Star Team

References:
1) indonesia/">http://www.wikipedia.com>Indonesia
2) Jensen M. M. and Wright D. N. (1993) Introduction to Micobiology for Health Sciences. Prentices-Hall Inc., new Jersey

Bacteria

Presence of bacteria is extremely high in unhygienic conditions. Individuals infected by bacteria will present symptoms that may be infectious to other uninfected individual thus spreading the infection. [1] The soldiers having their army stint in Indonesia are exposed to numerous bacteria e.g. Shigella species, Salmonella species, E. coli, Campylobacter species as hygiene conditions are often overlooked. In addition, they are exposed to more bacteria during their jungle warfare training as soil, streams, rivers, animal feces, uncooked food which contain abundant amounts of bacteria. [1]

Food borne bacteria
In the jungle, military personnel have to prepare food themselves, thus the chances of consuming un-cook or contaminated food is relatively high. Therefore, they are at risk of be infected with various bacteria that are found on food when not cooked or cleaned properly. [5, 6]

Result from undercooking of food especially meat, organisms that are common include:
Campylobacter jejuni
Salmonella spp
Shigella spp
Listeria monocytogenes
Clostridium perfringens
Clostridium botulinium

Symptoms include:
Nausea
Vomitting
Abdominal cramping
Diarrhea
Fever

Shigella [3]

Transmission: Most individuals are infected with shigellae when they ingest food or water contaminated with human fecal material. Spread is always from a human source.

Causes: Shigellosis

Signs and symptoms: Fever, abdominal cramping, diarrhea, dehydration, tenesmus (straining of stool)

Salmonella species [2]

Transmission: Salmonella is spread from the carcasses to humans when the poultry is eaten undercooked. Other sources of Salmonellae include milk products, food, and water contaminated with animal feces or urine.

Causes: Salmonellosis, enteritis, Typhoid fever, primary septicemia

Signs and symptoms: Nausea, vomiting, abdominal pain, diarrhea, fever, weight loss, anemia, anorexia, headache.

Campylobacter [1]

Transmission: Campylobacter is usually acquired from food (especially poultry), milk, or contact with infected animals (e.g. dogs) through the fecal-oral route.

Causes: Acute enteritis

Signs and symptoms: Bloody diarrhea, fever, abdominal pain, headache.


Animal vector bacteria [1,3]
As the soldiers are training in the jungle, they will encounter many different forms of wildlife. Bacteria carried by them may be transmitted to the soldiers. Common ones include:

Brucella spp (contact with infected animals e.g. dogs, pigs, cattle, horse)
Francisella sp. (contact with wild animals that carry ticks, mites and lice)
Yersinia spp (contact with rodents that carry fleas that carry the bacteria)
Pasteurella multocida (cat and dog bites)

Soil vector bacteria [2, 3]
In addition to the exposure to various fauna and flora living in the jungle, soldiers may also come in contact with soil. Soil is a place where many bacteria flourished thus they chances of getting an infection casued by a soil organism is also relatively possible

Clostrodium tetani [2]

Transmission: Soils or materials in contact with animal wastes are usually heavily contaminated with C. tetani and offer excellent sources of infections. Soil-contaminated wounds are most frequent source of infection.

Causes: Tetanus

Signs and symptoms: Spasmodic contractions (tetany) in involved muscles that are affected by the toxin and muscle spasms.

Mode of control: Vaccines

References

1. Levinson, W. (2006). Review of Medical Microbiology & Immunology. San Francisco, California: Lange Medical Books/McGraw-Hill publishing company

2. Elliott T., Hasting M. and Desselberger U. (1997). Lecture Notes on Medical Microbiology. Blackwell Science, Oxford

3. Walker, T.S. (1998). Microbiology. Philadelphia, Pennsylvannia: W.B Saunders Company

4. http://aggie-horticulture.tamu.edu/ > Search > food poisoning

5. http://www.medicinenet.com/ > Search > food poisoning

Done by: Star team

5. Brook G. F., Butel J. S. and Morse S. A. (1998). Jawetz, Melnick & Adelberg’s Medical Microbiology. Prentice Hall International Inc., United States of America

Introduction
Fungi are eukaryotic cells that lack chlorophyll. They have cell walls and filamentous structures and produce spores, or conidia. These organisms grow as saprophytes that obtain nutrients from dead organic matter. Most of the fungi that cause systemic infections may infect incidental hosts (i.e. humans and other animals) who are exposed to fungal elements or spores.

Systemic mycoses

Histoplasma capsulatum

Epidemiology
Dimorphic fungus (exists as mold in soil and yeast in tissue) that is found worldwide and in temperate, subtropical and tropical climates. Grows in soil and material contaminated with bat or bird droppings.

Causes
Histoplasmosis. Infection occurs resulting from inhalation of spores.

Transmission
Infection resulting from inhalation of spores of the fungi in their mold forms in soil. Within the lungs, the spores differentiate into yeasts.

