LABORATORY Flashcards
Specimen for diagnosing Enterobacteriaceae and Other Related Enterobacteria
Urine, blood, pus, spinal fluid, sputum
Gram Negative Enterobacteriaceae
Escherichia coli Shigella dysenteriae Salmonella typhi Klebsiella pneumoniae Proteus vulgaris
Cultural Method for Enterobacteriaceae spp
BAM MAC SSA EMB Broth culture 35-37 oC 24-48 hours
On MAC:
Typical strong lactose fermenters produce
Slow or weak lactose fermenters produce
Non-lactose fermenters produce
red colonies surrounded by a zone of precipitated bile.
light pink colonies or colonies that are clear at the periphery and have pink centers.
colorless colonies or transparent colonies
On EMB:
Typical strong lactose fermenters produce
Slow or weak lactose fermenters produce
Non-lactose fermenters produce
black with metallic sheen
pink to purple colonies
colorless colonies or transparent colonies
On SSA:
Slight growth of lactose fermenters that produce
Salmonella and Proteus colonies are
Shigella colonies are
pink or red colonies.
colorless with black centers.
colorless with no blackening.
stabbing the butt (or deep) to within 2-3 mm
streak over the agar surface (or slant) with a back-and-forth motion
Triple Sugar Iron Agar Reactions
stabbing to two-thirds of the distance to the
bottom in the center of the butt (or deep).
Sulfide-Indole-Motility Test
stabbing the butt (or deep) twice
streak over the agar surface (or slant).
Lysine Iron Agar Reactions
Incubation period for TSIA, SIM Test, and LIA Reactions
35-37oC for 18-24 hours
indicated by the retention of the original red color of the medium
Alkalinization or nonfermentation of the carbohydrates
designates alkaline reaction
“K”
indicated by a change in the color of the medium from red to yellow
Acid production from fermentation of carbohydrate(s)
designates acidic reaction
“A”
indicated by gas bubbles or spitting of the agar in the butt portion
Gas production
indicated by a black discoloration of the medium which first appears or remains confined in the butt portion
Hydrogen sulfide (H2S) production
alkaline slant and butt, no gas and H2S production
alkaline slant, acid butt, with gas and H2S production
acid slant and butt, with gas, no H2S production
K/K
K/Ag + H2S
A/Ag
Diffuse growth outward from the stab line or turbidity throughout the medium
Motility
Blackening of the medium (or along the stab line)
H2S production
To detect indole production, add 3-4 drops of Ehrlich’s or Kovac’s reagent, and observe for a pink to red color
Indole production
is detected in the butt by an alkaline (purple) slant
Lysine decarboxylation
is detected by a red slant.
Lysine deamination
detected by formation of a black precipitate.
Hydrogen sulfide (H2S) production
alkaline slant and butt, and NO H2S production
alkaline slant and butt butt, and H2S production
Red slant, acid butt, and NO H2S production
alkaline slant, acid butt, and NO H2S production
alkaline slant, acid butt, and H2S production
K/K
K/K + H2S
R/A
K/A + H2S
INDOLE TEST: Reagent
15 drops of Ehrlich’s + 1 ml of xylene or chloroform
Kovac’s reagent
INDOLE TEST: Positive result
Bright fuschia red color
*at the interface of the reagent and the broth (or xylene layer)
INDOLE TEST: Negative result
Yellow color or no color change
METHYL RED TEST: Reagent
5 drops of methyl red indicator
METHYL RED TEST: Positive result
Red color on the surface of the medium
METHYL RED TEST: Negative result
Yellow to orange color
VOGES-PROSKAUER TEST
- 6 ml of 5% alpha-naphthol
0. 2 ml of 40% KOH
VOGES-PROSKAUER TEST: Positive result
Red color
VOGES-PROSKAUER TEST: Negative result
Yellow color or no color change
SIMMON’S CITRATE UTILIZATION TEST: Reagent
no indicator
SIMMON’S CITRATE UTILIZATION TEST: Positive result
Prussian blue or deep blue color
SIMMON’S CITRATE UTILIZATION TEST: Negative result
No color change (retention of green color)
The presence of large capsules, which can be observed as colorless to light pink halo around the bacilli, is suggestive of
Klebsiella species
A basic fuchsin-methylene blue, ethyl alcohol phenol microscopic staining procedure.
