Diversity of Microbial World
Madam Noorulnajwa Diyana Yaacob
PPK BIOPROSES
April/May 2013
Course content ProkaryotesArchaeaBacteria Eukaryotes (microbial Protists)FungiAlgaeProtozoa Viruses
Introduction
Taxonomy is the science of the classification of organisms, with the goal of showing relationships among organisms.
Taxonomy also provides a means of identifying organisms.
consists of three separate but interrelated parts
classification – arrangement of organisms into groups (taxa; s., taxon)
nomenclature – assignment of names to taxa identification – determination of taxon to which an
isolate belongs
How would you classify?Types of classification: 1. natural2. polyphasic phenetic phylogenetic genotype
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Natural Classification arranges organisms into groups whose
members share many characteristics first such classification in 18th century
developed by Linnaeus based on anatomical characteristics
this approach to classification does not necessarily provide information on evolutionary relatedness
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Polyphasic Taxonomy
used to determine the genus and species of a newly discovered prokaryote
incorporates information from genetic, phenotypic, and phylogenetic analysis
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Phenetic Classification
groups organisms together based on mutual similarity of phenotypes (is the composite of an organism's observable characteristics or traits, such as its morphology, development, biochemical or physiological properties, phenology, behavior, and products of behavior)
can reveal evolutionary relationships, but not dependent on phylogenetic analysis i.e., doesn’t weigh characters
best systems compare as many attributes as possible
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Phylogenetic Classification also called phyletic classification
systems phylogeny
evolutionary development of a species usually based on direct comparison of
genetic material and gene productsWoese and Fox proposed using small
subunit (SSU) rRNA nucleotide sequences to assess evolutionary relatedness of organisms
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Genotypic Classification
comparison of genetic (inherited instructions it carries within its genetic code) similarity between organisms individual genes or whole genomes can be
compared70% homologous belong to the same
species
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Taxonomic Ranks - 1 microbes are placed in hierarchical
taxonomic levels with each level or rank sharing a common set of specific features
highest rank is domain Bacteria and Archaea – microbes only Eukarya – microbes and macroorganisms
within domain phylum, class, order, family, genus, species
epithet, some microbes have subspecies
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Species definition
collection of strains that share many stable properties and differ significantly from other groups of strains
also suggested as a definition of speciescollection of organisms that share the same
sequences in their core housekeeping genes
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Strains
descended from a single, pure microbial culture
vary from each other in many waysbiovars – differ biochemically and
physiologicallymorphovars – differ morphologicallyserovars – differ in antigenic properties
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Type Strain
usually one of first strains of a species studied
often most fully characterized not necessarily most representative
member of species
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Genus
well-defined group of one or more strains clearly separate from other genera often disagreement among taxonomists
about the assignment of a specific species to a genus
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Binomial System of Nomenclature
devised by Carl von Linné (Carolus Linnaeus) each organism has two names
genus name – italicized and capitalized (e.g., Escherichia)
species epithet – italicized but not capitalized (e.g., coli)
can be abbreviated after first use (e.g., E. coli) a new species cannot be recognized until it has
been published in the International Journal of Systematic and Evolutionary Microbiology
Techniques for Determining Microbial Taxonomy and Phylogeny
classical characteristicsmorphologicalphysiologicalbiochemicalecologicalgenetic
Ecological Characteristics
life-cycle patterns symbiotic relationships ability to cause disease habitat preferences growth requirements
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Molecular Approaches
extremely important because almost no fossil record was left by microbes
