Potentially Harmful Secondary Metabolites Produced by Indoor Chaetomium Species on Artificially and Naturally Contaminated Building Materials

Downloaded from orbit.dtu.dk on: Oct 02, 2021

Potentially harmful secondary metabolites produced by indoor Chaetomium species on artificially and naturally contaminated building materials

Dosen, Ina; Nielsen, Kristian Fog; Clausen, Geo; Andersen, Birgitte

Published in: Indoor Air

Link to article, DOI: 10.1111/ina.12290

Publication date: 2017

Document Version Peer reviewed version

Link back to DTU Orbit

Citation (APA): Dosen, I., Nielsen, K. F., Clausen, G., & Andersen, B. (2017). Potentially harmful secondary metabolites produced by indoor Chaetomium species on artificially and naturally contaminated building materials. Indoor Air, 27(1), 34–46. https://doi.org/10.1111/ina.12290

General rights Copyright and moral rights for the publications made accessible in the public portal are retained by the authors and/or other copyright owners and it is a condition of accessing publications that users recognise and abide by the legal requirements associated with these rights.

 Users may download and print one copy of any publication from the public portal for the purpose of private study or research.  You may not further distribute the material or use it for any profit-making activity or commercial gain  You may freely distribute the URL identifying the publication in the public portal

If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim.

Accepted Article chaetoglobosin chipwood, Keywords: Gypsum, indoorplywood, cochliodone, chaetoviridine,environment, [email protected] author, E-mail: *Corresponding Nils Koppels Allé, Technical University of Denmark, DK-2800 Kgs. Lyngby, Denmark TechnicalUniversity ofDenmar 2 1 Došen Ina materials Chaetomium Potentially harmful secondary metabolites produced by indoor Article type : Original Article 10-Feb-2016 : Date Accepted : 27-Jan-2016 Date Revised : 12-Oct-2015 Date Received This article This protectedis by Allcopyright. rights reserved. 10.1111/ina.12290doi: lead to differences betweenversionthis and the Versionof Record. Please cite this article as through thebeen typesetting,copyediting, pa This article hasbeen accepted for publicationand undergone fullpeer review but has not

International Centre for Indoor Environment and Energy, Department of of andCivil Engineering,Department Energy, Centre Environment for Indoor International of Systems Plads, Eukaryotic Biotechnology, Department Sectionfor Biology, Søltofts 1 , KristianFog Nielsen species onartificiallynaturallycontaminated and building k, DK-2800 Kgs. Lyngby,Denmark 1 , Geo Clausen , Geo gination and process, which proofreading may 2 , Birgitte Andersen , Birgitte 1 * Accepted Article building materials infected bybuilding materials both onall time first weredetected Cochliodones the production. for andmetabolite growth rate along with three different chaetoviridins/chaetomugilinsup to200mg/malong inconcentrations with three indoor environment. Chaetomium Indoor The presenceThe of the fungal genus Abstract This article This protectedis by Allcopyright. rights reserved. for biomarkers providespecific also We thisgenus. indoorspeciesby within compoundsproduced twomostcommon bioactive the Chaetomium risk studyIn assessment the this of weimprove water-damaged contaminated buildings with Practical implications globosum chaetoglobosins weredetected totalin to concentrations ofup 950mg/m different six strains. Onbuildingmaterials non-indoorreference to as comparison establish differences betweenmetaboliteproduction Pure agar inliterature. the described metabolites naturally bothartificially building all the screen and infected for materials weused mass to study, this spectrometrya liquid chromatography-high approach resolution thus detected In buildings. in and screened for, group, been chaetoglobosins, have compound from described currently been have 200 metabolites on humanhealth and About wellbeing. suspected impact anegative have environments is to Chaetomium and biomarkers. biomarkers. No sterigmatocystin bywas produced by providing ofcomprehensive a picture the secondary and metabolites C. elatum spp. preferred wood-based over gypsum,materials both in of terms for their identification. The presence of the common biomarker, C. globosum C. Chaetomium C. globosum cultures were also investigated in order to in investigated were also cultures and and secondaryits metabolites in indoor in vitro Chaetomium ,

as well as well as a common biomarker for C. elatum, elatum, C. and on building well as materials spp., but only bioactive spp., but the and are thus candidates and as Chaetomium 2 from from C. globosum Chaetomium spp. from C. 2 .

Accepted Article avoided (Craesia et al., 1987).(Craesia etal.,avoided are liver in the orremoval intestines inthe limited absorption the stomach, in by hydrolysis effects are observed after inhalation in comparison been haveeven fewer tested forEvidences inhalativetoxicity. suggest themost toxic that toxicologicallyandOnly 2013). metabolites been investigated al.,et have afew fungal (Beko absorption dermal by followed being one inhalation mostimportant the buildings, mouldy in routes2011; There 2013). areexposure severalMiller, et al., of Brewer metabolites tofungal role (Rand and a have byfungi indoorcausal trichothecens) could the produced macrocyclic suggested that fungal cell wall components (e.g. components wall cellsuggestedthat fungal component(s)fungal particular and health a effect has been notyet documented.has been It fatigue (Täubel et al., 2011; 2014). McMullin, Miller and The causality between a specific experiencealso negative effects,health suchas skin headaches, rashes, dizzinessand chronic 2009).Otherwisemay(WHO Europe, healthy for Regionalmoulds Office of presence people asthmatic and allergic people, whomay experience exacerbation ofthe theirillness in especially onmanypeople, negative effect a havehealth may growthindoors Fungal Introduction building. e.g. hiddencochliodone reveal , in dustcan This article This protectedis by Allcopyright. rights reserved. In mouldridden Danish buildings of species the morphology of andgrowthcharacteristics the to Due 2014). McMullin, and Miller 2011; Miller, and (Flannigan buildings North American and are also some of the mostfrequently found gypsummoulds (55-66 % of wallboard) in associated aresamples and withchipboard, linoleumet andconcrete2011), (Andersen al.,

Chaetomium Chaetomium the fungus, the isolation the frequency is to other routes ofexposureelimination routes as other to β -glucan) and bioactive metabolites (e.g. andbioactive -glucan) metabolites can be found on about 16 % of the 16 %of about on found canbe growth in awater-damaged Accepted Article More More than 200 biological active metabolites are known different from species of buildings. the (Greento be 2012) 2003; Madsen, etal., vehicle 2006)inmouldy likely are of exposure ormicro-pa µm) fragments (0.3-1.3 submicron-size withhairdust. may together fragments peritheciuminboth the These and bepresentair upperairways et al., mycelium andspore however, the unlikely dried-up is (Nielsen 1999), 2011). With Chaetomium spread by andmites (von al.,198 insects Arxet immediately airborne, but are extruded as a mature ascospores becomeThe do not results). (unpublished plywood and gypsum board (vonArx etal., 1986), for examplein the interface linoleum between and concrete or between (Samson et al., 2010). The perithecia are produced often in the dark in cracks and cavities underestimated,presumably as the spores are produced inthe withinasci a perithecium This article This protectedis by Allcopyright. rights reserved. Udagawaal., 1978; et al., 1979), butto our knowle intravenous toxicity twometabolites ofthese been1) have detected on(Fig. 1999; (Nielsen materials building Nielsen,al., et The 2003). chaetoglobosins (Udagava et al., 1979; Thohi 2011, etWang 2016).al., Both C. globosum thespecies environment indoor in are which common of most the (www.indexfungorum.org), 2012). 400 than Chen et al., More andcochliodones chaetoglobin(Jerram Aal.,et 1975; Ge al.,et 2008;Phonkerdet 2008;al., cochliodinol, chaetomugilins, like: andcompoundsinclude these 2012) al., (Zhang et Chaetomium canbe difficult detectto using orsampling air techniques dust (Andersenetal., and and C. elatum spores being large (8-12 µm) and notreadily airborne, deposition in spores µm) large being (8-12 (McGregor et al., (McGregor etal., 2008; et Samson 2010;al., Andersen al.,et C. globosum C. nung et nung et al., 2010), and chaetoglobosins A and C Chaetomium and sticky onto perithecialmass hair and may be was (Ohtsubo et tests demonstrated inanimal 6). Because this, of 6). visibleeven growth of C. elatum C. rticles (Gorny et al., 2002;rticles (Gorny al., Kildesø et etal., dge, no study toxicity inhalative of has species have been described described been have species have beenreported to produce Chaetomium

