Microbiology Final Exam Test Answers – Flashcards
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| Genetic information flow can be divided into three stages |
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| • Replication: DNA is duplicated • Transcription: information from DNA is transferred to RNA - mRNA (messenger RNA): encodes poly peptides - tRNA (transfer RNA): plays role in protein synthesis - rRNA (ribosomal RNA): plays role in protein synthesis • Translation:information in RNA Is used to build polypeptides |
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| mRNA (messenger RNA): |
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| encodes poly peptides |
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| tRNA (transfer RNA) |
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| plays role in protein synthesis |
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| rRNA (ribosomal RNA) |
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| plays role in protein synthesis |
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| The Double Helix |
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| All cells and some viruses have DNA in double-stranded molecule • Four nucleotides are found in DNA(Figure4.1): • Adenine (A) • Guanine (G) • Cytosine (C) • Thymine (T) • Backbone of DNA chain is alternating phosphates and the pentose sugar deoxyribose • Phosphates connect 3?-carbon of one sugar to 5?-carbon of the adjacent sugar • Two strands are antiparallel • Two strands have complementary base sequences • Adenine always pairs with thymine • Guanine always pairs with cytosine • Two strands form a double helix • Size of DNA molecule is expressed in base pairs • 1,000 base pairs = 1 kilobase pair = 1 kbp • 1 million base pairs = 1 megabase pair =1Mbp • E.coli genome = 4.64 Mbp • Each base pair takes up 0.34 nm of length along the helix • 10 base pairs make up 1 turn of the helix |
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| Supercoiled DNA |
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| DNA's further twisted to save space • Negative supercoiling: double helix is underwound • Positive supercoiling: double helix is overwound • Relaxed DNA: DNA has number of turns predicted by number of base pairs • Negative supercoiling is predominantly found in nature • DNA gyrase: introduces supercoils into DNA |
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| DNA gyrase |
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| : introduces negative supercoils into DNA by making double strand breaks (type II topoisomerase) |
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| Chromosome |
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| is a genetic element with "housekeeping" genes • Presence of essential genes is necessary for a genetic element to be called a chromosome -typical proks have a single, circular DNA chromosome containing all (or most) genes found inside the cell |
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| Plasmid |
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| is a genetic element that is expendable and rarely contains genes for growth under all conditions -replicate separately from chromosome -double stranded and circular, much smaller then chromosomes OTHER SLIDE: • Small circular or linear DNA molecules • Range in size from 1 kbp to >1 Mbp; typically less than 5% of the size of the chromosome • Carry a variety of nonessential, but often very helpful, genes • Abundance (copy number) is variable • A cell can contain more than one plasmid • Genetic information encoded on plasmids is not essential for cell function under all conditions BUT may confer a selective growth advantage under certain conditions |
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| Transposable elements |
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| • Segment of DNA that can move from one site to another site on the same or a different DNA molecule • Inserted into other DNA molecules -play important role in genetic variation • Three main types: • Insertion sequences • Tr a n s p o s o n s • Special viruses |
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| E. Coli |
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| Escherichia coli is a useful model organism for the study of biochemistry, genetics, and bacterial physiology • The E. coli chromosome from strain MG1655 has been mapped using conjugation, transduction, molecular cloning, and sequencing • Some features of the E. coli chromosome - Many genes encoding enzymes of a single biochemical pathway are clustered into operons - Operons are equally distributed on both strands -~5Mbpinsize - ~40% of predicted proteins are ofunknown function - Average protein contains ~300 amino acids - Insertion sequences (IS elements) |
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| R plasmids |
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| • Resistance plasmids; confer resistance to antibiotics and other growth inhibitors • Widespread and well-studied group of plasmids • Many are conjugative • In several pathogenic bacteria, virulence characteristics are encoded by plasmid genes • Virulence factors - Enable pathogen to colonize - Enable pathogen to cause host damage - Hemolysin: destroys red blood cells - Enterotoxin-toxin affecting intestines |
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| DNA replication |
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| is semiconservative • Each of the two progeny double helices have one parental and one new strand • Precursor of each nucleotide is a deoxynucleoside 5?- triphosphate (dNTP) • Replication ALWAYS proceeds from the 5? end to the 3? end |
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| DNA polymerases |
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| catalyze the addition of dNTPs • Five different DNA polymerases in E. coli - DNA polymerase III is primary enzyme replicating chromosomal DNA • DNA polymerases require a primer - Primer made from RNA by primase *DNA synthesis begins at the origin of replication in prokaryotes is unwound by DNA helicase and Extension of DNA Occurs continuously on the leading strand and Discontinuously on the lagging strand • Okazaki fragments are on lagging strand |
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| Replication fork |
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| : zone of unwound DNA where replication occurs |
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| DNA helicase |
