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‘Shrooms in rooms: What fungi in the built environment can
tell us about fungi and buildings Rachel Adams
UC Berkeley
Leviticus, Chapter 14: Verses 33-57 Step Verse Instruction 1 36 Evacuation 2 37 Professional inspection 3 38 Quarantine for 1 week 4 39 Re-inspection 5 40 Mold removal 6 41 Scrape affected area 7 42 Repair with new materials 8 43 If mold persists, call for new inspection 9 44 Professional inspection 10 45 If expert confirms that mold persists, demolish the home and dispose of all
materials outside the city 11 46 Quarantine anyone who was in the house 12 47 Clean clothes of anyone who was in the house 13 48 If expert finds that mold is gone, declare the house habitable again 14 49-53 Perform ritual cleaning, effect atonement for the house, and assume it is
now clean
• Damp and moldy buildings are consistently associated with negative health outcomes (Mendell et al. 2011)
• The strongest association with health effects is the perception of mold odor by the occupants (Quansah et al. 2012, Jaakkola et al. 2013)
The actual dampness-related causal agents – biological or chemical – for these health
effects are unknown
• Damp and moldy buildings are consistently associated with negative health outcomes (Mendell et al. 2011)
• The strongest association with health effects is the perception of mold odor by the occupants (Quansah et al. 2012, Jaakkola et al. 2013)
UC Berkeley’s University Village
Hypothesis: On a local scale, building characteristics, environmental conditions, and resident behaviors will
structure fungal communities.
5 hanging petri dishes • Kitchen • Living room • Bedroom • Bathroom • Outdoor (balcony)
Collection ~ 1 month, summer 2011 (n=11) and winter 2012 sampling (n=8)
Ceiling-suspended (empty) Petri dish
T and Φ Sensor Survey information on house characteristics and resident behavior
UC Berkeley’s University Village – sampling scheme
Amplicon-based approach: Pyrosequencing of the ITS region of fungal DNA to define OTUs/species
UC Berkeley’s University Village – sampling scheme
Order Number of OTUs Description Primary source 1 Agaricomycetes 238 Mushrooms, polypores Outdoor 2 Dothideomycetes 208 Molds Mixed 3 Tremellomycetes 111 Yeasts Mixed 4 Leotiomycetes 54 Plant pathogens, saprobes Outdoor 5 Eurotiomycetes 48 Molds Mixed 6 Sordariomycetes 39 Plant pathogens, saprobes Mixed 7 Microbotryomycetes 35 Yeasts Mixed 8 Saccharomycetes 19 Yeasts Mixed 9 Chytridiomycetes 9 Aquatic saprobes & parasites Outdoor
10 Wallemiomycetes 8 Molds Mixed 11 Lecanoromycetes 7 Lichenized fungi Outdoor 12 Pezizomycetes 6 Mushrooms, molds Outdoor 13 Agaricostilbomycetes 5 Yeasts Outdoor 14 Glomeromycetes 3 Plant root biotrophs Outdoor 15 Taphrinomycetes 3 Plant pathogens Outdoor 16 Orbiliomycetes 2 Saprobes Outdoor 18 Cystobasidiomycetes 1 Yeasts Outdoor 19 Ambiguous 44 20 Incertae sedis 16 21 Unclassified 130 Total 986
Fungi indoors are diverse, coming from outdoors
Adams et al., 2013a
