Yamamoto et al., Particle size distributions and seasonal
diversity of allergenic and pathogenic fungi in outdoor air
Supplementary Figure 1 Rarefaction curves of observed fungal OTUs by particle
aerodynamic diameter (da) and season. OTUs are based on 97% similarity.
Supplementary Figure 2 Relative abundances of allergenic and infectious pathogenic fungal
species by season. The allergenic (in red) and pathogenic (underlined) species are defined as
those listed in Supplementary Tables 3 and 4, respectively.
S1
Yamamoto et al., Particle size distributions and seasonal
diversity of allergenic and pathogenic fungi in outdoor air
Supplementary Figure 3 Relative abundances of known allergenic and pathogenic species in
each major fungal genus. The abundances are averaged over all particle sizes and seasons.
The allergenic (in red) and infectious pathogenic (underlined) species are defined based on the
lists of allergenic and pathogenic fungi in Supplementary Tables 3 and 4, respectively.
S2
Yamamoto et al., Particle size distributions and seasonal
diversity of allergenic and pathogenic fungi in outdoor air
Supplemental Table 1 Summary statistics of meteorological conditions
Spring
Summer
Fall
Winter
15:30
10:45
11:00
11:00
May 13, 2009
Aug 11, 2009
Oct 15, 2009
Jan 14, 2010
End
10:30
10:30
11:00
10:30
Jun 10, 2009
Sep 8, 2009
Nov 12, 2009
Feb 11, 2010
a
Temperature (C°)
Mean
16
23
10
0
Max. of daily average
21
28
17
8
Min. of daily average
12
17
4
-9
Max. of hourly average
28
32
21
13
Min. of hourly average
5
12
-3
-13
Relative humidity (%) a Mean
72
75
74
63
Max. of daily average
95
89
92
93
Min. of daily average
47
59
48
39
Max. of hourly average
100
97
100
100
Min. of hourly average
24
37
36
24
a
Wind velocity (m/sec)
Mean
2.4
2.2
3.2
3.3
Max. of daily average
4.0
4.6
7.0
7.0
Min. of daily average
1.0
0.8
0.2
0.9
Max. of 5 second speed
14.3
11.6
16.1
24.1
Max. of 2 minute speed
10.7
9.4
12.5
17.0
a
Precipitation (mm/day) Mean
3.59
1.09
4.17
1.51
a
The original data were obtained from the National Climatic Data Center of the National Oceanic and Atmospheric Administration
(http://www.ncdc.noaa.gov/oa/ncdc.html). The data obtained at the Tweed New Haven Airport Station (41°15'50"N 72°53'12"W) were used.
Sampling period
Start
S3
Yamamoto et al., Particle size distributions and seasonal
diversity of allergenic and pathogenic fungi in outdoor air
Supplemental Table 2 Sequences of primers and probes used for qPCR assays
Assay name
Fungal species
Aaltr a
Alternaria alternata
Afumi a
Aspergillus fumigatus,
Neosartorya fischeri
Cclad2 a
Cladosporium cladosporioides, svar. 2
Enigr a
Epicoccum nigrum
PenGrp3 a
Penicillium chrysogenum / griseofulvum
/ glandicola /coprophilum /expansum
and Eupenicillium
Universal Penicillium, Aspergillus
and Paecilomyces varioti
PenAsp1mgb a
ITS1F/ITS4 b
a
b
Universal fungi
Primer (probe)
name
AaltrF1
AltrR1-1
AaltrP1
AfumiF1
AfumiR1
AfumiP1
Cclad2F1
CcladR1
CcladP1
EnigrF1
EnigrR1
EnigrP1
PchryF1
PchryR1-1
PenP2
PenAspF1
PenAspR1
PenAspP1mgb
ITS1F
ITS4
Sequence 5’-3’
Method
GGCGGGCTGGAACCTC
GCAATTACAAAAGGTTTATGTTTGTCGTA
TTACAGCCTTGCTGAATTATTCACCCTTGTCTTT
GCCCGCCGTTTCGAC
CCGTTGTTGAAAGTTTTAACTGATTAC
CCCGCCGAAGACCCCAACATG
TACAAGTGACCCCGGCTACG
CCCCGGAGGCAACAGAG
CCGGGATGTTCATAACCCTTTGTTGTCC
TTGTAGACTTCGGTCTGCTACCTCTT
TGCAACTGCAAAGGGTTTGAAT
CATGTCTTTTGAGTACCTTCGTTTCCTCGGC
CGGGCCCGCCTTAAC
GAAAGTTTTAAATAATTTATATTTTCACTCAGAGTA
CGCGCCCGCCGAAGACA
CGGAAGGATCATTACTGAGTG
GCCCGCCGAAGCAAC
CCAACCTCCCACCCGTG
Adaptor A-Key-MID-CTTGGTCATTTAGAGGAAGTAA
Adaptor B-Key-TCCTCCGCTTATTGATATGC
TaqMan
Haugland and Vesper (2002).
