SARS-CoV-2 viral RNA levels are not “viral load”

The susceptibility of Asian tiger mosquitoes to DENV-2 in different seasons was observed in simulated field environments as a reference to design dengue fever control strategies in Guangzhou. The life table experiments of mosquitoes in four seasons were carried out in the field. The susceptibility of Ae. albopictus to dengue virus was observed in both environments in Guangzhou in summer and winter. Ae. albopictus was infected with dengue virus by oral feeding. On day 7 and 14 after infection, the viral load in the head, ovary, and midgut of the mosquito was detected using real-time fluorescent quantitative PCR. Immune-associated gene expression in infected mosquitoes was performed using quantitative real-time reverse transcriptase PCR. The hatching rate and pupation rate of Ae. albopictus larvae in different seasons differed significantly. The winter hatching rate of larvae was lower than that in summer, and the incubation time was longer than in summer. In the winter field environment, Ae. albopictus still underwent basic growth and development processes. Mosquitoes in the simulated field environment were more susceptible to DENV-2 than those in the simulated laboratory environment. In the midgut, viral RNA levels on day 7 in summer were higher than those on day 7 in winter (F = 14.459, P = 0.01); ovarian viral RNA levels on day 7 in summer were higher than those on day 7 in winter (F = 8.656, P < 0.001), but there was no significant difference in the viral load at other time points (P > 0.05). Dicer-2 mRNA expression on day 7 in winter was 4.071 times than that on day 7 in summer: the viral load and Dicer-2 expression correlated moderately. Ae. albopictus could still develop and transmit dengue virus in winter in Guangzhou. Mosquitoes under simulated field conditions were more susceptible to DENV-2 than those under simulated laboratory conditions.

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Persistence of SARS-CoV-2 Viral RNA in Nasopharyngeal Swabs after Death: An Observational Study

Microorganisms ◽

10.3390/microorganisms9040800 ◽

2021 ◽

Vol 9 (4) ◽

pp. 800

Author(s):

Francesca Servadei ◽

Silvestro Mauriello ◽

Manuel Scimeca ◽

Bartolo Caggiano ◽

Marco Ciotti ◽

...

Keyword(s):

Viral Load ◽

Observational Study ◽

Real Time ◽

Real Time Pcr ◽

Pcr Detection ◽

Viral Rna ◽

Clinical Documentation ◽

Post Mortem ◽

Medical Assistance ◽

Time Points

The aim of this study was to investigate the persistence of SARS-CoV-2 in post-mortem swabs of subjects who died from SARS-CoV-2 infection. The presence of the virus was evaluated post-mortem from airways of 27 SARS-CoV-2 positive patients at three different time points (T1 2 h; T2 12 h; T3 24 h) by real-time PCR. Detection of antibodies to SARS-CoV-2 was performed by Maglumi 2019-nCoV IgM/IgG chemiluminescence assay. SARS-CoV-2 viral RNA was still detectable in 70.3% of cases within 2 h after death and in 66,6% of cases up to 24 h after death. Our data showed an increase of the viral load in 78,6% of positive individuals 24 h post-mortem (T3) in comparison to that evaluated 2 h after death (T1). Noteworthy, we detected a positive T3 post-mortem swab (24 h after death) from 4 subjects who were negative at T1 (2 h after death). The results of our study may have an important value in the management of deceased subjects not only with a suspected or confirmed diagnosis of SARS-CoV-2, but also for unspecified causes and in the absence of clinical documentation or medical assistance.

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Dengue Viral RNA Levels in Peripheral Blood Mononuclear Cells Are Associated with Disease Severity and Preexisting Dengue Immune Status

PLoS ONE ◽

10.1371/journal.pone.0051335 ◽

2012 ◽

Vol 7 (12) ◽

pp. e51335 ◽

Cited By ~ 30

Author(s):

Anon Srikiatkhachorn ◽

Sineewanlaya Wichit ◽

Robert V. Gibbons ◽

Sharone Green ◽

Daniel H. Libraty ◽

...

