Download Airborne microorganisms in the African desert dust corridor over the

Aerobiologia (2006) 22:211–226
DOI 10.1007/s10453-006-9033-z
ORIGINAL PAPER
Airborne microorganisms in the African desert dust
corridor over the mid-Atlantic ridge, Ocean Drilling
Program, Leg 209
Dale W. Griffin Æ Douglas L. Westphal Æ
Michael A. Gray
Received: 9 March 2005 / Accepted: 24 May 2006 / Published online: 6 September 2006
Springer Science+Business Media B.V. 2006
Abstract The objective of this study was to
enhance our understanding of the fate and transAtlantic transport of dustborne microorganisms
from Northern Africa to the Caribbean and
Americas, and more specifically to determine if
culturable populations could be detected at a
mid-ocean site, closer to the source of dust relative to land-based Caribbean sites, during the
early summer months of May and June. Between
the dates of 22 May and 30 June 2003, daily air
samples were collected and evaluated for the
presence of culturable bacterial and fungal
colony-forming units (CFU). Here we report a
statistically significant correlation between daily
atmospheric CFU counts at a mid-ocean research
site (~15N, 45W) and daily desert dust concentrations as determined by the U.S. Navy’s Naval
Aerosol Analysis and Prediction System (NAAPS) Global Aerosol Model (Honrath et al.
(2004). Journal of Geophysical Research, 109;
Johnson et al. (2003). Global Biogeochemical
Cycles, 17, 1063; Reid et al. (2004). Geophysical
D. W. Griffin (&) Æ M. A. Gray
U.S. Geological Survey, 600, 4th St. South,
St. Petersburg, FL 33701, USA
e-mail: [email protected]
D. L. Westphal
Naval Research Laboratory, Aerosol and Radiation
Section, 7 Grace Hopper Avenue,
Stop 2, Monterey, CA 93943, USA
Research Letters, 31; Schollaert, Yoder, Westphal,
& O’Reilly (2003). Journal of Geophysical
Research, 108, 3191).
Keywords Aerobiology Æ Microbiology Æ
Atmospheric microbiology Æ Atmosphere Æ
Methods Æ Survival Æ Oceanic
1 Introduction
The phenomena known as ‘desert-dust storms’
moves an estimated 2.2 · 109 metric tons (2.2 ·
1015 grams) of soil and dried sediment through
the Earth’s atmosphere each year (Goudie &
Middleton, 2001). The largest of these events are
capable of dispersing large quantities of dust
across oceans and continents (Husar et al., 2001;
Moulin, Lambert, Dulac, & Dayan, 1997). Since a
gram of desert soil may contain as many as
1 · 109 bacterial cells (Whitman, Coleman, &
Wiebe, 1998), the presence of airborne desert
dust should correspond with increased concentrations of airborne microorganisms.
Research projects undertaken to address the
movement of microorganisms in association with
dust clouds emanating from the Gobi and Takla
Makan deserts of China, have documented elevated aerial concentrations at downwind locations
(Korea and Taiwan) when the regions are impacted by airborne desert dust (Wu, Tsai, Li,
123
212
Lung, & Su, 2004; Yeo & Kim, 2002). Research has
also demonstrated an association between African
dust events and airborne microorganisms in Africa
and the Caribbean (Griffin et al., 2003; Kellogg
et al., 2004). Using a combination of satellite
imagery and visual assessment of impaired atmospheric visibility in the U.S. Virgin Islands, elevated concentrations of bacteria and fungi above
background levels were only noted during African
dust events in late July and August of 2000 and
2001 (no association noted during events in May,
June, and early July) (Griffin et al., 2003).
This research project was undertaken to enhance our understanding of the fate and transAtlantic transport of dustborne microorganisms
from Northern Africa to the Caribbean and
Americas, and more specifically to determine if
Fig. 1 Panel A = Heavy
African desert dust
activity forms an
atmospheric bridge across
the Atlantic Ocean.
Marked research site
(~15 N, 45 W). This
26 May 2003 image is
courtesy of the SeaWiFS
Project, NASA/Goddard
Space Flight Center and
ORBIMAGE.
Panel B = NAAPS dust
optical depth for 26 May
2003. Contoured at unitless values of 0.1, 0.2, 0.4,
0.8, 1.6, and 3.2
123
Aerobiologia (2006) 22:211–226
culturable populations could be detected at a
mid-ocean site, closer to the source of dust relative to land-based Caribbean sites, during the
early summer months of May and June.
2 Methods
2.1 Research site
The drill sites for Ocean Drilling Program (ODP)
Leg 209 were located along the mid-Atlantic
Ridge (~45W) between 1443¢ N and 1544¢ N
(Fig. 1) (Keleman et al., 2004). While on station,
the Joint Oceanographic Institutions for Deep
Earth Sampling (JOIDES) vessel Resolution was
positioned with its bow to the east (into the trade
Aerobiologia (2006) 22:211–226
winds). Samples were collected from the
bridge deck located near the bow of the JOIDES
Resolution.
2.2 Atmospheric conditions
Table 1 lists daily measured atmospheric parameters including wind speed, wind direction, air
temperature, and percent relative humidity.
These parameters were measured to account for
any extreme atmospheric conditions and for
comparison with future or other studies. African
dust transport across the Atlantic was monitored
daily aboard ship using satellite imagery provided
by NASA’s SeaWiFS Project (Fig. 1a).
2.3 Air samples for isolation of culturable
bacteria and fungi
Air samples were collected from the bow of the
ship atop the right deck located in front of the
bridge (~10 m above sea level) using a portable
membrane-filtration apparatus (110 V vacuum
pump, Fisher Scientific, PVC two-place-manifold
and housing, assembled in house) between 0630
and 1845 h (local time). Presterilized filter housings containing 47-mm-diameter, 0.2-lm pore-size
cellulose acetate filter membranes were used to
collect all air samples (Fisher Scientific, Atlanta,
GA). The filter housings were removed from their
respective sterile bags and placed on an analytical
filter manifold. The housing lid was removed and
vacuum applied using a vacuum pump. Two to
three air samples (small-volume replicates, smallvolume replicates plus a large-volume sample, or
one small- and one large-volume sample, see
Table 1) were collected each day. Filtration-flow
rates ranged from 1.9 to 17.4 l min–1, and the total
volume of air filtered ranged from 89.3 l to
12,179.9 l (avg. ~2,270 l). The low end flow rates
utilized from 22 May through 30 May, and 25
June through 29 June, were due to a pump line
restriction and a decrease in pump diaphragm
efficiency respectively. Use of the two-placemanifold resulted in a normal flow rate of
8.7 l min–1 when simultaneously collecting two
samples or 17.4 l min–1 when collecting a single
sample. To control for handling contamination
(once each day), an additional filter housing was
213
removed from its bag, placed on the manifold,
and allowed to sit for 1 min without removing the
lid or turning on the vacuum. After filtration, the
housings and lids were sealed with parafilm, replaced in their respective bags, sealed with tape,
and immediately transported to the microbiology
laboratory for processing. R2A medium (Fisher
Scientific, Atlanta, GA) (Reasoner & Geldreich,
1985), was utilized in the following manner for
microbial culture. The sample filters were placed
whole on R2A medium plates, sample side up,
and were incubated in the dark at room temperature (~23C). Bacterial and fungal CFU were
enumerated at 48 h, 96 h, and between 96 h and
336 h of incubation and separated by isolation
streaking onto fresh plates of R2A. Isolated colonies were grown in Tryptic Soy Broth overnight
at room temperature on a tabletop rocker set at
low speed (Fisher Scientific, Atlanta, GA). The
following day, 0.5 ml of each culture was
transferred to a sterile cryogenic storage tube
containing 200 ll of sterile glycerol and stored at
–70C.
