Medical Microbiology-dPBL Package 2

There are outbreaks of viral, fungal and protozoa diseases among platoons of army soldiers in Indonesia. Soldiers reported sick after 2 weeks of jungle warfare training. It is of concern to the ministry that there are also sporadic reports of avian flu in the nearby villages. In view of these outbreaks, you have been tasked to conduct a pre-mission briefing with blogs and poster to educate future batches of soldiers.

‘Army soldiers’: Army soldiers are not able to bathe frequently. They are also always perspiring due to the constant training, therefore they are always damp and dirty.

‘Jungle training’: This implies that the environment that the soldiers are in is humid, wet and warm. The army soldiers are also exposed to all sorts of insects and animals. In addition, there is also soil around the place where the soldiers are training. Therefore, there could be any possible vectors to the various infections.

Various protozoa diseases are listed down in the table below:
Table 1: Protozoa

Reasons for identifying these protozoa:

Plasmodium falciparum/vivax: These protozoa is carried by the vector, female anopheles mosquitoes that can be found in tropical areas. Soldiers training in the jungle may be bitten by these female mosquitoes, thereby releasing the protozoa into the bloodstream of the soldiers where they multiple and cause malaria.

Toxoplasma gondii: Soldiers are prone to eating meat that is not cooked thoroughly due to the lack of proper facilities. In addition, soldiers might ingest contaminated water by the river etc. Contamination is due to the presence of of infected cat faeces by the protozoan. Toxoplasmosis can be transmitted to the soldiers due to frequent visits to the neighbouring villages where cats etc may be present.

Leishmania: This protozoa is spread by the bite of sandflies. Sandflies are found in sandy area and sandy areas are a common sight in jungles. As such, soldiers might be bitten by this vector and thus contract leishmaniasis.

Giardia lamblia: This is usually found in soil, food, water, or surfaces that have been contaminated with the feces from infected humans or animals. With unsanitary conditions and lack of proper facilities in the jungle, food might not be properly cooked and this could cause the transmission of Giardia lamblia. In addition, outbreaks among military personnel could also be caused by various infected animal such as birds, dogs and cats that can be found in the neighbouring area.

Entamoeba histolytica: This is found in many tropical countries. The presence of Entamoeba histolytica is due to the unsanitary conditions. In jungle warfare, the condition that the soldiers are in is generally unhygenic, thereby promoting the growth of this protozoan and the transmission of disease such as amoebic dysentery. Due to the unhygeneic conditions, the spread of this protozan is sanitary conditions in the jungle, amoebic dysentery is most commonly spread by water or contaminated, uncooked food or from carriers.

Trypanosoma brucei: This flagellated protozoan enters the blood-stream via the bite of bloodsucking male and female tse-tse. However the parasites are found mainly in Africa therefore it is excluded from being a potential pathogen that could be found in the jungles of Indonesia.

Cyclospora cayentanesis: This protozoa is common in tropical countries as the warm and moist environment found in such country is required for the protozoa (oocytes) sporulate into their infective forms. Thereby infecting the soldiers via contaminated water or food.

Cryptosporidium parvum: This protoza contaminate water supply and due to the lack of proper supplies, soldiers undergoing training may also ingest food that is contaminated. With the close interaction of the soldiers training together, infected soldiers are thus able to transmit the disease cryptosporidiosis to the healthy soldiers.

Various fungal diseases are listed down in the table below:

Table 2: Fungi

Reasons for identifying these fungi:

Ring Worm: Dermatophytes feeds on keratin which is the material found on the outer layer of the skin, hair and nails. These fungi thrive best on moist and hot skin that is hidden from the light. Soldiers undergoing training tends to perspire alot and due to the lack of constant and proper wash-up, the condition of the skin encourage the growth of these fungi. This fungus can exist anyway on the body.

Trichophyton rubrum: Soldiers can experience Athlete's foot (Tenea pedis) as they tend to put on their shoes for very long period of times. These fungi involved attack the feet due to the encouraged growth in the presence of a warm, dark and humid environment. In addition, spreading of fungi can take plac if the feet are not washed adequately with soap and water.

Candida Albicans: Such fungus easily "invade" the body through cuts. Soldiers are prone to injuries such as cuts or abrasion while having training in the jungle, thus increasing their exposure to Candida Albicans.

Cryptococcus neoformans: Cryptococcus neoformans var gattii grows in tropical area in the litter around certain eucalyptus trees. This fungus is airborne and the can be breathed in by the soldiers undergoing training in the forest.

Aspergillus fumigatus: Aspergillus fumigatus is a type of fungus found in soil. During training, the soldiers might have to do crawling on the ground, having close contact with the soil. inhalation of the fungi is therefore made possible, resulting in Allergic bronchopulmonary aspergillosis.

Histoplasma Capsulatum: Histoplasma capsulatum is a soil-borne, dimorphic fungus that causes histoplasmosis in human. It is found throughout the world but is most prevalent in countries favoring a warm, moist, and humid climate.

Various viral diseases are listed down in the table below:

Table 3: Virus

Reasons for identifying these virus:

Rabies virus: This virus could infect soldiers through other infected animals such as bats and monkeys. These two animals are commonly found in the jungles of Indonesia. The virus could be spread through the animal bites or due to aerosols from mucous membranes.

Japanese Encephalitis: The virus is transmitted through a mosquito vector known as Culex tritaeniorhynchus. Soldiers in the jungle could acquire the virus from mosquito bites. Disease from the virus is most prevalent in South East Asia.

Hepatitis A virus: Unsanitary conditions easily allow contamination of food and water. Due to the poor hygiene of the soldiers, the virus could be present in dirty utensils that are washed in the contaminated lakes or rivers. HAV is transmitted through the fecal oral route and thus it can be transferred from unwashed hands after the soldiers visited the toilets. In addition, during the training, soldiers will have to be in close contact with one another, therefore infected soldiers can in turn infect other healthy soldiers.

