The Importance of Diagnostic Testing in SARS-CoV-2 Adverse Outcomes
SARS-CoV-2 (COVID-19), a highly contagious disease, primarily manifests as an acute respiratory illness, however, for those with health complications, including: autoimmune diseases, asthma, heart disease and diabetes, the risk of developing serious illness and adverse outcomes is much greater. It is estimated that 1 in 6 will experience adverse outcomes that could be life-threatening 1. The spread and devastation of COVID-19 highlights the vital role laboratory diagnostics plays in the diagnosis and management of suspected and affected patients. As the COVID pandemic continues, it is imperative that fast and accurate diagnostic testing strategies are implemented for effective risk stratification and monitoring of treatment and recovery.
In this article, we will review how Randox Reagents can aid in diagnosing and managing SARS-CoV-2 adverse outcomes.
The immune system activates a pro-inflammatory response to enhance host immunity against viruses and decrease colonisation and infection, but only if the pro-inflammatory response is controlled. Uncontrolled pro-inflammatory responses can result in a cytokine storm 2. A cytokine storm is a serious complication associated with SARS-CoV-2, which can trigger life-threatening pneumonia, acute respiratory distress syndrome (ARDS) and multiple organ failure 3, 4. Cytokine storms occur in 5% of severe COVID-19 cases, with several inflammatory cytokines observed at high levels. Due to the elevation of several pro-inflammatory and anti-inflammatory cytokines, a multiplex immunoassay approach can offer several advantages over the widely utilised single ELISA tests. The simultaneous detection of multiple cytokines from a single patient sample will provide clinicians with a comprehensive overview of cytokine markers and complete patient profile, facilitating a personalised treatment plan to be implemented 5, 6.
In COVID-19 patients, CRP testing has proved to perform well in discriminating disease severity and predicting adverse outcomes 7. Elevated CRP levels have been identified in 86% of patients admitted to hospital. CRP measurements are useful in diagnosis, prognosis and monitoring for clinical improvements or deterioration. Moreover, the acute phase reactant, ferritin, has been observed to increase in approximately 60% of COVID-19 patients. In the critically ill COVID-19 patients, extremely elevated ferritin concentrations were recorded, which could be attributed to a cytokine storm and secondary haemophagocytic lymphohistiocytosis (a hyper-inflammatory syndrome associated with multi-organ failure) 8.
Acute kidney injury (AKI) is a common complication in diabetic patients who test positive for COVID-19. Regardless, the National Institute for Health and Care Excellence (NICE) recommend AKI testing in all COVID-19 patients upon hospital admission and their condition monitored throughout their stay 9.
The most commonly utilised screening test for renal impairment is serum creatinine (SCr); however, it is important to consider the accuracy and reliability of the method. Two commercially available methods exist for SCr determination: Jaffe and enzymatic. Whilst the Jaffe method is less expensive, it is more susceptible to interferences which can lead to the misdiagnosis of patients, which isn’t ideal in the current pandemic 7. Moreover, the sensitivity of SCr, regardless of method, in the early detection of renal disease is poor, as SCr is insensitive to small changes in glomerular filtration rate (GFR). Up to 50% of renal function can potentially be lost before significant SCr levels become detectable 8, 9. In comparison, cystatin C (CysC) is a superior marker of renal function and is useful in the determination of the extent of renal damage, as well as distinguishing those with severe and mild COVID-19 10.
Abnormal liver function tests significantly increases a COVID-19 patients risk of developing severe disease and complications such as pneumonia 11. Bilirubin levels, 3 times the upper limit have been observed in COVID-19 patients 11, 12. Whilst the diazo method is commonly utilised in bilirubin testing, superior methods exist. The vanadate oxidation (VO) method offers several advantages particularly in haemolytic and lipemic samples. These advantages are particularly evident in neonatal and infant populations where haemolysis is extremely common. Moreover, the VO method offers a wider analytical measurement range for the comfortable detection of clinically important results 13.
