The microbiome of the upper airways: focus on chronic rhinosinusitis

14 Pages·2015·0.97 MB·English

by  Thanit Chalermwatanachai

Chalermwatanachaietal.WorldAllergyOrganizationJournal (2015) 8:3 journal
DOI10.1186/s40413-014-0048-6
REVIEW Open Access
The microbiome of the upper airways: focus on
chronic rhinosinusitis
Thanit Chalermwatanachai1,2, Leydi Carolina Velásquez1,3 and Claus Bachert1,4*
Abstract
Upper airway diseases including allergic rhinitis, chronic rhinosinusitis withor without polyps, and cystic fibrosis are
characterized by substantially different inflammatoryprofiles. Traditionally, studies ontheassociationof specific
bacterial patterns with inflammatory profiles of diseaseshad been dependent on bacterial culturing. In thepast
30 years, molecular biology methods have allowed bacterialculture freestudies of microbial communities, revealing
microbiota much more diverse than previously recognized including those found in theupper airway.
Atpresence,thestudyofthepathophysiologyofupperairwaydiseasesisnecessarytoestablishtherelationshipbetween
themicrobiomeandinflammatorypatternstofindtheirclinicalreflectionsandalsotheirpossiblecausalrelationships.
Suchinvestigationsmayelucidatethepathtotherapeuticapproachesincorrectinganimbalancedmicrobiome.
Inthereviewwesummarizedtechniquesusedandthecurrentknowledgeonthemicrobiomeofupperairwaydiseases,
thelimitationsandpitfalls,andidentifiedareasofinterestforfurtherresearch.
Introduction peopleand people withdiseasessuchas asthma [6-8]and
It is generally believed that exposure to microorganism cystic fibrosis (CF) [8,9]. Unfortunately, with limited stu-
compromises health. Reduced exposure to microbiota re- dies currently available, it cannot be concluded with the
sults in decrease of incidence of infectious diseases but samedegreeforchronicrhinosinusitis(CRS)[10].
may adversely increase the incidence of allergic disorders Research on microbiome in CRS is therefore needed
[1-3]. Recent developments of culture-independent tools to elucidate pathophysiology of this disease such as; 1)
make it possible to identify microbial species not previ- the relationship between the microbiome and inflamma-
ouslydetectedbyconventionalmethods.Unbeknownstto tory patterns, 2) possible causal relationships between
us,manhadbeenlivingwiththesemicrooraganismssince microbe and CRS,3)investigation ofthe microbiome re-
thedawnoftime. garding possible therapeutic properties. Dysregulation of
The human body harbors from 10 to 100 trillion mi- the interactions between the immune system and com-
crobeswhichgreatlyoutnumberourownhumancells[4]. mensal bacteria is a contributing factor to the develop-
This bacterial assemblage has been coined, “the human mentandchronicityofanumberofinflammatorydiseases
microbiota” [4]. Subsequently, a project called “Human [11].Microorganismsinthegutmayplayasignificantrole
Microbiome” was established to investigate the flora in in regulating T helper cells (Th cells), regulatory T cells
healthy volunteers and their relationship tohuman health (Tregs)anddendriticcellsaswellasToll-likereceptorex-
and disease [5]. The study of the host-microbe relation- pression in sentinel cell (macrophage and dendritic cells)
ship has shown that microbes play a major role in our which are relevant to airway illnesses such as asthma and
well-being [4,6]. Alterations of microbial composition allergicdiseases[10].
have been linked to several human diseases [4]. There is
also evidence showing that, in the respiratory system, Techniquesinmicrobiotastudy
composition of airway microbiota varies between healthy Principal approaches to analyze human microbiota are:
culture-dependent and culture-independent techniques.