Accepted Article infected buildings. infected naturallybuilding andmaterials from materials artificially inoculated on produced be can are Therequired. are analysis volatile of biom risks analyses, comprehensive more fo methods Chaetomium study purposeof this The to wastherefore orbuildings.materials culture,in pure butthey notdid investigate the occurrence of these metabolites onbuilding biologically active for analysed authors 2013) the detected. etal., al., 2013;McMullin papers (McMullin In metabolites et recent numerousthe identify of many couldnot detection HPLCand spectroscopy with for Oldermetabolites. literature (e.g. Nielsen al., et 1999) only lessused the sensitive UV/Vis described the many of few a these onlymethods standards,targeted reference of environment (Vishwanathindoor etal., 2009;Poliz the metabolites in variety different of used spectrometry a for been mass (LC-MS/MS) have screening me multi-analyte targeted Several basedfungi onuse (V is ofGC-MS techniques In order to identify relevant relevant toidentify In order sterigmatocystin (Rank et al., 2011; Sekita et al., 1981a). published. Other been This article This protectedis by Allcopyright. rights reserved. species are pureable toproduce in agar Chaetomium Chaetomium species have reported to produce the carcinogenic been Chaetomium produced compounds for produced fu arkers (MVOCs) produced by relevant indoor arkersrelevant by (MVOCs) produced thods basedon chromatographyliquid tandem an al., Lankeret 2008;Polizzietal., 2012). map all secondary mapall metabolites different r their detection and possible quantification detection and r their ziet al., 2009). However, duetothe lack metabolites from indoor environments environments indoor from metabolites cultures anddetermine cultures whichofthose ture exposures/health Chaetomium

Accepted Article at -20°C. used for external calibration and matrix effect ev Liestal, CH), chaetoglobosin A (Enzo,Exeter bacterial origin.stock5 %of Standard solutions and of chaetoviridin A (AdipogenAG, origin fungal being of them 95% of with consistscollection of approximately 1500standards (Kildgård 2014; 2011). al., projects etal., metabolite in-house Nielsen This different et either sources, gi from as commercial years, collected over past inlibrary secondary included been 30 the of have metabolites Standards MA) Bedford, and autoclaved.(Millipore, if notstatedGermany), otherwise.used Water chemicals were analyticalgrade purchased Sigma-Aldrich andwere from (Steinheim, grade; LC-MS were analysis, LC-MS waterfor including study, this in used All solvents and standards Chemicals methods and Materials This article This protectedis by Allcopyright. rights reserved. Chaetomium used: were strains reference inoculation artificial and profiling Univ Technical Systems Biology, of Department All 23 Fungal andstrains cultivation C. homopilatum purposes,comparison nineadditional elatum (IBT 41944, indoor)and (IBT41944, Chaetomium strains isolated from indoor samples were also inoculated on agar plates inoculatedagarplates for samples werealso on isolatedstrains indoor from , two strains used in this study were from the IBT Culture Collection at the at Culture Collection IBT study the inthis strains werefrom used C. longicolleum C. globosum C. virescens on building material, two indoor andtwo two indoor on building material, Chaetomium and one and (IBT 7029, reference; and IBT 41801, indoor), indoor), reference; and IBT41801, (IBT7029, fts from other research groups or purified from research groupsfts other orpurified from (IBT 26237, reference). For metabolite profile metabolite For reference). (IBT26237, , UK) andchaetoglobosin C , (Sigma-Aldrich), in fungal work was purified on Milli-Q systemin fungal workwaspurified onMilli-Q aluation, were inprepared methanol andkept C. malaysiense ersity of Denmark (Table 1). For metabolite For metabolite 1). (Table Denmark of ersity strains were used: used: four were strains (Table 1). Furthermore, ten Furthermore, 1). (Table C. globosum Chaetomium , two C.

Accepted Article suspensions of four four suspensions of (drywall/ weregypsum plywood, types ofused: materials chipboard, board different building Five materials building inoculated Artificially (2014). wasperformed using(1963) Amesspecies level incubated in darkness 25 °Cat for 2 weeks. Theidentification of fungal strains and isolates to et(Samson, Agar(PDA) Dextrose Potato Sucrose(YES), agarExtract agar Oatmeal Malt (OAT), Agar (MEA), agar juice (V8) V8 and Collection.Culture Five different agar media were used for inoculation: Yeast Extract IBT our in identification species consequently included profiling, were and metabolite This article This protectedis by Allcopyright. rights reserved. blocks wereweighedblocks with one individually streak-inoculated and the streakwere used of5 inoculation for total). (20samples materials Boxes with in sample using40sterilised 1x kGy sample Each block was inplaced aplastic box (22.5×17.5×4.5 cm) with lid, a sealed and DE). Düsseldorf, (Henkel, Direct ControlPattex Universal giving treatment), inoculation four (no surface +nonwoven adhesive wallpaper w wall-covering adhesive, 3)wallpaper (wp1),2)only wallpaper adhesivewallpaper woodchip +nonwoven section re 2).Each cm)(Fig. (13×4.5 sections oblongequal divided in four block eachfurther and blocks was sample (13×18cm) into pa unblemished New, (MDF). density fibreboard autoclaved, double-distilled water autoclaved, water double-distilled was to added box.blocks The each were left to absorb of vertical inthe lines each middle the indivi

plasterboard), masonite (hardboard/high- masonite plasterboard), Chaetomium strains (see 1)prepared Table 10-day-old V8from cultures γ -irradiation (Sterigenics, Espergærde, Denmark). Spore (Sterigenics, -irradiation dual segment (Fig.1). Afdual segment (Fig.1). al. 2010). Onceal. inoculated,were 2010). strains all presented one of four different conditions:1) different presented oneof four sites per block. Wallpaper adhesive used was block. used Wallpaper sitesadhesive per ith pattern (wp2) and 4) non-treated material and 4)non-treated pattern (wp2) ith nels (90×240 cm) of each material were cut were material of each nels (90×240cm) , von Arx et al. (1986) and Wang et al. density fibreboard (HDF)) and medium (HDF)) fibreboard density Chaetomium ter inoculation 100 mLter inoculation100 strain along strain Accepted Article samples arrived at the lab or when discovered in the water-damaged building. water-damaged the in discovered when or lab the at arrived samples the when immediately, either extraction was performed metabolite for Sampling profiling. methods (see“Fungalstrains morphological contaminants the from naturally contaminated sampleswas performed by classic 2010).of tospecies al., Identification the (Samsonet Shear’s mounting level fluid surfacetothe infected tapeon amicroscope drop ofadhesive mounting it slidein a and This directly from taken was donethe area. bymould-infected gently transparentpressing pha preparationsfor tape materials, ten the gypsum a board, ceiling In and cardboard). tile order to verify collected shelf, were (chipboard water-damaged Four buildings from asamples indoor extraction wasperformed 6months after first fungal growthwasobserved. was was and After it produced. determined(macro- microscopically) the that fungal contaminant 5weekand incubated room for at temperature newresearch projects. The cleanmaterials ha gypsum(chipboard, (3), samples board material Ten samples, naturally contaminated with buildingmaterials contaminated Naturally not analysed due tothe absenceof growth. for inoculation oncerecorded aweek. metabolite wasSampling for extraction done weeks four after at temperatureincubated room in The darkness. were blocks inspected and growth progress and re-weighed were boxes the was removed; water excess any which 24 hafter for water This article This protectedis by Allcopyright. rights reserved. Chaetomium C. globosum C. spp ., the samples were donated to this project. Sampling for metabolite metabolite Sampling for project. this donatedto were samples the and C. elatum, while inoculated with material Chaetomium se contrast microscopy and (200x, 400x) were d been submerged in sterile water for 24 hours d submergedin sterilewater for been and cultivation”) and secondary metabolite andsecondary and cultivation”) concrete and OSB) other from originated s, resulting substantial in biomassfungal spp ., Chaetomium were analysed. Sixbuilding contamination on contamination C. virescens C.