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| unwinds the DNA |
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| DNA gyrase |
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| replaces supercoils ahead of ribosome |
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| single stranded binding proteins |
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| prevent single strands from annealing |
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| DNA synthesis in prokaryotes |
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| is bidirectional in prokaryotes • Two replication forks moving in opposite direction -DNA loops out allowing the replisome to move smoothly along both strands (DNA, not DNA polymerase is actually moving) -in addition to RNA Pol III, 1) DNA gyrase removes supercoils, 2) DNA helicase and primase (the primosome) unwind the DNA and 3) SSBP prevent the template strands from rewinding |
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| Replisome |
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| : complex of multiple proteins involved in replication of proks • DNA pulled through the replisome |
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| Proofreading |
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| • DNA replication is extremely accurate • Proofreading helps to ensure high fidelity • Mutation rates in cells are 10–8 to 10–11 errors per base inserted • Polymerase can detect mismatch through incorrect hydrogen bonding • Proofreading occurs in prokaryotes, eukaryotes, and viral DNA replication systems |
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| Transcription |
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| • Only one of the two strands of DNA is transcribed by RNA polymerase for any gene • Genes are present on both strands of DNA, but at different locations • Promoter: site of initiation of transcription • Recognized by sigma factor of RNA polymerase • Transcription stops at specific sites called transcription terminators • Unlike DNA replication, transcription involves smaller units of DNA • Often as small as a single gene • Allows cell to transcribe different genes at different rates |
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| Promoter: |
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| site of initiation of transcription |
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| sigma factor |
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| recognize two highly conserved regions of promoter • Two regions within promoters that are highly conserved: -Pribnow box (TATAAT box): located 10 bases before the start of transcription (–10 region) - –35 region: located ~35 bases upstream of transcription |
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| transcription terminators |
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| Termination of RNA synthesis is governed by a specific DNA sequence • Intrinsic terminators: transcription is terminated without any additional factors • Rho-dependent termination: Rho protein recognizes specific DNA sequences and causes a pause in the RNA polymerase |
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| Unit of transcription |
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| : unit of chromosome bounded by sites where transcription of DNA to RNA is initiated and terminated • Most genes encode proteins, but some RNAs are not translated (i.e., rRNA, tRNA) - Three types of rRNA:16S,23S,and5S - rRNA and tRNA are very stable - tRNA cotranscribed with rRNA or other tRNA • mRNAs have short half-lives (a few minutes) • Prokaryotes often have genes clustered together • These genes are transcribed all at once as a single mRNA • An mRNA encoding a group of cotranscribed genes is called a polycistronic mRNA (Figure 4.25) • Operon: a group of related genes cotranscribed on a polycistronic mRNA -Allows for expression of multiple genes to be coordinated |
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| Operon |
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| : a group of related genes cotranscribed on a polycistronic mRNA -Allows for expression of multiple genes to be coordinated |
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| Archaea RNA polymerase |
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| The Archaea contain only a single RNA polymerase •Resembles eukaryotic polymerase II • Archaea have a simplified version of eukaryotic transcription apparatus -Archaea Promoters and RNA polymerase similar to eukaryotes -Regulation of transcription has major similarities with Bacteria |
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| Eukaryotic genes have coding and noncoding regions |
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| • Exons are the coding sequences (STAYS IN) • Introns are the intervening sequences (CUT OUT) - Are rare in Archaea - Are found in tRNA and rRNA genes of Archaea • Archaeal introns excised by special endonuclease |
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| Proteins |
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| Proteins play a major role in cell function • Catalytic proteins (enzymes) • Structural proteins • Proteins are polymers of amino acids • Amino acids are linked by peptide bonds to form a polypeptide |
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| Translation |
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| : the synthesis of proteins from RNA 1) Initiation 2) elongation 3) termination |
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| Genetic code |
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| : a triplet of nucleic acid bases (codon) encodes a single amino acid • Specific codons for starting and stopping translation • Degenerate code: multiple codons encode a single amino acid • Anticodon on tRNA recognizes codon • Wobble: irregular base pairing allowed at third position of tRNA |
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| Degenerate code |
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| : multiple codons encode a single amino acid |
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| Wobble |
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| : irregular base pairing allowed at third position of tRNA |
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| Stop codons |
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| : terminate translation (UAA, UAG, and UGA) |
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| Start codon |