2.4 Evaluation of R2A medium for use
in aeromicrobiology studies
R2A medium was selected for use in the simultaneous recovery of potentially stressed (UVdamage, desiccation, temperature, etc.) airborne
bacteria and fungi due to the need for an effective
and efficient (cost and time) sampling assay. In
the UV-induced stress protocols outlined below, a
higher UV dose was selected for the fungi due to
the protective nature of their spores relative to
the bacteria (non-spore forming fraction). Airflow
rates between the fungi and bacteria studies
differed due to the number of samples simultaneously collected.
To determine if R2A could be used to effectively
recover airborne fungi (in comparison to traditional
fungal substrates), 20 sets of samples (three samples
each set) were collected in a courtyard in St.
Petersburg, Florida. Flow rates through each sample/filter were 5.8 l min–1 over a 20 min sample
time for a total volume of 116 l sample–1. Each filter
was then cut in half using sterile scissors and each
half was plated on Potato Dextrose Agar (PDA,
Becton Dickinson, Sparks, MD), Sabouraud Agar
123
123
A
B
A
B
A
B
A
B
A
B
A
B
A
B
A
B
A
B
A
B
A
B
A
B
A
B
A
B
A
B
A
B
A
B
A
B
C
828
828
743
743
746
746
751
751
729
729
727
727
737
737
723
723
716
716
721
721
736
736
718
718
734
734
746
746
754
754
729
729
749
749
743
743
1039
43
43
41
41
41
41
40
40
48
48
44
44
40
40
40
40
40
40
51
51
46
46
43
43
40
40
41
41
48
48
40
40
43
43
50
50
176
2.8
2.8
2.8
2.8
2.8
2.8
2.8
2.8
2.8
2.8
2.8
2.8
2.8
2.8
2.8
2.8
2.8
2.8
8.7
8.7
8.7
8.7
8.7
8.7
8.7
8.7
8.7
8.7
8.7
8.7
8.7
8.7
8.7
8.7
8.7
8.7
17.4
120.4
120.4
114.8
114.8
114.8
114.8
112
112
134.4
134.4
123.2
123.2
112
112
112
112
112
112
443.7
443.7
400.2
400.2
374.1
374.1
348
348
356.7
356.7
417.6
417.6
348
348
374.1
374.1
435
435
3062.4
5.8
5.8
6.4
6.4
8.2
8.2
5.6
5.6
5.1
5.1
5.3
5.3
5.1
5.1
5.6
5.6
2.2
2.2
3.1
3.1
6.7
6.7
6.1
6.1
6.1
6.1
7.8
7.8
5.4
5.4
5
5
6.2
6.2
5.7
5.7
5.7
113
113
112
112
82
82
102
102
97
97
87
87
90
90
58
58
48
48
60
60
110
110
110
110
67
67
78
78
70
70
75
75
70
70
70
70
70
27.3
27.3
26.1
26.1
25.8
25.8
25.1
25.1
25.8
25.8
25.8
25.8
25.4
25.4
25.2
25.2
25.5
25.5
24.9
24.9
25.4
25.4
26.1
26.1
25.6
25.6
25.6
25.6
25.6
25.6
25.5
25.5
26.4
26.4
25.6
25.6
25.6
70.7
70.7
76.2
76.2
70.9
70.9
81.8
81.8
68.1
68.1
75.5
75.5
73.6
73.6
77.5
77.5
74.8
74.8
75.3
75.3
69.5
69.5
65.7
65.7
68.3
68.3
68.2
68.2
68.8
68.8
79.4
79.4
73.9
73.9
71.9
71.9
71.9
0
0
0
0
0
0
2
1
1
0
1
0
1
0
0
0
0
0
0
0
1
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
3
1
1
1
0
0
1
0
0
0
1
0
0
0
0
0
0
0
0
0
0
0
0
0
2
0
0
0
0
0
2
0
0
0
0
0
0
44.7
17.8
14.9
7.4
8.1
0
17.8
0
0
0
8.9
0
0
0
2.5
0
0
0
0
0
0
0
0
0
5.7
0
0
0
0
0
0.7
8.2
5.5
4.9
47.4
110.9
76.6
65.3
25.5
6.6
5.7
8.8
8.2
3.5
2.0
2.1
3.4
3.9
3.4
0
0
0
31.3
11.2
4.1
8.9
0
4.5
0
1.3
0
0
0
0
2.9
0
0.2
Avg. total
NAAPS dust
Start
End Total time Flow rate Total
Wind Wind
Temp. % Hum. Bacteria Fungi Total
time
time (min)
(l/min)
volume (l) speed direction (C)
CFU
CFU CFU/m3 CFU/m3/day at 10 m/day
of air
(lg/m3)
(local time)
(m/s) degrees
5/22/03 745
745
5/23/03 702
702
5/24/03 705
705
5/25/03 711
711
5/26/03 641
641
5/27/03 643
643
5/28/03 657
657
5/29/03 643
643
5/30/03 636
636
5/31/03 630
630
6/1/03 650
650
6/2/03 635
635
6/3/03 654
654
6/4/03 705
705
6/5/03 706
706
6/6/03 649
649
6/7/03 706
706
6/8/03 653
653
743
ID Date
Table 1 Mid-Atlantic aeromicrobiology, atmospheric parameters and NAAPS dust data for the study period 22 May through 30 June 2004
214
Aerobiologia (2006) 22:211–226
6/25/03
6/24/03
6/23/03
6/22/03
6/21/03
6/20/03
6/19/03
6/18/03
6/17/03
6/16/03
6/15/03
6/14/03
6/13/03
6/12/03
6/11/03
6/10/03
737
737
1417
736
736
1350
734
734
733
733
741
741
1641
715
715
1705
751
1835
900
1655
739
1838
734
1845
735
1650
854
1705
725
1830
724
1710
725
1820
751
1821
739
1746
40
40
396
52
52
374
40
40
36
36
49
49
540
42
42
595
36
680
43
518
66
725
49
617
47
602
48
539
47
712
49
643
49
704
54
684
57
607
8.7
8.7
17.4
8.7
8.7
8.7
8.7
8.7
8.7
8.7
8.7
8.7
17.4
8.7
8.7
17.4
8.7
17.4
8.7
17.4
8.7
17.4
8.7
17.4
8.7
17.4
8.7
17.4
8.7
17.4
8.7
17.4
8.7
17.4
8.7
17.4
3.9
3.9
348
348
6890
452.4
452.4
6507.6
348
348
313.2
313.2
426.3
426.3
9396
365.4
365.4
10353
313.2
11518.8
374.1
8639.1
574.2
12040.5
426.3
10309.5
408.9
10065.9
417.6
8961
408.9
12179.9
426.3
11361.9
426.3
11823.3
469.8
11431.8
222.3
2367.3
5
5
5
4.5
4.5
4.5
6.2
6.2
6.7
6.7
5.8
5.8
5.8
6.3
6.3
6.3
6.1
6.1
5.4
5.4
4.7
4.7
4.9
4.9
3.9
3.9
4.8
4.8
6.7
6.7
4.9
4.9
6.6
6.6
4.6
4.6
6.5
6.5
110
110
110
76
76
76
67
67
78
78
82
82
82
95
95
95
78
78
87
87
105
105
70
70
65
65
65
65
65
65
85
85
90
90
75
75
62
62
25.3
25.3
25.3
25.9
25.9
25.9
25.9
25.9
25.2
25.2
26.2
26.2
26.2
25.6
25.6
25.6
25.8
25.8
26.2
26.2
25.7
25.7
25.9
25.9
25.9
25.9
29.6
29.6
26.1
26.1
26
26
25.9
25.9
26
26
25.8
25.8
71.4
71.4
71.4
71.4
71.4
71.4
78.3
78.3
82.8
82.8
68.8
68.8
68.8
70.9
70.9
70.9
68.1
68.1
75.4
75.4
75
75
72.9
72.9
79.6
79.6
70.7
70.7
74.6
74.6
74.3
74.3
76.4
76.4
75
75
67.4
67.4
0
0
0
0
0
1
0
0
0
3
0
0
0
0
0
0
0
2
0
0
0
7
0
0
0
0
0
0
0
0
0
0
0
0
0
1
0
0
0
0
0
0
0
5
0
1
0
0
0
0
0
0
0
0
0
11
0
11
1
7
0
6
2
7
0
3
0
1
0
2
0
0
0
0
0
0
0
0
0
0
0
0.9
0
2.8
0
9.6
0
0
0
0
0
0
0
1.1
0
1.3
1.7
1.2
0
0.6
4.9
0.7
0
0.3
0
0.1
0
0.2
0
0
0
0.1
0
0
0
0.1
0
0.1
0.1
0.2
2.8
0.3
1.5
0.7
0.6
0
0
4.8
1.4
0.3
0
6.3
7.4
7.2
5.3
14.7
20.1
32.9
23.7
21.4
15.3
11.9
5.0
5.7
15.5
17.2
7.4
5.1
657
657
741
644
644
736
654
654
657
657
652
652
741
633
633
