Monday, May 13, 2013

MICROBIAL GROWTH


Requirements for Growth
•Physical
•Chemical

Physical Requirements
Temperature 
•psychrophiles   (cold loving microbes )
•range            0 C  -  20 C
•mesophiles    (moderate temp. loving microbes)
•range            20 C   -  40 C
•thermophiles     (heat loving microbes)
•range            40 C  -  100 C

Bacteria According the Oxygen Demand

Obligate Aerobes

Obligate Anaerobes

Facultative Aerobes and Anaerobes

Microaerophilic



Do You Know 
HOW TO CULTURE BACTERIA?


from Slant agar to Plate media culture


 


from Broth agar to Broth culture




PRINCIPLES OF STAINING 

1. Simple stains – increase contrast
2. Differential stains – differentiate   between organisms or structures
3. Gram stain – differentiates between two kinds of cell wall structures


Preparing smears for staining

1. Bacteria on slide
2. Air Dry
3. Bacteria are HEAT FIXED to the slide
4. Stain is applied

Bacteria are slightly negative, so are attracted to the positive chromophore of the BASIC DYE
Common Basic Dyes
•crystal violet
•methylene blue
•safranin
•basic fuchsin


Gram Staining



1884 Hans Christian Gram
most important stain used in Bacteriology

Divides all Bacteria into 2 groups:
Gram (+)
Gram (-)

drop Crystal Violet

Grams Iodine (mordant)

Alcohol

Safranin  (Counterstain)

Identification of a Bacteria 

Result

Gram (+)  PURPLE
Gram (-)   RED
Difference - due to structure of cell wall
Gram (+)      Thick cell wall
Gram (-)       Thin cell wall



This is the pictures of result :




GRAM (-)          and        GRAM (+)

GRAM (+)
GRAM (-)






The Discovery of Microbiology

Do you know him??


He is Girolamo Fracastoro.
and He stated that disease caused by invisible living organism

HOW ABOUT HIM?


Antoni  van Leeuwenhoek 

He was a man that have been succeed in that era to finding MICROSCOPE


THE GOLDEN AGE OF MICROBIOLOGY


     •1857-1914:ƒ Beginning with Pasteur’s work, discoveries included the relationship between    microbes and disease, immunity, and antimicrobial drugs
       1876: Robert Koch
         prove that a bacterium causes anthrax
         provided the experimental steps
         prove that a specific microbe causes a postulates, to prove that a specific microbe causes a specific disease.





GENETIC MATTER TRANSFERRING IN BACTERIA

GENETIC MATTER TRANSFERRING IN BACTERIA
By
Elok M S and Na’afi Aisya



CHAPTER I
INTRODUCTION

A.  Background
Bacteria is one example of microorganism that learned. Bacteria is group of prokaryote according to the non-existed of nuclei membrane. Bacteria have a plasmid as a genetic extra chromosome.
Bacteria as one example of the microorganisms that live closely related to human life. Often more is known of the role that bacteria negative role in human life is the transfer agent of disease. So many human diseases caused by bacteria. Actually, in addition to the negative role of bacteria cause harm to human life so much positive role of bacteria.
To determine the role of bacteria both adverse and beneficial to human life required more knowledge about bacteria. How is the structure of the constituent bacteria that affect the role of bacteria for life as well as how the process or mechanism of bacteria as agents of diseases that can harm human life. Therefore, in this paper we will discuss transferring the genetic matter that has to do with the structure of plasmids which are owned by the bacteria, which then affect the role of bacteria in human life.

B.   States of Problem
a.     How about the mechanism of genetic transformation in bacteria?
b.     How about the mechanism of transduction in bacteria?
c.     How about the mechanism of conjugation in bacteria?

C.   Purpose
a.     To know the mechanism of genetic transformation in bacteria.
b.     To know the mechanism of transduction in bacteria.
c.     To know the mechanism of conjugation in bacteria.

CHAPTER II
CONTENT
A.  Genetic Transformation
A kind of process which free DNA is incorporated into a recipient cell and brings about genetic change called genetic transformation. It led to experiments proving without a doubt that DNA was the genetic material. Both Gram-negative and Gram-positive species are a number of bacteria that have been found to be transformable.
Competent is a cell that is able to take up a molecule of DNA and be transformed. Competence in most naturally transformable bacteria is regulated, and special proteins play a role in the uptake and processing of DNA. Competence-specific proteins may include a membrane associated DNA-binding protein, a cell wall autolysin, and various nucleases. For example in Bacillus, about 20 percent of the cells become competent and stay that way for several hours whereas in Streptococcus, 100 percent of the cells can become competent just only for a few minutes during the growth cycle.
Uptake of DNA
The form which DNA is taken up in bacteria has differences. In gram-negative, only double-stranded DNA is taken up into the cell despite the fact, only single-stranded segments actually become incorporated into the genome by recombinant. On the other hand, in gram-positive, only a single DNA strands is taken up, while the complementary strand is simultaneously degraded. Actually, the more effectively binds to the cell is double-stranded DNA.
During the transformation process, competent bacteria first bind DNA reversibly soon, however the binding becomes irreversible. Competent cells bind much more DNA than do non-competent cells as much as 1000 times more. The sizes of the transforming fragments are much smaller than that of the whole genome and this DNA further degraded during the uptake process.