Keyword(s):

Disease Severity ◽

Peripheral Blood Mononuclear Cells ◽

Peripheral Blood ◽

Mononuclear Cells ◽

Immune Status ◽

Viral Rna ◽

Peripheral Blood Mononuclear ◽

Blood Mononuclear Cells ◽

Rna Levels

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Quantitative measurement of infectious virus in SARS-CoV-2 Alpha, Delta and Epsilon variants reveals higher infectivity (viral titer:RNA ratio) in clinical samples containing the Delta and Epsilon variants.

10.1101/2021.09.07.21263229 ◽

2021 ◽

Author(s):

Hannah W Despres ◽

Margaret G Mills ◽

David J Shirley ◽

Madaline M Schmidt ◽

Meei-Li Huang ◽

...

Keyword(s):

Quantitative Measurement ◽

Infectious Virus ◽

Viral Rna ◽

Clinical Samples ◽

Rt Pcr ◽

Genome Copy ◽

Live Virus ◽

High Degree ◽

The Relationship ◽

Rna Levels

ABSTRACT Background Novel SARS-CoV-2 Variants of Concern (VoC) pose a challenge to controlling the COVID-19 pandemic. Previous studies indicate that clinical samples collected from individuals infected with the Delta variant may contain higher levels of RNA than previous variants, but the relationship between viral RNA and infectious virus for individual variants is unknown. Methods We measured infectious viral titer (using a micro-focus forming assay) as well as total and subgenomic viral RNA levels (using RT-PCR) in a set of 165 clinical samples containing SARS-CoV-2 Alpha, Delta and Epsilon variants that were processed within two days of collection from the patient. Results We observed a high degree of variation in the relationship between viral titers and RNA levels. Despite the variability we observed for individual samples the overall infectivity differed among the three variants. Both Delta and Epsilon had significantly higher infectivity than Alpha, as measured by the number of infectious units per quantity of viral E gene RNA (6 and 4 times as much, p=0.0002 and 0.009 respectively) or subgenomic E RNA (11 and 7 times as much, p<0.0001 and 0.006 respectively). Conclusion In addition to higher viral RNA levels reported for the Delta variant, the infectivity (amount of replication competent virus per viral genome copy) may also be increased compared to Alpha. Measuring the relationship between live virus and viral RNA is an important step in assessing the infectivity of novel SARS-CoV-2 variants. An increase in the infectivity of the Delta variant may further explain increased spread and suggests a need for increased measures to prevent viral transmission.

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mTORC1 Restricts Hepatitis C Virus Replication Through ULK1-mediated Suppression of miR-122 and Facilitates Post-replication Events

10.1101/446757 ◽

2018 ◽

Author(s):

Manish Kumar Johri ◽

Hiren Vasantrai Lashkari ◽

Dhiviya Vedagiri ◽

Divya Gupta ◽

Krishnan Harinivas Harshan

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Viral Infection ◽

Virus Replication ◽

Viral Rna ◽

Hcv Rna ◽

Significant Drop ◽

Mechanistic Target Of Rapamycin ◽

Mtorc1 Activation ◽

Rna Levels

ABSTRACTMechanistic target of rapamycin (mTOR) is an important kinase that assimilates several upstream signals including viral infection and facilitates appropriate response by the cell through two unique complexes mTORC1 and mTORC2. Here, we demonstrate that mTORC1 is activated early during HCV infection as antiviral response. Pharmacological inhibition of mTORC1 promoted HCV replication as suggested by elevated levels of HCV (+) and (-) RNA strands. This was accompanied by significant drop in extracellular HCV RNA levels indicating defective post-replication stages. The increase in viral RNA levels failed to augment intracellular infectious virion levels, suggesting that mTORC1 inhibition is detrimental to post-replication steps. Lower infectivity of the supernatant confirmed this observation. Depletion of Raptor and ULK1 accurately reproduced these results suggesting that mTORC1 imparted these effects on HCV through mTORC1-ULK1 arm. Interestingly, ULK1 depletion resulted in increased levels of miR-122, a critical host factor for HCV replication, thus revealing a new mechanism of regulation by ULK1. The binary effect of mTORC1 on HCV replication and egress suggests that mTORC1-ULK1 could be critical in replication: egress balance. Interestingly we discover that ULK1 depletion did not interfere with autophagy in Huh7.5 cells and hence the effects on HCV replication and post-replication events are not resultant of involvement of autophagy. Our studies demonstrate an overall ULK1 mediated anti-HCV function of mTORC1 and identifies an ULK1-independent autophagy that allows HCV replication in spite of mTORC1 activation.