Other liver function markers are known to be elevated in COVID-19 patients including both AST and ALT, with markers like albumin decreased.
The Importance of Lp(a) Testing
Lipoprotein(a) / Lp(a), a strong independent marker of cardiovascular disease risk has recently been identified as a key risk marker of cardiovascular complications in COVID-19 patients. Those with either baseline elevated or elevated levels of Lp(a) following COVID-19 infection may be at a significantly increased risk of developing thromboses. Consideration should be given to measurement of Lp(a) and prophylactic anticoagulation of infected patients to reduce the risk. Elevated Lp(a) levels may also cause acute destabilization of pre-existing but quiescent, atherosclerotic plaques, which could induce an acute myocardial infarction (AMI) or cerebrovascular accident (CVA) (stroke) 14.
The size heterogeneity of apo(a) isoforms represents the biggest challenge faced by laboratories in accurately measuring Lp(a). The variable numbers of repeated KIV-2 units in act as multiple epitopes, and so standardisation across calibrators is vital. Unless the calibrants have the same range of isoforms as test samples, those with higher numbers of the KIV-2 repeat, will represent with an overestimation in Lp(a) concentrations and those with smaller numbers of the KIV-2 repeat, will represent with an underestimation. The smaller isoforms are strongly associated with higher Lp(a) concentrations 15.
Lp(a) assays that are standardised to the WHO/IFCC (World Health Organization/International Federation of Clinical Chemistry) reference material, transferring values from mg/dl to nmol/l are more uniform. The assay considered the most reliable commercially available Lp(a) assay is so because: 15
1. The isoform size variations are reduced as a range of calibrators from separate pools of serum used, which covered a range of Lp(a) concentrations.
2. The isoform size and concentrations are inversely correlated, better matching calibrants with test samples.
3. Methods are calibrated in nmol/l and traceable to WHO/IFCC reference material and give acceptable bias compared with the Northwest Lipid Metabolism and Diabetes Research Laboratory (NLMDRKL) gold standard method.
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 Complications coronavirus can cause. https://www.webmd.com/lung/coronavirus-complications#1 (accessed 22 July 2020).
 Sinha P, Matthey MA, Calfee CS. Is a “cytokine storm” relevant to COVID-19? JAMA 2020; (): 1-3. https://jamanetwork.com/journals/jamainternalmedicine/fullarticle/2767939#:~:text=Why%20has%20the%20%E2%80%9Ccytokine%20storm,and%20associated%20with%20adverse%20outcomes.&text=Several%20early%20case%20series%20in,elevated%20above%20the%20normal%20range. (accessed 22 July 2020).
 Del Valle DM, Kim-Schulze S, Huang HH, Beckmann M, Nirenberg S, et al. An inflammatory cytokine signature helps predict COVID-19 severity and death. The Preprint Server for Health Sciences https://www.medrxiv.org/content/10.1101/2020.05.28.20115758v1.full.pdf (accessed 24 July 2020).
 The Royal College of Pathologists. Guidance on the use and interpretation of clinical biochemistry tests in patients with COVID-19 infection. https://www.rcpath.org/uploads/assets/3f1048e5-22ea-4bda-953af20671771524/G217-RCPath-guidance-on-use-and-interpretation-of-clinical-biochemistry-tests-in-patients-with-COVID-19-infection.pdf (accessed 14 June 2020).
 Dhungana N, Morris C, Krasowski MD. Operational impact of using a vanadate oxidase method for direct bilirubin measurements at an academic medical center clinical laboratory. Practical Laboratory Medicine 2017; 8(2017): 77-85.
 Moriarty PM, Gorby LK, Stroes ES, Kastelein JP, Davidson M, et al. Lipoprotein(a) and its potential association with thrombosis and inflammation in COVID-19: a testable hypothesis. Current Atherosclerosis Reports 2020; 22(48): 1-8.