Effect of heat inactivation for the detection of severe acute respiratory syndrome-corona virus-2 (SARS-CoV-2) with reverse transcription real time polymerase chain reaction (rRT-PCR): evidence from Ethiopian study
Abstract Background Coronavirus disease 2019 (COVID-19) has been a major public health importance and its specimen needs to be handled safely due to concerns of potential transmissibility to health care workers. Heat inactivation of the sample before nucleic acid isolation might permit safe testing...
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| Format: | Article |
| Language: | English |
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BMC
2022-02-01
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| Series: | BMC Infectious Diseases |
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| Online Access: | https://doi.org/10.1186/s12879-022-07134-7 |
| _version_ | 1828429289210511360 |
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| author | Belete Woldesemayat Gebremedihin Gebremicael Kidist Zealiyas Amelework Yilma Sisay Adane Mengistu Yimer Gadissa Gutema Altaye Feleke Kassu Desta |
| author_facet | Belete Woldesemayat Gebremedihin Gebremicael Kidist Zealiyas Amelework Yilma Sisay Adane Mengistu Yimer Gadissa Gutema Altaye Feleke Kassu Desta |
| author_sort | Belete Woldesemayat |
| collection | DOAJ |
| description | Abstract Background Coronavirus disease 2019 (COVID-19) has been a major public health importance and its specimen needs to be handled safely due to concerns of potential transmissibility to health care workers. Heat inactivation of the sample before nucleic acid isolation might permit safe testing processes. Hence, it is important to assess the effect of heat inactivation on SARS-CoV-2 RT-PCR detection in resource limited settings. Methods An experimental study was conducted at Ethiopian Public Health Institute (EPHI) from September 25 to October 15, 2020. A total of 188 Oro-pharyngeal swabs were collected from COVID-19 suspected cases, referred to EPHI for SARS COV-2 testing. One batch of the sample was inactivated at 56 °C heat for 30 min, and the other batch was stored at 4 °C for a similar period of time. RNA extraction and detection were done by DAAN Gene kit protocols. Abbott m2000 RT-PCR was used for amplification and detection. Data analysis was done by using SPSS version 23.0; Chi-square and Pearson correlation test for qualitative and semi-quantitative analysis were used. p-value < 0.05 was considered as statistically significant. Results Out of 188 total samples, 119 (63.3%) were positive and 69 (36.7%) were negative in the non-inactivated group. While, 115 (61.2%) of samples were positive and 73 (38.8) were negative in heat inactivated sample batch. Rate of positivity between groups did not have statistically significant difference (p > 0.05). The mean Cycle threshold (Ct) value difference between the two groups of ORF1a/b gene and N gene was 0.042 (95% CI − 0.247–0.331; t = 0.28; p = 0.774) and 0.38 (95% CI 0.097–0.682; t = 2.638; p = 0.010) respectively. Conclusion Heat inactivation at 56 °C for 30 min did not affect the qualitative rRT-PCR detection of SARS-CoV-2. However, the finding showed that there was statistically significant Ct value increment after heat inactivation compared to untreated samples. Therefore, false-negative results for high Ct value (Ct > 35) samples were found to be the challenge of this protocol. Hence alternative inactivation methods should be investigated and further studies should be considered. |
| first_indexed | 2024-12-10T17:32:08Z |
| format | Article |
| id | doaj.art-afeca9a749e4424e8a84f6f4db416d87 |
| institution | Directory Open Access Journal |
| issn | 1471-2334 |
| language | English |
| last_indexed | 2024-12-10T17:32:08Z |
| publishDate | 2022-02-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Infectious Diseases |
| spelling | doaj.art-afeca9a749e4424e8a84f6f4db416d872022-12-22T01:39:40ZengBMCBMC Infectious Diseases1471-23342022-02-012211910.1186/s12879-022-07134-7Effect of heat inactivation for the detection of severe acute respiratory syndrome-corona virus-2 (SARS-CoV-2) with reverse transcription real time polymerase chain reaction (rRT-PCR): evidence from Ethiopian studyBelete Woldesemayat0Gebremedihin Gebremicael1Kidist Zealiyas2Amelework Yilma3Sisay Adane4Mengistu Yimer5Gadissa Gutema6Altaye Feleke7Kassu Desta8HIV/AIDS Disease Research Team, TB and HIV/AIDS Disease Research Directorate, Ethiopian Public Health InstituteHIV/AIDS Disease Research Team, TB and HIV/AIDS Disease Research Directorate, Ethiopian Public Health InstituteHIV/AIDS Disease Research Team, TB and HIV/AIDS Disease Research Directorate, Ethiopian Public Health InstituteHIV/AIDS Disease Research Team, TB and HIV/AIDS Disease Research Directorate, Ethiopian Public Health