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| : translation begins with AUG |
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| Reading frame |
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| triplet code requires translation to begin at the correct nucleotide |
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| Shine–Dalgarno sequence |
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| : ensures proper reading frame |
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| Open reading frame (ORF) |
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| : AUG followed by a number of codons and a stop codon in the same reading frame |
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| Codon bias |
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| : multiple codons for the same amino acid are not used equally - Varies with organism - Correlated with tRNA availability - Cloned genes from one organism may not be translated by recipient organism because of codon bias • Some organelles and a few cells have slight variations of the genetic code (e.g., mitochondria of animals, Mycoplasma, and Paramecium) |
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| Transfer RNA |
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| : at least one tRNA per amino acid - Bacterial cells have 60 different tRNAs - Mammalian cells have 100–110 different tRNAs • Specific for both a codon and its cognate amino acid • tRNA and amino acid brought together by aminoacyl-tRNA synthetases • ATP is required to attach amino acid to tRNA • tRNA is cloverleaf in shape |
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| Ribosomes |
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| : sites of protein synthesis • Thousands of ribosomes per cell • Composed of two subunits (30S and 50S in prokaryotes) • S = Svedberg units • Combination of rRNA and protein • E. coli has 52 distinct ribosomal proteins |
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| Initiation |
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| : two ribosomal subunits assemble with mRNA • Beings at an AUG start codon • Shine Delgarno sequence helps placement of 16S rRNA |
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| Elongation |
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| : amino acids are brought to the ribosome and are added to the growing polypeptide • Occurs in the A and P sites of ribosome • Translocation: movement of the tRNA holding the polypeptide from the A to the P site |
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| Termination |
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| :occurs when ribosome reaches a stop codon • Release factors (RF): recognize stop codon and cleave polypeptide from tRNA • Ribosome subunits then dissociate • Subunits free to form new initiation complex and repeat process |
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| Polysomes |
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| :a complex formed by ribosomes simultaneously translating mRNA(multiple ribosomes translating a single mRNA molecule at the same time) *LOOKS LIKE CHRISTMAS TREE |
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| Protein Folding |
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| • Once formed, a polypeptide folds to form a more stable structure. • Secondary structure: Interactions of the R groups force the molecule to twist and fold in a certain way (Alpha helices and Beta-Sheets) • Tertiary structure: Three-dimensional shape of polypeptide • Quaternary structure: Number and types of polypeptides that make a protein |
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| Denaturation |
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| • Occurs when proteins are exposed to extremes of heat, pH, or certain chemicals - Causes the polypeptide chain to unfold - Destroys the secondary, tertiary, and/or quaternary structure of the protein - The biological properties of a protein are usually lost when it is denatured - Most polypeptides fold spontaneously into their active form - Some require assistance from molecular chaperones or chaperonins for folding to occur - They only assist in the folding; they are not incorporated into protein -Can also aid in refolding partially denatured proteins |
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| Signal sequences |
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| : found on proteins requiring transport from cell • 15–20 residues long • Found at the beginning of the protein molecule • Signal the cell's secretory system (Sec system) • Prevent protein from completely folding |
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| Secretion of folded proteins |
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| : the Twin Arginine Translocase • Tat system • Proteins that fold in the cytoplasm are exported by a transport system distinct from Sec, called the Tat protein export system • Iron–sulfur proteins • Redox proteins • Metabolic proteins - Other secretion systems: • Secretion of proteins: types I through VI (Figure 4.43) • All are a large complex of proteins that form channels through membranes • Types II and V depend on Sec or Tat • Types I, III, IV, and VI do not require Sec or Tat |
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| Genome |
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| • Entire complement of genetic • Includes genes, regulatory sequences, and noncoding DNA -Majority of genes encode proteins |
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| Genomics |
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| • Discipline of mapping, sequencing, analyzing, and comparing genomes • Field driven by cost/genome • More expensive to analyze data than to re-sequence *first human genome costed 3 billion dollars! • RNA virus MS2- First genome; sequenced in 1976 - 3,569 bp • Haemophilus influenzae- First cellular genome sequenced in 1995 -1,830,137 bp • Human Genome - Rough draft in 2000 - Polished draft in 2003 - ~3billion bp |
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| DNA sequencing using the Sanger method |
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| add DNA pol to mix of all four deoxyribonucleotide triphosphates, and separate into 4 reaction tubes. reaction products are separated by gel electrophoresis on gel and identified by autoradiography (largest fragments on top and shortest on bottom). In automated sequencing, each base has its own fluorescent dye. |