Signs & symptoms
Fever
Headache
Malaise
Myalgia (muscle pain)
Abdominal pain
Chills

Mode of control
No means of prevention except avoiding exposure in areas of endemic infection.


Dermaphytes

Trichophyton species
Trichophyton ajelloi (geophilic)
Trichophyton concentricum (anthrophilic)
Trichophyton flavescens (geophilic)
Trichophyton gloriae (geophilic)
Trichophyton interdigitale (anthrophilic)
Trichophyton megnini (anthrophilic)
Trichophyton mentagrophytes (zoophilic, anthrophilic) (rodents)
Trichophyton phaseoliforme (geophilic)
Trichophyton rubrum (anthrophilic) (most common)
Trichophyton schoenleinii (anthrophilic)
Trichophyton simii (zoophilic) (monkey, chicken)
Trichophyton soudanese (anthrophilic)
Trichophyton terrestre (geophilic)
Trichophyton tonsurans (anthrophilic)
Trichophyton vanbreuseghemii (geophilic)
Trichophyton violaceum (anthrophilic)
Trichophyton yaoundei (anthrophilic)

Epidermophyton floccosum

Epidemiology
Found worldwide, more common in countries with hot and humid climates with crowding or debilitation

Cause
Dermatophytosis
Tinea corporis (ringworm)
Tinea cruris (jock itch)
Tinea pedis (athlete’s foot)
Tinea manuum (hand)
Tinea favosa (scalp)
Folliculitis (inflammation of hair follicles)
Tinea capitis (hair infection)
Onychomycosis (nail infection)
Invasive infection for immunocompromised host

Transmission
Direct or indirect contact with infected host (human or animal)
Direct or indirect contact with skin or hair
Geophilic species are acquired via contact with spores in the soil
Skin trauma (burns, cuts), high temperature and humidity and increase susceptibility to infection

Signs & symptoms
Itching, stinging and burning between the toes, on the soles
Excessive dryness of the skin on the bottom of the feet
Thick, crumbly, discoloured, brittle and ragged nails
Blisters
Hair loss
Scabs
Tinea of the nails

Mode of control
No means of prevention except avoiding exposure in areas of infection

Microsporum species
Microsporum audouinii (anthrophilic)
Microsporum canis (zoophilic) (cats and dogs)
Microsporum cooeki (geophilic, zoophilic) (fur from cats, dogs and rodents)
Microsporum ferrugineum (anthrophilic)
Microsporum gallinae (zoophilic) (chicken)
Microsporum gypseum (geophilic)
Microsporum nanum (geophilic, zoophilic) (pig)
Microsporum persicolor (zoophilic) (rodents)

Epidemiology
Found worldwide, more common in countries with hot and humid climates with crowding or debilitation

Cause
Dermatophytosis
Tinea corporis (ringworm)
Tinea cruris (jock itch)
Tinea pedis (athlete’s foot)
Tinea manuum (hand)
Tinea favosa (scalp)
Tinea capitis (scalp)
onychomycosis (rare)
Invasive infection for immunocompromised host
Note: Microsporum persicolor does not infect hair

Transmission
Direct or indirect contact with infected host (human or animal)
Direct or indirect contact with skin or hair
Geophilic species are acquired via contact with spores in the soil
Skin trauma (burns, cuts), high temperature and humidity and increase susceptibility to infection

Signs & symptoms
Itching, stinging and burning between the toes, on the soles
Excessive dryness of the skin on the bottom of the feet
Thick, crumbly, discoloured, brittle and ragged nails
Blisters
Hair loss
Scabs
Tinea of the nails

Mode of Control
No means of prevention except avoiding exposure in areas of infection

Malassezia furfur

Epidemiology
Worldwide, part of normal skin flora, infections likely to occur in hot and humid conditions

Cause
Pityriasis versicolor (skin rash)
Seborrheic dermatitis (flaky scalp, face, trunk)
Folliculitis (inflammation of hair follicles)
Blepharitis (inflammation of the eyelids)
Tinea Blanca (hair)
Severe systemic infections in immunocompromised host

Transmission
Normal skin flora, that causes infection when it becomes an opportunistic infection

Signs and Symptoms
Itching
Scaling
However sometimes the infection is asymptomatic


Sporothrix schenckii

Epidemiology
Worldwide, isolated from soil, living and decomposing plants, wood and moss

Cause
Sporotrichosis (Rose handler’s disease)
Invasive sinusitis
Meningitis
Osteoarticular infection
Bursal infection

Transmission
Infection by entry of infecting fungus through the skin via minor trauma (eg. cuts)

Signs & symptoms
Large ulcerations
Small painless skin bump on the arms, fingers or hand
Boil-like lesions
Firm skin nodules
Suppurative nodules


EXCLUSION OF FUNGI

1. Opportunistic mycoses
These fungi cause infections only in immunocompromised people.

Candida species
Candida albicans
Candida guilliermondii
Candida parapsilosis
Candida tropicalis

Epidemiology
This fungus is found worldwide. For some of the fungi, they are part of the normal microbial flora of the mucous membranes of the mouth and intestinal tract.