*Correspondence:[email protected] Culture-dependentmethodsinvolveisolationandculturing
1DepartmentofOto-Rhino-Laryngology,TheUpperAirwaysResearch of microorganisms prior to their identification accord-
Laboratory(URL),GhentUniversityHospital,Ghent9000,Belgium
ing to morphological, biochemical or genetic characteris-
4DivisionofENTDiseases,Clintec,KarolinskaInstitutet,Stockholm,Sweden
Fulllistofauthorinformationisavailableattheendofthearticle tics. These methods are time-consuming, due to culture
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Chalermwatanachaietal.WorldAllergyOrganizationJournal (2015) 8:3 Page2of14
andbias,ascertainmediaandgrowthconditionsfavorthe (FISH), and quantitative PCR (qPCR), and are based on
growthofsomebacteriaoverothers[12].Inaddition,this oligonucleotideprobesandprimersthattargettheriboso-
approachmaynotprovideatruereflectionofthediversity mal RNA sequences or other genes in different hybridi-
of microbes in a sample. A “no growth” result does not zation procedures. Thus these techniques require a prior
necessarily imply that a sample is sterile. It is estimated knowledge of the microbial DNA sequence. A DNA mi-
that up to 99% of microorganisms observable in nature croarray(alsocommonlyknownas DNAchiporbiochip)
typically cannot be cultured by standard techniques [13]. isacollection ofmicroscopic DNA spots(oligonucleotide
Un-cultivability is a widespread condition that includes: probes) attached to a solid surface. It is usually used for
(i) organisms for which the specific growth requirements gene expression analysis or screening of single nucleotide
(nutritional, temperature, aeration, etc.) are not fulfilled; polymorphisms. The FISH technique uses fluorescent
(ii)slow-growingorganismsareout-competedinthepres- probes that bind to only those parts of the chromosome
ence of fast-growing microorganisms and (iii) disfavored with which they show a high degree of sequence com-
organisms, which cannot stand the stressful conditions plementarity. It detects and localizes the presence of
imposed by cultivation [13,14]. This approach camou- specific DNA sequences on chromosomes. qPCR or
flagesthetruebacterialcommunity.Thereneededtobea real-timePCRfollowsthegeneralprinciple ofPCRwith
betterapproachtoanalyzethesemicroorganisms. the advantage of detecting the amount of initial DNA
Since the 1980s,the application ofmolecular detection in samples using either fluorescent DNA-binding dyes
methods has allowed culture-independent investigations or fluorescence-tagged oligonucleotide probes [15].
ofthemicrobialcommunities[15].Moleculartechniques The introduction of next generation sequencing chan-
have proven effective in characterizing complex micro- ged the history of genomic research as it increased se-
bial assemblages in environmental samples [16]. How- quencing throughput, and did not require prior cloning
ever, an important usefulness of molecular techniques is steps [25]. These technologies are not only changing our
the ability to detect genetic materials of non-viable mi- genome sequencing approaches and the associated time-
croorganisms [17,18]. Culture-independent methods are lines and costs [26], but also developing many excit-
based on the direct analysis of bacterial DNA (or RNA) ing fields such as metagenomics, metatranscriptomics
without culturing. Due to the sensitivity of these tech- and single-cell genomics [15]. Three platforms for high
niques, special care and attention is required for proce- throughput parallel DNA sequencing are in reasonably
dures that include sample collection and handling, DNA widespreaduseatpresent:theRoche/454FLX,theIllumina
extraction, amplification of gene fragments, distinction (MiSeq,HiSeq,andNextSeq),andtheIonTorrent.
of different fragments, identification of microorganism At presence, researchers have a large choice in formu-
andanalysis ofthemicrobialcommunity [15]. lating methodological strategies: depending on the access
For bacterial identification, the predominant gene tar- tothetechnology,budget,andobjectivesofresearch.Each
get for amplification has been the 16S ribosomal RNA culture-independent methodology has its own limitations
gene (or 16S rRNA) [19,20], which is a component of and biases, investigators must take additional measures;
the 30S small subunit of prokaryotic ribosomes [21]. It for example one may use more than one molecular tech-
hasbeenwidelytargeted because of(i)itspresenceinal- nique or a culture-dependent approach in parallel to
most all bacteria, often existing as a multigene family, or provide additional validation of results and reduce the
as operon; (ii) the conservation of the 16S rRNA gene, possibility of false findings due to methodological errors
suggesting that random sequence changes are a measure and biases. Although the culture-independent techniques
of time (evolution) rather than a reflection of different have the ability to detect more microbes than culture
bacteria; and (iii) the size of 16S rRNA genes (1,500 bp) technique,theculture-dependentmethodssofarremaina
being large enough for informatic purposes [22]. More- better means of obtaining individual isolates contributing
over, there are several available reference sequences and andobtainingisolatesforfurtherassays.
