Most pathogenic: Mycoplasma haemofelis (formerly Haemobartonella felis)
• Moderately pathogenic: Candidatus Mycoplasma turicensis
• Least pathogenic: Candidatus Mycoplasma haemominutum

Because of these differences in pathogenicity, the Feline Hemotropic Mycoplasma (FHM) RealPCR™ tests and profiles help you determine the significance of an FHM infection, which may affect case management, including the pursuit of additional diagnostics and treatment.

Visual slide review cannot distinguish FHM species, but real-time PCR can. Compare these two methods below to see how real-time PCR detects infections that a manual slide review can miss:²

Feline Hemotropic Mycoplasma (FHM) Tests and Profiles

Specimen requirements:

2 mL EDTA whole blood (LTT); keep refrigerated; collect sample prior to antibiotic administration.

• When ordering an FHM profile that includes a CBC, chemistry or T₄, also submit 2 mL serum, 1 mL EDTA whole blood (LTT) and 2 blood smears.
• When ordering an FHM profile that includes a urinalysis, also submit 5 mL urine in a sterile container.

The IDEXX Feline Immunodeficiency Virus (FIV) RealPCR™ Test is the first highly specific diagnostic that detects the presence of FIV nucleic acid to confirm active infection. However, a negative FIV RealPCR Test result does not rule out infection. Rather, it can indicate one of three situations: the cat may not be infected; it may be infected, but nucleic acid is below limit of detection; or the cat may be infected with an uncommon strain not currently detected by PCR.

Additional Considerations for the FIV RealPCR Test

• FIV serology testing, as with the SNAP® FIV/FeLV Combo Test and the SNAP® Feline Triple® Test, remains the first-line diagnostic because of its high sensitivity and specificity.
• FIV RealPCR Test should only be considered in serologically positive cats when vaccination history is unknown or when cats have been vaccinated for FIV but infection is still suspected.
• Although the FIV RealPCR Test is highly specific for infection, it cannot “rule out” infection or determine FIV vaccination status.
• Given the limitations of interpretation, the FIV RealPCR Test should not be used to alter decisions regarding FIV vaccination protocol.
• FIV PCR is not recommended as a screening for FIV.

Feline Immunodeficiency Virus (FIV) Test

Specimen requirements:

2 mL EDTA whole blood (LTT); keep refrigerated.

Polymerase chain reaction (PCR) is a simple yet elegant process that enables the production of virtually unlimited copies of genetic material in the laboratory.

Real-time PCR is a safer, faster and more robust variation of PCR. Conventional PCR harbors more risk for cross-contamination, is more time consuming and does not allow a quantitative assessment of the quality of the DNA extracted from the sample material.

For most of the inactivated vaccines, real-time PCR does differentiate between vaccination and infection. Real-time PCR does not detect the presence of antibodies; therefore, a positive result indicates presence of the pathogen. For this reason, PCR has a higher association with disease than with antibody detection. Detection of viral DNA/RNA may occur within a few weeks after vaccination with a modified live vaccine (MLV) while the vaccine virus is replicating. We recommend a three-week interval after the last vaccination to prevent detection of any DNA originating from the vaccine.

A positive result confirms the presence of the pathogen. 

A negative result means the pathogen is either not present, below the limit of detection, not detectable by the current PCR assay or that the sample quality was low.

Detection limit of a method is defined in various ways. The most accepted definition is the target nucleic acid concentration or number of molecules that can still be amplified in 95% of the experiments.

The diagnostic sensitivity describes the probability of a true positive result when the PCR is tested in a positive population. In other words, higher sensitivity means higher probability that you’ll correctly identify all positive samples and won’t miss a diagnosis. The diagnostic specificity is described as the probability of a true negative result when the PCR is tested in a negative population. Higher specificity is the probability that the test is really detecting what you’re testing for and not something unrelated. Therefore, higher specificity increases the probability that a positive result is due to the agent.

This depends on the presence or absence of clinical signs, the pathogen(s) suspected and assay being used. In general, a two- to four-week waiting period is recommended.

Yes. PCR is a complementary diagnostic procedure and should be run parallel with other diagnostic assays. Depending on the pathogen, this could include complete blood analysis, serology, antigen detection systems (such as SNAP® tests), etc.

Samples can be stored in a refrigerator (4°C) for up to three days (over the weekend).

Fluids (peritoneal/pleural fluid, whole blood, feces) and swabs can be stored for three days (over the weekend) in a refrigerator (4°C).

Fresh tissues should be sent on ice within 24 hours. Tissues can be stored in a standard freezer at -20°C for up to one week. Paraffin-embedded tissues are acceptable.

If you use a courier pickup, samples can be submitted with your other laboratory samples. If you use FedEx, all samples should be shipped according to applicable government and IATA regulations. Send samples in an insulated package with ice packs. Samples should be placed in a secondary container within the package, in case of damage during shipping. Absorbent material should be included in the secondary container in case of leaks or breaks in the primary container.

The sample type is specific for each pathogen. Please refer to the specimen requirements for each test on the RealPCR Tests tab.

Samples will be kept up to three days after receipt for reanalysis. The extracted DNA is stored for one year and can be used for reanalysis or further testing.

Ideally, separate samples should be submitted for PCR.

Check on the status of your sample online at www.vetconnect.com or call IDEXX at 1-888-433-9987. PCR testing is run daily at our Sacramento facility and you can expect results in 1–3 days.

For more information about IDEXX RealPCR™ panels and stand-alone tests, call our Internal medicine specialists at IDEXX Reference Laboratories at 1-888-433-9987, option 4, option 2. IDEXX’s board-certified internal medicine specialists are available to help answer your questions, such as:

• When to use a PCR assay
• How to interpret results
• Treatment recommendations

The 2009 Directory of PCR Services (PDF) lists all the RealPCR tests available from IDEXX.

The original samples are stored for three days. However, the extracted DNA from the sample is stored for one year. Additional tests can be requested on stored DNA when appropriate. For example, if originally only a distemper PCR was ordered, a full canine respiratory panel can be performed on the stored DNA if the original sample was a conjunctival swab and pharyngeal swab and not whole blood.

PCR tests for all pathogens are run daily Monday through Friday. Turnaround time for a sample is 24 hours after receipt of the sample at the West Sacramento facility. Samples delivered on Saturday or Sunday are processed on Monday. Including shipping time, you can expect results in 1–3 working days.