DIAGNOSIS OF MENINGITIS SKILL BASED DIAGNOSIS OF MENINGITIS

8/7/2019 DIAGNOSIS OF MENINGITIS SKILL BASED DIAGNOSIS OF MENINGITIS

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DIAGNOSIS OF MENINGITISSKILLS FOR RESIDENTS

Dr.T.V.Rao MD

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Why Skill Based Learning for Residents2

The Indian Medical Curriculum is in for Rapid changes

for making the MBBS Doctors competent to performseveral life saving procedures learned to the greater

perfection. This programme is created that young

residents to learn the life saving diagnosis ofMeningitis by doing a Lumbar puncture and simple

observation in emergency hours with interactive

observation. Dr.T.V.Rao MD



What is Meningitis Meningitis is an infection

of the coverings aroundthe brain and spinal cord.

The infection occurs mostoften in children, teens,

and young adults. Also atrisk are older adults andpeople who have long-term health problems, such

as a weakened immunesystem.

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Why Diagnosing Meningitis is

Important

Diagnosing Meningitis is top priority in clinicalMedicine, in particular Bacterial meningitis, can be

a life threatening condition , the need for

appreciate antibiotic therapy at the earliest is a

priority.

Even with Minimal Diagnostic faculties if done withprecision can reduce morbidity and mortality

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On suspicion of Meningitis

Every patient suspected of having Meningitis should have aspecimen of CSF examination inthe laboratory to establish the

infection and to rule out infection.

5



Basic Understanding on

Meningitis

On a broad basis

Meningitis is

classified as1 Purulent

Meningitis

2 AsepticMeningitis

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What is Purulent Meningitis

The CSF appearstypically turbid due to

the presence of

Leucocytes 100 toseveral thousands / mm3

most of which are

Polymorph nuclear leucocytes

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Neonates and Infants

Meningitis

There is specific affinity of some pathogens infectingNeonates and Infants

1 Coli forms

2 ß hemolytic streptococci

3 Pseudomonas

4 Salmonella and Listeria Monocytogenes

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Iatrogenic Meningitis

Carelessly performed Lumbarpuncture

Accidental wound infectionin neurosurgical wounds

Pyogenic StaphylococcusStreptococci

Coli form bacilli

Anaerobic cocciBacteriods

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Aseptic Meningitis

In these conditions CSF isclear or only slightlyturbid contain moderatenumber of leucocytes

10 – 500 / mm3Majority of cells arelymphocytes, except inearly stages. majority

are caused by viruses

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Etiological agents of Aseptic Meningitis

Enteroviruses

ECHO virusesCoxsackie virus

Polio virus

Mumps virus

moderately infectiveHerpes simplex

Varicella zoster

Measles – Adenovirus

Arboviruses

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CSF resembles - Aseptic Meningitis in

Several other Infections

Few conditions associatedwith other etiologicalagent resemble asepticmeningitisLeptospirosis

( Serovars Canicolaicterohaemorrhagea ) Fungi ( Cryptococcusneoformans )

Amoeba – Naegleria,Harmanella.

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Confusing CSF appearance When early treatment is

given in Bacterialmeningitis the Clinico

pathological appearance

appears as Viral meningitis In viral Encephalitis

moderate Lymphocyte

exduate is found as it inViral meningitis

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Tuberculosis Meningitis

On many occasions Tuberculosis

present as Aseptic meningitis,results from Pulmonary ormesenteric tuberculosis

Can be associated with Miliarytuberculosis.

Cell counts on CSF will reveal 100– 500 leucocytes / mm3

Majority are Lymphocytes

May form veil clot when CSF isallowed to stand in a undisturbedstate.

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WHY MICROBIOLOGICAL DIAGNOSIS IS

LIFE SAVINGInformation derived from the results has impact on :

Diagnosis of infectious diseasesAntibiotic prescribing

Formulation of local antibiotic policyPublic health impact eg

Meningococcal infection.

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Why specimen collection is Important

in Microbiology

Specimen collection in Microbiology to isolate and

identify the causative agents forms back bone of the investigative procedures.

In developing world, lack of awareness and casualattitude among junior staff hampers the definitivediagnosis.

Specific procedures in collecting specimens willcertainly improve the quality of services of Microbiology Departments

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Some tips better Diagnosis Laboratory investigation should start as early as possible

Specimens obtained early, preferably prior toantimicrobial treatment likely to yield the infective

pathogen

Before doing anything, explain the procedure to patient

and relatives

When collecting the specimen, avoid contamination Take a sufficient quantity of material

Follow the appropriate precautions for safety19



An Ideal Request formAn Ideal Request formAn Ideal Request formAn Ideal Request form

Name xxxx Age Sex IP/ OP No xyz Time Date

Ward xx123 Urgent / Routine

Nature of specimen CSF Investigation needed xxxx

xxxx

Doctor/Staff

Contact No 1234567

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Why Proper written RequestWhy Proper written RequestWhy Proper written RequestWhy Proper written Request Your request is a legal document.