Ross Fever virus: This disease is carried and transmitted by the Southern Saltmarsh mosquito or Aedes camptorhyncus. As most other diseases mentioned, mosquitoes thrive in damp places such as the jungle. This then allows possible infection when an infected mosquito bites a soldier.

Human Immunodeficiency Virus (HIV) : HIV infection is is caused by the transmission of virus through sexual contact or through blood or blood product route. The soldiers could have gone to the village and had sexual intercourse with the female villagers. Also, medical equipments such as syringe might be shared among them, thereby transmitting the virus to each other.

Various viral fever are listed down in the table below:

Table 4: Virus (Fever)

Reasons for identifying these virus:

Yellow fever virus: As soldiers have their trainings in the Indonesia's forests, getting mosquitoes' attacks are inevitable. Aedes aegypti are found in forests. It is caused by Flaviviridae, a positive single-stranded RNA virus whereby soldiers get infected after deposition of viral particles through the skin in infected arthropod saliva(bite).

Dengue fever virus: Dengue, which is caused by the Aedes aegypti mosquito are commonly found in tropical climates e.g. Indonesia thus soldiers have a higher tendency of contracting dengue fever especially during the day as growth of those mosquitoes are enhanced in the presence of a warm and humid environment e.g. Indonesia.

Rift valley virus: As the soldiers have their trainings in the forest, they are bound to have direct or indirect contacts with infected wild animals or consumed contaminated food e.g. chicken. RVF is a viral zoonosis causing fever. It also can be caused by infected mosquitoes. RVF is able to affect primarily domestic livestock and passes down to humans.

Chikungunya virus: Similarly to the cause of Dengue Fever, soldiers are able to get infected with Chikungunya from mosquito bites such as the Aedes and Culex. It is a viral fever caused by an alphavirus, whereby the mosquitoes are commonly found in warm and humid climates e.g. Indonesia.

Avian flu is listed down in the table below:

Table 5: Virus (Avian Flu)

Reason for identifying this virus:

Avian influenza virus: This virus is found in tropical countries like Indonesia where it is warm and humid. Soldiers, during their free time will have the chance to visit the nearby village where they will be exposed to birds such as chickens. Consuming of infected chickens, thus increase the chance of the soldiers contracting this disease. In addition, the recent outbreak of this disease in the village has further increase the chance of the soldiers contracting this disease. In the jungle, the soldiers are also exposed to various kinds of birds species that could be infected by the virus as well.

References

Fungal Diseases
http://canadiancpd.medscape.com>viewarticle>503661
www.ops-oms.org>>English>AD>DPC>CD>atlanta_july2000.doc
http://cancerweb.ncl.ac.uk>cgi-bin>omd?epidemic+polyarthritis
http://dermnetnz.org>fungal>cryptococcosis.html
http://en.wikipedia.org>wiki>Histoplasmosis http://www.ahc.sa.gov.au>site>page.cfm?u=608
http://www.botany.utoronto.ca>courses>bot405>notes>Lecture%2010.pdf
http://www.candidasupport.org>
http://www.cyh.com>HealthTopics>HealthTopicDetails.aspx?p=114&np=303&id=1907
http://www.dermnetnz.org>fungal>tinea-pedis.html
http://www.faetc.org>PDF>Primary_Care_Guide>Chapter_19-Fungal_Infections.pdf http://www.healthscout.com>ency>68>312>main.html
http://www.histopathology-india.net>CHIKV.htm
http://www.mycology.adelaide.edu.au>Mycoses>Subcutaneous>Lobomycosis>index.html
http://www.nlm.nih.gov>medlineplus>ency>article>000070.htm
http://www.phac-aspc.gc.ca>msds-ftss>msds11e.html
http://www.who.int>mediacentre>factsheets>fs207>en>

Protozoa Diseases
http://deploymenthealthlibrary.fhp.osd.mil>products>Staying%20Healthy%20Guide-%20Soldiers%20Guide%20in%20Indonesia%20and%20Malaysia%20(Tri-fold)%20(125).pdf
http://en.wikipedia.org>wiki>Giardia
http://en.wikipedia.org>wiki>Intestinal_parasite
http://en.wikipedia.org>wiki>Leishmania
http://en.wikipedia.org>wiki>Malaria
http://focosi.altervista.org>pathoprotozoa.htm
http://infectiouspeople.blogspot.com>2007>01>protozoal-infections.html
http://rds.yahoo.com>_ylt=A0oGkm3iYZlH5SsAV8lXNyoA;_ylu=X3oDMTFhNWE2YThkBHNlYwNzcgRwb3MDNQRjb2xvA3NrMQR2dGlkA01BUDAwM185NARsA1dTMQ-->SIG=1214b237g>EXP=1201320802>**http%3a>>iai.asm.org>cgi>content>full>69>9>5940
http://www.cdc.gov>ncidod>dpd>parasites>cyclospora>factsht_cyclospora.htm#symptoms
http://www.cdfound.to.it>HTML>khan.htm#Current%20Prevention
http://www.ndsu.nodak.edu>instruct>brewer>brewer>entomology>topics>disease.htm

Viral Diseases
http://attra.ncat.org>attra-pub>soilborne.html
http://en.wikipedia.org>Ascariasis
http://en.wikipedia.org>wiki>HIV
http://en.wikipedia.org>Tetanus
http://www.cdc.gov>flu>avian>gen-info>facts.htm
http://www.mayoclinic.com>health>bird-flu>DS00566>DSECTION=8
http://www.mayoclinic.com>health>hiv-aids>DS00005>DSECTION=8
http://www.medicinenet.com>bird_flu>article.htm
http://www.medicinenet.com>bird_flu>page5.htm
http://www.metrokc.gov>health>prevcont>yellow.htm#schedule
http://www.umm.edu>patiented>articles>what_causes_encephalitis_000096_2.htm
http://www.who.int>immunization>topics>rabies>en>index.html

http://www.wordtravels.com>Travelguide>Countries>Indonesia>Health

Medical Microbiology- dPBL- Case 1 to 6- Microorganisms Identifications

Case 1 (Suat Fang, 0503328G)

