Download Характеристика бактерий

Физиология
микроорганизмов
Игнатов А. Н.
1
Подсчет живых клеток
Метод разведения
2
Агаризованная среда
Первооткрыватель Frau Franny Eilshemius , супруга коллеги Роберта
Коха.
Dr. Walther and Fanny Hess
Водоросль Gracilaria – красная водоросль Gleidium
purpurascens
3
Оценка роста бактерий

Прямой подсчет клеток:
микроскопия,
проточная цитометрия
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Подсчет колоний на
среде
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Оценка сухой биомассы

Мутность среды
4
1, 2, 4, 8, 16, 32, 64, 128, 256, ….
Время удвоения
популяции
 Escherichia coli:12.5 min
 Vibrio cholerae : 13 min (can kill a man within 12 h)
 Mycobacterium tuberculosis : 24 h (develop
symptom after months)
5
Рост и выживание
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Лаг – фаза - Lag phase: adaptation to the
environment
Логарифмический рост - Exponential logarithmic
growth: machine in full rum
Стационарная фаза - Stationary phase: nutrition
exhausted, toxin increased
Отмирание - Decline: cell die (steady biomass) or
lysis (decrease biomass)
Покой - Dormant as spore, non-viable state
6
略
+ 37oC, pH 5.1 ; + 45oC, pH 6.2
Рост в биореаторе при различной температуре
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Факторы, влияющие на рост
бактерий
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Питательные вещества
pH (Нейтрофилы, ацидофилы, алкалифилы)
Температура (психрофилы, мезофилы,
термофилы)
Аэрация
Давление
Сила ионного раствора и осмотическое
давление (галофилы, осмофилы)
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Метаболизм – тип реакций, позволяющих
аккумулировать питательные вещества для
производства энергии для роста и размножения
Оптимум для жизни от +27 до
40.5оС.
Оптимум pH от 6.5 до 8.5.
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Термофилы
70o-110o
Мезофилы
10o-50o
10
Термофилы
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Горячие термальные источники
Источник термостабильных
ферментов для индустрии
Taq polymerase
Distinct membrane
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Жизнь на ранней Земле без O2
Аэробное дыхание производит свободные радикалы:
Каталаза: H2O2 --- H20 and 02
Супероксид-дисмутаза Superoxide dismutase (SOD): oxygen radical -- H20 and O2
Тип роста
1.
2.
3.
4.
5.
Облигатный аэроб -Obligate aerobe
Облигатный анаэроб - Obligate anaerobe
Микроарофил - Microaerophile
Аэротолерантный анаэроб - Aerotolerant anaerobe
Факультативный анаэроб - Facultative anaerobe/aerobe
略
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Требования к
кислороду - Air
requirement:O2
Aerobe: A microorganism whose growth requires the
presence of air or free oxygen
Anaerobe: A microorganism that grows only or best in the
absence of free oxygen. Organisms utilize bound oxygen
Microaerophile: A microorganism that grows best in the
presence of low concentrations of oxygen
Facultative anaerobe/aerobe: A microbe that adjusts its
metabolism to depending on the oxygen concentration in
which it is growing
Aerotolerant anaerobe: an organism that always grows in
an anaerobic mode -- it ignors the presence of oxygen.
Capneic microbe: An organism that requires 3 to 10%
CO2 for growth
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Рост- выращивание
略
Plating technique of Robert Koch
for single colony isolation
Microbiol. Mol. Biol. Rev. 64:847-867 (2000)
The majority of microbes persist
attached to surfaces within a
structured biofilm ecosystem
and not as free floating
organisms.
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Рост бактерий в биопленках
Метаболизм
Устойчивость
16
略
Два типа биопленок
Природные
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Symbioses
Termite, ruminant
digestion
Sewage treatment
bioreactors
Water pipes
Dental units
Contact lens cases
патогенные
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Dental plaque
Endocarditis
Cystic Fibrosis
Otitis media
Urinary catheter
Implants
17
略
Непрерывное
культивирование
микроорганизмов
18
Живые но не культурабельные клетки
19
http://nihroadmap.nih.gov/hmp/
Секвенирование микробиоты человека.
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6 задач микробов
1.
Поиск питательных веществ
2.
Метаболизм – синтез и катаболизм
3.
Производство энергии
4.
Выделение отходов-токсинов
5.
Размножение
6.
Адаптация - эволюция
21
Метаболизм
a very short summary
Секреция экзоферментов.
 Поглощение питательных веществ адсорбцией

22
Синтез и катаболизм
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Гликолиз

