Labs that cannot afford expensive multiplex tests for GI infections are turning to 24-hour reference labs using a testing platform that can run hundreds of samples per day.

By Anami Patel, Ph.D MB (ASCP)CM DLM CM

Infectious diarrhea also known as gastroenteritis, is a leading cause of death worldwide and a serious health risk to the growing population of vulnerable patients in the U.S. While the lack of potable water and inadequate sanitation are often to blame in many parts of the world, the primary cause in the U.S. is foodborne pathogen transmission. According to the U.S. Food and Drug Administration (FDA), 48 million cases of foodborne illness occur annually in the U.S., impacting one in every six Americans1. These illnesses result in an estimated 128,000 hospitalizations and 3,000 deaths in the U.S. every year1.

In the last two decades, rapid advancements in nucleic acid amplification testing (NAAT) via PCR for microbial detection have revolutionized the science of gastroenteritis diagnosis. Prior to NAAT, the detection of gastrointestinal (GI) pathogens was primarily done via stool culture, which is very time consuming and inaccurate. Time-to-results often takes several days or more with a high rate of false-negative results. This leads to delayed diagnosis and treatment2. These delays contribute to antimicrobial misuse and the development of antimicrobial resistance3.

The introduction of NAAT improved diagnosis of GI infections significantly. The PCR assays for GI infections improved accuracy and turnaround time. Prior to the development of syndromic panels, these tests only looked for a single pathogen and were unable to provide any information on co-infections.

Multiplex Testing for GI Infections

In the last decade, clinical microbiology has witnessed the burgeoning of multiplex syndromic NAAT panels for the detection of GI, respiratory, and bloodstream infections. These tests simultaneously detect and amplify nucleic acids from an array of bacteria, virus, and parasites that typically cause serious infections of these systems. These tests provide extremely rapid results with a level of accuracy that was impossible 20 years ago.

Some manufacturers have introduced multiplex syndromic molecular panels that provide results for a single patient sample in under an hour. In the hospital setting, these tests have had a dramatic, positive impact on the diagnosis and treatment of patients who are particularly vulnerable to opportunistic infections due to underlying illnesses, such as cancer. However, these kits typically are expensive and require the investment into a molecular testing platform, which can be costly.

There has emerged a need for multiplex syndromic molecular panels in the out-patient setting where near-immediate results are often not required. In truth, for many patients with suspected GI infections in the out-patient setting, multiplex syndromic molecular test results when delivered within 24 hours are more than sufficient to treat these patients accurately and in a timely manner. Considering the high entry costs of rapid test platforms, out-patient clinics are turning to reference laboratories for this service.

The Path Forward to Diagnosing GI Infections

Having participated as a clinical investigator in the validation study for the Applied BioCode Gastrointestinal Pathogen Panel (GPP) test in my previous role as a hospital laboratory director, I was eager to acquire this platform for PathAI, the reference lab where I currently serve as CSO of Molecular Diagnostics & Genomics. So, in 2018 as soon as this test was granted FDA 510(k) marketing clearance, we acquired it for our molecular lab.

PathAI is a large reference laboratory based in Memphis, Tenn., which provides results in under 24 hours to all 50 states. Its network includes all major payers plus over 100 commercial payer organizations, covering more than 260 million patients nationwide. Our molecular lab includes more than 3,000 square feet and is staffed by 16 clinical lab scientists. Our focus is on gastrointestinal infections, dermatopathology, urology, and women’s health. We resolve 90% of our cases within 72 hours.

The GPP test runs on the MDx-3000 system and detects 17 most common bacteria, viruses, and parasites that cause infectious diarrhea (see table below). The system automates the PCR amplification, hybridization, target capture, and detection steps of molecular diagnostic testing. The workflow supports processing 188 samples in an 8-hour shift.

Every day, PathAI receives 80-90 stool samples from around the country. In many cases, our GI testing is faster than a local hospital-based laboratory because we are really generating results in less than 24 hours. If a patient sample is collected at an early morning appointment or late in the day, these physicians receive results mid-afternoon the following day, affording them the ability to appropriately prescribe antimicrobial treatment the day they receive the results.

Even if physicians have access to molecular testing locally, many chose our service because they can avoid the common bottlenecks from staff shortages and maintenance issues so common to small laboratories. Customers return to PathAI lab services for our quality of service, speed, and broad clinical testing menu. In other words, we fill a need that many of these physicians and clinics don’t have locally.

Our physicians rely on the GPP test because they don’t have to choose the pathogens to test for, based on symptoms and clinical presentation. Using a syndromic panel, they get results from the 17 most likely GI pathogens.

The GI test process begins each morning with samples that can be processed as fresh stool or Cary Blair samples. We begin with an automated extraction step using the Roche MagNA Pure for extraction and purification. Samples and PCR reagents are then pipetted into a 96-well PCR plate. Once done, the PCR amplification, hybridization, and detection are completed in a fully automated process by the MDx-3000.

