Toxoplasma gondii Quantitative PCR

Toxoplasma gondiiToxoplasma gondii is highly problematic when primary infection occurs during the first trimester of pregnancy, often leading to fetal death.  When Toxoplasma gondii reactivation occurs in immunocompromised patients it can lead to life-threatening disease, including encephalitis and extracerebral toxoplasmosis.  Onset of disease in most patients is preceded by an increase in parasite load measured in peripheral blood specimens using quantitative PCR.  ViraCor’s quantitative PCR assay is used to detect and monitor blood levels of Toxoplasma gondii DNA, as this correlates well with treatment.

See below for additional Toxoplasma gondii assay and pathogen-specific information.  For online ordering methods click here or contact us .

Assay Sheet

Test ID

2200 Toxoplasma gondii Real-time qPCR

CPT Code

87799

Clinical Utility

Toxoplasma gondii is highly problematic when primary infection occurs during the first trimester of pregnancy, often leading to fetal death or when reactivation occurs in immunocompromised patients, leading to life-threatening disease including encephalitis and extracerebral toxoplasmosis. Onset of disease in most patients is preceded by an increase in parasite load measured in peripheral blood specimens using quantitative PCR. Monitoring blood levels of Toxoplasma gondii DNA correlates well with treatment.

Procedure

Extraction of Toxoplasma gondii DNA from blood, CSF, other biological fluids, or tissues followed by amplification and detection using real-time, quantitative PCR. An internal control is added to ensure the extraction was performed correctly and the PCR reaction was not inhibited.

Specimens

Whole Blood: 3-5 ml submitted in an EDTA tube; ship ambient.
CSF: 1 ml fluid frozen; submitted in a sterile, leakproof tube; ship on dry ice.
Amniotic Fluid: 1-4 ml submitted in a sterile, leakproof tube; ship ambient.
BAL: 1-4 ml submitted in a sterile, leakproof tube; ship ambient.
Other: Please inquire.

Specificity

The primers and probes used in this assay are specific for known Toxoplasma gondii strains based on similarity search algorithms. Additionally, no cross reactivity was detected with any viral or protozoa pathogens.

Toxoplasma gondii Assay Range

500 copies/ml to 1 x 1010 copies/ml

Turnaround Time

Within 24 hours of receiving specimen

pdf DOWNLOAD ASSAY SHEET

PCR tests are performed pursuant to a license agreement with Roche Molecular Systems, Inc.
This assay was developed and the performance characteristics were determined at ViraCor Laboratories. This test is performed in a CLIA certified laboratory. FDA approval is not required for the performance of this test.
AS17-0108

 

Pathogen Overview

ABOUT THE TOXOPLASMA GONDII ORGANISM

Toxoplasma gondii is an obligate intracellular protozoan ubiquitous in birds and mammals. Toxoplasmosis is the disease that occurs when T. gondii invades and multiplies asexually as tachyzoites within the cytoplasm of nucleated cells. When host immunity develops, multiplication of tachyzoites ceases and tissue cysts form, which remain latent, especially in the brain and muscle. Sexual reproduction of T. gondii occurs only in the intestinal tract of cats; the resultant oocysts passed in the feces remain infectious up to a year in soil, depending upon the temperature and moisture content.

The two major routes of transmission in humans are oral and congenital. Humans become infected with T. gondii through direct contact with oocysts in cat feces or through eating meat contaminated with the extraintestinal form of T. gondii. Transmission of the infection from mother to fetus occurs almost solely in women who acquire the infection during pregnancy.

Nearly one quarter of adults and adolescents in the United States have been infected with T. gondii; the incidence of seropositivity increases with increasing age. The diagnosis of toxoplasmosis is most critical in four groups of patients: pregnant women who acquire infection during gestation, fetuses and newborns who are congenitally infected, immunocompromised patients and patients with chorioretinitis.

TOXOPLASMA GONDII CLINICAL MANIFESTATIONS

Infections in healthy individuals are typically asymptomatic or associated with self-limited symptoms, such as fever, malaise, and lymphadenopathy, which usually resolves within weeks to months and does not require treatment. Infections in pregnant women, also asymptomatic, can be transplacentally transmitted to the fetus. If the fetus is infected in the first trimester, the result ranges from severe disease to spontaneous abortion or stillbirth; if infection occurs afterwards, disease manifestations can include encephalitis, mental retardation, blindness, and epilepsy.

Immunocompromised patients are at risk for primary infection or reactivation of latent infection. Transmission of T. gondii by organ transplantation from a seropositive donor to a seronegative recipient is an important potential cause of primary infection in solid organ transplant patients. Hematopoietic stem cell transplant patients and AIDS patients are at most risk for reactivation of a latent infection. Primary toxoplasmosis has been documented between day 1 and 13 months post-transplant; reactivations have been documented up to 7 years post-transplant. Interestingly, toxoplasmosis is sometimes seen only after prophylaxis for pneumocystis pneumonia (PCP) ceases, since the two diseases share therapeutic regimens. Common reactivated infections in transplant patients include disseminated toxoplasmosis, cerebral toxoplasmosis, pulmonary toxoplasmosis, and occasionally, combined ocular and cerebral toxoplasmosis.

