Courtney Dever and Jennifer Lopez
Classification
Kingdom: Bacteria
Phylum: Firmicutes
Class: Bacilli
Order: Bacillales
Family: Paenibacillaceae
Species
|
NCBI: Taxonomy |
Genus: Brevibacillus species: laterosporus [2]
Habitat Information
Describe the location and conditions under which the organism was isolated.
Two tablespoons of an unknown soil organism were collected at a depth of approximately 1” below the soil surface on January 29, 2016 at 11:53 am from (latitude 30.280954, longitude -97.760251) and placed in a plastic Ziploc baggie at room temperature. Below is a picture of the area, and also a picture of the exact location where the soil was taken at the time of retrieval.
Weather conditions on January 29, 2016 at 11:53 am were as follows [5]
Wind: S14
Visibility: 10
Weather: A few clouds
Sky conditions: FEW 200
Temperature:
• Air temperature: 73°F
• Dewpoint: 34
• 6 hour max: 73
• 6 hour min: 32
Relative humidity: 24%
Pressure:
• Altimeter (in): 30.06
• Sea level (mb): 1017.8
• Precipitation (in): 0
Weather conditions also included [6]
ET0 or PET: 0.10 in.
Temperature max: 76 °F
Temperature min: 36°F
RH min: 17%
Solar radiation: 17.55 MJm2
Rain: 0 in.
Wind 4 am: 0.12 mph
Wind 4 pm: 6.13 mph
Soil conditions included [7]
Map Unit Name: Travis soils and urban land, 1 to 8 percent slopes
Acres in AOI: 0.1
Percent of AOI: 100%
Description and Significance
Describe the appearance (colonial and cellular), possible antimicrobial activity etc. of the organism, and why the organism might be significant.
We chose colony #1 from our Master Patch Plate on E.coli because it showed a small clearing around the colony and hinted at antimicrobial properties.
Colony morphology:
-Size: small
-Margin: round/uneven
-Elevation:convex
-Surface: smooth
-Pigment produced: none.
-Color: Whitish and opaque.
Gram stain: positive.
Endospore stain: positive.
Capsule stain: negative.
Our organism is a G+ rod (bacillus) that forms short chains of 2. It also forms endospores.
Brevibacillus laterosporus has been shown to have antimycobacterial properties in a Brazilian study. [3]
Brevibacillus laterosporus has been shown to have pesticidal uses against insects and nematodes and mollusks. [4]
Brevibacillus laterosporus is a broad spectrum antimicrobial species due to its antimicrobial uses against phytopathogenic bacteria and fungi. [4]
Genome Structure
Describe the size and content of the genome. How many chromosomes? Circular or linear? Other interesting features? What is known about its sequence? Include S Ribosomal sequence that you obtained from PCR and sequencing here.
The genome of Brevibacillus laterosporus is small. It has one chromosome (5,106,578 bp) and 2 circular plasmids. [8]
BLAST Search from DNA sequencing results:
>CD-Forward_G10.ab1
GATGGAGCAACGCCGCGTGAACGATGAAGGCTTTC
GGGTCGTAAAGTTCTGTTGTTAGGGAAGAAACAGTGCTATTTAAATAAGATAGCACCTTGACGGTACCTAACGAGAAAGC
CACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTGTCCGGAATTATTGGGCGTAAAGCGCGCG
CAGGTGGCTATGTAAGTCTGATGTTAAAGCCCGAGGCTCAACCTCGGTTCGCATTGGAAACTGTGTAGCTTGAGTGCAGG
AGAGGAAAGTGGTATTCCACGTGTAGCGGTGAAATGCGTAGAGATGTGGAGGAACACCAGTGGCGAAGGCGACTTTCTGG
CCTGTAACTGACACTGAGGCGCGAAAGCGTGGGGAGCAAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGATGA
GTGCTAGGTGTTAGGGGTTTCAATACCCTTAGTGCCGCAGCTAACGCAATAAGCACTCCGCCTGGGGAGTACGCTCGCAA
GAGTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCT
TACCAGGTCTTGACATCCCACTGACCGCTCTAGAGATAGAGCTTCCCTTCGGGGCAGTGGTGACAGGTGGTGCATGGTTG
TCGTCAGCTCGTGCCGTGANATGTCATA
>JL-Reverse_H10.ab1
ACCACCTGTCACCACTGCCCCGAAGGGAAGCTCTATCTCTAGAGCGGTCAGTGGGATGTCAAGACC
TGGTAAGGTTCTTCGCGTTGCTTCGAATTAAACCACATGCTCCACCGCTTGTGCGGGCCCCCGTCAATTCCTTTGAGTTT
CACTCTTGCGAGCGTACTCCCCAGGCGGAGTGCTTATTGCGTTAGCTGCGGCACTAAGGGTATTGAAACCCCTAACACCT
AGCACTCATCGTTTACGGCGTGGACTACCAGGGTATCTAATCCTGTTTGCTCCCCACGCTTTCGCGCCTCAGTGTCAGTT
ACAGGCCAGAAAGTCGCCTTCGCCACTGGTGTTCCTCCACATCTCTACGCATTTCACCGCTACACGTGGAATACCACTTT
CCTCTCCTGCACTCAAGCTACACAGTTTCCAATGCGAACCGAGGTTGAGCCTCGGGCTTTAACATCAGACTTACATAGCC
ACCTGCGCGCGCTTTACGCCCAATAATTCCGGACAACGCTTGCCACCTACGTATTACCGCGGCTGCTGGCACGTAGTTAG
CCGTGGCTTTCTCGTTAGGTACCGTCAAGGTGCTANCTTATTTAAATAGCACTGTTTCTTCCCTAACAACAGAACTTTAC
GACCCGAAAGCCTTCATCGTTCACGCGGCGTTGCTCCATCAGACTTTCGTCCATTGTGGAAAATTCCCTACTGCTGCCNC
CCGTA
Cell Structure, Metabolism and Life Cycle
Interesting features of cell structure; how it gains energy; what important molecules it produces.
An interesting feature of B. laterosporus is that it forms a canoe-shaped lamellar body on one side of its spores. [4]
From our biochemical tests, namely Oxidase test, we were able to determine that our organism, B. laterosporus is an aerobe, uses oxygen as the final electron acceptor in the electron chain transport system. From the Phenol Red broth tests, we know that B. laterosporus ferments glucose for energy.
Physiology and Pathogenesis
Biochemical characteristics, enzymes made, other characteristics that may be used to identify the organism; contributions to environment (if any).
If relevant, how does this organism cause disease? Human, animal, plant hosts? Virulence factors, as well as patient symptoms.
Biochemical Characteristics:
Phenol Red Broth:
•Glucose fermentation: (+) result; broth was yellow indicating glucose fermentation.
•Lactose fermentation: (-) result; broth was red—no change in broth.
•Sucrose fermentation: (-) result; broth was red—no change in broth.
problems encountered during procedure: no problems occurred during procedure.
Starch Hydrolysis: (-) result; clearing was not present on starch agar media; soil organism does not produce the α-amylase enzyme.
Casein Hydrolysis: (-) result; no clearing in skim milk agar; soil organism does not produce the casease enzyme.
Gelatin Hydrolysis: (+) result; solid broth turned to liquid; soil organism produces the gelatinase enzyme.
DNA Hydrolysis: (+) result; clearing in agar; soil organism produces the DNase enzyme.
Lipid Hydrolysis: (-) result; no clearing in Tributyrin agar; soil organism does not produce the Lipase enzyme.
Methyl Red: (-) result; broth is orange in color; soil organism does not use the mixed acid fermentation pathway, and does not produce mixed acid end-products.
Voges Proskauer (+) result; broth is red in color; soil organism utilizes the butylene glycol pathway, produces acetoin, and produces neutral end-products.
Citrate Test: (-) result, slant is green in color; soil organism does not utilize citrate as a carbon source.
SIM Tests:
•Motility (+) result; soil organism is motile.
•Indole (+) result; soil organism is able to produce indole. *indole result is questionable as top of media was pink
•Sulfur (-) result; soil organism cannot reduce sulfur to hydrogen sulfide.