Wayson staining for Yersinia species
Useful alternative for Gram staining.
Wayson staining for Yersinia species
show bipolar purple staining with a central vacuole giving a characteristic “safety pin” appearance.
Yersiniae
Used as an enrichment medium for the isolation of Salmonella from feces, urine, water, food and other materials.
Selenite broth
Selenite broth: inhibitor
Sodium selenite
Incubation for Selenite Broth - because coliforms or other intestinal flora may overgrow the pathogens within a few hours.
8-12 hours at 35 oC
Overheating of Selenite Broth may produce a visible [?], making it unsatisfactory for use.
precipitate
Used as an enrichment medium for the recovery of Salmonella and Shigella from clinical and nonclinical specimens.
GN (Hajna) broth
[?], in a higher concentration than glucose, enhances the growth of Salmonella and Shigella.
Mannitol
GN (Hajna) broth: inhibitor
Sodium citrate and sodium desoxycholate
Incubation for GN (Hajna) broth - because of relatively low concentration of desoxycholate, it is less inhibitory to E. coli and other coliforms.
4-6 hours at 35 oC,
Used as a selective enrichment for Salmonella species.
Tetrathionate broth, with iodine-iodide solution
formed in the medium by the addition of iodine-iodide solution
tetrathionate
tetrathionate broth: inhibitor
inhibits the normal intestinal flora of fecal specimens.
bile salt, tetrathionate
Incubation for Tetrathionate broth, with iodine-iodide solution
18-24 hours at 35 oC
heat the broth base to [?], add the
iodine solution
boiling
Colonies are usually large, white or gray, smooth, shiny, circular, raised colonies which may or may not be hemolytic.
Blood agar medium (BAM)
Used for selective isolation of G- enteric bacilli by incorporation of agent/s inhibitory to G+ bacteria; may be slightly selective, moderately selective, or highly selective.
SELECTIVE MEDIA
Are also generally differential plating media to distinguish between coliforms (lactose-fermenting) from noncoliforms (non-lactosefermenting) enteric bacilli
SELECTIVE MEDIA
MacConkey Agar (MAC): inhibitor
Crystal violet dye and bile salts i
MAC: pH indicator
neutral red
is a selective and differential medium for the detection of sorbitol-nonfermenting Escherichia coli serotype O157:H7 associated with hemorrhagic colitis that results from the action of a shiga-like toxin
(SLT)
MacConkey Agar with Sorbitol
On standard MacConkey Agar containing [?], this strain is indistinguishable from other lactose-fermenting E. coli.
lactose
Unlike most E. coli strains, E. coli O157:H7 ferments [?] slowly or not at all.
sorbitol
efficacy of MacConkey Agar containing sorbitol instead of lactose as a [?] for the detection of E. coli O157:H7 in stool cultures was determined.
differential medium
Salmonella Shigella Agar (SSA): inhibitor
Bile salt salts, brilliant green and citrates
SSA: pH indicator
neutral red
SSA: Sulfur Source
Sodium thiosulfate
SSA: H2S indicator
ferric citrate - black precipitate
During preparation, heat to boiling to completely dissolve the agar.
SSA
XLD: inhibitor
Bile salts (sodium desoxycholate)
XLD: pH indicator
phenol red
is fermented by practically all enterics except for the shigellae. This property enables differentiation of Shigella.
Xylose
initially produce yellow colonies due to xylose fermentation, results in delayed red colonies due to alkaline amines produced.
Lysine decarboxylation by Salmonella
XLD: H2S indicator
Sodium thiosulfate and ferric ammonium citrate - black centers
HEA: inhibitor
Bile salts
HEA: Carbohydrate for acid production
Lactose, sucrose (saccharose), and salicin
acid fuchsin react with bromthymol blue producing a
yellow color
HEA: Sulfur Source
Sodium thiosulfate
HEA: HS2 indicator
Ferric ammonium citrate - black centers
BSA: inhibitors
Bismuth sulfite and brilliant green
BSA: sulfur source
bismuth sulfite
BSA: H2S production
ferrous sulfate - brown to black color with metallic sheen
BGA: inhibitor
Brilliant green dye
BSA: pH indicator
Phenol red
BSA
produce white to red colonies surrounded by red zones in the medium.
