An Exploration of Endophytes Within Known Medicinal Plants ...An Exploration of Endophytes Within...
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An Exploration of Endophytes Within Known Medicinal Plants in Pursuit of Novel Antibiotic Synthesis Zaid Hatem 1 , Ethan Lilly 2 , Emily Rotich PhD 1 , Matthew Heard PhD 1 1 Belmont University, 2 Montgomery Bell Academy Question One: Diversity of Endophytes Question Three: Interactions with other Microorganisms Results Methods Methods Question Two: Characteristics of the Bacteria Acknowledgements Introduction Future Directions Discussion Big Questions Approximately 33,000 people die every year from infections caused by antibiotic resistant bacteria 1 . To fix this problem, we need new antibiotics. Because all our current antibiotics are derived from naturally produced compounds 2 and because many antibiotics like penicillin and streptomycin are derived from microorganisms, we decided to look for microbes that we could explore for novel antibiotic synthesis. Also, because many plants have been used in traditional medicine, we selected plants that are known to have medicinal values in order to explore their endophytes (microbes living within the plant) 3 . We selected Mentha spicata (spearmint), Mentha piperita (peppermint), Rosmarinus officinalis (rosemary), and Ginkgo biloba (gingko) to investigate due to their availability and known antibiotic properties 4, 5, 6 . In order to determine how each plant’s environment affects the diversity of these endophytes, we took samples from monocropped specimens (plants grown by themselves) and an intercropped sample (a plant grown with other plants) of each plant species except G. biloba. 1. How diverse are the fungal and bacterial endophytic communities in these four plants? Do we find differences in the diversities of endophytes between the monocropped and intercropped plants? 2. What characteristics do these endophytes have? 3. How do these endophytes interact with clinically-relevant microorganisms? Continuations in this work include... ● DNA extraction and sequencing of each isolate in order to identify the endophytic species for phylogenetic analysis ● exploring seasonal changes and their effects on endophytic diversity ● isolating alkaliphiles (bacteria that grow in basic conditions) ● sampling the same plant species from different locations to explore the endophytic diversities/communities ● exploring the in vitro interaction of the endophytes from the same microbiome ● determining the interaction between the endophytic isolates with other clinically-relevant microorganisms. We would like to give special thanks to the following people and organizations for their help, support, and/or funding of this project: Belmont University SURF, Montgomery Bell Academy, the Wilson family, Nicholas Ragsdale PhD, Rabab K. Sager PhD, Jonathan Creamer PhD, Jim Dickens PhD, Afaf Shamma MSc, Luma S. Hachim MSc, and our families. References ● Our isolates showed more diverse endophytic microbes in intercropped species than monocropped species. ● Although we have not identified the bacterial isolates to the species level, the gram test displayed a gram positive majority. ● The antibiotic tests showed some of the isolates to be resistant to some antibiotics. This result suggests that these endophytic isolates have an antibiosis mode of action against other microorganisms. ● The dominance of some isolates over the E. coli concludes that these endophytes could be a source of antibiotics, which needs to be explored further. Figure 1: The number of bacterial isolates. Blue represents the isolates from monocropped plants while orange represents isolates from intercropped plants. No monocropped G. biloba was used. Figure 2: The number of fungal isolates. Blue represents the isolates from monocropped plants while orange represents isolates from intercropped plants. No monocropped G. biloba was used. Figure 5: ‘A’ shows that B23 resists penicillin, novobiocin, and erythromycin. ‘B’ shows a bacteria (B9) susceptible to all four antibiotics. Figure 8: Shows the antibiotic resistance of select bacterial endophytic isolates to P10, E15, NB30, and C15. The standard deviation error bars were used because the experiment was done in triplicates. Figure 6: B1 isolate, one of 30 different plates, shows the results of the E. coli interaction tests. The growth of the bacteria from the disc into the E. coli shows that this bacterial strain competes for resources, an antibiotic agent property. Figure 7: Fungal isolate CF4 showing the interaction with E. coli. While the zone of clearance is not very large, this fungus could be a candidate for further interaction tests. This fungus has a bactericidal effect on E. coli. Gram and Shape M. spicata M. piperita R. officinalis G. biloba I M I M I M I Bacilli + 2 2 10 2 0 1 5 Bacilli − 0 0 1 0 0 0 1 Cocci + 0 0 0 0 0 0 0 Cocci − 0 0 2 3 0 0 1 Table 1: The results of the Gram test and the shape of 45% of the bacterial isolates. “I” represents the the samples taken from intercropped hosts while “M” represents the samples that were taken from monocropped hosts. Results Figure 3: Working under the laminar hood on antibiotic resistance tests. Figure 4: B9 isolate showing gram negative bacillus. A B B
Transcript of An Exploration of Endophytes Within Known Medicinal Plants ...An Exploration of Endophytes Within...