Two molecules of ATP are use to
phosphorylate glucose and start
glycolysis.
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The phosphorylated molecule is then
broken down in a series of reactions into
two three carbon molecules (lysis).
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Пентоза-фосфатный шунт
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When reducing power are needed
Sensitive to the level of NADP+
Did not generate ATP
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Идентификация бактерий по метаболическому профилю
Some microbes can metabolize certain molecules while
others can’t. When molecules are metabolized, specific
waste products are created such as acids. Those waste
products can be labeled by color.
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Классификация Бактерий
Taxonomy and Characteristics
Prokaryotes
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Domain Archaea
[ancient bacteria]
Domain Eubacteria
[true bacteria]
make up the old
Kingdom Monera
Prokaryotes
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Most abundant group
Widely distributed
Oldest organisms
Eukaryotes vs. Prokaryotes
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Eukaryotes
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DNA
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Prokaryotes
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Membrane, nucleus
Multiple chromosomes
Associated with histones
Organelles
Cell Walls
Division
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DNA
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Sexual: Meiosis
Asexual: Mitosis
Nonmembrane, nucleiod
One circular chromosome
No histones
Organelles
Cell Walls: peptidoglycan
Division
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Asexual
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Motility: Flagella whips
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Binary Fission
Snapping
Budding
Motility: Flagella spins
Domain Archaea
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Evolved from earliest
cells
Methanogens
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Largest group
Use CO2
Extreme environment
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Halophiles
Thermophiles
Psychrophiles
Acidophiles
Alkaliphiles
Eubacteria
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Prokaryotic
Unicellular
Shapes
Vary in size
Taxonomy: Bergey’s Manual
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Low G+C Gram Positive
High G+C Gram Positive
Gram Negative Proteobacteria
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Nonpathogenic: environmental contributions
Pathogenic
Bacterial Shapes
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Result of Asexual Reproduction
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Binary Fission
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Snapping Division
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Cross wall divides
Daughter cells +/- separate
Inner cell wall divides
Daughter cells hinged
Budding
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Outgrowth of original cell
Budding
Snapping Division
Binary Fission
Binary Fission Results
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Cocci
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Pairs
Chains
Tetrads
Cubes
Clusters
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Bacillus
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Separate
Pairs
Chains
Bacterial Shapes
Cocci: Pairs
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Division in one plane
 Diplococci
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Neisseria
Cocci: Chains
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Division in 2 Planes
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Streptococcus
Cocci: Tetrads
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Division in three planes
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Micrococcus
Cocci: 8-cell group
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Divides in 3 planes
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Sarcina
Cocci: Clusters
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Division in 3 planes
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Staphylococcus
Cocci Summary
Rods: Straight
E. coli
Rods: Club-Shaped
Corynebacterium
Rods: Branching
Actinomyces
Rods: Comma form
Vibrio
Rods: Spore Formers
Gram Negative Rod Summary
Gram Positive Rod Summary
Spiral Forms
Pleomorphic
Bacterial Shapes Review
Motility
Low G+C Gram Positive Organisms
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Rods
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Clostridia
Mycoplasmas
Bacillus
Listeria
Lactobacillus
Cocci
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Streptococcus
Enterococcus
Staphylococcus
High G+C Gram Positives
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Rods
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Corynebacterium
Mycobacterium
Actinomycetes
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Corynebacterium
Actinomyces
Nocardia
Streptomyces
Nocardia
M.tb
Gram Negative Alpha Proteobacteria
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Pathogenic
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Rickettsia
Brucella
Ehrlichia
Ehrlichia
Gram Negative Beta Proteobacteria
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Pathogenic
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Neisseria
Bordetella
Spirillum
Burkholderia
Gram Negative Gamma Proteobacteria
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Pathogenic
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Legionella
Coxiella
Pseudomonads
Enterobacteriaceae
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E. coli
Salmonella
Shigella
Proteus
Yersinia
Enterobacter
Serratia
Pseudomonas
Gram Negative Epsilon Proteobacteria
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Pathogenic
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Campylobacter
Helicobacter
Helicobacter
Campylobacter
Other Bacteria
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Chlamydia
Pathogenic
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Chlamydia
Spirochetes
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Borrelia
Treponema [syphilis]
Borrelia [Lyme ds]
Treponema
Scientific Nomenclature
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Binomial genus and specific epithet
(species). Is used world wide
Is always underlined
Rules for naming are set by international
committee’s
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International Code of Zoological Momenclature
International Code of Botanical Nomenclature
Bacteriological Code and Bergey’s Manual
Scientific Names
Scientific
Binomial
Klebsiella
pneumoniae
Pfiesteria piscicida
Salmonella
typhimurium
Streptococcus
pyogenes
Penicillium
chrysogenum
Trypanosoma cruzi
Source of Genus
Source of
Name
Specific Epithet
Honors Edwin
Klebs
Honors Lois
Pfiester
Honors Daniel
Salmon
Chains of cells
(strepto-)
Tuftlike (penicill-)
Corkscrew-like
(trypano-, borer;
The disease
Disease in fish
Stupor (typh-) in
mice (muri-)
Forms pus (pyo-)
Produces a
yellow (chryso-)
pigment
Honors Oswaldo
Cruz
Species Definition
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Eukaryotic species:
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Prokaryotic species:
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A group of closely related organisms that breed among
themselves
A population of cells with similar characteristics
Clone: Population of cells derived from a single cell
Strain: Genetically different cells within a clone
Culture: grown in the lab
Viral species:

Population of viruses with similar characteristics that
occupies a particular ecological niche
Is it as easy to classify Microbes as it is
Macrobes?
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How to classify
What do we have to look at?
Identifying Bacteria
Applications, p. 283
Phylogenetic Relationships of
Prokaryotes
Figure 10.6
Of more than 2600 species identified
so far

Only about 250 or
10% are pathogens
Classification and Identification
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Classification: Placing organisms in groups
of related species. Lists of characteristics of
known organisms.
Identification: Matching characteristics of an
“unknown” organism to lists of known
organisms.

Clinical lab identification
Bergey’s Manual of Systematic
Bacteriology
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Morphological characteristics
Presence of various enzymes
Serological tests
Phage typing
Fatty acid profiles
DNA finger printing
Sequence of ribosomal RNA
Is still very difficult

Morphological
Identification
characteristics:
Methods
Useful for
identifying
eukaryotes
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Differential
staining: Gram
staining, acidfast staining
Biochemical
tests:
Determines
presence of
bacterial
enzymes
A dichotomous key
Figure 10.8
A clinical microbiology lab report form
Figure 10.7
Identifying a Gram – Negative,
Oxidase – Negative Rod
Figure 10.8
•Unknown enteri
inoculated into tube
•After incubation the
15 tests are observed
•A numerical score is
assigned
•The species is
assigned
•This may be of is the
strain has changed
somehow. More tests
are required

Design a rapid test for a Staphylococcus
aureus. 10-14
Figure 6.10
Serology study of serum and its immune responce
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Combine known
antiserum +
unknown bacterium
Slide agglutination
ELISAp288, 514
Western blotp289
Southern Blot p292
DNA chip p293
Figure 10.10
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Strains with different antigens are called
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Serotypes
Serovars
biovars
DNA Hybridization
Phage Typing
Determining a
strains
suceptability to
certain phage or
bacterial viruses
Figure 10.13
The Western Blot
Flow Cytometry
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Uses differences
in electrical
conductivity
between species
Fluorescence of
some species
Cells selectively
stained with
antibody plus
fluorescent dye
Figure 18.12
A typical
dichotomous
key See
appendix H in
your lab book
Genetics
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DNA base
composition
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DNA fingerprinting
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Guanine + cytosine
moles% (GC)
Electrophoresis of
restriction enzyme
digests
rRNA sequencing
Polymerase Chain
Reaction (PCR)p251
Figure 10.14
Nucleic Acid Hybridization: DNA
DNA Chip
chip
Technolog
y
Figure 10.17
Differentiate between
classificaiton and
identification
Figure 10.5
Fluorescent
in situ
hybridization
 Add DNA
probe for S.
aureus

Figure 10.18
Dichotomous keys are used for
identification of organisms
Cladograms show phylogenetic
relationships among organisms
Using Bergies manual
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Used to Identify bacteria not classify
Features that are used to differentiate various
organism often have little to do with arranging
the orgs in taxonomic groups
4 major groups
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Domain Bacteria
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Gram-negative Eubacteria that have cell walls.
Proteobacteria
Non proteobacteria Gram negative bacteria
Gram positive Eubacteria that have cell walls
Domain archaeobacteria
Some groups by identificaiton
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Spirochetes
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Aerobic/microaerophilic, motikle, helical/vibroid gram negative
bacteria
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Genus lactococcus, streptococcus, Staphylococcus
Endospore forming gram positive rods and cocci

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Genus Enterobater, Escherichia, Klebsiella, Serratia, Shigella, Yersinia, Eikenella
Gram positive Cocci
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Genus Agrobacterium, Alcaligenes, Pseudomonas
Facultatively anaerobic gram negative rods
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Geneus Camphylobacter
Gram negative aerobic/microaerophilic rods and cocci
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Genus Borrelia, Leptospira, Treponema
Genus bacillus, clostridium
Regular, nonsporing gram positive rods
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Genus Lactobacillus, listeria