was was Accepted Article sterile Q-tip or by cutting pieces (1 Q-tip orbycm sterile pieces cutting only qualitatively analysed. supernatant was directly used for chemical anal acetonitrile: mixtur MilliQ water (75:25) evaporated to dryness in a gentle stream dryness N stream of to evaporated gentle ina by sonication for 60 min was performed. The extract was transferred to cleana 2 vial,ml (75:25 v/v) mixture acetonitrile:water mL of scalpel, if scraping was not possible. Fungal biomass was placed in a 2 screwml top vial, 1 The metabolite The profiling was done on the 15-days-old MEA and PDA cultures using a micro- extraction Metabolite bi fungaleither byswabbing/scraping off Sampling from both artificially inoculated and naturally contaminated materials was done extraction Metabolite This article This protectedis by Allcopyright. rights reserved. Sant Technologies, (Agilent 1290 UHPLCsystem on anAgilent mass wasInfinity performed flight spectrometry (UHPLC-DAD-QTOFMS) chromatogra liquid performance Ultra-high analyses UHPLC-DAD-QTOFMS “Metabolitebuilding extraction materials”. of were Theextracts 60min. for by sonication plugs the were extracted vial and added wasto each acid, 1%formic containing vol/vol/vol) 1.0top vial. ml of extraction solvent, ethyl acetate/dichloromethane/methanol (3:2:1, onecolonyagar cut across agar from plugs (6 mm were media placed ID) and in a2 ml screw for modified method scale extraction of pure cultures cultures of pure materials of building Chaetomium omass from an area of approximately 1cm an areaof fromomass 2 ) ofinfectedsurface material disposable with a e and centrifuged (15 min,15000g).The phy-diode array detection-quadrupole time of time phy-diode array detection-quadrupole with 1% formic acid was addedandextraction acid formic with1% ysis. Samples received as a bulk material were 2 , re-dissolved in400 further treated asdescribed previouslyfurther treated in a Clara, California,with aa Clara, USA) equipped metabolites (Samson, 2010). al. metabolites et Three μ L 1 % formic acidinL % formic 1 2 with a with Accepted Article [M+NH MS/HRMS) can be found in (Kildgård et al.,2014). samples be et werein (Kildgård All MS/HRMS)found analysed can onlyESI in Other MSincluding parameters, information source, usinghexakis-(2,2,3 ion electrospray was 6550 performed Agilent on QTOFMS withMS Jetan detection Stream equipped Dual et al., 2014). developed previously methodology, for allin house (Kildgårdfungal andpublished extracts arraydiode scanning in detector 200-640 range nm and 20 times/s, using standard This article This protectedis by Allcopyright. rights reserved. created by extracting Antibase2012 database for all compounds having allcompounds database for extracting Antibase2012 bycreated aggressive list al., used (Klitgårdwas search approach in the et 2014) dereplication The 2014). al.,(Kildgårdet fungal metabolites secondary 95%are which of containing 1500compounds MS/HRMS spectraThe wasdoneagainst our searching library obtained all internal of and3) UV/Vis detection of that known been butnotcompounds intheliterature described have available standardsas of searching usingdereplication HRMS datawhere the full possible of was performed lists containing all fungal secondary metabolites (~ inMS/HRMS library matching MS/HRMS data searchof 1) direct and approaches: following the of combination performed using was a of secondary metabolites Identification duemode, to the nature ofanalysed compounds. fragments included [M+H] were: fragments in function this search of(235 compounds which 10were available as reference standards). Adducts and common thewell as compoundsdescribed in source as 4 ] + . All All were analysed treatedbeing as ions . poorly ionizingcompounds. ,3-tetrafluoropropoxy)phosphazene as lock,3-tetrafluoropropoxy)phosphazene mass. on automated data-dependant MS/HRMS (Auto- automated on 1500), (Kildgård et al., 2014), 2) aggressive literature and notincluded inAntibase2012 single charged; thearea charged; single cut-off to wasset + , [M+Na] Chaetomium Chaetomium + , [M+H-H 2 O] spp. as a + and +

Accepted Article by poor water absorption ability (5 % w/w) by the material (Table 2) providing unfavourable unfavourable 2) providing (Table the material by (5 %w/w) ability absorption water by poor beexplained this the caseof inoculation. MDF, could In sections, months after several even that noneof microscopyrevealed and strains the four by any of masonite MDF or weeksoneither four after wasnovisible growth There globosum wallpaper. with sections to adhesive compared faster and denser in non-treated sections (no su wood-based materials supported highestthe biomass.amount of Visually, growth was rate production peritheciaproduction on of plywood andchipboard resultingwas rapid, in densely overgrown strains of MDF,is pr masonite and board, chipboard, different materials, gypsum plywood, of evaluationfungal growthon five The Fungal growth on artificia Results and discussion selecting time the most ion.abundant 2015). The method for screening included all QuantitativeMasHunter QTOF(versionB.06.00)for Analysis was used (Nielsen andLarsen, semi-quantification purposes,samples andfor of largerscreening number of For fast overloadingthe detector (Kildgård et al.,2014). avoid to in order height peak the of belowrecorded 10% was spectrum mass the 10,000, and This article This protectedis by Allcopyright. rights reserved. and gypsum-based materials proved to be good substrates for material after weeks,four while growth on gypsum was visually less dense. Both wood-based on chipboard, butless densegrowth on plywood gypsum. and C. globosum C. was observed two after weeks. Once started, the growth and lly inoculated esented inTable esented 2. First sporadichyphal growth of both materials Chaetomium C. elatum rface treatment)in sections and with treated the spores had germinated in any of four any four in the sporeshadgerminated of showed similar growth pattern to growth pattern similar showed compoundswithretention known Chaetomium growth, butthe growth, C. Accepted Article Metabolite production 2014). Moon, and (Ryu growth and germination mould limiting in advantageous be to structures to that, arecentaddition paper reports research wallpapers with flatter or irregular surface in betweenpolystyrene patternnever lines surfaces on the (Fig.and itself 2). In pattern regardlesspattern), of inoculated strain orspecies, with the growth appearing only on polystyrene, a with covering wall WP2 (non-woven observed on was inhomogeneous growth WP1, whileWP2 showed to be poorest in wallpapers Comparing type 1 andtype(WP1) suggestingthat its habitat natural very isdifferent indoorenvironments.from strain, reference The substrate for for conditions This article This protectedis by Allcopyright. rights reserved. C, (B, F D, G, chaetoglobosins or V) (III III or and prochaetoglobosins Analysis of the Chaetoglobosins chaetoglobosin A(Fig. 1). andchaetoviridins andchaetomugilins,groups: cochliodones chaetoglobosins, chemical All detectedmetabolites, regardlessmaterials. of Table 3, is presented tablesin while 4and5sh metaboliteproduction of The previously by inpurecultures Thohinung et Chaetomium Chaetomium C. elatum C. virescens, virescens, C. , despite the good absorption ability (17 %w/w) the of material. extracts from inoculated materials didnot detect ofthe any growth. On the other hand, masonite showedalso tobe poor C. elatum did not grow on any type of material thisstudy,did notgrowusedon any in type material of and C. globosum al. (2010). al. (2010). otherhand,On the chaetoglobosins ow metabolites detected on naturally infected infected naturally on detected metabolites ow 2 (WP2),growth was more pronounced on sample type, belong to one of the following one of belongtype, to sample supporting the growth (Table 2). An growth supporting2). (Table the on artificially inoculated materials ed ), reported reported ), Accepted Article Four compounds with with compositionelemental (C Four compounds inoculated with building andchaetomugilins only were found on chaetoviridins materials materials, building artificially inoculated of extracts In 1). (Fig. group of specific members as identified elemental composition andhence, duetothe tobeunavailability needed of standards, same the with example compounds metabolite group of of a consisting another is This Chaetoviridins/chaetomugilins with On contaminated materials chaetoglobosinsnaturally weredetectedinall samples infected assessment. risk current notpose should compoundsabstaining these aproblem differences for suggesting any from onconducted workhasbeen toxicological group onlyvery limited a specific (Fig. Since 1). referenceavailability of standards, theonlypossibility leftwas to assign asmembers them of retention timetheir to compared the logD was toovalues speculative. Hence, without the distinguishing by compounds UV/Vis was these was observed with chaetoglobosins EandF. similar Due totheir UV chromophores, remaining peaks specificto chaetoglobosins was notpossible without standards. The same standards, unequivocally identified as chaetoglobosins AandC, while exact assigningtheof least at peaksfour identifiedall as chaetoglobosin A(Fig. 3).Twoofthose were, by use of significant differencesinMS/MS their spectra norUV MS/HRMS spectra. libraryresulted in or chaetomugilins were or were chaetomugilins detected building on any ofthe The (Table materials absence 3). of no chaetoviridins IBT 7029 sections whilst all materials, on inallfour for and chaetomugilins were detected on materials inoculated with two were detected onmaterials inoculated with This article This protectedis by Allcopyright. rights reserved. elatum C. globosum C. (Table 4 and 5). 5). 4and (Table C. globosum (with exception of cellular concrete) and in nosamples infected with indoor strain IBT 41801. This stThis 41801. IBT strain indoor

