Group Post: MMIC dPBL-1

Biochemical and Culture Testing

CASE 1

Possible Organisms	Biochemical Tests & Results	Culture agars
Escherichia coli	Gram Stain: - Oxidase: - Lactose: + Lysine: + TSI slant: + Indole: + Methyl red: + VP: - Citrate: -	5% Sheep Blood Agar Eosin-Methylthionine Blue Agar MacConkey Agar
Pseudomonas aeruginosa	Gram stain: - Oxidase: + Pyocyanin: + Fluoreseein: + Citrate: + Nitrase: + Lipase: + Ureases: +/- Indole: - TSI: -	Pseudomonas Agar P Pseudomonas Agar F MacConkey agar plate Eosin-Methylthionine Blue agar Cetrimide agar
Staphylococcus aureus	Gram Stain: + Catalase: + Coagulase: +	5% Sheep Blood Agar Mannitol Salt Agar
Staphylococcus saprophyicus	Gram Stain: + Catalase: + Coagulase: - Oxidase: - Phosphatase: - Urease: + Lipase: + 2-hour PYR broth hydrolysis: - D-trehalose; +	5% Sheep Blood Agar Mueller-Hinton Agar Lawn with Urine
Enterococcus faecalis	Gram Stain: + Catalase: -	5% Sheep Blood Agar Chocolate Agar 6.5% NaCl Concentration Agar (halotolerant) Bile Esculin Agar (BEA) Slants
Proteus-Providencia-Morganella e.g. Proteus mirabilis	Gram Stain: - Lactose: - Oxidase: - Urease: + Indole: - Nitrogen Reductase: + Methyl Red: + Vogues-Proskauer: - Catalase: + Phenylalanine Deaminase: +	MacConkey Agar CLED
Klebsiella-Enterobacter-Serratia e.g. Klebsiella oxytoca	Gram Stain: - Oxidase: - Lactose: - Urease: - Nitrate: + Indole: + Sucrose: + Dextrose: - Methyl Red: - Vogues-Proskauer: + Gelatin: - Amylase: + Lipase: - Catalase: + Citrate: + H2S: -	5% Sheep Blood Agar MacConkey Agar Trypticase Soy Agar
Klebsiella-Enterobacter-Serratia e.g. Klesiella pneumoniae	Indole: - Sucrose: + Dextrose: - Methyl Red: - Vogues-Proskauer: + H2S: - Lysine Decarboxylase: +	5% Sheep Blood Agar MacConkey Agar

Done by Lizzie

CASE 2

Campylobacter jejuni	Oxidase (+) Sensitivity to nalidixic acid	Blood agar with antibiotics to other fecal microbes Fail to grow at 25C, must grow at 42C Microaerophilic (grow at 5% oxygen + 10% carbon dioxide)
Shigella species	TSI (alkaline slant and acidic butt, with no gas and no H2S)	MacConkey's or EMB agar (colourless colonies) Anaerobe
Escherichia coli	TSI (acidic slant and acidic butt, with gas ,but no H2S) Indole test (+)	MacConkey's (pink colonies) or EMB agar (green sheen colonies) Anaerobe
Clostridium perfringens	ELISA to detect Clostridium perfringens Enterotoxin	TSC agar containing egg yolk Neomycin blood agar Strict anaerobe
Clostridium difficile	Cytotoxicity test on human cell-line ELISA to detect Clostridium perfringens exotoxin	Cycloserine cefoxitin fructose egg yolk agar (CCFA)
Vibrio Cholerae	Oxidase (+) TSI (acidic slant acidic deep; no gas) Agglutination of bacteria by polyvalent O1 antibody	TCBS (yellow colonies), Blood agar
Vibro parahemolyticus	Oxidase-positive Grow in 10% NaCl	TCBS (green colonies)
Bacillus cereus	Lecithinase (+) Motile (swarming on agar) Catalase (+) Indole (-) Nitrate (+) Urease (-) Citrate (+) MR (-) VP(+) TSI (Alkaline slant, acidic deep) Oxidase (-) Lactose fermenting; sucrose non-fermenting	Sheep’s blood Agar (beta hemolytic), large greenish colonies), Mannitol (grey colonies)
Yersinia enterocolitica	Motile at 25 deg MR (+) VP (-) Urease (-) Indole (-) Citrate (-) Catalase (+) TSI (acidic slant acidic deep; no gas)	MacConkey Agar (small lactose negative colonies) EMB
Staphylococci aureus	Catalase (+) Citrate (-) Coagulase (+) Indole (-)ME (+) Oxidase (-) Urease (-) VP (-) TSI (acidic slant acidic deep)	Sheep blood agar (beta hemolytic, yellow colonies) Mannitol salt agar (yellow colonies) 7.5% NaCl media

