The Culturing of Staphylococcus epidermidis and Streptococcus salivarius with Escherichia coli Does Not Increase Antibiotic Resistance in Escherichia coli

By: Sarah Weber

INTRODUCTION

• Antibiotic resistance is important -- antibiotics are used to treat bacterial infections

• When testing effectiveness of an antibiotic -- performed on individual strains

• Not realistic -- hundreds of strains interact in the environment

• Bacteria intermingle and transfer genes between each other in order to survive

INTRODUCTION

• Escherichia coli– Member of a large and diverse group of gram

negative bacteria– Normally live in human and animal intestines– Most strains are harmless, some are pathogenic– Can be used to indicate if water is contaminated– Rod-shaped (bacillus) bacteria

INTRODUCTION

• Streptococcus salivarius – Gram positive– Principal bacterium living in human oral cavity

without causing harm – Pioneer in colonizing dental plaque– Can cause disease if it enters the blood stream– Spherical shaped (coccus) bacteria

INTRODUCTION

• Staphylococcus epidermidis– Gram positive– Part of normal skin flora in humans– Leading cause of hospital born infections in

immune compromised patients– Spherical shaped (coccus) bacteria in grape-like

clusters

INTRODUCTION

• Penicillin– One of the earliest discovered and widely used

antibiotics– Used to treat many different types of infections

caused by bacteria– Kills bacteria by interfering with the ability to

synthesize the cell wall

INTRODUCTION

• Erythromycin– Macrolide antibiotic– Used to treat many different types of infections

caused by bacteria– Slows the growth of sensitive bacteria – Reduces the production of important proteins

needed by the bacteria to survive

INTRODUCTION

• Streptomycin– Aminoglycoside antibiotic– Used to treat many different kinds of bacterial

infections– Cannot be given orally; injected intramuscularly– Prevents growth of bacteria by protein synthesis

inhibition

INTRODUCTION

• Mutation rate– Bacteria mutate at a rapid rate, some faster than

others– Bacteria that survive in the presence of antibiotics

acquire resistance through:• Resistance genes• Recombination with foreign DNA

– Presence of antibiotics induces mutations causing a slow mutation rate

AIM

• The aim of the research was to study the speed at which bacterial resistance occurred in a pure culture versus mixed cultures.

HYPOTHESIS

• I hypothesized that the antibiotic resistance in the E. coli with S. epidermidis and E. coli with S. salivarius would increase faster than in the E. coli alone.

METHODS• Cultures made of parent strains • Parent strains plated to see zone of inhibition • Plates were made of E. coli only, mixtures of E. coli

with S. salivarius, and mixtures of E. coli with S. epidermidis

• Test plates of individual strains– Left to Right: E. coli, S. epidermidis, S. salivarius

METHODS• Every other day a culture was made using the

previous growth plate to make a new generation using only the E. coli with S. salivarius and E. coli with S. epidermidis plates

• The cultures were incubated, then plated onto blood agar with one disk each of penicillin, erythromycin, and streptomycin

E. coli S. salivarius S. epidermidis0

0.5

1

1.5

2

2.5

3

3.5

4

Diameter of the Zone of Inhibition for the Parent Generation

PenicillinErythromycinStreptomycin

Parent Generation

Zon

e of

Inh

ibiti

on(m

m)

• Generation Number 1‾ Left to right: E. coli, E. coli with

S. salivarius, E. coli with S. epidermidis

• Generation Number 16‾ Left to right: E. coli, E. coli with

S. salivarius, E. coli with S. epidermidis

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 170

0.2

0.4

0.6

0.8

1

1.2

1.4

Diameters of the Zone of Inhibition for E. coli Con-trol

PenicillinStreptomycin

Generations

Zon

e of

Inh

ibiti

on (m

m)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 170

0.2

0.4

0.6

0.8

1

1.2

1.4

1.6

Diameters of the Zone of Inhibition for S. salivarius and E. coli

PenicillinErythromycinStreptomycin

Generations

Zon

e of

Inh

ibiti

on (m

m)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 170

0.5

1

1.5

2

2.5

Diameters of the Zone of Inhibition for S. epidermidis and E.coli

PenicillinErythromycinStreptomycin

Generations

Zon

e of

Inh

ibiti

on (m

m)

RESULTS

• An ANOVA (General Linear Model) was performed• No significant difference between the two different

bacterial combinations on the rate of change in the zone of inhibition (p> 0.05)

• Significant difference in generations (p< 0.05)

Generation Mean (mm)

1 0.5875a

17 0.3167c,d

RESULTS

• Significant difference with respect to inhibition of bacterial growth (p< 0.05)

Antibiotic Mean (mm)

Streptomycin 0.4371a

Erythromycin 0.3400a,b

Penicillin 0.2961b

CONCLUSION

• Hypothesis of antibiotic resistance in the E. coli with S. epidermidis and E. coli with S. salivarius would increase faster than in the E. coli alone was rejected

• Significance: two bacteria grown together did not selectively mutate any faster than individual strains

• Change in the zone over time showed the antibiotics are not as effective as when first introduced

LIMITATIONS OF RESEARCH

• Too many similarities between the two treatment bacteria (S. salivarius and S. epidermidis)

• Too many similarities in the zones of inhibition• The number of generations was insufficient • The recombination of the two bacteria did not

occur

FURTHER RESEARCH

• Allow for more generations to determine how many are needed for the bacteria to become completely desensitized to the antibiotics

• Change the bacteria used • Change the antibiotics used

QUESTIONS