Larena et al. (1999); Manter and Vivanco, (2007).
S4
TaqMan
TaqMan
TaqMan
TaqMan
TaqMan
Pyrosequencing
Yamamoto et al., Particle size distributions and seasonal
diversity of allergenic and pathogenic fungi in outdoor air
Supplemental Table 3 List of allergenic fungi (Simon-Nobbe et al., 2008)
Allergenic fungi
ASCOMYCOTA
Alternaria alternata
Alternaria argyranthemi
Alternaria brassicicola
Alternaria blumeae
Alternaria brassicae
Alternaria capsici
Alternaria carotiincultae
Alternaria cetera
Alternaria cheiranthi
Alternaria cinerariae
Alternaria conjuncta
Alternaria crassa
Alternaria cucumerina
Alternaria dauci
Alternaria dumosa
Alternaria eryngii
Alternaria ethzedia
Alternaria euphorbiicola
Alternaria japonica
Alternaria limoniasperae
Alternaria longipes
Alternaria macrospora
Alternaria metachromatica
Alternaria mimicula
Alternaria mouchaccae
Alternaria oregonensis
Alternaria petroselini
Alternaria photistica
Alternaria porri
Alternaria pseudorostrata
Alternaria radicina
Alternaria solani
Alternaria smyrnii
Alternaria sonchi
Alternaria tagetica
Alternaria tenuissima
Aspergillus flavus
Aspergillus fumigatus
Aspergillus nidulans
Aspergillus niger
Aspergillus oryzae
Beauveria bassiana
Candida albicans
Candida boidinii
Cladosporium herbarum
Cladosporium cladosporioides
Curvularia lunata
Embellisia allii
Embellisia indefessa
Embellisia novae-zelandiae
Embellisia telluster
Epicoccum purpurascens
Epicoccum nigrum
Fusarium culmorum
Fusarium solani
Nimbya caricis
Penicillium brevicompactum
S5
Penicillium chrysogenum
Penicillium notatum
Penicillium citrinum
Penicillium oxalicum
Pleospora herbarum
Stemphylium botryosum
Saccharomyces cerevisiae
Stachybotrys chartarum
Stemphylium callistephi
Stemphylium vesicarium
Thermomyces lanuginosus
Trichophyton mentagrophytes
Trichophyton rubrum
Trichophyton schoenleinii
Trichophyton tonsurans
Ulocladium alternariae
Ulocladium atrum
Ulocladium botrytis
Ulocladium chartarum
Ulocladium cucurbitae
BASIDIOMYCOTA
Coprinus comatus
Malassezia furfur
Malassezia sympodialis
Psilocybe cubensis
Rhodotorula mucilaginosa
Yamamoto et al., Particle size distributions and seasonal
diversity of allergenic and pathogenic fungi in outdoor air
Supplemental Table 4 List of pathogenic fungi (Makimura, 2001)
Pathogenic fungi a
ASCOMYCOTA
Issatschenkia orientalis
Cryptococcus neoformans
Ajellomyces capsulatus
Madurella grisae
Filobasidiella neoformans
Ajellomyces dermatitidis
Microsporum canis
Schizophyllum commune
Arthroderma benhamiae
Microsporum fulvum
Trichosporon asahii
Arthroderma fulvum
Microsporum gypseum
Trichosporon cutaneum
Arthroderma gypseum
Nectria haematococca
Trichosporon inkin
Arthroderma incurvatum
Paecilomyces variotii
Trichosporon mucoides
Arthroderma otae
Paracoccidioides brasiliensis
Arthroderma vanbreuseghemii
Penicillium marneffei
ZYGOMYCOTA
Aspergillus flavus
Pichia anomala
Absidia corymbifera
Aspergillus fumigatus
Pichia guilliermondii
Cunninghamella sp.
Aspergillus niger
Pneumocystis carinii
Mucor circinelloides
Blastomyces dermatitidis
Pseudallescheria boydii
Rhizopus oryzae
Candida albicans
Scedosporium apiospermum
Candida glabrata
Sporothrix schenckii
Candida guilliermondii
Trichophyton mentagrophytes
Candida krusei
Trichophyton rubrum
Candida parapsilosis
Trichophyton verrucosum
Candida tropicalis
Trichophyton violaceum
Candida pelliculosa
Cladophialophora carrionii
BASIDIOMYCOTA
Coccidioides immitis
Malassezia furfur
Epidermophyton floccosum
Malassezia globosa
Exophiala dermatitidis
Malassezia obtusa
Fonsecaea pedrosoi
Malassezia pachydermatis
Fusarium solani
Malassezia restricta
Geotrichum candidum
Malassezia slooffiae
Histoplasma capsulatum
Malassezia sympodialis
Hortaea werneckii
Rhodotorula rubra
a The fungal species are selected from the Alphabetical List of Pathogenic Fungi ver.1.2.7. in Pathogenic
Fungi Database ver. 1.9.6.1 (http://timm.main.teikyo-u.ac.jp/pfdb/cover/alphabetical_list.html).
