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MIKROBIOLOGI – 11
Ordering Organisms
Bacterial Classification • Taksonomi: ilmu tentang identifikasi, klasifikasi
dan tatanama (nomenclature)
– Takson: suatu kategori/tingkatan, seperti phylum,
genus, atau species
– Dasar: fenotip
• Filogeni: sejarah evolusi dan hubungan genetik
antarorganisme
– Dasar: genotip

Evolution of Classification Schemes

Taxonomic Grades
Plants Animals
“Kingdoms of life”
Rank Example

Protists
Plants Animals Kingdom Procaryotae
Clear differences in Division Gracilicutis
cellular structure
recognized
Prokaryote Eukaryote Class Scotobacteria
Subdivision of eukaryotes
Order Spirochaetales
Protists
Family Leptospiraceae
Monera Plants Fungi Animals

Discovery of Archaea Genus Leptospira

Species L. interrogans
Archaea Bacteria Eukaryote
Figure 15.1 from Microbe

What is a species? Phylogeny of bacteria

• “A species is a group of actually or potentially interbreeding populations
that are reproductively isolated from other such groups”
• Typological species Includes
– A group of organisms in which individuals are members of the species if Staphylococci,
they sufficiently conform to certain fixed properties. Streptococci,
• Morphological species Listeria, B.
– A population or group of populations that differs morphologically from anthracis
other populations.
• Phylogenetic (Cladistic) / Evolutionary / Darwinian species
– A group of organisms that shares an ancestor; a lineage that maintains
its integrity with respect to other lineages through both time and space.
• Ecological species Includes E. coli,
– A set of organisms adapted to a particular set of resources, called a Salmonella,
niche, in the environment. Helicobacter,
• Genetic species Vibrio,
Pseudomonas,
– based on similarity of DNA of individuals or populations. Brucella
– Similar to phylogenetic, but doesn’t take into account what the
ancestors were or where DNA came from

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Nomenclature Further Subdivisions

• Standard
– Italicized, with genus name capitalized • Biovar (biotype) - special biochemical property
– sp.= singular; spp.= plural • Serovar (serotype) - antigenic properties
• Uncultivated • Pathovar (pathotype) - host specificity or other
– 1995 Proposal to use “Candidatus” as temporary pathogenic property
designation • Phagovar (phagotype) - bacteriophage susceptibility
• Must have some sequence information
• Must have some phenotypic/ecological information • Morphovar (morphotype) - morphological properties
• Subcategories

Traditional Bacterial Taxonomy

Traditional Bacterial Taxonomy
Strengths Weaknesses
• Classification based on Morphology Easy -Too broad
– Morphology & Gram Stain -Some bacteria change shape

• Rod, coccus, spiral, branched Ecology Addresses the biology of Organisms occupying very
the organism different niches may still be
– Ecology related
• Thermophilic, halophilic, soil, anaerobe, primary Biochemistry -Good means of -Single mutation can cause
pathogen, etc. differentiating between change
closely related organisms -Possibility for unrelated
– Biochemistry -Easily defined organisms to have similar
• Carbon source, nitrogen source, presence of profiles
specific enzymes (eg. urease, catalase) -Not useful for nonculturable

Standard Identification Scheme

Biochemical Characterization
Only works for culturable

Can be misleading

Shapes can vary

• Used clinically
• Can be automated Subject to point mutation,
can be dependent on
growth conditions

API strips

Courtesy of www.bact.wisc.edu/Microtextbook

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Fatty Acid
Chemotaxonomy
Analysis
• Analysis of cellular components including: • FAME= fatty acid methyl ester
– Cell wall
• Primarily for G+ • Fatty acids differ in chain length and presence
• Peptidoglycan and teichoic acid types of double bonds, ring structures, branched
– Lipids chains, hydroxy groups
• LPS and other membrane components • Analysis by gas chromatography
– Polyamines
• Useful above genus level • Precise growth conditions critical
• Precise growth conditions critical

Fatty Acid Analysis Fatty Acid Analysis

(FAME) (FAME) – 2

Development of new
Serology
species
• Heavily used for identification of medically important
• Higher eukaryotes
strains of E. coli, Salmonella, Legionella, and other
– Frequent genetic recombination due to mating
bacteria.
• Only between closely related organisms (<2% divergence)
• Antibodies are primarily raised against surface antigens,
– Mutation
such as flagella and LPS
– Reproductive isolation
• Gives no information about relatedness
• Bacteria
• Differences in serotype can result from minor amino – Most gene transfer is vertical
acid or chemical changes that may have little – Horizontal gene transfer is promiscuous
physiological relevance
• More useful for diagnosis than classification

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Use of DNA for

Classification DNA hybridization
• GC ratios
– Ranges from 20-80%
– Identical GC ratios may be meaningless
– GC ratios differing by >5% correlates with low DNA homology
• DNA hybridization (similarity, re-association)
• Sequence analysis
– Ribosomal
– Whole genome
– Collection of genes
• <100 widely conserved
From Brock Biology of Microorganisms, 10th ed.