Diagnosis is especially difficult in immunocompromised patients, as clinical presentation varies and is often nonspecific. The most common symptoms are fever and altered mental state; pneumonitis and myocarditis are less common presentations. Patients with disseminated toxoplasmosis present with fever that is unresponsive to antibiotics, weakness and fatigue. Chorioretinitis or multiorgan involvement presenting with acute respiratory failure and haemodynamic abnormalities similar to septic shock may also occur. Cerebral toxoplasmosis, the most common form of infection, presents as mental confusion, seizures and typical deep-seated lesions of the brain. Disseminated cerebral infections may present with a radiological image of small miliary lesions. Pulmonary toxoplasmosis presents as respiratory distress and lesions in the lungs.

TOXOPLASMA GONDII LABORATORY DIAGNOSIS

T. gondii cannot be cultured in the clinical diagnostic setting, so serological testing has been the mainstay of toxoplasmosis diagnosis, though the method has many significant limitations. For example, IgG antibody detection can be used to identify immunocompromised patients at risk of reactivation, but not for diagnosis of current toxoplasmosis, since any exposure to T. gondii at any point in time will result in positive antibodies. Discrimination between recent and more distant infections is not possible using serological methods. In immunocompromised patients, absence of specific antibodies does not rule out active disease since these patients may not be able to mount an appropriate humoral immune response. Risk for congenital toxoplasmosis may go undetected if the pregnant mother was tested during the active phase of infection, when IgG or IgM antibodies may not be detectable. An additional diagnostic method is microscopic examination of tissue, which can be very challenging to obtain. In cases of disseminated toxoplasmosis, the infected tissue could be in almost any organ.

Quantitative real-time PCR overcomes the limitations of serological testing and is a recent and very promising option. It has been shown to be a highly sensitive, specific and rapid method to detect T. gondii DNA from a wide variety of specimen sources. In addition, monitoring levels of T. gondii DNA in patients correlates well with treatment, enabling physicians to track response to therapy over time and assess outcomes.

TOXOPLASMA GONDII TREATMENT

Transplant recipients most likely to acquire infection via the allograft should be tested for baseline toxoplasma IgG antibodies, along with the donor. A seropositive donor and seronegative recipient are at the highest risk; therefore, the transplant recipient should receive prophylaxis.

A combination of pyrimethamine and sulfonamides is the best evaluated therapy and is recommended as first line therapy. However, not all patients can tolerate this combination, therefore, pyrimethamine and clindamycin in combination is recommended as second line therapy. After treatment of the acute phase infection, secondary prophylaxis should be administered as maintenance therapy, which usually consists as the same regimen at half doses until underlying immunosuppression has ceased.
Management of maternal and fetal infection varies widely. Spiramycin is often administered after diagnosis of a recently acquired maternal infection.

Selected References

Jones J, Kruszon-Moran D, Wilson M. Toxoplasma gondii infection in the United States, 1999–2000. Emerg Infect Dis. 2003;9(11):1371-1374.

Jones J, Lopez A, Wilson M, et al. Congenital toxoplasmosis: a review. Obstet Gynecol Surv. 2001;56(5):296-305.

Lin M, Chen T, Kuo T, et al. Real-time PCR for quantitative detection of Toxoplasma gondii. J Clin Mirobiol. 2000;38(11):4121-4125.

Martino R, Bretagne S, Einsele H, et al. Early detection of toxoplasma infection by molecular monitoring of Toxoplasma gondii in peripheral blood samples after allogeneic stem cell transplantation. Clin Infect Dis. 2005;(40):67-78.

Montoya J, Liesenfeld O. Toxoplasmosis. Lancet. 2004;(363):1965-1976.

Remington J, Thulliez P, Montoya J. Recent developments for diagnosis of toxoplasmosis. J Clin Microbiol. 2004;2(3):941-945.

Wulf M, Crevel R, Portier R, et al. Toxoplasmosis after renal transplantation: implications of a missed diagnosis. J Clin Microbiol. 2005;43(7):3544-3547.

