A nested real-time PCR assay has an increased sensitivity suitable for detection of viruses in aerosol studies

Perrott, Phillipa; Smith, Greg; Ristovski, Zoran; Harding, Robert M.; Hargreaves, Megan

doi:10.1111/j.1365-2672.2008.04119.x

Cited by 24 publications

(20 citation statements)

References 19 publications

(22 reference statements)

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“…1). Protocols that are commonly used in bioaerosol studies include the 16S rRNA gene for total bacteria (Blais Lecours et al, 2012;Masclaux et al, 2013;Nehme et al, 2008;Oppliger et al, 2008), total archaea (Blais Lecours et al, 2012), influenza (Perrott et al, 2009), Saccharopolyspora rectivirgula (Blais Lecours et al, 2012) and bacteriophages (Verreault et al, 2011).…”

Section: Molecular Tools For Detection and Quantification Of Noncultumentioning

confidence: 99%

Non-culturable bioaerosols in indoor settings: Impact on health and molecular approaches for detection

Blais-Lecours

Perrott

Duchaine

2015

Atmospheric Environment

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h i g h l i g h t sNon-culturable fraction of bioaerosols is important but still misunderstood. Airborne non-culturable agents can be hazardous for human respiratory health. Molecular tools allow the detection of airborne non-culturable microorganisms. Culture-dependent and independent tools should be used for bioaerosol studies. Several research needs relating to non-culturable agents must be addressed. a b s t r a c tDespite their significant impact on respiratory health, bioaerosols in indoor settings remain understudied and misunderstood. Culture techniques, predominantly used for bioaerosol characterisation in the past, allow for the recovery of only a small fraction of the real airborne microbial burden in indoor settings, given the inability of several microorganisms to grow on agar plates. However, with the development of new tools to detect non-culturable environmental microorganisms, the study of bioaerosols has advanced significantly. Most importantly, these techniques have revealed a more complex bioaerosol burden that also includes non-culturable microorganisms, such as archaea and viruses. Nevertheless, air quality specialists and consultants remain reluctant to adopt these new researchdeveloped techniques, given that there are relatively few studies found in the literature, making it difficult to find a point of comparison. Furthermore, it is unclear as to how this new non-culturable data can be used to assess the impact of bioaerosol exposure on human health. This article reviews the literature that describes the non-culturable fraction of bioaerosols, focussing on bacteria, archaea and viruses, and examines its impact on bioaerosol-related diseases. It also outlines available molecular tools for the detection and quantification of these microorganisms and states various research needs in this field.

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Section: Molecular Tools For Detection and Quantification Of Noncultumentioning

confidence: 99%

Non-culturable bioaerosols in indoor settings: Impact on health and molecular approaches for detection

Blais-Lecours

Perrott

Duchaine

2015

Atmospheric Environment

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“…For these reasons, the coliphage MS2 (Golmohammadi et al 1993), a small non-enveloped bacterial virus (Leviviridae family) with a capsid diameter of 25 nm and a genome made of a single-stranded RNA (ssRNA) molecule (3569 nucleotides), has been used as an airborne viral model in many aerosol studies (Barker and Jones 2005;Burton et al 2007;Eninger et al 2009;Grinshpun et al 2007;Hogan et al 2005;Hogan et al 2006;Hogan et al 2004;Holton and Webb 1994;Kettleson et al 2009;Li et al 2009;Perrott et al 2009;Tseng and Li, 2005;Walker and Ko, 2007;Wang and Brion 2007). The Pseudomonas syringae virulent phage phi 6 is another well characterized bacterial virus that has some interesting features as a surrogate for RNA viruses, although it has been used only sporadically in aerosol studies (Ellis and Schlegel 1974).…”

Section: Introductionmentioning

confidence: 99%

Evaluation of Filters for the Sampling and Quantification of RNA Phage Aerosols

Gendron

Verreault

Veillette

et al. 2010

Aerosol Science and Technology

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This study exploits the virulent bacteriophages phi 6 (dsRNA) and MS2 (ssRNA) as surrogates for airborne RNA viruses. Two different filter types, polytetrafluoroethylene (PTFE) and polycarbonate (PC), were tested for their efficiency in collecting aerosolized RNA phages. Two commercial kits were tested for total RNA isolation. Also, heat shock treatments were performed in three different media to obtain the most favorable conditions for reverse transcription assays of dsRNA. Our findings suggest that PC filters are more suitable to recover infectious airborne RNA viruses as determined by plaque assays. Both types of filters were equally efficient in recovering RNA from aerosolized phage phi 6 as established by qRT-PCR. Viral samples should be treated with QIAamp Viral RNA Mini Kit and a 5 min heat shock treatment at 110• C in TE buffer before RT-PCR to maximize detection of phage phi 6. Overall, the infectivity of the recovered phages was severely affected by the aerosolization/air sampling process and the presence of RNA viruses in air samples should be determined by qRT-PCR.

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“…For diagnosis, more sensitive methods might be necessary to detect respiratory viruses present in EBC since it is unpredictable how diluted the viral particles in the specimen are. Recently, nested qRT-PCR assays have been developed to allow a more sensitive detection of viruses in aerosols [16].…”

Section: Discussionmentioning

confidence: 99%

Exhaled breath condensate sampling is not a new method for detection of respiratory viruses

Houspie

Coster

Keyaerts

et al. 2011

Virol J

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BackgroundExhaled breath condensate (EBC) sampling has been considered an inventive and novel method for the isolation of respiratory viruses.MethodsIn our study, 102 volunteers experiencing upper airway infection were recruited over the winter and early spring of 2008/2009 and the first half of the winter of 2009/2010. Ninety-nine EBCs were successfully obtained and screened for 14 commonly circulating respiratory viruses. To investigate the efficiency of virus isolation from EBC, a nasal swab was taken in parallel from a subset of volunteers. The combined use of the ECoVent device with the RTube™ allowed the registration of the exhaled volume and breathing frequency during collection. In this way, the number of exhaled viral particles per liter air or per minute can theoretically be estimated.ResultsViral screening resulted in the detection of 4 different viruses in EBC and/or nasal swabs: Rhinovirus, Human Respiratory Syncytial Virus B, Influenza A and Influenza B. Rhinovirus was detected in 6 EBCs and 1 EBC was Influenza B positive. We report a viral detection rate of 7% for the EBCs, which is much lower than the detection rate of 46.8% observed using nasal swabs.ConclusionAlthough very promising, EBC collection using the RTube™ is not reliable for diagnosis of respiratory infections.

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A nested real-time PCR assay has an increased sensitivity suitable for detection of viruses in aerosol studies

Cited by 24 publications

References 19 publications

Non-culturable bioaerosols in indoor settings: Impact on health and molecular approaches for detection

Non-culturable bioaerosols in indoor settings: Impact on health and molecular approaches for detection

Evaluation of Filters for the Sampling and Quantification of RNA Phage Aerosols

Exhaled breath condensate sampling is not a new method for detection of respiratory viruses

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