allows for the collection of a large and accurate data set from many organisms
phylogenetic inferences based on these provide the best analysis of microbial evolution currently available
Molecular Characteristics
nucleic acid base composition nucleic acid hybridization nucleic acid sequencing genomic fingerprinting amino acid sequencing
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Nucleic Acid Base Composition
G + C contentMol% G + C =
(G + C/G + C + A + T)100usually determined from melting
temperature (Tm)variation within a genus usually <10%
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Figure 17.2
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Nucleic Acid Hybridization DNA-DNA hybridization
measure of sequence homology common procedure
bind nonradioactive DNA to nitrocellulose filter
incubate filter with radioactive single-stranded DNA
measure amount of radioactive DNA attached to filter
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Table 17.4
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Nucleic Acid Sequencing
Small subunit rRNAs (SSU rRNAs)sequences of 16S and 18S rRNA most
powerful and direct method for inferring microbial phylogenies and making taxonomic assignments at genus level
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Comparative Analysis of 16S rRNA Sequences oligonucleotide signature sequences found
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 2 organisms, their relatedness is represented by percent sequence homology 70% is cutoff value for species definition
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Figure 17.3
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Genomic Fingerprinting used for microbial classification and
determination of phylogenetic relationships
requires analysis of genes that evolve more quickly than rRNA encoding genesmultilocus sequence analysis (MSLA)
the sequencing and comparison of 5 to 7 housekeeping genes is done to prevent misleading results from analysis of one gene
multilocus sequence typing (MLST) – discriminates among strains
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Restriction Fragment Length Polymorphism (RFLP) uses restriction enzymes to recognize
specific nucleotide sequencescleavage patterns are compared
ribotyping similarity between rRNA genes is determined
by RFLP rather than sequencing
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Figure 17.4
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Single Nucleotide Polymorphism (SNP)
Looks at single nucleotide changes, or polymorphisms, in specific genes
16S rRNA focuses on one specific gene Regions targeted because they are
normally conserved, so single changes in a base pair reveal evolutionary change
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Figure 17.5
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Amino Acid Sequencing amino acid sequences reflect mRNA
sequence and therefore of the gene which encodes that protein
amino acid sequencing of proteins such as cytochromes, histones, and heat-shock proteins has provided relevant taxonomic and phylogenetic information
cannot be used for all proteins because sequences of proteins with different functions often change at different rates
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Figure 17.6
The prokaryotes:
Domain Archaea
and
Domain Bacteria
1923,David Bergey (prof of bacteriology) published a classification of bacteria for identification of bacterial (and archaea) species.
Bergey’s Manual categorizes bacteria into taxa based on rRNA sequences.
Bergey’s Manual lists identifying characteristics such as Gram stain reaction, cellular morphology, oxygen requirements, and nutritional properties.
Bergey’s Manual of Systematic Bacteriology
The Archaea
Archaea
Scientist identified archaea as a distinct type of prokaryotes based on its unique rRNA sequence
Reproduce by : binary fusion, budding or fragmentation
Cells shape : cocci, bacilli, spiral, lobed, cuboidal etc
Not causing disease to humans/animals Cell wall contain proteins, glycoproteins,
lipoproteins, polysaccharides
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Archaeal Cell Surfaces cell envelopes
varied S layers attached to plasma membrane
pseudomurein (peptidoglycan-like polymer) complex polysaccharides, proteins, or
glycoproteins found in some other species only Ignicoccus has outer membrane
flagella closely resemble bacterial type IV pili
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Archaeal Membrane Lipids
differ from Bacteria and Eukarya in having branched chain hydrocarbons attached to glycerol by ether linkages
polar phospholipids, sulfolipids, glycolipids, and unique lipids are also found in archaeal membranes
