scholarly journals Correlation between Cycle Threshold (Ct) Value and IL-6 and D-Dimer in Chip-based RT-PCR Positive COVID-19 Cases: Study from a Stand-alone Laboratory

Author(s):  
Chhavi Gandhi ◽  
H. N. Ravikumar ◽  
Vani Ravikumar ◽  
C. Vani

Background: Novel Coronavirus (SARS-COV-2) is a leading cause of morbidity and mortality since the beginning of 2020 leading to range of symptoms from mild flu to respiratory distress, which is called COVID-19. RTPCR being the main diagnostic test can confirm the presence of the virus in the clinical samples, while various studies have defined Interleukin-6 and D-dimer as potent biomarker for severity. In this study, we have attempted to correlate the severity of COVID-19 with the presence of IL-6 and D-dimer and the Cycle threshold (Ct vlue) as determined by chip based RTPCR. Aim: The study aims to correlate the Cycle threshold value obtained after chip-based RT-PCR with markers such as IL-6 and D-Dimers.                                                      Methodology: It is a retrospective, observational study done in 799 subjects  in a span of three months (August 2020 to October 2020) at R V Metropolis Diagnostic and Healthcare Pvt Ltd. All symptomatic patients who tested positive in the Laboratory for COVID-19 by chip-based RT-PCR were included. Chip based RTPCR or Truenat test was performed on Nasopharyngeal swabs of the suspected subjects. Interleukin-6 was determined by Electrochemiluminiscence assay while D-dimer was done on the principle of Chemiluminiscence. Statistical Analysis Used: SPSS 12.0 version. Results: Total number of subjects enrolled were 799, with mean age of the subjects being 46.80± 17.55 years. In the study, males were found to be affected by COVID-19 more than females with ratio of male to female being 1.65:1. 498 (62.3%) of males presented with COVID-19 while it was observed in 301 (37.6%) females. Out of 799 subjects, 289 (36.2%) were symptomatic and out of 289 subjects, 140 (17.5% of total subjects) required hospitalisation. Cycle threshold values of both screening as well as confirmatory genes were determined separately in the cases of symptomatic and asymptomatic cases and there was no significant difference between the Ct values in cases of symptomatic and asymptomatic patients. Symptomatic patients were subcategorised under hospitalised and non-hospitalised and Again, no significant difference was seen between the two subset of patients in terms of Ct-value and, indirectly, the viral load of their clinical sample. The results convey that IL-6 and D-Dimer was significantly high (p=0.001 and <0.001 respectively) in case of symptomatic patients.D-Dimer was significantly high (p= <0.001) in the patients who needed hospitalisation. IL-6 was significantly raised as well (p=0.02). Screening and confirmatory gene were found to have no significant relationship with IL-6 and D-Dimer, neither any correlation was observed with them. Conclusion: Biomarkers such as Interleukin-6 and D-dimer can very well help in determining the severity and need for hospitalisation in a COVID-19 affected patient, but they have been found to have no relationship with cycle threshold value of RTPCR in our study.

Author(s):  
Andargachew Mulu ◽  
Dawit Hailu Alemayehu ◽  
Fekadu Alemu ◽  
Dessalegn Abeje Tefera ◽  
Sinknesh Wolde ◽  
...  

AbstractBackgroundThe coronavirus disease (COVID-19) pandemic has revealed the global public health importance of robust diagnostic testing. To overcome the challenge of nucleic acid (NA) extraction and testing kit availability efficient method is urgently needed.ObjectivesTo establish an efficient, time and resource-saving and cost-effective methods, and to propose an ad hoc pooling approach for mass screening of SARS-CoV-2MethodsDirect clinical sample and NA pooling approach was used for the standard reverse transcriptase polymerase chain reaction (RT-PCR) test of the SARS CoV-2 targeting the envelop (E) and open reading frame (ORF1ab) genomic region of the virus. In this approach, experimental pools were created using SARS CoV-2 positive clinical samples spiked with up to 9 negative samples prior to NA extraction step to have a final extraction volume of 200 μL (maximum dilution factor of 10). Viral NA was also subsequently extracted from each pool and tested using the SARS CoV-2 RT-PCR assay.ResultsWe found that a single positive sample can be amplified and detected in pools of up to 7 samples depending on the ct value of the original sample, corresponding to high, medium, and low SARS CoV-2 viral copies/reaction. However, to minimize false negativity of the assay with pooling strategies and with unknown false negativity rate of the assay under validation, we recommend poling of 4 in 1 using the standard protocols of the assay, reagents and equipment. The predictive algorithm indicated a pooling ratio of 4 in 1 was expected to retain accuracy of the test irrespective of the ct value (relative RNA copy number) of the sample spiked and result in a 237% increase in testing efficiency.ConclusionsThe approaches showed its concept in easily customized and resource-saving manner and would allow expanding of current screening capacities and enable the expansion of detection in the community.