Cryptococcus species
Cryptococcus neoformans
Cryptococcus gattii

Epidemiology
Cryptococcus neoformans is found in soil, especially those enriched by pigeon droppings
Cryptococcus gattii can be found in eucalyptus trees and decaying wood forming hollows in living trees

Aspergillus species
Aspergillus fumigatus
Aspergillus niger
Aspergillus flavus
Aspergillus terreus
Aspergillus nidulans

Epidemiology
Fungus is ubiquitous in the environment, growing in soil and decaying vegetation. The transmission of fungus is by airborne conidia.

2. Coccidioides immitis

Epidemiology
This fungus is found in arid regions of USA and South America

3. Blastomyces dermatitidis

Epidemiology
This fungus is endemic in North & South America, Africa and in the Middle East.


References
Martin
http://www.emedicine.com > search > Cryptococcosis
http://www.emedicine.com > search > Histoplasmosis
http://www.doctorfungus.org > search > Cryptococcus
http://www.mycology.adelaide.edu.au > search > Candida
Levinson, W. (2006). Review of Medical Microbiology & Immunology. San Francisco, California: Lange Medical Books/McGraw-Hill publishing company.
Murray, P.R. (2002). Medical Microbiology. St. Louis, Missouri: Mosby Inc.
Virella, G. (1997). Microbiology and Infectious Diseases. Pennsylvania, USA: Williams & Wilkins.

Randall
http://www.doctorfungus.org > search > Trichophyton
http://www.wrongdiagnosis.com > T > tinea
http://www.mayoclinic.com/ > health > search > athlete’s foot
http://www.emedicine.com/aaem/topic277.htm
http://www.doctorfungus.org > search > Epidermophyton
http://www.doctorfungus.org > search> Microsporum
http://www.doctorfungus.org > search > Malassezia furfur
http://www.doctorfungus.org > search > Sporothrix schenckii
http://www.wrongdiagnosis.com > S > sporotrichosis
Walker, T.S. (1998). Microbiology. Philadelphia, Pennsylvannia: W.B Saunders Company
Levinson, W. (2006). Review of Medical Microbiology & Immunology. San Francisco, California: Lange Medical Books/McGraw-Hill publishing company.

Done by: Martin Ng & Randall Chua (TG02)

Parasites
Parasites occur in two distinct forms: single celled protozoa and multi-cellular metazoan called helminthes. [1]

Helminthes
Metazoan are subdivided into two groups: Platyhelminthes and the Nemathelminthes.
The phylum Platyhelminthes contains two important classes: Cestodes and Trematoda. [1]

Cestodes are tapeworms. It consists of two main parts: a rounded head called the scolex which has specialized means of attaching to the intestinal wall and a flat body of multiple segments called proglottids. The oldest proglottids at the distal end produce many eggs which are excreted in the feces and transmitted to various intermediate hosts such as cattle, pigs and fish. Humans usually acquire the infection when undercooked flesh containing the larvae is ingested. [1,2]

Taenia solium: T. solium is found worldwide. Because pigs are intermediate hosts of the parasite, completion of the life cycle occurs in regions where humans live in close contact with pigs and eat undercooked pork. Taeniasis and cysticercosis are very rare in predominantly Muslim countries, as Islam forbids the consumption of pork.[1]

Taenia saginata: T. saginata, also known as Taeniarhynchus saginata or the Beef tapeworm, is a parasite of both cattle and humans, but which can only reproduce in humans. T. saginata occurs where cattle is raised, human feces is improperly disposed of, meat inspection programs are poor, and where meat is eaten without proper cooking.[1]

Echinococcus multilocularis: The definitive hosts are mainly foxes and the intermediate hosts are various rodents. Humans are infected by accidental ingestion of food contaminated with fox feces. [2]

Trematodes are transmitted to humans either via penetration of the skin by the free-swimming cercariae of the schistosomes or via ingestion of cyst in undercooked fish or crabs in Clonorchis and Paragonimus infection, respectively. [1]

Schistosoma japonicum: It causes schistosomiasis. It is found only in Asia and domestic animals such as water buffalos and pigs acts as reservoirs. It is common in tropical and subtropical climates. Most patients are asymptomatic, but chronic infections may become symptomatic. [1]

Nemathelminthes are also known as nematodes. They are commonly classified under: intestinal nematodes and tissue nematodes. Infections with nematodes are associated with conditions of poor hygiene. Such infections are extremely common throughout the tropics and subtropics. [1]

Ancylostoma duodenale and Necator americanus: Humans are infected when filariform larvae in moist soil penetrate the skin. They are found especially in tropical areas. [2]

Strongyloides stercoralis: They are found in the soil. It penetrates the skin and occurs primarily in the tropics, especially SEA.[1]

Wuchereria bancrofti : It occurs in tropical and subtropical areas and is endemic in central Africa, along the Mediterranean coast, and in many parts of Asia, including China, Korea, Japan and the Philippines. [1]


Protozoa

Protozoa are eukaryotic cells and have many of the intracellular components of characteristic of higher forms of life. Most protozoa have some form of active locomotion. Protozoa are found in soil, in most bodies of water, and in many higher forms of life. Relatively few protozoa are able to cause diseases in humans and most are beneficial contributors to the various biological cycles in nature.