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Archaeal Lipids and Membranes
Bacteria/Eukaryotes
fatty acids attached to glycerol by ester linkages
Archaea branched chain
hydrocarbons attached to glycerol by ether linkages
some have diglycerol tetraethers
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Figure 18.4
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Archaeal Taxonomy
two phyla based on Bergey’s ManualEuryarchaeotaCrenarchaeota
16S rRNA and SSU rRNA analysis also shows Group I are ThaumarchaeotaGroup II are Korachaeota
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Figure 18.1
Very high/low temp/pH, concentrated salts or completely anoxic (extreme environments)
Archae are either gram +ve or gram –ve Classified into two phylum :
1) Crenarchaeota
2) Euryarchaeota – 5 major physiologic groups (the metanogens, the halobacteria, the thermoplasms, extremely thermophilic S°-reducers and sulfate-reducing)
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Phylum Crenarchaeota most are extremely thermophilic
hyperthermophiles (hydrothermal vents) most are strict anaerobes some are acidophiles many are sulfur-dependent
for some, used as electron acceptor in anaerobic respiration
for some, used as electron source
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Figure 18.9
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Figure 18.10
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Crenarchaeota… include organotrophs and lithotrophs
(sulfur-oxidizing and hydrogen-oxidizing)
contains 25 genera two best studied are Sulfolobus and
Thermoproteus
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Genus Thermoproteus long thin rod, bent or branched
cell walls composed of glycoprotein thermoacidophiles
70–97 °C pH 2.5–6.5
anaerobic metabolism lithotrophic on sulfur and hydrogen organotrophic on sugars, amino acids, alcohols,
and organic acids using elemental sulfur as electron acceptor
autotrophic using CO or CO2 as carbon source
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Genus Sulfolobus irregularly lobed, spherical shaped
cell walls contain lipoproteins and carbohydrates
thermoacidophiles70–80°CpH 2–3
metabolism lithotrophic on sulfur using oxygen (usually)
or ferric iron as electron acceptororganotrophic on sugars and amino acids
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Phylum Euryarchaeota consists of many classes, orders, and
families often divided informally into five major
groupsmethanogenshalobacteria thermoplasmsextremely thermophilic S0-metabolizerssulfate-reducers
The Methanogens - strict anaerobes- obtain energy by converting CO2, H2, methanol to methane or methane & CO2- eg. Methanobacterium, Methanococcus
- methanogenesis* last step in the degradation of organic compounds
*occurs in anaerobic environments e.g., animal rumens e.g., anaerobic sludge digesters e.g., within anaerobic protozoa
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Ecological and Practical Importance of Methanogens important in wastewater treatment can produce significant amounts of
methane can be used as clean burning fuel and energy
source is greenhouse gas and may contribute to
global warming can oxidize iron
contributes significantly to corrosion of iron pipes
The Halobacteria- extreme halophiles- aerobic chemoorganotrophs (use organic compound as energy sources)- dependent on high salt content- cell wall dependent on NaCl, they disintegrated when [NaCl] < 1.5M- dead sea
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Strategies to Cope with Osmotic Stress increase cytoplasmic osmolarity
use compatible solutes (small organics) “salt-in” approach
use antiporters and symporters to increase concentration of KCl and NaCl to level of external environment
acidic amino acids in proteins
The Thermoplasms- lack cell walls - but plasma membrane strengthen by diglycerol tetraether, lipopolysaccharides, and glycoproteins- grow best at 55-59°C, pH1-2- eg. Thermoplasma
Extremely Thermophillic So-Reducers- strictly anaerobic- can reduce sulfur to sulfide- grow best at 88-100°C- motile by flagella- eg. Thermococcus
Sulfate-reducing
- irregular garm –ve coccoid cellscell walls consist of glycoprotein subunits
- extremely thermophilicoptimum 83°C isolated from marine hydrothermal vents
- obtain their energy by oxidizing organic compounds or H2 while reducing sulfates to sulfides. In a sense, they "breathe" sulfate rather than oxygen
- eg. Archaeoglobus
Bacteria
Domain Bacteria
Bacteria are essential to life on Earth.
We should realize that without bacteria, much of life as we know it would not be possible.
In fact, all organisms made up of eukaryotic cells probably evolved from bacterialike organisms, which were some of the earlist forms of life.