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| Virtually all genomic sequencing projects use shotgun sequencing |
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| • Entire genome is cloned, and resultant clones are sequenced • Much of the sequencing is redundant • Generally 7-to 10-fold coverage • Computer algorithms are used to look for replicate sequences and assemble them |
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| Genome assembly |
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| consists of connecting the DNA fragments in the correct order • Occasionally assembly is not possible • Closure can be pursued using PCR to target areas of the genome • Closed vs. draft genome - Closed genome relies on manpower - More expensive - More information |
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| Annotation |
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| : converting raw sequence data into a list of genes present in the genome • Annotation is "bottleneck" in genomics -net result is just "a list of parts" (thats why functional genomics is more useful) |
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| Bioinformatics |
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| • Science that applies powerful computational tools to DNA and protein sequences • For the purpose of analyzing, storing, and accessing the sequences for comparative purposes |
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| Functional ORF |
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| : an open reading frame that encodes a protein • Computer algorithms used to search for ORFs • Look for start/stop codons and Shine–Dalgarno sequences • ORFs can be compared to ORFs in other genomes STEPS- 1) comp finds possible start codons 2)comp finds possible stop codons 3) comp counts codons between start and stop 4) comp fins possible ribosomal binding site, if found the correct distance in front of the reading frame-the probability of a genuine ORF is stronger 5)comp calculates codon bias in ORF 6) comp decides if ORF is likely to be genuine 7) list of probable ORF's |
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| Hypothetical proteins |
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| : uncharacterized ORFs; proteins that likely exist but whose function is currently unknown • 40-70% of genomes are hypothetical! • Likely encode nonessential genes • In E. coli, many predicted to encode regulatory or redundant proteins |
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| Noncoding RNA: |
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| RNA that does not code for protein • Lack start codons and have multiple stop codons • Examples • TransferRNA(tRNA) • RibosomalRNA(rRNA) • Noncoding regulatory RNA molecules |
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| Genome Size and Content |
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| • Complement of genes in a particular organism defines its biology, but genomes are also molded by an organism's lifestyle • positive correlation between genome size and ORFs • On average, a prokaryotic gene is 1,000 bp long • ~1,000 genes per megabase (1 Mbp = 1,000,000 bp) • As genome size increases, gene content proportionally increases Smallest cellular genomes belong to parasitic or endosymbiotic prokaryotes • Obligateparasitesrangefrom 490 kbp (Nanoarchaeum equitans) to 4,400 kbp (Mycobacterium tuberculosis) • Endosymbiontscanbe smaller (e.g., 160-kbp genome of Carsonella ruddii) • Estimates suggest the minimum number of genes for a viable cell is 250–300 genes • Largest prokaryotic genomes comparable to those of some eukaryotes • Sorangium cellulosum (Bacteria) • Largest prokaryotic genome to date at 12.3 Mbp • Largest archaeal genomes tend to be smaller (~5 Mbp) |
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| Genome reconstruction |
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| • Identify genes in metabolic pathways • Many genes can be identified by sequence similarity to genes found in other organisms (comparative analysis) • Comparative analyses allow for predictions of metabolic pathways and transport systems • Example:Thermotoga maritima |
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| Known chloroplast genomes |
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| • Circular DNA molecules • Typically 120–160 kbp • Contain two inverted repeats of 6– 76 kbp • Many genes encode proteins for photosynthesis and autotrophy • Introns common; primarily of self- splicing type |
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| Known mitochondrial genomes |
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| • Diverse structures; some linear • Typically smaller than chloroplast genomes • Primarily encode proteins for oxidative phosphorylation • Usesimplifiedgeneticcodes rather than "universal" code • Some contain small plasmids • Mammalian mitochondria encode 13 proteins |
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| Transcriptome |
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| • The total RNA produced in an organism under a given set of conditions • measure gene expression • Microarrays: small solid supports to which genes or segments of genes are fixed and arrayed spatially in a known pattern; often called gene chips • RNAseq (RNA sequencing) • What can be learned from RNA experiments- Global gene expression - Expression of specific groups of genes under different conditions - Expression of genes with unknown function; can yield clues to possible roles - Comparison of gene content in closely related organisms - Identification of specific organisms |
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| Proteomics |
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| • Genome-wide study of the structure, function, and regulation of an organism's proteins - methods in proteomics • Two-dimensional (2-D) polyacrylamide gel electrophoresis - Technique for separating, identifying, and measuring all proteins present in a sample - In first (horizontal) dimension, proteins are separated by differences in isoelectric points - In second(vertical) dimension, proteins are separated by size • Computational techniques - Sequence the genome of the organism - Compare to genomes of other organisms - Identify similar genes - Different DNA sequence may not change protein sequence • Proteins with >50% sequence similarity typically have similar functions • Proteins with >70% sequence similarity almost certainly have similar functions • Protein domains - Distinct structural modules within proteins - Have characteristic functions that can reveal much about a protein's role, even in the absence of complete sequence homology |