Particulars of patient

Name: Khong Fay Seah Sex: Female
HRN: OPD 009 IC No. : S00055X
Date of birth : 1/12/80 Age : 27 years
Ward/Clinic : Clinic M Bed No : -------

Clinical diagnosis

Complains: Fever, chills and dysuria
Diagnosis: Urinary Tract Infection
Specimen: Urine

Suspected Microorganisms Identifications



References

http://en.wikipedia.org>wiki>Enterococcus
http://en.wikipedia.org>wiki>Escherichia_coli
http://en.wikipedia.org>wiki>Klebsiella_pneumoniae
http://en.wikipedia.org>wiki>Proteus_mirabilis
http://en.wikipedia.org>wiki>Pseudomonas_aeruginosa
http://en.wikipedia.org>wiki>Staphylococcus_saprophyticus
http://www.austincc.edu>microbugz>html http://www.bact.wisc.edu>themicrobialworld>E.coli.html
http://www.bd.com>ds>technicalCenter>inserts>L007359(08)(1206)
http://www.umdnj.edu>micrsweb>case2gramnegatives>intro.html

Case 2 (Sharifah, 0503189C)
Particulars of patient

Name : Kwan Siew Yan Sex : Female
HRN : OPD001 IC No. : S000123X
Date of birth : 16/6/78 Age : 23 years
Ward/Clinic : Clinic X Bed No : -------

Clinical diagnosis

Complaints: Diarrhea
Diagnosis: Enterocolitis
Specimen: Stool

Suspected Microorganisms Identifications

References
http://en.wikipedia.org>wiki> Salmonella
http://en.wikipedia.org>wiki> Escherichia_coli
http://en.wikipedia.org>wiki> Campylobacter
http://en.wikipedia.org>wiki> Vibrio
http://en.wikipedia.org>wiki> Yersinia
Levinson, W. (2004). Review of Medical Microbiology and Immunology. 9th edition. McGraw-Hill.

Case 3 (Royston, 0503289A)
Particulars of patient

Name : Maisy Hong Sex : Female
HRN : 006789T IC No. : S000111Y
Date of birth : 1/12/40 Age : 67 years
Ward/Clinic : AB2 Bed No : -------
(Patient is in-patient, thus more susceptible to nosocomial infection)

Clinical diagnosis

Complaints: Fever, chills, bladder distension (stretching); on indwelling catheter
Diagnosis: Urinary Tract Infection
Specimen: Urine

Suspected Microorganisms Identifications

References

http://www.wikipedia.org>agar_plates
http://www.wikipedia.org>Enterococcus
http://www.wikipedia.org>Escherichia_coli
http://www.wikipedia.org>Klebsiella
http://www.wikipedia.org>Proteus
http://www.wikipedia.org> Pseudomonas_aeruginosa
http://www.wikipedia.org>Staphylococcus_saprophyticus

Case 4 (Najib, 0503217B), (Charmaine, 0503186I)

Particulars of patient

Name : Tong Wei Hong Sex : Male
HRN : OPD 004 IC No. : S000444X
Date of birth : 1/12/39 Age : 68 years
Ward/Clinic : Clinic M Bed No : -------

Clinical diagnosis

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

Suspected Microorganisms Identifications

References

http://www.blinn.edu>natscience>phillips>Micro%20Pictures.htm
http://www.hpa-standardmethods.org.uk>documents>bsopid>pdf>bsopid11.pdf http://www.mc.maricopa.edu~johnson>labtools>Dbiochem>opto.html
http://www.pubmecentral.nih.gov>articlerender.fcgi?artid=379757
http://www.sigmaaldrich.com>img>assets>13860>75744.pdf

Case 5 (Jeremy, 0503168G)
Particulars of patient

Name : Wong Fei Hong Sex : Male
HRN : OPD 0010 IC No. : S210444X
Date of birth : 1/12/70 Age : 37 years
Ward/Clinic : Clinic S Bed No : -------

Clinical diagnosis

Complaints: Fever, swelling around operation wound
Diagnosis: Wound infection
Specimen: Wound swab

Suspected Microorganisms Identifications

References
Warren Levinson. Review of medical microbiology and immunology (9^thed).
http://users.stlcc.edu/kkiser/biochem.html
http://www.oxoid.com

Case 6 (Natalie, 0503275J)
Particulars of patient

Name : Ong Fei Fei Sex : Female
HRN : OPD 0013 IC No. : S210334X
Date of birth: 1/12/70 Age: 37 years
Ward/Clinic: Clinic T Bed No: -------

Clinical diagnosis

Complaints: Fever, pain during urination, virginal discharge
Diagnosis: UTI
Specimen: Virginal Discharge

Suspected Microorganisms Identifications

Escherichia coli (E.coli)

Test: Gram-Stain
Biochemical Tests: Lactose fermentation test, Indole test, Lysine test
Treatment: Antibiotics which may be used to treat E. coli infection include (but are not limited to) amoxicillin as well as other semi-synthetic penicillins, many cephalosporins, carbapenems, aztreonam, trimethoprim-sulfamethoxazole, ciprofloxacin, nitrofurantoin and the aminoglycosides

Chamydia

Test: Gram-Stain/DNA-based test
Identification: Chlamydia species are readily identified and distinguished from other chlamydial species using DNA-based tests. Most strains of C. trachomatis are recognized by monoclonal antibodies (mAbs) to epitopes in the VS4 region of MOMP
Treatment: It may be treated with any of several antibiotics such as azithromycin, erythromycin or doxycycline/tetracycline.