taxonomies databases such as Greengenes, SILVA and
theRibosomaldatabaseproject[23].However,amplifica- Microbiotainallergicrhinitis
tion of target genes using polymerase chain reaction As the gate into our body, the respiratory tract itself
(PCR) has made it impossible to completely avoid PCR- harbors a heterogeneous microbiota that decreases in
basedbiasesandchimeraproduction.Itthusmay distort biomass from upper to lower tract [27]. Even in health,
thelevelofdiversityandbacterialcompositioninasample recent findings indicated that direct exposure to bacter-
because of the amplification of pseudogenes [24]. There- ial communities in the airways may provide an explan-
fore, other technologies are often used as complementary ation for how commensal bacteria can regulate chronic
approaches to 16S rRNA gene sequencing for reducing airway inflammation [11]. Since the observation that in-
distortion of bacterial diversity and composition. They fections within households in early childhood have a role
are DNA microarray, fluorescence in situ hybridization inpreventingallergicrhinitis[3],numerousepidemiologic
Chalermwatanachaietal.WorldAllergyOrganizationJournal (2015) 8:3 Page3of14
andexperimentalstudieshavesoughttoclarifyandextend suggesting that each CFairway infection is unique, with
the so-called hygiene hypothesis with respect to asthma relatively stable and resilient bacterial communities [44].
and other allergic and autoimmune disorders. The evi- The diversity and species richness of fungal and bacterial
dence supporting the hygiene hypothesis established the communities were significantly lower in patients with de-
“microbiota (microflora) hypothesis”. This concept pro- creased lung function and poor clinical status [39]. The
poses that perturbations in gastrointestinal bacteria have authors observed a strong positive correlation between
disruptedthemechanismsofmucosalimmunologictoler- low species richness and poor lung function [37]. These
ance, which has led to an increase in the incidence of al- findingsshowthecriticalrelationshipbetweenairwaybac-
lergic airway disease [28]. Independent studies found that terialcommunitystructure,diseasestage,andclinicalstate
a reduced diversity of the gut microbiota in infancy is as- atthetimeofsamplecollection[42].
sociatedwithanincreasedriskofallergicmanifestationat The main microorganisms found in CFairways are the
school age [29-31]. The association between the compos- genera Haemophilus, Pseudomonas, Staphylococcus and
ition of microbiota in the intestine, asthma and allergic Stenotrophomonas. Less common are gram-negatives,
disease is nowadays of high interest [32,33], although the Streptococcus and Mycobacterium spp [45]. Most bac-
exact mechanism of the interaction of intestinal-systemic teriaofCFairwaysaredifficultto cultureusing conven-
immunity is still not defined [34,35]. There are several tional clinical methods; therefore, molecular approaches
publicationsreviewingtherelationshipsbetweenintestinal mayconfirmorrevealnovelbacteriathatmightberelated
microbes and asthma [7,10,36]. Suggesting that the intes- to the pathogenesis of cystic fibrosis. Examples of interest
tinal microbiome contributes to the regulation of local are the Streptococcus milleri group (Streptococcus angino-
andsystemicinflammatoryresponsesvia short-chainfatty sus, Streptococcus intermedius, Streptococcus constellatus)
acids,aproduct offermentation of dietaryfibersbyintes- [46], Pseudomonas intermedia [46], and Gemella species
tinal bacteria [11]. Following this model, it is likely that [47] (Table 1). Further experiments suggested that these
therespiratorymicrobiotamayalsohaveanimpactonair- bacteria could act as co-pathogens or enhance the viru-
way inflammation in allergic responses [1,11], however, lenceofconventionalCFpathogens[48].
thisneedsfurtherinvestigation.