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| Interactome |
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| • Complete set of interactions among molecules • Data expressed in the form of network diagrams |
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| Metabolome |
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| • The complete set of metabolic intermediates and other small molecules produced in an organism • Mass spectrometry is one of the primary techniques for monitoring metabolites • MALDI:Matrix assisted laser desorption ionization • TOF:Time of flight |
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| Systems biology |
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| • Integrationof different fields of research (Figure 6.24) • Genomics • Proteonomics • Transcriptonomics • Metabolonomics • Other • Compares data and builds a computer model of the system being studied |
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| Metagenome |
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| • The total gene content of the organisms present in an environment • Several environments have been surveyed by large-scale metagenome projects • Examples: acid mine runoff waters, deep-sea sediments, fertile soils |
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| Homologous |
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| :related sequence that implies common genetic ancestry |
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| Gene families |
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| groups of gene homologs |
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| Paralogs |
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| : genes within an organism whose similarity to one or more genes in the same organism is the result of gene duplication (think 'parallel' to each other because there is two of the same genes) |
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| Orthologs |
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| : genes found in one organism that are similar to those in another organism but differ because of speciation (think 'straight' from the same ancestor) |
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| Gene Duplication |
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| • Gene duplications thought to be mechanism for evolution of most new genes • Deletions can eliminate gene no longer needed • Gene analysis in the three domains of life suggests that many genes present in all organisms have common evolutionary roots |
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| Horizontal gene transfer (happens in proks) |
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| • The transfer of genetic information between organisms, as opposed to vertical inheritance from parental organism(s) • May be extensive in nature • May cross phylogenetic domain boundaries • Detecting horizontal gene flow - Presence of genes typically found only in distantly related species - Presence of a DNA with Guanosine/Cytosine content or codon bias that differs significantly from remainder of genome • Horizontally transferred genes typically do not encode core metabolic functions |
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| Transposons |
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| (most common form of mobile DNA) :pieces of DNA that can move between chromosome, plasmids, and viruses between different host DNA molecules. (can transfer genes like resistance to antibiotics or toxin production) • Transposons may transfer DNA between different organisms • Transposons may also mediate large-scale chromosomal changes within a single organism • Presence of multiple insertion sequences (IS) • Recombination among identical IS can result in chromosomal rearrangements • Examples: deletions, inversions, or translocations |
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| Insertion sequences |
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| —pieces of transposable DNA whose genes encode only transposition -contributed to the evolution of several bacterial pathogens |
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| Chromosomal islands |
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| :Region of bacterial chromosome of foreign origin that contains clustered genes for some extra property such as virulence or symbiosis - Pathogenicity islands: chromosomal islands containing genes for virulence • Chromosomal islands contribute specialized functions not essential to growth -Virulence -Biodegradation of recalcitrant compounds -For example, hydrocarbons and herbicides Symbiosis |
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| Pan and Core genomes |
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| genomes of bacterial species consist of two components |
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| Core genome |
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| : shared by all strains of the species |
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| Pan genome |
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| : includes all the optional extras present in some but not all strains of the species |
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| Isolation |
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| • The separation of individual organisms from the mixed community |
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| Enrichment cultures |
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| • Select for desired organisms through manipulation of medium and incubation condition • Can prove the presence of an organism in a habitat • Cannot prove that an organism does not inhabit an environment ** The ability to isolate an organism from an environment says nothing about its ecological significance |
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| Inoculum |
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| • The sample from which microorganisms will be isolated |
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| Enrichment bias |