Klebsiella pneumoniae

Test: Gram Stain
Biochemical Tests: Indole test, citrate test, urease test, motility test, malonate test, Phenylalanine slant test
Treatment: carbenicillin, ampicillin, quinolones, and ceftazidime

Pseudomonas aeruginosa

Test: Gram Stain
Biochemical Tests: Triple Ion Sugar (TSI), Oxidase test, Indole test, Citrate test
Treatment: Aminoglycosides, Quinolones, Cephalosporins, Ureidopenicillins

Followings are the additional microorganisms of other causative agents of UTI:

Candida albicans (yeast)
Candida albicans is a diploid fungus (a form of yeast), which is capable of mating but not of meiosis, and a causal agent of opportunistic oral and genital infections in humans. C. albicans is among the gut flora, the many organisms which live in the human mouth and gastrointestinal tract. Under normal circumstances, C. albicans lives in 80% of the human population with no harmful effects, although overgrowth results in candidiasis.

Test: Calcofluor- white Gram Stain followed by direct microscopy, Culturing
Treatment: Amphotericin B, Ketoconazole

Trichomonas vaginalis (protozoan)
Trichomonas vaginalis, an anaerobic, parasitic flagellated protozoan, is the causative agent of trichomoniasis, and is the most common pathogenic protozoan infection of humans in industrialized countries.

Test: Pap smear, Culturing
Treatment: Metronidazole/ Tinidazole

Gardnerella vaginalis
Gardnerella is a genus of gram-variable bacteria of which Gardnerella vaginalis is the only species. Gardnerella vaginalis can cause bacterial vaginosis in some women.

G. vaginalis is an aerobic, non-motile, slow growing coccobacillus. It grows as small, circular, convex, gray colonies on chocolate agar; it will also grow on HBT agar. A selective medium for G. vaginalis is colistin-oxolinic acid blood agar

Test: Microscopy of Clue cells, Amine test
Treatment: metronidazole

Illustrations for the types of Biochemical tests used and their results

1. Citrate Utilisation Test: The citrate tube is used to determine if an organism is capable of utilizing citrate.

2. Urease test: The urea agar slant allows detection of urease activity of both rapidly urease pos organisms as well as enterobacteriaceae family.

3. Motility Test: Used to detect the motility of organisms in a semi-solid gelatin medium.

4. Indole Test: Can be detected by its ability to combine with certain aldehyde to form a coloured compound.

5. Phenylalanine Slant Test: Determine the ability of an organism to deaminate phenylalanine to phenylpyruvic acid enzymatically with resulting acidity.

6. Malonate Broth Test: Determine the ability of an organism to use sodium molante as the sole carbon with resulting alkanity and it is used to differentiate enterobact family.

Triple Sugar Iron (TSI) Test: Used to differentiate enterics based on the ability to reduce sulfur and ferment carbohydrates.

All pictures extracted from: http://users.stlcc.edu/ >kkiser>biochem

Laboratory Investigations (Bacteria Identification)

The specimen (virginal discharge) will be cultured so as to isolate the microorganisms short listed above. Next, a series of biochemical tests will be carried out and incubated for a day before verifying the results for the final identification of the specific microorganism. Lastly, antibiotic susceptibility testing is performed to achieve a cure for the infection.

1) Gram-staining followed by microscopy examination (of unknown bacteria)
i.e. Gram Negative: E. Coli, P. Aeruginosa, and Enterobacter-Klebsiella-Serrtia Family

2) Culturing of bacteria
MacConkey’s Agar: To check for Lactose Fermenters
Positive: E.Coli and Enterobacter-Klebsiella-Serrtia Family
Negative: P. Aeruginosa
Nutrient Agar: To observe for P. Aeruginosa
Eosin- methylene blue Agar: E. Coli

3) For gram-neg bacteria, biochemical tests are carried out (i.e. different tests + F12). F12 is of antimicrobial agents such as penicillin, it tells us the sensitiveness and susceptibility of the bacteria towards different types of antibodies.

Example of Biochemical Tests:

1. Simmons citrate


2. Urease


3. Motility (& OF) test


4. Indole


5. Phenylalanine slant


6. Malonate test

References

http://en.wikipedia.org>wiki>Escherichia_coli
http://en.wikipedia.org>wiki>Chlamydia_trachomatis
http://en.wikipedia.org>wiki>Pseudomonas_aeruginosa
http://en.wikipedia.org>wiki>Klebsiella_pneumoniae
http://en.wikipedia.org>wiki>Trichomonas_vaginalis
http://en.wikipedia.org>wiki>Gardnerella
http://en.wikipedia.org>wiki>Candida_albicans
http://kidney.niddk.nih.gov>Kudiseases>pubs>utiadult/
http://whitewolf.newcastle.edu.au>techinfo>proc_bacto_biochem
http://www.healthscout.com>Gardnerella Vaginalis>Symptoms

Medical Microbiology- dPBL- Case 1 to 6

Case 1 (Suat Fang, 0503328G)
Particulars of patient

Name : Khong Fay Seah Sex : Female
HRN : OPD 009 IC No. : S00055X
Date of birth : 1/12/80 Age : 27 years
Ward/Clinic : Clinic M Bed No : -------

Clinical diagnosis

Complains: Fever, chills and dysuria
Diagnosis: Urinary Tract Infection
Specimen: Urine

Introduction

The lower urinary tract, which contains the bladder and urethra, and the upper urinary tract, that contains two kidneys and the ureters, makes up the 2 sections of urinary tract.

UTI is the infection of one or more components of the urinary tract due to bacteria that enter the opening of the urethra. Urine does not normally contain microorganisms. When bacteria get into the bladder or kidney and multiply in the urine, they cause a UTI. UTI is more common in women because their urethra is shorter and closer to the anus.

Suspected microorganisms

o Escherichia coli (E.coli),
o Klebsiella pneumoniae,
o Proteus mirabilis
o Pseudomonas aeruginosa
o Staphylococcus saprophyticus and
o Enterococcus spp.

Escherichia coli (E.coli)
· gram-negative bacterium
· found in the digestive tract
· Present on the skin around the rectal area
· typically ferment lactose
· grows well on MacConkey agar
· Most common cause of UTI.

