Microbiology practical Medical.

Students 3level lectures Genius University

Clostridium tetani

Specimen collection:
To diagnose Clostridium tetani, specimens that can be collected for laboratory analysis include:
1. Wound swabs or tissue samples: If the patient has a wound that may be infected with C. tetani, a swab
or tissue sample can be collected. The specimen should be collected using sterile technique and
transported to the laboratory as soon as possible.
2. Blood: A blood sample may be taken to look for the presence of antibodies against the tetanus toxin.

 Morphology:
 Gram-positive bacilli, motile
 Capsulated: They produce capsule in the patient’s tissue.
 Spores forming . 4S: site : terminal , shape: round, size: spherical projecting , stain: not stain by gram
stain. (drum stick appearance).
 They are found in the soil and the intestine of man and animals.
 Culture:
 Anaerobes
 Growth on simple media
 Growth rapidly on Robertson cooked met broth, incubated at 37 C for 24 hrs. non saccrolytic &
nonprotolytic after 6 hrs from growth in Robertson cooked met broth act sub culture on blood
agar . swarming and β hemolysis after incubated at 37 C for 24 hrs
 Toxin detection

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 Microbiology practical Medical. Students 3level lectures Genius University

Robertson cooked met broth

Blood agar
Anaerobe Jar

Listeria monocytogenes
L monocytogenes is important as a cause of a wide spectrum of disease in animals and humans. L
monocytogenes is capable of growing and surviving over a wide range of environmental conditions.
It can survive at refrigerator temperatures (4°C), under conditions of low pH and high salt conditions.
Therefore, it is able to overcome food preservation and safety barriers, making it an important foodborne
pathogen.
Diseases: abortion, meningitis, gastroenteritis
neonatal sepsis, pustular lesions
Specimen collection:
Blood , CSF, stool
Morphology and Identification :
L.monocytogenes is a short, gram-positive, non–spore-forming rod. It is catalase positive and has a
tumbling end-over-end ,motility at 22–28°C but not at 37°C; the motility test rapidly differentiates
Listeria from diphtheroids .

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 Microbiology practical Medical. Students 3level lectures Genius University
Material :
1- Blood agar plate General character of Listeria spp
2- Gram stain 1- G + ve rods
3- oxidase reagent. 2- Short, 0.4 – 0.5 X 0.5 – 2.0 μm
4- Loop
with rounded ends
5- Slid
6- Listeria selective agar ( Oxford) 3- Aerobe or facultative anaerobes
7- Hydrogen peroxide 3% ( catalase reagent) 4- Motile by peritrichous flagella
8- Bunsen burner 5- Non – capsulated
9- Marker 6- Fastidious
10- Microscope 7- Catalase positive
11- Autoclave 8- Oxidase negative
12- Incubator
Procedures :
1- Preparation of blood agar and Listeria selective agar ( Oxford)
2- Inoculated patient specimen
3- The organisms grows best at 35 – 37"C and growth is examined after 16 – 48 hr
Colonial appearance :
• On the blood agar :
Colonies are 0.5 – 1.5 mm in diameter , smooth, translucent
characteristic small zone of β hemolysis(1-2mm) around and under colonies.
• On the Listeria selective agar ( Oxford) :
Black colonies approximately 1mm in diameter that surrounded by black halos, after 24 hr.
These colonies typically become 2 – 3 mm in diameter after 48 hr, remaining black with black halo,
but colonies develop sunken center. This characteristics blacking is caused by the indicator
(aesculin). L.monocytogenes will hydrolyse aesculin and associated reaction with the ferric
ammonium citrate give rise to black precipitate around the colonies

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 Microbiology practical Medical. Students 3level lectures Genius University

Microscopic Appearance :
o Gram stain :
G + ve rods, Short, with rounded ends, single or pairs (diplococcic), non – capsulated , non-spore,
Biochemical tests :
 Catalase test : positive
 Oxidase test : negative

Motility test = motile

Treatment :
Ampicillin ± Aminoglycoside.
TMP-SMZ (Tri-methoprim- sulfamethoxazoleis)

Viral Meningitis
Meningitis is a serious condition that can be caused by various infectious agents, including bacteria, viruses,
and fungi. Some of the viral infections that can cause meningitis include:
1. Enteroviruses: These are a group of viruses that can cause a range of infections, including meningitis.
Some examples of enteroviruses that can cause meningitis include Coxsackie virus and Echovirus.
Lab diagnosis: Throat swab, Stool, CSF, and Conjunctival swab = RT – PCR
Cell culture

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 Microbiology practical Medical. Students 3level lectures Genius University
Neutralization
Detection of rising antibody titres
Polio virus : Throat swab, stool = EM or PCR. Tissue Culture (Cell death), Rising titer

2. Herpesviruses: Herpes simplex virus (HSV) and

Varicella-zoster virus (VZV) are two types of
herpesviruses that can cause meningitis.
3. Mumps virus: Mumps is a viral infection that can cause
meningitis, and it is especially common in children.
4. Rabies virus
5. Influenza virus: While rare, the influenza virus can cause meningitis in some cases.
Viral diagnosis: ELISA and PCR
Detection of specific viral Ag by (ELISA)
Detection of Nucleic acid of the virus by PCR

Fungi
Cryptococcus neoformans
Coccidoides immitis
Histoplasma capsulatum

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 Microbiology practical Medical. Students 3level lectures Genius University
Blastomyces dermatitides
Candida spp.

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