Sunday, 9 December 2007

MMIC - Follow up on Specimen 3.

Specimen 3:
Preliminary Information:

Patient: Maisy Hong (67 years)
Sample: Urine
Symptoms: Fever, chills, bladder distension; on indwelling catheter
Bladder distension: Inability to urinate. May be due to obstruction.
Indwelling catheter: prevent catheter from being pulled out of the bladder

Diagnosis: Urinary Track Infection

From week 1:

Possible causative agents: catheter- associated UTI

Most probable agents: according to incidence seen in UTI patients
1) Escherichia coli – 53-72%
2) Klebsiella spp. (pneumoniae) – 6-12%
3) Proteus spp. (mirabilis) – 4-6%
4) Enterococcus spp.(faecalis) – 2-12%

Additional suspected agents:
5) Pseudomonas spp. (aeruginosa)
6) Enterobacter spp. (aerogenes)
7) Serratia spp. (marcescens)
8) Candida spp. (albicans): may cause UTI, associated with use of catheter

Note: Chlamydia and Mycoplasma were excluded as these are typically sexually transmitted, highly unlikely to be the cause in a 67 year old woman with a catheter

Preliminary tests:
Specimen: Urine (dried on slide)

Fungal stain
* Periodic acid-Schiff (PAS) can be used to detect if causative agent is fungi.
Gram stain
*Used to detect if bacteria is
- gram negative
- Escherichia coli (rod)
- Proteus Species (rod)
- Enterobacter Species (bacilli)
- Klebsiella Species (bacilli)
- Pseudomonas Species (bacilli)
*gram positive
- Enterococcus Species (cocci)

After the above test have been done, it would be clearer as to whether it is a fungi or bacteria. If it is a bacteria, it would have been narrowed down to gram type. Further screening as shown below for each individual suspected agent will then be performed if agent falls under the initial testing results.

Individual tests:

Escherichia coli
Morphology (microscopy): Gram negative bacilli, singly.
Pathogenesis: Adherence factor P fimbria binds specific galatose disaccharide found on uroepithelial cells surfaces. Siderophores produce cytotoxic alpha and beta hemolysins for lysis of urinary track cells for iron acquisition.
Key Characteristics: Rapidly ferment lactose, positive indole test, positive B-glucaronidase (using substrate MUG), facultative anaerobic (mixed-acid fermentation)
1) EMB (Eosin Methylene Blue). Detect lactose fermenting colonies. Contains eosin and methylene blue.
· Black colonies with green sheen (positive, only E.coli can produce this colour due to acid production)
2) MacConkey Agar. Contains bile salts, crystal violet dye (to inhibit Gram-positive bacteria), neutral red dye (which stains microbes fermenting lactose).
· Red colonies (positive)
3) Indole test. Measures the ability of the microorganism to degrade tryptophan into indole, ammonia and pyruvic acid. Pure bacterial culture must be grown in sterile tryptophan or peptone broth for 24-48 hours prior to test. Following incubation, add 5 drops of Kovac's reagent.
· Red or red-violet color (positive)
4) Methyl Red (MR).
· Red (positive)
5) Citrate test. Utilizes Simmon's citrate media to determine if a bacterium can grow utilizing citrate as its sole carbon and energy source
· Green (negative)
6) MUG (on nutrient agar). Nutrient Agar with methylumbelliferyl-ß-D glucuronide (MUG) detects Escherichia coli (E.coli).
· Colonies with bright blue fluorescence
7) Blood agar plates (β-hemolytic)
· Hemolysis (Positive)
8) Urease Test
· Orange (Negative).
9) Triple Sugar Iron (TSI). Contains phenol red, high concentrations of lactose and sucrose, and a low concentration of glucose as well as sodium thiosulfate and ferric citrate . Provides aerobic and anaerobic conditions. Detects lactose fermentative bacteria – any bacteria that can ferment any of the three sugars.
· Red agar turns yellowish (positive). Acidic slant & butt (lactose and glucose fermenter)
· Stormy fermentation – production of hydrogen gas
10) Antibiotic Susceptibility :
· Amoxicillin, cephalosporins, carbapenems, aztreonam, trimethoprim-sulfamethoxazole, ciprofloxacin, nitrofurantoin and the aminoglycosides
11) Serology. Detection of O & H antigens (ELISA)
· Slide agglutination – Positive
· Tube agglutination - Positive
Biochemical summary:

Klebsiella spp. (pneumoniae)
Morphology (microscopy): Encapsulated gram negative rod (pH 5, acidic) may be cocci in more alkaline conditions (pH 7)
Pathogenesis: Type 1 fimbriae consisting of building block protein (FimA) extend beyond the capsular matrix and mediate adhesion to host epithelial cells.
Capsular polysaccharide matrix encases cell surface to provide resistance against host defence mechanisms.
Key characteristics: Negative indole test. Positive Voges Proskauer (Vi). Lactose fermenting, facultative anaerobic.
1) MacConkey Agar.
· Red colonies (positive)
2)Triple Sugar Iron (TSI).
· Red agar remains red (negative). Alkaline slant & butt. (non-fermenter)
3) Indole test.
· Yellow (negative)
4) Voges-Proskauer (VP) test. Determines if neutral products such as ethyl alcohol, acetoin and butanediol are formed
· Pink-burgundy color (positive), 30mins to develop colour
5) Methyl Red (MR) test:
· Yellow (negative)
6) Citrate test. Utilizes Simmon's citrate media to determine if a bacterium can grow utilizing citrate as its sole carbon and energy source
· Prussian blue color (positive)
7) Urease Test.
· Orange (Negative).
8) Antibiotic Susceptibility :
· Third generation cephalosporin
9) Serology. Capsular serotyping
· K antigens
Biochemical summary:

Proteus spp. (mirabilis)
Morphology (microscopy): Gram negative bacilli.
Key Characteristics: Mostly positive indole test, except P. mirabilis. Deaminate phenylalaine. Grow on potassium cyanide medium. Ferment xylose. Motile, swarming effect on agar plates. urease-positive. Facultative anaerobe.
1) Oxidase test
· Negative
2) Triple Sugar Iron (TSI).
· Black precipitation (H2S production), red slant (alkaline). Glucose fermenter, non-lactose fermenter.
3) Indole Test.
· Most strains positive (red), mirabilis negative (yellow).
4) Methyl Red Test
· Red (positive)
5) Voges-Proskauer (VP) test.
· Negative.
6) Citrate test.
· Prussian blue color (positive)
7) Urease Test.
· Pink (Positive).
8) MacConkey Agar
· Light/colourless colonies (negative)
9) OILM medium. Test for ornithine decarboxylase utilization.
· Green/blue upper portion (Positive).
10) Blood agar plates
· Swarming observed
11) Antibiotic Susceptibility
· Penicillin derivatives, cephalosporins, quinolones, aminoglucosides
Biochemical summary:

Enterococcus spp.(faecalis)
Morphology (microscopy): Gram positive cocci. (diplococci)
Key Characteristics: catalase negative, colonies appear grey. Group D streptococci. gamma hemolytic (no hemolysis)
1) Catalase test. Identifies organisms which produce the catalase enzyme; this enzyme converts hydrogen peroxide to water and oxygen gas
· Negative
2) Blood plate agar
· No hemolysis
3) Bile-Esculin Hydrolysis Test. Determine the ability of an organism to hydrolyze the glycoside esculin to esculatin and glucose in the presence of bile (10 - 40%).
· Black to dark brown slant (Positive)
4) 6.5% salt broth. Test salt tolerance.
· Positive, growth.
5) PYR (pyroglutamyl aminopeptidase) disc
· Positive
6) Xylose fermentation test. performed with D-xylose tablets.
· Positive
7) MacConkey Agar. Without crystal violet for inhibition of gram positive bacteria.
· Pink (positive)
8) Serology. Western blot (IgG from patient’s sera). ELISA.
· Polysaccharide antigen extracted from bacterial cell walls detected
9) Antibiotic Susceptibility
· Quinupristin, ampicillin and vancomycin

Pseudomonas spp. (aeruginosa)
Morphology (microscopy): Gram negative bacilli.
Key Characteristics: Aerobic (classified facultative anaerobe), rod-shaped bacterium with unipolar motility (due to single polar flagellum), opportunistic pathogen (seen in immunocompromised individuals). Pearlescent appearance and grape-like odor
1) MacConkey Agar
· Colourless colonies - Negative (non glucose fermenter)
2) Catalase test. Identifies organisms which produce the catalase enzyme; this enzyme converts hydrogen peroxide to water and oxygen gas
· Positive
3) Blood plate agar
· swarming
4) Oxidase test
· Positive
5) Urease Test
· Positive
6) Indole Test
· Negative
7) Methyl Red
· Negative
8) Voges Proskauer Test
· Negative
9) Citrate test.
. Prussian blue color (positive)
10) TSI
· red slant (alkaline), red butt (alkaline) . Non fermenter of glucose
11) Antibiotic Susceptibility
· Aminoglycosides, Quinolones, Cephalosporins
12) Serology. Detect A, B , O antigens on cell wall
· Slide agglutination – Positive
· Tube agglutination - Positive
Biochemical summary:

Enterobacter spp. (aerogenes)
Morphology (microscopy): Gram negative bacilli.
Key Characteristics: Facultative anaerobe, oxidase negative, catalase positive. Typically caused by catheter insertions
1) MacConkey Agar
· Pink colonies - Positive, may be weak (glucose fermenter)
2) Catalase test. Identifies organisms which produce the catalase enzyme; this enzyme converts hydrogen peroxide to water and oxygen gas
· Positive
3) Oxidase test
· Negative
4) Urease Test
· Variable
5) Indole Test
· Negative
6) Methyl Red
· Negative
7) Voges Proskauer Test
· Positive
8) Citrate test.
· Prussian blue color (positive)
9) TSI
· red slant (alkaline), yellow butt (acidic) . Fermenter of glucose
10) Antibiotic Susceptibility
. Ciprofloxacin, Tazobactem
Biochemical summary:

Serratia spp. (marcescens)
Morphology (microscopy): Gram negative bacilli.
Key Characteristics: Produces lactose-fermenting colonies on differential agar, but is a late lactose fermenter may seem like a negative reaction (considered negative). Forms red-pigmented colonies. Able to perform casein hydrolysis and degrates tryptophan and citrate.
1) MacConkey Agar
· Red colonies. Negative (initial)
2) Catalase test. Identifies organisms which produce the catalase enzyme; this enzyme converts hydrogen peroxide to water and oxygen gas
· Positive
3) Oxidase test
· Negative
4) Urease Test
· Positive
6) Indole Test
· Negative
7) Methyl Red
· Negative
8) Voges Proskauer Test
· Positive
9) Citrate test.
· Prussian blue color (positive)
10) TSI
· red slant (alkaline), yellow butt (acidic) . Fermenter of glucose
11) Antibiotic Susceptibility
. Aminoglycoside , Amikacin, Gentamicin, Tobramycin and Quinolones
Biochemical summary:

Candida spp. (albicans)
Morphology (microscopy): chlamydospores.
Pathogenesis: Produces extracellular proteinases from SAP genes, phospholipases B enzymes and lipases that are secreted for host infection.

