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Antifungal Properties of Cutaneous Bacteria Found on Rana catesbeiana (North American Bullfrog) and Bufo boreas halophilus (California Toad). Kathy Szick-Miranda.

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Presentation on theme: "Antifungal Properties of Cutaneous Bacteria Found on Rana catesbeiana (North American Bullfrog) and Bufo boreas halophilus (California Toad). Kathy Szick-Miranda."— Presentation transcript:

1 Antifungal Properties of Cutaneous Bacteria Found on Rana catesbeiana (North American Bullfrog) and Bufo boreas halophilus (California Toad). Kathy Szick-Miranda California State University, Bakersfield 2010

2 anshu04.jpg  Amphibians Introduction /R9Ny0PfOPQU/s1600/bullfrog.jpg 3fHERaN40H4/Tc1frxaD5xI/AAAAAAAAAhI/HkEBZF5HGBg/s160 0/newts+3+of+them.jpg frogs/colorful-frogs02.jpg

3  Massive decline in the number of amphibian species worldwide.  One disease is chytridiomycosis, caused by the fungal pathogen Batrachochytrium dendrobatidis (Bd). Introduction pg?335

4  In the Southern San Joaquin valley, Bullfrogs (Rana catesbeiana) and California Toads (Bufo boreas halophilus) seem to be doing just fine! Introduction R. catesbeiana B. b. halophilus Charles M. Lane Kerry Kriger technology.com/projects/san_joaquin/ima ges/sjvec.jpg

5  Cutaneous bacteria act as a protective barrier. Marsh and Selwyn 1977; Al-Admawy and Noble 1981; McFall-Ngai et al  Cutaneous bacteria of some amphibians produce antibiotics that protect their hosts from pathogenic fungi. Austin 2000; Brucker et al. 2008a, 2008b; Harris et al Introduction Hypothesis: Bullfrogs and California Toads possess cutaneous bacteria that will inhibit the growth of some fungi.

6 Catch frogs and toads Streak plate Purify cutaneous bacterial isolates Methods Fig. 1AFig. 1BFig. 1CFig. 1D Fig. 1E Fig. 1AFig. 1BFig. 1CFig. 1D Fig. 1E Rinse with sterile water Swab frogs/toads

7 DNA extraction from fungi PCR amplification DNA sequencing to identify fungi Methods/Results Fig. 1AFig. 1BFig. 1CFig. 1D Fig. 1E Fig. 1AFig. 1BFig. 1CFig. 1D Fig. 1E  Five fungi identified: 2 distinct Aspergillus sp. Cochlibolus sp Eupenicillium sp. Galactomyces geotrichum Plate water samples Purify fungal isolates Collect water from frog/toad environment

8 Methods Fig. 1AFig. 1BFig. 1CFig. 1D Fig. 1E Fig. 1AFig. 1BFig. 1CFig. 1D Fig. 1E Cutaneous bacteria challenged against environmental fungi

9 Results  233 pure bacterial isolates challenged against 5 environmental fungi 5 isolates were positive against 4 fungi 5 isolates were positive against 3 fungi 16 isolates were positive against 2 fungi 43 isolates were positive against 1 fungus  Additional results: Cochlibolus sp. inhibited by 87% of the positive isolates. Eupenicillium sp. inhibited by 20% of the positive isolates. Aspergillus sp. (a) inhibited by 20% of the positive isolates. Galatomyces geotrichum inhibited by 22% of the positive isolates Aspergillus sp. (b) inhibited by 9% of the positive isolates.

10 Question Hypothesis #2: Bullfrogs and California Toads possess cutaneous bacteria that will inhibit the growth of known amphibian and human pathogens.  Data support the original hypothesis. Challenge positive isolates against known pathogens Approach:

11 Pathogens Basidiobolus ranarum: known human and amphibian pathogen causes skin and GI lesions found worldwide Candida albicans: normally found in low levels in the human body causes yeast infections Cryptococcus neoformans: affects immunocompromised patients causes lung infections

12 Results 85% of isolates inhibited the growth of B. ranarum 39% of isolates inhibited the growth of C. albicans 76% of isolates inhibited the growth of C. neoformans

13 Conclusions  Bullfrogs and toads possess cutaneous bacteria that inhibit the growth of some fungi.  Some cutaneous bacteria isolated from bullfrogs and toads inhibit the growth of known pathogenic fungi.  Implications: Improvement in amphibian conservation Advances in the treatment of fungal pathogens

14  Complete challenge assays with known pathogens.  Identify positive bacterial isolates.  Challenge the positive isolates against Bd and other known human pathogens.  Determine which metabolites in each of the bacterial species exhibit antifungal activity.  Examine cutaneous bacterial diversity of frogs and toads. flickriver.com 2012 bioweb.uwlax.edu 2008Impactlab.com 2009 Future Work

15 Dr. Antje Lauer – CSUB Chevron REVS-UP Program – CSUB CSUPERB-Faculty Seed Development Grant Chevron REVS-UP participants Student Researchers: Amanda Payne Ashley Nunez Lauren Dowel Christine Hluza David Tate Kathryn Hubert Esther Ibarra RJ Jimenez scientificamerican.com 2011 Acknowledgements


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