S6
Yamamoto et al., Particle size distributions and seasonal
diversity of allergenic and pathogenic fungi in outdoor air
Supplemental Table 5 Summary statistics of fungi ITS sequences
Season
Aerodynamic
Number of
Number of sequences determined to each taxonomic level (no. of taxa identified)
diameter, da (μm) sequences
Phylum
Class a
Order a
Genus
Species
Spring
>9.0
831
824 (2)
724 (11)
658 (34)
556 (119)
476 (145)
5.8-9.0
766
766 (2)
718 (11)
698 (29)
584 (105)
542 (144)
4.7-5.8
739
737 (2)
696 (11)
676 (28)
610 (100)
576 (132)
3.3-4.7
677
677 (2)
662 (7)
595 (21)
547 (81)
510 (102)
2.1-3.3
433
433 (2)
420 (7)
351 (15)
319 (64)
296 (80)
Summer >9.0
664
664 (2)
658 (9)
651 (20)
588 (54)
535 (68)
5.8-9.0
918
918 (2)
906 (13)
881 (34)
737 (149)
633 (186)
4.7-5.8
955
948 (2)
934 (12)
916 (32)
780 (151)
689 (203)
3.3-4.7
518
517 (2)
515 (8)
494 (22)
423 (106)
372 (137)
2.1-3.3
315
315 (2)
314 (8)
291 (17)
268 (79)
237 (99)
Fall
>9.0
875
874 (2)
840 (10)
712 (27)
661 (100)
522 (127)
5.8-9.0
749
739 (2)
703 (10)
655 (32)
587 (142)
465 (201)
4.7-5.8
501
500 (2)
485 (10)
475 (28)
439 (121)
384 (168)
3.3-4.7
777
774 (2)
762 (11)
751 (26)
729 (124)
627 (180)
2.1-3.3
311
310 (2)
309 (7)
299 (15)
294 (56)
280 (64)
Winter
>9.0
487
482 (2)
454 (10)
423 (34)
363 (101)
306 (138)
5.8-9.0
916
914 (2)
817 (10)
802 (35)
764 (130)
665 (174)
4.7-5.8
2062
2062(2)
1983 (12)
1970 (26)
1895 (68)
1593 (90)
3.3-4.7
893
893 (2)
885 (8)
866 (26)
837 (76)
742 (101)
2.1-3.3
939
938 (2)
935 (9)
897 (19)
883 (63)
830 (77)
Total
15326
15285 (2)
14720 (19)
14061 (68)
12864 (558)
11280 (1172)
a
Fungi that are categorized as incertae sedis at these taxonomic levels are excluded from counting the numbers of identified taxa.
S7
Yamamoto et al., Particle size distributions and seasonal
diversity of allergenic and pathogenic fungi in outdoor air
Supplemental Table 7 Diversity parameters for sampled atmospheric fungi based on 97%
OTU similarity
Season
Spring
Summer
Fall
Winter
Aerodynamic
diameter, da (μm)
> 9.0
5.8-9.0
4.7-5.8
3.3-4.7
2.1-3.3
> 9.0
5.8-9.0
4.7-5.8
3.3-4.7
2.1-3.3
> 9.0
5.8-9.0
4.7-5.8
3.3-4.7
2.1-3.3
> 9.0
5.8-9.0
4.7-5.8
3.3-4.7
2.1-3.3
Number of
observed OTUs
336
300
368
374
173
203
385
544
268
194
256
359
280
545
195
185
418
350
283
358
S8
Chao1
Shannon index
2387
1686
1983
1942
908
746
2482
4273
2662
2044
5157
3320
1650
3579
1010
892
1563
808
1086
1117
6.7
7.4
7.6
7.7
6.7
6.0
6.9
8.0
7.0
7.1
5.2
6.8
7.4
8.7
7.4
6.8
7.7
6.0
7.1
7.5
Yamamoto et al., Particle size distributions and seasonal
diversity of allergenic and pathogenic fungi in outdoor air
Supplementary References
Haugland R, Vesper S (2002). Method of identifying and quantifying specific fungi and
bacteria. U.S. Environmental Protection Agency: USA.
Larena I, Salazar O, Gonzalez V, Julian MC, Rubio V (1999). Design of a primer for
ribosomal DNA internal transcribed spacer with enhanced specificity for ascomycetes. J
Biotechnol 75: 187-194.
Makimura K (2001). Alphabetical List of Pathogenic Fungi Ver.1.2.7. Pathogenic Fungi
Database (PFDB) Ver. 1.9.6.1.
Manter DK, Vivanco JM (2007). Use of the ITS primers, ITS1F and ITS4, to characterize
fungal abundance and diversity in mixed-template samples by qPCR and length
heterogeneity analysis. J Microbiol Meth 71: 7-14.
Simon-Nobbe B, Denk U, Poll V, Rid R, Breitenbach M (2008). The spectrum of fungal
allergy. Int Arch Allergy Immunol 145: 58-86.
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