1) Southern Blot. Detect SAP genes using SAP1 probe.
· Positive.
2) Serology. Detect cell wall antigens - ELISA (CWP, PPM, CW)
· Positive.
3) Oxoid OBIS albicans Test. Rapid card-based chromogenic test that detects the presence of two enzymes specific to C. albicans and C. dubliniensis: b-galactosaminidase and L-proline aminopeptidase
· Magenta colour (Positive).

References: > e.coli.html > userguides > tech1 > p35681 > johnson > labtools > Dbiochem > imvic.html > cgi > content > abstract > 73 > 8 > 4626 > cgi > reprint > 28 > 4 > 534.pdf > eccmid15 > abstract.asp?id=36567 > ClassFiles-LA > Microbiology > entero.htm > microbugz > html > catalase_test.html

Debra, TG02

Case 5 follow up

WFH's case of wound infection.

Revised list of suspected microorganisms:
1. S. aureus
2. S. pyrogenes
3. Clostridium perfringens
4. Clostridium botulinum
5. Pseudomonas aeruginosa
6. S. faecalis (enterococcus)
7.Proteus mirabilis


Key characteristics

Lab diagnosis

S. aureus

Produce coagulase enzyme

Coagulase positive (agglutination)

Produce catalase

Catalase positive [Frothing when react with H2O2 ]

Ferments mannitol

Yellow colonies on mannitol salt agar

Lyses rbc

B haemolysis on BA

B lactamase

B lactam resistant. Sensitivity to naficin, methicillin, vancomycin

Gram positive (purple) cocci, “grape-like”

S. pyogenes

Lyses rbc

B haemolysis on BA

Does not produce catalase

Catalase negative

Bacitracin sensitive

Hippurate and cAMP negative

Gram positive cocci in chains

C. perfringens

Double zone of haemolysis

B haemolysis on BA

Alpha lecithinase

Lecithinase positive on egg yolk agar

Gram positive, large pink rods

Proteolysis on CM (meat turn black)

C. botulinum


Swarming effect on agar

Lipase positive

Lecithinase and protease neg

Meat at the bottom of CM remain brown

Gram positive, oval and subterminal spores, bacilli

P. aeruginosa

Non fermentative

Pink colonies on macconkey agar

Oxidase positive

Pyocyanin pigment

Blue on pseudomonas agar P

Pyoverdin pigment

Fluorescein on Pseudomonas agar F

Utilizes citrate

Citrate + (green to blue on citrate agar slant)


Unable to hydrolyse and deaminate tryptophan

Indole -

Does not utilize carbohydrates


S. faecalis

Non haemolytic

Gamma haemolysis on BA

Tolerant to 6.5% NaCl

Growth on mannitol salt agar

Resistant to bile

Hydrolyses esculin

Blackening of esculin agar

Does not produce B lactamase

B lactam sensitive

Gram positive cocci in chains

P. mirabilis

Unable to metabolise lactose

Pink colonies on MAC

Distinct ordor

Utilises urea and citrate

Citrate +

H2S production

Black precipitate on TSI

Unable to hydrolyse and deaminate tryptophan

Indole -

Nitrogen reductase -

Mixed acid fermentation

Methyl red +

Unable to ferment glucose

Voges- Proskauer -

Produce catalase

Catalase +

Oxidase -

Phenylalanine deaminase +

Yeng Ting
Tg 02

MMIC blog 2 - case 4

Probable microorganisms:

Streptococcus pneumoniae
- gram-positive
- alpha-haemolytic diplococcus bacterium
- optochin sensitive

- produce IgA protease that enhances organism’s ability to colonize the mucosa of the upper respiratory tract. It multiply in tissue and causes inflammation.

Lab investigation:
- Gram stain
- Culture on blood agar plates
- catalase negative
- antibiotic susceptibility – susceptible to penicillins and erythromycin.

Moraxella catarrhalis
- gram-negative
- aerobic
- diplococcus
- produce beta-lactamases

- cause respiratory tract-associated infection in humans

Lab investigation:
- Gram stain
- oxidase negative
- antibiotic susceptibility – susceptible to cephalosporins, resistant to penicillin

Haemophilus influenzae
- gram-negative
- coccobacilli
- non-motile
- generally aerobic, but can grow as a facultative anaerobe.

- Their capsule allows them to resist phagocytosis and complement-mediated lysis in the non-immune host. Unencapsulated strains are less invasive, but they are able to induce an inflammatory response that causes disease.