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Mesalamine Reduces Intestinal ACE2 Expression Without Modifying SARS-CoV-2 Infection or Disease Severity in Mice.

10.1101/2021.07.23.453393 ◽

2021 ◽

Author(s):

David M Alvarado ◽

Juhee Son ◽

Larissa B. Thackray ◽

Michael Diamond ◽

Siyuan Ding ◽

...

Keyword(s):

Viral Entry ◽

Post Infection ◽

Viral Rna ◽

Standardized Mortality Ratio ◽

Intestinal Loop ◽

Health Crisis ◽

Qrt Pcr ◽

Gi Symptoms ◽

Rna Levels

Introduction: Coronavirus Disease 2019 (COVID-19) is an ongoing public health crisis that has sickened or precipitated death in millions. The etiologic agent of COVID-19, Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), infects the intestinal epithelium, and can induce GI symptoms similar to the human inflammatory bowel diseases (IBD). An international surveillance epidemiology study (SECURE-IBD) reported that the standardized mortality ratio trends higher in IBD patients (1.5-1.8) and that mesalamine/sulfasalazine therapy correlates with poor outcome. The goal of our study was to experimentally address the relationship between mesalamine and SARS-CoV-2 entry, replication, and/or pathogenesis. Methods: Viral infection was performed with a chimeric vesicular stomatitis virus expressing SARS-CoV-2 spike protein and EGFP (VSV-SARS-CoV-2) and SARS-CoV-2 virus derived from an infectious cDNA clone of 2019n-CoV/USA_WA1/2020. Primary human ileal spheroids derived from healthy donors were grown as 3D spheroids or on 2D transwells. We assessed the effect of 10 mM mesalamine (Millipore Sigma) on viral RNA levels, as well as the expression of the SARS-CoV-2 receptor angiotensin II-converting enzyme 2 (ACE2), Transmembrane Serine Protease 2 (TMPRSS2), TMPRSS4, Cathepsin B (CTSB) and CTSL by qRT-PCR. 8-12 week old K18-ACE2 were treated orally with PBS or mesalamine at 200 mg/kg daily. Mice were inoculated intranasally with 1x10^3 FFU of SARS-CoV-2. Mice were weighed daily and viral titers were determined 7 days post infection (dpi) by qRT-PCR. For the intestinal viral entry model, VSV-SARS-CoV-2 was injected into a ligated intestinal loop of anesthetized K18-ACE2 mice and tissues were harvested 6 hours post-infection. Results: We found no change in viral RNA levels in human intestinal epithelial cells in response to mesalamine. Expression of ACE2 was reduced following mesalamine treatment in enteroids, while CTSL expression was increased. Mice receiving mesalamine lost weight at similar rates compared to mice receiving vehicle control. Mesalamine treatment did not change viral load in the lung, heart, or intestinal tissues harvested at 7 dpi. Pretreatment with mesalamine did not modulate intestinal entry of the chimeric VSV-SARS-CoV-2 in K18-ACE2 mice. Conclusions: Mesalamine did not alter viral entry, replication, or pathogenesis in vitro or in mouse models. Mesalamine treatment reduced expression of the viral receptor ACE2 while concurrently increasing CTSL expression in human ileum organoids.