716
715
715
817
817
633
633
648
648
648
648
806
806
638
638
633
633
636
636
657
657
636
739
A
B
C
A
B
C
A
B
A
B
A
B
C
A
B
C
A
B
A
B
A
B
A
B
A
B
A
B
A
B
A
B
A
B
A
B
A
B
6/9/03
Avg. total
NAAPS dust
Start
End Total time Flow rate Total
Wind Wind
Temp. % Hum. Bacteria Fungi Total
time (min)
(l/min)
volume (l) speed direction (C)
CFU
CFU CFU/m3 CFU/m3/day at 10 m/day
time
of air
(lg/m3)
(local time)
(m/s) degrees
ID Date
Table 1 continued
Aerobiologia (2006) 22:211–226
215
123
123
10.1
24.4
45.3
21.8
17.3
0.6
0.3
15.9
5.7
0
0
1.2
0
0.5
28.5
3.2
11.2
0.2
0
0
0
3
0
1
1
4
1
2
0
0
0
0
0
0
5
0
0
0
0
0
79.2
79.2
81.2
81.2
76.4
76.4
67.6
67.6
74.8
74.8
24.8
24.8
24.8
24.8
26
26
26.6
26.6
25.8
25.8
110
110
77
77
78
78
75
75
75
75
6.5
6.5
5.1
5.1
6.5
6.5
5.3
5.3
3.9
3.9
119.7
2403.6
144.4
2108.6
210.6
1250.2
89.3
9338.5
417.6
11623.2
1.9
3.9
1.9
3.9
3.9
1.9
1.9
17.4
8.7
17.4
63
647
76
570
54
658
47
645
48
692
831
1815
832
1646
735
1835
757
1755
726
1810
6/26/03 728
728
6/27/03 716
716
6/28/03 641
737
6/29/03 710
710
6/30/03 638
638
A
B
A
B
A
B
A
B
A
B
ID Date
Avg. total
NAAPS dust
Start
End Total time Flow rate Total
Wind Wind
Temp. % Hum. Bacteria Fungi Total
time
time (min)
(l/min)
volume speed direction (C)
CFU
CFU CFU/m3 CFU/m3/day at 10 m/day
of air
(lg/m3)
(local time)
(l)
(m/s) degrees
Aerobiologia (2006) 22:211–226
Table 1 continued
216
(SA, Becton Dickinson, Sparks, MD), or R2A.
One set of the plated filter halves were then
exposed to UV-C light for 5 min (exposure
rate = 95.5 lW cm–1), to simulate atmospheric UV
induced stress. All filters were then incubated in the
dark at room temperature (~23C) and fungal CFU
enumerated at 96 and 144 h.
To determine if R2A could be effectively used
to recover airborne bacteria in comparison to
nutrient-rich substrates, 20 sets of samples (two
samples each set) were collected in a courtyard in
St. Petersburg, Florida. Flow rates through each
sample/filter were 8.7 l min–1 over a 20 min
sample time for a total volume of 174 l sample–1.
Each filter was then cut in half using sterile scissors and each half was then plated on Blood Agar
(Sheep’s Blood Agar Base with 5% Sheep’s
Blood), and R2A, respectively. One set of the
plated filter halves were then exposed to UV-C
light for 5 min (exposure rate = 49.5 lW cm–1),
to simulate atmospheric UV induced stress. All
samples were then incubated in the dark at room
temperature (~23C) and bacterial CFU enumerated at 72 and 144 h.
2.5 Evaluation of air volume filtered
on recovery of airborne bacterial
and fungal CFU
To determine the effect of air volume assayed
when using membrane filtration to collect airborne bacteria and fungi, 22 air samples were
collected using a flow rate of 6.3 l min–1 over
sample times that ranged from 56 to 2165 min
(356.7–13,791.1 l air). To account for temporal
flux in airborne microbial concentrations small to
medium volume air samples ( < 6,000 l) were
collected at different times of the day. Large
volume air samples (>6,000 l) were collected over
a 24 h period starting and ending at approximately 0900 h. All sample filters were then plated
on R2A, incubated in the dark at room temperature (~23C) and bacterial and fungal CFU
enumerated at 96 h.
2.6 Genetic identification of bacteria
Genetic identification of bacterial and fungal isolates was performed post-cruise. The polymerase
Aerobiologia (2006) 22:211–226
chain reaction (PCR) was used to amplify a 1538base pair segment of the 16S rRNA gene using a
universal prokaryote primer set (Upstream = 5¢CCGAATTCGTCGACAA CAGAGTTTGAT
CCTGGCTCAG-3¢, Downstream = 5¢-CCCGG
GATCCAAGCTTACGGCT ACCTTGTTACG
ACTT-3¢) (Grasby et al., 20 03). Three-prime
ends of primers correspond to Escherichia coli 16S
rDNA nucleotide numbers 27 and 1492, respectively. For DNA extraction, bacterial isolates
were recovered from cryogenic storage via
streaking onto R2A followed by 48 h of incubation at room temperature. A sterile pipette tip was
then used to pick and inoculate cells into 180 ll of
lysis buffer recommended for extraction of DNA
from Gram positive bacteria in a DNeasy Tissue
kit (Qiagen Inc., Valencia, CA). The DNeasy
Tissue kit protocol was followed, and purified
DNA was eluted in 100 ll of the kit elution buffer.
Ten microliters of purified DNA were used for
PCR. The PCR master mix recipe per reaction
was: 25 ll of HotStarTaq Master Mix (Qiagen
Inc., Valencia, CA, catalog #203445), 1 ll each of
10-nM upstream and downstream primer (synthesized by Operon Technologies, Inc., Alameda,
CA), and 13 ll of HotStarTaq Master Mix H2O.
The Hot-Start PCR amplification profile was: one
step for 15 min at 95C, 40 cycles of (30 s at 94C,
30 s at 45C, 2 min at 72C), one step for 10 min at
72C and hold at 4C. PCR amplicons were
cleaned and directly sequenced using the listed
downstream primer (one strand, one reaction) by
Northwoods DNA, Inc. (Becida, MN). Sequence
lengths ranged from 441 to 751 base pairs.
GenBank Blast search (http://www.ncbi.nlm.nih.
gov/BLAST/) was used for sequence/isolate
identification.
217
REDExtract-N-Amp Plant PCR kit (SigmaAldrich Corp. St. Louis, MO) DNA-extraction
solution in a 1.5-ml microfuge tube. A Freeze/
thaw technique was then used to enhance lysis of
the fungi cells. This process was repeated
10 times using a crushed dry-ice/ethanol and
boiling-water bath. After completion of the
freeze/thaw protocol, the sample was processed
as outlined in REDExtract-N-Amp kit protocol.