Themicrobiomeinchronicrhinosinusitis
Microorganismsintheairwaysofcysticfibrosispatients Specimen collection is one of the most important steps
Cystic fibrosis (CF) is an autosomal recessive genetic in the analysis of remote areas such as the sinuses. An
disorder that affects among other organs the lungs and appropriate collection of the samples is the first step to
sinuses. It is characterized by abnormal transport of perform a meaningful, high quality analysis. It must not
chloride and sodium across the epithelium, leading to be biased by interference from the nares. Specimen can
thick, viscous secretions. This leads todefective mucocili- be tissue, nasal secretions [55] or material sampled by a
aryclearanceandchronicairwayinfectionwithacomplex swab. The use of an endoscope for the sampling during
microbiota [37]. Lung disease in cystic fibrosis results sinus surgery is advisable [17,18,56], although simple
from chronic airway infection and inflammation leading swabs are often used [57] in both healthy and diseased
to progressive bronchiectasis and respiratory failure [38]. patients [18]. Samples can be collected from various ana-
SpecimensformolecularmicrobialanalysisinCFhavefor tomicallocationsinthenosesuchastheinferiorturbinate
the most part all derived from sputum [37,39-42], while [55], the middle-meatus [56], the ethmoidal sinuses, the
swabs from the middle meatus in patients with sinusitis sphenoid [18], and the anterior nasal cavity [58]. Mucosal
were not used until now. Thus, no conclusions can be surfaces of the lateral, central, and medial portions of the
made for the upper airways yet. Samples consisted of maxillary sinus are also collected from locations in the
serial collections of more than six patients in most of nose [17]. The use of middle-meatus swabs for DNA-
the studies [37,42-44]. based bacterial assays is appropriate for the detection of
Previous studies indicated that exacerbations might be multiplebacterialspecies,includinganaerobes,whichmay
associated with changes in microbial densities and the be undetected when swabs are used solely for culture.
acquisitionofnew microbialspecies[37].Bacterialpatho- Based on available cultivation-based studies, the micro-
gens, including Pseudomonas aeruginosa, Staphylococcus biology of the middle meatus correlates well with patho-
aureus, and Burkholderia cepacia are known contribu- genicorganismofCRS,whereasswabsofthenareswould
torsto such exacerbations.Recent studies using the latest not be appropriate as a replacement of middle meatus
culture-dependent techniques and culture-independent swabs in investigations of CRS pathogen [59]. Swabs
molecular techniques have broadened our view of CF should not be contaminated by the microorganism of the
airway bacterial communities [38]. Each CF patient pre- nares during insertion/retraction from the middle meatus
sented a unique microbiome [40]. The species present or sinuses [56]. To avoid contamination by the nasal
tended to vary more “between” than “within” subjects, vestibule, researchers often use appropriate protective
Table1Summaryofcysticfibrosismicrobiotastudies;typeofsample,techniqueusedandgenusidentified C
h
a
Author Year n Sample Techinque Achromobacter Actinomyces Atopobium Bacteroidetes Burkholderia le
rm
SalipanteSJ* 2013 60CF Sputum 16srRNApyrosequencing w
a
ZemanickET 2013 37CF Sputum Conventionalculture16SrRNA x x tan
a
pyrosequencing ch
a
i
DelhaesL 2012 8CF Sputum Conventionalculture16SrRNA e
t
pyrosequencing a
l.
W
FodorAA 2012 23CF Sputum 16SrRNApyrosequencing x o
rld
StressmannFA** 2012 14CF Sputum ConventionalcultureanalysisT-RFLP A
lle
ZhaoJ 2012 6CF Sputum 16srRNApyrosequencing rg
y
O
GussAM 2011 4CF Sputum 16srRNApyrosequencing x x rg
a
n
SibleyCD 2011 6CF Sputum ConventionalcultureT-RFLP16SrRNA x x x x iz
a
pyrosequencing tio
n
CoxMJ*** 2010 63CF SputumThroatswab Microarray Jo
u
JHarrisJK**** 2007 28CF14healthy Bronchoalveolarlavagefluid DGGE/TGGE rna
l
RogersGB***** 2004 34CF Sputum T-RFLP (20
1
5
) 8
:3

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4
Table1Summaryofcysticfibrosismicrobiotastudies;typeofsample,techniqueusedandgenusidentified(Continued) C
h
a
Author Campylobacter Capnocytophaga Craurococcus Fusobacterium Granulicatella Haemophilus Lactobacillus Leptotrichia Ochrobactrum lerm
SalipanteSJ* x w
a
ta
ZemanickET x x x x x x na
c
h
DelhaesL x x a
i
e
t
FodorAA a
l.
