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33356 - NeXGen™ Fungal / AFB NGS Assay

Test Code: 33356

NeXGen Fungal / AFB Next Generation Sequencing NGS Assay

Next Generation Sequencing (NGS) testing has gained significant importance in the medical clinical laboratory in the United States over the past decade. In recent years, the field of molecular diagnostics has witnessed significant advancements, particularly in the detection of infectious diseases. One such breakthrough is the application of hybrid-capture next-generation sequencing (NGS) technology. NGS technology allows for the rapid and cost-effective analysis of large amounts of genetic information, enabling various applications in clinical diagnostics and research. The utilization of NGS testing in the United States has been driven by advancements in technology, decreasing costs, and increased understanding of the human genome.

Hybrid-capture NGS is a powerful technique that combines the specificity of target capture with the high-throughput capabilities of NGS. It involves the enrichment of specific DNA targets using complementary probes, followed by sequencing and analysis of the captured targets. This approach allows for the detection and identification of a wide range of pathogens, including fungi and acid-fast bacteria, in a single assay.

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Clinical and Procedure
Clinical Utility

While there are differing versions of Next Generation Sequencing technologies in use, hybrid-capture NGS is the better choice for informative, actionable, and relevant diagnostics.  This method offers several advantages over traditional diagnostic techniques, such as culture-based, serological, and PCR methods. Hybrid-capture NGS can detect a wide range of clinically relevant species, including those that are difficult to culture or identify using conventional methods. It also provides a higher sensitivity and specificity, enabling the detection of low-level infections and the differentiation of closely related species. Invasive Fungal disease and Acid-Fast Bacteria testing using hybrid-capture NGS has the potential to improve patient outcomes by enabling early and accurate diagnosis, guiding appropriate (personalized medicine) therapy, and improved disease management strategies.

Procedure

Invasive fungal disease (IFD) and Acid-Fast Bacteria (AFB) testing using next-generation sequencing (NGS) is a diagnostic approach that utilizes advanced genetic sequencing technologies to detect and identify pathogens in clinical samples. NGS allows for the simultaneous analysis of multiple fungal species, providing a comprehensive and accurate assessment of the organism’s composition in a given sample.

  1. During the testing process, DNA is extracted from the clinical specimen and prepared for sequencing.
  2. A hybrid-capture step then selectively identifies specific DNA targets (sequences) that they want to study. This selection is based on what they are looking for, such as specific genes or regions associated with certain disease-causing microorganisms.
  3. These targets are "captured" or isolated from the rest of the DNA and then are amplified or copied many times to create more copies.
  4. This amplification helps to make sure there is enough material for the next step of the assay. The “captured” and “amplified” targets are then sequenced using a powerful technology called Next Generation Sequencing. This technology reads the genetic code of each captured DNA molecule, letter by letter, and generates a huge amount of data.
  5. The data that is generated is analyzed by specialized software to interpret the genomic information contained. This analysis helps scientists and healthcare professionals understand if there are any suspected disease-causing microorganisms present.

This method offers several advantages over traditional diagnostic techniques, such as culture-based methods or microscopy. NGS can detect a wide range of clinically relevant species, including those that are difficult to culture or identify using conventional methods. It also provides a higher sensitivity and specificity, enabling the detection of low-level fungal infections and the differentiation of closely related species.

Invasive fungal disease and Acid-Fast Bacteria testing using NGS has the potential to improve patient outcomes by enabling early and accurate diagnosis, guiding appropriate therapy, and monitoring treatment response.

Specificity

96% Specificity. 24 species validated in the wet-lab. 143 species validated in-silico.

Turnaround Time

<72 hours after receipt of sample by laboratory

Specimen Information
Specimen Type Test Code CPT Code NY Approved Volume Assay Range Special Instructions
whole blood 33356 No

10 ml Streck Cell-Free DNA BCT (Cat #218962) -or-

5 mL Streck Cell-Free DNA BCT (Cat #230637) -or-

4 mL K2-EDTA BCT. (Fisher Scientific Cat # 02-683-99C).

Streck Cell-free DNA BCT

10mL (Cat #218962) or

5mL (Cat #230637) tube

  • Tubes should be completely filled.
  • Store at ambient temperature.
  • Stable at ambient temperature for 7 days.

Refrigerated or frozen samples are not acceptable.