Done by Kent and Ye Tun

CASE 3

E. coli	Oxidase - Catalase + Indole + Methyl red + Voges-Prokauer - Citrate Utilization - Urease - TSI; H2S production Acid slant/ acid butt & gas produced; +	MacConkey agar Lactose fermenters; (red colonies) Blood agar β-hemolytic
Klebsiella sp (Klebsiella pneumuniae, Klebsiella oxytoca)	Oxidase - Catalase + Indole + Methyl red - Voges-Prokauer + Citrate Utilization + Urease - TSI; H2S production Alkaline slant/ acid butt; -	MacConkey agar Lactose fermenters; (pink colonies) Blood agar Mucoid colonies formed
Enterobacter (Enterobacter aerogenes)	Oxidase - Catalase + Indole - Methyl red - Voges-Prokauer + Citrate Utilization + Urease TSI; H2S production	MacConkey agar Blood agar
Serratia	Oxidase - Catalase + Indole Methyl red - Voges-Prokauer + Citrate Utilization + Urease TSI; H2S production	MacConkey agar Blood agar
Proteus mirabilis	Oxidase - Catalase + Indole + Methyl red + Voges-Prokauer - Citrate Utilization Urease + TSI; H2S production Alkaline slant/ acid butt; +	MacConkey agar Non-lactose fermenters; (white colonies) Blood agar Swarming observed
Morganella morganii	Oxidase - Catalase + Indole Methyl red Voges-Prokauer Citrate Utilization Urease + TSI; H2S production	MacConkey agar Non-lactose fermenters; (white colonies) Blood agar
Pseudomona aeruginosa	Oxidase + Catalase + Indole - Methyl red Voges-Prokauer Citrate Utilization + Urease Coagulase TSI; H2S production	MacConkey agar Non-lactose fermenters; (white colonies) Blood agar
Staphylococcus aureus	Catalase + Methyl red Voges-Prokauer Citrate Utilization Coagulase +	MacConkey agar - Blood agar β-hemolytic
Staphylococcus saprophyticus	Catalase + Methyl red Voges-Prokauer Citrate Utilization Urease + Coagulase -	MacConkey agar Non-lactose fermenters; (white colonies) Blood agar -
Enterococcus sp	Catalase Indole Methyl red Voges-Prokauer Citrate Utilization Urease Coagulase TSI; H2S production	MacConkey agar Blood agar

Done by Joan

CASE 4

Possible Organisms	Biochemical Tests & Results	Culture agars
Streptococcus pneumoniae	Gram Staining +ve Catalase -ve	2. 5% Sheep Blood agar with Optocin sensitivity Alpha-hemolysis with sensitivity to optocin
Klebsiella-Enterobacter-Serratia (Klebsiella sp.)	Gram Staining -ve Oxidase -ve Indole +ve/-ve MR-VP +/- or -/+ Citrate +ve H2S Production -ve Lysine Decarbocylase +ve	Lactose Fermentation (MacConkey agar) +ve
Bordetella pertussis	Gram Staining -ve Oxidase +ve Urease -ve Nitrase -ve Citrate -ve	Borde+ - Gengou agar BCYE agar