W
StressmannFA** o
rld
ZhaoJ x A
lle
GussAM x rg
y
O
SibleyCD x rg
a
CoxMJ*** niz
a
JHarrisJK**** x tio
n
RogersGB***** x x Jou
rn
a
l
(2
0
1
5
) 8
:3

P
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4
Table1Summaryofcysticfibrosismicrobiotastudies;typeofsample,techniqueusedandgenusidentified(Continued) C
h
a
Author Peptostreptococcus Porphyromonas Prevotella Pseudomonas Rhizobium Rothia Staphylococcus Stenotrophomonas Streptococcus Veillonella Ref. lerm
SalipanteSJ* x x x [49] w
a
ta
ZemanickET x x x x x x x [35] na
c
h
DelhaesL x x x x x [39] a
i
e
t
FodorAA x x x x x [37] a
l.
W
StressmannFA** x x x x [44] o
rld
ZhaoJ x x x x x [41] A
lle
GussAM x x x x x [50] rg
y
O
SibleyCD x x x x x x [51] rg
a
CoxMJ*** x x [52] niz
a
JHarrisJK**** x x x [53] tio
n
RogersGB***** x x x x x [54] Jou
rn
*Stenotrophomonasmaltophilia,Streptococcusagalactiae,Haemophilusinfluenzae,Pseudomonasaeruginosa. al
**P.aeruginosa,Stenotrophomonasmaltophili’,Staphylococcusaureus,StreptococcusGroupF. (2
0
***S.aureus,P.aeruginosa. 1
5
****S.aureus,S.maltophilia,P.aeruginosa,Streptococcusmitis,H.influenza. ) 8
*****P.aeruginosa,Porphyromonasendodontalis,P.gingivalis,Prevotelladenticola,P.melaninogenica,P.nigrescens,P.veroralis,P.intermedia,P.loescheii,P.salivae,P.buccae,P.oris,Craurococcusroseus,Rhizobiumloti, :3

Ochrobactrumanthropi,Peptostreptococcusanaerobius.
Asterisksindicatestudiesinwhichmentionedspecieswereidentified.
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Chalermwatanachaietal.WorldAllergyOrganizationJournal (2015) 8:3 Page7of14
devices such as a sterilized Killian nasal speculum with detected,thosewithinthe generaPseudomonas,Citrobac-
long leaves [58]. ter, Haemophilus, Propionibacterium, Staphylococcus, and
Before the era of culture-independent methods, con- Streptococcus were found numerically dominant, with
ventionalcultureshaveimplicatedStaphylococcusaureus Pseudomonas aeruginosa being the most frequently de-
and coagulase-negative Staphylococcus as principal pa- tected species [55]. Another prospective study collected
thogens in chronic rhinosinusitis (CRS) [60]. The deve- mucosalbiopsiesfrom18patientsundergoingendoscopic
lopment of culture-independent molecular techniques sinus surgery for CRS and 9 control patients with healthy
allowedthedetectionofmorebacteria[60]andrevealed sinuses (indication: pituitary adenomas) compared swab
greater biodiversity than conventional culture [56]. culture with bTEFAP (bacterial tag-encoded FLX ampli-
Thus, the etiology of CRS may be polymicrobial [55] con pyrosequencing). Standard cultures mainly showed
and the role of anaerobe bacteria may be more promin- Staphylococcus aureus and Coagulase-negative Staphylo-
entthanpresumed;however,itislikelythatthebacteria coccus aureus, whereas the molecular analysis identified
detected by culture-dependent techniques still are of up to 20 predominant organisms per sample. Staphylo-
clinicalrelevance[60]. coccus aureus was nevertheless detected in about 50%;
Usingcomparative microbiome profiling in a cohort of moreover, they disclosed anaerobic species with so far
a small number of not further defined CRS patients and unknown impact in CRS, Diaphorobacter and Peptoni-
healthy subjects, it was proposed that the sinus micro- philus. Interestingly, Diaphorobacter is described as a
biota of CRS patients exhibit significantly reduced bac- strong biofilm creator [55,60].