PAO-16-08007 PCR tests are performed pursuant to a license agreement with Roche Molecular Systems, Inc.

pdf DOWNLOAD PATHOGEN OVERVIEW

Collection & Shipping

 

 Specimen Source
 Collection Procedure
 Transport Procedure
 Blood
Plasma
2-3 ml separated from whole blood collected in EDTA (lavender top) tube.
 Ship at ambient temperature Monday-Friday
Whole Blood
3-5 ml collected in EDTA (lavender top) tube. Do not freeze.
 Ship at ambient temperature Monday-Friday
 ImmuKnow® Specimens- Whole Blood
2-3 ml collected in a sodium heparin (green top) tube. Maintain temperature by shipping the specimen in 2 inch thick styrofoam with specimen surrounded by ambient temperature gel packs.
 Ship ambient for priority overnight delivery  Monday‐Friday
Specimen must arrive at ViraCor within 30 hours of collection.
 Hepatitis Specimens- Whole Blood
7-10 ml in EDTA, ACD Solution A, or PPT sterile tube. Minimum specimen requirement is 2 ml plasma. Separate plasma from cells within 4 hours of collection and freeze. To remove plasma from cells, centrifuge at 1000 xg for 10-15 minutes. Do not clarify by filtration or further centrifugation. If specimen was collected in PPT tube, the entire tube can be frozen if desired following centrifugation.
 Ship ambient or frozen
Monday-Friday
 Body fluid other than blood or urine
Collect 2-3 ml in a sterile screw-cap tube.
 Ship at ambient temperature Monday-Friday
 Bone Marrow
1-2 ml, collected in an EDTA (lavender top) tube. Do not freeze.
 Ship at ambient temperature Monday-Friday
 Bronchial Lavage/Bronchial Wash
2-3 ml, collected in sterile screw-cap tube.
 Ship at ambient temperature Monday-Friday
 CSF
1-1.5 ml in sterile screw-cap tube. Freeze prior to shipment.
 Ship on DRY ICE
Monday-Friday
 Eye swab
Swab the inflamed conjunctiva or corneal lesions. Place swab in 1-2 ml sterile saline or viral transport media in sterile screw-cap tube.
 Ship at ambient temperature Monday-Friday
 Fecal
Sterile swab (plastic shaft only) or very small (pea size) fecal sample placed in 1-2 ml sterile saline or viral transport in sterile screw-cap tube.
 Ship at ambient temperature Monday-Friday
 Nasopharyngeal Aspirate/Tracheal Aspirate
2-3 ml collected in sterile saline in sterile screw-cap tube.
 Ship at ambient temperature Monday-Friday
 Nasopharyngeal Swab
Sterile swab (flexible shaft) placed in 1-2 sterile saline or viral transport media in sterile screw-cap tube. Do not use calcium alginate swab.
 Ship at ambient temerpature Monday-Friday
 Swab
Sterile swab (plastic shaft only) placed in 1-2 ml sterile saline or viral transport media in sterile screw-cap tube. Do not use calcium alginate swab. 
Ship at ambient temperature Monday-Friday
 Tissue
Place in a sterile screw-top container, add a small amount of saline to keep moist. 
Ship at ambient temperature Monday-Friday Frozen tissue is acceptable 
 Urine
5 ml sample collected in a sterile urinalysis container. Transfer to a 15 ml sterile screw-cap tube for shipment. 
 Ship at ambient temperature Monday-Friday
 Vesicular Lesion
Collect fluid and cellular material from the base of several fresh vesicles. Place swab in 1-2 mil sterile saline or viral transport media in sterile screw-cap tube. Do not use calcium alginate swab.
Ship at ambient temperature Monday-Friday  
 Other Specimen
Please inquire.

Shipping

  • All specimens must be labeled with patient's name and collection date.
  • A ViraCor Test Request Form must accompany each specimen.
  • Ship specimens FedEx Priority Overnight to: ViraCor Laboratories | 1001 NW Technology Dr | Lee's Summit MO 64086

PCR tests are performed pursuant to a license agreement with Roche Molecular Systems Inc.
ImmuKnow is a registered trademark of Cylex Incorporated.
Respiratory Viral Panel is a product of Luminex Corporation.

Abstracts & Publications

Jones J, Kruszon-Moran D, Wilson M. Toxoplasma gondii infection in the United States, 1999–2000. Emerg Infect Dis. 2003;9(11):1371-1374.

Jones J, Lopez A, Wilson M, et al. Congenital toxoplasmosis: a review. Obstet Gynecol Surv. 2001;56(5):296-305.

Lin M, Chen T, Kuo T, et al. Real-time PCR for quantitative detection of Toxoplasma gondii. J Clin Mirobiol. 2000;38(11):4121-4125.

Martino R, Bretagne S, Einsele H, et al. Early detection of toxoplasma infection by molecular monitoring of Toxoplasma gondii in peripheral blood samples after allogeneic stem cell transplantation. Clin Infect Dis. 2005;(40):67-78.

Montoya J, Liesenfeld O. Toxoplasmosis. Lancet. 2004;(363):1965-1976.

Remington J, Thulliez P, Montoya J. Recent developments for diagnosis of toxoplasmosis. J Clin Microbiol. 2004;2(3):941-945.

Wulf M, Crevel R, Portier R, et al. Toxoplasmosis after renal transplantation: implications of a missed diagnosis. J Clin Microbiol. 2005;43(7):3544-3547.