Protozoa and other parasitic diseases are common in underdeveloped, tropical, and sub tropical area. The overall effects of these diseases on the general well-being of the inhabitants of these areas are of major importance.

The diagnosis, treatment, and control of protozoal diseases differ in some ways from those used for other microbial diseases. Because of the large size, distinct shapes of the protozoa and that protozoa are difficult to grow on media, direct microscopy is the routine method of diagnosis. The symptoms of many protozoal diseases are very general and often are not used solely as a basis for a specific diagnosis.
Control of protozoal disease is often difficult since human are not the primary host, effective vaccines are not currently available, anti- protozoal drugs are often toxic to the host and close contact between humans and the disease reservoir is fostered by poverty and inadequate education.

Some disease-producing protozoa have a life cycle that involves more than one type of host. The parasite goes through sexual reproduction in the definitive host, whereas asexual reproduction occurs in the intermediate human host. The cell types which are present in a host may be the growing trophozoite (damages tissues) and the dormant, resistant cyst (the form which survives outside a host perpetuating transmission by indirect means.


Reasons for inclusion:

Indonesia is a tropical country with high humidity. In places with high humidity, Food and water can be easily contaminated with fecal products of humans and animals. In the jungle, lakes and water bodies are common. Wild animals’ and human feces may contaminate water and food that the soldiers eat thus infecting them. Swimming in lakes and water bodies may also cause infections as some protozoa can be transmitted through mucosal membranes, trauma and ingestion. Usually in the army, personal hygiene is always overlooked, combined with the poor sanitation of the jungle, it can cause severe infection. During military training, there is also personal time. Soldiers may explore neighboring villages. The soldiers may get infected with diseases during sexual intercourse with women in neighboring villages. In the villages, they may also come into contact with domestic animals.


Entamoeba histolytica [1]

Epidemiology: It is a cosmopolitan intestinal parasite. Organism is acquired by ingestion of cysts that are transmitted primarily by the fecal-oral route in contaminated food and water. Anal- oral transmission, e.g., among male homosexuals, also occurs. There is no animal reservoir. Infection is found worldwide, but occur most frequently in tropical countries, especially in areas with poor sanitation. There are no animal reservoirs.

Causes: Amebic dysentery and liver abscess

Signs and Symptoms:
Asymptomatic infection is common.

Acute intestinal amebiasis: Dysentry (bloody, mucus-containing diarrhea) accompanied by lower abdominal discomfort, flatulence, and tenesmus

Chronic amebiasis: occasional diarrhea, weight loss, and fatigue. Granulomatous lesion called an ameboma may form in the cecal or rectosigmoid areas of the colon.

Amebic abscess (liver): right upper-quadrant pain, weight loss, fever, and a tender, enlarged liver. Right lobe abscesses can penetrate the diaphragm and cause lung disease. Aspiration of the liver abscess yields brownish-yellow pus with the consistency of anchovy-paste.

Mode of Control: Improving basic hygiene and sanitation practices is most effective way of controlling outbreaks of amebiasis as E. histolytica thrives in conditions where standards of sanitation and hygiene are poor. Avoid fecal contamination of food and water and observing good personal hygiene such as hand washing. Water supplies should be purified before drinking. Vegetables should be cooked as it may have been contaminated with human feces.


Giardia lamblia [1.2]

Epidemiology: This organism is a cosmopolitan parasite, common in temperate latitudes and in developed countries, as well as in the tropics. Transmission occurs by ingestion of the cyst in food and water contaminated by feces. The source of infection may be infected human carriers or wild animals that act as reservoirs of infection and contaminate water with the cysts passes in their feces. They pass cyst in the stool which then contaminates the water supplies. Outbreaks are usually related to contaminate water supplies. Drinking of untreated stream water will lead to infection.

Causes: Giardiasis

Signs and Symptoms: Non bloody, foul- smelling diarrhea, nauseam anorexia, flatulence, and abdominal cramps that persist for weeks or months

Mode of Control: Water should be boiled, filtered, or treated with iodine. Frequent hand washing and separation of feeding areas from sanitization area. No vaccine or preventive drug is available.


