Diagnostic Accuracy of Polymerase Chain Reaction for Mycobacterium Tuberculosis Using EBUS-TBNA Samples

July 16, 2015 updated by: Jung Seop Eom, Pusan National University Hospital

Comparison of the Diagnostic Accuracy of Nested and Real-time Polymerase Chain Reaction for Mycobacterium Tuberculosis Using EBUS-TBNA Samples in Patients With Isolated Intrathoracic Lymphadenopathy

The purpose of this study is to compare the diagnostic efficacy of nested and realtime polymerase chain reaction for Mycobacterium tuberculosis using EBUS-TBNA samples in patients with isolated intrathoracic lymphadenopathy.

Study Overview

Status

Unknown

Conditions

Intervention / Treatment

Detailed Description

Although endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) has been widely used to perform mediastinal lymph node sampling, little information is available on polymerase chain reaction for Mycobacterium tuberculosis (TB-PCR) using EBUS-TBNA samples in patients with isolated intrathoracic lymphadenopathy. In addition, two methods of TB-PCR (nested PCR and realtime PCR) are available for the detection of Mycobacterium tuberculosis and which method is superior for detection of Mycobacterium tuberculosis has not been established in patients with isolated intrathoracic lymphadenopathy.

Thus, this study was design to compare the diagnostic efficacy of nested and realtime TB-PCR using EBUS-TBNA samples in patients with isolated intrathoracic lymphadenopathy in South Korea, where the incidence of tuberculosis is intermediate (97/100,000 per year).

Study Type

Interventional

Enrollment (Anticipated)

100

Phase

  • Not Applicable

Contacts and Locations

This section provides the contact details for those conducting the study, and information on where this study is being conducted.

Study Locations

Participation Criteria

Researchers look for people who fit a certain description, called eligibility criteria. Some examples of these criteria are a person's general health condition or prior treatments.

Eligibility Criteria

Ages Eligible for Study

18 years and older (Adult, Older Adult)

Accepts Healthy Volunteers

No

Genders Eligible for Study

All

Description

Inclusion Criteria:

  • Patients with isolated intrathoracic lymphadenopathy

Exclusion Criteria:

  • Suspicion of primary lung cancer on CT or PET scan
  • Suspicion of pulmonary tuberculosis on CT or PET scan
  • Suspicion of sarcoidosis of lung parenchyma on CT or PET scan

Study Plan

This section provides details of the study plan, including how the study is designed and what the study is measuring.

How is the study designed?

Design Details

  • Primary Purpose: Diagnostic
  • Allocation: Non-Randomized
  • Interventional Model: Single Group Assignment
  • Masking: None (Open Label)