Order Number of OTUs Description Primary source 1 Agaricomycetes 238 Mushrooms, polypores Outdoor 2 Dothideomycetes 208 Molds Mixed 3 Tremellomycetes 111 Yeasts Mixed 4 Leotiomycetes 54 Plant pathogens, saprobes Outdoor 5 Eurotiomycetes 48 Molds Mixed 6 Sordariomycetes 39 Plant pathogens, saprobes Mixed 7 Microbotryomycetes 35 Yeasts Mixed 8 Saccharomycetes 19 Yeasts Mixed 9 Chytridiomycetes 9 Aquatic saprobes & parasites Outdoor
10 Wallemiomycetes 8 Molds Mixed 11 Lecanoromycetes 7 Lichenized fungi Outdoor 12 Pezizomycetes 6 Mushrooms, molds Outdoor 13 Agaricostilbomycetes 5 Yeasts Outdoor 14 Glomeromycetes 3 Plant root biotrophs Outdoor 15 Taphrinomycetes 3 Plant pathogens Outdoor 16 Orbiliomycetes 2 Saprobes Outdoor 18 Cystobasidiomycetes 1 Yeasts Outdoor 19 Ambiguous 44 20 Incertae sedis 16 21 Unclassified 130 Total 986
Fungi indoors are diverse, coming from outdoors
Adams et al., 2013a
Surfaces – including foreheads – are collectors of airborne fungi
Culture of my forehead: 4 morphospecies
10x that are hitchhiking on foreheads
Adams et al., 2013b
Fungi indoors are not influenced by residents
Categorical Unit Room type Unit age Floor number Number of bedrooms Number of bathrooms Number of residents Frequency of cleaning
Continuous Geographic distance Temperature Relative humidity Temperature variance Relative humidity variance
Adams et al., 2013a
Fungi indoors are not influenced by residents
Categorical Unit Room type Unit age Floor number Number of bedrooms Number of bathrooms Number of residents Frequency of cleaning
Continuous Geographic distance Temperature Relative humidity Temperature variance Relative humidity variance
Adams et al., 2013a
Dispersal limitation and endemism in fungi
Release and dispersal of basidiospores from Amanita muscaria var. alba and their infiltration into a residence. Li (2005) Mycol. Res. 109 (11): 1235–1242
Dispersal limitation and endemism in fungi
30cm 5.2m ~10m
Release and dispersal of basidiospores from Amanita muscaria var. alba and their infiltration into a residence. Li (2005) Mycol. Res. 109 (11): 1235–1242
Dispersal limitation and endemism in fungi
30cm 5.2m ~10m
Release and dispersal of basidiospores from Amanita muscaria var. alba and their infiltration into a residence. Li (2005) Mycol. Res. 109 (11): 1235–1242
Day 3: 41,000 spores/m3 1,000 spores/m3 12 spores/m3
Dispersal and indoor fungi
• In these non-water damaged buildings, airborne composite fungal samples were dominated by outdoor air
• The source strength of deposition from the air to surfaces was much greater than the inverse
• The outdoor pools varied in space (hundreds of meters) and seasonally
• Surveys of the built environment support growing evidence for endemism in fungi
Dispersal and indoor fungi
• In these non-water damaged buildings, airborne composite fungal samples were dominated by outdoor air
• Do these long-term composite samples mask (interesting) short-term dynamics?
• The source strength of deposition from the air to surfaces was much greater than the inverse
• Does the same pattern hold on the short term?