The 20 ll PCR master mix recipe was: 10 ll of
REDExtract-N-Amp PCR mix, 1 ll each of
10 nM upstream and downstream primer (synthesized by Operon Technologies, Inc., Alameda, CA), 4 ll of extracted DNA, and 4 ll of
0.02-lm filter sterilized and autoclaved Milli-Q
water (Millipore, Bedford, MA). PCR reaction
conditions as listed in the REDExtract-N-Amp
Plant PCR kit protocol were followed using an
annealing temperature of 45C. Amplicons were
visualized using gel electrophoresis stained with
ethidium bromide. For efficient sequencing
reactions, amplicons were cloned into a plasmid
vector using the QIAGEN PCR Cloning Plus kit
(Qiagen, Valencia, CA). Overnight ligation of
amplicon and plasmid vector was used. PCR
plasmids were cleaned and sequenced (sequence
length ranged from 207 to 716 bp) using the
vector primer M13R (one strand, one reaction)
by Northwoods DNA, Inc. (Becida, MN). GenBank Blast search (http://www.ncbi.nlm.nih.
gov/BLAST/) was used for sequence/isolate
identification.
2.8 Genbank accession numbers
Accession numbers for the bacterial and fungal
DNA sequences included in this report are
AY857646 through AY857737.
2.7 Identification of fungi
Fungi were identified to the genus level using
PCR. Universal fungal primers (EF3 and EF4,
1556bp amplicon) were utilized for amplification
(Smit, Leeflang, Glandorg, Elsas, & Wernars,
1999). For DNA extraction, cryogenically stored
fungal isolates were streaked onto R2A followed
by 7 days of incubation at room temperature. A
sterile pipette tip was used to harvest < 3 mg of
mycelia/spores that was inoculated into 100 ll of
2.9 NAAPS model dust concentration
estimates
Daily atmospheric dust concentrations reported
as lg m–3 at 10 m above sea level were provided
by the Naval Research Laboratory (Monterey,
CA) post-cruise (see Table 1 and Fig. 1b). The
concentration values listed in Table 1 are the
averages of three daily estimates (600, 1200,
1800 h). The NAAPS model uses a first principles
123
218
Aerobiologia (2006) 22:211–226
approach to predict the distribution of mineraldust aerosol. The modeled processes include
mobilization, vertical and horizontal mixing,
advection, wet removal, and dry deposition. The
model carries a bulk-dust component with properties that are representative of dust particles
subject to long-range transport, i.e., particles with
radii less than 5 lm. The dry-deposition velocity
depends on the surface-layer turbulence, ocean
roughness, and the mean particle-sedimentation
velocity (Honrath et al., 2004; Johnson et al.,
2003; Reid et al., 2004; Schollaert, Yoder,
Westphal, & O’Reilly, 2003). Additional NAAPS
information can be found at http://www.nrlmry.
navy.mil/aerosol/Docs/globaer_model.html.
2.10 Statistical analysis
SPSS 13.0 for Windows (SPSS Inc., Chicago, IL)
was used for statistical analyses. Mid-Atlantic
airborne microbial population size and NAAPS
dust concentration data were tested for normality
using a One-Sample Kolmogorov-Smirnov Test.
(Dytham, 1999), which demonstrated that the
data were not normally distributed (N = 40,
a = 0.01, 2-tailed P < 0.001, 0.019, for microbial
and dust data, respectively). Whereas a log
transformation of the dust concentration values
normalized the distribution of the data, it did not
normalize the distribution of the values for
microbial concentration. The correlation between
the microbiology data and the NAAPS dust
concentration data was then evaluated using
Spearman’s rank-order correlation. (Dytham,
1999).
In the study to compare recovery efficiency of
the different culture media, population sizes
of fungi recovered on R2A, PDA, and SA
media (N = 20 set–1) were normally distributed
(P range = 0.088–0.881). Hence, population sizes
on R2A were compared with each of the other
substrates using an Independent t-test. For bacterial population sizes, Independent t-tests were
utilized when the data were normally distributed
and the Mann–Whitney test when they were
not. The membrane filtration air-volumeCFU-recovery data were evaluated using Pearson’s correlation a = 0.01 (2-tailed) as the data set
was normally distributed.
3 Results
3.1 Evaluation of R2A and membrane
filtration sample volumes
When R2A was compared to PDA and SA for
recovery of fungi (Table 2), analysis of the data
sets (96 h of incubation—no UV, 96 h—UV,
144 h—no UV, and 144 h—UV) demonstrated
Table 2 Evaluation of different culture media for use in cultivation of non-stressed and ultraviolet-C stressed airborne
fungi and bacteria
Fungal CFU counts
Medium
96 h incubation—
no UV treatment
R2A
9.7 (5.3)
PDA
9.6 (4.2)
SA
8.2 (3.9)
Bacterial CFU counts
Medium
72 h—no UV
R2A
4.7 (6.1)
Blood Agar
3.6 (3.1)
96 h incubation—
UV treatment
4.0 (2.8)
4.3 (2.2)
4.0 (2.3)
144 h incubation—
no UV treatment
9.9 (3.6)
10.1 (4.8)
9.5 (4.5)
144 h incubation—
UV treatment
4.8 (2.3)
4.4 (2.3)
4.6 (1.9)
72 h—UV
3.0 (1.9)*
1.5 (1.2)
144 h—no UV
5.6 (5.7)
4.8 (4.7) (N = 16)
144 h—UV
3.6 (2.2)
2.7 (1.6) (N = 19)
Values represent the mean number of colony forming units (CFU) growing on cellulose acetate filters (20 media–1) through
which outdoor air was filtered using a portable membrane filtration apparatus for 20 min at a flow rate of 5.8 l min–1 for
fungi and 8.7 l min–1 for bacteria. After sampling, half of the filters were exposed to UV light for 5 min (95.5 lW cm–1 for
fungi and 49.5lW cm–1 for bacteria) prior to plating on media
R2A = Reasoner’s 2 Agar, PDA = Potato Dextrose Agar, SA = Sabouraud Agar, (##) = Standard deviation,
* = Statistically significant difference, Mann–Whitney test, P = 0.009, UV = Ultraviolet-C light. (N = #) = number of
plates used for analysis due to the loss of the remaining plates to overgrowth of fungi
123
Aerobiologia (2006) 22:211–226
that none of the data sets were significantly
different (P range—0.581 to 1.0). When the R2A
versus Blood Agar CFU count experiment was
conducted (Table 2), analysis of the data sets
(72 h of incubation—no UV, 72 h—UV,
144 h—no UV, and 144 h—UV) demonstrated
that R2A at all time points produced higher CFU
counts and significantly higher counts at 72 h of
incubation with UV exposure (P = 0.009).
For evaluating the influence of sample volume
on membrane filtration CFU recovery rate, CFU
count data were normalized by dividing the counts
by the total liters of air sampled. Data were then
converted and expressed at CFU m–3 (Fig. 2).
Bacterial counts ranged from 1.5 to 48.9 CFU m–3
(avg. 16.4), fungal counts ranged from 15.7 to
100.9 CFU m–3 (avg. 35.0), and total counts ranged from 17.6 to 131.8 CFU m–3 (avg. 51.4). Data
analysis demonstrated significant negative correlations (microbial concentrations versus air
volume sampled) for total and bacterial CFU
counts (r = –0.578, P = 0.005, and r = –0.776,
P < 0.001, respectively), and fungal CFU counts
showed a negative correlation that was not
significant (r = –0.280, P = 0.206).