Salmonella species (other than S. Typhi/Paratyphi)
shows no growth to trace growth on BGA.
S. typhi/ paratyphi
For fecal specimens or rectal swabs: INOCULATING MEDIUM
enrichment broth
- Selenite broth
- GN broth
For fecal specimens or rectal swabs:
incubation for:
- Selenite broth
- GN broth
- 35 ± 2°C for 8-12 hours
- 35 ± 2°C for 4-6 hours
For fecal specimens or rectal swabs:
- Then, subculture into:
- BAM
- One medium from each of the groups of selective media (MAC, EMB; SSA, XLD, HAE; BSA, BGA)
For fecal specimens or rectal swabs:
- Incubate at
35 ± 2°C for 18-24 hours.
For non-fecal specimens: INOCULATING MEDIUM
- BAM, and
- MAC or EMB
If negative after 24 hours, reincubate an additional [?]. Select suspicious colonies for definitive biochemical or serologic testing.
24 hours
definitive biochemical or serologic testing
Most common Yersinia species isolated from humans acquired from contaminated food.
Yersinia Enterocolitica
Specimen for Isolation of Yersinia Enterocolitica
Feces
Cold enrichment of feces by incubating cultures at [?] in [?] enhances recovery of Y. enterocolitica.
4 oC for 1-3 weeks
buffered saline
• Subculture for the Isolation of Yersinia Enterocolitica
lactose-negative colonies, flat, colorless, or pale pink, 1-2 mm diameter
On MAC
• Subculture for the Isolation of Yersinia Enterocolitica
[?]
Fermentation of mannitol in the presence of [?] results in a characteristic [?] colony, colorless with red center. Selective inhibition of G+ and G- bacteria is obtained by means of [?[, [?], and [?]. Supplementation with [?] and [?] improves inhibition of normal enteric bacteria
On CIN (Cefsulodin-Irgasan-novobiocin) Agar
- neutral red
- “bull’s-eye”
- crystal violet, sodium desoxycholate and Irgasan (triclosan)
- cefsulodin and novobiocin
The incorporation of four protein derivatives makes TSI nutritionally very rich.
beef extract
yeast extract
peptone
proteose peptone
The lack of inhibitors permits the of all but the most fastidious bacterial species (excluding the obligate anaerobes). For this reason, TSI agar can be used only when testing a bacterial species selected from a single colony recovered on [?] or [?] agar plates.
primary or selective
There are two reaction chambers within the same tube.
The [?] exposed throughout its surface to atmospheric oxygen, is aerobic;
the [?], is protected from the air and is relatively anaerobic.
It is important when preparing the media that the slant and the deep are kept equal in length, approximately [?] each
upper slant
lower butt or the deep
3 cm (1.5 in.)
TSI Reactions: Acid production
[?] are evenly distributed throughout both the slant and butt (deep) portion of the tube. However, glucose is present in a [?] concentration of each of the lactose and sucrose.
Glucose, lactose, and sucrose
1/10
TSI Reactions: Acid production
The phenol red indicator is yellow below a pH of
6.8.
TSI Reactions: Acid production
Because the pH of the uninoculated medium is buffered at [?], relatively small quantities of acid production result in a visible color change.
7.4
The organism ferments glucose, and lactose and/or sucrose producing large amounts of acids, so when examined at the end of 18-24 hours, both the slant and the butt appear yellow.
A/A Acid slant / Acid butt
Glucose-fermenting organism that cannot utilize lactose and sucrose produces only a small quantity of acid from the [?] concentration of glucose in the medium.
0.1%
Initially, during the first [?] hours of incubation (i), acid may be sufficient to convert both the deep and the slant color to yellow.
8 to 12
When the glucose supply is exhausted, and the bacteria begin oxidative degradation of the amino acids within the slant of the tube where oxygen is present.
