PRESENTED TO:-

Dr. AMRITA SRIVASTAVA

ASSISTANT PROFESSOR

DEPARTMENT OF LIFE SCIENCE

CENTRAL UNIVERSITY OF BIHAR

PRESENTED BY:-

MONALISA

ROLL NO.- 06

M.SC. LIFE SCIENCE (2014-16)

CENTRAL UNIVERSITY OF BIHAR

There are many approaches used in classifying

microorganisms.

Broadly these are divided into two categories.

Classical Characteristics

Morphological

Physiological

BiochemicalEcological

Genetic

Easy to study and analyze

Variable characters, adaptations plays great role

Features used in classification

cell shape, cell size, colonial morphology, staining behavior, cilia and flagella, colour, endospore shape, etc.

Directly related to nature and activity of microbial enzymes and proteins

Analysis provides indirect comparison of microbial genomes

Features used in classification

Carbon-nitrogen sources, mechanism of energy conversion , nutritional type, pH optimum, salt tolerance, temperature, etc.

Features used in classification

Pigment produced, fermentation products, sensitivity to metabolic inhibitors and antibiotics, etc.

Not involved in growth and metabolism but commercially useful

Ecological characteristics

Features used in classification

Life cycle patterns, nature of symbiotic relationship, ability to cause disease, habitat preferences, etc.

Microbes similar in many respects can inhabit different ecological niche, suggesting they are not be closely related as first suspected.

Genetic characteristics

• Chromosomal gene exchange process- transformation,

conjugation.

• Occur between prokaryotic species but rarely between genera.

• Provides evidence of close relationship.

• Plasmids carry genes coding for phenotypic traits.

• Degree of relatedness can be misunderstood if that phenotypic

character is present on chromosomes used in classification.

Molecular characteristics

nucleic acid base

composition

nucleic acid hybridization

nucleic acid sequencing

genomic fingerprinting

amino acid sequencing

•Estimated by determining the melting temperature (Tm ) of the

DNA

•Higher G + C gives a

higher melting temperature

GC content

•Higher animals and plants : 30% to 50%

•Prokaryotes : 25% to 80%

•Useful in determining prokaryotic genera: variation in genera

less than 10% , content may vary greatly

Measure amount of radioactive DNA attached to nylon filter.

The degree of similarity is expressed as the % of experimental DNA

radioactivity retained on the filter as compared to other species of the

same genus under the same conditions.

Usually less than 5 % difference in melting point ( T m ) is considered as

members of same species.

Nucleic acid sequencing

•Most ideal method for comparing microbial genomes

•Sequences of 16S (prokaryotes) and 18S (eukaryotes) small

subunit ribosomal RNA (SSU rRNA) are used most often in

phylogenetic studies•Oligonucleotide signature sequences are short conserved sequences specific for a phylogenetically defined group of organisms

•Either complete or, more often, specific rRNA fragments can be compared

•When comparing rRNA sequences between two organisms, their relatedness is represented by an association coefficient or Sab

value

•The higher the Sab value, the more closely related the organisms

•Sab value= 1.0 ; identical organism

•DNA sequences can also be used to determine species strains in addition to genus

•It requires analysis of genes that evolve more quickly than rRNA encoding genes

•Multilocus sequence typing (MLST), the sequencing and comparison of 5 to 7 housekeeping genes instead of single gene is done.

•This is to prevent misleading results from analysis of one gene.

Genomic fingerprinting

•Repetitive sequences amplified by the polymerase chain reaction

•Amplified fragments run on agarose gel, with each lane of gel corresponding to one microbial isolate

•pattern of bands analyzed by Gel Document system•comparison of bands on basis of restriction fragment length polymorphism (RFLP)

Such as, 154 bp BOX, 124-127 ERIC, 30-40 bpREP

•It allows for identification to species, subspecies and often allows strain level identification

Amino acid sequencing

•The amino acid sequence of a protein is a reflection of the mRNA sequence and therefore of the gene which encodes that protein

•Amino acid sequencing of proteins such as cytochromes, histonesand heat-shock proteins has provided relevant taxonomic and phylogenetic information

•Provides more information

•Slow and expensive method.