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Rapid POCT for streptococcal infection


30 May, 2014  
A novel point-of-care test for Group A streptococcal infections reduces the reliance on bacterial culture for diagnosis of these conditions
 
Jukka Vakkila MD PhD
Docent in Pediatrics,
University of Helsinki;
and Chief of Pediatrics Clinic for Children
and Adolescents,
Mehiläinen Ltd, Finland
 
Streptococcus pyogenes, also known as Group A streptococcus (GAS) has been associated with variety of diseases. These include infections of the throat (pharyngitis, tonsillitis), skin (erysipelas, impetigo, perianal cellulitis) and, less commonly, suppurative arthritis, pneumonia, pericarditis, osteomyelitis and some other well described diseases.
 
In addition, rheumatoid fever, toxic shock syndrome, scarlet fever and necrotising fasciitis are caused by GAS. Although many of the above-mentioned infections are simple to treat and not life threatening, GAS may lead into rapid death of the patient if left untreated. Thus, as such, it is a very important pathogen in humans and significant efforts and costs are consumed for the diagnosis and treatment of GAS-infections worldwide.
 
Diagnosis
In theory, the diagnosis of streptococcus A disease is relatively straightforward. In typical cases, the symptoms and signs are easily recognisable by an experienced clinician and diagnosis can be confirmed either by streptococcal culture or rapid antigen tests that are commonly available. However, in practice the situation is more complex and the sensitivity and specificity of GAS diagnosis is far from perfect. Atypical cases are common and not all doctors are experienced clinicians.
 
Thus, even with the help of the most sensitive laboratory tests, a significant proportion of the patients remain undiagnosed because they will not be tested for GAS. The threshold for testing is naturally higher the longer it takes to receive the test results. By contrast, the specificity of the clinical diagnosis of GAS is not poor. Decisions by an experienced clinician are in line with the laboratory diagnoses in 70–80% of cases.(1) However, even with such expertise the frequency of misdiagnosed patients is much too high to be accepted since it would result in that 20–30% of patients would unnecessarily be treated by antibiotics.
 
Thus, a prerequisite for accurate diagnosis and therapy of GAS is a sensitive and specific laboratory test by which the presence or absence of S. pyogenes can be proven. For decades, the gold standard in GAS diagnosis has been bacterial culture of throat swabs on selective sheep blood plates. The method has some inherent theoretical or potential advantages over the other laboratory tests; dead bacteria or cell debris will not be detected, and bacteria grown on the plates are enriched and further investigations for the subsetting and strain identification are possible if regarded necessary.
 
The major obstacles include the lack of a universally accepted procedure for the culture and variations in sensitivity depending on whether the culture is performed at the clinician’s office or at the diagnostic laboratory. However, the biggest disadvantage relates to the delay in obtaining the results, that is, 24–72 hours may be required before the results are available. Patients and physicians are usually reluctant to hold the treatment decisions that long because of, for example, pressures to return to work as soon as possible. 
 
Rapid qualitative antigen detection
Rapid qualitative antigen detection tests have been developed to overcome the problem related with the delays in obtaining results. The fastest immunoassay tests can be read within five minutes from the addition of the reagents and sample. The specificity of rapid antigen tests is usually excellent. However, the sensitivity of the assays is inferior if compared with culture (53–97%) and therefore, it has been recommended that negative test results should be confirmed with a bacterial culture.(2) In practice, this recommendation is a complex and costly. Rapid tests relying on the use of molecular biology methods are at best as sensitive and specific as culture, but require special skills, facilities and equipment and, as such, do not represent ideal point-of-care (POC) tests. 
 