K/A Alkaline slant / Acid butt
In the deep (anaerobic portion) of the tube, however, [?] is
insufficient to counteract the acid formed, and the medium remains yellow
amino acid degradation
This indicates a lack of acid production and failure of the test organism to
ferment any of the sugars present.
K/K Alkaline slant / Alkaline butt.
This results in the release of [?] that counteract the small quantities of acid present in the slant; by 18 to 24 hours (ii), the entire slant reverts to an alkaline pH, and the color returns to red.
amines
Presence of cracks in the medium or the “pulling away” of the medium from the walls of the test tube.
TSI Reactions
Gas (H2 + CO2) production
source of sulfur atoms used for
hydrogen sulfide production
Sodium thiosulfate
incorporated in the culture media then react with hydrogen sulfide to produce an insoluble black precipitate (ferrous sulfide).
Iron salts (ferrous sulfate)
An acid environment is required for an organism to produce [?] and, therefore, a source of hydrogen ions must be provided. Because the butt of the TSI tubes becomes [?] with glucose fermentation (hydrogen ions increase), the blackening is often first seen or confined there, particularly with non–lactose-fermenting bacteria. Thus, it follows that a black deep should be read as acid even if the usual yellow color is obscured by the black precipitate.
hydrogen sulfide
acidic
The formula of KIA is similar to TSI, EXCEPT that it lacks [?]and the H2S indicator is [?] instead of ferrous sulfate.
sucrose
ferric ammonium citrate
When inoculating a series of tubes of differential culture media with an unknown organism, it is important that the [?] be streaked first to prevent carryover of [?] from the other media.
citrate medium
proteins or carbohydrates
The MR-VP broth culture may be incubated up to
2-4 days
Because indole is [?], xylene or chloroform should be added to the test medium before adding Ehrlich’s reagent
soluble in organic compounds
makes extraction step is less critical for Kovac’s reagent
is used for the diluent (ethyl alcohol is used with Ehrlich’s reagent
amyl alcohol
Because other organisms may produce smaller quantities of acid from the test substrate, an intermediate orange color between yellow and red may develop. This does not indicate a positive test
Methyl red (MR) test
A positive VP test is represented by the development of a red color in [?] or more after addition of the reagents.
15 minutes
The test should not be read after standing for over 1 hour because negative VP cultures may produce a [?] potentially resulting in a false
ositive interpretation
Name the test
copper-like color
Voges-Proskauer (VP) test
sources of carbon used to detect citrate utilization by test bacteria
protein and carbohydrates
When inoculating a series of tubes of differential culture media with an unknown organism, it is important that the citrate medium be streaked first to prevent [?] from the other media
carryover of proteins or carbohydrates
Bacteria that possess the enzyme [?] are capable of cleaving tryptophan in tryptone, thereby producing [?]
tryptophanase
indole, pyruvic acid, and ammonia.
Indole can be detected in tryptophan test medium by observing the development of a red color after adding a solution containing [?] (e.g., Ehrlich’s or Kovac’s reagent)
p-dimethylaminobenzaldehyde
Bacteria that metabolize [?] formed from the fermentation of glucose via the mixed acid fermentation pathway produce sufficient strong acid to maintain a pH [?], the acid color breakpoint of the [?] indicator to change into a red color
pyruvate
below 4.4
methyl red
Bacteria that metabolize pyruvate formed from the fermentation of glucose via the butylene glycol pathway results in the production of [?], a as the chief end product and form smaller quantities of mixed acids.
acetoin (acetyl methyl carbinol)
In the presence of atmospheric oxygen and [?], acetoin is converted to [?], and α-naphthol serves as a catalyst to bring out a red complex
40% potassium hydroxide
diacetyl
Bacteria that utilize [?] as the sole source of carbon can also use [?] as the sole source of nitrogen.
citrate
ammonium phosphate
Extraction of nitrogen from the ammonium salts in the medium produce [?]and [?], which results in an alkaline pH.
sodium bicarbonate (NaHCO3) and ammonia (NH3)
Alkalinization in the medium of SCU Test is represented by development of blue color with [?]—yellow [?] and blue [?]—as the indicator
bromthymol blue
below pH 6.0
above pH 7.6
The ingredients in [?] Medium enable the determination of three activities by which enteric bacteria can be differentiated.
