OLEH SUDRAJAT FMIPA UNMUL 2009. Klasifikasi Bakteri Somewhat different: a clinical rapid ID is often...
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Transcript of OLEH SUDRAJAT FMIPA UNMUL 2009. Klasifikasi Bakteri Somewhat different: a clinical rapid ID is often...
Klasifikasi BakteriSomewhat different: a clinical rapid ID is
often important when trying to find causative agent of a disease
Bergey’s manual: Manual is in lab for a reference when doing unknown
Developed on 1940’s for grouping bacteria according to standard diagnostic lab techniques available at the time (such as Grams stain)
Klasifikasi BakteriThe manual divides bacteria into 4 groups or
divisions on the basis of their Cell Wall1. Gram + (stain violet)2. Gram - (destain, and are counterstained
pink or reddish color)3. Bacteria that lack a cell wall (mycoplasma)4. organisms that have a cell wall lacking
“peptidoglycan” (archaeobacteria – Now called “Archaea”
Klasifikasi BakteriMore modern methods now used:1. DNA studies, genomics, gene probes2. using Bacterial viruses “bacteriophages”3. Serology – antibody – antigen reactions
(like blood typing)Examples: Salmonella typhiSalmonella typhimuriumAzotobacter vinelandii 12837E. coli 0157H7 nasty strain of E. coli
Genotypic Characteristics for Identifying Prokaryotesthe use of genotypic testing has increased
with the availability of technologygenotypic testing is particularly useful in
the case of organisms that are difficult to identify
several techniques includegene probesPCRsequencing rRNA
Genotypic Characteristics for Identifying Prokaryotesgene probes
single stranded DNA that has been labeled with a identifiable tag, such as a fluorescent dye
are complementary to target nucleotide sequences unique in DNA of pathogen
Microbe gene probed
Genotypic Characteristics for Identifying Prokaryotes
If there is a suspicion, based on symptoms or other environmental parameters that indicates that the organism to be identified may be “ organism A”, a single strand of “organism A’s” DNA is introduced with a tag attached (such as fluorescent dye). If the introduced DNA binds to the unknown organism, then it is identified as “organism A”. If it does not bind to the unknown organism, then the unknown is not “organism A”.
Genotypic Characteristics for Identifying ProkaryotesPCR: polymerase chain reaction
used to detect small amounts of DNA present in a sample (blood, food, soil)
the PCR chain reaction is used to amplify the amount of DNA present
Genotypic Characteristics for Identifying Prokaryotessequencing ribosomal RNA
of particular use for identifying prokaryotes impossible to grow in a culture
focus is place on the 16S molecules of the RNA because of it’s size approximately 1500 nucleotides
once the 16S molecule is sequenced, it can then be compared to the sequences of known organisms
Machine usedto pick coloniescontaining wantedDNA
Difficulties in Classifying Prokaryoteshistorically prokaryotes have been
grouped according to phenotypic attributesproblems with this approach include
mutation resulting in phenotypic changes “just because they look alike, does not
mean that they are even closely related according the prokaryotics”
new molecular approaches are providing better insight to the relatedness of microorganismsthe more similar the nucleotide
sequence, the more closely related DNAextraction
Genotypic Characteristics used in Classifying Prokaryotescomparison of nucleotide sequences
differences in DNA sequence can assist in determination of divergence of evolutionary path for organisms
DNA hybridizationsingle strands of DNA anneal
16S ribonucleic acidcomparing sequence of ribosomal RNA
relatedness to other organisms can be determined using numerical taxonomydetermined by the percentage of characteristics two
organisms have in commonThe more you have in common phenotypically with
another organism the closer related you are to that organism.
Genotypic Characteristics for Identifying ProkaryotesPCR: polymerase chain reaction
used to detect small amounts of DNA present in a sample (blood, food, soil)
the PCR chain reaction is used to amplify the amount of DNA present
Bacterial TaxonomyCharacteristics used to classify organisms
TraditionalSize, shape, gram reaction, need for O2
Ability to metabolize sugarsMetabolic end products
Supplemented byComparison of 100-300 characteristicsNucleic acid sequence of ribosomal RNA
General Groupings used in Taxonomy
Aerobic (grows in air), obligate if must have O2. Capnophilic if needs CO2.
Facultative anaerobe (grows in air, and can grow without oxygen).
Anaerobe (grows without oxygen, and most species do not grow well in air as O2 is toxic for them).
Microaerophilic (grows in a low concentration of oxygen, but not in its absence or in air).