Stain: gram staining
Biochemical Test: Oxidase test, MRVP

Klebsiella pneumonia
· gram-negative
· non-motile
· lactose fermenting
· facultative anaerobic
· rod shaped bacterium
· found in the normal flora of the mouth, skin, and intestines
· Second common cause of UTI.

Stain: gram staining
Biochemical Test: Oxidase test, Indole-Test

Proteus mirabilis
· gram-negative
· facultative anaerobic bacterium
· shows swarming, motility, and urease activity
· rod shaped bacterium
· Has the ability to produce high levels of urease. Urease hydrolyzes urea to ammonia (NH3) and thus makes the urine more alkaline.

Stain: gram staining
Biochemical Test: Oxidase test, indole test

Pseudomonas aeruginosa
· gram-negative
· motile
· aerobic rod shape bacteria
· oxidase-positive
· do not ferment lactose
· common inhabitants of soil and water
· tend to cause disease in humans with abnormal host defenses.

Stain: gram staining
Biochemical Test: Triple Ion Sugar (TSI)

Staphylococcus saprophyticus
· gram-postive
· facultative anaerobes
· coagulase-negative species of Staphylococcus bacteria
· catalase-positive
· reside in the urinary tract and bladder of sexually active females.
· phosphatase-negative
· urease and lipase positive.

Stain: gram staining
Biochemical Test: Catalase test, coagulase test

Enterococcus faecalis
· gram-positive Streptococci
· spherical bacterium which forms pairs or chains during growth

Stain: gram staining
Biochemical Test: Catalase test, PYRase activity test

1. The urine sample will be cultured on blood agar and Cystine-Lactose-Electrolyte Deficient (CLED) agar and incubated.
2. Gram-staining will then be done to differentiate between gram-positive and gram-negative microorganisms.
3. If the gram-stain showed gram-positive cocci, the catalase test can be done to differentiate between Streptococcus and staphylococcus bacterium.
4. Coagulase test will be performed if catalase test shows positive results to determine the type of Staphylococcus bacterium.
5. PYRase activity test can be done for negative results catalase test to determine the type of Streptococcus bacterium.

References

http://en.wikipedia.org>wiki>Enterococcus
http://en.wikipedia.org>wiki>Escherichia_coli
http://en.wikipedia.org>wiki>Klebsiella_pneumoniae
http://en.wikipedia.org>wiki>Proteus_mirabilis
http://en.wikipedia.org>wiki>Pseudomonas_aeruginosa
http://en.wikipedia.org>wiki>Staphylococcus_saprophyticus
http://kidney.niddk.nih.gov>Kudiseases>pubs>utiadult>
http://www.healthassist.net>conditions>uti.shtml
http://www.medicinenet.com>urine_infection>article.htm

Case 2 (Sharifah, 0503189C)
Particulars of patient

Name : Kwan Siew Yan Sex : Female
HRN : OPD001 IC No. : S000123X
Date of birth : 16/6/78 Age : 23 years
Ward/Clinic : Clinic X Bed No : -------

Clinical diagnosis

Complaints: Diarrhea
Diagnosis: Enterocolitis
Specimen: Stool

Introduction

- Definition
‘Enterocolitis’ is the combination of two words ‘Enteritis’ which is the inflammation of the small intestine and ‘Colitis’ which is the inflammation of the large intestine, specifically, the colon.

- Characteristics
It is characterized by the inflammation of the epithelial and subepithelial tissue at the small and large intestines. Symptoms include presence of blood in feces, abdominal pain and diarrhea (as complained by this patient).

Suspected Micro-organisms

- Salmonella
- Shigella
- Campylobacter
- Escherichia
- Vibro
- Yersinia

Salmonella
- Gram negative rod-shaped bacterium
- Motile
- Produces Hydrogen Sulphide
- Non- lactose fermenter
- Has a high infectious dose (must have a high dose to cause infection)
- Is an invasive organism
- Typical species that causes enterocolitis (however, other species are known to have causes said disease as well) : Salmonella typhimurium

Shigella
- Gram negative rod-shaped bacterium
- Non-motile
- Do not produce Hydrogen Sulphide
- Non- lactose fermenter
- Has a low infectiouse dose
- Is an invasive organism
- Usually involved in bacillary dysentery
- Typical species that causes enterocolitis : Shigella dysenteriae, Shigella sonnei

Campylobacter
- Gram negative rod, comma or S-shaped bacterium
- Motile
- Microaerophilic (grows best in 5% Oxygen)
- Usually involved in bacillary dysentery
- Typical species that causes enterocolitis : Campylobacter jejuni

Escherichia
- Gram negative rod, comma-shaped bacterium
- Motile
- Do not produce Hydrogen Sulphide
- Lactose fermenter
- Is an invasive organism
- Typical species that causes diarrhea : Enteropathogenic Escherichia coli (EPEC), Enterotoxigenic Escherichia coli (ETEC)

Vibrio
- Gram negative rod, comma-shaped bacterium
- Motile
- Is a slow lactose fermenter
- Found in marine organisms
- Halophiles
- Is not an invasive organism
- Typical species that causes diarrhea : Vibrio cholerae, Vibrio parahaemolyticus

Yersinia
- Gram negative rod-shaped bacterium
- Motile
- Non- lactose fermenter
- Is not an invasive organism
- Transmitted by fecal contaminations by domestic animals
- Typical species that causes enterocolitis : Yersinia enterocolitica

Types of Preliminary Tests to be Performed

- Microscopy (Gram Stain)
- Leukocyte Count
- Stool Occult Blood
- Culture
o MacConkey
o Blood
o XLD or DCA
o TCBS
- Enrichment Broth
o Selenite

Types of Secondary Tests to be Performed

- TSI Slant
- Oxidase Test
- Indole Test
- Fermentation of Sugars (e.g. Glucose, Lactose, Mannitol etc.)

