This presentation provides information about the different methods of classification of bacteria.

1. Classification of Bacteria
Presented By:-
Zeeshan Mazhar
Md. Mahtab Rashid
M.Sc Ag (Mycology & Plant Pathology)
I.Ag.Sc , B.H.U.

2. Why do we need Bacterial Classification?
• A reliable classification system is a prerequisite for scientists and professionals
dealing with microorganisms in order to keep track of their tremendous variety.
• The ultimate objective of biological classification is the characterization and orderly
arrangement of organisms into groups.
• Classification is often confused with identification but, as a matter of fact,
classification is a prerequisite for identification.
• Many bacteriologists think that Bergey's Manual represents the official classification
but this is a misunderstanding. The editors of Bergey's Manual try to provide a
classification that is as accurate and up-to-date as possible but it is not official, in
contrast to bacterial nomenclature where each taxon has one valid name. The
closest to an official classification system is the one that is widely accepted by the
community

3. History of Classification of Bacteria & Archea
• The history of the classification of bacteria clearly demonstrates that changes were caused
by the availability of new techniques.
• The late 19th century was the beginning of bacterial taxonomy and Ferdinand Cohn in
1872 was the first to classify six genera of bacteria (as members of the plants) mainly
based on their morphology.
• At the beginning of the 20th century more and more physiological and biochemical data
were used, in addition to morphology, as important markers for the classification and
identification of microorganisms.
• The first edition of Bergey's Manual of Determinative Bacteriology classified the Bacteria in
1923 on the basis of these phenotypic properties as “typically unicellular plants”, the so-
called Schizomycetes. Even in the 7th edition of Bergey's Manual, published in
1957, Bacteria were still classified as members of plants (Protophyta, primitive plants).
Based on the partial sequences of 16S ribosomal RNA (rRNA) genes, Archaea (originally
named Archaebacteria) were first classified as a separate kingdom in 1977.
• In the 8th edition of Bergey's Manual, which was published in 1974, Bacteria were no
longer considered as plants and were recognized as members of the kingdom Procaryotae.

4. Time span Classification mainly based on
Late 19th century Morphology, Growth Requirements, Pathogenic potential
1900–1960 Morphology, Physiology, Biochemistry
1960–1980 Chemotaxonomy, Numerical Taxonomy,
DNA–DNA Hybridization
1980–today Genotypic Analyses, Multi-locus Sequence Analyses,
Average Nucleotide Identity, Whole Genome Analysis

5. Basis of Classification
A. Phentoypic Classification
i. Gram stain and bacterial morphology: Of all the different classification systems the Gram stain has withstood
the test of time. Discovered by H.C. Gram in 1884 it remains an important and useful technique to this day. It
allows a large proportion of clinically important bacteria to be classified as either Gram positive or negative based
on their morphology and differential staining properties. Slides are sequentially stained with crystal violet, iodine,
then destained with alcohol and counter-stained with safranin. Gram positive bacteria stain bluepurple and Gram
negative bacteria stain red. The difference between the two groups is believed to be due to a much larger
peptidoglycan (cell wall) in Gram positives. As a result the iodine and crystal violet precipitate in the thickened
cell wall and are not eluted by alcohol in contrast with the Gram negatives where the crystal violet is readily
eluted from the bacteria. As a result bacteria can be distinguished based on their morphology and staining
properties. Some bacteria such as mycobacteria (the cause of tuberculosis) are not reliably stained due to the
large lipid content of the peptidoglycan. Alternative staining techniques (Kinyoun or acid fast stain) are therefore
used that take advantage of the resistance to destaining after lengthier initial staining.
ii. Growth Requirements: Microorganisms can be grouped on the basis of their need for oxygen to grow.
Facultatively anaerobic bacteria can grow in high oxygen or low oxygen content and are among the more versatile
bacteria. In contrast, strictly anaerobic bacteria grow only in conditions where there is minimal or no oxygen
present in the environment. Bacteria such as bacteroides found in the large bowel are examples of anaerobes.
Strict aerobes only grow in the presence of significant quantities of oxygen. Pseudomonas aeruginosa, an
opportunistic pathogen, is an example of a strict aerobe. Microaerophilic bacteria grow under conditions of
reduced oxygen and sometimes also require increased levels of carbon dioxide. Neisseria species (e.g., the cause
of gonorrhea) are examples of micraerophilic bacteria.

6. iii. Biochemical reactions: Clinical microbiology laboratories typically will identify a
pathogen in a clinical sample, purify the microorganism by plating a single colony of
the microorganism on a separate plate, and then perform a series of biochemical
studies that will identify the bacterial species.
iv. Serologic systems: Selected antisera can be used to classify different bacterial
species. This may be based on either carbohydrate or protein antigens from the
bacterial cell wall or the capsular polysaccharide. (Group A streptococcal M proteins or
O and H polysaccharide antigens of salmonella).
v. Environmental Reservoirs: When considering likely pathogens it is also important to
know which of the different species are found in different locations. Environmental
reservoirs are generally divided into those that are endogenous (i.e., on or within the
human body) and exogenous (somewhere in the environment). When considering the
likely cause of an infection the likely source of the infection is important in your
differential diagnosis. For example an abscess that develops after large bowel surgery
is likely caused by an anaerobic organism that is resident in the large bowel. A skin rash
developing in a hiker with a history of multiple tick bites is more likely to be borrelia,
the agent of Lyme disease. An outbreak of food poisoning traced to imported
unpasteurized cheese might be due to listeria. Endogenous reservoirs account for a
large proportion of human infections. Many parts of the body have their own normal
flora. S. epidermidis is found on the skin. Viridans streptococci are a part of the normal
oropharyngeal flora and S. aureus is a commensal of the anterior nares.