219
3.2 SeaWiFS image data
The complete SeaWiFS image data set for the
study period showed an almost continuous
transmission of dust off the northwest coast of
Africa toward the research site. Figure 1 panel A,
a 26 May 2003 SeaWiFS image, shows an atmospheric bridge of African dust spanning the
Atlantic Ocean, which coincided with the heaviest daily concentration of dust at the research site
as determined by the NAAPS Global Aerosol
Model (Fig. 3). The averaged NAAPS dust
concentrations ranged from 2.0 to 110.9 lg m–3
with an average of 18.3 lg m–3day–1.
3.3 Mid-Atlantic data
Wind direction and speed measurements for the
duration of the aeromicrobiology study ranged
from 48.0 to 113.0 relative to North (avg.
81.7) and 2.2 to 8.2 m s–1 (avg. 5.6 m s–1),
(Table 1). Relative humidity and temperature,
two factors that can influence airborne microbial survival (Choi et al., 1997; Gloster & Alexandersen, 2004; Peccia & Hernandez, 2001),
Fig. 2 Determination of
recovery efficiency of
bacterial and fungal
colony forming units
(CFU) on R2A medium
relative to air volume
sampled. Panel A, fungal
CFU. Panel B, bacterial
CFU
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220
Aerobiologia (2006) 22:211–226
Fig. 3 Mid-Atlantic
airborne dust and total
bacterial and fungalcolony-forming unit
concentrations
(normalized data for
concentrations m–3 air)
for each sample date, 22
May through 30 June 2003
averaged 73.6% (range 65.7–81.8%) and 25.8C
(range 24.8–29.6).
A total of 26 bacterial and 83 fungal CFU were
recovered on 24 of 40 sample dates (no CFU
noted on 16 of the sample dates). Total (bacterial
and fungal) concentrations ranged from 0.1–
31.3 CFU m–3. All reported CFU were recovered
between 96 and 336 h of incubation (no CFU
noted at 48 h of incubation). Of the 26 bacterial
strains isolated, 25 were recovered from cryogenic
storage (1 of the stored strains was no longer
viable) and identified by 16S rDNA sequencing,
only two were Proteobacteria (Pseudomonas sp.
and Novosphingobium subarticum. GenBank
Blast nearest neighbor. Table 3). Of the 23 Grampositive bacteria, 12 were high G+C content
(species of the genera Brevibacterium, Cellulomonas, Frigoribacterium, Gordonia, Kocuria,
Lechevalieria, Leifsonia, and Lentzea), seven
were spore-forming low G+C content (Bacillus
sp.), and the remaining four were non-sporeforming low G+C content (Staphylococcus sp.).
Of the 83 fungal strains collected, 62 were
recovered from cryogenic storage and 18S rDNA
sequences obtained. Seventeen different genera
were identified from this group of sequences
and included species of Alternaria, Aureobasidium, Cladosporium, Dendryphion, Emericella,
Lojkania, Lithothelium, Massaria, Myriangium,
Neotestudina, Penicillium, Phoma, Preussia,
Setosphaeria, Stachybotrys, Trichophyton, and
Ulocladium
(Table 3).
Statistical
analysis
123
demonstrated a statistically significant correlation
between the NAAPS dust and total microbialCFU-concentration data (rs = 0.608, P < 0.001),
i.e., higher model-predicted dust concentrations
correlated with higher CFU concentrations.
4 Discussion
In all cases R2A produced higher average counts
than those observed with Blood Agar use and
significantly higher counts when recovering UV
stressed bacteria at 72 h of incubation. Another
side benefit of R2A is that the low nutrient concentrations in this medium limited overgrowth by
fast growing fungi which allows simultaneous
recovery of fungal and bacterial CFU on a single
substrate and filter. Additionally, if utilizing spore
morphology to identify fungal isolates, low
nutrient agars enhance sporulation due to the
growing fungal colonies quickly exhausting required nutrients, a condition known to induce
sporulation. (Morton, 1961). Although the data
was not statistically significant, R2A outperformed PDA and SA in 2 of the 4 fungal CFU
recovery study parameters (96 h—no UV, and
144 h—UV) and preformed as well or better than
at least one of the substrates in the other 2 data
sets. The influence of sample volume on membrane filtration CFU recoveries demonstrated
that longer sample times (larger volumes) at an
air flow rate of 6.3 l min–3 resulted in lower
1
1
1
1
1
1
10
1
1
1
1
5
Bacteria
3
3
2
1
1
1
3
1
1
1
1
1
1
1
2
1
1
Fungi
1
# of isolates
Lithothelium septemseptatum
Massaria platani
Myriangium duriaei
Myrothecium sp., Letendraea helminthicola,
Cucurbidothis pityophila,
Paraphaeosphaeria sp. (pilleata or michotii)
98 (555/565)
Alternaria brassicae, Embellisia sp.,
Pezizomycotina sp.
Alternaria dauci
Alternaria sp.
Aureobasidium pullulans or
Discosphaerina fagi
Cladosporium sp.
Cladosporium sp. or
Trimmatostroma macowanii
Dendryphion sp.
Emericella nidulans
Lojkania enalia
96 (597/616)
99 (577/580)
99 (373/375), (568/570), (574/575),
(574/576), (594/596), 98 (613/625),
(666,674), (694/704), 698/704), (575/576)
98 (579/586)
97 (508/523)
98 (511/521)
99 (318/321)
99 (309/310)
99 (667/671)
100 (584/584), (438/438), 99 (597/600,
(671/676), 98 (695/704)
100 (436/436)
99 (617/623)
100 (353/353), 99 (633/634), 98 (704/716)
99 (640/643), (666/668), (667/668)
98 (685/696), 98 (625/633)
98 (606/615)
97 (655/669)
98 (621/629)
100 (685/685), 99 (668/669), (670/673)
100 (628/628)
97 (615/634)
98 (588/595)
97 (229/235)
95 (479/503)
99 (728/735)
98 (710/724)
100 (609/609), 99 (568/570)
100 (542/542)
100 (612/612)
% Homology (sequence length)
Bacillus aminovorans
Bacillus benzoevorans
Bacillus sp.
Brevibacterium casei
Cellulomonas terrae
Frigoribacterium sp.
Gordonia terrae
Kocuria rosea
Lechevalieria fradiae
Leifsonia xyli or L. poae
Lentzea sp.
Novosphingobium sp.
Pseudomonas sp.
Staphylococcus epidermidis
Staphylococcus pasteuri
Staphylococcus sp.
Unknown
GenBank closest relative
AY584662
AF164363
AY016347
AB195634
AY382475
AB008403
AY016346 (·10)
AF548071
AY251121
AY741243*t
AY741246*t
AY030322, AY137509 (·4)
AY741243*t
AY211147*m, AY211154 (·2)*m
AY043085 (·3)
AY21131*m, DQ129237
X76564
AY884570
AF157479
AY771333 (·3)
AY211167*m
AY114175
AE016822
DQ008601
SSU37347
AJ575816
AJ717377
AB009944 (·2)
DQ113448
AY345421 (Actinomyces)
GenBank closest relative #(s)
Table 3 Bacterial and fungal tropical mid-Atlantic (~15 N, 45 W) atmospheric isolates collected between 22 May and 30 June 2003
Aerobiologia (2006) 22:211–226
221
123
123
Neotestudina rosatii
Penicillium chrysogenum
Penicillium glabrum
Penicillium sp. or
Talaromyces leycettanus
Penicillium sp., Uscovopsis sp.,
Thysanophora sp.,
Chromocleista malachitea,
Talaromycis leycettanus, or
Eupencillium sp.
Phoma herbarum
Pleosporaceae sp.
Preussia terricola
Setosphaeria monoceras,
Pleospora herbarum,
or Embellisia sp.
Setosphaeria monoceras,
S. rostrata, or Cochliobolus sativus
Setosphaeria rostrata
Stachybotrys kampalensis
Trichophyton mentagrophytes,
or T. rubrum
Ulocladium botrytis
Ulocladium botrytis,
Clathrospora diplospora,
or Alternaria sp.