C. globosum 32 not possible, and identification based on identification possible,not and H 36 N 2 O 5 ) were detected showing neither neither showing detected were ) strains(IBT 7029and41801). rain produced rain produced chaetoviridins C. Accepted Article analysis ofanalysis artificially building inoculated material extracts of both al. 1999) as an unidentified et (Nielsen, previously was reported thisstrain inoculated with chaetoviridin Aonmaterials This article This protectedis by Allcopyright. rights reserved. matched threepeaksmatched all as cochliodonesthe A However, the presence of all proposed fragments (Supplementary information, Fig S1) in the the Fig in S1) proposed fragments information, (Supplementary ofpresence all the However, compounds, suggests NMR datathat is necessary unequivocal for identification.their knowledge chaetospirone 2005))biosynthetic(Bitzer, the of andlimited these pathways of elemental same e.g. composition, compounds withliterature (several the available explain presence of three peaks with its axialstereoce around structure explained by hemiacetal reversible rotation ring formation. Furthermore, of the biaryl- among The them. peaks eluting at9.5 11.4 and min also showed splitting, could bewhich of peaks three all showed pattern of no significantMS/MSfragmentation spectra difference Comparison stereoisomers of 2008). (PhonkerdA al., andBare cochliodones et 11.39 minutes,11.39 sharingthe same [M+H] elatum indoor beenpreviouslyhave to linked chaetoviridins andchaetomugilins detected were materials infected onall with C. globosum Cochliodones inoculated with no and Chaetoviridins were chaetomugilins also and none infected with other showed withthe threecommonconsecutive peaks 9.53 times 7.74, and of retention and C. elatum C. C. elatum (Table 3). incommon, whichseemed tohave of few metabolites a none Chaetomium Chaetomium nter easily threestereoisomers, could could which provide m/z +

strains strains UHPLC-DAD-QTOFMS or environments. m/z matching A/B. cochliodone Inconsistency in the metabolite. On naturally infected materials, spp., including ofAggressive 4). dereplication 639.2436 (Fig. t detected from any materials artificially materials any from detected t and B.Existing lite C. elatum C. C. globosum (Table 4 and 5). 4and (Table rature reports that that reports rature C. globosum C. and and C.

Accepted Article A, an azaphilone alkaloid in previously reported dimer Additional Chaetoglobin A C. elatum detectedchaetoglobin Awas byinfected ondifferent materials both globosum while gypsum, none onplywood chipboard, or but on four sections chaetoglobin Ainall available pattern in supplementary(fragmentation FigS2). information, detected onall materials infected with either all materials in all sections. Onnaturally in not support enough biomass for extraction. extraction. for biomass enough support not treatment sectionsurface (no on non-treated only sections. Ongypsum 3were in 2 chipboard 1, plywoodfour and all found and cochliodones On artificially inoculated materials, and that time first the this is our knowledge, byPhonkerd etisolated (2008) al. asoilfrom derived of strain cochliodone 2(9.53min), min)and cochliodone 3(11.39min).were The first cochliodones textthesefollowing compoundsgiven been have names provisional the 1(7.74cochliodone spectra, cochliodone strongly obtained the suggested This article This protectedis by Allcopyright. rights reserved. C. elatum. was to produceable it onplywood.also it When naturalcomes to contamination, (1of 1)(Tables 4 and5)

metabolite was found to be common for

C. elatum the cochliodones have been reported in the cochliodones reportedin been have C. globosum C. elatum produced cochliodones 1, the 3onboth produced 2 cochliodones and fected buildingfected materials, cochliodones were ) and WP1 whilst the two other sections did sections two other the WP1whilst ) and C. globosum C. globosum structure as the correct structure. In the structure. structure the correct as In or produced cochliodones 1, 2 and 3 on 2 and 1, cochliodones produced C. globosum C. globosum C. cochliodes and C. globosum C. elatum C. (Table 4 and 5). 5). 4 and (Table C. elatum C. (Ge et al., 2008) al., et (Ge . To the best of : chaetoglobin : chaetoglobin C. globosum (4 of 6) and produced C.

Accepted Article hair, while Morphologically species togetherused in classical identification morphologicalwith methods. were study collected this during samples indoor isolated from of strains profiles Metabolite Metabolite profiles of indoor and This article This protectedis by Allcopyright. rights reserved. extracts of and3-O-methyl sterigdihydrosterigmatocystin sterigmatocystin/aflatoxin path biosynthetic inposit resulted MS/HRMS library-matching tometabolites common indoorspecies not toobtain their and metabolite profiles.full C. malaysiense six none-indoor reference The but was not detected on any of the artificially inoculated building materials. was cochliodinol, which was produced by both exceptionThe byIBT 41944. inoculated building materials werefound on also cultures pure Two indoor loweraproduction incomparison to thestrains. other withinany this theHowever, strain species. S1). Table (Supplementary Thereinformation, was no metabolite exclusively by produced analysed strains in chaetoglobosins, chaetoviridinswere identified all and chaetomugilins profiles 14 chemical of all The reference of case profiles betweenmetabolite reference and indoor chemical analyses are shown Table 6.in Th C. elatum C. elatum C. C. virescens C. (1) and (1) C. globosum C. globosum has straight dichotomously branched terminal hair. Theresults from strains had identical metabolite profiles. All All in compounds profiles. produced hadidenticalmetabolite strains C. virescens C. , C. malaysiense C. strain 7029). (IBT differ from Chaetomium reference strains incultures pure reference strains C. globosum (1) were primarily screened for thepresence of C. elatum C. species, C. globosum ways 2011):sterigmatocystin, etal., (Rank and e results showed significant differences in matocystin. Sterigmatocystinmatocystin. was found in C. elatum ive hits for three important products of products important ive three for hits Chaetomium C. longicolleum C. strains were very similar. Several Several very similar. were strains byhaving unbranched,coiled terminal C. longicolleum, reference strain reference (IBT 7029) showed and and strains (Fig. 5) except in the in 5)except (Fig. strains C. globosum (2) whilst 3-O-methyl 3-O-methyl whilst , C. homopilatum C. , inpure cultures (2),

Accepted Article amount of chaetoviridins ofµg/cm amount (19 (Polizzi etal., 2009)). Same trend was seen wi and lowest on gypsum (upto10 µg/cm pointed pointed towards high (up concentration to 100µg/cm S2) Table information (Supplementary metabolites individual for estimates Concentration relatively high amounts, with the highest(up to95µg/cm total high the relativelyamount amounts, with in chaetoglobosinsmaterials onHPLC-UV/Vis found on were analyses, 1999) based TableSupplementary information, S2).As expect in ispresented metabolites are found 6 for individual materials (Results Table in of theQuantification metabolite major groups found on artificiallybuilding inoculated compounds onbuilding and bioactive materials of metabolites Quantification environment can be onlylinked to 15 with indoorcommon the Thisshows strains. that presence of sterigmatocystin inindoor accurate mass. reference six spec Altogether longirosterones (S presence of the indicated 1976). two the AnalysisChen, of and S3) (Kingston Fig. information longicolleum 5). (Fig.by and retention (RT) was confirmed characteristic UVspectra time in was foundsterigmatocystin only This article This protectedis by Allcopyright. rights reserved. 5µg/cm significantly lower amounts (the highest found concentration otherchaetoglobosins wasfor in group each Other metabolites cochliodone within werepresent A 3. and A, chaetoviridin has previously been found for chaetoglobosin A (up to 13.8µg/cm 2 for C. elatum and C. malaysiense C. ) were also onnon-treated found (nosurface treatment) chipboard. A. versicolor also produced several versicolorins (Suplementary (Suplementary several versicolorins produced also C. virescens C. 2 ) and cochliodones (21 µg/cm 2 ). These). concentrations were inaccordance what with upplementary information Table S1) based on TableS1) information upplementary th two other groups of metabolites: two groupsthehighestth other of

ies any of metabolites nopresence showed and not to extracts. ed from our previoused our from study et (Nielsen al., 2 ) of three metabolites: ) ofthreemetabolites: chaetoglobosin C. globosum