2021 ◽  
pp. 24-30
Author(s):  
Aditi Munmun Sengupta ◽  
Diptendu Chatterjee ◽  
Bibhuti Saha

The real-time reverse transcription-polymerase chain reaction (RT-PCR) is considered as the sensitive proof for detecting the viral infection of the SARS-CoV-2 virus obtained from respiratory samples. The quantitative values for the analysis are benecial for estimating the transmissibility of people who test positive for SARS-CoV-2. This can be further achieved by analyzing the samples by semiquantitative means through the interpretation of the cycle threshold (Ct) values of RT-PCR that represent the rst cycle of PCR at which a detectable signal appears during the assays. The Ct value shows a correlation between high viral load and disease infectiousness, which is observed with other respiratory viruses, including the inuenza B infection and rhinovirus infection. Hence, the present study aims to analyze the surveillance of COVID-19 to monitor longer-term epidemiologic trends and trends in deaths due to COVID-19. In order to achieve this aim, the present review was reported to the preferred reporting items for systematic reviews and meta-analysis statements (PRISMA) for analyzing the Ct value-based epidemic predictions and to monitor long-term epidemiologic trends of SARS-CoV-2 virus. Total 33 studies have been nalized for nding out the results of the study. The epidemiologic parameter and a representative of the surveillance data for reporting to the World Health Organization were fullled by analyzing the systematic review and metaanalysis of the selected study. Moreover, the evaluation of the impact of the pandemic on the health care system and society was achieved by analyzing the studies mentioned here.


PLoS ONE ◽  
2021 ◽  
Vol 16 (2) ◽  
pp. e0247767
Author(s):  
Andargachew Mulu ◽  
Dawit Hailu Alemayehu ◽  
Fekadu Alemu ◽  
Dessalegn Abeje Tefera ◽  
Sinknesh Wolde ◽  
...  

Background The coronavirus disease 2019 (COVID-19) pandemic has revealed the global public health importance of robust diagnostic testing. To overcome the challenge of nucleic acid (NA) extraction and testing kit availability, an efficient method is urgently needed. Objectives To establish an efficient, time and resource-saving and cost-effective methods, and to propose an ad hoc pooling approach for mass screening of SARS-CoV-2. Methods We evaluated pooling approach on both direct clinical and NA samples. The standard reverse transcriptase polymerase chain reaction (RT-PCR) test of the SARS CoV-2 was employed targeting the nucleocapsid (N) and open reading frame (ORF1ab) genomic region of the virus. The experimental pools were created using SARS CoV-2 positive clinical samples and extracted RNA spiked with up to 9 negative samples. For the direct clinical samples viral NA was extracted from each pool to a final extraction volume of 200μL, and subsequently both samples tested using the SARS CoV-2 RT-PCR assay. Results We found that a single positive sample can be amplified and detected in pools of up to 7 samples depending on the cycle threshold (Ct) value of the original sample, corresponding to high, and low SARS CoV-2 viral copies per reaction. However, to minimize false negativity of the assay with pooling strategies and with unknown false negativity rate of the assay under validation, we recommend pooling of 4/5 in 1 using the standard protocols of the assay, reagents and equipment. The predictive algorithm indicated a pooling ratio of 5 in 1 was expected to retain accuracy of the test irrespective of the Ct value samples spiked, and result in a 137% increase in testing efficiency. Conclusions The approaches showed its concept in easily customized and resource-saving manner and would allow expanding of current screening capacities and enable the expansion of detection in the community. We recommend clinical sample pooling of 4 or 5 in 1. However, we don’t advise pooling of clinical samples when disease prevalence is greater than 7%; particularly when sample size is large.


2020 ◽  
Author(s):  
Andargachew Mulu ◽  
Dawit Hailu Alemayehu ◽  
Fekadu Alemu ◽  
Dessalegn Abeje Tefera ◽  
Sinknesh Wolde ◽  
...  