8
1
1
1
6
1
4
99 (573/576), (660/667)
99 (581/583)
99 (436/438)
99 (620/622)
97 (397/409)
99 (571/576)
99 (371/373),
99 (578/583), (616/623),
(617/621), (619/622)
AY864822 (·5)
97 (399/409), (472/483)( · 2),
(472/484), (489/504), (567/584)
99 (571/576), (576/580), (580/583),
(643/649), (656/662), (674/679)
AF548106 (·2)
AF548106
SRU42487*f
AF548099*c
AY083226
AY016352
AB195634 (·2),
AY642526 (·3),
AF212309
AY544726
AF212309
AY640998
AF290082
AF548090
AY526487
L76623
GenBank closest relative #(s)
99 (437/438)
99 (561/566), (562/567), (565/567),
(565/568), (612/615), (646/649),
(647/650), 97 (561/565)
98 (496/503)
98 (556/562)
99 (637/643)
% Homology (sequence length)
Where multiple genus/species are listed only the first GenBank Blast # match is listed. *m = Closest GenBank relative a Bamako, Mali air isolate, *t = Closest
GenBank relative an Erdemli, Turkey air isolate, *f = species recognized as a Florida plant/tree pathogen. *c = Closest GenBank relative a Caribbean air isolate
2
1
1
1
1
1
6
1
GenBank closest relative
# of isolates
Table 3 continued
222
Aerobiologia (2006) 22:211–226
Aerobiologia (2006) 22:211–226
recovery rates. These data also demonstrated that
the fungi (spore formers) are less susceptible to
desiccation stress than non-spore forming
bacteria.
While no single culture medium will result the
growth of all viable cells in any environmental
setting (soil, air water, serum, etc.), R2A is an
efficient medium for utilization in single medium
studies. R2A was selected for use in this and
previous studies primarily because of its ability to
simultaneously recover stressed and non-stressed
bacteria and fungi using a single filter (Griffin
et al., 2003; Kellogg et al., 2004). While UV light
may be attenuated up to 50% in large dust plumes
(Herman, Krotkov, Celarier, Larko, & Labow,
1999), a typical dust cloud transit (across the
Atlantic) time of 5–7 days can result in significant
microbial UV exposure. Desiccation stress from
wind transport (Lighthart, 1997), which averaged
5.6 m s–1 for this study period, and collection via
membrane filtration as demonstrated can further
limit CFU recovery. Employment of a low nutrient substrate such as R2A limits nutrient-shock
which may be experienced by stressed cells plated
on a nutrient rich substrate and numerous studies
have shown that R2A gives superior recovery of
CFU in comparison to high nutrient substrates in
many different settings (body fluids, water,
etc.) (Guerrero, 1987; Horgan, Matheson,
McLoughlin-Borlace, & Dart, 1999; Lesne, Berthet, Binard, Rouxel, & Humbert, 2000; Massa,
Caruso, Trovatelli, & Tosques, 1998; Reasoner,
2004; Reasoner & Geldreich, 1985; van der Linde,
Lim, Rondeel, Antonissen, & de Jong, 1999). A
number of studies have successfully utilized R2A
to recover bacteria from soils, the lower atmosphere, and extreme atmospheric altitudes (Bodour, Drees, & Maier, 2003; Gentry, Newby,
Josephson, & Pepper, 2001; Griffin, 2004; Griffin
et al., 2003; Kellogg et al., 2004; Ovreas &
Torsvik, 1998).
Membrane filtration was chosen for collection
of culturable bacterial and fungal CFU due to
cost, ease of use, and the ability of membranes to
efficiently capture cells. To limit viability loss due
to filtration desiccation, low flow rates and amount
of air volumes filtered were selected. The pump
air flow rate over the filter membrane was
0.4 m s–1 in contrast to the prevailing wind speed
223
which averaged 5.6 m s–1 and turbulence. Given
this high-wind and thus desiccating environment,
we believed that additional cell stress due to filtration will have minimal impact on surviving
culturable populations relative to losses due to
other types of collection stressors such as those
seen with impaction (Stewart et al., 1995). The
large volume air samples utilized after 6 June 2003
were chosen to increase the probability of recovering microbial CFU after a noted drop off in the
recovery of CFU with the small volume samples
following the elevated dust period of 25–29 May,
2003. It should be noted that the filtration volume
CFU recovery data presented demonstrated statistically significant lower bacterial recovery with
elevated volumes filtered and any bacterial CFU
reported in Table 1 for the large volume samples
should be considered conservative in nature.
These experiments demonstrate that choice of
methodology does make a difference in rates of
recovery of microorganisms and this is well known
in the various fields of microbiology.
Of the identified bacteria recovered in this
current study, 2 were Proteobacteria. Proteobacteria (Gram-negative bacteria) are the most prevalent bacterial group in marine waters (Sherr &
Sherr, 2000), and during periods of sea-spray
generation (severe weather, high surface activity,
etc.), these organisms may become the dominant
air isolate in marine environments, as previously
observed (Griffin et al., 2003). All of the nonspore-forming bacteria recovered were pigmented. Pigmentation, high G+C content, and
spores impart resistance to UV inactivation and
as a result enhance atmospheric survival
(Riesenman & Nicholson, 2000; Sundin & Jacobs,
1999). Although 16S rDNA sequences alone do
not provide the resolution needed to determine
relatedness (Stackebrandt & Goebel, 1994) or
point of origin, two of the isolates (one Bacillus
aminovorans, 353/353 bp, and Kocuria rosea, 628/
628 bp) matched at 100% sequence similarity to
two dust-borne isolates previously collected from
the atmosphere in Bamako, Mali, Africa (Kellogg
et al., 2004). Kocuria rosea has also been isolated
from the atmosphere in the U.S. Virgin Islands
when African desert dust was present, on several
occasions, and is a known human pathogen
(Altuntas et al., 2004; Griffin, Garrison, Herman,
123
224
& Shinn, 2001; Griffin et al., 2003). The remaining
two isolates of B. aminovorans and the Bacillus
sp. isolate listed in Table 3 also matched
(GenBank closest relative) to Bamako, Mali isolates at a sequence identity range of 98–99%
(633–704 bp). Three of the isolates were identified as Gordonia terra, and this species is capable
of causing disease in humans (Lesens et al., 2000).
Gordonia terrae, in addition to various species of
Bacillus and Staphlyococcus, have also been
identified in atmospheric samples collected in
Bamako, Mali, Africa (Kellogg et al., 2004).
The most prevalent fungal species isolated
were 10 colonies of Lojikania enalia. Two of the
currently available isolates of Lojikania enalia
(American Type Culture Collection, ATTC 16472
and 16473) were isolated in Liberia, Africa, from
an aerial root and a piece of driftwood, respectively. A 569 bp alignment of all the Lojikania
enalia isolates listed in Table 3 with the ATCC
16473 Lojikania enalia isolate resulted in a
sequence identity range of 99.3–99.8%. Twenty
percent of the fungal isolates identified at the
species level are recognized pathogens to humans,
animals, plants, and trees (Alternaria dauci,
Massaria platani, four isolates of Neotestudina
rosatii and Ulocladium botrytis). The three
Alternaria sp. isolates listed in Table 3 all matched to isolates (each matched to a previous
GenBank submission as listed in Table 3) we
have recovered from atmospheric samples in
Erdemli, Turkey which is a dominant dust-borne
microorganism at that coastal location (unpublished data). Spores of various species of Alternaria are potent allergens. Given that ~50% of
the particles in Florida’s atmosphere each summer are African in origin (Prospero, 1999), it is
interesting to note that Massaria platani (aka
Splanchnonema platani), a causative agent of
Florida Sycamore (Plantanus occidentalis) canker
(Alfieri, Langdon, Wehlburg, & Kimbrough,
1984), and Alternaria dauci, an identified pathogen of Florida carrots (Dugdale, Mortimer, Isaac,
& Collin, 2000), were isolated during this project.