Relatedness of
Enterobacteriaceae Use of rRNA (rDNA)
Sequence
• Escherichia and Salmonella
share 50% homology by
• 23S = 3300 nt; 16S = 1650 nt; 5S = 120 nt
DNA:DNA hybridization • Conserved regions allow PCR amplification with
• Escherichia and Shigella share “universal” primers
70-90+% homology by
DNA:DNA hybridization
• rRNA is abundant and relatively stable
• Escherichia and Proteus are • Relatively easy to align sequences
more distantly related

Courtesy of Jeffrey Lawrence

Advantages of using DNA for Problems with using DNA

classification information exclusively
• Important phenotypic differences can result from
• Not dependent upon growth conditions relatively small differences between genomes
• Contains information on the organism’s • DNA hybridization (reassociation)
– Cumbersome
evolutionary history – Much lab-to-lab variability
– Genes in the process of being lost – Many comparisons necessary
– Islands resulting from lateral transfer – Mostly useful for culturable organisms
• Ribosomal DNA sequence
• Organisms are truly defined by their DNA – Not enough information

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Classification of Exceptions are the

Bacteria Rule
• Genes on plasmids (extrachromosomal DNA) are no longer
considered when determining phylogeny • Bacillus anthracis and Bacillus thuringiensis are
• One species definition: “…strains with approximately 70% or very closely related, but are accepted as
greater DNA-DNA relatedness and with 5°C or less Δ Tm…”
Wayne et al., 1987 different species due to unique ecological niches
– If applied to eukaryotes, all primates would belong to • Helicobacter pylori strains differ by up to 7%,
same species
• Another definition: 16s sequence differs by more than 3% from but are considered the same species because
all other sequences
they occupy the same niche
– 16s sequence not always sensitive enough for speciation
• Genus definition: 16s sequence differs by more than 5% from all
other sequences, or 25% genomic hybridization

Nomenclature of
the
genus Salmonella S. enterica is divided into 7 subspecies: (1) subsp. arizonae,
A single species (Salmonella enterica) was proposed in 1987, but clinicians (2) subsp. diarizonae, (3) subsp. enterica (4) subsp.
objected houtenae, (5) subsp. indica, (6) subsp. salamae, (7) subsp.
The genus Salmonella recently encompassed three species: “VII”
• Salmonella bongori
• Salmonella subterranea subsp. enterica is further divided into >150 serovars,
• Salmonella enterica including Typhimurium, Dublin, Enteriditis, Gallinarum,
Paratyphi A
Currently (according to NCBI) the species are: • Each serovar has been defined according to O group, Somatic O
• S. bongori antigen, Flagellar (H) antigen phase 1, and Somatic (H) antigen
phase 2
• S. subterranea
• Names are often shortened for convenience (if not clarity): S.
• S. choleraesuis enterica subsp. enterica serovar Dublin = S. dublin
• S. enteritidis
• Typhimurium, Choleraesuis, and Enteritidis appear as both
• S. typhi species and serovars???!!!
• S. typhimurium
• S. enterica

Culture Resource
Collections s
• Live specimens of all species should be deposited in a • International Journal of Systematic and Evolutionary
recognized public culture collection
Microbiology
– The official journal for publishing new bacterial
• ATCC: American type culture collection strains
• NTCC: National type culture collection (London) • New official names found at www.bacterio.cict.fr
• DSMZ: Deutsche Sammlung von mikroorganismen und • National Center for Biotechnology Information
zellculturen (Germany) (NCBI) Taxonomy browser (TaxBrowser)

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Reference Guidelines for Recognizing new

Books species
• Bergey’s Manual of Systematic Bacteriology
– Highly detailed phenotypic and genotypic information
– First edition published in 1923 1. Collect an adequate number of strains
– Second edition 2. Use 16S rRNA to find the closest related taxon
• Volume 1: 2001
• Volume 2: 2005 3. Do not use 70% DNA similarity as an absolute limit
• Volumes 3-5: coming soon
4. Characterize the phenotype
• Bergey’s Manual of Determinative Bacteriology
– Only contains phenotypic data for identification of culturable 5. Take your time doing analyses
organisms
• The Prokaryotes 6. Follow nomenclature rules
– 1992 edition has 4 volumes
– 3rd edition online:
http://141.150.157.117:8080/prokPUB/index.htm Based on FEMS Microbiol Rev, 2001, 25(1):39-67