Done by Charmaine

CASE 5

Staphylococcus aureus	Gram Staining + Catalase + Coagulase +	Mannitol Salt Agar with 7-9% NaCl Mannitol fermentation Sheep Blood Agar Hemolysis of blood agar
Streptococcus pyogenes	Gram Staining + Catalase - PYR (Pyrrolidonearylamidase) Test +	Sheep Blood Agar Beta Hemolysis
Enterococci	Gram Staining + Bile- esculin test -	Sheep Blood Agar Gamma Hemolysis 6.5% NaCl Agar Can grow with 6.5% Nacl
Escherichia coli	Gram Staining - Indole + Methyl Red + Voges Prokauer - Citrate - TSI Acid slant and butt with gas production but no H2S in TSI	MacConkey Lactose fermentative (deep red colony) Eosin Methylene Blue( EMB) Green metallic sheen on EMB
Pseudomonas aeruginosa	Gram Staining - Oxidase + Catalase + Indole + Citrate + Urease - TSI - Pyocyanin - Fluroscein -	MAC Non- lactose fermentative (colourless colony on MAC) Pseudo F agar PositiveFluorescein pigment in Pseudo F agar

Done by Ai Tee

CASE 6

Gardnerella vaginalis	Catalase - Oxidase - Vaginal pH test pH of vaginal fluid >5.0 Whiff test Release of bad-smelling odor when discharge is mixed with KOH DNA Probe (Affirm™ VP III) High concentrations of G. vaginalis in VP III test	Chocolate agar Small, circular, convex, gray colonies HBT agar Colistin-oxolinic acid blood agar Beta-hemolysis
E. coli	Triple Sugar Iron (TSI) test TSI test show acid slant, acid butt with gas. Indole test + Methyl red test + Voges-Proskauer (VP) test - Citrate test -	EMB agar Black colonies with greenish-black metallic sheen on EMB agar MacConkey agar Deep red colonies on MacConkey agar (lactose fermentor) TSI agar
Pseudomonas aeruginosa	Oxidase test + Catalase test + Nitrase tese + Lipase test + TSI test Metallic sheen growth on the surface of TSI agar	EMB agar Mac Conkey’s agar (sterile specimen) Cetrimide agar (non-sterile specimen) Colonies appear flat, large, and oval. Secretes blue-green pigment (pyocyanin) on Cetrimide agar TSI agar No colour change on TSI medium (K/K/g-/H2S- profile)
Neisseria gonorrhoeae	Oxidase test + Direct immunofluorescence	Thayer-Martin chocolate agar Growth of spherical shape colonies on TM chocolate agar
Chlamydia Trachomatis	Direct immunofluorescence Presence of monoclonal antibodies (mAbs) to epitopes in the VS4 region of MOMP ELISA Presence of Chlamydia antigens in ELISA test PCR DNA-based tests.	Blood agar Mac Conkey’s agar
Trichomonas vaginalis	Vaginal pH test pH of vaginal fluid >4.5 PCR Enzyme immunoassay Presence of T. vaginalis antigen in enzyme immunoassay Direct Immunofluorescence.	Blood agar Mac Conkey’s agar
Mobiluncus	Catalase test - Indole test - Oxidase test -	Blood agar Clear, colourless colonies, around 2mm. Mac Conkey’s agar
Enterobacter-Klebsiella-Serrtia	-	Blood agar Mac Conkey’s agar Deep red colonies on MacConkey agar (lactose fermentor)
Proteus-Providencia-Morganella	Urease test + Indole test +	Blood agar Appear as swarming on blood agar. Mac Conkey agar Colourless colonies on MacConkey agar (Non-lactose fermentor)
Enterococcus faecalis	Catalase test -	Blood agar Alpha, beta, or non-hemolytic on blood agar. Mac Conkey agar
Candida albicans	-	Blood agar Sabouraud’s agar Presence of white, butyrous colonies.
Staphylococcus saprophyticus	Catalase test + Coagulase test - Urease test + Lipase test +	Blood agar Mac Conkey’s agar Spherical, irregular grape-like cluster in culture.