It is tempting to speculate that transatlantic
transport of dust could be a vector to renew
reservoirs of some plant and animal pathogens in
North America and could also be a source of new
diseases.
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Aerobiologia (2006) 22:211–226
As Fig. 3 illustrates, the majority of bacterial
and fungal CFU was recovered during periods of
elevated dust concentration. The three highest
total CFU counts relative to air volume assayed
occurred on 25 May 2003, 26 May 2003, and
6 June 2003 (31.3, 11.2, and 15.9 CFU m–3,
respectively). These counts corresponded with the
two highest periods of dust activity as depicted by
the NAAPS model (47.4–110.9 lg m–3, 25–28
May 2003, and 45.3 lg m–3, respectively).
Figure 3 also illustrates that the highest recovery
of culturable microorganisms occurred on 25 May
2003, one day before the model derived dust
concentration peak on 26 May 2003. This could be
due to the lighter dust particles moving out ahead
of heavier dust cloud particulates, due to preferential wind-transport.
Dust-borne aeromicrobiology research conducted in Bamako, Mali, Africa (February 3, 2001
to March 1, 2002) demonstrated bacterial and
fungal population sizes of 3,548 and 129 CFU m–3,
respectively (Kellogg et al., 2004). In this
mid-Atlantic study, bacterial and fungal populations averaged (all samples) between 0.1 and
0.4 CFU m–3. These data indicate a dust-borne
reduction of culturable bacterial and fungal
CFU m–3 of ~104 and 102, respectively, from the
continent of Africa to this mid-Atlantic research
site during the early Northern Hemispheres
trans-Atlantic dust season. This noted difference is
probably due to the synergistic effects of particle
fall-out during transport, atmospheric dilution, and
cell stress (desiccation, UV exposure, temperature,
humidity, etc.).
It should be emphasized that microbial ecology studies have shown that what is typically
cultured from a given volume of soil represents
< 1.0% of the actual total prokaryote population
present (Torsvik, Salte, Sorheim, & Goksoyr,
1990). Since African dust moving through the
atmosphere is a component of African soil, the
dust-associated CFU counts presented in this
study may only represent a small fraction of the
prokaryote community that was actually present
in any of the given atmospheric samples. This
study presents evidence of early summer survival
and transport of microorganisms from North
Africa to a mid-Atlantic research site. Additionally, the statistical correlation between the
Aerobiologia (2006) 22:211–226
NAAPS model dust concentrations and microbial CFU counts observed demonstrates the potential for using models to address the global
atmospheric dispersion of microorganisms (nonpathogenic and pathogenic) and other soil-associated constituents, since they pertain to ecological evolution and ecosystem and human
health.
Acknowledgements This research was supported by the
U.S. Geological Survey (2090-OJ503) and the Ocean
Drilling Program. Thanks to Norman Kuring and Gene
Feldman of NASA’s SeaWiFS Project for providing daily
satellite imagery while aboard ship, and Cindy Morris of
INRA Avignon, Montfavet, France for several thorough
edits and suggestions. Also, special thanks to ODP Leg 209
shipboard scientific party and crew. Any use of trade
names is for descriptive purposes only and does not imply
endorsement by the U.S. Government.
References
Alfieri, S. A. J., Langdon, K. R., Wehlburg, C., & Kimbrough, J. W. (1984). Index of plant diseases in Florida,
revised edition. Florida Department of Agriculture
and Consumer Serv., Div. Plant Ind.
Altuntas, F., Yildiz, O., Eser, B., Gundogan, K., Sumerkan, B., & Cetin, M. (2004). Catheter-related bacteremia due to Kocuria rosea in a patient undergoing
peripheral blood stem cell transplantation. BMC
Infectious Diseases, 4, 62.
Bodour, A. A., Drees, K. P., & Maier, R. M. (2003). Distribution of biosurfactant-producing bacteria in
undisturbed and contaminated arid Southwestern
soils. Applied and Environmental Microbiology, 69,
3280–3287.
Choi, D. S., Park, Y. K., Oh, S. K., Yoon, H. J., Kim, J. C.,
Seo, W. J., & Cha, S. H. (1997). Distribution of airborne microorganisms in yellow sands of Korea.
Journal of Microbiology, 35, 1–9.
Dugdale, L. J., Mortimer, A. M., Isaac, S., & Collin, H. A.
(2000). Disease response of carrot and carrot somaclones to Alternaria dauci. Plant Pathology, 49, 57–61.
Dytham, C. (1999). Choosing and using statistics, A biologist’s guide. Oxford: Blackwell Science.
Gentry, T. J., Newby, D. T., Josephson, K. L. & Pepper,
I. L. (2001). Soil microbial population dynamics following bioaugmentation with a 3-chlorobenzoatedegrading bacterial culture. Bioaugmentation effects
on soil microorganisms. Biodegradation, 12, 349–357.
Gloster, J., & Alexandersen, S. (2004). New directions:
airborne transmission of foot-and-mouth disease
virus. Atmospheric Environment, 38, 503–505.
Goudie, A. S., & Middleton, N. J. (2001). Saharan dust
storms: nature and consequences. Earth-Science
Reviews, 56, 179–204.
225
Grasby, S. E., Allen, C. C., Longazo, T. G., Lisle, J. T.,
Griffin, D. W., & Beauchamp, B. (2003). Supra-glacial
sulphur springs and associated biological activity in
the Canadian High Arctic – signs of life beneath the
ice. Astrobiology, 3, 583–598.
Griffin, D. W. (2004). Terrestrial microorganism at an
altitude of 20,000 m in Earth’s atmosphere. Aerobiologia, 20, 135–140.
Griffin, D. W., Garrison, V. H., Herman, J. R., & Shinn,
E. A. (2001). African desert dust in the Caribbean
atmosphere: microbiology and public health. Aerobiologia, 17, 203–213.
Griffin, D. W., Kellogg, C. A., Garrison, V. H., Lisle, J. T.,
Borden, T. C., & Shinn, E. A. (2003). African dust in
the Caribbean atmosphere. Aerobiologia, 19, 143–157.
Guerrero, J. J. (1987). Comparaciâon de la eficacia de dos
medios de cultivo en la recuperaciâon de bacterias
heterotrâoficas injuriadas con cloro. Revista Argentina
de microbiologı́a, 19, 161–164.
Herman, J.R., Krotkov, N., Celarier, E., Larko, D., &
Labow, G. (1999). The distribution of UV radiation at
the Earth’s surface from TOMS measured UV-backscattered radiances. Geophysical Research, 104,
12059–12076.
Honrath, R. E., Owen, R. C., Val Marti’n, M., Reid, J. S.,
Lapina, K., Fialho, P., Dziobak, M. P., Kleissl, J., &
Westphal, D. L. (2004). Regional and hemispheric
impacts of anthropogenic and biomass burning emissions on summertime CO and O3 in the North
Atlantic lower free troposphere. Journal of Geophysical Research, it 109, doi:10.1029/2004JD005147
Horgan, S. E., Matheson, M. M., McLoughlin-Borlace, L.,
& Dart, J. K. (1999). Use of a low nutrient culture
medium for the identification of bacteria causing
severe ocular infection. Journal of Medical Microbiology, 48, 701–703.
Husar, R. B., Tratt, D. M., Schichtel, B. A., Falke, S. R., Li,
F., Jaffe, D., Gasso, S., Gill, T., Laulainen, N. S., Lu,
F., Reheis, M. C., Chun, Y., Westphal, D., Holben, B.