Abstract Background: The coronavirus disease (COVID-19) pandemic has revealed the global public health importance of robust diagnostic testing. To overcome the challenge of nucleic acid (NA) extraction and testing kit availability efficient method is urgently needed.Objectives: To establish an efficient, time and resource-saving and cost-effective methods, and to propose an ad hoc pooling approach for mass screening of SARS-CoV-2 Methods: Direct clinical sample and NA pooling approach was used for the standard reverse transcriptase polymerase chain reaction (RT-PCR) test of the SARS CoV-2 targeting the envelop (E) and open reading frame (ORF1ab) genomic region of the virus. In this approach, experimental pools were created using SARS CoV-2 positive clinical samples spiked with up to 9 negative samples prior to NA extraction step to have a final extraction volume of 200μL (maximum dilution factor of 10). Viral NA was also subsequently extracted from each pool and tested using the SARS CoV-2 RT-PCR assay.Results: We found that a single positive sample can be amplified and detected in pools of up to 7 samples depending on the ct value of the original sample, corresponding to high, medium, and low SARS CoV-2 viral copies/reaction. However, to minimize false negativity of the assay with pooling strategies and with unknown false negativity rate of the assay under validation, we recommend poling of 4 in 1 using the standard protocols of the assay, reagents and equipment. The predictive algorithm indicated a pooling ratio of 4 in 1 was expected to retain accuracy of the test irrespective of the ct value (relative ribonucleic acid RNA copy number) of the sample spiked and result in a 237% increase in testing efficiency. Conclusions: The approaches showed its concept in easily customized and resource-saving manner and would allow expanding of current screening capacities and enable the expansion of detection in the community.


Author(s):  
Neetu John ◽  
Rani Soni ◽  
Dev Jyoti Majumdar

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread globally and is a major public health issue. Procedures that have been established to decrease the spread of the virus depend on the careful and precise detection of infected individuals using quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). There have been many ambiguous concerns among the public regarding the severity of infection and its connection with the Cycle threshold (Ct) value and there was forceful need to inform the values to the public especially those with symptoms. The main objective of this study was to determine the association between the E (Envelop) gene Ct values of the symptomatic and asymptomatic COVID 19 patients. The study was conducted at the Virus Research and Diagnostic Laboratory (VRDL), Jagdalpur, Chhattisgarh. Between March 2020 and June 2020, samples were collected from the Bastar region as per the Indian Council of Medical Research (ICMR) guidelines. A total of 29228 clinical samples were tested by qRT-PCR targeting the E and RdRp (RNA-dependant RNA polymerase) genes as well as the ORF (open reading frame) gene that encode polyproteins of SARS-CoV-2. Of the 29228 samples tested, 75 were tested positive and 29153 were tested negative. In addition, the Ct values varied between the symptomatic and asymptomatic patients. It was observed that, the Ct values ranged from 15 to 32 in the asymptomatic patients and between 13 to 34 in the symptomatic patients. E gene Ct value analysis showed no significant difference between the asymptomatic and symptomatic COVID-19 patients. Thus, we observed that there was no association between the Ct values of symptomatic and asymptomatic patients of COVID-19.


2021 ◽  
Vol 30 (9) ◽  
pp. 11-17
Author(s):  
Hoang Vu Mai Phuong ◽  
Ung Thi Hong Trang ◽  
Nguyen Vu Son ◽  
Le Thi Thanh ◽  
Nguyen Le Khanh Hang ◽  
...  

From January to August 2020, Northern Viet Nam faced a COVID-19 outbreak, up to September 2020, there were 1122 confrmed cases of SARS-CoV-2, of which 465 cases were imported from Europe, America and Asia, 657 cases were identifed domestically. A total of 30,686 samples were collected during the SARS-CoV-2 outbreak in Northern Viet Nam and examined by Real-time RT-PCR using primers and probe from Charite - Berlin protocol. This study showed the initial results of SARS-CoV-2 detection and RNA quantitative in positive samples. The positive rate was 0.8%, ranging from 0.4 to 3.5% according to collection sites. Out of 251 positive samples, the mean Ct value was 28 (IQR: 22.3-32; range 14 - 38). The positive samples had a Ct value below 30 was 68.5%, there was no signifcant difference between the Ct value of the group ≤ 30 and > 30. The mean of the RNA copies/µl was 8.4.107, (IQR: 2.29.106 - 1.83.109 RNA copies/µl, range: 1.95.103 – 4.95.1011). In the group of imported COVID-19 cases, the rate of virus at low level was 29%, an average was 56% and at high level was 15%. In the community groups, the viral load data showed that the average rate at low, intermediate and high level were 20%, 63% and 17% respectively. The proportion of high-level viral load may raise an alert to start the quarantine process to reduce the transmission of SARS-CoV-2


Author(s):  
Ajitha Pottirayil ◽  
Shankar A. S. ◽  
Shaji Kannoth ◽  
Poorna Prasad ◽  
Sharath Kumar B. Jaikar ◽  
...  

