Transcript Variances seen in Bacterial Analysis for Water and Waste

Variances seen in
Bacterial Analysis for
Water and Waste Water
Sampling
Gretchen Hathaway
Whatman Sales Representative
July 19, 2007
Topics
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M-Endo (LTB, BGB, & ECB)
M-FC & M-FC with rosolic acid
MI Broth and Agar
ColiCheck & ColiCheck with MUG
M-Endo
M-Endo
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M-Endo Broth is used for the enumeration of coliforms by
membrane filtration.
Ideal for drinking water and waste water contaminant
testing, M-Endo Coliform Broth is used to differentiate
lactose from non-lactose fermenting intestinal organisms.
M-Endo : Description
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M-Endo is a red colored media, which needs
to be stored in the dark to prevent
discoloration of the media.
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Gram-positive bacteria are inhibited on this
media by the desoxycholate and lauryl
sulfate. The addition of ethanol increases the
antibacterial nature of the formulation.
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Lactose fermenting organisms form
aldehydes, which react with Schiffs reagent
(basic fuchsin and sodium sulfite) to give red
colored zones around the colonies.
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Coliform colonies are therefore red with a
characteristic metallic sheen.
M-Endo: Interpretation
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Production of both acid and
aldehyde by lactose fermenters,
such as Escherichia coli, produce
deep red colonies that color the
surrounding medium and have a
green metallic sheen.
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Non-lactose fermenters form
colorless, translucent colonies
M-Endo Verification
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The chart to the side
demonstrates the need of
verification steps due to
the type of growth.
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Verification is done in
durham tubes with Lauryl
Tryptose Broth, Brilliant
Green Broth (to confirm
Total coliform) and EC
Broth (to confirm Fecal
coliform).
ColiCheck and
ColiCheck with MUG
ColiCheck and ColiCheck with MUG
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ColiCheck is used for the presumptive
identification of coliforms in water samples by a
presence/absence technique.
Two product types are available, ColiCheck that
identifies coliforms and ColiCheck with MUG
that checks for the presence/absence of
Escherichia coli.
ColiCheck and ColiCheck with MUG
Description
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Bromocresol purple is a pH indicator which demonstrates
a color change from purple to yellow in the presence of
acid.
Lactose fermenting organisms produce acid which initiates
the color change.
The presence of coliforms is detected with greater
sensitivity by use of a relatively large sample volume
(100ml) in a single bottle.
The addition of MUG (4-methyl-umbellrifryl-βglucuronide) which is a fluorogenic enzyme allows the
media to selectively identify E. coli.
MUG is hydrolyzed by the E. coli specific enzyme βglucuronidase to release 4-Methylumbelliferone which
fluoresces under UV light (366nm).
ColiCheck and ColiCheck with MUG
Interpretation
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Yellow color in the sample
after 24 hours incubation at
35° C indicates the presence
of coliform organisms.
A red sample after 24 hour
incubation at 35° C should be
incubated for further 24 hours.
A red sample at 48 hours
incubation at 35° C indicates
no coliform organisms present
in the sample.
ColiCheck w/ MUG:
Florescence under UV Light is
specific for E. coli.
M-FC broth
M-FC
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M-FC media (membrane Fecal Coliform
media) is used for the detection of fecal
coliforms as an index of water pollution.
M-FC: Description
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Allows the development
of fecal coliforms at
elevated temperatures
(44.5° C).
M-FC: Interpretation
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Bile salts included in the
medium inhibit the
growth of gram-positive
bacteria.
Fecal coliforms ferment
lactose at elevated
temperatures and
produce blue colonies.
Other organisms form
grey to cream colonies.
M-FC with Rosolic
Acid
M-FC with Rosolic Acid
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M-FC Broth with Rosolic Acid is used in
waste water analysis. M-FC (Membrane Fecal
Coliform) media is designed for the detection of
fecal coliforms as an index of water pollution.
M-FC with Rosolic Acid: Description
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M-FC with Rosolic Acid
acts and functions in the
same way as m-FC
Broth.
Rosolic acid inhibits
bacterial growth in
general, except for fecal
coliforms.
M-FC with Rosolic Acid:
Interpretation
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Bile salts inhibit nonenteric bacteria.
Aniline blue indicates the
ability of fecal coliforms
to ferment lactose to acid
that causes a pH change
in the medium.
Other organisms form
grey to cream colonies.
MI Broth and Agar
MI Broth and Agar
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Used for the simultaneous detection of total
coliforms and Escherichia coli in water
according to the Surface Water Treatment
Rule (USEPA) and the Total Coliform Rule
(USEPA), which applies to drinking water.
MI Broth and Agar: Description
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MI Broth detects the presence of coliform
bacteria by the production of β-galactosidase,
which cleaves the substrate MUGal to
produce 4- Methylumbelliferone, which
fluoresces on exposure to UV light (366nm).
Non-coliforms do not produce this enzyme
and therefore do not fluoresce on the
medium.
Escherichia coli is detected by the compound
IBDG.
The β-glucuronidase produced by E. coli
cleaves the substrate to produce a blue
indigo color in the colonies.
As E. coli is also a total coliform, and also
produces β-galactosidase it will also
fluoresce.
The antibiotic cefsulodin is added to inhibit
the growth of gram-positive bacteria and
some non-coliform gram negative bacteria
that can cause false positive reactions.
Fewer False Positives are seen
with MI Broth.
MI Broth and Agar: Interpretation
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Fluorescent blue colonies are
Escherichia coli.
Colonies that demonstrate
blue/white fluorescence where the
colonies are clear, cream or pale
yellow in color are other coliform
organisms.
Clear colonies that do not fluoresce
are non-coliforms.
Short incubation period… Positive
control for E. coli ATCC 25922 and
Enterobacter aerogenes ATCC 13048
can be achieved in 18-24 hours at
35°C.
Negative control… Complete
inhibition of Pseudomonas
aeruginosa ATCC 10145 occurs at
35°C after 24 hours.
Other Products offered by Whatman
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Monitors/ Analytical Filter Funnels
Membrane Systems and Membrane Butler
Dilution Bottles
Petri dishes with pads
Filter papers
Glass Fiber for Total Suspended and Volatile Solids, &
TCLP
pH papers
Syringe filters
The End