SIM
are indicators of hydrogen sulfide production of SIM Test
Sodium thiosulfate and ferrous ammonium sulfate
The ferrous ammonium sulfate reacts with H2S gas to produce [?], a black precipitate.
ferrous sulfide
The [?] is rich in tryptophan, which is attacked by certain microorganisms resulting in the production of indole.
casein peptone
The [?] is detected by the addition of chemical reagents following the incubation period.
indole
detection is possible due to the semisolid nature of the medium.
from the central stab line indicates that the test organism is motile
Motility
Growth radiating out
Observe for [?] first (and for ?) before the addition of Ehrlich’s or Kovac’s reagent for the detection of indole production.
Such reagent may diffuse into the medium making it [?] which may result in false motility interpretation.
motility
H2S production
hazy or cloudy
Other media for motility testing:
Motilility Test Medium
Motility Indole Ornithine (MIO) Medium
Motility Indole Lysine Sulfide (MILS)
Medium, MIL (Motility-Indole-Lysine)
Lysine is the substrate for use in detecting the enzymes, [?] and [?]
lysine decarboxylase and lysine deaminase
Lysine decarboxylation (LDC+) by bacteria produces alkaline-reacting amine, [?], which causes the pH indicator, [?] — yellow at or [?] and purple [?]. — to change its color to purple.
cadaverine
bromocresol purple
below pH 5.2
at or above pH 6.8
As decarboxylation only occurs in an acidic medium (?), the culture medium must first be acidified by [?]
below pH 6.0
glucose fermentation
LDC+ reaction
purple butt
LDC- reaction
yellow butt
LDC- bacteria ferment [?] and cause the entire culture medium to turn yellow. On prolonged incubation of the culture medium slant may occur, resulting in a color change to violet
glucose
alkalinization
[?] are indicators of hydrogen sulfide formation in LIA.
Cultures of [?] that produce hydrogen sulfide cause blackening of the medium
Ferric ammonium citrate and sodium thiosulfate
enteric bacilli
Lysine deamination (LDA+) gives [?]; this compound reacts with the [?] near the surface of the medium, under the influence of [?], to form reddishbrown compounds
ketocarboxylic acid
iron salt
oxygen
LDA+ reaction
red lant
LDA- reaction
purple slant
LIA reactions
- K/K
Escherichia coli
Klebsiella, Enterobacter (now Klebsiella) aerogenes
Hafni
Serrati
- K/K+ H2S
Salmonella
Edwardsiella
- K/A
Shigella
Citrobacter koseri
Enterobacter cloacae S. Paratyphi A
- K/A+ H2S
Citrobacter freundii
- R/A
Morganella
Providencia Proteus
o may NOT blacken LIA since acid in the butt may suppress H2S
- R/A + H2S
Proteus
Decarboxylase Test: Medium,Component
Moeller Decarboxylase Broth
Urease Test: Medium
Stuart’s Urea Broth, or Christensen’s Urea Agar
Phenylalanine Deaminase (PAD) Test: Medium
Phenylalanine Agar
Phenylalanine Deaminase (PAD) Test: Reagent
10% Ferric Chloride
ONPG Test: Reagents
a. Sodium phosphate buffer, 1 M, pH 7.0
b. o-Nitrophenyl-β-D-galactopyranoside (ONPG), 0.75 M (Buffered ONPG tablets are commercially available.)
c. Physiologic saline
d. Toluene
MUG Test: Reagent
4- methylumbelliferyl-ßD-glucuronide
Decarboxylases react with the [?] (COOH) group of specific amino acids, forming [?] as a second product.
are the three amino acids routinely tested in the identification of the Enterobacteriaceae
carboxyl
alkaline-reacting amines, carbon dioxide
Lysine, ornithine, and arginine
The specific amine products are as follows:
Lysine:
Ornithine:
Arginine:
Lysine → Cadaverine
Ornithine → Putrescine
Arginine → Citrulline
The conversion of [?] to [?] is a dihydrolase, rather than a decarboxylase reaction, in which an [?] group is removed from arginine as a first step.