Plasmodium Species [1]

Epidemiology: The four species are P. vivax, P. falciparum, P. ovale and P. malariae. P. vivax and P. falciparum are the most common causes. The vector and definitive host is the female Anopheles mosquito. Disease is transmitted primarily by the bite of the mosquito. This occurs mainly in the tropical and subtropical areas, especially in Asia.

Causes: Malaria

Signs and Symptoms: Abrupt onset of fever and chills, accompanied by headache myalgias (muscle pain), and aethralgias (joint pains). Fever can reach up 41°C. It is accompanies by shaking chills, nausea, vomiting and abdominal pain. Splenomegaly and hepatomegaly is commonly seen and anaemia is prominent.

P. falciparum can cause extensive brain (cerebral malaria) and kidney (blackwater) damage. (large scale intravascular lysis)

Mode of Control: Chemoprophylaxis of malaria. The use of mosquito netting, window screens, protective clothing and insect repellent can lower the probability of mosquito bites. Elimination of breeding site includes drainage of stagnant water in swamps and ditches, removal of standing water and improvement in land drainage.


Toxoplasma gondii [2]

Epidemiology: Infection by T. gondii occurs worldwide. It is an obligate intracellular parasite widespread among wild and domestic animals. The definitive host is the domestic cat and other felines; humans and other mammals are intermediate host. Cockroaches, flies and earthworms serve as non-infected transport host. Infection begins with the ingestion of cyst in undercooked meats or from contact with cat feces.

Causes: Toxoplasmosis

Signs and Symptoms: Resemble infectious mononucleosis (fever, muscle soreness, sore throat, fatigue)

Mode of control: Control is difficult because of the diversity of animal reservoir. To prevent toxoplasmosis, cook meat thoroughly to kill cyst. Cat should not be fed raw meat as it is the definitive host of the obligate intracellular parasite.


Acanthamoeba castellanii [2]

Epidemiology: They are free-living amoebae. They are found in warm freshwater lakes and in soil. Infection can occur when swimming in the lakes and when there is skin trauma

Causes: Meningeoncephalitis

Signs and Symptoms: eye infection

Mode of Controls: Do not swim in lakes. Avoid traumatized skin contact with lake water or soil.


Naegleria fowleri [1,2]

Epidemiology: N. fowleri thrives in warm water at lower oxygen tensions. It is often found in the mud at the bottom of puddles, ditches, and lakes. It enters the body trough mucous membranes or inhalation while an individual is swimming.

Causes: Meningeoncephalitis

Signs and Symptoms: headache, fever and deteriorates rapidly into coma

Mode of Controls: Do not swim in lakes. Avoid traumatized skin contact with lake water or soil.


Cyclospora cayetanensis [2]

Epidemiology: It is distributed worldwide. Infection appears to be acquired by ingestion of oocytes in water or with contaminated fruits or vegetables.

Causes: “Traveler’s diarrhea”

Signs and Symptons: Water diarrhea with low grade fever and abdominal cramps

Mode of control: Treatment of water supplies with chlorination and filtration. Improving basic hygiene and sanitation practices is most effective way of controlling outbreaks.


Cryptosporidium parvum [1]

Epidemiology: Infection is usually water borne or acquired from animals. Oocytes are present in animal feces and can be transmitted to the human through the fecal- oral route.

Causes: cryptosporidiosis

Signs and Symptoms: Diarrhea leading to fluid loss and malnutrition

Mode of control: Improving basic hygiene and sanitation practices is most effective way of controlling outbreaks. Swimming in lakes or any bodies of water is not advisable.


Trichomonas vaginalis [1,2]

Epidemiology: The primary location is the vagina and the prostate. Transmission of this organism is by sexual contact. It is predominantly a vaginal parasite, although urethritis may occur in the male consorts of infected women. Infection in women is more severe than in men.

Causes: Urinary tract infection

Signs and Symptoms: Pain during urination, discharge

Mode of Control: Abstinence or no unprotected sex


Reasons for exclusion:

Trypanosoma cruzi (Chagas’ disease) [1]
Vector is the reduviid bug. Animal reservoirs include wild species such as the armadillo, raccoon and rat. Rats are commonly found in jungles. However, this disease is common in America only.

Trypanosoma brucei rhodesiense (sleeping sickness) [1]
Wild animals as reservoirs for the tsetse fly, Glossina. Wild animals are abundant in jungles. But this disease occurs primarily in West Africa therefore it is ruled out.

Leishmania donovani [1]
Lifecycle involves the sandfly as the vector and a variety of mammals such as dogs, foxes and rodents as reservoirs. However, this is common in Middle East, Africa, Russia, India and China

Babesia [1,2]
Human infection is rare. Cases reported in Africa, New York, and Southern Europe. In immunocompetent individuals the disease is usually self limiting, so no specific treatment is required.

Isopora belli [2]
It causes self limiting diarrhea in immunocompetenet individuals, but in more severe cases, particularly in patients that are immunocompromised (e.g. AIDS), they will cause severe diarrhea. Medical screening is usually done before enlistment to determine if the individual is healthy. If the individual is immunocompromised, it can be detected during the screening.

