Characterization and Isolation of Bacterial Colonies by Plate Streaking

Al Jay T. Mejos
Institute of Biology, College of Science, University of the Philippine, Diliman, Quezon City

Abstract

The ubiquity of microorganisms makes them ineluctable factors that affect human affairs. This study has demonstrated not only the omnipresence
but also the abundance of microorganisms in the environment from which they were collected. The microorganisms were inoculated into Nutrient
Agar tubes, Nutrient Broth tubes, streaked on Potato Dextrose Agar and pure cultures were obtained using Plate Streaking method on NA plates.
A selected pure culture on NA plate was characterized being 4 cm2 in area, having a white pigmentation, translucent opacity, circular form, entire
margin and convex elevation. The colony also formed a pellicle on NB and did not grow on PDA, identifying it as non-fungal. Since the gross colony
morphology and characteristics are very variable bases for comparison, the descriptions obtained were not enough for bacteria identification.

Introduction plate was left in the locker, and then refrigerated, and then
Microorganisms are everywhere and their activities affect the most afterwards observed.
common of human affairs (Betsy & Keogh, 2005). Therefore,
knowledge of microbial activities and how to manipulate them is Results
very important and it is in this regard that this paper is going to Medium NA slant NB PDA plate NA Plate
demonstrate the ubiquity of microorganisms, particularly in the
men’s restroom on the second floor of the Institute of Biology.
Media refers to the growth medium in which bacteria cultures are
grown and maintained (Talaro, 2008). They could also be used in the Sketch
characterization of bacteria and in the analyses of their activities. In
this study four types of media were used: Nutrient Agar Slant,
Nutrient Broth, Potato Dextrose Agar and Nutrient Agar Plate. Spreader Pellicle Several
Nutrient Broths are liquid media used for the observation of growth formation No growth colonies
Description
pattern was was observed. were
bacteria motility and the formation of aggregate layers such as mats observed. observed. isolated
and pellicles. PDAs are solid, selective media for the growth of fungi Table 1: Observed Colony Morphology
and yeasts. NA media are solid, nonselective, chemically-defined
growth media made up of beef extract or peptone and agar as a Aspect Size Color Opacity Form Margin Elevation
solidifying agent. They are often prepared in slants for maintaining
4
cultures or plates for experiments in bacterial growth observations Type white translucent circular Entire convex
cm2
and colony morphology. Growth is the increase in the number of Table 2: Observed Colony Characteristics
cells while colony morphology is the physical appearance of a
bacterial colony (http://a-s.clayton.edu, 2009). Since bacteria cannot Discussion
be typically seen by the naked eye, colony morphology is the one Microorganisms are fastidious in different degrees and aspects. Most
firstly observed in the characterization of bacteria (Bailey & Scott, may be very adaptive like the isolated colony yet it was still unable
1966). to grow on PDA plate due to the absence of cellulase that could
In characterizing bacteria, it is also necessary that pure strains be have allowed it to digest PDA, therefore it is most probably not a
obtained and therefore plate streaking is used to separate individual fungus (Stevens, 1981).
bacteria cells, allowing them to form colonies of genetically identical The use of colony morphology, expected relative abundance in a
bacteria eliminating contaminations (Ruangpan & Tendencia, 2004). given habitat and bacterial motility may aid in the preliminary
Contaminations are unwanted bacteria or impurities that are a identification of a bacterial colony yet it can only do so little since so
hindrance in obtaining reliable experimental results. They are many bacteria share so many common traits like pellicle-forming,
reduced to a minimum by the employment of Aseptic Techniques. growth pattern, colony form and margin (Old & Duguid, 1970) and
The pure culture obtained was characterized based on colony even these bases of classification can be very variable that it can not
morphology, formation of bacterial matting and streak shape. be impeccable to rely only on gross morphology in identification of a
given colony.
Methodology
Media Preparation References
Commercially prepared Nutrient Agar powder, Nutrient Broth
powder and PDA powder were dissolved in different flasks of hot Bailey WR, Scott EG. 1966. Diagnostic Microbiology. 2nd Edition. Toppan
distilled water. 5 mL increments of the NA solution was poured into Company Ltd., Japan. 342
test tubes and plugged with cotton. The same was done with the NB
Betsy T, Keogh J. 2005. Microbiology Demystified. McGraw-Hill Co. Inc. p. 1
solution. The tubes, flask and Petri plates were autoclaved. The
remaining solutions in the flasks were poured into the plates inside Old DC, Duguid JP. 1970. Selective Outgrowth of Fimbriate Bacteria in
the laminar flow hood, which was previously wiped with 70% Static Liquid Medium. Journal of Bacteriology, Aug. 1970. Vol. 103,
ethanol and exposed to UV for 30 minutes. The tubes and plates no. 2 pp. 447-456
were allowed to solidify inside the hood.
Ruangpan L, Tendencia, EA. 2004. Laboratory Manual of Standardized
Bacteria Collection Methods for Antimicrobial
An NA plate was opened and exposed to the environment inside Sensitivity Tests for Bacteria Isolated from Aquatic Animals
the male restroom on the second floor in IB for 5 minutes. The and Environment. http://rfdp.seafdec.org.ph. Date Accessed: July 12,
plate was then incubated at 37°C for 24 hours and refrigerated. 2009.
Bacteria Isolation and Characterization Stevens RB, editor. 1981. Mycology Guidebook. University of Washington Press,
The morphology of a single isolated colony was characterized and Seattle.
inoculated into the NB tube, NA slant using single streak method
and to the PDA and NA plates using the quadrant streaking method. Talaro KP. 2008. Foundations in Microbiology: Basic Principles. 6th ed.
The subcultures were then incubated for 24 hours, while the PDA McGraw-Hill. 62-71

http://a-s.clayton.edu. Colonial Morphology. Date Accessed: July 4, 2009.