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The Relationship between Simian Immunodeficiency Virus RNA Levels and the mRNA Levels of Alpha/Beta Interferons (IFN-α/β) and IFN-α/β-Inducible Mx in Lymphoid Tissues of Rhesus Macaques during Acute and Chronic Infection

Journal of Virology ◽

10.1128/jvi.76.16.8433-8445.2002 ◽

2002 ◽

Vol 76 (16) ◽

pp. 8433-8445 ◽

Cited By ~ 72

Author(s):

Kristina Abel ◽

Michelle J. Alegria-Hartman ◽

Kristina Rothaeusler ◽

Marta Marthas ◽

Christopher J. Miller

Keyword(s):

Virus Replication ◽

Acute Phase ◽

Rhesus Macaques ◽

Viral Rna ◽

Lymphoid Tissues ◽

Mrna Levels ◽

Immunodeficiency Virus ◽

Siv Infection ◽

Alpha Beta ◽

Rna Levels

ABSTRACT To define the role of alpha/beta interferons (IFN-α/β) in simian immunodeficiency virus (SIV) infection, IFN-α and IFN-β mRNA levels and mRNA levels of Mx, an antiviral effector molecule, were determined in lymphoid tissues of rhesus macaques infected with pathogenic SIV. IFN-α/β responses were induced during the acute phase and persisted in various lymphoid tissues throughout the chronic phase of infection. IFN-α/β responses were most consistent in tissues with high viral RNA levels; thus, IFN-α/β responses were not generally associated with effective control of SIV replication. IFN-α/β responses were differentially regulated in different lymphoid tissues and at different stages of infection. The most consistent IFN-α/β responses in acute and chronic SIV infection were observed in peripheral lymph nodes. In the spleen, only a transient increase in IFN-α/β mRNA levels during acute SIV infection was observed. Further, IFN-α and IFN-β mRNA levels showed a tissue-specific expression pattern during the chronic, but not the acute, phase of infection. In the acute phase of infection, SIV RNA levels in lymphoid tissues of rhesus macaques correlated with mRNA levels of both IFN-α and IFN-β, whereas during chronic SIV infection only increased IFN-α mRNA levels correlated with the level of virus replication in the same tissues. In lymphoid tissues of all SIV-infected monkeys, higher viral RNA levels were associated with increased Mx mRNA levels. We found no evidence that monkeys with increased Mx mRNA levels in lymphoid tissues had enhanced control of virus replication. In fact, Mx mRNA levels were associated with high viral RNA levels in lymphoid tissues of chronically infected animals.

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Hydrogen peroxide vapour treatment inactivates norovirus but has limited effect on post-treatment viral RNA levels

Infectious Diseases ◽

10.1080/23744235.2018.1546056 ◽

2019 ◽

Vol 51 (3) ◽

pp. 197-205 ◽

Cited By ~ 4

Author(s):

Torsten Holmdahl ◽

Inga Odenholt ◽

Kristian Riesbeck ◽

Patrik Medstrand ◽

Anders Widell

Keyword(s):

Hydrogen Peroxide ◽

Post Treatment ◽

Viral Rna ◽

Limited Effect ◽

Rna Levels

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Viral Kinetics and Resistance Development in Children Treated with Neuraminidase Inhibitors: The Influenza Resistance Information Study (IRIS)

Clinical Infectious Diseases ◽

10.1093/cid/ciz939 ◽

2019 ◽

Vol 71 (5) ◽

pp. 1186-1194 ◽

Cited By ~ 3

Author(s):

Rueshandra Roosenhoff ◽

Vaughan Reed ◽

Andy Kenwright ◽

Martin Schutten ◽

Charles A Boucher ◽

...