The Proteobacteria Largest group of bacteria. More than 500 genera gram-negative, some motile using flagella Most are facultative/obligate anaerobes Share common 16s rRNA sequence 5 distinct classes of proteobacteria (α,β, ε, ɣ,δ) :
- Alphaproteobacteria - Betaproteobacteria- Gammaproteobacteria- Deltaproteobacteria- Epsilonproteobacteria
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Figure 20.1
Alphaproteobacteria Gram -ve Most are oligotrophic (capable of growing at low nutrient
levels) Example of alphaproteobacteria ;
1) Most purple nonsulfur phototrophs are in this group (use light energy and CO2 and do not produce O2)
2) Nitrifying bacteria e.g. Nitrobacter (oxidize NH3 to NO3 by a process called nitrification)3) Pathogenic bacteria eg. Rickettsia (typhus), Brucella (brucellosis), Ehrlichia (ehlichiosis)4) Beneficial bacteria eg. Acetobacter and Caulobacter (synthesize acetic acid); Agrobacterium (used in genetic recombination in plants)
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Purple Nonsulfur Bacteria with one exception (genus
Rhodocyclus) all are a-proteobacteria metabolically flexible
normally grow anaerobically as anoxygenic photoorganoheterotrophs
possess bacteriochlorophylls a or b in photosystems located in membranes that are continuous with plasma membrane
some can oxidize sulfide, but not elemental sulfur, to sulfate
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Purple Nonsulfur Bacteria…
Rhodospirillum industrial importanceproduces H2 novel biodegradable plasticoxidize carbon monoxide to carbon dioxide
morphologically diversemost motile by polar flagella
found in mud and water of lakes and ponds with abundant organic matter and low sulfide levels; some marine species
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Nitrifying Bacteria
very diverse chemolithoautotrophsnitrification – gain electrons from oxidation
of ammonium to nitrate or nitrite nitrite further oxidized to nitrate
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Nitrification ammonia nitrite nitrate conversion of ammonia to nitrate by
action of two generae.g., Nitrosomonas – ammonia to nitritee.g., Nitrobacter – nitrite to nitrate
fate of nitrateeasily used by plants lost from soil through leaching or
denitrification
Betaproteobacteria Gram –ve oligotrophic (capable of growing at low nutrient
levels) Differ with alphaproteobacteria in rRNA sequence Example of betaproteobacteria :
1) nitrifying bacteria eg. Nitrosomonas
2) pathogenic species, Neisseria (gonorrhea), Bordetella (whooping cough)
3) Thiobacillus (ecologically important), Zoogloea (sewage treatment)
Gammaproteobacteria – largest class purple sulfur bacteria – obligate anaerobes that
oxidize hydrogen sulfide to sulfur intracellular pathogens (Legionella, Coxiella), methane oxidizers (Methylococcus), facultative anaerobes that utilize glycolysis and
the pentose phosphate pathway (Escherichia coli),
pseudomonads –aerobes that catabolize carbohydrates (Pseudomonas, and Azomonas)
Deltaproteobacteria Sulfate reducing microbes Eg. Desulfovibrio
(important in the sulfur cycle) Myxobacteria – gram negative, soil-dwelling
bacteria , dormant myxospores; common worldwide in the soils having decaying plant material or dung
Epsilonproteobacteria Gram-negative rods, vibrios, or spiral Include important human pathogens Eg. Campylobacter (causes blood poisoning)
The Gram Positive Bacteria
In Bergey’s Manual, gram-positive bacteria (able to form endospore) are divided into those that have :- low G + C ratio (base pair in genome below 50%)- high G + C ratio
Low G + C gram-positive bacteria include 3 groups clostridia, mycoplasms, Gram-positive Bacilli and Cocci
High G + C gram-positive bacteria include mycobacteria, corynebacteria, and actinomycetes.
Clostridia Eg. Clostridium – anaerobic, form
endospores, rod shape, gram +ve pathogenic bacteria causing gangrene,
tetanus, botulism, and diarrhea
Mycoplasmas Facultative or obligate anaerobes lack cell walls Gram +ve (previously under gram negative
category until nucleic acid sequences proved similarity with gram positive organisms)
When culture on agar, form ‘fried egg’ appearance bcoz cell in the center of the colony grow into the agar while those around the spread outward
Usually associated with pneumonia and urinary tract infections
Fried egg appearance
Gram positive Bacilli and Cocci Eg. Bacillus – form endospores, flagella
(B.licheniformis synthesis antibiotic. B.anthracis cause anthrax)
Eg. Lactobacillus – nonsporing rods, nonmotile, produce lactic acid as fermentation product. Mostly found in human mouth, intestinal tract, stomach. Protect body from pathogens
Streptococcus – nonmotile, cocci associated in pairs and chain. Cause pneumonia, scarlet fever
StreptococcusBacillus
High G+C gram-positive bacteria Include Corynebacterium, Mycobacterium and
Actinomycetes that have a G+C ratio > 50% in the phylum Actinobacteria, which have species with rod-shaped cells
Corynebacterium store phosphates in metachromatic granules. C. diptheria causes diphtheria
Mycobacterium cause tuberculosis and leporosy. It has unique resistant cell walls containing mycolic acids. Hence, acid fast stain (for penetrating waxy cell walls) is used for its identification
Actinomycetes resemble fungi as they produce spores and form filaments; important genera: Actinomyces found in human mouths; Nocardia useful in degradation of pollutants; and Streptomyces produces antibiotics
THE EUKARYOTES :FUNGI, ALGAE,
PROTOZOA
FUNGI Organisms in kingdom fungi include molds,
mushrooms, yeasts Fungi are aerobic or facultatively anaerobic
(yeast), chemoheterotrophs, spore-bearing, lack chlorophyll
Most fungi are decomposers, and a few are parasites of plants and animals
Some fungi – cause disease (mycoses) Some fungi – essential to many industries
(bread, wine, cheese, soy sauce)
Characteristics of Fungi Body/vegetative struc. of fungi – Thallus Thalli of yeast – small, globular, single cell Thalli of mold – large, composed of long,
branched, threadlike filaments of cell called hyphae that form mycelium
Hyphae - septate- Aseptate (coenocytic)
Fungi grow best in the dark, moist habitats
Acquire nutrients by absorption. Secrete enzyme to break large organic mol. Into simple mol.