terial diversity compared to those of healthy controls. Table2providesa summaryofpreviousstudiesrelated
Abreu et al., found a depletion of multiple phylogenet- to the microbiome in chronic rhinosinusitis, including
ically distinct lactic acid bacteria coincident with an in- sample size, type of sample, technique used and genus
crease in the relative abundance of a single species, found.
Corynebacterium tuberculostearicum [17]. These mi- Comparisons of molecular analyses suggest that the
crobe caused goblet cell hyperplasia and mucin hyper- detection of microorganisms by Fluorescence in-situ hy-
secretion in a murine model of sinus infection. In this bridization (FISH) and culture-dependent techniques is
model, Lactobacillus sakei represented a potentially related to the abundance of an organism, furthermore,
protective species [17]. However, the finding of this single cultivation tends to give advantage to rapidly growing
species has not been confirmed by others [55,56] (see bacteria [18]. The investigators employed conventional
Table2).Inalargerstudy,StaphylococcusaureusandPro- cultivation, molecular diagnostics and FISH to detect
pionibacterium acnes were the most common organisms Staphylococcus aureus as a standard. They found that
in CRS (mostly CRSwNP) and controls, respectively [18]. FISH analysis had a sensitivity of 78% with a specificity
Recently,theinvestigatorsdetectedStaphylococcusaureus, of 93% compared to the molecular technique [18]. Evi-
Staphylococcus epidermidis and Propionibacterium acnes dence from high-sensitivity techniques demonstrates that
as the most prevalent and abundant microorganisms in thehealthysinusisclearlynotsterile[18],butshowshigh
healthysinuses[61]. diversityoftheresidentmicrobiota[17].Thenasalmicro-
Using culture-independent (qPCR and 16S rRNA gene biotaofhealthysubjectsmainlyconsistofmembersofthe
sequencing) methodologies for pathogen identification phylum Actinobacteria (e.g., Propionibacterium spp. and
in chronic rhinosinusitis patients, among 57,407 pyrose- Corynebacteriumspp.),whereasthephylaFirmicutes(e.g.,
quences were generated. The most prevalent ones were Staphylococcus spp.) and Proteobacteria (e.g. Enterobacter
from coagulase-negative staphylococci (100%), 21/21 spe- spp) are less frequent [55,56,60,63]. It appears that the
cimens, Corynebacterium spp (not specifically Coryne- prevalence and abundance of organisms is critical in de-
bacterium tuberculostearicum) (85.7%) 18/21, P. acnes termininghealthyconditions[18].
(76.2%), 16/21, and Staphylococcus aureus (66.7%) 14/21. Thus, similar to CF, findings in CRS have pointed out
Althoughtheseauthorsfoundsignificantlydifferentdistri- that the microbiome is unique for each individual patient
butions of 16S rRNA sequences recovered from CRS vs. [42-44]andthecommunityofmicrobesisdiversified[10].