Lab investigations:
- Gram stain
- culture on blood agar
- catalase positive
- oxidase positive

Mycoplasma pneumoniae
- causes infectious bronchitis

Lab investigations:
- lack of bacteria in a gram stained sputum sample
- lack of growth on blood agar
- positive blood test for cold hemagglutinins.
- Antibiotic susceptibility – susceptible to erythromycin

Chlamydia pneumoniae
- causes infectious bronchitis

Lab investigations:
- Giemsa stain
- Serologic test for antibody in patient’s serum.
- Antibiotic susceptibility – susceptible to tetracycline such as doxycycline

Bordetella pertussis
- gram-negative
- coccobacillus
- aerobic
- non-motile

Lab investigation:
- Culture on Bordet-Gengou agar plate with added cephalosporin select for the organism
- Oxidase positive
- urease negative
- nitrate negative
- citrate negative
- Antibiotic susceptibility – susceptible to erythromycin

Klebsiella pneumoniae
- gram-negative
- facultative anaerobic
- non-motile
- lactose fermenting

Lab investigations:
- Culture – MacConkey’s agar with lactose fermenting colonies.
- oxidase negative
- TSI: slant & butt yellow (acidic)
- H2S negative
- indole negative
- methyl-red negative
- Voges-Proskauer positive
- citrate positive
- urease positive
- Antibiotic susceptibility – susceptible to cephalosporin, resistant to penicillin

References:> Lung and Airway Disorders> Bronchitis> Streptoccocus pneumoniae> Moraxella catarrhalis> Haemophilus influenzae


Saturday, 8 December 2007

Cheng Hong: MMic PBL Kuan Siew Yan follow up

There are some other organisms and viruses that are able to cause diarrhea like:

Clostridium perfringens
Staphylococcus aureus
§ Rotavirus
§ Norovirus
§ Vibrio cholerae

Clostridium perfringens
Characteristics: Rigid, thick walled cell, gram positive, spore forming anaerobic rods

Pathogenesis: Causes gas gangrene and food poisoning. C. perfringens is a normal flora in the colon but not found in the small bowels. If it is in the small bowels, the enterotoxin (super antigen on the C. perfringens) will cause diarrhea.

Why is not likely in this case: The diarrhea will only last for 24hrs and the disease will be resolved after 24hrs. This bacteria do not cause enterocolitis.

Staphylococcus aureus
Characteristics: Rigid, thick walled cell, gram positive cocci

Pathogenesis: Causes food poisoning. The enterotoxin causes food poisoning (vomiting/ watery, non-bloody diarrhea). The enterotoxin acts like a super antigen and stimulate the release of interleukin-1 and 2. May be transmitted through improperly cooked food as S. aureus is quite heat resistant.

Biochemical features: Coagulase positive, positive latex test, plate on Mannitol salt agar which acts as a selective medium and differential medium

Others: Smears will show gram positive cocci in clusters (purple).

Characteristics:Reovirus family, non-enveloped, ssRNA virus, Icosahedral capsid

Pathogenesis: Causes diarrhea in young children, transmitted through fecal oral route. The virus multiplies in the small intestine and causing salt, glucose, water to be loss through diarrhea.

Diagnosis: Using ELISA techniques/ radioimmunology/ Rapid testing kits

Why it is not likely in this case: As Rotavirus infection mostly occurs in babies and very young children. Adults rarely will be infected as at the age of 6, children will have antibodies against at least 1 serotype of rotavirus.

Characteristics: Calicivirus family, non-enveloped, ssRNA virus, Icosahedral capsid

Pathogenesis: Transmitted through fecal oral route, or ingesting contaminated seafood/water. Virulent (low infectious dose). May cause vomiting, fever, diarrhea.

Diagnosis: PCR method or ELISA method

Vibrio cholerae
Characteristics: Rigid, thick walled cell, Gram negative, Facultative curved rods (comma shape)

Pathogenesis: Transmitted through fecal contamination in water and food, found in shellfish/ oysters. It causes watery diarrhea by causing the cells in the gut to lose water and ion. It does not cause bloody diarrhea or abdominal pain.

Biochemical features: oxidase positive, acid slant and acid butt, but no gas or H2S on TSI. Can be confirmed usingagglutination test (polyvalent O1 or non-O1 antiserum.

Please click the link below to view the biochemical test. (Maximise the table for better viewing)

References from:Review of Medical Microbiology and Immunology(9th edition). WARREN LEVINSON
:Color Atlas of Medical Bacteriology. ASM PRESS

MMIC Blog 2 - Case 6

[A follow-up on my patient, Ong Fei Fei's investigation]

Laboratory Diagnosis

Since the patient has been previously diagnosed with UTI, the microbe could have move down and infect the vagina as well, hence urethra microbes are considered in this case too, for examples:

E. coli
Proteus mirabilis
S. saprophyticus
Group B streptococci (Alpha & non-hemolytic)
Coagulase-negative Staphylococci (S. aureus)
Klebsiella species
Proteus species
Pseudomonas aeruginosa

(Follow how UTI microbes are identify in other cases - see debra's and/or elaine's)

As for more vaginal-specific microbes, they are the followings:

Trichomonas Vaginalis

  • Wet mount preparation then Direct microscopy

  • Motile characteristic

  • Greater sensitivity: Fixed stained preparation (Giemsa/Papanicolaou) then Direct Immunofluorescence

  • Culture (Diamond’s medium) then incubate at 37oC à microscopy

  • Antibody detection techniques: high in false positive and false negative results

  • Enzyme immunoassay for detection of T. vaginalis antigen

Candida Albicans

  • Gram’s stain then Direct Microscopy

  • Sabouraud’s agar then Incubate 24-48 hours. white, butyrous colonies observed if positive
Bacterial Vaginosis (anaerobic/non-specific)

  • Clue cells (Vaginal epithelial cells with edges darkened by presence of numerous small bacteria adhering to their surface)

  • pH (>4.5)

