phage presentation- the isolation and characterization of mycobacteriophage musamodel from tropical...
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ISOLATION OF A NOVEL MYCOBACTERIOPHAGE
Lizbeth Pérez Castro
Javier Zavala Ayala
Mónica Del Moral
Félix Vallés
WHAT IS A BACTERIOPHAGE?
• A bacteriophage is any virus that infects any bacterium.
• Mycobacteriophages are viruses that infect mycobacteria.
• Found in different bacterial colonies.
• Mostly found in the soil and sea water.
• Resistant to antibiotics.
http://news.bbcimg.co.uk/media/images/66413000/jpg/_66413229_m0900092-bacteriophage_virus-
spl.jpg
BACTERIAS FOR HOSTING
1. M. smegmatis (Smeg)
• fast growing rate and non-pathogenic behaviour
• Aerobic organisms
• Found in water, soil and plants
2. Bacillus
• Obligate aerobes or facultative anaerobes
• Well understood- used as a model for differentiation, gene regulation, and cell cycle events in bacteria.
• Common in soil
Both are:
Gram positive
Acid fast bacteria
Morfology- rodshape
BACTERIOPHAGE STRUCTURE
• Phages consist of 3 basic components:
• Capsid-contains the genetic material
• Genetic material- DNA
• Tail-serves to attach to bacteria and DNA passes through tail and into bacterium
http://en.wikipedia.org/wiki/Bacteriophage
PHAGE LIFE CYCLES
• Temperate-Lysogenic:
• Replicate and lyse or dormant state- incorporate its genetic material into DNA of host.
• Virulent-Lytic:
• Straightforward cycle by lysing all bacteria they infect.
http://biomedfrontiers.org/infection-2014-3-28/
INTRODUCTION
• Problem: Can novel bacteriophages be isolated from tropical soil of Puerto Rico?
• Hypothesis: Unique bacteriophages will be isolated and characterized from a collected soil sample of Puerto Rico.
http://news.bbcimg.co.uk/media/images/66413000/jpg/_664
13229_m0900092-bacteriophage_virus-spl.jpg
PROCEDURE
httpwww.teachthechildrenwell.comscience.h
tml
http://www.exportersin
dia.com/swastikaelectr
ic/products.htm
http://quizlet.com/6760545/lab-3-streak-plate-
technique-flash-cards/
Soil
EnrichmentFiltration
Purification
HTPLTen Plate Preparation
MY BACTERIOPHAGE
• Name: Nitidusvenutus
• Located in: Carretera 943 Sector Los Chinos, Gurabo
• Coordinates: 18º 16’ 7’ N, 65º 58’ 32’ W
• Collected: February 24, 2014
• It was found in a rural area next to cow feces on a template day.
LOCATION
MY BACTERIOPHAGE
RESULTS
http://faculty.plattsburgh.edu/donald.slish/lambdatitre.html
MY BACTERIOPHAGE: MUSAMODEL
• Name- Musamodel
• Located in: 1-9 calle Barcelona, Urb.Terralinda, Caguas P.R
• Coordinates: 88˚14’6”N , 66˚2’39”W
• Collected: March 16, 2014
• Time: 7:51pm
• Temperature: 79˚
• About 3 inches deep and 1 foot closeto concrete
• Found next to a plantain tree
• Soil was moist and loose
LOCATION
SAMPLES COLLECTED1st 2nd 3rd 4th 5th 6th 7th
Date 2/6/14 2/18/14 2/24/14 3/3/14 3/10/14 3/16/14 3/16/14
Time 7:53 am 7:06 am 9:55 pm 9:38 am 6:26 am 7:45 pm 7:51 pm
Coordinates 18˚14’6”N,
66˚2’40”W
18˚14’13”N,
66˚2’35”W
18˚14’7”N,
66˚2’39”W
18˚14’18”N,
66˚2’42”W
18˚14’7”N,
66˚2’40”W
18˚14’6”N,
66˚2’39”W
18˚14’17”N,
66˚2’42”W
Temperature 75˚ 75˚ 79˚ 81˚ 73˚ 79˚ 79˚
Soil
description
Moist, 3
inches deep
Moist, 1 inch
deep
Moist, 1
inch deep
Moist, with
moss, 1 inch
deep
Moist, 1
inch
Moist,
loose, 5
inches
deep
Moist, loose,
3 inches
deep
Site
description
Urban, 5 feet
close to
concrete
Urban, 1 foot
from
concrete
Urban, 2
feet from
concrete
Urban, ½
foot from
concrete
Urban, 3
feet away
from
concrete
Urban, 1
foot close
to
concrete
Next to a
plantain
plant, 1 foot
close to
concrete
RESULTS
CONCLUSION
• Two mycobacteriophages were found:
1. Nitidusvenutusa. Located in Carretera 943 Sector Los Chinos, Gurabo.
b. Based on the clear plaques, we can assume it has a lytic life cycle.
2. Musamodela. Located in 1-9 calle Barcelona, Urb. Terralinda, Caguas P.R.
b. Up to now, we believe it has a lytic life cycle because of clear plaques.
c. Three purifications were completed.
FUTURE PLANS
• Nitidusvenutus;• Isolation of the bacteriophage’s DNA• Bioinformatics analyses and sequencing.
• Musamodel;• 3rd purification- to have more isolated plaques and measure diameter• Spot test• Make phage stocks-
• Empirical test
• 10 plate preparation
• Analyze • Bioinformatics and sequencing
ACKNOWLEDGEMENT
• Dr. Michael Rubin
• Juan Apiz
• Eduardo Correa
• Gustavo Martínez
• Giovanni Cruz
• Edwin Alvarado