Reproduction of fungi – sexual & asexual
Asexual reproduction
Several ways :
1) Transverse fission - Parent cell undergo mitosis, divide into daughter cell by formation of new cell wall
2) Budding – after mitosis, one daughter nucleus is sequestered in a small bleb that is isolated from parent cell by formation of cell wall
3) Asexual spore formation - filamentous fungi produce asexual spores through mitosis and subsequent cell division.several types of asexual spores :1) Sporangiospores form inside a sac called sporangium2) Chlamydospores form with a thickened cell wall inside hyphae3) Conidiospores (conidia) produced at the tip or side of hyphae, not within sac4) Blastospores produced from vegetative mother (hyphae) cell by budding5) Arthrospores hyphae that fragment into individual spores
sporangiospores
conidiospores
Chlamydiospores
Blastospores
Arthrospores
chlamydospores
sporangiospores
conidiospores
4) Sexual reproduction in fungi
Fungal mating type designated as + and –.
4 basic steps :
1) Haploid (n) cells from + and – thallus fuse, form dikaryon (cell with both +&- nuclei)
2) pair of nuclei within a dikaryon fuse to form one diploid (2n) nucleus
3) meiosis of the diploid restores the haploid state
4) haploid nuclei partitioned into + and - spores
Classification of fungi
1) Zygomycota Coenocytic molds – zygomycetes produce sporangiospores (asexual) and
zygospores (sexual) e.g. Black bread mold Rhizopus nigricans
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usually reproduce asexually by spores that develop at the tips of aerial hyphae
sexual reproduction occurs when environmental conditions are not favorable requires compatible opposite mating types hormone production causes hyphae to produce
gametes gametes fuse, forming a zygote zygote becomes zygospore
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Figure 24.4
2) Ascomycota Septate hyphae Form ascospores within sac-like structure
call asci (sexual) Form conidiospores in asexual
reproduction Eg Penicillium
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Ascomycota ascomycetes or sac fungi
found in freshwater, marine, and terrestrial habitats
red, brown, and blue-green molds cause food spoilage
some are human and plant pathogenssome yeasts and truffles are ediblesome used as research tools
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Figure 24.6
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Ascomycota Yeast Life Cycle alternates between haploid and diploid
in nutrient rich, mitosis and budding occurs at non-scarred regions
stops after entire mother cell is scarred
nutrient poor, meiosis and haploid ascus containing ascospores formed
haploid cells of opposite mating types fuse tightly regulated by pheromones
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Figure 24.7
3) Basidiomycota septate hyphae produce basidiospores (sexual), some produce
conidiospores (asexual) Eg mushrooms, puffballs, stinkhorns
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Figure 24.12
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Human Impact Basidiomycota decomposers edible and non-edible mushrooms
toxins are poisons and hallucinogenic pathogens of humans, other animals,
and plantse.g., Cryptococcus neoformans –
cryptococcosissystemic infection, primarily of lungs
and central nervous system
Protozoa
Eukaryotic, unicellular and lack of cell wall Motile (cilia, flagella, pseudopodia) Grow in moist habitats Some are in group of Planktonic (floating free in
lakes, ocean and form the basis of aquatic food chain)
Some protozoa can produce a cyst that provides protection during adverse environmental conditions
Asexuall reproduction by binary fission, schizogony/multiple fission
Sexually reproduction by conjugation
1)Nucleus undergoes mitosis2)Cytoplasm divides by cytokinesis
Schizogony/multiple fission
Classification of protozoa
Grouping based on locomotive structure do not reflect genetic relationship.