[?] is next converted to ornithine, which then undergoes decarboxylation to form [?]
arginine to citrulline
NH2
Citrulline
putrescine
The amino acid to be tested is added to the [?] before inoculation with the test organism.
decarboxylase base
A control tube, consisting of only the base without the amino acid, must also be set up in parallel. Both tubes are [?] incubated by overlaying with [?]
anaerobically
mineral oil
During the initial stages of incubation, both tubes turn yellow, owing to the fermentation of the small amount of [?] in the medium. If the amino acid is decarboxylated, [?] are formed and the medium reverts to its original purple color
glucose
alkaline amines
[?] enzyme possessed by many species of microorganisms hydrolyzes urea with the release of [?]. The ammonia reacts in solution to form [?], (NH4)2CO3, resulting in alkalinization of the medium
Urease
ammonia and carbon dioxide
ammonium carbonate
is a compound structurally similar to lactose except that the glucose has been replaced by an o-nitrophenyl group
o-Nitrophenyl-β-D-galactopyranoside
On hydrolysis, through the action of the enzyme [?], ONPG cleaves into two residues, [?]
β-galactosidase
galactose and o-nitrophenol
is a colorless compound
ONPG
yellow compound that provides visual evidence of hydrolysis
o-nitrophenol
Lactose-fermenting bacteria possess both [?], two enzymes required for the production of acid in the lactose fermentation test
lactose permease and βgalactosidase
The [?] is required for the lactose molecule to penetrate the bacterial cell where the β-galactosidase can cleave the galactoside bond, producing [?]
permease
glucose and galactose
Non–lactose-fermenting bacteria are devoid of both enzymes and are incapable of producing acid from lactose. Some bacterial species appear to be non–lactosefermenters because they lack [?], but do possess [?] and give a positive ONPG test
permease
βgalactosidase
So-called late lactose fermenters may be delayed in their production of acid from lactose because of [?]. In these instances, a positive ONPG test may provide a rapid identification of [?]
weak permease activity
delayed lactose fermentation
is a compound structurally similar to lactose except that the glucose has been replaced by an o-nitrophenyl group
Nitrophenyl-β-D-galactopyranoside
ONPG
On hydrolysis, through the action of the enzyme [?], ONPG cleaves into two residues, galactose and o
itrophenol
β-galactosidase
he MUG test is based on the hydrolysis of MUG (4-methylumbelliferyl-βD
lucuronide) by [?], an enzyme produced by most strains of Escherichia coli and other Enterobacteriaceae. The end product of hydrolysis, 4-methylumbelliferone (4- MU), fluoresces blue under [?]
β-glucuronidase
long wavelength ultraviolet light
From a well-isolated colony of the test
organism previously recovered on primary isolation agar, inoculate two tubes of Møller decarboxylase medium, one containing the [?] to be tested and the other to be used as a control tube devoid of amino acid
amino acid
Overlay both tubes with sterile mineral oil to cover about [?] of the surface
1 cm
Incubation for Decarboxylase Test, Urease Test, Phenylalanine Deaminase (PAD)
Test
35°C for 18–24 hours
Incubation for ONPG Test
37°C
Incubation for MUG Test
35°-37°C for 30 minutes
The [?] is inoculated with
a loopful of a pure culture of the test organism; the surface of the agar slant is streaked with the test organism
broth medium
Inoculate the [?] of the medium with a single colony of the test organism isolated in pure culture of primary plating agar
agar slant
After incubation, add [?] directly to the surface of the agar. As the reagent is added, the tube is rotated to dislodge the surface colonies
4 or 5 drops of the ferric chloride
Emulsify a loopful of bacterial growth in [?] to produce a heavy suspension. Bacteria grown in medium containing lactose, such as TSI or KIA, produce optimal results in the ONPG test
0.5 mL of physiologic saline
Add one drop of [?] to the suspension and vigorously mixed for a few seconds to release the enzyme for the bacterial cells
Add an equal quantity of [?] to the suspension
toluene
buffered ONPG solution
Place the mixture in a 37°C water bath.