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| • Microorganisms cultured in the lab are frequently only minor components of the microbial ecosystem • Reason: the nutrients available in the lab culture are typically much higher than in nature • Dilution of inoculum is performed to eliminate rapidly growing, but quantitatively insignificant, weed species |
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| The Winogradsky column |
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| • Anartificial microbial ecosy stem (Figure 18.2) • Servesasalong- term source of bacteria for enrichment cultures • Named for Sergei Winogradsky • First used in late 19th century to study soil microorganisms |
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| Pure cultures |
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| contain a single kind of microorganism • Can be obtained by streak plate, agar shake, or liquid dilution |
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| Axenic culture |
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| can be verified by • Microscopy • Observation of colony characteristics • Tests of the culture for growth in other media |
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| Agar dilution tubes |
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| are mixed cultures diluted in molten agar • Useful for purifying anaerobic organisms |
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| Most-probable-number technique |
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| • Serial 10x dilutions of inoculum in a liquid medium • Used to estimate number of microorganisms in food, wastewater, and other samples • Can also count cell numbers with flow cytometry |
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| Fluorescent staining using DAPI, acridine orange(AO), or SYBR Green I (SYBR) |
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| • DAPI-stained cells fluoresce bright blue (Figure 18.6a) • AO-stained cells fluoresce orange or greenish orange (Figure 18.6b) • SYBR-stained cells fluoresce green (Figure 18.6c) • DAPI, AO, and SYBR fluoresce under UVlight • DAPI, AO, and SYBR are used for the enumeration of microorganisms in samples • DAPI, AO, and SYBR are nonspecific and stain nucleic acids • Cannot differentiate between live and dead cells |
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| Viability stains |
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| : differentiate between live and dead cells • Two dyes are used • Based on integrity of cell membrane • Green cells are live • Redcellsaredead • Can have issues with nonspecific staining in environmental samples |
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| Green fluorescent protein |
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| can be genetically engineered into cells to make them autofluorescent • Can be used to track bacteria • Can act as a reporter gene |
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| Nucleic acid probe |
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| is DNA or RNA complementary to a sequence in a target gene or RNA |
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| FISH: fluorescence in situ hybridization |
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| • FISH can be used to measure gene expression in organisms in a natural sample • Phylogenetics of microbial populations • Fluorescingnucleotides complementary to rRNA sequence (Figure 18.10) • FISH technology can also employ multiple phylogenetic probes • Used in microbial ecology, food industry, and clinical diagnostics |
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| CARD-FISH |
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| • FISH can be used to measure gene expression in organisms in a natural sample • A FISH method that enhances the signal is called catalyzed reporter deposition FISH (CARD-FISH) |
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| culture independent Genetic Analyses of Microbial Communities |
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| • Specific genes can be used as a measure of diversity -Techniques used in molecular biodiversity studies • DNA isolation and sequencing • PCR • Restriction enzymedigest • Electrophoresis • Molecular cloning |
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| PCR Methods of Microbial Community Analysis |
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| • DGGE: denaturing gradient gel electrophoresis separates genes of the same size based on differences in base sequence • Denaturant is a mixture of urea and formamide • Strands melt at different denaturant concentrations • Next-generation DNA sequencers do not require a cloning step • PCR products can be used directly for sequencing • Allows for the detection of minor phylotypes • Results of PCR phylogenetic analyses • Several phylogenetically distinct prokaryotes are present • rRNA sequences differ from those of all known laboratory cultures • Molecular methods conclude that fewer than 0.1% of bacteria have been cultured |
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| Environmental genomics (metagenomics) |
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| • DNA is cloned from microbial community and sequenced • Detects as many genes as possible • Yields picture of gene pool in environment • Can detect genes that are not amplified by current PCR primers • Powerful tool for assessing the phylogenetic and metabolic diversity of an environment |
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| Metatranscriptomics |
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| • Analyzes community RNA • Reveals genes in a community that are actually expressed • Reveals level of gene expression |
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| Metaproteomics |
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| • Measures the diversity and abundance of different proteins in a community |
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| (Single-cell genomics) Multiple displacement amplification (MDA) |
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| • Links metabolic function to individual cells • Amplifies DNA from a single organism • Uses cell sorting • Uses bacteriophage DNA polymerase |
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| What we know since books was printed: |
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| •Most microorganisms are helpful – Nearly all contribute to your health, minimally protecting you from disease *Flora is plants – Do not use this word!!! – Bacteria,archaea,viruses,and fungi are microbes *Humans are likely colonized before birth – The placenta and the meconium both contain microbes |