The identity ofthese metabolites 2 C. homopilatum 2 for and 2 on various substrates on various ) being onchipboard C. globosum C. elatum C. virescens, C. virescens, C. . strains and and Accepted Article In cases like this, with the combination of old water damage, of absence any visible growth or indoor Hence,thesecompounds of byagar medium. other common were probably one produced the presenceshowed 3, cochliodone which of wasnot detectedthis species from growingon any onto V8agar revealed the presence of viable concrete in our experiments, proved inourexperiments, concrete notto be very susceptible to fungal growth theto due specific metabolites were notdetected. This might be explained by thetype material, of as concrete sample (Table 4), the contaminant was identified as and chaetomugilins were exclusively produced by chaetoglobosins, as all basedchaetoviridins onsecondaryperformed profile be metabolite For samples. infected in naturally identification contaminant fungal in was explored profiles metabolite pure culture useof The Chaetomium material treated withadhesive compared to surfaceswithwallpapers. treatment) and (nosurface material on andhigh production blank gypsum among materials, Highdensity. productionwas metabolite found onchipboard andplywood compared to betweeninoculated artificially different chipboard). Using of these three metabolitesconcentrations to metabolite productioncompare 6.6 µg/cm This article This protectedis by Allcopyright. rights reserved. where no damage, visible water old of agood is example 5) (Table tile ceiling Thewater-damaged analyses. chemical characteristics (e.g. structurethe of terminal hair perichecia)theon the combined to be with of metabolitesabsence characteristic The for oflow concentrations thereforenutrients, biom Chaetomium 2 , 1.7 µg/cm species present in indoor environments present inindoor environments species spp. on the sampled spp. material of absence onthe despitethe their viable spores. 2 for chaetoviridin C1 and µg/cm C1 6.8chaetoviridin C2 and and for Chaetomium C. globosum, growth present. scrapingsHowever, was thetile of materials corroborated the findings on growth on findingsthe corroborated materials ass sampled might have not been sufficient. sufficient. been not have might sampled ass C. erectum identification to the species level could easily could level species to the identification C. globosum this species. In of thecellular the species. case this and subsequentanalysis and chemical C. globosum necessitates morphological 2 for cochliodone 2, all on all 2, cochliodone for , however, species Accepted Article before effects of individual metabolites are known. Until more exposure known. Untilcomprehensive individualmetabolites are of effects before However, these intoaccount. taken to be have combinedalso effects beassessed cannot microbial comp components other and of fungal should not onlymetabolites studiedindividually. be Additive and possible synergistic effects remainsand/or discomfort to bedetermined thebeen mostfrequently found indoor Theabsencespecies. of common indoor theiridentification ofal.,2009) base 2009; Polizzi et et al., Previouspapers (Vishwanath offacilitates identification indoorcontaminants. presence verychallenging.the Therefore, of fungal several of biomass viable presence and This article This protectedis by Allcopyright. rights reserved. chaetoviridin Aand 3seem cochliodone to be highest of to importance theirhighdue in exposures screened. possible Regarding Chaetomium usedindoor as canbe compounds These on different buildingmaterials.types of detected is This firsttime,the to our knowledge, that cochliodones and chaetoglobin A have been Conclusion indoor contaminants and what they ofrelevance Therefore, fungal metabolites onin buildings. thorough all studies frequent the on made no be of are conducted, effects metabolites the conclusions and their can studies Whether these from being detected. ormarkers biomarker forspecies, other however, prevents speciesother than Chaetomium biomarkers,was presence sincetheir the found in any not strains of non-indoor Chaetomium spp.on metabolites only found in metabolites play a role in building dampness-related illness dampness-related building play rolein metabolites a produce in buildings are needed. chemical biomarkers, such as thecochliodones, species, detection ofis indoorcontaminants . Certainly, effects of bioactive effects secondary. Certainly, chaetoglobosinindoor environment, A, ounds present inthe indoor wet environment C. globosum . So far, C. globosum C. globosum Chaetomium has has

Accepted Article Acknowledgements References References TechnologiesAgilent for theLeader Thought Donationof the UHPLC-QTOF system. and project, the supportof financial for FONDEN VILLUM tothank like would authors The (Täubel etal., 2011),which is relevant the for overall exposure assessment. Settled dust metabolites. is easy sample to and in terms of microbiota diversity indoor of the presence ofthe allin these settledmetabolites dust asdust seemsto accuratelyrepresent the remain healtheffects reported the inthis studyfound toxicological significance have and act can as somecausal factors for of themore of one Whether onbuilding or found materials. quantities This article This protectedis by Allcopyright. rights reserved. G., Beko, Weschler, Langer, S.,C. J., Callesen, Toftum, M., J.,and G. Clausen, (2013) Ames, L.M. (1961)Amonograph the Chaetomiaceae,of Andersen,B., Frisvad, J.C., homes anddaycare centers, ingestion, from dust toestimates compared phatalateChildren's intakes and resultant cumulative exposures estimated urine from and Development Series Environ. Microbiol., Association between fungal species and water-damaged building materials,

77 , 4180-4188 , 2 Søndergård, I.,Søndergård, Rasmussen,I. , 1-125. PLoS ONE, s to bedetermined. The nextstep would beto investigate . 8 , e62442. to airbornebybecome disruption mechanical presence of spores, and spores, micro-particles of presence inhalation and dermal absorbtion in their in absorbtion dermal and inhalation The United StatesArmy Research S., and Larsen,L.S. (2011) S., and Chaetomium