Balantidium coli [1,2]
Domestic animals, especially pigs are the main reservoir for the organism. It is commonly associated with pig farming. Humans are infected after ingestion of the cyst in food or water contaminated with animal or human feces. However, it rarely causes disease in humans. Most individual are asymptomatic; diarrhea rarely occurs.


References:
1)Levinson, W. (2006). Review of Medical Microbiology and Immunology. San Francisco,California: Lange Medical Books/ McGraw-Hill Medical Publishing Division

2)Murray, P.R. (2002). Medical Microbiology. St. Louis, Missouri: Mosby Inc.


Done by:
Foo Yong Yang
Loh Mun Jo-anne
TG02

Name: Wong Fei Hong (Case 5)
Sex: Male
Age: 37 years old
Type of Specimen: Wound swab

Clinical diagnosis
Symptoms: Fever, swelling around operation wound
Diagnosis: Wound infection

Approach for the diagnosis of wound infection [1]

1. A wound swab was obtained


2. The specimen is gram-stained. The shape, size, arrangement and whether they are gram-positive or gram-negative should be observed.


3. The specimen is cultured on appropriate media, to obtain ‘pure culture’. Plates should be incubated in aerobic or anaerobic environment.


4. Identification of the organism by means of biochemical tests.
   
5. Perform antibiotic susceptibility tests.
   
   Note: Wound infection could be due to multiple organisms, so it’s important to culture the specimen on several different media under different atmospheric conditions e.g. aerobic or anaerobic. Gram staining provides useful information about the range of possible causative agents. [1]

Short listed bacteria [1, 2]:
Gram-positive cocci:
1) Streptococci Species (Facultative anaerobes)
- Enterococcus faecalis
- Streptococcus pyogenes

2) Staphylococcus aureus (Facultative anaerobes)

Gram-positive rod:
1) Bacillus anthracis (Strict aerobes; spore formation)

2) Clostridium perfringens (Stric anaerobes; spore formation)

Gram-negative rod:
1) Pseudomonas aeruginosa (Strict aerobes)

2) Enterobacteriaceae species (Facultative anaerobes)
- Escherichia coli
- Enterobacter cloacae
- Serratia marcescens
- Proteus mirabilis
- Klebsiella pneumoniae


Differential diagnosis for gram-positive cocci:

Streptococci Species
1] Enterococcus faecalis
2] Streptococcus pyogenes

Sheep blood agar:[1]

- Beta-hemolytic colonies: Streptococcus pyogenes
- Gamma-hemolytic or lack of change in the medium around colonies: Enterococcus faecalis

Catalase test: Both Negative [2]
This test differentiates Streptococci Species and Staphylococcus aureus.

- A drop of hydrogen peroxide is added to a sample from the bacterial colony
- Presence of catalase converts hydrogen peroxide into water and oxygen
- Results in the production of visible bubbles

Bile-esculi agar: [2]

- Positive: E. faecalis
- Negative: Strep. Pyogenes

E. faecalis hydrolyse esculin, which results in black discoloration of the agar.

Hypertonic (6.5%) NaCl solution:

- E. faecalis grows, while Strep. Pyogenes doesn’t

Antibody Susceptibility: [1]
1] Enterococcus faecalis:
- Penicillin
- Erythromycin
- Bacitracin
- Tetracycline

2] Streptococcus pyogenes:
- Penicillin
- Erythromycin
- Bacitracin
- Tetracycline

Staphylococcus aureus [1]

Sheep blood agar:
- Beta-hemolytic colonies indicate Staphylococcus aureus

Catalase test: Positive

Coagulase test: Positive

Coagulase test differentiates S. aureus, from other staphylococcus species such as S. epidermidis present in the normal flora.

Mannitol Salt Agar: Yellow colonies

Differential test for mannitol fermentors and non-fermentators; S. aureus ferments mannitol.

Antibody susceptiblity:
- Vancomycin
- Gentamicin
- Nafcillin
- Methicillin

Picture 1: Beta-hemolysis represented by beta sign and gamma-hemolysis represented by gamma sign
Picture: micrsweb>lab3grampostitive>lab3introtomethods.html/">rsweb>lab3grampostitive>lab3introtomethods.html/">http://www3.umdnj.edu>micrsweb>lab3grampostitive>lab3introtomethods.html

Picture 2: Catalase positive reaction indicating S. aureus infection
Picture:html>stitch.phps=72880656991974&id=39006255044608/">html>stitch.phps=72880656991974&id=39006255044608/">html>stitch.phps=72880656991974&id=39006255044608/">html>stitch.phps=72880656991974&id=39006255044608/">http://www.cfkeep.org>html>stitch.phps=72880656991974&id=39006255044608

Differential diagnosis for spore forming gram-positive rods:

Bacillus anthracis: [1, 2]

Blood agar (Culture aerobically): Gamma-hemolytic ;large and irregular colonies

Egg yolk agar: Lecithinase negative

In addition, it can be diagnosed by means of antigen and molecular tests.