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| Microbiome |
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| – The genes of all bacteria, archaea, fungi and viruses in the site of interest |
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| Microbiota |
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| – All bacteria, archaea, fungi and viruses in the site of interest |
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| Commensal |
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| – an organism that benefits from it’s host but has no impact on the host |
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| Beneficial |
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| – An organism that contributes to the host’s health |
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| Microflora |
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| – Not a thing, the microbiome is not made up of tiny plants! |
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| Microbiome changes over time |
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| NEWBORN- initial gut bacteria depends upon delivery mode - vaginal: vertical inheritance from mother - C-section: higher susceptibility to certain pathogens, higher risk of atopic diseases EARLY CHILDHOOD-new strains outcompete only ones, rapid increase in diversity, early microbiota development=high instability, shifts in response to diet, illness ADULT- highly distinct differentiated microbiota, microbial community may change, but at a slower rate then in childhood ELDERLY- substantially different gut communities then in younger adults |
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| Human microbiota |
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| • 1.3 microbes per human cells • This is a dynamic number • Approximately 39 trillion bacterial cells on the body – ~30 trillion human cells • 84% are red blood cells **does not count fungi, viruses, and archaea -The colon houses most of our microbiome |
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| Where do the microbes live? |
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| In mucus above epithelial cells |
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| What do gut microbes eat? |
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| • Fiber! • Protein • Sugarsubstitutes • Whatever you don’t absorb~~ Corn ...and if you don’t feed your gut microbiome it eats you or dies out |
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| A good microbiome |
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| • Regulates insulin levels • Keeps you from storing too much fat • Helps you feel full for longer periods of time • Feeds the cells of your colon and keeps them healthy • Keeps your cholesterol low |
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| A lacking microbiome |
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| • Diabetes • Obesity • Depression • Autism • Crohn’s disease • Colitis • Colon cancer • high blood pressure • heart attacks • Opportunities for pathogens to move in |
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| What fermentation products influence our health and how? |
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| • Acetate – increases cholesterol synthesis • Propionate –decreases cholesterol synthesis – increases carbohydrate tolerance • Butyrate – Prevents colon cancer – Suppresses weight gain – Alleviates diabetes – Anti-inflammatory – Decreases epithelial permeability |
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| Dysbiosis |
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| -the lacking microbiome • A disruption in the microbiome resulting in open niches and/or antagonistic relationship with the host. • Often a loss of diversity HAPPENS AS A RESULT OF Clostridium difficile |
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| Clostridium difficile |
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| causes life-threatening diarrhea. These infection mostly occur in people who have had both recent medical care and antibiotics. Resistance to the antibiotic used to treat it is not yet a problem, but the bacteria rapidly spreads because it is naturally resistant to many other drugs used to treat other infections. kills most people it infects. 90% of deaths were elderly. -1st: C. difficile spore exposure -gets into gut, and germinates, now its vegetative -colonizes in you gut, the disease developed and toxins are produces causing inflammation -then Spore shedding and infects others FECAL TRANSPLANTS are used to treat this |
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| How do we keep our microbiome happy? |
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| • Feed it – Withwhat• Stabilize it – repopulation • fermented foods – yogurt – sauerkraut – kimchi • Don’t disrupt it – limit antibiotics – avoid antimicrobials –particularly hand sanitizers and soaps with triclosan • it is killing you! • What should you eat – high fiber – low sugar – few processed foods |
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| Can we manipulate the gut microbiome? |
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| • 20 students • Increased resistant starch in their diet – unmodified potato starch • Changes in micro biome structure and function -Change in butyrate concentration during increased consumption of resistant starch -Resistant Starch-degrading organisms increased -Butyrate producers increase in the high butyrate group |
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| Virulence |
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| is the relative ability of a pathogen to cause disease • Pathogens use various strategies to establish virulence • Measuring virulence - Virulence can be estimated from experimental studies of the LD50 (lethal dose50) - The amount of an agent that kills 50% of the animals in a test group - Highly virulent pathogens show little difference in the number of cells required to kill 100% of the population as compared to 50% of the population (the more virulent the organic-the less cells that needed to be injected to kill the mouse) |
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| Attenuation |
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| • The decrease or loss of virulence • Many pathogens use a combination of toxins, invasiveness, and other virulence factors to enhance pathogenicity |