N., Gueymard, C., McKendry, I., Kuring, N.,
Feldman, G. C., McClain, C., Frouin, R. J., Merrill, J.,
DuBois, D., Vignola, F., Murayama, T., Nickovic, S.,
Wilson, W. W., Sassen, K., Sugimoto, N., & Malm,
W.C. (2001). Asian dust events of April 1998. Journal
of Geophysical Research-Atmospheres, 106, 18317–
18330.
Johnson, K. S., Elrod, V. A., Fitzwater, S. E., Plant, J. N.,
Chavez, F. P., Tanner, S. J., Gordon, R. M., Westphal,
D. L., Perry, K. D., Wu, J., & Karl, D. M. (2003).
Surface ocean-lower atmosphere interactions in the
Northeast Pacific Ocean Gyre: aerosols, iron and the
ecosystem response. Global Biogeochemical Cycles,
17, 1063, doi:1010.1029/2000JC000555
Keleman, P. B., Kikawa, E., Miller, D. J., Abe, N., Bach,
W., Carlson, R. L., Casey, J. F., Chambers, L. M.,
Cheadle, M., Cipriani, A., Dick, H. J. B., Faul, U.,
Garces, M., Garrido, C., J. S., G., Godard, M. M.,
Graham, D. W., Griffin, D. W., Harvey, J., Ildefonse,
B., Iturrino, G. J., Josef, J., Meurer, W. P., Paulick, H.,
Rosner, M., Schroeder, T., Seyler, M., & Takazawa, E.
123
226
(2004). Leg 209 summary. In Proceedings of the Ocean
Drilling Program, Initial Reports – Leg 209, pp. 1–139.
College Station, TX: Ocean Drilling Program.
Kellogg, C. A., Griffin, D. W., Garrison, V. H., Peak, K.
K., Royall, N., Smith, R. R., & Shinn, E. A. (2004).
Characterization of aerosolized bacteria and fungi
from desert dust events in Mali, West Africa. Aerobiologia, 20, 99–110.
Lesens, O., Hansmann, Y., Riegel, P., Heller, R., BenaissaDjellouli, M., Martinot, M., Petit, H., & Christmann,
D. (2000). Bacteremia endocarditis caused by a
Gordonia species in a patient with a central venous
catheter. Emerging Infectious Diseases, 6, 382–385.
Lesne, J., Berthet, S., Binard, S., Rouxel, A., & Humbert,
F. (2000). Changes in culturability and virulence of
Salmonella typhimurium during long-term starvation
under desiccating conditions. International Journal
of Food Microbiology, 60, 195–203.
Lighthart, B. (1997). The ecology of bacteria in the
alfresco atmosphere. FEMS Microbiology and Ecology, 23, 263–274.
Massa, S., Caruso, M., Trovatelli, F., & Tosques, M. (1998).
Comparison of plate count agar and R2A medium for
enumeration of heterotrophic bacteria in natural
mineral water. World Journal of Microbiology &
Biotechnology, 14, 727–730.
Morton, A. G. (1961). The induction of sporulation in
mould fungi. Proceedings of the Royal Society
of London. Series B, Biological Sciences, 153, 548–569.
Moulin, C., Lambert, C. E., Dulac, F., & Dayan, U. (1997).
Control of atmospheric export of dust from North
Africa by the North Atlantic Oscillation. Nature, 387,
691–694.
Ovreas, L., & Torsvik, V. (1998). Microbial diversity and
community structure in two different agricultural soil
communities. Microbial Ecology, 36, 303–315.
Peccia, J., & Hernandez, M. (2001). Photoreactivation in
airborne Mycobacterium parafortuitum. Applied and
Environmental Microbiology, 67, 4225–4232.
Prospero, J. M. (1999). Long-term measurements of the
transport of African mineral dust to the southeastern
United States: implications for regional air quality.
Journal of Geophysical Research, 104, 15,917–15,927.
Reasoner, D.J. (2004). Heterotrophic plate count methodology in the United States. International Journal
of Food Microbiology, 92, 307–315.
Reasoner, D. J., & Geldreich, E. E. (1985). A new medium
for the enumeration and subculture of bacteria from
portable water. Applied and Environmental Microbiology, 49, 1–7.
Reid, J. S., Prins, E. M., Westphal, D. L., Schmidt, C. C.,
Richardson, K. A., Christopher, S. A., Eck, T. F.,
Reid, E. A., Curtis, C. A., & Hoffman, J. P. (2004).
Real-time monitoring of South American smoke
particle emissions and transport using a coupled
remote sensing/box-model approach. Geophysical
Research Letters, 31, doi:10.1029/2203GL018845
123
Aerobiologia (2006) 22:211–226
Riesenman, P. J., & Nicholson, W. L. (2000). Role of the
spore coat layers in Bacillus subtilis spore resistance
to hydrogen peroxide, artificial UV-C, UV-B, and
solar UV radiation. Applied and Environmental
Microbiology, 66, 620–626.
Schollaert, S. E., Yoder, J. A., Westphal, D. L., & O’Reilly, J. E. (2003). The influence of dust and sulfate
aerosols on ocean color spectra and chlorophyll-a
concentrations derived for SeaWiFS of the U.S.
Coast. Journal of Geophysical Research, 108, 3191,
doi:3110.1029/2000JC000555
Sherr, E., & Sherr, B. (2000). Marine microbes: an overview. In D. L. Kirchman (Ed.), Microbial ecology
of the oceans (pp. 1–542). New York: Wiley-Liss Inc.
Smit, E., Leeflang, P., Glandorg, B., Elsas, J. D. V., &
Wernars, K. (1999). Analysis of fungal diversity in the
wheat rhizosphere by sequencing of cloned PCRamplified genes encoding 18S rRNA and temperature
gradient gel electrophoresis. Applied and Environmental Microbiology, 65, 2614–2621.
Stackebrandt, E., & Goebel, B. M. (1994). Taxonomic
note: a place for DNA–DNA reassociation and 16S
rRNA sequence analysis in the present species
definition in bacteriology. International Journal of
Systematic Bacteriology, 44, 846–849.
Stewart, S. L., Grinshpun, S. A., Willeke, K., Terzieva, S.,
Ulevicius, V., & Donnelly, J. (1995). Effect of impact
stress on microbial recovery on an agar surface. Applied and Environmental Microbiology, 61, 1232–1239.
Sundin, G. W., & Jacobs, J. L. (1999). Ultraviolet radiation
(UVR) sensitivity analysis and UVR survival strategies of a bacterial community from the phyllosphere
of field-grown peanut (Arachis hypogeae L.). Microbial Ecology, 38, 27–38.
Torsvik, V., Salte, K., Sorheim, R., & Goksoyr, J. (1990).
Comparison of phenotypic diversity and DNA heterogeneity in a population of soil bacteria. Applied and
Environmental Microbiology, 56, 776–781.
van der Linde, K., Lim, B., Rondeel, J., Antonissen, L., &
de Jong, G. (1999). Improved bacteriological surveillance of haemodialysis fluids: a comparison between
Tryptic soy agar and Reasoner’s 2A media. Nephrology Dialysis Transplantation, 14, 2433–2437.
Whitman, W. B., Coleman, D. C., & Wiebe, W. J. (1998).
Prokaryotes: the unseen majority. Proceedings of the
National Academy of Sciences of the United States
of America, 95, 6578–6583.
Wu, P. C., Tsai, J. C., Li, F. C., Lung, S. C., & Su, H. J.
(2004). Increased levels of ambient fungal spores in
Taiwan are associated with dust events from China.
Atmospheric Environment, 38, 4879–4886.
Yeo, H. G., & Kim, J. H. (2002). SPM and fungal spores in
the ambient air of west Korea during the Asian dust
(Yellow sand) period. Atmospheric Environment, 36,
5437–5442.