Antibody susceptibility: [1]
- Ciprofloxacin
- Doxycycline

Clostridium perfringens: [2]

Blood agar (Culture anaerobically): Double zone of beta-hemolysis

Egg yolk agar: Lecithinase positive

Antibody susceptibility: [1]
- Penicillin

Picture 3: Left plate: Positive test for lecithinase; Right plate: Negative for lecithinase
Picture: kkiser>lecithpn.jpg/">kkiser>lecithpn.jpg/">kkiser>lecithpn.jpg/">http://users.stlcc.edu>kkiser>Lecithpn.jpg

Differential diagnosis for gram-negative rods:[1]

MacConkey agar: selective agar used for the isolation and differentiation of lactose-fermenting and non-fermenting gram-negative rods. Alternatively, Eosin Methylene Blue Agar (EMB) can be used.

Lactose fermenters: Escherichia coli, Klebsiella pneumoniae, and Enterobacter cloacae; form red to pink colonies.

Non-fermenters: Proteus mirabilis, Pseudomonas aeruginosa and Serratia marcescens; form colourless colonies

Oxidase test: [1]
- Positive: Pseudomonas aeruginosa
- Negative: Enterobacteriaceae species

This test determines the presence of oxidase enzyme. An oxidase paper is touched onto colony, in the presence of oxidase, paper changes to purple (Pseudomonas aeruginosa).

Triple sugar iron agar: [1]

Biochemcial tests: Indole test, Methyl red test, Voges-Proskauer reaction test and Citrate utilization test (IMViC test)

Pseudomonas aeruginosa: [1]
- Colonies: Fluoresce under ultraviolet light; give fruity odor and grow at 42° C
- Indole test: Negative
- Methyl red test: Positive
- Voges-Proskauer reaction test: Negative
- Citrate utilization test: Positive
- Urease test: Negative

Antibody susceptibility: [1]
- Penicillin e.g. ticarcillin or piperacillin
- Aminoglyoside e.g. gentamicin or amikacin

Enterobacteriaceae species:

1] Escherichia coli [1]
- Motile
- Indole test: Negative
- Methyl red test: Positive
- Voges-Proskauer reaction test: Negative
- Citrate utilization test: Negative
- Urease test: Negative

Antibody susceptibility: [1]
- Cefotaxime
- Gentamicin
- Ampicillin
- Loperamide

2] Enterobacter cloacae [1]
- Indole test:Negative
- Methyl red test: Negative
- Voges-Proskauer reaction test: Positive
- Citrate utilization test: Positive
- Urease test: Negative

Antibody susceptibility: [1]
- Imipenem
- Gentamicin
- Cefotaxime

3] Serratia marcescens [1]
- Indole test: Positive
- Methyl red test: Negative
- Voges-Proskauer reaction test: Positive
- Citrate utilization test: Positive
- Urease test: Negative

Antibody susceptibility: [1]
- Imipenem
- Gentamicin
- Cefotaxime

4] Proteus mirabilis [1]
- Indole test: Positive
- Methyl red test: Positive
- Voges-Proskauer reaction test: Negative
- Citrate utilization test: Positive
- Urease test: Positive

Antibody susceptibility: [1]
- Ampicillin
- Cefotaxime

5] Klebsiella pneumoniae [1]
- Indole test: Positive
- Methyl red test: Positive
- Voges-Proskauer reaction test: Positive
- Citrate utilization test: Positive
- Urease test: Positive

Antibody susceptibility: [1]
- Imipenem
- Gentamicin
- Cefotaxime

Refereneces:

1)Levinson, W. (2006). Review of Medical Microbiology and Immunology. San Francisco,California: Lange Medical Books/ McGraw-Hill Medical Publishing Division

2)www.textbookofbacteriology.net>search

Foo Yong Yang
TG02
0503196H

http://www.yahoo.com/> Image search> MacConkey agar

Eosin-Methylene Blue Agar
Eosin-methylene blue agar is a selective, differential medium used for the isolation and identification of gram-negative bacteria. Eosin Y and methylene blue dyes inhibit the growth of gram-positive bacteria and allows the growth of gram-negative bacteria. Lactose and sucrose are incorporated to allow differentiation of isolates based on lactose fermentation. Fermentation is detected by colour changes and precipitation of dye such as pH drops. Sucrose is incorporated as it serves as another alternative source of carbohydrates for slow lactose fermenters.