Background: COVID-19 patients experience cytokine storm which cause pulmonary and extra-pulmonary complications even with currently available of standard of care. Additional antiviral and immune boosters are the need of hour to treat COVID-19 and to prevent post covid complications.Methods: In this study we enrolled 40 asymptomatic to mild COVID-19 patients to receive azadvir herbal steam inhaler along with standard of care. We evaluated the benefits of azadvir herbal steam inhaler by assessing RT-PCR conversion, clinical outcomes and improvement in immune markers (LDH, CRP, D-DIMER).Results: At the end of the study the immune markers improved significantly in study patients. In mild symptomatic cases IL-6 was 23.2 pg/ml on day 0 and 21.8 pg/ml on day 14. Reduction in IL-6 in mild symptomatic patients was statistically highly significant (p=0.0056). Mean IL-6 in asymptomatic patients was 22.3 pg/ml on day 0 and 21.1 pg/ml on day 14. Reduction in IL-6 in asymptomatic patients was statistically highly significant (p=0.0035).  Mean D-dimer was showing decreasing trend from day 0 to day 14 in mild symptomatic patients. In asymptomatic patients D dimer was 0.8 µg/ml on day 0 and 0.6 µg/ml on day 14. D-dimer decreased significantly from day 0 to day 14 (p value =0.0013). Mean LDH values on day 0 in mild symptomatic patients was 319.4 U/l and 219.3 on day 14. The reduction in LDH values in mild symptomatic patients is statistically significant (p value <0.0122). In asymptomatic patients mean LDH values on day 0 was 237 U/l and 194 U/l on day 14. The reduction in LDH values in asymptomatic group was statistically significant. Mean CRP values in mild symptomatic patients on day 0 was 12.2 mg/l and 3.8 mg/l on day 14. There was significant reduction in CRP values in mild symptomatic group which was statistically significant (p value =0.0546). Mean CRP values in asymptomatic patients on day 0 was 4.9 mg/l and 2.8 mg/l on day 14. There was significant reduction in mean CRP in asymptomatic patients which was statistically significant (p value =0.0446). In the present study all 40 patients (100%) cleared the virus and became negative for RT PCR test within 6 days. None of the patients progressed to severe COVID-19 and none of the patients succumbed to the disease.Conclusions: Azadvir accelerated recovery of COVID-19 patients by RT-PCR conversion, early improvement in clinical symptoms and immune markers in this study. This study results clearly indicates that azadvir has antiviral, immune booster activity and has definitive role in the management of asymptomatic to mild COVID-19 patients along with standard of care (CTRI no. CTRI/2020/06/026181).


2022 ◽  
Vol 13 (1) ◽  
pp. 13-18
Author(s):  
Jayasri Helen Gali ◽  
Manasa Musku ◽  
Devireddy Pallavi Reddy ◽  
Vimala Thomas ◽  
Ehsan Ahmed Khan

Background: Emergence of variants with specific mutations in key epitopes in the spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) raises concerns pertinent to its severity. Aims and Objectives: To assess the demographics, clinico-radiological pattern, and outcome of reverse transcription-polymerase chain reaction (RT-PCR) positive isolated coronavirus disease 2019 (COVID-19) patients arrived from the United Kingdom (UK). Materials and Methods: A cross-sectional, observational study was conducted in RT-PCR positive COVID-19 patients arrived from the UK, from December 2020 to February 2021. Nasopharyngeal samples of all patients were sent for whole-genome sequencing of SARS-CoV-2. The aimed parameters were compared between the B.1.17 positive and non-B.1.1.7 groups, among the people, arrived from the UK. All statistical tests with P<0.05 were considered significant. Results: A total of 59 SARS-CoV2 infected patients, who arrived from the UK, were isolated from December 2020 to February 2021 at Telangana Institute of Medical Sciences hospital, Hyderabad, were enrolled in the study. Of these, 27 patients (mean age-31.81±11.28 years) were infected with the B.1.1.7, diagnosed by whole genomic sequencing. Males were predominant in our study. Personal habits such as smoking, alcohol intake were higher among the sequenced group with a significant P<0.05. The most common symptoms observed in the sequenced group were cough (22.22%), sore throat (22.22%), cold (11.11%), fever (11.11%), and in the unsequenced group were cough (22.22%), cold (6.25%), fever (6.25%). In the sequenced group, chest X-ray posteroanterior view was normal in 74%, patchy ground-glass opacities was observed in 25.92% patients, whereas in the unsequenced group it was 84.37% and 15.62%, respectively. Asymptomatic patients, observed in the sequenced and unsequenced group were 48.18% and 78.12%, respectively, and symptomatic patients were 51.85% and 21.87%, respectively. We found a statistically significant difference between sequenced and unsequenced patients in the asymptomatic group with a significant P<0.05. In the sequenced and unsequenced group, mild cases were 48.18% and 18.75%, moderate cases were 3.7% and 3.12%, respectively. We found no evidence of an association between disease severity and lineage B.1.17. Conclusion: Our data, within the context and limitations of a real-world study, provide initial reassurance that severity in hospitalized patients with B.1.1.7 is not markedly different from severity in those without B.1.1.7.