metabolites metabolites Appl. Accepted Article Brewer, J.H., Thrasher, J.D. andHooper, D. (2005)Isolerung J. Strukturaufk Bitzer, und This article This protectedis by Allcopyright. rights reserved. Green, B. J., Tovey, E. R., Sercombe, J. K., Blachere, F. M., Beezhold, D. H., and M., Boissier, Schmechel, and E., D., Robine, Reponen,K., T., Willeke, L., R. Gorny, Ge, H.M., W.Y., Zhang, Song, Saparpakorn, G., Ding, Y.C.,P., Hannongbua, andTan, S., Flannigan, B. and Miller, J.D. (2011) Microbi Wannemacher, G., C. York, L., M. III, Nealley, L. J. Jones, D., J. Thurman, A., D. Craesia, Chen, G.-D., Li, Y.-J., Gao, H., Chen, Y.,Li, X.-X., Li, J., Guo, L-D., Cen,Y.-Z., Yao,and 44 D.Schmechel, (2006) Airborne fungal andfragments allergenicity, Microbiol., as biocontaminants, (2002)Fungal fragmentsindoor air Grinshpun, S.A. Göttingen. endophytischen Pilzen durch chemisches Screening, PhD Thesis, University of and mycotoxins: is naso-sinus fungal biofilm the culprit, Chaetomium globosum R.X. (2008)Chaetoglobins A two B, and unusual alkaloids from endophytic FL, CRC Press,Ed. 2, pp 57-107. control, investigation healthBoca Raton, and Diversity, Environments. impacts, Work R.A.Flannigan, Samson J.and DavidMill Fundam. Appl. Toxicol., W. J.,R. D. andBunner,inhalationAcutein mice, T-2 toxicity L.(1987) of mycotoxin elatum X.-S. (2012)New azaphilones and phenolic chlorinated glycosides from (Supl 1) with caspase-3 withactivity, inhibitory caspase-3 , S245-255. 68 , 3522-3531. culture, culture, 8 , 230-235. Chem. Commun.,?, al growthin indoor In: B.environments. (2013) Chronic illness (2013) Chronicillness associated with mould lärung neuer Naturstoffe aus BakterienNaturstoffe lärung neuer und Planta Med, er (eds)er Microorganisms in Home and Indoor 5978-5980. 78 Toxins (Basel) , 1683-1689. , 6 Appl. Environ. Med. Mycol., Mycol., Med. , 66–80. Chaetomium Accepted Article McGregor H., Miller J.D., Rand T., Solomon J. (2008). J. Solomon Miller J.D.,Rand T., H., McGregor Madsen, A.M., Matthiesen, C.B., Frederiksen, M.W., Frederiksen, M., Frankel, M., Spilak, Klitgård, A., Iversen, A., Andersen, M.R., Larsen, T.O., Frisvad, J.C., and Nielsen, K.F. D.G.I.,Kingston, Chen, P.N. Vercellotti, and (1976)Metabolites J.R. of Kildgård, S., Månsson, M., Došen, I., Klitgård, Frisvad,A., J.C., Larsen, T.O., and Kristian Kildesø, J., Wurtz, H., Nielsen,Kruse,F., K. P., Wilkins, K., Thrane,U.,Gravesen, S., Jerram, W. A., McInnes, A. G., Maass, W. S. G., Smith, D. G., Taylor, A., and Walter, J. A. A. J. Walter, and A., Taylor, G., D. Smith, G., S. W. Maass, G., A. McInnes, A., W. Jerram, This article This protectedis by Allcopyright. rights reserved. Hygiene Association,VA, USA. of indoormold from certain moldy building materials Environ. Monit. endotoxin, ofbacteria, settlingpotential dust, fungi inflammatory indoor and M., Gunnarsen, L., Timm, M. (2012) Sampling, extraction and measurement of (2013) Agressive using dereplication screening UHPLC-DAD-QTOF: extracts up for to Drugs, marine-derivedUHPLC-DAD-QTOFMS fungiLibrary, by andaMS/HRMS of (2014) dereplication K.F. Nielsen, Accurate F. bioactive secondaryfrom metabolites 3000 fungal secondary metabolites, secondary 3000 fungal wet building material, material, building wet Nielsen, P. A., andSchneider, T. (2003) De 53 dihydroxy-6-methoxy-1-methylxanthone, versicolor (1975) The chemistry of cochliodinol, a cochliodinol, (1975) of chemistrymetabolite The of , 727-737. 12 , : 6,8-di-O-methylnidrufin, griseofulvin, dechlorogriseofulvin, and dechlorogriseofulvin, 3,8- griseofulvin, : 6,8-di-O-methylnidrufin, 3681-3705. , Eds.: Prezant B., Prezant Weekes D.M.,, Eds.: a , 14 , 3230-3239. Indoor Air, 13 Anal. Bioanal. Chem. , 148-155. , 43-51, IN: , 43-51, Phytochemistry, Phytochemistry, termination termination of fungal spore release from Recognition, evaluation and control evaluation Recognition, nd Miller nd Miller J.D., American Industrial Mold ecology: offungi Recovery Chaetomium , 15 406 , 1037-1039. , 1933-1943. spp., Can. J. Chem., J. Can. Aspergillus Aspergillus Mar. Mar. J. J. Accepted Article McMullin, D. R.,McMullin, M. Sumarah, W., Miller, Chaetoglobosins J.and (2013) D. andazaphilones Miller, J. D. and McMullin, D.D. J. R.(2014)Fung Miller, and McMullin, Nielsen, K. F., and Larsen, T. O. (2015) The (2015)The T.and Larsen, O. Nielsen, F., K. Nielsen, K. F. (2003) Mycot New(2013) J. D. andMiller, A., Blackwell, B. M. W., D. R.,Sumarah, McMullin, This article This protectedis by Allcopyright. rights reserved. Nielsen, F., K. Gravesen, S., Nielsen, P.A., Andersen, B., Thrane, U., Frisvad,and J.C. Nielsen, K. F., Månsson, M., Rank, C., Frisvad, J.C., and Larsen, T.O. (2011)Dereplication T.O. andLarsen, C., J.C., Rank, Månsson, Frisvad, F., Nielsen, K. M., Polizzi, V., Delmulle, B., Adams, A.,Picco, A.M., Rosseel, Y.,Kindt, R., Van Bocxlaer, J., Phonkerd, N., Kanokmedhakul, S., Kanokmedhakul, K., Soytong, K., Prabpai,S., and Ohtsubo, K., M., Saito, S.,Yoshihira,Sekita, K environment, produced by of strains canadian in fungalsecondary metabolite analysis, 117. contaminants:10 years on, Tetrahedron Lett., azaphilones from materials, materials, building naturally of onartificially infested mycotoxins (1999) Production and of microbial naturalof microbialproducts byLC-DAD-TOFMS, De Kimpe, N., VanPeteghem, and DeC., Chaetomium cochliodes Kongsearee, P.(2008) Bis-spiro-azaphi on mice andrats, chaetoglobosin Aanew cytochalasan bycompound produced Mycopathologia, Mycotoxin Res., Jpn. J. of Exp. Med., Med., Exp. J. of Jpn. 54 Chaetomium globosum , 568-572. oxin production by indoor moulds, byindoor moulds, production oxin VTh01 and Appl. Microbiol. Biotechnol. 145 29 , , 43-56. 47-54. Chaetomium globosum C. cochliodes 48 importance of massof spectrometricdereplication importance Front. Microbiol., al secondaryal metabolites as indoorairharmful , 105-110. ., and Natori, S. (1978) Acute toxic toxic of effects Acute S.(1978) ., andNatori, lones andazaphilones from the fungi isolated from thebuild from isolated environment, Saeger, S. (2009) JEM spotlite: Fungi, spotlite: (2009)JEM Saeger,S. CTh05, J. Nat. Prod., , 98 Fungal Genet. Biol., Tetrahedron, Tetrahedron, 6 , 9953-9966. isolated from theindoor isolated , 1-15. Chaetomium globosum 74 , 2338-2348. 64 , 9636-9645. 39 , 103-

Accepted Article Ryu, S.H., and Moon, (2014) H.J. germination on the Mould and growth wallpapersrate Varga,Nielsen, C., T.O.,Rank, K.F., Larsen, Rand, T.G. and Miller, J.D. (2011) Analysis for toxins and inflammatory compounds. In: B. Polizzi, Adams, V., A., Malysheva,S.V., DeSaeger,S.,Van Peteghem, C.,Moretti, A., This article This protectedis by Allcopyright. rights reserved. M.,Täubel, Sulyok, M., Vishwanath, V., Bloom,E., Turunen, M., Järvi, K., Kauhanen, E. andSugiyama Kurata Udagawa H., S., Y., K., Muroi S.,Yoshihira S., T., Natori Sekita Samson, R.A., Houbraken J., Thrane, U., Frisvad, J.C., Andersen, B.and (2010) 116 with different physicalwith different properties andthe surface structures, Distribution of sterigmatocystin in filamentous fungi, FL, CRC Press,Ed. 2, pp 291-306. control, investigation healthBoca Raton, and Diversity, Environments. impacts, Work R.A.Flannigan, Samson and J. DavidMillerMicroorganisms (eds)Home in and Indoor damagen indoor environments, of(2011) Co-occurence bacterial toxic f and J. Microbiol. by production Umeda M., (1981a).Mycotoxin CBSIndoor fungi. Laborat 171-179. growthfungal andchemotaxonomic classification of Picco, A.M., andDe Kimpe, N.(2012) volatile markersIdentification of for indoor buildings, damaged microbialmycotoxins and volatile organic compounds inmoldy water from interiors , 941-953. , 27 , 766-772. J. Environ. Monit., ory Manualory Series 2., Indoor Air, 11 J., Samson, R.A., and Frisvad, J.C. R.A.,and Frisvad, Samson, J., (2011) , 1849-1858.