Arms and Interventions

Participant Group / Arm
Intervention / Treatment
Experimental: Nested PCR for formalin-fixed tissues
In all patients, nested polymerase chain reaction for Mycobacterium tuberculosis is performed with formalin-fixed paraffin-embedded specimens obtained by endobronchial ultrasound-guided transbronchial needle aspiration.
Device: Nested PCR for formalin-fixed tissues
For nested PCR, formalin-fixed paraffin-embedded tissues were incubated in 1 mL of xylene at 60°C for 30 min and then centrifuged for 10 min. Paraffin and the supernatant were removed from the samples after centrifugation. The same procedures were repeated until deparaffinization was complete. After adding 1 mL of alcohol, the samples were centrifuged for 5 min, and the supernatant was removed. The sample was then air-dried as a pellet. DNA was extracted using QIAamp (Qiagen, Valencia, CA, USA). PCR amplifications were performed using Mycobacterium tuberculosis IS6110 primers. The first round using outer primers and the second round using inner primers amplified a 256- and 181-bp fragment, respectively. Finally, the PCR products were visualized in a 2% agarose gel.
Other Names:
  • MTB-PCR kit; Biosewoom, Seoul, South Korea
Experimental: Nested PCR for fresh tissues
In all patients, nested polymerase chain reaction for Mycobacterium tuberculosis is performed with specimens in sterile saline obtained by endobronchial ultrasound-guided transbronchial needle aspiration.
Device: Nested PCR for fresh tissues
For nested PCR with specimens in sterile saline, DNA was extracted using QIAamp (Qiagen, Valencia, CA, USA). PCR amplifications were performed using Mycobacterium tuberculosis IS6110 primers. The first round using outer primers and the second round using inner primers amplified a 256- and 181-bp fragment, respectively. Finally, the PCR products were visualized in a 2% agarose gel.
Other Names:
  • MTB-PCR kit; Biosewoom, Seoul, South Korea
Experimental: Real-time PCR for fresh tissues
In all patients, real-time polymerase chain reaction for Mycobacterium tuberculosis is performed with specimens in sterile saline obtained by endobronchial ultrasound-guided transbronchial needle aspiration.
Device: Real-time PCR for fresh tissues
For real-time PCR, specimens in sterile saline were filtered and 1 mL of phosphate based saline was added. After centrifugation for 3 min, the supernatant was removed. Next, 50 μL of extraction buffer was added to the sample, and the sample was heated at 100°C for 20 min. After centrifugation for 3 min, the supernatant was used in PCR. Real-time PCR was performed using the AdvanSure TB/NTM real-time PCR kit. Three channels were used in the real-time PCR reaction (Mycobacterium tuberculosis complex, mycobacteria, and internal control). Signals for FAM, HEX, and Cy5 were measured in each channel. Mycobacterium tuberculosis was considered present if the cycle threshold of rpoB was less than 35 on each signal and greater than or equivalent to that of IS6110.
Other Names:
  • AdvanSure TB/NTM real-time PCR kit; LG, Seoul, South Korea

What is the study measuring?

Primary Outcome Measures

Outcome Measure
Measure Description
Time Frame
Number of participants who detected each polymerase chain reactions for Mycobacterium tuberculosis
Time Frame: 6 months
In all participants with isolated intrathoracic lymphadenopathy, three modalities of TB-PCR (nested PCR for formalin-fixed paraffin-embedded tissues, nested PCR for fresh tissues and real-time PCR for fresh tissues) are performed using EBUS-TBNA samples. The numbers of participants who reported as positive for each TB-PCR modalities are collected. From these data, sensitivity, specificity and diagnostic accuracy of each TB-PCR modalities to diagnose tuberculous lymphadenitis will be calculated.
6 months

Collaborators and Investigators

This is where you will find people and organizations involved with this study.

Sponsor

Pusan National University Hospital

Investigators

  • Principal Investigator: Jeong Ha Mok, Master, Pusan National University Hospital
  • Principal Investigator: Jung Seop Eom, Master, Pusan National University Hospital

Publications and helpful links

The person responsible for entering information about the study voluntarily provides these publications. These may be about anything related to the study.

General Publications

Study record dates

These dates track the progress of study record and summary results submissions to ClinicalTrials.gov. Study records and reported results are reviewed by the National Library of Medicine (NLM) to make sure they meet specific quality control standards before being posted on the public website.

Study Major Dates

Study Start

July 1, 2015

Primary Completion (Anticipated)

December 1, 2018

Study Completion (Anticipated)

December 1, 2018

Study Registration Dates

First Submitted

July 8, 2015

First Submitted That Met QC Criteria

July 13, 2015

First Posted (Estimate)

July 14, 2015

Study Record Updates

Last Update Posted (Estimate)

July 20, 2015

Last Update Submitted That Met QC Criteria

July 16, 2015

Last Verified

July 1, 2015

More Information

Terms related to this study

Keywords

Additional Relevant MeSH Terms

Other Study ID Numbers

  • PNUH-P-1

Drug and device information, study documents

Studies a U.S. FDA-regulated drug product

No

Studies a U.S. FDA-regulated device product

No

product manufactured in and exported from the U.S.

No

This information was retrieved directly from the website clinicaltrials.gov without any changes. If you have any requests to change, remove or update your study details, please contact register@clinicaltrials.gov. As soon as a change is implemented on clinicaltrials.gov, this will be updated automatically on our website as well.

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