Analytical validation of a lymphocytic choriomeningitis virus real-time RT-PCR assay

Cordey, Samuel; Sahli, Roland; Moraz, Marie-Laurence; Estrade, Christine; Morandi, Laurence; Cherpillod, Pascal; Charrel, Rémi N.; Kunz, Stefan; Kaiser, Laurent

doi:10.1016/j.jviromet.2011.06.018

Cited by 23 publications

(20 citation statements)

References 22 publications

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“…Polymerase chain reaction has been utilized as a means of detecting LCMV mRNA in an infected infant 41 . The use of PCR offers exciting possibilities for both prenatal and postnatal detection of LCMV infection 42 . However, LCMV is not known to induce persistent infection in humans, and the time course of viral clearance from an infected human fetus is unknown.…”

Section: Diagnosismentioning

confidence: 99%

Lymphocytic Choriomeningitis Virus: An Underrecognized Cause of Neurologic Disease in the Fetus, Child, and Adult

Bonthius

2012

Seminars in Pediatric Neurology

164

155

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Lymphocytic choriomeningitis virus (LCMV) is an important cause of neurologic disease in humans. Carried and secreted principally by wild mice, LCMV covers a large geographic range and infects great numbers of people. Humans acquire LCMV disease when they come into contact with the secretions of infected mice. Because it has a strong neurotropism, the clinical signs and symptoms of LCMV infection are mostly neurologic. When the virus is acquired postnatally by children or adults, the clinical manifestations are usually those of aseptic meningitis. Most people who acquire LCMV infection during childhood or adulthood are moderately symptomatic for several weeks, but have a full recovery. A much more severe disease ensues when the infection occurs prenatally. LCMV can infect the fetal brain and retina, where it leads to substantial injury and permanent dysfunction. The possibility of LCMV infection should be considered in all babies with evidence of congenital infection, especially those with prominent neurologic signs, such as microencephaly, periventricular calcifications, and hydrocephalus.

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Section: Diagnosismentioning

confidence: 99%

Lymphocytic Choriomeningitis Virus: An Underrecognized Cause of Neurologic Disease in the Fetus, Child, and Adult

Bonthius

2012

Seminars in Pediatric Neurology

164

155

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“…Recently, several studies illustrating improved methods for detecting LCMV, especially real time RT-PCR for viral RNA, have been reported [9], [10], [11]. It has been shown that RNA detection is a rapid, sensitive method for identifying LCMV viral presence in murine tissues and clinical samples, which has several advantages over the standard plaque assay ( Table 2 ).…”

Section: Discussionmentioning

confidence: 99%

“…There are several factors that limit the value of RNA analysis for viral quantification in research as opposed to clinical usage. Quantification is generally expressed in RNA copies and only one study [11] represented RNA copies and LOD values in terms of PFU/ml, the standard experimental measure for LCMV and most other infections. Additionally, especially for lower limits of detection, the presence of viral RNA does not necessarily correlate with active viral infection.…”

Section: Discussionmentioning

confidence: 99%

“…Several promising techniques, including Real-Time RT-PCR for GP and NP transcripts, have been developed and utilized with some frequency for in vivo and in vitro detection of LCMV. Quantitation of LCMV RNA is highly reproducible and sensitive, detecting as few as five RNA copies or the equivalent of 10 PFU/ml of virus [9], [10], [11]. However, one potential drawback of quantification of viral titers in tissues and serum by Real-time RT-PCR is that the number of viral RNA copies present cannot be directly correlated with infectious virus, particularly in light of the well-characterized presence of defective interfering virions in LCMV infection [12], [13].…”

Section: Introductionmentioning

confidence: 99%

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Accelerated and Improved Quantification of Lymphocytic Choriomeningitis Virus (LCMV) Titers by Flow Cytometry

Johnson

Homann

2012

PLoS ONE

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Lymphocytic choriomeningitis virus (LCMV), a natural murine pathogen, is a member of the Arenavirus family, may cause atypical meningitis in humans, and has been utilized extensively as a model pathogen for the study of virus-induced disease and immune responses. Historically, viral titers have been quantified by a standard plaque assay, but for non-cytopathic viruses including LCMV this requires lengthy incubation, so results cannot be obtained rapidly. Additionally, due to specific technical constraints of the plaque assay including the visual detection format, it has an element of subjectivity along with limited sensitivity. In this study, we describe the development of a FACS-based assay that utilizes detection of LCMV nucleoprotein (NP) expression in infected cells to determine viral titers, and that exhibits several advantages over the standard plaque assay. We show that the LCMV-NP FACS assay is an objective and reproducible detection method that requires smaller sample volumes, exhibits a ∼20-fold increase in sensitivity to and produces results three times faster than the plaque assay. Importantly, when applied to models of acute and chronic LCMV infection, the LCMV-NP FACS assay revealed the presence of infectious virus in samples that were determined to be negative by plaque assay. Therefore, this technique represents an accelerated, enhanced and objective alternative method for detection of infectious LCMV that is amenable to adaptation for other viral infections as well as high throughput diagnostic platforms.

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“…LCMV infection is considered to be underdiagnosed ( 6 , 7 ), although the virus was not detected in several studies that prospectively tested for LCMV in large numbers (>90) of clinical encephalitis or meningitis patients ( 8 – 10 ). Despite infrequently causing aseptic meningitis or mild disease in immunocompetent persons, LCMV has the potential to cause severe disease and should be considered for patients with a compatible illness and potential rodent exposure.…”

mentioning

confidence: 99%

Lymphocytic Choriomeningitis Virus Infections among American Indians

Knust¹,

Holman²,

Redd³

et al. 2013

Emerg. Infect. Dis.

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Analytical validation of a lymphocytic choriomeningitis virus real-time RT-PCR assay

Cited by 23 publications

References 22 publications

Lymphocytic Choriomeningitis Virus: An Underrecognized Cause of Neurologic Disease in the Fetus, Child, and Adult

Lymphocytic Choriomeningitis Virus: An Underrecognized Cause of Neurologic Disease in the Fetus, Child, and Adult

Accelerated and Improved Quantification of Lymphocytic Choriomeningitis Virus (LCMV) Titers by Flow Cytometry

Lymphocytic Choriomeningitis Virus Infections among American Indians

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