Contribution of Leaf-Surface Fungi to the Air Spora

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Contribution of Leaf-Surface Fungi to the Air
Spora

• Estelle Levetin and Kip Dorsey

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Air Spora and Leaf Surfaces

• Phylloplane fungi considered major contributors
  to air spora
• In Tulsa leaves appear by mid-April and remain on
  the trees till November
• High concentration of airborne spores occur
  during this same period
• Few studies have compared taxa on leaf surfaces
  with those in the atmosphere

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Leaf Surfaces

• Leaf surfaces colonized by many types of bacteria
  and fungi
• Both saprobic and pathogenic species
• Leaf surfaces can also serve as traps for
  airborne fungi that settle on surface
• Both colonizers and settled spores may become
  entrained (or reentrained) into the atmosphere

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Purpose of Study

• Examine the phylloplane fungi on two common trees
  in the Tulsa area
• Look for patterns in the communities on leaves
  throughout the growing season
• Compare these taxa to those appearing in the
  atmosphere
• Look for possible parallels in changes of air
  spora and leaf surface fungi

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Daily Air Sampling

• Air sampling was carried out with a Burkard Spore
  Trap on the roof of biology building on the
  University of Tulsa campus
• Sample drums were changed every 7 days and cut
  into 24 hour segments
• Slides were examined at 1000x using the single
  longitudinal transverse method

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Tree Types

• Quercus (Oak) and Ulmus (Elm) are two dominant
  tree types in eastern Oklahoma
• Quercus palustris and Ulmus americana were chosen
  for this studies

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Leaf Surface Fungi

• Leaves collected weekly from Apr. 18 - Nov. 23
• Three leaves from each tree were aseptically
  picked, placed in sterile plastic bags and
  immediately taken to the Aerobiology Lab
• Various methods tested for fungal isolation from
  leaf surfaces
• 4cm2 areas on both adaxial and abaxial surfaces
  were separately wiped with sterile cotton swabs
  that were dampened with sterile distilled water
• Swabs were each put in a sterile tube with 1ml of
  sterile distilled water and vortexed for 30
  seconds
• Swabs were removed and 0.5ml of each spore
  suspension was pipetted onto petri dishes
  containing malt extract agar plus streptomycin

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Leaf Surface Fungi

• 12 plates were prepared each week total 360
  plates
• Cultures were incubated at room temperature for 5
  to 7 days
• Colonies were counted and fungi identified to
  genus level with standard keys

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Airborne Fungal Spore Concentration
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23 Taxa identified Cladosporium, Ascospores,
Basidiospores, and Alternaria Comprised 90 of
Total
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Results of leaf surface cultures

• 21,624 colonies isolated from leaf surfaces
• 23 taxa identified
• 15 taxa appeared on first sample, April 18
• Most prevalent taxa (over 90)
• Yeast 62.3
• Phoma-type 12.3
• Cladosporium 10.8
• Alternaria 4.7

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Fungi Identified from Leaf Surfaces
Acremonium Drechslera Pithomyces
Alternaria Epicoccum Rhizopus
Arthrinum Fusarium Seimatosporium
Aspergillus Geotrichum Sporothrix
Aureobasidium Hyalodendron Trichoderma
Choanephora Nigrospora Yeast
Cladosporium Penicillium Non-sporulating
Curvularia Phoma-type
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Leaf Surface Concentrations

• Generally there was a greater number of colonies
  isolated from the abaxial surface but no
  significant difference
• Generally there was a greater number of colonies
  isolated from elm leaves but no significant
  difference between oak and elm
• Mean concentrations of leaf surface fungi
  (CFU/cm2)

Oak Adaxial Oak Abaxial Elm Adaxial Elm Adaxial
22.3 31.7 32.7 34.6
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Yeasts

• A general term to describe unicellular fungi that
  reproduce by budding
• No attempt was made to identify yeast other than
  Aureobasidium
• Several different types of yeasts were routinely
  isolated based on pigmentation and cell shape
• Yeasts generally dispersed by rain splash
  however, Taylor recently reported that
  Aureobasidium was the most abundant taxa
  identified on Burkard air samples from a southern
  California location