  • Amine test: add few drops of KOH and presence of amines gives a fishy smell

  • Gram’s stain

  • Culture: for Gardnerella vaginalis and Mobiluncus species

1. Gardnerella vaginalis

Slow growing
Catalase and oxidase negative
Beta-hemolysis on human blood agar but not on sheep blood agar
Selective blood agar: add gentamicin, nalidixic acid and amphotericin B
Hippurate hydrolysis: positive
Starch fermentation: positive
Metronizadole 50ug disc: sensitive
Sulphonamide 1000ug dsc: resistant

2. Mobiluncus species

Divided into: M. curtisii and M. mulieris
Fastidious; slow growing
Typically motile, catalase, oxidase, indole negative
Clear, colourless colonies (2mm) after 5 days incubation
Gas-liquid chromatography distinguish species
Commercial kit: detect enzyme activity (praline aminopeptidase and alpha-D-glucosidase)

3. Neisseria Gonorrhoea

Die readily outside human body
Use of rich media supplemented with yeast extract or iso-viatalex and blood (Chocolate agar)
Antibiotic inhibitors: vancomycin, nystatin, colistin, trimethoprim
Direct Microscopy (less sensitive for women – 50%)
Other technique: Direct immunofluorescence

4. Chlamydia trachomatis

[Direct examination of smear with fluorescein-conjugated monoclonal antibodies(Ab) (use of commercial kits)]

Roll specimen gently on slide
Fix with methanol for 4 min
2 Ab (one directed to the outer membrane – species specific, one specific to the genus lipopolysaccharide)
Subjective/ false positive
Suitable for small sample numbers and rapid screening


Uses McCoy’s cells treated with cyclohexamide
Centrifuged and incubate for 72 hours
Detection by Giemsa stain/iodine/fluorescein-labeled monoclonal Ab
Sensitivity: ~80%

[Enzyme-linked immunoabsorbent assays (ELISA)]

Uses polyclonal and monoclonal antibodies against lipopolysaccharide (Antigen detection)
Sensitivity: ~97%; Specificity: ~92.5% (improves by blocking tests)
False positive due to cross reactivity with other bacteria

[Nucleic acid probes]

DNA hybridization
Highly specific but lack sensitivity


Complement fixation tests are insensitive; difficult to differentiate the serotypes
Micro-immunofluorescence test detect specific IgG/IgM; difficult in sexually active populations

Summary Diagrams

Other Investigation required

Urinalysis - for screening purpose; inexpensive and easy to perform (previously confirmed UTI)
Urine culture - for accurate diagnosis of infection to determine complications, such as antimicrobial susceptibility of infecting bacteria (previously confirmed UTI)

References > TrendsInUGSH > Features > sfp > 23 > 232> articles > e232136.html > uti_lower_women > in_depth > goals_and_outcome_measures > mmpe > sec17 > ch231 > ch231b.html > afp > 20020415 > 1589.pdf medical > firstaidfortheboards > pdf > 0071443363 > 0071443363_282.pdf son > nrsg835 > gyninfect.htm

Posted by: Pei Shan, TG02

Friday, 7 December 2007

Elaine Blog2: case 1 Khong Fay seah

The entrobacteriaceae are heterogeneous group of gram-negative rods whose natural habitat is the intestinal tract of humans and animals. The family includes: Escherichia, shigella, salmonella, enterobacter, klebsiella, serratia.

E.coli form circular, convex, smooth colonies with distinct edges. Klebsiella colonies are large and very mucoid and tend to coalesce with prolonged incubation.
Salmonella and shigellae produce colonies similar to E.coli but they do not ferment lactose. However, salmonella and shigellae do not appear in the urinary tract.

E.coli is the most common cause of UTI and accounts for approximately 90% of first urinary tract infections in young women. The symptoms and signs include urinary frequency, dysuria, hematuria, and pyuria. However, none of these symptoms or signs is specific for E.coli.


  • Urine collected from clean-catch mid-stream or one obtained by bladder catherterization or suprapubic.

  • common causative: E.coli; Enterobacteriaceae; other gram-negactive rods

  • usual microscope: gram-negative rods seen on stained smear of uncentrifuged urine indicate more than 105/mL

  • culture on blood agar and macConkey

  • Comments: gray colonies that are B-hemolytic and give a positive spot indole test are usually E.coli; other require further biochemicsl tests

E.coli--> colonies show metallis on EMB agar

Klebsiella pneumoniae--> has large mucoid capsule and hence viscous colonies

Proteus mirabilis--> motility causes "swarming" on agar; produce urease

Pseudomonas aeruginosa--> blue-green pigment and fruity odor produced; causes nonsocomial infections and often drug-resistant

If unknown-form of colonies are observed (other than E.coli) , biochemical tests should be done to differentiate each micro-organism from each other.

Diagnosis: Biochemical Reaction:

IMViC (Indole Methy Re€d Voges-Proskauer simmons's Citrate) plus lactose fermentation & TSI


  1. E.coli ++--+ acid slant/acid butt with gas production

  2. Pseudomonas Aeruginosa -+--- alkaline slant/alkaline butt plus positive oxidation fermentation

  3. Klebsiella Pneumoniae +-+++ acid slant/acid butt with gas production

  4. Proteus Mirabilis -+++- alkaline slant/acid butt with H2S production

Other possible bacteria and tests required:

5. Enterococcus Faecalis

  • growth in the presence of bile, hydrolyze esculin

  • Alpha-hemolytic

  • growth in 6.5% NaCl, PYR-positive

6. Staphylococcus Saprophyticus

  • -ve mannitol

  • -ve coagulase

  • -ve Novobiocin sensitivity

  • -ve alpha toxin

7. Neisseria Gonorrhoeae

  • +ve glucose

  • -ve maltose

  • -ve Lactose

  • -ve Sucrose

8. Mycoplasma Genitalium

  • culture is difficult.