21 ungal secondary metabolites in moisture- , 368-375. Chaetomium Utrecht: CBS. Aspergillus Fungal Biol., spp and related fungi. Indoor Built Indoor Built Environ. species, 115 , 406-420. Fungal Biol. Food and Food Can. , 23 , , Accepted Article Van Lanker, F., Adams, Delmulle,A., DeB., Saeger, S., Moretti, A., VanPeteghem,C., and Muroi,Udagawa, S.,Kurata, T., H.,Sekita, S., Yoshihira,Natori, K., and Umeda, S., M. Thohinung, S., Kanokmedhakul, S., Kanokmedhakul, K., Kukongviriyapan, V., Tusskorn,O., This article This protectedis by Allcopyright. rights reserved. X.W., Wang Wang X.L., Liu F.J.,X.M., Zhao, Li,J.,Cai,(2014) Phylogenetic L. assessment Wang, X.W., Lombard, L., Groenewald, J.Z., Videira, S.I.R., Samson, R.A., Liu, X.Z. and Von Arx, J.A., Guarro, J. and Guarro, (1986) genusVon Figueras, J. M.J. and J.A., TheAscomycete Arx, Vishwanath, V.,Labuda, Sulyok,R., M., Bicker Zhang, Q.,Li, H.Q., Zong,S.C., Gao, J.M., and Zhang, Chemical (2012) A.L. and bioactive WHO (2009) of of complex, reassessmentthe of Phylogenetic (2016) Crous P.W. substrates, grown ondifferent bymolds produced indoor volatiles of the analysis the headspacefor SPME-GC-MS of (2008)Use N. De Kimpe, and chaetocin by of chaetoglobosins, ster production (1979) The Chaetomium elatum from (2010) the fungusand Soytong, 10-(indol-3-yl)-[13]cytochalasans K., Cytotoxic and epitypification of epitypification and 1127-1133. diversities of the genus genus the of diversities World Health Organisation. Beiheftezur Nova Hedwigia chromatography/tandem mass spectrometry, by indoormatrices liquid in metabolites 186fungal andbacterial of determination Chaetomium indicum Dampness and Mould Persoonia Chaetomium , ChE01, 36 C. funicola Chaetomium , 83-133. and allied species, with the introduction of three species, ofnew species three andallied withtheintroduction , Arch. Pharm. Res., Pharm. Arch. 84 spp. andrelated fungi, – , 1-162. and WHO Guidelines for Indoor Air Quality; secondary metabolites, C. indicum C.

, Krska,W.,, and R.(2009)Simultaneous Anal. Bioanal. Chem., igmatocystin. O-methylsterigmatocystin,

33 , Mycol. Progress,Mycol. , 1135-1141. Can. J.Microbiol., Chaetomium globosum Med. Chem. Med. J. Environ.J. Monitor.

395 13 , 719–732. , , 1355-1372. 25 12 Chaetomium , , 127-148. 170-177. species species Geneva, Geneva, , 10 , , Accepted Article Chaetomium Chaetomium Chaetomium Genus Table 1. This article This protectedis by Allcopyright. rights reserved. c b a Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Chaetomium Strains isolated from naturallyinfected materialsduringthis study. Strainsused in artificial inoculation experiment. Strains used only for metabolite profiling. onlyprofiling. Strains metabolite used for

Chaetomium

Species homopilatum homopilatum elatum elatum erectum globosum globosum virescens malaysiense longicolleum longicolleum homopilatum globosum globosum globosum globosum globosum globosum globosum globosum globosum globosum globosum globosum globosum globosum globosum

species and strains used in the study. study. the in used strains and species

IBT no. IBT 26237 41560 42179 41944 42278 41865 41578 41567 41566 41564 42279 41904 41801 41800 7029 42302 42301 42303 42300 42299 42298 42296 41766 a CBS 148.51 Stored cotton, USA Stored cotton, USA CBS 148.51 a b b b b b c c c c c c c c b b a b b c c a G3Gpu or G, DK Gypsum board Gypsum board E, DK Gypsum board D, DK Gypsum board A, DK G-3 air, Indoor DK E-1 D-2 A-1 V8 VKR 057 CS5 77 Wheat straw, India CBS 547.75 Carpet, overdoo front DK Dirt on curtain, DK Dust EK1-V8-Chae Vab VKR 081 A BAHome CS6 98 Soil, Japan Soil, Madagascar Dung, CBS USA 669.82 CBS Japan Soil, 119.57 CBS 2260 NHL = 167.61 CBS Japan scobs, Wood 103.79 2259 NHL = 337.68 CBS board, skirting DK Wooden on Dust floor, DK Plywood construction, wall Greenland Linoleum, KFV-TH-CHI-V8 068 MRO DK EK2-V8-Chae 7b GR11BA - ysmbadA, DK Gypsum board G, DK Gypsum board Gypsum board E, DK A-3 G-2 E-2 photo from albu Cardboard 3009 BASample

Other no. Other Origin r , DK m , DK Accepted Article MDF Chipboard Materials blank withno treatment (see Fig. 1). with building two materials types of (wp1wallpaper and 2) and adhesiveadhesive, alone and Table 2. This article This protectedis by Allcopyright. rights reserved. a covered; ++++: 80 - 100 %covered. Gypsum Plywood Masonite Growth evaluation: growth;NG: 5 ofno +: -20 % material covered; ++:20-50 % covered; +++: 50 -80 %

Growth evaluation of different different evaluation Growth of

blank NG NG blank NG p2 NG wp 2 deie NG adhesive p1 NG 5 wp 1 NG blank p2 NG wp 2 p1 0 + ++ + NG +++ +++ +++ 20 wp 1 p2 + + NG + ++ NG ++ adhesive +++ blank +++ wp 2 NG + + ++++ +++ NG adhesive p2 + + ++ NG +++ +++ +++ ++ 15 wp 1 blank ++++ wp 2 ++++ ++++ ++++ adhesive ++++ ++++ NG Growth NG p2 + G NG NG NG + + NG ++ + 17 ++ wp 1 17 ++ blank ++ NG NG + ++ wp 2 ++ NG adhesive wp 1 Water content Water (% w/w) (% w/w) C. globosum Chaetomium IBT 7029 7029 IBT a

+++ ++ NG ++ +++ spp.artificially ondifferent inoculated C. globosum IBT 41801 NG NG NG NG NG NG NG

IBT 41944 IBT 41944 C. elatum elatum C. NG NG NG NG NG NG NG NG

C. virescens C. IBT 26237 NG NG NG NG NG NG NG NG

Accepted Article

osin III/III osin Prochaetoglob II osin Prochaetoglob I osin Prochaetoglob n D/S Chaetomugili C/ Chaetoviridin n C Chaetomugili A/ Chaetoviridin U/V G/D/ n Chaetoglobosi n E/F Chaetoglobosi n C Chaetoglobosi n A Chaetoglobosi A Chaetoglobin 3 Cochliodone 2 Cochliodone 1 Cochliodone Metabolite with contaminated Table 3. This article This protectedis by Allcopyright. rights reserved. sections these in observed growth insufficient analysed, not NA: +/-: compound only byone produced strainof thestrains of two wallpaper adhesive;– wp2nonwoven wallcovering with pattern +wallpaper adhesive; adhesive; blank –non-treated material. wallpaper + wallpaper woodchip nonwoven wp1- material: of type each on sections Four IV osin Prochaetoglob

Chipboard Plywood Gypsum Gypsum Plywood Chipboard

ed Secondary found metabolites in buildingmaterialsartificially wetted artificially

C /- /- /- /- + - + - + - - - + + - + + + - + - + - + -+ + - +- + + - + -+ + - + - + -+ + - +- + + - + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + g ------wp 1 wp C e - - - - + - + - + -+ + - +- + - Chaetomium globosum C / /- /- /- /- /- + + + + g adhes - ive wp 2 blank wp 1 wp blank 2 wp ive C e ------C /- /- /- /- + + + + g C e - + + - - - - C /- /- /- /- /- /- /- + + + + g C e - + - + - + - + ------+- - ++ -+ - - - C /- /- /- /- + + + + g (Cg) and (Cg) C e - + - - + - - - C /- /- /- /- /- /- /- + + + + g adhes ive wp 2 blank wp 1 wp blank 2 wp ive C e - - - - C. elatum C. elatum

C. globosum C /- /- /- /- /- + + + + + g C e - + - - - + - - - C /- /- /- + + + g (Ce) (Ce) (IBT 7029 or IBT 41801); IBT 41801); or 7029 (IBT C e - - -

C / /- /- /- /- /- /- + + + + + + + g - C e ------adhesi C /- /- /- /- /- + + + + + g ve wp wp ve 2 blank C N A N A N A N A N A N A N A N A N A N A N A N A N A N A e /- /- /- /- /- /- C + + + + + + + + + + g - - - - C N A N A N A N A N A N A N A N A N A N A N A N A N A N A e /- /- /- /- /- /- C + + + + + - + + + + + g - - - - C e ------

a with contaminated Table 5. Metabolitesbold in in arefound the highest amounts. c b a Metabolitesbold in in arefound the highest amounts. of contaminated Table 4. with contaminated Eachdifferent was sample Sampled andon analyzed both face andreverse cardboard. The fungus identified usingwas tapepreparation direct AcceptedTheusing wasidentified fungus preparation tape directly Article Gypsum board 3 (7) 3 board Gypsum OSB (4) Gypsum board (3) board Gypsum Gypsum board 2 (4) 2 board Gypsum Chipboard (3) Chipboard (1) 1 board Gypsum