7 taxa of protozoa : alveolates, cercozoa, radiolaria, amoebozoa, eglenozoa, diplomonads, and parabasalids
Alveolates Have small membrane cavities called
alveoli beneath cell surface. 3 groups : ciliates (have cilia),
apicomplexans (pathogen to animal), dinoflagellates (have flagella)
Cercozoa Unicellular, called amoeba Move & feed by pseudopodia Have snail-like shells of calcium carbonate
Radiolaria Amoeba that have ornate shells composed of
silica
Amoebozoa Have lobe-shaped pseudopodia, no shell Eg Acanthamoeba, Naegleria
Eglenozoa move by means of flagella and lack sexual
reproduction; they include Trypanosoma
Diplomonad Lack mitocondria, golgi bodie Have 2 nuclei and multiple flagella
Giardia
Algae
Simple eukaryotic, phototrophic organisms, like plants
Carry out photosynthesis using chlorophyll Most live in aquatic environments
Characteristic of Algae
Unicellular or simple multicellular (thalli) Thallus of seaweed (large marine algae)
are complex, with holdfast (attached to rock), stemlike stipes and leaflike blades
Algae reproduce sexual and asexual (fragmentation & cell division)
Classification of algae Classify according to their structure and
pigment :
- red algae
- brown algae – cell wall composed of cellulosa & alginic acid
- green algae
- diatoms – silica cell wall composed of two halves called frustules that fit together like petri dishes
DIATOMS
frustule
VIRUSES
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Virus Classification
classification based on numerous characteristicsnucleic acid typepresence or absence of envelopcapsid symmetrydimensions of viron and capsid
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Figure 25.2
Characteristics of Viruses Viral disease – SARS, AIDS, influenza, herpes, common
cold Viruses – miniscule, infectious agent with simple acellular
organization and pattern of reproduction Viruses can exist – extracellular or intracellular Virion (complete virus particle) consist of :
- nucleocapsid (composed of 1 or more DNA or RNA, held within capsid)- in some viruses – envelope (phospholipid membrane)
Capsid - build by few types of protein = protomer - 3 types – helical, icosahedral, complex
symmetry
Virus structure
Different types of virus
Helical Helical Icosahedral Complex symmetry
Types of capsids
Virus size range 10-1000nm Most virus infect only particular host’s cells
Eg. HIV only infect T lymphocytes (a type of white blood cell)
Some viruses infect many kinds of cells in many different hosts
Eg. Rabies can infect most mammals Viruses are obligatory intracellular parasites.
They multiply by using the host cell’s synthesizing machinery to cause the synthesis of specialized elements that can transfer the viral nucleic acid to other cells
Viral replication
Virus cannot reproduce themselves bcoz:- have no genes for all enzyme needed for replication- have no ribosomes for protein synthesis
Viruses dependent of host’s organelles and enzymes to replicate
Virus replication – Lytic replication
Lytic replication of bacteriophage Consist of 5 stages – attachment, entry,
synthesis, assembly, release
1) Attachment – structure responsible for attachment to host = tail fiber. Attachment is dependent on chemical attraction and precise fit between T4 tail and protein receptor on E.coli cell wall
2) Entry – T4 release lysozyme to weaken peptidoglycan of E.coli cell wall. T4 inject genome into E.coli, leaving T4 coat outside.
3-4) Synthesis – viral enzyme degrade the bacterial DNA. E.coli start synthesis new viruses. T4 DNA is transcribed, producing mRNA which is translated to T4 protein (component of tail and head, lysozyme)
5) Assembly – T4 components are assemble in spontaneous manner to form mature virion
6) Release – newly assembled virions are released from the cell as lysozyme completes its work on the cell wall
Lytic replication takes about 25min can produce 100-200 new virions each cycle
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