InstituteHIV/AIDS Disease Research Team, TB and HIV/AIDS Disease Research Directorate, Ethiopian Public Health InstituteHIV/AIDS Disease Research Team, TB and HIV/AIDS Disease Research Directorate, Ethiopian Public Health InstituteHIV/AIDS Disease Research Team, TB and HIV/AIDS Disease Research Directorate, Ethiopian Public Health InstituteHIV/AIDS Disease Research Team, TB and HIV/AIDS Disease Research Directorate, Ethiopian Public Health InstituteDepartment of Medical Laboratory Sciences, College of Health Sciences, Addis Ababa UniversityAbstract Background Coronavirus disease 2019 (COVID-19) has been a major public health importance and its specimen needs to be handled safely due to concerns of potential transmissibility to health care workers. Heat inactivation of the sample before nucleic acid isolation might permit safe testing processes. Hence, it is important to assess the effect of heat inactivation on SARS-CoV-2 RT-PCR detection in resource limited settings. Methods An experimental study was conducted at Ethiopian Public Health Institute (EPHI) from September 25 to October 15, 2020. A total of 188 Oro-pharyngeal swabs were collected from COVID-19 suspected cases, referred to EPHI for SARS COV-2 testing. One batch of the sample was inactivated at 56 °C heat for 30 min, and the other batch was stored at 4 °C for a similar period of time. RNA extraction and detection were done by DAAN Gene kit protocols. Abbott m2000 RT-PCR was used for amplification and detection. Data analysis was done by using SPSS version 23.0; Chi-square and Pearson correlation test for qualitative and semi-quantitative analysis were used. p-value < 0.05 was considered as statistically significant. Results Out of 188 total samples, 119 (63.3%) were positive and 69 (36.7%) were negative in the non-inactivated group. While, 115 (61.2%) of samples were positive and 73 (38.8) were negative in heat inactivated sample batch. Rate of positivity between groups did not have statistically significant difference (p > 0.05). The mean Cycle threshold (Ct) value difference between the two groups of ORF1a/b gene and N gene was 0.042 (95% CI − 0.247–0.331; t = 0.28; p = 0.774) and 0.38 (95% CI 0.097–0.682; t = 2.638; p = 0.010) respectively. Conclusion Heat inactivation at 56 °C for 30 min did not affect the qualitative rRT-PCR detection of SARS-CoV-2. However, the finding showed that there was statistically significant Ct value increment after heat inactivation compared to untreated samples. Therefore, false-negative results for high Ct value (Ct > 35) samples were found to be the challenge of this protocol. Hence alternative inactivation methods should be investigated and further studies should be considered.https://doi.org/10.1186/s12879-022-07134-7COVID-19Ct valueHeat inactivationrRT-PCRSARS-CoV-2 |
| spellingShingle | Belete Woldesemayat Gebremedihin Gebremicael Kidist Zealiyas Amelework Yilma Sisay Adane Mengistu Yimer Gadissa Gutema Altaye Feleke Kassu Desta Effect of heat inactivation for the detection of severe acute respiratory syndrome-corona virus-2 (SARS-CoV-2) with reverse transcription real time polymerase chain reaction (rRT-PCR): evidence from Ethiopian study BMC Infectious Diseases COVID-19 Ct value Heat inactivation rRT-PCR SARS-CoV-2 |
| title | Effect of heat inactivation for the detection of severe acute respiratory syndrome-corona virus-2 (SARS-CoV-2) with reverse transcription real time polymerase chain reaction (rRT-PCR): evidence from Ethiopian study |
| title_full | Effect of heat inactivation for the detection of severe acute respiratory syndrome-corona virus-2 (SARS-CoV-2) with reverse transcription real time polymerase chain reaction (rRT-PCR): evidence from Ethiopian study |
| title_fullStr | Effect of heat inactivation for the detection of severe acute respiratory syndrome-corona virus-2 (SARS-CoV-2) with reverse transcription real time polymerase chain reaction (rRT-PCR): evidence from Ethiopian study |
| title_full_unstemmed | Effect of heat inactivation for the detection of severe acute respiratory syndrome-corona virus-2 (SARS-CoV-2) with reverse transcription real time polymerase chain reaction (rRT-PCR): evidence from Ethiopian study |
| title_short | Effect of heat inactivation for the detection of severe acute respiratory syndrome-corona virus-2 (SARS-CoV-2) with reverse transcription real time polymerase chain reaction (rRT-PCR): evidence from Ethiopian study |
| title_sort | effect of heat inactivation for the detection of severe acute respiratory syndrome corona virus 2 sars cov 2 with reverse transcription real time polymerase chain reaction rrt pcr evidence from ethiopian study |
| topic | COVID-19 Ct value Heat inactivation rRT-PCR SARS-CoV-2 |
| url | https://doi.org/10.1186/s12879-022-07134-7 |
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