Integration of transforming DNA
a.     Transforming DNA is started by DNA-binding protein which made it bound at the cell surface,
b.     The DNA associates with a competence-specific protein which remains attached to the DNA, presumably preventing it from nuclease attack, until it reaches the chromosome where RecA protein takes over.
c.     The DNA is integrated into the genome of the recipient by re-combinational process.
d.     During replication of this hybrid DNA, one parental and one recombinant DNA molecule is formed.
Upon segregation at cell division, the letter will be present in the transformed cell, which is now genetically altered as compared to parental type. The transformation of plasmid DNA generally occurs in the absence of recombination between the plasmid and bacterial chromosome.
Electroporation
The cell membranes produce small pores which exposed to pulsed electrical fields. If DNA molecules are present outside the cells during the electrical pulse, the can then enter the cells through these pores. So that, the entering process of DNA molecules to the cells through the pores by electrical pulse called electroporation.
This technique has now been used to transport DNA into a large number of different species of bacteria, both Gram-negative and Gram-positive. Electroporation allows an experimenter to transfer a plasmid directly from one cell to another if both are present during electroporation.
B.   Transduction
Transduction is a process where DNA is transferred from cell to cell through the agency of viruses. Genetic transfer of host genes by viruses can occur in two ways, such as:
a.     Generalized transduction, host genes derived from virtually any portion of the host genome become a part of the DNA of the mature virus particle in place of virus genome.
b.     Specialized transduction, occurs only in some temperate viruses; a specific group of host genes is integrated directly into the virus genome usually replacing some of the virus genes and is transferred to the recipient during lysogenization.
Both specialized and generalized transduction is defective as a virus because bacterial genes have replaced some necessary viral genes.
a.     Generalized transduction
Virtually any genetic marker can be transferred from donor to recipient. Generalized transduction was first discovered in the bacterium Salmonella typhimurium with phage P22, known occur in Escherichia coli and another bacteria.







(Brock : 2012)
When the population of sensitive bacteria is infected with a phage, the events of the phage lytic cycle may be initiated. During a lytic infection, the enzymes responsible for packaging viral DNA into the bacteriophage sometimes accidentally package host DNA. The resulting particle is called a transducting particle. Upon lysis of the cell, these particles are released a long with normal virions, so that the lysate contains a mixture of normal and transducing virions. Because transducing particles cannot initiate a normal viral infection (they contain no viral DNA), they are said to be defective. When this lysate is used to infect a population of recipient cells, most of the cells become infected with normal virus. A small proportion of the population receives transducing particles, whose DNA can now undergo genetic recombination with the host DNA. Since only a small proportion of the particles in the lysate are of the defective transducting type and since each of those contains only a small fragment of donor DNA, the probability of transducting particle containing a particular gene is quite low.
Phages that form transducing particles can be either temperate or virulent, the main requirements being that they have a DNA packaging mechanism that permits accidental recognition of host DNA and that packaging occurs before the host genome is completely degraded. The detection of transduction is most certain when multiplicity of phage to host is low, so that a host cell is infected with only a single phage particle; with multiple infection, the cell may be killed by the normal particles.

b.   Specialized transduction
Specialized transduction is transfer which efficient process by phage of a specific set of host genes can be arranged. The example involves transduction of the galactose genes by the temperate phage lambda of E.coli. When the cell lysogenized into the host DNA at a specific site, the region in which lambda integrates is immediately adjacent to the cluster of host genes that control the enzymes involved in galactose utilization and lambda is inserted into the host DNA at that site and the replication of viral DNA is under host control.



(Brock : 2012)
Upon induction, viral DNA separates from host DNA by process integration reverse. When the lysogenic cell is induced, lambda DNA is excised as a unit. In rare condition, the phage genome excised incorrectly. Then both normal and rare conditions are in detached DNA replication. The phage synthesis occur until its completed. The last the cell lyses and releases. For the normal condition it occurred with normal phage whereas the rare condition cell lyses and releases defective phage capable of transducing galactose genes.


C.   Conjugation
Conjugation is a mechanism of transfer of genetic information (DNA) from donor cell to a recipient cell that occur as a result of cell-cell contact. Conjugation is controlled by genes carried by certain plasmids (such as the F plasmid) and involves transfer of the plasmid from a donor cell to a recipient cell. Plasmid DNA transfer involves replication via the rolling circle mechanism.
Bacterial conjugation was first discovered by Lederberg and Tatum in 1946. They combine the two mutant strains of Escherichia coli distinct unable to synthesize one or more essential growth factors and give him a chance to mate.
In the process of conjugation, donor cells (males) included some of the DNA into a recipient cell via phili sex owned by the male cells. Once the donor DNA into the recipient cell, enzymes that act on DNA cutting recipient of a DNA fragment recipient. Then the donor DNA integrated into chromosomal DNA in the recipient. This mechanism actually takes place also in the activities of transformation and transduction.
With the conjugation process, the specific genes for traits such as resistance to the drug can move from population to population of resistant bacteria resistant bacteria. Therefore, if it occurs in a population of bacteria can develop multi-drug resistance.

CHAPTER III
CONCLUSION

a.     Transduction is the transfer of host genes from one bacterium to another by a bacterial virus. In generalized transduction, defective virus particles randomly incorporate fragments of the cell’s chromosomal DNA, but the transducing efficiency is low. In specialized transduction, the DNA of a temperate virus excises incorrectly and takes adjacent host genes along with it; the transducing efficiency here may be very high.
b.     Transformation is a genetic transfer process by which free DNA is incorporated into a recipient cell and brings about genetic change.
c.     Conjugation is a mechanism of transfer of genetic information (DNA) from donor cell to a recipient cell that occur as a result of cell-cell contact.


References
Brock, et all. 1994. Biology of Microorganism Seventh Edition. United States of America : Prentice –Hall International, Inc.
Brock, et all. 2012. Biology of Microorganism Thirteenth Edition (e-book). San Francisco : Pearson Education, Inc.
Prescott, Lansing M. 2002. Microbiology 5th Edition. USA : The McGraw−Hill
Companies.