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Phoma

• A large genus (223 spp) of anamorphic fungi in
  the Ceolomycetes that are characterized by
  conidia formation in a pycnidium spore
  dispersal by rain splash
• Many common ascomycetes have a Phoma anamorph
  including species in
• Leptosphaeria
• Didymella
• Mycosphaerella
• Pleospora
• Phaeosphaeria
• Diaporthe
• Phoma - like genera Phomopsis, Plenodomus,
  Peyronellaea, etc
• Some Epicoccum isolates have a Phoma synanamorph

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Concentrations of Leaf Surface Fungi and Airborne
Spores
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Correlation of Airborne Spores and Leaf Surface
Fungi

• When individual genera examined there was no
  significant correlation between leaf surface
  fungi and airborne concentration
• Phoma did show a significant correlation with
  airborne ascospore levels
• Nine fungal taxa were found on both leaf surface
  cultures and the airborne samples
• The mean concentrations for the season were
  compared using a Spearman Correlation
• There was a significant correlation between leaf
  surface fungi and airborne fungi (r0.74,
  p0.035)

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Comparison of Airborne Ascospore Concentration to
Leaf Surface Phoma
Positive correlation r 0.41, plt0.05
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Correlation with meteorological variables

• Temperature was the most important factor of
  airborne spore levels
• Airborne spore concentrations were significantly
  related to average daily temperature during the
  April to November study period
• Rainfall was the most important factor for leaf
  surface fungal concentrations
• Several taxa showed significant correlations with
  weekly rainfall totals

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Comparison of total weekly rainfall and number
of colonies from leaf surfaces
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Correlation of Leaf Surface Fungi and Rainfall
Spearman R P Value
Total Colonies 0.557 0.001
Yeast 0.554 0.001
Phoma 0.481 0.007
Cladosporium 0.131 0.489
Alternaria -0.232 0.217
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Leaf Surface Significance

• The leaf surface concentrations
• Oak 27.00 CFU/cm2
• Elm 33.68 CFU/cm2
• Approximate surface area of elm and oak leaves
• Oak 50 cm2
• Elm 23 cm2
• Total number of leaves per tree estimated using
  average branch-to-branchlet technique
• Oak 100,000 leaves
• Elm 325,000 leaves
• Surface area estimate for trees
• Oak 5.0 x 106 cm2
• Elm 7.5 x 106 cm2
• Estimate of CFU per tree
• Oak 1.35 x 108 CFU
• Elm 2.53 x 108 CFU

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Possible Atmospheric Output on Peak Leaf Surface
Days
Fungi Peak Date Conc. CFU/cm2 Possible Oak total Possible Elm total
Cladosporium 6-Sep 6.67 3.34 x 107 5.00 x 107
Alternaria 13-Nov 3.58 1.79 x 107 2.69 x 107
Curvularia 22-Aug 0.78 3.9 x 106 5.85 x 106
Drechslera 18-Nov 1.00 5.0 x 106 7.5 x 106
Epicoccum 1-Nov 1.04 5.2 x 106 7.8 x 106
Pithomyces 25-Jul 1.08 5.4 x 106 8.1 x 106
Penicillium 22-Aug 1.13 5.65 x 106 8.48 x 106
Aspergillus 18-Aug 0.33 1.65 x 106 2.48 x 106
Nigrospora 3-Oct 0.50 2.5 x 106 3.75 x 106
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Conclusion

• Leaf surface fungi include taxa with airborne
  dispersal and those with rain splash dispersal
• Leaf surface fungi with airborne dispersal can be
  major contributors to the air spora
• Questions remain about
• Population changes on leaf surface
• Contribution of Phoma-complex and yeast to the
  air spora

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Acknowledgments

• Assistance of Claudia Owens for help with air
  sample analysis is greatly appreciated.