Done by Adrian

Group Post: MMIC dPBL 1

List of possible organisms for each case

	Diagnosis	Possible bacteria species	Morphology & Microscopy
Case 1	Urinary Tract Infection (UTI)	Staphylococcus aureus Staphylococcus saprophyticus Enterococcus faecalis	Gram Positive, Spherical-shaped, immobile and form grape-like clusters Gram Positive, Cocci-shaped occur singly and in pairs, short chains, and grape-like clusters Gram Positive, Cocci-shaped in pairs and chains
Case 2	Enterocolitis	Campylobacter jejuni Shigella species Salmonella species Escherichia coli (entero-pathogenic; EPEC) Clostridium perfringens Clostridium difficile Vibrio Cholerae Vibro parahemolyticus Bacillus cereus Yersinia enterocolitica Staphylococci aureus	Gram-negative rods Gram-negative rods Gram-negative rods Gram-negative rods Gram-positive rods Gram-positive rods Gram-negative rods Gram-negative rods Gram-positive rods Gram-negative rods Gram-positive cocci
Case 3	Urinary Tract Infection (UTI)	E. coli Klebsiella sp (Klebsiella pneumuniae, Klebsiella oxytoca) Enterobacter (Enterobacter aerogenes) Serratia Proteus mirabilis Morganella morganii Pseudomona aeruginosa Staphylococcus aureus Staphylococcus saprophyticus Enterococcus sp.	Gram negative bacilli Gram negative bacilli Gram negative bacilli Gram negative bacilli Gram negative bacilli Gram negative bacilli Gram negative bacilli Gram positive cocci Gram positive cocci Gram positive cocci
Case 4	Bronchitis	Strep. Pneumoniae Klebsiella-Enterobacter-Serratia (Klebsiella sp.) Bordetella pertussis	Gram positive, diplococci, lancet-shaped Gram negative rods Gram negative, coccibacilli
Case 5	Wound Infection	Beta Haemolytic Streptococci (Streptococcus pyogenes) Enterococci (Enterococcus faecalis) Staphylococci (Staphylococcus aureus/MRSA) Clostridium Pseudomonas aeruginosa Enterobacter species Escherichia coli Klebsiella species Proteus species Bacteroides	Gram-positive facultative anaerobes cocci Gram-positive facultative anaerobes cocci Gram-positive facultative anaerobes cocci Gram-positive anaerobe rod Gram-negative aerobic rods Gram-negative facultative rods Gram-negative facultative rods Gram-negative facultative rods Gram-negative facultative rods Gram negative Anaerobes rods
Case 6	Urinary Tract Infection (UTI)	Gardnerella vaginalis Chlamydia trachomatis Neisseria gonorrhoeae Ureaplasma urealyticum Staphylococcus saprophyticus Enterobacteriaceae (E. coli, serratia, klebsiella, enterobacter, citrobacter) Enterococci	Gram-negative rod with a gram-positive cell wall Gram-negative bacteria, coccoid or rod-shaped Gram negative cocci Gram-negative bacteria Gram positive, globular shaped, colonies resemble grape-like clusters Gram-negative, rod-shaped Gram-positive cocci which often occur in pairs (diplococci)

Authors: Lizzie, Kent & Ye Tun, Joan, Charmiane, Ai Tee and Adrian (according to case no.)

Brief description of diagnosis:

1. Urinary Tract Infection (UTI)

A urinary tract infection (UTI) is a bacterial infection that affects any part of the urinary tract. The urinary tract is made up of the kidneys, ureters, bladder, and urethra, and each plays a role in removing liquid waste from the body. Although urine contains a variety of fluids, salts, and waste products, it usually does not have bacteria in it. When bacteria get into the bladder or kidney and multiply in the urine, they cause a UTI.

Cystitis: bladder infection.

Pyelonephritis: kidney infection is much more serious.

Urethritis: urethra infection.

Each type of UTI may result in more specific signs and symptoms, depending on which part of your urinary tract is infected. The symptoms shown in acute pylonephritis are closest to what is given in the case study, whereby it is an infection of your kidneys may occur after spreading from an infection in your bladder. Kidney infection can cause upper back and flank pain, high fever, shaking chills, and nausea or vomiting.