http://www.yahoo.com/> Image search> Eosin-methylene blue agar


Blood Agar
Blood agar is used to cultivate a wide variety of fastidious bacterium. An infusion agar of tryptic soy agar base can be enriched by the addition of 5-10% of sheep’s, rabbit’s blood. The incorporation of blood not only provides enrichment for growth but also allows the detection and characterization of haemolytic patterns

micrsweb>lab3grampostitive>lab3introtomethods.html/">http://www3.umdnj.edu>micrsweb>lab3grampostitive>lab3introtomethods.html/

-alpha-hemolysis represented by alpha sign
alpha-hemolysis is the partial clearing of the blood around the colony that results in a green discoloration pf the medium


-Beta-hemolysis represented by beta sign
Beta-hemolysis is the complete clearing of blood around the bacterial colonies. This is due to the complete haemolysis of the surrounding RBCs

-Gamma-hemolysis represented by gamma sign
Gamma-hemolysis occurs when the RBCs surrounding the colonies are unaffected and there is no change in the colour of the bacterial colonies


Mannitol Salt Agar
Differential test for mannitol fermentors and non-fermentators; S. aureus ferments mannitol producing yellow colonies.

Left: Non- fermented mannitol Right: Fermented mannitol (S. aureus)

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Triple Sugar Iron Agar Test
TSI contains glucose, lactose and sucrose as well as a pH-sensitive color indicator. It also contains an iron ingredient for detecting hydrogen sulfide production, which blackens the medium if it occurs. Fermentation of the sugars by the test organism is interpreted by color changes in the butt
and the slant of the mediums.

Interpretation
Acid (yellow) butt – alkaline (pink) slant (K/A)
- glucose fermented
- sucrose or lactose not fermented

acid butt - acid slant (A/A)
- glucose fermented
- lactose and/or sucrose fermented

alkaline butt - alkaline slant (K/K)
- neither glucose, lactose or sucrose fermented
- organism probably not an enteric bacillus

Gas production - indicated by bubbles in the butt. With large amounts of gas production, the agar may be broken or pushed upward.
Hydrogen sulfide production - indicated by any amount of blackening in the butt.

TSI agar

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Interpretation Positive test: bright pink colour at the interface between the broth and the reagent Negative test: No colour change or a slight yellow ring

Methyl Red test
Methyl red is an acid sensitive dye that is yellow at pH above 4.5 and red at pH below 4.5. This test is used to indicate whether glucose has been broken down completely to highly acidic end products or only partially to less acidic end products.

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Interpretation
Positive - strong red color will develop
Negative - no color change


Citrate test
Organisms that are capable of utilizing citrate as their sole carbon source also utilize ammonium salts as their sole nitrogen source. This results in an alkaline environment that turns the indicator bromothymol blue to an intense blue when pH is above 7.6. This characteristic can used to differentiate members of the family Enterobacteriaceae and other gram-negative rods.

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Oxidase Test
Use to determine the presence of oxidase enzymes. An oxidase paper is touched onto colony, in the presence of oxidase, paper changes to purple indicating Pseudomonas aeruginosa. The absence of oxidase result in no colour change, and is indicative of Enterobacteriaceae family


Coagulase Test
Coagualse test is performed to test the ability of the microorganism to clot plasma by the action of coagulase

Rabbit plasma is incubated with staphylococcus colony If staphylococci isolated from infection are able to clot blood, then they are coagulase positive S. aureus.

Coagulase test differentiates S. aureus, from other staphylococcus species such as S. epidermidis and S. saprophyticus present in the normal flora.

Right: Coagulase negative; Left: Coagulase positive

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DNase Test
Extacellular DNases are only present in a few bacterial species such as Serratia marcescens, Pseudomonas aerugionsa. The biochemical and morphology of other Enterbacteriaceae are similar especially in Klebsiella-Enterobacter supp. The only difference which separates S. marcescens from Klebsiella-Enterobacter supp is the DNases that is produced.


8% sodium chloride broth
Test is done to differentiate vibrio cholerae and vibrio parahaemolyticus.
Unlike vibrio parahaemolyticus, vibrio cholerae cannot grow at 8% sodium chloride broth.


Spore stain (Bartholomew and mitters)
In spore stain test, hot malachite green is used to stain spores, while the safranin is used as a counterstain. In this test, spores would be stain green and the rest of the cell appear pink/red

References
http://www.mc.maricopa.edu/~johnson/labtools.html> Optochin sensitivity & bile solubility

http://en.wikipedia.org/wiki/Main_Page > search > catalase

http://en.wikipedia.org/wiki/Main_Page > search > oxidase

Mahon C. R. and Manuselis G, Jr (1995). Textbook of Diagnostic Microbiology. W.B. Saunders Company

MacFaddin J. F. (2000). Biochemical Test for Indentification of Medical Bacteria. Lippincott Williams & Wilkins


search>pseudomonas/">www.textbookofbacteriology.net>search>pseudomonas


Levinson, W. (2006). Review of Medical Microbiology and Immunology. San Francisco,California: Lange Medical Books/ McGraw-Hill Medical Publishing Division