Goal: Use quantity and composition of microbial communities to inform role of occupancy of airborne bioaerosols
Controlled Environment Chamber
Goal: Use quantity and composition of microbial communities to inform role of occupancy of airborne bioaerosols Treatment 0 people 1 person sitting 2 people sitting 2 people walking 8 people sitting
Flooring Carpet Sheeting
Location
Indoor Outdoor
Replicates
3 x x x
Controlled Environment Chamber
1. Extraction negatives Lab contaminant (Boletus) and possible human
contaminant (Pyrenochaeta unguis-hominis). 2. Mock community
-- Extracted DNA of 18 taxa pooled and amplified along with rest of samples. -- Zero-ing all reads with an abundance less than 10 results in community of 20 or 21 taxa, recovery of all taxa
3. Taxonomy-based filtering
Remove all taxa that do not match to “fungus”
Quality control issues with amplicon sequencing
1. Extraction negatives Lab contaminant (Boletus) and possible human
contaminant (Pyrenochaeta unguis-hominis). 2. Mock community
-- Extracted DNA of 18 taxa pooled and amplified along with rest of samples. -- Zero-ing all reads with an abundance less than 10 results in community of 20 or 21 taxa, recovery of all taxa
3. Taxonomy-based filtering
Remove all taxa that do not match to “fungus”
Quality control issues with amplicon sequencing
1. Extraction negatives Lab contaminant (Boletus) and possible human
contaminant (Pyrenochaeta unguis-hominis). 2. Mock community
-- Extracted DNA of 18 taxa pooled and amplified along with rest of samples. -- Zero-ing all reads with an abundance less than 10 results in community of 20 or 21 taxa, recovery of all taxa
3. Taxonomy-based filtering
Remove all taxa that do not match to kingdom fungi
Quality control issues with amplicon sequencing
High occupancy – modest human signature – see a signal of the flooring in the indoor air
Low occupancy – similar to outdoor air
Hypotheses: Occupancy increases shedding & resuspension rates and structures airborne bioaerosols
Expectations for microbial composition
Fungal composition shows high indoor/outdoor overlap
Factor p
Variance explained
Date 0.01 0.39
Occupancy 0.05 0.14
Time 0.01 0.08 Floor -- --
Occupancy does not affect indoor-outdoor distance
Carpet Sheet
0.65
0.70
0.75
0.80
0.85
0.90
0.95
Com
mun
ity d
ista
nce
0 1 2 2w 8
0.65
0.70
0.75
0.80
0.85
0.90
0.95
p>0.05
Factor p Variance explained
Date 0.01 0.39
Occupancy 0.05 0.14
Time 0.01 0.08
Floor -- --
Predictors of fungal composition
Indoor concentration generally tracks outdoor concentration
0 5000 10000 15000 20000 25000 30000
05000
10000
20000
Outdoor - GCN/m3
Indo
or -
GC
N/m
3
Outdoor
Indoor concentration generally tracks outdoor concentration
0 5000 10000 15000 20000 25000 30000
05000
10000
20000
Outdoor - GCN/m3
Indo
or -
GC
N/m
3
Against this variable background, difficult for us to identify differences due to occupancy
Outdoor
Carpet Indoor Outdoor Battarrea stevenii 0 160217 222 Trametes versicolor 140 47085 46720 UC1859899 voucher 34 31633 22716 Aureobasidium pullulans 6216 25089 53291 Cladosporium cladosporioides 5993 16128 42522 Cladosporium_sp_1_GP_2013 3145 13928 100781 Cladosporium cladosporioides 1797 11353 27356 Cladosporium sp. 1097 10107 44716 YNP777 voucher 0 9841 0 Rhodotorula mucilaginosa 1931 8689 4339 Pisolithus 73 7721 8406 Sistotremastrum 0 7685 7959 Cortinarius 0 7622 5000 Fungi 2428 6865 19330 Penicillium_sp_RM1_8 57 6852 11947 Cladosporium 1031 6528 30887 Cladosporium cladosporioides 1877 6300 22418 Cladosporium dominicanum 2113 6143 16402 Cryptococcus albidosimilis 510 5894 3753 Fungal_endophyte_sp_AP053 2371 5719 25075