Chipboard (3) Chipboard Cellular concrete (4) of samples) (number type Sample This article This protectedis by Allcopyright. rights reserved. Ceiling tile (3) (5) album) (photo pages Cardboard sample) pr analyses of (number type Sample Secondary building found metabolites in water-damaged materialsnaturally Secondary found metabolites in buildingmaterialsartificiallywetted naturally chaetoglobosin A

chaetoglobosin A

Chaetomium

chaetoglobosin A chaetoglobosin A a

Chaetomium A/chaetomugilin C chaetoglobosin E/F, chaetoviridin C/chaetomugilin D/S, A/chaetomugilin C chaetoglobosin E/F, chaetoviridin C/chaetomugilin D/S, A/chaetomugilin C chaetoglobosin E/F, chaetoviridin C/chaetomugilin D/S, cochliodones 1, 3 2and cochliodones cochliodones 1, 3 2and cochliodones chaetoviridin C/chaetomugilin D/S, II, I, prochaetoglobosin prochaetoglobosin E/F, chaetoglobosin A chaetoglobin 1, 2 and 3, cochliodones Identified metabolites

chaetoviridin H C/ chaetomugilin D/S, chaetoviridin E/F, chaetoglobosin G/D/U/V, chaetoglobosin C, chaetoglobosin 2, cochliodone 1, Cochliodone Cochliodone 1, cochliodone 2, cochliodone 1, Cochliodone Cochliodone 3 2, cochliodone 1, Cochliodone Identified metabolites spp. spp. spp. , chaetoglobosin C, chaetoglobosin G/D/U/V, G/D/U/V, chaetoglobosin C, , chaetoglobosin , chaetoglobosin C, chaetoglobosin G/D/U/V, chaetoglobosin C, chaetoglobosin , , chaetoglobosin C, chaetoglobosin G/D/U/V, G/D/U/V, chaetoglobosin C, , chaetoglobosin , chaetoglobosin C, chaetoglobosin G/D/U/V, G/D/U/V, chaetoglobosin C, , chaetoglobosin , , cochliodones 1, 2 and 3, chaetoglobin A A chaetoglobin 3, 2and 1, cochliodones , , cochliodones 1, 2 and 3 and 2 1, cochliodones cochliodones 1 C. globosum

chaetoviridin A/chaetomugilin C lyon material, but notviablespecies for identification. on material, butviable not for species identification. strain, 7 strains isolated in total in isolated strains 7 strain, chaetoviridin A/chaetomugilin C , 2 and 3 cochliodone 3 cochliodone 3 cochliodone 3 , chaetoglobin A A chaetoglobin , , chaetoglobin A A chaetoglobin , , , chaetoglobin A chaetoglobin , A chaetoglobin , chaetoglobosin A, chaetoglobosin chaetoviridin chaetoviridin chaetoviridin , chaetoviridin E, E, chaetoviridin , . ,

C. globosum C. C. globosum globosum C. C. globosum C. Chaetomium C. globosum globosum C. globosum C. isolated Species C. globosum Chaetomium Chaetomium C. erectum C. elatum material on Species c sp.

sp. a

Concentration range (µg/cm CHIPBOARD Materials

Accepted Article PLYWOOD GYPSUM Table 6. Table 6. This article This protectedis by Allcopyright. rights reserved. on artificially inoculated building materials wp type 1 type wp wp type 1 type wp wp type 2 type wp 2 type wp 2 type wp wp type 1 type wp Estimation of range(µg/cm concentration Estimation adhesive adhesive adhesive blank blank blank .goou C ltm .goou C ltm .goou C ltm .goou C. elatum C. globosum C. elatum globosum C. C. elatum C. globosum elatum C. C. globosum .98.2N 15-91 N 1.9-21.04 NA 2.54 0.03-2.62 1.59-19.19 3-3.19 0.69-81.72 NA .83.2N 03-41 N 00-45 .518 N NA 0.02-14.57 NA 0.36-14.11 NA 0.55-1.880.78-36.32 NA .73.4N 07-33 N 0.21-11.74 NA 0.74-13.38 0.04-2.990.67-39.84 NA NA 0.11-0.25 0.04-4.46 NA 0.07-3.72 1.44-1.77 1.02-95.432.33-2.332.97 NA .427 A .405 N 0.71-4.01 NA NA 0.54-0.55 0.19-2.852.24-2.72 NANA 0.2-12.89 NA NA 1.26-15.261.15-62.4 NA 0.62-2.03 0.02 .328 A .427 N 0.09-4.36 NA 1.26 0.64-2.75 0.32-4.810.33-2.88 NA 0.97 .-06 A .993 N 14-.500-. N NA 1.44-7.05 NA NA 0.07-3.5 0.19-9.32 0.2-10.64 NA .425 A .938 N 03-.6N N NA NA 0.34-3.46 NA NA 0.29-3.88 2.24-2.56 NA 0.02-3.68 NA 0.47-8.49 0.86-2.651.07-6.52 NA 2.02-6.99 2.17 .226 A .- N 0.09-4.75 NA NA 0.05-1.68 NA 0.1-6 2.32-2.64 NA -.2N .-.5 A00-.5N N NA NA 0.07-6.756-9.920.2-8.85 NA NA NA Chaetoglobosins Chaetoviridins Cochliodones Cochliodones Chaetoglobin Chaetoviridins Chaetoglobosins A 2) for four major metabolite groups metabolite major found four for 2 ) Accepted Article

This article This protectedis by Allcopyright. rights reserved. Fig. 1Fig. Chemical structures of same elemental composition infound

chaetoglobosins, chaetoviridins andchaetomugilins with the

C. globosum

Accepted Article From left right: to From Fig. 2 This article This protectedis by Allcopyright. rights reserved. wallpaperadhesive, wallpaper adhesive + nonw Sections fromleft toright:Sections wallpaper +adhesive nonwoven woodchipwallpaper(wp1), Artificially inoculatedbuilding blocks with inoculationpattern specificand growthin fungal growthon pattern ofwallpaper typewallpaper two(redwith blueleaves) C. elatum and C. globosum different sectionsdifferent treated material material treated on chipboard showing characteristic absence of

oven wall-covering with pattern (wp2), non-

Accepted Article pseudomolecular ionof chaetoglobosin A, C, G,D, and6; of and m/z 531.2853 tocorresponding pseudomolecularionof chaetoglobosin E and F

This article This protectedis by Allcopyright. rights reserved. (peaks 1(peaks and MS 4). spectraof identified chaetoglobosin A (peak 5)and C 6)were(peak Fig. 3 Combined extracted ion chromatograms of 529.2697m/z correspondingto presented below

U and/or UV resulting peaks: in four 2, 3, 5

Accepted Article

This article This protectedis by Allcopyright. rights reserved. Fig. 4 Fig. cochliodone 2cochliodone (Rt9.52 min) – 3and cochliodone 3(Rt11.39min) togetherMS/MS with Chromatogramthree peaks (A)of identifiedas 1– cochliodone 1(Rt7.74 min), 2– spectrum ofspectrum cochliodone 3 20at (B) eV

Accepted Article

This article This protectedis by Allcopyright. rights reserved. globosum globosum Fig. 5 Fig. cochliodone 2, 5– chaetoglobosin6 A, – chaetoglobosin C, 7– cochliodone 3;B: 1 – cochliodone 1, 2 – chaetoglobosin G/D/U/V, 3 – chaetoglobosin G/D/U/V, 4 – Comparison of combined extracted ion chromatograms (6-12 min) of indoor ( , IBT,41801) andreference strain ( virescens 1 – 3-O-methylsterigmatocystin, 2 – sterigmatocystin sterigmatocystin 2 – – 3-O-methylsterigmatocystin, 1 C. virescens

, IBT26237) MEAextracts: A: C. C. C.