2021 ◽  
Vol 10 (Supplement_2) ◽  
pp. S10-S11
Author(s):  
John Strutner ◽  
Nanda Ramchandar ◽  
Shruti Dubey ◽  
Mary Gamboa ◽  
Michelle K Vanderpool ◽  
...  

Abstract Background Understanding viral kinetics of SARS-CoV-2 is important to assess risk of transmission, manage treatment, and determine the need for isolation and protective equipment. Children have been noted to have less severe illness than adults and may have less transmission potential. We sought to determine whether children deemed to be asymptomatic had a difference in the PCR cycle threshold (Ct) value of respiratory samples from symptomatic children with SARS CoV-2 infection. Methods This was a retrospective cross-sectional study to compare PCR Ct values of 728 children who tested positive for SARS-CoV-2 by respiratory samples collected over a 4-month period. The study was a single center review of patients who tested positive for SARS-CoV-2 by RT-PCR from a respiratory specimen at a large tertiary care children’s hospital. Inclusion criteria included children 0–18 years of age who tested positive for SARS-CoV-2 by RT-PCR from a respiratory specimen for whom clinical information was available in the electronic medical record. Results We analyzed 728 children who tested positive for SARS-CoV-2 by RT-PCR from a respiratory sample over a 4-month period and for whom data was available in the electronic medical record. Overall, 71.2% of infected children were symptomatic. The mean Ct value for symptomatic patients (Ct mean 19.9, SD 6.3) was significantly lower than asymptomatic patients (Ct mean 23.5, SD 6.5) (P value &lt; 0.001, CI95th 2.6 - 4.6). The mean PCR Ct value was lowest in children less than 5 years of age. Conclusions and Relevance In this retrospective review of children who tested positive by RT-PCR for SARS CoV-2, the mean Ct was significantly lower in symptomatic children and was lowest in children under 5 years of age.


Author(s):  
Grace Lai-Hung Wong ◽  
Terry Cheuk-Fung Yip ◽  
Vincent Wai-Sun Wong ◽  
Yee-Kit Tse ◽  
David Shu-Cheong Hui ◽  
...  

Abstract Background Liver injury in patients with COVID-19 is common and prognostic. Direct viral tropism of SARS-CoV-2 for angiotensin-converting enzyme 2 receptors in hepatocytes may be one of the mechanisms of liver injury. We aimed to determine the role of viral persistence of SARS-CoV-2, based on cycle threshold (Ct) value, in liver injury in COVID-19. Methods This was a territory-wide retrospective cohort study of all public hospitals in Hong Kong. Laboratory-confirmed COVID-19 were identified. Serial liver biochemistries and Ct value of SARS-CoV-2 RNA were analyzed. Results We identified 7,622 COVID-19 patients (mean age 47 years, 48.2% male) diagnosed from 24 March to 1 January 2021 who had serial liver biochemistries and Ct values. 1,363 (17.9%) COVID-19 patients had ALT/AST elevations with two temporal patterns – early (within first 14 days from symptom onset) and late (after 14 days from symptom onset). COVID-19 patients with ALT/AST elevations had a lower Ct value at admission (23 vs. 25; P&lt;0.001), day 5 (24 vs. 26; P&lt;0.001) and day 20 (31 vs. 32; P&lt;0.001) after admission, compared to those without ALT/AST elevations. COVID-19 patients with ALT/AST elevations had a longer duration from first positive to first negative RT-PCR of SARS-CoV-2 (13 vs. 9 days; P&lt;0.001). ALT/AST elevation and presence of diabetes were the independent risk factors of viral persistence. Conclusions Liver injury in COVID-19 is linked to a higher SARS-CoV-2 viral load during the early phase of infection, signifying a possible direct viral injury to liver. Prolonged viral persistence of SARS-CoV-2 is associated with liver injury.


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