mariPOC
A recently developed rapid and automated near-patient test system, the mariPOC, (ArcDia International, Turku, Finland) is based on two-photon excitation fluorometry and concentration of antigen on microspheres by antigen–antibody reaction.(3) The technology utilises microvolume reaction chambers and separation-free fluorescent measurements. The technology allows real-time follow up of reaction kinetics and in this application the test reactions are read at 20 minutes and at two hours from the beginning of the reactions. Accordingly, strong positive samples can be revealed very rapidly and even the lowest positive samples can be detected at the point-of-care, all in the same single automated test. mariPOC was initially validated for eight respiratory viruses from nasopharyngeal aspirate and swab samples(4,5) and an application is available also for detection on GAS from throat swabs. We tested the applicability of the mariPOC GAS test at our clinics (Mehiläinen Ltd, Töölö and Turku, Finland) and compared its performance with bacterial culture both on symptomatic cases and asymptomatic controls.
 
A reporting level of 300CFU/ml resulted in 5.5% of healthy controls (n=109) and 26% of patients (n=121) being positive with the mariPOC system; the respective frequencies were 1.8% and 14% with bacterial culture. The group of patients (n=16) that were GAS-positive with mariPOC  but negative in culture was evaluated further. Available additional patient data were collected and confirmed the clinical significance of the mariPOC result in 13 out of 16 patients. These data confirmed that the GAS diagnosis varied in different patients and included, for example, clinical signs and symptoms typically found in acute and complicated GAS-diseases, a positive result with another rapid antigen test, recent GAS-positive bacterial culture of another family member, or conversion of a patient’s negative bacterial culture into a positive one within four weeks of the mariPOC testing date. 
 
Based on the above mentioned results, it appears that the mariPOC GAS test will be challenging the bacterial culture as the gold standard in diagnosis of GAS diseases. The specificity was similar to that found with culture but, in addition, several new GAS-positive cases were found with mariPOC that were not reported by culture. Additional data from these patients were convincing that they represented real clinical cases and not just asymptomatic carriers. 
 
The common problem for all present tests is that they do not distinguish different GAS strains, but report them equally. It may be, however, that GAS strains in asymptomatic carriers are different from those of symptomatic patients, and therefore the knowledge of the frequency of asymptomatic carriers may have little to do with clinical decision making. After all, in real life there are few patients who visit a physician’s office who would be tested for GAS without any GAS-related symptoms. We also need to bear in mind that the whole issue of asymptomatic carrier status is still not understood. It is not such a long time ago when there were no real concerns about the presence of Helicobacter pylori in gastric epithelia. 
 
Conclusions
Clinical diagnosis for patients with signs and symptoms suggesting a GAS-related disease is, in most cases, inaccurate and leads into excessive use of antibiotics without confirmation by laboratory assay. The gold standard for decades has been bacterial culture on selective plates, but in spite of being cheap and specific the results are delayed and therefore, more rapid POC tests are required. Our pilot trial, in which we compared the fully automated mariPOC GAS-test with bacterial culture provided data suggesting that the new rapid POC test is at least as sensitive as traditional culture. In addition, there were several clinically significant patients that could be found only by the mariPOC test, indicating that the sensitivity of mariPOC is better than that of culture. This may result in better understanding of asymptomatic carrier status in the future.
 
References
  1. Breese BB, Disney FA. The accuracy of diagnosis of beta streptococcal infections on clinical grounds. J Pediatrics 1954;44:670–3. 
  2. Gerber MA, Shulman ST. Rapid diagnosis of pharyngitis caused by group A streptococci. Clin Microbiol Rev 2004;17:571–80. 
  3. Koskinen JO et al Rapid method for detection of influenza A and B virus antigens by use of a two-photon excitation assay technique and dry chemistry reagents. J Clin Microbiol 2007;45:3581–8. 
  4. Ivaska L et al. Identification of respiratory viruses with a novel point-of-care multianalyte antigen detection test in children with acute respiratory tract infection. J Clin Virol 2013;57:136–40. 
  5. Tuuminen T, Suomala P, Koskinen JO. Evaluation of automated multianalyte point-of-care mariPOC test for the detection of influenza A virus and respiratory syncytial virus. J Med Virol 2013;85:1598–601.