References

Levinson, W. (2004). Review of Medical Microbiology and Immunology. 9th edition. McGraw-Hill.
http://en.wikipedia.org>wiki>Salmonella
http://en.wikipedia.org>wiki>Campylobacter
http://en.wikipedia.org>wiki>Escherichia_coli
http://en.wikipedia.org>wiki>Vibrio
http://en.wikipedia.org>wiki>Yersinia

Case 3 (Royston, 0503289A)
Particulars of patient

Name : Maisy Hong Sex : Female
HRN : 006789T IC No. : S000111Y
Date of birth : 1/12/40 Age : 67 years
Ward/Clinic : AB2 Bed No : -------
(Patient is in-patient, thus more susceptible to nosocomial infection)

Clinical diagnosis

Complaints: Fever, chills, bladder distension (stretching); on indwelling catheter
Diagnosis: Urinary Tract Infection
Specimen: Urine

Introduction

Given the symptoms from the case, the UTI is most like to be upper urinary tract infections like pyelonephritis, which is an ascending urinary tract infection that has reached the pelvis of the kidney.

Background Info: Catheter-associated UTI
Causes
A catheter is a hollow tube that is used to drain urine from the bladder. An indwelling catheter stays in place for long periods of time. The presence of a catheter within the urinary tract increases the likelihood of urinary tract infection. As the urinary catheter is left in place for long periods of time, bacteria will inevitably grow in it. A harmful infection may occur if the number of bacteria becomes large or if specific pathologic bacteria grow in the urinary tract.

Signs and Tests
1. A dipstick test to detect the presence of nitrites and substances produced by bacteria that caused UTIs. Hence, a positive test indicates that an infection is present before urine cultures are performed.
2. A urinalysis may show white blood cells (WBCs) or red blood cells (RBCs).
3. A urine culture maybe performed to determine the type of bacteria in the urine and the appropriate antibiotic for treatment.

Collection of urine specimen: As an indwelling catheter is in place, the urine should be obtained by sterile aspiration of the catheter with needle and syringe but not from the collection bag.

Guidelines for indwelling catheter urine specimen
1. Do not collect urine from the drainage bag because growth of bacteria outside the catheter may have occurred at this site.
2. Clean the catheter with an alcohol pad.
3. Use a sterile needle and syringe to puncture the tubing. Aspirate the urine directly from the tubing.
4. Transfer the urine to a sterile specimen container.
5. Urine catheter tip cultures are not acceptable.

Suspected Micro-organisms
· Enterobacteriaceae species
o E. coli
o Klebsiella-Enterobacter-Serratia
o Proteus-Providencia-Morganella
· Staphylococcus saprophyticus
· Streptococci species (Entercocci)
· Pseudomonas aeruginosa

Key characteristics of micro-organisms
1) Enterobacteriaceae species
• Gram negative rods
• Facultative anaerobes
• Catalase positive
• Oxidase negative

1a) E. Coli
• Rapidly ferment lactose
• Beta-hemolytic
• Produce positive indole test
• Positive for b-glucoronidase using the substrate
• Ferments mannitol

1b) Klebsiella-Enterobacter-Serratia

Klebsiella species
• Non-motility
• Lysine carbohydrate positive
• Citrate positive
• Have large polysaccharide
• Voges-Proskauer positive
• Rapidly ferment lactose

Enterobacter species
• Motile
• Citrate positive
• Ornithine decarboxylase positive
• Voges-Proskauer positive
• Rapidly ferment lactose

Serratia
• Produces Dnase, lipase and gelatinase
• Voges-Proskauer positive
• Slow fermenter of lactose

1c) Proteus-Providencia-Morganella
• Does not ferment lactose
• Motile
• Grow on potassium cyanide medium
• Ferment xylose
• Urease positive for Proteus species and Morganella morganii
• Urease negative for Providencia species

2) Staphylococcus saprophyticus
• Gram positive cocci arranged in grape-like clusters
• Catalase positive
• Coagulase negative
• Phosphatase negative
• Urease & lipase positive

3) Streptococci species (Entercocci)
• Gram positive cocci arranged in pairs
• Facultative anaerobes
• Catalase negative
• Non-hemolytic
• Bile-esculin positive
• Able to grow in 6.5% NaCl

4) Pseudomonas aeruginosa
• Gram negative motile rods as single/pairs/occasionally short chains
• Oxidase positive
• Does not ferment lactose

References

Geo FB, Janet SB & Stephen AM. (2004). Jawetz, Melnick, & Adelberg’s Medical Microbiology. 23rd edition. McGraw-Hill.
http://en.wikipedia.org > search
http://www3.umdnj.edu/ >micrsweb>case2gramnegatives>intro.html

Case 4 (Najib, 0503217B), (Charmaine, 0503186I)
Particulars of patient

Name : Tong Wei Hong Sex : Male
HRN : OPD 004 IC No. : S000444X
Date of birth : 1/12/39 Age : 68 years
Ward/Clinic : Clinic M Bed No : -------

Clinical diagnosis

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

Introduction

Bronchitis is a respiratory disease where the mucous membrane of the lungs’ bronchial passage is inflamed. The swollen membrane narrows and shut off the tiny airways in the lungs, causing cough that is accompanied with thick phlegm and breathlessness.

The disease comes in two forms: acute (lasting less than 6 weeks) and chronic (reoccurring frequently for more than two years) bronchitis. Acute bronchitis is commonly caused by lung infections where 90% of the infections are of viral origin and the remaining 10% of bacterial origin. Chronic bronchitis may be caused by one or more factors and this include repeated attacks of acute bronchitis which will weaken and irritate bronchial airways over time.