non-CRS cases, neither richness nor evenness indices As a general principle, a decreasing bacterial diversity
showed statistically significantdifferences [56].In another is correlated with disease severity in CF [37-39,42,44],
approachusing16SrRNAgeneclonesequencinginater- whereas CRS patients were characterized by an altered
minalrestrictionfragmentlengthpolymorphism(T-RFLP) microbial composition and greater abundance of Staphy-
analysis, the bacteria present in 70 clinical samples from lococcus aureus [56]. There was no single common mi-
43 CRS patients undergoing endoscopic sinus surgery crobiota profile among patients with similar clinical
were characterized; a total of 48 separate bands were de- conditions in the studies performed so far, although
tected. Species belonging to 34 genera were identified Staphylococcus aureus was prominent in most studies
as present by clone sequence analysis. Of the species [10,11,68]. Thus, there is a clear need for larger series
Table2Summaryofchronicrhinosinusitismicrobiotastudies;typeofsample,techniqueusedandgenusidentified C
h
a
Author Year n Sample Techinque Aureobacterium Alicycliphilus Burkholderia le
rm
RamakrishnanVRetal.* 2013 28healthy Middlemeatusswab qPCR16SrRNApyrosequencing w
a
AuroraRetal. 2013 30CRS12healthy Superiormiddlemeatuslavage 16SrRNApyrosequencing x tan
a
c
BoaseSetal.** 2013 38CRS6healthy MucusMiddlemeatusswab ConventionalcultureIbisT5000 ha
i
e
AbreuNAetal. 2012 10CRS10healthy Brushsamplesofmucosalsurfaces PhyloChipanalysis x t
a
ofthelateral,central,andmedial l.
W
portionsofthemaxillarysinus o
rld
FeazelLMetal.** 2012 15CRS5healthy Middlemeatusswabs Conventionalculture16SrRNA A
pyrosequences llerg
y
StressmannFAetal.*** 2011 43CRS. Polypandinferiorturbinatetissue. 16SrRNApyrosequencingT-RFLP O
Mucin(ifpresent) rga
n
FrankDNetal.**** 2010 26S.aureuscarriers Nostrilswabs 16SrRNApyrosequencing iza
16non-carriers5 tio
n
healthy Jo
u
StephensonMFetal.***** 2010 18CRS9healthy MucosalBiopsy 16srRNApyrosequencing rn
a
l
HealyDYetal.****** 2008 11CRS3healthy Mucosasamples FISHDNAprobesHaemophilusinfluenzae (2
0
StreptococcuspneumophiliaStaphylococcus 1
5
aureusPseudomonasaeruginosa ) 8
:3

SandersonARetal.******* 2006 18CRS5healthy Biopsiesofthesinusmucosa FISHDNAprobesHaemophilusinfluenzae
StreptococcuspneumophiliaStaphylococcus
aureusPseudomonasaeruginosa
LinaGetal.******** 2003 216healthy Nasalvestibuleswabs Conventionalculture
RasmussenTTetal.********* 2000 10healthy Nasalwashes ConventionalcultureCapillarysequencing x
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Table2Summaryofchronicrhinosinusitismicrobiotastudies;typeofsample,techniqueusedandgenusidentified(Continued) C
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Author Carnobacterium Citrobacter Cloacibacterium Corynebacterium Curtobacterium Cyanobacterium Diaphorobacter Enterobacter Enterococcus lerm
RamakrishnanVRetal.* x x w
a
ta
AuroraRetal. x x x x n
a
c
h
BoaseSetal.** a
i
e
t
AbreuNAetal. x x x a
l.
W
FeazelLMetal.** x o
rld
StressmannFAetal.*** x A
lle
FrankDNetal.**** x x x rg
y
O
StephensonMFetal.***** x rg
a
HealyDYetal.****** niz
a
SandersonARetal.******* tio
n
LinaGetal.******** x Jou
rn
RasmussenTTetal.********* x al
(2
0
1
5
) 8
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Table2Summaryofchronicrhinosinusitismicrobiotastudies;typeofsample,techniqueusedandgenusidentified(Continued) C
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Author Fusobacterium Haemophilus Helicobacter Lachnospira Lactobacillus Micrococcus Moraxella Mycobacterium Neisseria lerm
RamakrishnanVRetal.* x x x x w
a
ta
AuroraRetal. n
a
c
h
BoaseSetal.** a
i
e
t
AbreuNAetal. x x x x a
l.
W
FeazelLMetal.** o
rld
StressmannFAetal.*** x A
lle
FrankDNetal.**** rg
y
O
StephensonMFetal.***** rg
a
HealyDYetal.****** x niz
a
SandersonARetal.******* x tio
n
LinaGetal.******** x Jou
rn
RasmussenTTetal.********* al
(2
0
1
5
) 8
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