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| Toxicity |
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| • Organism causes disease by means of a toxin that inhibits host cell function or kills host cells • Toxins can travel to sites within host not inhabited by pathogen • Many pathogens use a combination of toxins, invasiveness, and other virulence factors to enhance pathogenicity |
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| Invasiveness |
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| • Ability of a pathogen to grow in host tissue at densities that inhibit host function • Can cause damage without producing a toxin • Many pathogens use a combination of toxins, invasiveness, and other virulence factors to enhance pathogenicity |
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| Adherence |
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| • Bacteria and viruses that initiate infection often adhere specifically to epithelial cells through interactions between molecules on the surfaces of the pathogen and the host cell • Bacterial adherence can be facilitated by • Extracellular macromolecules that are not covalently attached to the bacterial cell surface • Examples: slime layer, capsule • Fimbriae and pili |
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| Invasion, Infection, and Virulence Factors |
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| • The initial inoculum of a pathogen is insufficient to cause host damage • The pathogen must multiply and colonize the tissue • The availability of nutrients is most important in affecting pathogen growth • Pathogens may grow locally at the site of invasion or may spread throughout the body |
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| Bacteremia |
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| : the presence of bacteria in the bloodstream |
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| Septicemia |
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| : bloodborne systemic infection • May lead to massive inflammation, septic shock, and death |
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| Infection |
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| : any situation in which a microorganism (not a member of the local flora) is established and growing in a host |
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| Invasion, Infection, and Virulence Factors (on teeth) |
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| • Infection requires attachment to surface as well as growth • Attachment and growth have been well studied in the formation of biofilms on tooth surfaces • Acidic glycoproteins from saliva form a thin film • Streptococci colonize thefilm • Streptococcussobrinus and Streptococcus mutans are common agents in tooth decay • Extensive growth of oral microorganisms, especially streptococci, results in a thick bacterial layer (dental plaque; Figure 23.16) • As plaque continues to develop, anaerobic bacterial species begin to grow • As dental plaque accumulates, the microorganisms produce high concentrations of acid, resulting in decalcification of the tooth enamel (dental caries) |
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| Pathogens produce enzymes that: |
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| • Enhance virulence by breaking down or altering host tissue to provide access to nutrients • Example: hyaluronidase • Protect the pathogen by interfering with normal host defense mechanisms • Example: coagulase Salmonella species encode a large number of virulence factors (Figure 23.17) • Several genes that direct invasion are clustered together on the chromosome as pathogenicity islands • Another Salmonella pathogenicity island contains genes that promote a more systemic disease • Salmonella also contains resistance plasmids (R plasmids) |
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| Exotoxins |
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| • Proteins released from the pathogen cell as it grows • Three categories • Cytotoxins • AB toxins • Superantigen toxins |
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| Cytotoxins (cytolytic toxins) |
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| • Work by degrading cytoplasmic membrane integrity, causing cell lysis and death • Toxins that lyse red blood cells are called hemolysins (Figure 23.18) • Staphylococcal ?-toxin kills nucleated cells and lyses erythrocytes |
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| AB toxins |
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| • Consist of two subunits, A and B • Work by binding to host cell receptor (B subunit) and transferring damaging agent (A subunit) across the cell membrane (Figure 23.20) • Examples: diphtheria toxin, tetanus toxin, botulinum toxin • Clostridium tetani and Clostridium botulinum produce potent AB exotoxins that affect nervous tissue • Botulinum toxin consists of several related AB toxins that are the most potent biological toxins known; tetanus toxin is also an AB protein neurotoxin |
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| Enterotoxins |
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| • Exotoxins whose activity affects the small intestine • Generallycause massive secretion of fluid into the intestinal lumen, resulting in vomiting and diarrhea • Example: cholera toxin |
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| Endotoxin |
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| • The lipopolysaccharide portion of the cell envelope of certain gram-negative Bacteria, which is a toxin when solubilized • Generally less toxic than exotoxins • Presence of endotoxin can be detected by the Limulus amoebocyte lysate (LAL) assay |
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| Innate Resistance to Infection |
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| • Hosts have innate resistance to most pathogens (Figure 23.25) • Natural host resistance • Tissue specificity • Physical and chemical barriers |
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| Compromised host |
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| • One or more resistance mechanisms are inactive • The probability of infection is increased • Age is an important factor • Very young and very old individuals are more susceptible • Stress can predispose a healthy individual to disease • Diet plays a role in host susceptibility to infection • Certain genetic conditions can compromise a host |