Suspected microorganisms

Influenza A and B
Parainfluenza virus
Moraxella catarrhalis
Haemophilus influenzae
Chlamydia pneumoniae
Pseudomonas aeruginosa
Streptococcus pneumoniae

4 bacteria were short listed for this case:

Streptococcus pneumoniae (S. pneumoniae):
S. pneumoniae are Gram-positive, lancet-shaped cocci (elongated cocci with a slightly pointed outer curvature). They are usually seen as diplococci, but they may also occur singly and in short chains. Individual cells are between 0.5 and 1.25 micrometers in diameter. They do not form spores, and they are non-motile. They lack catalase and ferment glucose to lactic acid

Stain: Gram stain
Biochemical Test: MR-VP test

Pseudomonas aeruginosa (P. aeruginosa):
P.aeruginosa is a Gram-negative, aerobic and rod-shaped bacterium with no particular arrangement. Although it is classified as an aerobic organism, P.aeruginosa is considered by many as a facultative anaerobe as it is well adapted to proliferate in conditions of partial or total oxygen depletion. Adaptation to anaerobic environments is essential for certain lifestyles of P. aeruginosa, like during lung infection in cystic fibrosis patients where thick layers of alginate surrounding bacterial mucoid cells can limit the diffusion of oxygen.

Stain: Gram stain
Biochemical Test: Triple sugar iron (TSI)

Moraxella catarrhalis (M. catarrhalis):
M. catarrhalis is a gram negative, aerobic, oxidase-positive diplococcus which may colonise and cause respiratory tract associated infections in humans.

Stain: Gram stain

Haemophilus influenzae (H. influenzae):
H. influenzae is a non-motile Gram-negative coccobacillus. It is generally aerobic, but can grow as a facultative anaerobe. The organism is also catalase and oxidase positive.

Stain: Gram stain

Investigation

The sputum will be cultured so as to isolate the microorganisms short listed above. Then the biochemical tests specific to each bacteria will be performed so as to identify them.

References

http://en.wikipedia.org>wiki>Haemophilus_influenzae
http://en.wikipedia.org>wiki>Moraxella_catarrhalis
http://en.wikipedia.org>wiki>Pseudomonas_aeruginosa
http://en.wikipedia.org>wiki>Streptococcus_pneumoniae
http://www.medicinenet.com>bronchitis>page3.htm
http://www.nlm.nih.gov>medlineplus>bronchitis.html

Case 5 (Jeremy, 0503168G)
Particulars of patient

Name : Wong Fei Hong Sex : Male
HRN : OPD 0010 IC No. : S210444X
Date of birth : 1/12/70 Age : 37 years
Ward/Clinic : Clinic S Bed No : -------

Clinical diagnosis

Complaints: Fever, swelling around operation wound
Diagnosis: Wound infection
Specimen: Wound swab

Introduction

For a wound to be considered as a surgical site infection, it must fufill the following criteria:
- Infection must occur within 30 days of surgery
- Infection must involve only the skin and subcutaneous tissue
- Must be at least one of the following:
o Purulent discharge from a superficial infection OR
o Organisms isolated from aseptically wound culture
- Must have at least one of these signs:
o Pain or tenderness
o Localised swelling
o Redness or heat

Surgical wound infection is caused by endogenous or exogenous.
An example of endogenous infection is due to poor surgical technique while an example of exogenous infection is due to improper sterilization of instruments.

Suspected microorganisms
Staphylococcus aureus (facultative anaerobe, gram-positive cocci)
Enterococcus faecalis (facultative anaerobe, gram-positive cocci)
Streptococcus pyogenes (facultative anaerobe, gram-positive cocci)
Escherichia coli (facultative anaerobe, gram-negative bacilli)
Pseudomonas aeruginosa (aerobic, gram-negative bacilli)
Clostridium species (anaerobic, gram-positive bacilli)
Enterobacter species (facultative anaerobe, gram-negative bacilli)
Proteus mirabilis (facultative anaerobe, gram-negative bacilli)
Klebsiella pneumoniae (facultative anaerobe, gram-negative bacilli)

5 bacteria were short listed for this case:

Staphylococcus aureus (S.aureus):
S. aureus is a Gram-positive, cluster-forming cocci. Human are the major reservoirs of S.aureus. They are non-motile, non-spore forming facultative anaerobes. They are able to ferment mannitol, and are catalase and coagulase positive. S.aureus is the common microorganism present in surgical-wound infection.

Stain: Gram stain
Biochemical Test: Catalase test, coagulase test, mannitol, DNase test

Enterococcus faecalis (E.faecalis):
E.faecalis is a Gram-positive, facultative anaerobic cocci. Along with E.coli, they are indicators for faecal contamination. E.faecalis have also emerged as a significant, antibiotic-resistant, nosocomial pathogen.

Stain: Gram stain
Biochemical Test: MRVP

Streptococcus pyogenes (S.pyogenes):
S.pyogenes is a Gram-positive, facultative anaerobe cocci. They are non-motile, non-sporeforming cocci that occur in chains or in pairs of cells. They are normal flora of the body but can cause infection after penetrating the host defence.

Stain: Gram stain
Biochemical Test: Catalase test, coagulase test, mannitol

Escherichia coli (E.coli):
E.coli is a Gram-negative, facultative anaerobe bacillus. E.coli can grow in the presence or absence of O2. Under anaerobic conditions it will grow by means of fermentation, producing characteristic "mixed acids and gas" as end products. However, it can also grow by means of anaerobic respiration, since it is able to utilize NO3, NO2 or fumarate as final electron acceptors for respiratory electron transport processes. E.coli is a normal flora of the body, and can be found in intestines and feces of human.

Stain: Gram stain
Biochemical Test: MRVP

Pseudomonas aeruginosa (P.aeruginosa)
P.aeruginosa is a motile, Gram-negative, aerobic bacillus. P.aeruginosa is an opportunistic pathogen in human. They exploit any break of defense in human to cause an infection. It is primary a nosocomial pathogen.

Stain: Gram stain
Biochemical Test: Oxidase test

Investigation

The swab will be cultured so as to isolate the microorganisms short listed above. Then the biochemical tests specific to each bacteria will be performed so as to identify them.