Pyelonephritis is an inflammation of the kidney and upper urinary tract that usually results from noncontagious bacterial infection of the bladder (cystitis). In acute pylonephritis, bacteria begin colonising the tubules and connective tissue of the kidney itself. Small abscesses and streaks of pus begin to appear in the renal cortex and medulla respectively. Pyelonephritis most often occurs as a result of urinary tract infection, particularly in the presence of occasional or persistent backflow of urine from the bladder into the ureters or kidney pelvis (vesicoureteric reflux).

Urinary catheterization is the insertion of a catheter into a patient's bladder through the urethra, to drain urine from the bladder into an attached bag or container. Indwelling catheters should be restricted to patients whose incontinence is caused by urinary tract obstruction that can not be treated, and for which alternative therapy is not feasible. However, the catheter may introduce bacteria into the urethra and bladder, resulting in urinary tract
infection. The risk for developing a UTI increases when long-term catheterization is required.

Bacteria causing UTI may be isolated from the vaginal discharge. UTI can also result from sexual intercourse where bacteria from the vaginal area is transferred to the urethra and bladder.

Authors: Lizzie, Joan and Adrian


2. Enterocolitis

Enterocollitis is the inflammation of the small intestine and colon. Disease-causing bacteria usually invade the small intestines and colon and cause inflammation (blood or pus in the stool, fever) and abdominal pain and diarrhea. Campylobacter jejuni is the most common bacterium that causes acute enterocolitis in the U.S. Other bacteria that cause enterocolitis include Shigella, Salmonella and EPEC (E. Coli Enteropathogenic). These bacteria usually are acquired by drinking contaminated water or eating contaminated foods such as vegetables, poultry, and dairy products.

Enterocolitis caused by the bacterium Clostridium difficile is unusual because it often is caused by antibiotic treatment. It is also the most common nosocomial infection (infection acquired while in the hospital) to cause diarrhea.

E. coli O157:H7 produces a toxin that causes hemorrhagic enterocolitis (enterocolitis with bleeding). There was a famous outbreak of hemorrhagic enterocolitis in the U. S. traced to contaminated ground beef in hamburgers (hence it is also called hamburger colitis).

Bacterial overgrowth of the small intestine.

Because of small intestinal problems, normal colonic bacteria may spread from the colon and into the small intestine. When they do, they are in a position to digest food that the small intestine has not had time to digest and absorb. The mechanism that leads to the development of diarrhea in bacterial overgrowth is not known.

Authors: Kent and Ye Tun

3. Bronchitis

Bronchitis is an inflammation of the bronchi of the lungs. There are two main types of bronchitis: acute and chronic bronchitis.

Acute bronchitis can be caused by viruses such as influenza A and B, parainfluenza virus, respiratory syncytical virus, coronavirus, adenovirus, and rhinovirus. Bacterias are also able to cause acute bronchitis. Air pollutants such as cigarette smoke, dusts, or fumes of chemicals are also able to cause acute bronchitis.

The symptoms of acute bronchitis include:

* Sore throat

* Chest congestion

* Sinus fullness

* Breathlessness

* Wheezing

* Slight fever and chills

* Overall malaise

* Cough

Chronic bronchitis is where by the signs and symptoms become prolonged, and is defined clinically as a
persistent cough that produces sputum (phlegm) and mucus, for at least three months in two consecutive years. Chronic bronchitis is part of chronic obstructive pulmonary disease (COPD).

Author: Charmaine

4. Surgical Site Infection (SSI)

There are different levels of SSI:

• Superficial incisional, affecting the skin and subcutaneous tissue.

• Deep incisional, affecting the facial and muscle layers.

• Organ or space infection affecting any part of the anatomy opened or manipulated during the operation.

Symptoms: Pain, fever, inflammation, swelling and pus formation



Causes of infection:

-Microbes contamination (most commonly by S. aureus, S. pyogenes or P. aeruginosa

-Migration of patients’ own bacterial normal flora

-The materials or equipment used in the operational procedures (for e.g.: poor surgical techniques)

Author: Ai Tee