7. B. Genotypic Classification
i. Ribosomal RNA (rRNA) sequence analysis: This has emerged as a major
method for classification. It has been used (as described above) to establish a
phylogenetic tree. In addition it is now also used to rapidly diagnose the
pathogen responsible for an infection, to help select appropriate therapy and to
identify noncultivatable microorganisms.
ii. Universal Phylogenetic Tree: Woese has developed a “universal phylogenetic
tree” for all living organisms that establishes a tripartite division of all living
organisms– bacteria, archaea and eucarya. His work is based on a comparison of
16s ribosomal RNA sequences. These sequences are highly conserved and
undergo change at a slow, gradual and consistent rate. They are therefore useful
for making comparisons among the different living organisms.
iii. DNA-DNA hybridization
iv. G+C content

8. Morphological classification
• Bacteria can be classified into six major groups on morphological basis.
1. TRUE BACTERIA
• Cocci – These are spherical or oval cells. On the basis of arrangement of
individual organisms they can be described as
– Monococci (Cocci in singles) – Monococcus spp.
– Diplococci (Cocci in pairs) – Streptococcus pneumoniae
– Staphylococci (Cocci in grape-like clusters) – Staphylococcus aureus
– Streptococci (Cocci in chains) – Streptococcus pyogenes
– Tetrad (Cocci in group of four) - Micrococcus spp.
– Sarcina (Cocci in group of eight)

9. • Bacilli – These are rod-shaped bacteria. On the basis of arrangement of
organisms, they can be described as
– Diplobacilli
– Streptobacilli
– Palisades
– Chinese-letter form
– Coccobacilli
– Comma-shaped

12. 2. ACTINOMYCETES (actin- ray, mykes-fungus)
These are rigid organisms like true bacteria but they resemble fungi in that
they exhibit branching and tend to form filaments.
They are termed such because of their resemblance to sun rays when seen in
tissue sections.

13. 3. Spirochaetes
These are relatively longer, slender, non-
branched microorganisms of spiral shape
having several coils.

14. 4. Mycoplasmas
These bacteria lack in rigid cell wall (cell
wall lacking) and are highly pleomorphic
and of indefinite shape.
They occur in round or oval bodies and
in interlacing filaments.
5. Rickettsiae and Chlamydiae
These are very small, obligate parasites,
and at one time were considered closely
related to the viruses. Now, these are
regarded as bacteria.

15. Based on Anatomical features
• Capsule
• Capsulate– Streptococcus pneumoniae
• Non-capsulate – Viridans streptococci
• Flagella
• Flagellate –
• Monotrichous
• Lophotrichous
• Amphitrichous
• Peritrichous
• Aflagellate – Shigella spp.
• Spore
• Spore-forming – Bacillus spp.
• Non-sporing – Escherichia coli
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16. Based on Staining reaction
• GRAM’S STAIN
• Gram-positive cocci – Staphylococcus aureus
• Gram-negative cocci – Neisseria gonorrhoeae
• Gram-positive rods – Clostridium spp.
• Gram-negative rods – E. coli
• ACID FAST STAIN
• Acid-fast bacilli –Mycobacterium tuberculosis
• Non-acid-fast bacilli – Staphylococcus aureus

17. Based on Cultural characteristics
• Extra growth factors requirements
• Fastidious – Hemophilus influenzae
• Non-fastidious – Escherichia coli
• Hemolysis on Sheep Blood Agar
• Alpha-hemolysis – Streptococcus pneumoniae
• Beta-hemolysis – Streptococcus pyogenes
• Utilization of carbohydrates
• Oxidative - Micrococcus
• Fermentative – Escherichia coli
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18. Based on Cultural characteristics
• Growth rate
• Rapid growers– Vibrio cholerae
• Slow growers – Mycobacterium tuberculosis
• Pigment production
• Pigment producer – Staphylococcus aureus
• Pigment non-producer – Escherichia coli
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Based on Nutrition
• Autotrophs
• Heterotrophs

19. Based on environmental factors
A. Temperature
• Psychrophiles (15-200C) – Pseudomonas fluorescens
• Mesophiles (20-400C) – Escherichia coli, Salmonella enterica,
Staphylococcus aureus
• Thermophiles (50-600C)- Bacillus stearothermophilus
• Extremely thermophiles (as high as 2500C)
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20. B. Oxygen dependence
• Aerobe (grow in ambient temperature, which contains 21% O2 and
a small amount of CO2, 0.03%)
• Obligate aerobes – Strictly require O2 for their growth
(Pseudomonas aeruginosa)
• Micro-aerophilic (grow under reduced O2, 5-10% and increased
CO2, 8-10%)- Campylobacter jejuni, Helicobacter pylori
• Facultative anaerobe (capable of growing either in presence or
absence of O2)- E. coli
• Obligate anaerobe – Clostridium spp.
• Capnophilic (require increased concentration of CO2, i.e., 5-10%) –
H. influenzae,
N. gonorrhoeae
• Aerotolerant
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21. 21
C. pH
• Acidophiles (Lactobacillus acidophilus)
• Alkaliphiles (Vibrio)
• Neutralophiles (pH 6-8)
Majority of the medically important bacteria grow best at neutral or slightly
alkaline reaction (pH 7.2-7.6)
D. Salt concentration
• Halophiles
• Non-halophiles

22. Other ways of classification
• Motile/Non-motile
• Pathogenic/Non-pathogenic
• Sensitive/Resistant (to particular antibiotic/ chemicals)
• Lactose fermenter/Lactose non-fermenter
• Bergey’s Manual of Determinative Bacteriology
• Gram-negative eubacteria that have cell walls
• Gram-positive eubacteria that have cell walls
• Cell wall-less eubacteria: Mycoplasma
• Archaeobacteria
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26. Thanks for having patience with me.