An Exploration of Endophytes Within Known Medicinal Plants in Pursuit of Novel Antibiotic Synthesis
Zaid Hatem 1, Ethan Lilly 2, Emily Rotich PhD 1, Matthew Heard PhD 11 Belmont University, 2 Montgomery Bell Academy
Question One: Diversity of Endophytes Question Three: Interactions with other Microorganisms
ResultsMethods
Methods
Question Two: Characteristics of the Bacteria
Acknowledgements
Introduction
Future Directions
Discussion
Big Questions
Approximately 33,000 people die every year from infections caused by antibiotic resistant bacteria 1. To fix this problem, we need new antibiotics. Because all our current antibiotics are derived from naturally produced compounds 2 and because many antibiotics like penicillin and streptomycin are derived from microorganisms, we decided to look for microbes that we could explore for novel antibiotic synthesis. Also, because many plants have been used in traditional medicine, we selected plants that are known to have medicinal values in order to explore their endophytes (microbes living within the plant) 3. We selected Mentha spicata (spearmint), Mentha piperita (peppermint), Rosmarinus officinalis (rosemary), and Ginkgo biloba (gingko) to investigate due to their availability and known antibiotic properties 4, 5, 6. In order to determine how each plant’s environment affects the diversity of these endophytes, we took samples from monocropped specimens (plants grown by themselves) and an intercropped sample (a plant grown with other plants) of each plant species except G. biloba.
1. How diverse are the fungal and bacterial endophytic communities in these four plants? Do we find differences in the diversities of endophytes between the monocropped and intercropped plants?2. What characteristics do these endophytes have?3. How do these endophytes interact with clinically-relevant microorganisms?
Continuations in this work include...● DNA extraction and sequencing of each isolate in
order to identify the endophytic species for phylogenetic analysis
● exploring seasonal changes and their effects on endophytic diversity
● isolating alkaliphiles (bacteria that grow in basic conditions)
● sampling the same plant species from different locations to explore the endophytic diversities/communities
● exploring the in vitro interaction of the endophytes from the same microbiome
● determining the interaction between the endophytic isolates with other clinically-relevant microorganisms.
We would like to give special thanks to the following people and organizations for their help, support, and/or funding of this project: Belmont University SURF, Montgomery Bell Academy, the Wilson family, Nicholas Ragsdale PhD, Rabab K. Sager PhD, Jonathan Creamer PhD, Jim Dickens PhD, Afaf Shamma MSc, Luma S. Hachim MSc, and our families.
References
● Our isolates showed more diverse endophytic microbes in intercropped species than monocropped species.
● Although we have not identified the bacterial isolates to the species level, the gram test displayed a gram positive majority.
● The antibiotic tests showed some of the isolates to be resistant to some antibiotics. This result suggests that these endophytic isolates have an antibiosis mode of action against other microorganisms.
● The dominance of some isolates over the E. coli concludes that these endophytes could be a source of antibiotics, which needs to be explored further.
Figure 1: The number of bacterial isolates. Blue represents the isolates from monocropped plants while orange represents isolates from intercropped plants. No monocropped G. biloba was used.
Figure 2: The number of fungal isolates. Blue represents the isolates from monocropped plants while orange represents isolates from intercropped plants. No monocropped G. biloba was used.
Figure 5: ‘A’ shows that B23 resists penicillin, novobiocin, and erythromycin. ‘B’ shows a bacteria (B9) susceptible to all four antibiotics.
Figure 8: Shows the antibiotic resistance of select bacterial endophytic isolates to P10, E15, NB30, and C15. The standard deviation error bars were used because the experiment was done in triplicates.
Figure 6: B1 isolate, one of 30 different plates, shows the results of the E. coli interaction tests. The growth of the bacteria from the disc into the E. coli shows that this bacterial strain competes for resources, an antibiotic agent property.
Figure 7: Fungal isolate CF4 showing the interaction with E. coli. While the zone of clearance is not very large, this fungus could be a candidate for further interaction tests. This fungus has a bactericidal effect on E. coli.
Gram and Shape
M. spicata M. piperita R. officinalis G. biloba
I M I M I M I
Bacilli + 2 2 10 2 0 1 5
Bacilli − 0 0 1 0 0 0 1
Cocci + 0 0 0 0 0 0 0
Cocci − 0 0 2 3 0 0 1
Table 1: The results of the Gram test and the shape of 45% of the bacterial isolates. “I” represents the the samples taken from intercropped hosts while “M” represents the samples that were taken from monocropped hosts.
Results
Figure 3: Working under the laminar hood on antibiotic resistance tests.
Figure 4: B9 isolate showing gram negative bacillus.
ABB
Morgan Stevenson
