Transcript Team 8 Presentation

Identification and
Characterization of
Enterococcus spp. in
Local Surface Waters
Team Microbiology
Advisor: Dr. June Middleton
Assistant: Alex Kohl
Neha Bansal, Laura Decker, Kevin Huang,
Ashwinee Ragam, Angela Sekerke, Reema Shah,
Sarah Song, Pallavi Yerramilli, Eddie Zhang,
Jeremy Zornow
Enterococcus spp. History
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Closely related to Streptococcus
Separate genus formed in 1984
16 different species
Located in intestines of warmblooded animals, plants, soil, and
water
Characteristics
• Gram-positive cocci
• Group D glycerol teichoic acids
• 6.5% NaCl
• Esculin
Esculin test
Medical Significance
• E. faecium and E. faecalis
• Nosocomial infections
– Urinary tract infection
– Wound infection
– Bacterial endocarditis
E. faecium
E. faecalis
Antibiotics: antimicrobial
compounds that terminate or
impede the proliferation of
bacteria derived from certain
microorganisms
Selective Toxicity: the ability
of an antibiotic to target
bacteria without harming the
host cells
Antibiotic Resistance
the potential of a
microorganism to
resist and endure
the effects of an
antibiotic
Vancomycin-Resistant
Enterococcus spp.
Virulence Factors
• The products of any genes that help
bacteria invade and colonize a host.
• Virulence factors work synergistically
to help the cell invade a host.
• The more virulence factors a
bacterium expresses, the more
dangerous it is to our health.
Virulence Factors
• Hemolysins
– Compounds that destroy red blood cells.
• Bacteriolysins
– Compounds that kill “rival” bacteria, leading to
the dominance of an area by one strain or
species.
• Gelatinase
– Disassembles gelatins and collagen – the
bacterial cell will then recycle the peptide
segments for use among the colonizing
bacteria.
Sample Collection
Burnham Pond, Drew Pond, Loantaka
Brook, and Whippany River
Culture Isolation
Whippany River
1 mL
Drew Pond
1 mL
Whippany River
5 mL
Drew Pond
5 mL
Whippany River
25 mL
Drew Pond
10 mL
Speciation
Yellow = Fermentation
(Positive)
Red = Negative
Sugars Tested:
arabinose
mannitol
methyl
α-D-glucopyranoside (MGP)
ribose
sorbose
sorbitol
Antibiotic Resistance
Antibiotics Tested:
 cephalothin (30µg/ml)
 chloramphenicol (30µg/ml)
 ciprofloxacin (5µg/ml)
 erythromycin (15µg/ml)
 gentamycin (10µg/ml)
 nitrofurantoin (300µg/ml)
 penicillin (10µg/ml)
 streptomycin (10µg/ml)
 tetracycline (30µg/ml)
 vancomycin (30µg/ml)
Virulence Factors
α hemolysis
Virulence Factors Tested:
 Gelatinase
 Hemolysins
 Bacteriocins
~ Tested
against:
Streptococcus mitis PI 519
Enterococcus faecalis ATCC
29212
Enterococcus mundtii O
Enterococcus mundtii P
Enterococcus mundtii R
β hemolysis
Counts
Comparison of Total Enterococci Levels in Local Water Sources
10000
3150
1100
880
log Colonies per 100 mL
1000
240
100
10
1
Whippany River (W)
Loantaka Brook (LB)
Drew Pond (DREW)
Water Source
Burnham Pond (BP)
Relative Abundance
Relative Abundance of Enterococcal Species in Local Water Sources
60
E. faecium
E. faecalis
50
E. durans/hirae
E. gallinarum
Percent of Total Isolates
E. casseliflavus
40
E. mundtii
30
20
10
0
Whippany River (n=46)
Drew Pond (n=48)
Loantaka Brook (n=41)
Water Source
Burnham Pond (n=33)
Relative Abundance con’t
• Low concentration of Enterococcus
faecalis
• High concentration of Enterococcus
casseliflavus
• Varying concentrations of
Enterococcus faecium
Antibiotic Resistance
Relative Antibiotic Resistance of Total Enterococci in Local Water Sources
100
Percen Total Isolates
80
Penicillin
Cephalothin
Tetracycline
60
Streptomycin
Gentamycin
Ciprofloxacin
40
Vancomycin
20
0
Whippany River (n=46)
Burnham Pond (n=33)
Drew Pond (n=48)
Water Source
Loantaka Brook (n=41)
Antibiotic Resistance con’t
• High resistance to Penicillin,
Cephalothin, Streptomycin
• Moderate resistance to Gentamycin
and Vancomycin
• Low resistance to Tetracycline and
Ciprofloxacin
Virulence
• Majority of Enterococcus from
Whippany River lysed all five of the
bacteria
• Majority of Enterococcus from Drew
Pond and Loantaka Brook lysed none
of the bacteria
• Burnham Pond possessed no
colonies of virulent bacteria
Conclusions
• Maximum level of Enterococcus
allowed for drinking water is 7
isolates per 100 mL and for bathing
water is 100 isolates per 100 mL
• All of the water sources we tested
were unsuitable for drinking or
bathing
Conclusions con’t
• E. faecalis and E. faecium are
nosocomial pathogens (causes of
secondary infections in hospitals)
• Only pathogenic species of
Enterococcus
Conclusions con’t
• Overuse of common antibiotics has led to
high bacterial resistance
• Three antibiotics virtually ineffective in
curbing Enterococcus spp. growth
– Penicillin
– Cephalothin
– Streptomycin
• Three antibiotics displayed 100%
effectiveness against Enterococcus spp.
– Chloramphenicol
– Erythromycin
– Nitrofurantoin
Conclusions con’t
• E. faecalis and E. faecium were found
to have similar resistance patterns
• One main exception – E. faecium
displayed susceptibility to
vancomycin
Conclusions con’t
• Whippany River - large concentration
of E. faecalis and E. faecium
• Enterococcus found in Drew Pond
and Loantaka Brook possess lower
virulence capability
• Burnham Pond –no isolates of E.
faecalis or E. faecium found in pond
Fin
Paul Quinn the Bacterial Contamination!!!!!