  • Data obtained from PCR, molecular probes, serologic tests

9. Ureaplasma Urealyticum

  • It required 10% urea for growth

Antibiotic Susceptibility Testing:

Escherichia coli (gram -ve rods enteric and related organisms)
drug choice: Cefotaxime, Fluoroquinolones, nitrofurantoin

Pseudomonas aeruginosa (gram -ve rods aerobic)
drug choice: Aminoglycoside, pencillin

klebsiella pneumoniae (gram -ve rods enteric and related organisms )
drug choice: A cephalosporin

proteus mirabilis (gram -ve rods enteric and related organisms)
drug choice: Ampicillin

Staphylococcus saprophyticus (gram +ve cocci)
drug choice: penicillin, tetracycline, vancomycin, gentamicin, rifampin

Neisseria gonorrhoeae (gram -ve cocci)
drug choice: ceftriaxone, ciprofloxacin, gatifloxacin,

Enterococcus faecalis (Pos cocci)
drug choice: Ampicillin, gentamicin

Mycoplasma genitalium (wall-less cells)
drug choice: erythromycin (ERY), clarithromycin (CLR), ciprofloxacin hydrochloride (CIP)

Ureaplasma urealyticum
drug choice: ciprofloxacin and ofloxacin, tetracycline and doxycycline, roxithromycin, erythromycin

Monday, 3 December 2007

MMIC BLOG 1 - Case 6

Preliminary Information

Name: Ong Fei Fei
Sex: Female
Age: 37
Clinical Diagnosis: Urinary Tract Infection (UTI)
Symptoms: Fever, pain during urination, virginal discharge
Anitbiotic Treatment: none
Specimen: Vaginal discharge


  • More common in female than male
  • Infection occurs less frequently in men because the urethra is much longer and the distance between the anus and urethral meatus is greater than in women
  • Prostatic secretions also confer a degree of antibacterial activity
  • A single episode of UTI in women is usually uncomplicated
  • UTI infection is limited to the bladder - the responsible organisms are usually gram negative coliforms
  • Ascending infection from the bladder can cause acute pyelonephritis which typically presents with loin pain, fever and chills, costovertebral angle tenderness, nausea and vomiting
  • A woman with UTI symptoms without significant bacteriuria finding on culture = acute uretheral syndrome (in 30% cases; 70% bacterial infections)
  • Patients with acute uretheral syndrome can be either with pyuria on urinalysis (70%) or without (30%)
  • Those with pyuria have true microbial infection usually with chlamydia; those without pyuria have no known microbial cause and the dysuria (a burning sensation in the urethra during voiding) and frequency may be related to irritation from mechanical trauma
  • Mostly caused by Commensal colonic gram-negative aerobic bacteria (> 75%)E.g. E. coli strains (with specific attachment factors for transitional epithelium of the bladder and ureters), other enterobacteria, especially Klebsiella, Proteus mirabilis, and Pseudomonas aeruginosa.
  • Enterococci (group D streptococci) and coagulase-negative staphylococci (eg, Staphylococcus saprophyticus) are the most frequently implicated gram-positive organisms.
  • Infection Localization - Vaginitis is often distinguished by the presence of vaginal discharge, vaginal odor, and dyspareunia

[Vaginal Discharge]
  • It is a secretion produced from glands in the vaginal and the cervical lining
  • All women have a little discharge starting approximately a year before puberty and ending after the menopause
  • A normal physiological discharge is usually clear, creamy or very slightly yellow
  • Abnormalcy when there is suddenly a great amount of discharge or the color is brownish, reddish or greenish or it becomes smelly.
  • Pathogens implicated are bacterial, fungal and protozoan
  • May be associated with Sexually Transmitted Disease (STDs); due to hypoestrogenism


Trichomonas Vaginalis
  • Transmission: STD
  • Symptoms: Frothy green/yellow discharge, pruritus, urinary symptoms. May be asymptomatic (50%)
  • Protozoa found only in tropozoite stage, with 4 flagella
  • May be seen in urine
  • Offensive odour
  • Vagina may be inflamed (Complain of Vaginal/Vulval pruritis)
  • May have cervical erosions
Candida Albicans

  • A type of fungal (yeast) infection
  • Transmission: Increased with pregnancy, Diabetes Mellitus & antibiotics.
  • Symptoms: Discharge is white, resembles milk curds; Severe itching, dysuria, dyspareunia.

Bacterial Vaginosis (anaerobic/non-specific)

(Rarely presented with dysuria or dyspareunia)

1. Gardnerella vaginalis
Transmission: unknown; may be STD
Symptoms: Thin, watery, yellow-gray discharge with "fishy" odor

2. Mobiluncus species

3. Gonorrhoea
Transmission: STD (More easily transmitted from an infected man to womon ~90%)
Symptoms: Purulent vaginal discharge, dysuria, urinary frequency, inflammation, vulvar swelling and the cervix may be eroded. Most women are asymptomatic.

4. Chlamydia trachomatis (C. trachomatis, C. psittaci, C. pneumoniae)
Transmission: commonly STD
Symptoms (mild): May have thin/purulent discharge, urinary burning and frequency, lower abdominal pain and friable cervix. Women usually asymptomatic.