Keyword(s):

Viral Load ◽

Influenza A ◽

Antiviral Treatment ◽

Vaccination Status ◽

Viral Clearance ◽

Viral Rna ◽

Baseline Viral Load ◽

Influenza B ◽

Resistance Mutations ◽

Viral Burden

Abstract Background We studied the effect of age, baseline viral load, vaccination status, antiviral therapy, and emergence of drug resistance on viral shedding in children infected with influenza A or B virus. Methods Samples from children (aged ≤13 years) enrolled during the 7 years of the prospective Influenza Resistance Information Study were analyzed using polymerase chain reaction to determine the influenza virus (sub-)type, viral load, and resistance mutations. Disease severity was assessed; clinical symptoms were recorded. The association of age with viral load and viral clearance was examined by determining the area under the curve for viral RNA shedding using logistic regression and Kaplan-Meier analyses. Results A total of 2131 children infected with influenza (683, A/H1N1pdm09; 825, A/H3N2; 623, influenza B) were investigated. Age did not affect the mean baseline viral load. Children aged 1−5 years had prolonged viral RNA shedding (±1–2 days) compared with older children and up to 1.2-fold higher total viral burden. Besides, in older age (odds ratio [OR], 1.08; confidence interval [CI], 1.05–1.12), prior vaccination status (OR, 1.72; CI, 1.22–2.43) and antiviral treatment (OR, 1.74; CI, 1.43–2.12) increased the rate of viral clearance. Resistance mutations were detected in 49 children infected with influenza A virus (34, A/H1N1pdm09; 15, A/H3N2) treated with oseltamivir, most of whom were aged <5 years (n = 39). Conclusions Children aged 1−5 years had a higher total viral burden with prolonged virus shedding and had an increased risk of acquiring resistance mutations following antiviral treatment. Clinical Trials Registration NCT00884117.

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Discrepancies between Protease Inhibitor Concentrations and Viral Load in Reservoirs and Sanctuary Sites in Human Immunodeficiency Virus-Infected Patients

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.47.1.238-243.2003 ◽

2003 ◽

Vol 47 (1) ◽

pp. 238-243 ◽

Cited By ~ 125

Author(s):

Caroline Solas ◽

Alain Lafeuillade ◽

Philippe Halfon ◽

Stéphane Chadapaud ◽

Gilles Hittinger ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

Drug Resistance ◽

Lymph Node ◽

Viral Load ◽

Lymphoid Tissue ◽

Drug Penetration ◽

Hiv Rna ◽

Drug Concentrations ◽

Immunodeficiency Virus ◽

Rna Levels

ABSTRACT The variable penetration of antiretroviral drugs into sanctuary sites may contribute to the differential evolution of human immunodeficiency virus (HIV) and the emergence of drug resistance. We evaluated the penetration of indinavir, nelfinavir, and lopinavir-ritonavir (lopinavir/r) in the central nervous system, genital tract, and lymphoid tissue and assessed the correlation with residual viral replication. Plasma, cerebrospinal fluid (CSF), semen, and lymph node biopsy samples were collected from 41 HIV-infected patients on stable highly active antiretroviral therapy regimens to determine drug concentrations and HIV RNA levels. When HIV RNA was detectable, sequencing of the reverse transcriptase and protease genes was performed. Ratios of the concentration in semen/concentration in plasma were 1.9 for indinavir, 0.08 for nelfinavir, and 0.07 for lopinavir. Only indinavir was detectable in CSF, with a concentration in CSF/concentration in plasma ratio of 0.17. Differential penetration into lymphoid tissue was observed, with concentration in lymph node tissue/concentration in plasma ratios of 2.07, 0.58, and 0.21 for indinavir, nelfinavir, and lopinavir, respectively. HIV RNA levels were <50 copies/ml in all CSF samples of patients in whom HIV RNA was not detectable in plasma. HIV RNA was detectable in the semen of three patients (two patients receiving nelfinavir and one patient receiving lopinavir/r), and its detection was associated with multiple resistance mutations, while the viral load in plasma was undetectable. HIV RNA was detectable in all lymph node tissue samples. Differential drug penetration was observed among the three protease inhibitors in the sanctuary sites, but there was no correlation between drug levels and HIV RNA levels, suggesting that multiple factors are involved in the persistence of viral reservoirs. Further studies are required to clarify the role and clinical relevance of drug penetration in sanctuaries in terms of long-term efficacy and drug resistance.

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