Carpet Indoor Outdoor Battarrea stevenii 0 160217 222 Trametes versicolor 140 47085 46720 UC1859899 voucher 34 31633 22716 Aureobasidium pullulans 6216 25089 53291 Cladosporium cladosporioides 5993 16128 42522 Cladosporium_sp_1_GP_2013 3145 13928 100781 Cladosporium cladosporioides 1797 11353 27356 Cladosporium sp. 1097 10107 44716 YNP777 voucher (Yosemite) 0 9841 0 Rhodotorula mucilaginosa 1931 8689 4339 Pisolithus 73 7721 8406 Sistotremastrum 0 7685 7959 Cortinarius 0 7622 5000 Fungi 2428 6865 19330 Penicillium_sp_RM1_8 57 6852 11947 Cladosporium 1031 6528 30887 Cladosporium cladosporioides 1877 6300 22418 Cladosporium dominicanum 2113 6143 16402 Cryptococcus albidosimilis 510 5894 3753 Fungal_endophyte_sp_AP053 2371 5719 25075
Carpet Indoor Outdoor Battarrea stevenii 0 160217 222 Trametes versicolor 140 47085 46720 UC1859899 voucher 34 31633 22716 Aureobasidium pullulans 6216 25089 53291 Cladosporium cladosporioides 5993 16128 42522 Cladosporium_sp_1_GP_2013 3145 13928 100781 Cladosporium cladosporioides 1797 11353 27356 Cladosporium sp. 1097 10107 44716 YNP777 voucher (Yosemite) 0 9841 0 Rhodotorula mucilaginosa 1931 8689 4339 Pisolithus 73 7721 8406 Sistotremastrum 0 7685 7959 Cortinarius 0 7622 5000 Fungi 2428 6865 19330 Penicillium_sp_RM1_8 57 6852 11947 Cladosporium 1031 6528 30887 Cladosporium cladosporioides 1877 6300 22418 Cladosporium dominicanum 2113 6143 16402 Cryptococcus albidosimilis 510 5894 3753 Fungal_endophyte_sp_AP053 2371 5719 25075
Carpet Indoor Outdoor Battarrea stevenii 0 160217 222 Trametes versicolor 140 47085 46720 UC1859899 voucher 34 31633 22716 Aureobasidium pullulans 6216 25089 53291 Cladosporium cladosporioides 5993 16128 42522 Cladosporium_sp_1_GP_2013 3145 13928 100781 Cladosporium cladosporioides 1797 11353 27356 Cladosporium sp. 1097 10107 44716 YNP777 voucher 0 9841 0 Rhodotorula mucilaginosa 1931 8689 4339 Pisolithus 73 7721 8406 Sistotremastrum 0 7685 7959 Cortinarius 0 7622 5000 Fungi 2428 6865 19330 Penicillium_sp_RM1_8 57 6852 11947 Cladosporium 1031 6528 30887 Cladosporium cladosporioides 1877 6300 22418 Cladosporium dominicanum 2113 6143 16402 Cryptococcus albidosimilis 510 5894 3753 Fungal_endophyte_sp_AP053 2371 5719 25075
Air samples in a mycology classroom: a unique source distorts perceived species richness
Adams et al., 2013c
Mycology classroom appears to be less rich than other classrooms but also have higher biomass
0 2000 4000 6000 8000
0200
400
600
800
1000
B
AC
D
E
Individuals
Cha
o E
stim
ated
Ric
hnes
s
A B C D E
050
100
150
200
ClassroomP
enic
illiu
m s
pore
equ
ival
ents
Adams et al., 2013c
Composi?on of non-‐mycology classrooms are similar
AB
CD
E
Proportion
Classroom
0 20 40 60 80 100
Adams et al., 2013c
Mycology classroom dominated by a few taxa
AB
CD
E
Proportion
Classroom
0 20 40 60 80 100
Adams et al., 2013c
xx PuEalls dominate mycology classroom
Pisolithus, aka dog turd fungus Ba+arrea, tall s?ltball
Lycoperdon, common puEall
Implications for amplicon-based studies
Culturing greatly undersamples the diversity that is there, but the efficiency of current culture-independent techniques can make it difficult to identify true residents Differences in biomass across samples can skew community assessments and community comparisons
Implications for studying fungi in the built environment
The far majority of what we encounter in indoor air does not originate indoors
Different in other climates? May still have impacts on human health
Interpretation of indoor air samples are greatly aided when there is a comparable outdoor sample Short-term air sampling can be idiosyncratic Effect of human occupancy/activity is largely swamped by outdoor dynamics
BIMERC: Berkeley Indoor Ecology Research Consortium
Rachel Adams John Taylor Tom Bruns
Gary Anderson Steve Lindow Ed Arens
Allen Goldstein Bill Nazaroff Seema Bhangar