scholarly journals A comparison of four commercially available RNA extraction kits for wastewater surveillance of SARS-CoV-2 in a college population

Author(s):  
Megan O'Brien ◽  
Zachary C. Rundell ◽  
Michelle D. Nemec ◽  
Laura M. Langan ◽  
Jeffrey A. Back ◽  
...  
2021 ◽  
Author(s):  
Megan O'Brian ◽  
Zachary C Rundell ◽  
Michelle D Nemec ◽  
Laura M Langan ◽  
Jeffrey A Back ◽  
...  

Localized wastewater surveillance has allowed for public health officials to gain a broader understanding of SARS-CoV-2 viral prevalence in the community allowing public health officials time to prepare for impending outbreaks. Given variable levels of virus in the population through public health interventions, proper concentration and extraction of viral RNA is a key step in ensuring accurate detections. With many commercial RNA extraction kits and methodologies available, the performance of 4 different kits were evaluated for SARS-CoV-2 RNA detection in wastewater, specifically focusing on their applicability to lower population densities such as those at university campus dorms. Raw wastewater samples were collected at 4 sites on a college campus over a 24 hour period as a composite sample. Included in these sites was an isolation site that housed students that tested positive for Covid-19 via nasopharyngeal swabs. These samples were analyzed using the following kits: Qiagen All Prep PowerViral DNA/RNA kit, New England BioLabs Monarch RNA MiniPrep Kit, and Zymo Quick RNA-Viral Kit, and the Zymo Quick-RNA Fecal/Soil Microbe MicroPrep Kit. All four sites were processed according to the manufacturers guidelines. Extractions were then quantified with RT-qPCR one-step reactions using an N2 primer and a linearized plasmid standard. While the Zymo Quick-RNA Fecal/Soil Microbe MicroPrep Kit (also known as the Zymo Environ Water RNA Kit) only recovered approximately 73% (+/- 38%) SARS-CoV-2 RNA compared to the Zymo Quick-RNA Viral kit, it was the most time efficient kit to yield comparable results. This extraction kit had a cumulative processing time of approximately five hours compared, while the other three kits had processing times between approximately 9 and 9.5 hours. Based on the current research, the most effective kits for smaller population densities are pellet based and include a homogenization, inhibitor removal, and RNA preservation step.


2014 ◽  
Author(s):  
Linsey R. High ◽  
Nicholas K. Lim ◽  
Ruth S. O'Brien ◽  
Chelsea V. Mitchell ◽  
Samantha J. T. Ross ◽  
...  

2014 ◽  
Vol 1 (1) ◽  
pp. 68-71
Author(s):  
A. Gerilovych ◽  
B. Stegniy ◽  
A. Stegniy ◽  
M. Stegniy ◽  
K. Smietanka ◽  
...  

Objective. To research the molecular characteristics of two HPAI strains – A/Ch/Syvash/02/05/H5N1 and A/Ch/Krasnogvardeysk/58/08/H5N1, which were identifi ed as representatives of the highly pathogenic H5N1 viruses. Methods. RNA extraction, real-time polymerase chain reaction (PCR). Results. The phylogenetic studies revealed that the above mentioned strains belong to two various genetic lineages originated from the Eastern European strains isolated in 2005, but differ from the viruses introduced to the Central and Western Europe in 2005/2006, and also the lineages consisting of H5N1 viruses isolated in the Europe and Middle East in late 2007. Conclusions. Relying on experimental studies, it can be concluded that the strains of A/Ch/Syvash/02/05/H5N1 and A/Ch/Krasnogvardeysk/58/08/H5N1 are highly pathogenic.


2018 ◽  
Vol 9 (03) ◽  
pp. 20192-20203
Author(s):  
Dr Maghsoudi, Hossein ◽  
Samaneh Haj-allahyari

Osteoarthritis (arthritis) is biomechanical, biochemical and cellular phenomenon, and is not known as a degenerative disease. Arthritis is one of the common chronic diseases and the most important reason of physical disability in the world. According to its side effect such as peptic ulcers, gastrointestinal bleeding, liver toxicity and renal complications dueofprescribing current treatment contain corticosteroid and non-steroidal, we decided to evaluate possible effect of anti-inflammatory Esential oil of Fraxinus excelsior (EOFE) on biomarkers involved in disease. EOFE were prepared of genetic resources center. Bovine  articular cartilage derived from the metacarpophalangeal joints of 14–18-month-old animals (without any sign of inflammation and bleeding) sent to laboratory in sterile bags at 4ºC. Cells were cultured in appropriate condition and counted by hemocytometer, viability assessed by trypan blue. After LPS treatment, cytokine levels were assayed. Cells cultures again and were kept in 37C, 90% humidity in CO2 incubator and after RNA extraction, RT-PCR and PCR done. Also by Real-time PCR, gene expression was evaluated. E.E.F.E level cause down regulation of COX-2, IL-1β, TNF-α in LPS-stimulated cells.


2010 ◽  
Vol 26 (3) ◽  
pp. 401-405
Author(s):  
Dong-dong LUAN ◽  
Qing-hua SHI ◽  
Hong CHEN ◽  
Yu-zhi DONG

2020 ◽  
Vol 17 (6) ◽  
pp. 397-407
Author(s):  
Maryam Jarchi ◽  
Farah Bokharaei-Salim ◽  
Maryam Esghaei ◽  
Seyed Jalal Kiani ◽  
Fatemeh Jahanbakhsh ◽  
...  

Background: The advent of resistance-associated mutations in HIV-1 is a barrier to the success of the ARTs. Objective: In this study, the abundance of HIV-1 infection in Iranian children, and also detection of the TDR in naïve HIV-1 infected pediatric (under 12 years old) were evaluated. Materials: From June 2014 to January 2019, a total of 544 consecutive treatment-naïve HIV-1- infected individuals enrolled in this study. After RNA extraction, amplification, and sequencing of the HIV-1 pol gene, the DRM and phylogenetic analysis were successfully performed on the plasma specimens of the ART-naïve HIV-1-infected-children under 12 years old. The DRMs were recognized using the Stanford HIV Drug Resistance Database. Results: Out of the 544 evaluated treatment-naïve HIV-1-infected individuals, 15 (2.8%) cases were children under 12 years old. The phylogenetic analyses of the amplified region of pol gene indicated that all of the 15 HIV-1-infected pediatric patients were infected by CRF35_AD, and a total of 13.3% (2/15) of these children were infected with HIV-1 variants with SDRMs (one child harbored two related SDRMs [D67N, V179F], and another child had three related SDRMs [M184V, T215F, and K103N]), according to the last algorithm of the WHO. No PIs-related SDRMs were observed in HIV-1-infected children. Conclusion: The current study demonstrated that a total of 13.3% of treatment-naïve HIV-1-infected Iranian pediatrics (under 12 years old) were infected with HIV-1 variants with SDRMs. Therefore, it seems that screening to recognize resistance-associated mutations before the initiation of ARTs among Iranian children is essential for favorable medication efficacy and dependable prognosis.


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