References

http://www.surgical-tutor.org.uk>default-home.htm?principles>microbiology>wound_infection.htm
http://www.textbookofbacteriology.net>e.coli.html
http://www.textbookofbacteriology.net>normalflora.html
http://www.textbookofbacteriology.net>pseudomonas.html
http://www.textbookofbacteriology.net>staph.html
http://www.textbookofbacteriology.net>streptococcus.html

Case 6 (Natalie, 0503275J)
Particulars of patient

Name : Ong Fei Fei Sex : Female
HRN : OPD 0013 IC No. : S210334X
Date of birth : 1/12/70 Age : 37 years
Ward/Clinic : Clinic T Bed No : -------

Clinical diagnosis

Complaints: Fever, pain during urination, virginal discharge
Diagnosis: UTI
Specimen: Virginal Discharge

Introduction

Urinary Tract Infection (UTI) is commonly suspected in clinical practice and up to 50% of all women may suffer from symptomatic UTI at some time during their lives. UTI is considered to be complicated when it affects pregnant women, children, men or the elderly and if it affects kidney tissue (Upper UTI). While simple UTI is uncommon in men aged 20-50, prostatic enlargement in older men may cause urinary tract obstruction and thereby causing UTI.

Normally, urine is sterile. An infection occurs when tiny organisms, usually bacteria from the digestive tract, cling to the opening of the urethra and begin to multiply. The urethra is the tube that carries urine from the bladder to outside the body. Most infections arise from one type of bacteria, Escherichia coli (E. coli), which normally lives in the colon.

In many cases, bacteria first travel to the urethra. When bacteria multiply, an infection can occur. An infection limited to the urethra is called urethritis. If bacteria move to the bladder and multiply, a bladder infection, called cystitis, results. If the infection is not treated promptly, bacteria may then travel further up the ureters to multiply and infect the kidneys. A kidney infection is called pyelonephritis.

Suspected microorganisms

Escherichia coli (E.coli)
Chlamydia
Mycoplasma
Klebsiella pneumoniae
Proteus mirabilis
Psedonmonas aeruginosa,

4 bacteria were short listed for this case:

Escherichia coli (E.coli)
Escherichia coli (E. coli), is a Gram-negative, non-sporulating, facultative eubacterium that is commonly found in the lower gastrointestinal tract of warm-blooded animals. Peritrichous strains are motile, but some strains lack flagella. E.coli are not always confined to the intestine, and their ability to survive for brief periods outside the body make them an ideal indicator organism to test environmental samples for fecal contamination.

As Gram-negative organisms, E. coli are resistant to many antibiotics that are effective against Gram-positive organisms.

Test: Gram-Stain
Treatment: Antibiotics which may be used to treat E. coli infection include (but are not limited to) amoxicillin as well as other semi-synthetic penicillins, many cephalosporins, carbapenems, aztreonam, trimethoprim-sulfamethoxazole, ciprofloxacin, nitrofurantoin and the aminoglycosides

Chlamydia
Chlamydia trachomatis is one of three bacterial species in the genus Chlamydia, family Chlamydiaceae, class Chlamydiae, phylum Chlamydiae, domain Bacteria. C. trachomatis is a gram-negative bacteria. It comprises two human biovars: trachoma and lymphogranuloma venereum (LGV). Many, but not all, C. trachomatis strains have an extrachromosomal plasmid.

Test: Gram-Stain/DNA-based test
Identification: Chlamydia species are readily identified and distinguished from other chlamydial species using DNA-based tests.Most strains of C. trachomatis are recognized by monoclonal antibodies (mAbs) to epitopes in the VS4 region of MOMP
Treatment: It may be treated with any of several antibiotics such as azithromycin,erythromycin or doxycycline/tetracycline.


Klebsiella pneumoniae
Klebsiella pneumoniae is a Gram-negative, non-motile, encapsulated, lactose fermenting, facultative anaerobic, rod shaped bacterium found in the normal flora of the mouth, skin, and intestines. it is distinguished by being indole-negative and by its ability to grow on both melezitose and 3-hydroxybutyrate. It naturally occurs in the soil and about 30% of strains can fix nitrogen in anaerobic condition.

Test: Gram Stain
Biochemical Test: Indole-Test, Melezitose Test

Pseudomonas aeruginosa
Pseudomonas aeruginosa is a Gram-negative, aerobic, rod-shaped bacterium. Almost all strains are motile by means of a single polar flagellum. P. aeruginosa secretes a variety of pigments, including pyocyanin (blue-green), fluorescein (yellow-green and fluorescent, now also known as pyoverdin), and pyorubin (red-brown). P. aeruginosa is often preliminarily identified by its pearlescent appearance and grape-like odor in vitro. Definitive clinical identification of P. aeruginosa often includes identifying the production of both pyocyanin and fluorescein as well as its ability to grow at 42°C. Although classified as an aerobic organism, P. aeruginosa is considered by many as a facultative anaerobe as it is well adapted to proliferate in conditions of partial or total oxygen depletion. This organism can achieve anaerobic growth with nitrate as a terminal electron acceptor, and in its absence it is also able to ferment arginine by substrate-level phosphorylation.

Test: Gram Stain
Biochemical Test: Triple Ion Sugar (TSI)

Investigation

The specimen (virginal discharge) will be cultured so as to isolate the microorganisms short listed above. Next, a series of biochemical tests will be carried out and incubated for a day before verifying the results for the final identification of the specific microorganism.

References

http://en.wikipedia.org>wiki>Chlamydia_trachomatis
http://en.wikipedia.org>wiki>Escherichia_coli
http://en.wikipedia.org>wiki>Klebsiella_pneumoniae
http://en.wikipedia.org>wiki>Pseudomonas_aeruginosa
http://kidney.niddk.nih.gov>Kudiseases>pubs>utiadult>
http://www.medicinenet.com>urine_infection>article.htm