When using ONPG tablets, a loopful of bacterial suspension is added directly to the ONPG substrate resulting from adding [?] to a tablet in a test tube. This suspension is also placed in a 37°C water bath
1 mL of distilled water
a. Place a MUG disk in an empty sterile petri dish and add a drop of distilled water.
b. Smear 2-3 isolated colonies from
18-24 hour old culture on the disk. c. Place a piece of filter paper saturated with water in the lid of the petri dish to provide humid environment.
d. Incubate aerobically at 35°-37°C
for up to 30 minutes. e. Following incubation, examine the disk for fluorescence using long-wave ultraviolet light (360nm) in a darkened room.
Direct disk test
a. Add 0.25ml of distilled water to a clean glass tube.
b. Make a heavy suspension with 3-4 colonies of test isolate in the tube. c. Using a forcep, place a MUG disk in the tube and shake vigorously to ensure adequate elution of substrate in the surrounding liquid.
d. Incubate aerobically at 35°-37°C for up to 1 hour.
e. Following incubation, examine the disk for fluorescence using longwave ultraviolet light (360nm) in a darkened room
Tube test
(+) Purple color
(-) Yellow color
Decarboxylase Test
(+) Pink-red color
(-) Yellow color
Urease Test
(+) Green color
(-) No green color
Phenylalanine Deaminase (PAD) Test
The rate of [?] of ONPG to o-n itrophenol may be rapid for some organisms, producing a visible yellow color reaction within 5–10 minutes
hydrolysis
Most tests are positive within 1 hour; however, reactions should not be interpreted as negative [?]. The yellow color is usually distinct and indicates that the organism has produced onitrophenol from the ONPG substrate through the action of [?]
before 24 hours of incubation
βgalactosidase
Alternatively, the disk can be placed directly on the agar surface. In which case, it will not require the addition of water because moisture from the medium will rehydrate the disk
Mug Test
It is used to screen isolates of Escherichia coli, majority (97%) of which is [?]. Some strains of other organisms other than E. coli (e.g., [?], etc.) also possess the enzyme β-glucuronidase. Hence, the detection of the [?] enzyme is commonly employed in laboratories to identify and differentiate such organisms
MUG-positive
Salmonella, Shigella, Staphylococcus, Streptococcus
β-glucuronidase
It is used for the detection of E. coli serotype [?] strains. This specific serotype is [?]
O157:H7
MUG negative (and sorbitol negative)
False negative reactions may be reported when test colonies isolated from media containing [?] are used and hence it may make the interpretation difficult
dyes (EMB, MAC)
is the only species in the genus Plesiomonas
Plesiomonas shigelloides
oxidase- and catalase-positive
glucose-fermenting
gram-negative rod with polar flagella
Plesiomonas shigelloides
Because of phenotypic characteristics, the genus Plesiomonas was formerly in the family [?], however, recent molecular genetic evidence demonstrates that the genus Plesiomonas is most closely related to the genus [?]. Thus it has recently been moved into the [?] family as the only [?] member of the group
Vibrionaceae
Proteus
oxidase-positive
P. shigelloides is found in aquatic environments that are limited geographically by its minimum growth temperature of [?]. It may be found in fresh and estuarine water, usually in [?] countries
8° C
tropical
P. shigelloides has emerged as a potential cause of enteric disease in humans, most often after the consumption of [?] or [?]
undercooked seafood or untreated water
At least three major clinical types of gastroenteritis are caused by Plesiomonas:
‣ The more common [?
‣ A [?] that lasts from 14 days to 2 to 3 months
‣ A more invasive, dysenteric form that resembles [?]
watery or secretory diarrhea
subacute or chronic disease
colitis
Plesiomonads have also been isolated from a number of extraintestinal infections, most notably
meningitis in neonates, septicemia and shock
[?] can be a source of infections for veterinarians, zookeepers, aquaculturists, fish handlers, and athletes participating in water-related sports
Occupational exposure
More serious infections, such as [?] and [?], usually occur only in severely immunocompromised patients or neonates
bacteremia and meningitis