Possible Interpretation
Vaginal Discharge - suggests Vaginitis or urethritis (e.g. Sexually transmitted Disease (STD), candidiasis) due to hypoestrogenism: e.g. T. vaginalis or N. gonorrhoeae.

Investigation required
  • Urinalysis - for screening; inexpensive, easy to perform
  • Urine culture - for accurate diagnosis of infection to determine complications, such as antimicrobial susceptibility of infecting bacteria
  • Vaginal and urethral discharge - wet-mount preparation to detect Trichomonas vaginalis and Candida species
  • Gramstaining - detect Neisseria gonorrhoeae

References: > TrendsInUGSH > Features > sfp > 23 > 232> articles > e232136.html > uti_lower_women > in_depth > goals_and_outcome_measures > mmpe > sec17 > ch231 > ch231b.html > afp > 20020415 > 1589.pdf

Posted by: Pei Shan, TG02

MMIC PBL - Case 3

Specimen 3:
Preliminary Information:
Patient: Maisy Hong (67 years)
Sample: Urine
Symptoms: Fever, chills, bladder distension; on indwelling catheter
Bladder distension: Inability to urinate. May be due to obstruction.
Indwelling catheter: prevent catheter from being pulled out of the bladder
Diagnosis: Urinary Track Infection

- Women are more prone to UTI as the urethra is much shorter and closer to the anus than in males. Also, they lack the bacteriostatic properties of prostatic secretions.
- Use of urinary catheters (foreign body) in elderly may result in an increased risk of urinary tract infection.

Possible causative agents:

1. Bacteria: Causes up to 85% of all UTI cases.
(a) Escherichia coli: Most common causative agent of UTI, typically from bowels (colon).

(b) Enterococcus Species: Originate from colon.
E. faecalis

(c) Klebsiella Species: Opportunistic pathogens that causes nosocomial infections
K. pneumoniae

(d) Enterobacter Species: Opportunistic pathogen; Present in large intestine but also present in soil and water.
E. aerogenes
E. cloacae
E. taylorae

(e) Proteus Species: Have urease activity that raises urinary pH. Causes stone formation.
P. mirabilis

(f) Pseudomonas Species: Causes UTI primarily in patients with lowered host defences
P. aeruginosa

2. Fungi: Account for 40% of nosocomial UTI cases.

(a) Candida Species: may cause UTI in diabetic patients
C. albicans - causes vaginitis and chronic mucocutaneous candidiasis

As patient has a catheter, it is likely associated with contributing to UTI.
In this case, most incidences are caused by bacterial infection/colonization of the catheter, hence bacteria are suspected to be the cause.
Fungal stain may be used to rule out fungal infection in this case.

To narrow down type of bacteria, a gram stain should be done, to determine gram stain as well as morphology or bacteria:
Gram Stain:
Negative (pink): Escherichia coli (rod), Proteus Species (rod), Enterobacter Species (bacilli), Klebsiella Species (bacilli), Pseudomonas Species (bacilli).
Positive (purple): Enterococcus Species (cocci)

Gram negative strains are the most common causes; hence further biochemical test, urine cultures and antibiotic susceptibility must be employed to narrow down causative agent.

Most probable agents: according to incidence seen in UTI patients
Escherichia coli – 53-72%
Klebsiella pneumoniae – 6-12%
Proteus mirabilis – 4-6%
Enterococcus faecalis – 2-12%

References: > pointis > indwelling.html > medical > bladder_distension.htm > faculty > chamberlain > Website > lectures > lecture > uti.htm

Debra (TGo2)

MMIC - case 4

Name: Tong Wei Hong
Age: 68 yrs old
Sex: Male

Complaints: Fever, chills, excessive phlegm, breathing problems
Diagnosis: Bronchitis
Specimen: Sputum

Bronchitis is a respiratory disease in which the mucous membrane in the lungs' bronchial passages becomes inflamed. As the irritated membrane swells and grows thicker, it narrows or shuts off the tiny airways in the lungs, resulting in coughing spells accompanied by thick phlegm and breathlessness. There are two main types of bronchitis: acute and chronic.

- Several viruses cause bronchitis, including influenza A and B.

- A number of bacteria are known to cause bronchitis, such as Mycoplasma pneumoniae.

- Bronchitis also can occur when you inhale irritating fumes or dusts. Chemical solvents and smoke, including tobacco smoke, have been linked to acute bronchitis.

- People at increased risk both of getting bronchitis and of having more severe symptoms include the elderly, those with weakened immune systems, smokers, and anyone with repeated exposure to lung irritants.

Microorganisms commonly isolated from sputum include:
- Streptococcus pneumonia,
- Haemophilus influenzae
- Staphylococcus aureus
- Legionella pneumophila
- Mycoplasma pneumonia
- Klebsiella pneumoniae
- Pseudomonas aeruginosa
- Bordetella pertussis
- Escherichia coli

Cultures and tests are done on the sputum to help identify the bacteria that are causing an infection in the lungs or the airways (bronchi).
· Bacterial culture - gram stain or acid fast stain of the sputum done at the same time can help make the diagnosis.

· Fungal culture - The sputum sample is spread on special culture plates that will encourage the growth of mold and yeast. Different biochemical tests and stains are used to identify molds and yeast. Cultures for fungi may take several weeks.