The timeline for this process is as follows:

  • 30 minutes to set up samples
  • 60 minutes for the extraction process
  • 30 minutes for PCR setup and consumable loading
  • 3.5 hours for automatic PCR amplification, target capture, signal generation, and detection

All told, from sample preparation to results, the process takes less than 6 hours for 96 samples—most of which is hands-off and automated. This frees staff time to perform other critical testing while awaiting results, which promotes efficient workflow. Additional benefits of the MDx-3000 system include its ease of use, small footprint, and the ability to mask results for pathogens that are not needed. For example, if the physician strongly suspects a parasitic GI infection, the MDx-3000 software allows for the masking of the viral and bacterial results in the report. If the doctor’s suspicion is wrong, the results for the bacterial and viral pathogens detected by the GPP can be unmasked, without the need of collecting a new sample. In fact, these results can be shared with clinicians immediately if requested. This feature not only benefits patients, but it also provides a cost-saving option to physicians.

Rapid parasite detection is an excellent example of the molecular testing capability of the MDx-3000 system. Traditionally, parasitology orders come in as ova and parasite (O&P) examinations. This requires a concentrate of stools to improve sensitivity. In fact, the CDC recommends that three stool samples be collected on different days to make an accurate diagnosis4. Ideally, this process would take 72 hours, but can often take twice as long whereas the GPP test provides results same day or the next day.

A key feature of this GI test and platform is the use of barcoded magnetic beads (BMBs) for detection. BMBs combine a semi-conductor manufacturing process with molecular and immunochemistry methodologies into a digital technology. This platform is capable of detecting multiple analytes in one test, thereby increasing the throughput of detection by conventional assays. The BMBs are fabricated by encasing paramagnetic material with biocompatible polymers. This results in a stable surface chemistry where the paramagnetic material exhibits magnetic properties for ease of washing, separation, and recovery. BMBs offer a digital multiplexing solution for syndromic detection of infectious agents. These beads provide 4,096 unique digital barcodes and the barcode patterns are designed to give high contrast signals, enabling accurate identification of target analytes. The beads are functionalized for coupling with nucleic acids, proteins, or other probe molecules, allowing high-density multiplex assays to be carried out in heterogeneous media.

The GPP test does not detect antimicrobial-resistant pathogens, but it does support good antimicrobial stewardship for physicians and clinics. By providing extremely accurate and sensitive results, clinical partners can make optimal use of their own antibiogram data to select a therapy that is appropriate for a particular infection.

Lastly, report generation by the MDx-3000 system is intuitive and user friendly. The reports provide positive, negative and invalid results. The MDx-3000 software integrates easily with the lab information system (LIS) and requires no additional middleware or other patches. The system is connected bi-directionally so when an order is received in LIS, it goes directly to the MDx-3000. When the run is complete, those results are transferred automatically to the LIS system.

Charting a GI Diagnostic Sea Change

Clinical lab scientists who began their careers 20 or more years ago are well aware of the revolution that has occurred in the detection of GI pathogens. The improved speed and accuracy provided by NAAT technology is nothing short of a sea change, one that has undoubtedly saved the lives of an untold number of patients.

The focus of this technology has been on vulnerable hospitalized patients who can be exposed to great risk from certain GI infections. The need is greatest among these patients so it is appropriate and good that the vulnerable were given top priority. But that’s not to say that this technology cannot be very beneficial to the general population, including out-patients with no underlying conditions.

The GPP when combined with a reference laboratory that can provide results within 24 hours is an affordable path forward to bring the benefits of robust multiplex syndromic testing to a much greater percentage of the population who suffer from GI infections in the U.S. each year.


Anami Patel, PhD, MB (ASCP)CM, DLM CM, has worked at PathAI for over six years, where he currently serves as chief scientific officer of Molecular Diagnostics & Genomics in Memphis, TN. Prior to that, he spent 13 years at the Le Bonheur Medical Center as laboratory director; and four years at the St. Jude Children’s Research Hospital as a research lab specialist.


  1.  What you need to know about foodborne illness, US FDA,128%2C000%20hospitalizations%20and%203%2C000%20deaths.
  2.  C. Hogan et al. Clinical Outcomes of Treated and Untreated C. difficile PCR-Positive/Toxin-Negative Adult Hospitalized Patients: a Quasi-Experimental Noninferiority Study. Clinical Microbiology; 25 May 2022.
  3. Beal SG, Tremblay EE, Toffel S et al.  A gastrointestinal PCR panel improves clinical management and lowers health care costs. J Clin Microbiol 2017; 56: e01457-17
  4. Diagnosis of Parasitic Diseases, Centers for Disease Control and Prevention.,special%20containers%20with%20preservative%20fluid.