Nitrate Reduction Test: (-) result; soil organism is not able to reduce Nitrate to ammonia or molecular nitrogen; zinc was added the test tube turned red, showing a negative result for nitrate reduction.
Urea Hydrolysis: (-) result; broth is salmon in color; soil organism does not produce the urease enzyme. *result is questionable as the broth was not bright pink, nor was it orange or yellow.
Triple Sugar Iron: (K/A) result; slant is red in color, butt is orange in color; glucose fermentation with acid production; peptones/proteins catabolized aerobically (in the slant) with alkaline products (reversion).
Decarboxylation:
•Arginine (-) result; broth is yellow; soil organism does not produce arginine decarboxylase and does not ferment.
•Lysine (-) result; broth is yellow; soil organism does not produce lysine decarboxylase.
•Ornithine (-)result; broth is yellow; soil organism does not produce ornithine decarboxylase.
Oxidase Test: (+) result; soil organism can produce Cytochrome c oxidase.
Eosin Methylene Blue Agar : (-) result; organism didn't grow b/c it is G+.
Hektoen Enteric Agar (HE): (-) result; organism didn't grow b/c it is G+.
MacConkey Agar (MAC): (-) result; organism didn't grow b/c it is G+.
Phenylalanine Deaminase: (-) result; organism doesn't produce Phenylalanine deaminase.
Catalase Test: (-) result; organism doesn't produce catalase.
Blood Agar: (gamma) result; no clearing in the media; soil organism does not produce hemolysins.
Mannitol Salt Agar : (-) result; no growth and no fermentation.
Phenylethyl Alcohol Agar (PEA): (+) result; growth, but really slow growth.
Bile Esculin: (+) result; slant is dark brown in color; organism hydrolyzes esculin.
6.5% Salt Tolerance: (-) result; organism is not salt tolerant.
Bacitracin/Optochin: (-) result; organism produced no antibodies to A-disc or P-disc.
Antimicrobial Sensitivity/Kirby-Bauer Test:
•(-) to Triple Sulfa; organism is resistant to triple sulfa.
•(-) to Azlocillin; organism is resistant to azlocillin.
•(-) to Ceftazidime; organism is resistant to ceftazidime.
•(-) to Bacitracin; organism is resistant to bacitracin.
problems encountered during procedure: no problems occurred during procedure.
Disinfectant sensitivity:
•(-) to lavender; organism is resistant to lavender oil.
•(-) to clove; organism is resistant to clove oil.
•(-) to rosemary; organism is resistant to rosemary oil.
•(-) to oregano;organism is resistant to oregano oil.
problems encountered during procedure: no problems occurred during procedure.
Enzymes made: gelatinase, oxidase and DNase.
Other identifying characteristics: Gram + rod that produces endospores.
There are no known contributions to the environment. It is not known to cause disease.
References
http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi [1]
http://blast.ncbi.nlm.nih.gov/Blast.cgi [2]
http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1517-83822012000400036 [3]
Ruiu, Luca "Brevibacillus laterosporus, a Pathogen of Invertebrates and a Broad-Spectrum Antimicrobial Species". Insects 2013 Sep, Volume 4(3), 476-492; doi:10.3390/insects4030476
Retrieved from: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4553477/?tool=pmcentrez and
www.mdpi.com/2075-4450/4/3/476/pdf [4]
National Oceanic and Atmospheric Administration's National Weather Service at http://w1.weather.gov/ [5]
Texas A & M Agrilife Extension http://texaset.tamu.edu/ [6]
United States Department of Agriculture Natural Resources Conservation Service, Web Soil Survey
http://websoilsurvey.sc.egov.usda.gov/App/HomePage.htm [7]
Djuvic, Marvin, Anja Poehlein, Andrea Thürmer, and Rolf Daniel. "Genome Sequence of Brevibacillus laterosporus LMG 15441, a Pathogen of Invertebrates" Journal of Bacteriology, 2011, Volume 193, No. 19, pg 5535-5536.
Retrieved from: http://jb.asm.org/content/193/19/5535.full [8]
Author
Page authored by Courtney Dever and Jennifer Lopez, students of Prof. Kristine Hollingsworth at Austin Community College.