· Viral culture - sputum is mixed with commercially-prepared animal cells in a test tube. Characteristic changes to the cells caused by the growing virus help identify the virus.

bronchitis>causes/">>bronchitis>causes>Home > Encyclopedia > Encyclopedia Index S


Sunday, 2 December 2007

dPBL MMIC:cheng hong (enterocolitis)

Name: Kuan Siew Yan / 29yrs/ Female

Diagnosis: Enterocolitis

What is Enterocolitis?
Cause by organisms that invade the epithelial and the subepithelial tissue of the small and large intestine thus causing inflammation and diarrhea.

Possible Causative Agents
1.Salmonella species
2.Shigella species
3.Campylobacter jejuni
4.Escherichia coli O157

1. Salmonella
§ Characteristics: Rigid, thick walled cell, Gram negative, Facultative straight rods

§ Species: S.typhi, Salmonella choleraeuis, Salmonella enteritidis

§ Pathogenesis: Manifest as Enterocolitis (penetrates through mucosal cells into the lamina propria; need at least 100000 organisms to cause infection). Typhoid/enteric fevers or Septicemia.

§ Biochemical features: Non lactose fermenting colonies on MacConkey/ EMB agar
: Alkaline slant/ acidic butt, gas and H2S in butt in TSI
: Lysine, Aginine, Ornithine positive
: indole neg, methyl red pos, Voges-Proskauer neg, citrate neg

2. Shigella species

§ Charateristics: Rigid, thick walled cell, Gram negative, Facultative straight rods, non-motile

Species: 4 different genus groups (A,B,C,D)

Pathogenesis: virulent (only need 100 organism to cause the disease), transmitted through fecal-oral route, normally have bloody diarrhea

Biochemical features: non-lactose fermenting, no gas production from fermenting glucose, no H2S production, alkaline slant and acidic butt on TSI, indole neg, methyl red pos, vogues proskauer neg, citrate neg, lysine arginine ornithine neg

Other features: a methylene blue stain will differentiate if it is an invasive or toxin producing bacteria (neutrophils will show that it is an invasive infection; like Shigella, Salmonella, Campylobacter

3. Campylobacter jejuni
§ Characteristics: Rigid, thick walled cell, Gram negative, Facultative curved rods (S shape)

§ Species: C. jejuni, C. intestinalis

§ Pathogenesis: Transmitted through fecal oral route or food/water contaminated with Campylobacter, causes entrocolitis

§ Biochemical features: culture on blood plate containing antibiotics (vancomycin, trimethoprim, cephalothin, polymyxin, amphotericin B) and incubate at 42oC at 5%oxygen and 10% carbon dioxide, no H2S production on TSI, hippurate hydrolysis test pos for C. jejuni, susceptible to nalidixic

§ Others: hanging drop shows motility of Campylobacter

4. Escherichia coli O157
Characteristics: Rigid, thick walled cell, Gram negative, Facultative straight rods

Pathogenesis: Enterohemorrhagic infection due to ingesting undercookedmeat, the organism cause the disease by using its features (pili, capsule, endotoxin, 3 exotoxins)-results in watery/ bloody diarrhea

Biochemical features: Can be either lactose fermenting/ non lactose fermenting, appears green on EBM agar, produces indole from tryptophan, decarboxylates lysine, motile, O157:H7 does not ferment sorbitol, acidic slant acidic butt produces gas abd no H2S production

References from:Review of Medical Microbiology and Immunology(9th edition). WARREN LEVINSON

:Color Atlas of Medical Bacteriology. ASM PRESS

Cheng Hong


Name: Khong Fay Seah
Age: 27 yrs old
Sex: Female

Complaints: Fever, chills, dysuria (painful urination)
Diagnosis: UTI
Specimen: Urine

Test ordered (Main Lab): Urine FEME (microscopy, dipstick)


Step 1:
A drop of uncentrifuged urine placed into a KOVA chamber, and examined with restricted light intensity under the high-dry objective of an ordinary clinical microscope can reveal leukocytes, epithelial cells, and bacteria.

Finding 105 organisms per millililiter can conclude UTI.
Normal hematocrit/HB
Elevated WBC (>18,000/uL)
Innumerable white blood cells
A few RBC
Numerous bacteria

Positive Urine dipstick nitirite suggest strongly bacteria growth in the urine tract.

Step 3:
A gram-stained smear of uncentrifuged urine that shows gram-negative rods is diagnostic of UTI

Step 4: in the micro lab

· Culture urine using a bacteriologic loop calibrated to deliver 0.01 or 0.00mL to agar plates.
· Aerobic culture ( maconkey and blood agar plates) & incubated overnight @ 37 degree.

In active pyelonephritis, the number of bacteria in urine collected by uretheral catheter is relatively lo.

While accumulating in the bladder, bacteria multiply rapidly and soon reach >105/mL (not contamination). The presence of more than 105/mL of the same type of bacteria per milliliter, establish a diagnosis of active infection. E.coli might be causing bacteria.

If the growth of different types of bacteria fewer than 104/mL, suggest that organisms come from normal flora or contaminants.

This female patient with acute dysuria and UTI will have 102/mL-103/mL.

If the cultures are negative but clinical signs of UTI are present, ureteral obstruction, tuberculosis can be considered.

E.coli cause 80-90% of acute bacterial lower tract infection in young women. The patient with acute cystitis has negative urine cultures for bacteria. Other possible causing bacteria: Staphylococcus, Neisseria Gonorrhoeae, Klebsiella, Enterobacter.

If such infection is caused by E.coli, identification and susceptibility testing of bacteria are not necessary.