Microbio Midterm

A hypothetical pathogenic bacterium (disease-causing microorganism) is introduced into a susceptible animal model of infection, leading to no symptoms. Which of the following statements about Koch's Postulates is correct?

does not fulfill Koch’s postulates

What is not a feature of the structure and RNA expression of prokaryotic genomes?

introns

Removal of the Shine Dalgarno (RBS) sequence of the gene would inhibit what process?

translational initiation

If the -35 and -10 sequences were removed, the binding to the promoter of which of the following would be inhibited?

sigma factors

What is not an example of horizontal gene transfer?

regulation of plasmid copy number

An Ames test is performed to determine the mutagenicty of two compounds (A and B). Incubation of the Ames test bacteria with compound A led to 10,000 colonies on the resulting histidine- plate. Incubation with an equal amount of compound B with an equal amount of test bacteria led to 100 colonies on the histidine- plate. Which compound is more mutagenic?

A is more mutagenic

What is characteristic of Gram+ cell envelopes?

Lipoteichoic acid

What causes the expression (activation) of SOS genes?

RecA binding to ssDNA causes LexA to degrade itself

What forms part of the crosslinks between peptidoglycan monomers components in Gram-positive bacteria?

glycine

In PTS transport of glucose, why does the cell phosphorylate glucose?

Modification of glucose maintains a concentration gradient

What did Pasteur’s swan-neck flask experiment prove?

Disproved spontaneous generation (microbes come from other microbes)

List Koch’s postulates (in order)

Observe microbe → isolate → cause disease → reisolate

Major limitation of Koch’s postulates

→ Many microorganisms cannot be grown in pure culture

→ Some pathogens grow extremely slowly

→ Some microbes require host-specific growth factors

→ Ethical issues: cannot intentionally infect humans

→ Some diseases are caused by multiple organisms

→ Some infected individuals are asymptomatic carriers

→ Some pathogens only infect specific hosts (no animal model)

Who introduced agar for solid media?

→ Angelina and Walther Hesse

Why was agar important?

→ Allows isolation of single colonies

Gram stain steps in order

Crystal violet → iodine → alcohol → safranin

Why do Gram+ stain purple?

Thick peptidoglycan retains crystal violet

Why do Gram− stain pink?

Thin peptidoglycan loses crystal violet and takes safranin

Gram+ features

Thick peptidoglycan, teichoic acids, no outer membrane

Gram− features

Thin peptidoglycan, outer membrane, LPS, periplasm

Unique to Gram−

Outer membrane / LPS

What makes peptidoglycan rigid?

Peptide crosslinking of glycan chains

Gram+ peptide interbridge uses what amino acid?

glycine

cell envelope

everything surrounding the cytoplasm

It includes:

• Cell membrane

• Cell wall (peptidoglycan)

• Sometimes an outer membrane (Gram− only)

Primary active transport

Uses ATP directly

Secondary active transport

• Uses existing gradient

• moves solutes against their concentration gradient using energy stored in electrochemical gradients, rather than direct ATP hydrolysis

PTS system energy source

PEP

What happens to sugar in PTS?

it is phosphorylated during transport

Why phosphorylate glucose?

Maintains inward concentration gradient

Symporter

Two solutes move same direction

Antiporter

Two solutes move opposite directions

Silent mutation

No amino acid change

Missense mutation

Different amino acid

Nonsense mutation

Premature stop codon

Frameshift mutation

Reading frame shifted (insertion/deletion not multiple of 3)

What does Ames test detect?

Mutagens

What does more colonies mean in Ames test?

higher mutagenicity

What type of bacteria are used in Ames test?

his⁻ (histidine-requiring)

What is a revertant?

Mutant that regains ability to synthesize histidine

Primase

Makes RNA primers

RNase H

Removes RNA primers

DNA polymerase

Extends DNA

Topoisomerase

Relieves supercoiling

What activates SOS response?

RecA binding single-stranded DNA

Role of LexA

Represses SOS genes until cleaved

Promoter definition

DNA site where RNA polymerase binds

Conserved promoter regions

−35 and −10

Which protein recognizes promoter?

Sigma factor

Shine-Dalgarno sequence

Ribosome binding site

Shine-Dalgarno interacts with

16S rRNA

Removal of Shine-Dalgarno affects

Translation initiation

Rho-dependent termination

Rho helicase destabilizes RNA/DNA

Rho-independent termination

Hairpin + U-rich region

Transformation

Uptake of naked DNA

Transduction

Phage-mediated DNA transfer

Conjugation

Cell-to-cell DNA transfer via pilus

Generalized transduction

Random bacterial genes transferred

Specialized transduction

Only genes near prophage site

Not HGT example

Regulation of plasmid copy number

High copy plasmids advantage

Ensures inheritance

Low copy plasmids advantage

Lower metabolic cost

Rolling circle replication

Single strand nick → displacement → replication

Stringent plasmids

Replicate with chromosome

Relaxed plasmids

Replicate independently

Central dogma

DNA → RNA → Protein

What is homology?

Shared sequence from shared ancestry

Orthologous genes

Same gene in different species (speciation)

Paralogous genes

Gene duplication within same genome

What makes a good molecular clock?

Universal, conserved, slow constant mutation rate

Why is 16S rRNA a good molecular clock?

Universal, conserved, interacts with Shine-Dalgarno

What part of RNA polymerase recognizes promoters?

Sigma factor

What sugars make up peptidoglycan?

→ NAG and NAM

→ Contains glycan chains + peptides

What are transposons?

Mobile DNA elements moved by transposase

Why are transposons important?

Can spread antibiotic resistance

Two main bacterial double-strand break repair methods

NHEJ and RecA-mediated recombination

What is passive transport?

Movement down concentration gradient without energy

What is active transport?

Movement against concentration gradient requiring energy

Who first observed living microorganisms? (Microscope)

Antonie van Leeuwenhoek