 

K2 EDTA BCT 4mL tube

(Fisher Scientific, Cat # 02-683-99C)

  • Tubes should be filled completely.
  • Spin and isolate plasma within 24 hours.
  • Store isolated plasma frozen.
  • Stable frozen for 30 days.
Shipping

Ship Monday through Friday. Friday shipments must be labeled for Saturday delivery. All specimens must be labeled with patient's name and collection date. A Viracor Eurofins test requisition form must accompany each specimen. Multiple tests can be run on one specimen. Ship specimens FedEx Priority Overnight® to: Viracor Eurofins, 18000 W 99th St. Ste, #10, Lenexa, KS 66219.

Causes for Rejection

Whole blood frozen, specimens beyond their acceptable length of time from collection as listed in the specimen handling, or specimen types or containers other than those listed.

Disclaimer

Specimens are approved for testing in New York only when indicated in the Specimen Information field above. The CPT codes provided are based on Eurofins Viracor’s interpretation of the American Medical Association's Current Procedural Terminology (CPT) codes and are provided for informational purposes only. CPT coding is the sole responsibility of the billing party. Questions regarding coding should be addressed to your local Medicare carrier. Eurofins Viracor assumes no responsibility for billing errors due to reliance on the CPT codes illustrated in this material. 

NeXGen Fungal / AFB next-generation sequencing (NGS) detects medically relevant fungal, acid fast bacteria (AFB) and selected bacterial species which are clinically similar to fungal infections derived from circulating cell free DNA (cfDNA) in the specimen. This test has been validated for human plasma isolated from whole blood collected in Streck BCT tubes. If an organism has been detected, the organism and the gene target(s) are indicated. A complete list of organisms that can been reported with this test can be found on our website.

This test was developed, and its performance characteristics determined by Eurofins Viracor. It has not been cleared or approved by the U.S. Food and Drug Administration. Results should be used in conjunction with clinical findings and should not form the sole basis for a diagnosis or treatment decision.

References

1) Chen P, Sun W, He Y. Comparison of the next-generation sequencing (NGS) technology with culture methods in the diagnosis of bacterial and fungal infections. J Thorac Dis 2020;12(9):4924-4929. doi: 10.21037/jtd-20-930
2) Fungal sequencing and classification with the ITS Metagenomics Protocol. www.illumina.com/company/legal.html. Pub. No. 1270-2018-001-B QB6724
3) Liquid biopsy for infectious diseases: a focus on microbial cell-free DNA sequencing. Dongsheng Han, Rui Li, Jiping Shi, Ping Tan, Rui Zhang and Jinming Li.. http://www.thno.org
4) Tsang, C.-C.; Teng, J.L.L.; Lau, S.K.P.;Woo, P.C.Y. Rapid Genomic Diagnosis of Fungal Infections in the Age of Next-Generation Sequencing. J. Fungi 2021, 7, 636. https://doi.org/10.3390/jof7080636
5) Artificial Intelligence in Precision Health. https://doi.org/10.1016/B978-0-12-817133-2.00018-5
6) Invasive fungal infections in liver transplantation Xia Liu, Zongxin Ling, Lanjuan Li, Bing Ruan. International Journal of Infectious Diseases 15 (2011) e298–e304
7) Closing the Gap in Surveillance and Audit of Invasive Mold Diseases for Antifungal Stewardship Using Machine Learning. Diva Baggio, Trisha Peel, Anton Y. Peleg, Sharon Avery, Madhurima Prayaga, Michelle Foo, Gholamreza Haffari, Ming Liu, Christoph Bergmeir and Michelle Ananda-Rajah. J. Clin. Med. 2019, 8, 1390; doi:10.3390/jcm8091390
8) Clinical application of cell-free next generation sequencing for infectious diseases at a tertiary children’s hospital. Julianne Wilke, Nanda Ramchandar, Christopher Cannavino, Alice Pong, Adriana Tremoulet, Leidy Tovar Padua, Helen Harvey, Jennifer Foley, Lauge Farnaes and Nicole G. Coufal. Wilke et al. BMC Infectious Diseases (2021) 21:552. https://doi.org/10.1186/s12879-021-06292-4
9) Amplicon-Based Next-Generation Sequencing for Detection of Fungi in Formalin-Fixed, Paraffin-Embedded Tissues Correlation with Histopathology and Clinical Applications. Paige M.K. Larkin, Katy L. Lawson, Deisy A. Contreras, Catherine Q. Le, Marisol Trejo, Susan Realegeno, Evann E. Hilt, Sukantha Chandrasekaran, Omai B. Garner, Gregory A. Fishbein, and Shangxin Yang.The Journal of Molecular Diagnostics, Vol. 22, No. 10, October 2020
10) https://blog.varsomics.com/en/what-is-next-generation-sequencing
11) https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6688590/
12) https://journals.asm.org/doi/10.1128/mbio.01468-23#
13) https://www.nature.com/scitable/definition/microarray-202/

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