Lecture 3

Question 1

Before molecular biology methods were available. How were E.coli and Shigella distuniguished

Answer 1

Based on the basis of motility, metabolic profile and clinical manifestation

Question 2

What did serotyping studies demonstrate

Answer 2

That the O- antigen and H (flagellar) antigen and sometimes K (capsular) antigen are useful for distinguishing between strains

Question 3

What is serotyping

Answer 3

Typing based on immune recognition of cell surface antigens.

Bacteria of the same serotype cross-react to the same antibodies

Question 4

What’s an example of the E.coli serotype

Answer 4

O157:H7

Question 5

What serotypes were associated with outbreaks

Answer 5

The pathovar enteropathogenic E.coli. - Originally defined based on serology but is now defined by characteristic A/E lesions in the ileum.

They form pillars on the surface of the cell so eject into the proteins, causing pedestals to form

Question 6

Enterohaemorrhagic E.coli (EHEC)

Answer 6

Associated with haemorrhagic colitis and usally causes A/E lesions similar to EPEC. Also produces shiga toxin that can cause renal damage resulting in haemolytic uraemic syndrome e.g. O157:H7

Question 7

Enterotxogenic E.coli (ETEC)

Answer 7

Major causes of infantile diarrhoea in the developing world and travellers diarrhoea in visitors to those regions. Associated with heat-labile and heat stable enterotoxins

Question 8

What happens when infecting HEp-2 cells in vitro (EPEC and EAEC and DAEC)

Answer 8

EPEC strains form tight clusters as they stick to specific regions of the host cells

EAEC form characteristic stacked-brick pattern, cells adhere to each other to help survival in gut

DAEC are defined based on their diffuse adherence pattern (scattered around host cells)

Question 9

What was DNA-DNA hybridisation used for

Answer 9

Measuring the evolutionary relatedness between different bacyeria

Question 10

What happens in DNA-DNA hybridisation

Answer 10

DNA strands form two strains and are hybridised together and by measuring the temperature required to dissociate the hybrid DNA into seperate strand, it’s possible to estimate the degree of relatedness

Question 11

Milkman study (1973)

Answer 11

Quantitative study of E.coli population genetics by measuring the electrophoretic mobility of enzymes derived from different E.coli strains.

If two enzymes have strains with same mobility - marks them as the same allele/ gene and vice versa, can get a measure of how closely related the strains are

Question 12

What does multi locus enzyme electrophoresis involve

Answer 12

MLEE involves assessing the electrophoretic mobility of a series of purified enzymes. It produces quantitative molecular data which can be used to understand the evolutionary relationships between strains

Question 13

What did early studies show about seroytping

Answer 13

Does not correlate well with genetic diversity as measured using MLEE. Genetically similar strains can have different serotypes and distantly related strains can share the same serotype

Question 14

What did analysis of E.coli and Shigella strains using nucleotide data from genes thrB and thrC indicate

Answer 14

That there were multiple seperate shigella lineages within the diversity of E.coli. Suggesting that shigella strains had evolved on multiple distinct occasions with convergent evolution of their defining characteristics such as loss of motility

Question 15

What is multi locus sequence typing

Answer 15

Rather than sequencing one or two genes, you sequence a few from the same loci (around 400bp sections from several different genes)

Question 16

What does MLST sequencing involve

Answer 16

Sequencing 400bp sections from 7-8 house keeping genes from different chromosomal regions

Question 17

E.coli what did they identify

Answer 17

5 “cryptic clades”

Question 18

Differences between e.coli and shigella

Answer 18

Shigella spp. are non-motile, obligate pathogens that cannot ferment lactose and are lysine decarboxylase and indole-negative

E.coli are usually motile, lactose-, lysine decarboxylase and are indole - positive are suggested to be mostly commensal organisms

Question 19

Enteroaggregative E.coli (EAEC)

Answer 19

Show a characterisitic stacked brick pattern of aggregation wherease diffuse adherent E.coli are characterised by a diffuse pattern of adherence with little aggregation

Question 19

Enteroinvasive E.coli

Answer 19

Share many characterisitcs with shigella and exhibit similar invasive phenotypes but elicit milder symptoms compared to diarrhoeagenic E.coli

Question 20

Extraintenstinal pathogenic E.coli (ExPEC)

Answer 20

Associated with several clinical conditions including UTIs caused by UPEC and neonatic meningitis

Question 21

What did the quantitiative study of E.coli population genetics begin with

Answer 21

The measurement of variations in electrophoretic mobility of enzymes derived from different e.coli strains

Question 22

ECOR strain collection

Answer 22

An initial phylogeny obtained through cluster analysis of MLEE data for 35 loci defined 6 phylogenetic groups - constructed using UPGMA

Question 23

Phylogenetic analysis using nucleotide sequence data

Answer 23

Data derived from enzyme electrophoresis are not ideal for the purposes of phylogenetic analysis- enzymes with little sequence similarity may exhibit similar electrophoretic mobility

Question 24

What do nucleotide and amino sequence provide

Answer 24

A platform for sophisticated phylogenetic analyses- provide disecrete character state information for many loci and unlikely to suffer convergence