Identifies all the outcome of test.

No interchange of results.

Short forms are dangerous

Signature of the Doctor / Nurse is essential in

legible form, can help to contact in case ofresults which can save a patient.

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Specimen collection for

CSF Examination

Lumbar puncture to

collect the CSF forexamination to becollected by Physician

trained in procedurewith asepticprecautions to prevent

introduction of Infection.

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Procedure to collect CSF

The trained physician willcollect only 3-5 ml into alabeled sterile container

Removal of large volume ofCSF lead to headache,

The fluid to be collected at the rateof 4-5 drops per second.

If sudden removal of fluid isallowed may draw down

cerebellum into the Foramenmagnum and compress the Medullaof the Brain

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CSF needs a New and Sterile container

Fresh sterile screw cappedcontainer to be used.

Reused containers, not tobe used, contamination

from the previousspecimens misrepresentthe present specimen.

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Lumbar puncture for CSF collection

The best site for puncture is interspace between 3 and 4 lumbarvertebrae

( Corresponds to highest point of iliac crest )

The Physician should wear sterile

gloves and conduct theprocedure with sterileprecautions, The site ofprocedure should be disinfectedand sterile occlusive dressingapplied to the puncture site afterthe procedure.

25





Preservation of CSF

It is important when there isdelay in transportation ofspecimens to Laboratorydo not keep in

Refrigerator, which tendsto kill H. Influenza If delay is anticipated leave

at Room Temperature.

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Laboratory Examination of CSF

The specimens should beexamined with naked eye

Look for Turbidity

Contamination

with BloodNormal CSF appears likewater

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Specimen Examination

CSF to be examined for

Cell counts

Gram staining

Culturing

Estimation of

protein and glucose

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Cell counts in CSF Microscopic examination of

uncentrigured, well mixedCSF is done in slidecounting chamber.

Count the number of

Polymorphs

Lymphocytes

Erythrocytes

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Normal cell counts31

CSF normally contains 0- 5 leucocytes / mm3

Mainly LymphocytesNewly born children contain up to 30/mm3

Mainly polymorphsIn purulent Meningitis there are usually 100 – 300

leucocytes/mm3

In aseptic meningitis there are usually 10 – 500leucocytes/mm3

Mostly lymphocytes, though polymorphs maypredominate in the earliest stage of the illness.

In Tuberculosis meningitis there are usually 100 – 500leucocytes/mm3



Care in Counting the Cells

When counting the cells,care must be taken toidentify the RBC and

rare presence ofyeasts, amoebashould not be

mistaken forleukocytes

32



Differential Leukocyte counts

If there is any difficulty indifferentiating polymorphs andlymphocytes in the countingchamber

Make a film of cellular depositafter specimen has beencentrifuged

Stain withMethylene blueleishmans or Carol thionineand examined under oilimmersion to asses therelative number of twotypes of leucocytes

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Gram Staining of CSF

The CSF to be centrifuged to deposit the cells andbacteria

The film made from the deposit to be stained withGram’s method

Make a thick smear with of area spread 10 mm indiameter encircle by a scratch on the surface of theslide

If the CSF appears turbid make a thin film All the smears are dried and fixed on heat

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Gram Staining technique35



Gram Staining Procedure

36



Examination of Gram Stained smear

A careful search for

Bacteria to be made inparticular where there

are plenty of leucocytes

At least keen observationto be done for 10 mt

before reporting a

negative smear.

37



Observe for the Presence ofOne should be familiar with the

following bacteria for successful

reportingMeningococci

Pneumococci

Haemophilus

Coli form bacilliStreptococci

Listeria

All the results are promptly reported totreating Physician

When variety of bacteria are foundspecimens may be contaminated.

May need a fresh specimen forexamination

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Culturing of CSF The deposited sediment

plated on culture media

Blood agar,Chocolate agar

incubated with 5-10%Carbon dioxide

A part of the specimeninoculated into Robertson'scooked medium

In suspected cases of Brainabscess Bacteroides and

anaerobic cocci arecultured in anaerobicmedium

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Direct antibiotic sensitivity detection

When the organisms arenumerous on Gram stainedfilm CSF can be directlyinoculated into Blood agarand Chocolate agar

The commonly used effectiveantibiotic disks are tested

with sensitivity pattern, Commonly we can test BenzylPenicillin, andChloramphenicol

The antibiotic sensitivity

pattern can be reported atthe earliest

40



Biochemical testing for Infections CSF should be tested for quantization

of

Glucose and ProteinNormal CSF contain

2.2 to 4mmol/liter correlates to 60%of the plasma levels

Protein is present at concentration of

0.15 to 0.4 grams/literIt can be higher in neonates can be upto 1.5 grams / liter

In pyogenic meningitis Proteinconcentration is increased and

Glucose concentration decreased.In aseptic meningitis Glucoseconcentration is normal and proteinconcentration raised

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Tests for Bacterial antigen DetectionCo agglutination Tests

There are several testkits availablecommercially fordetection antigens of

MeningococciPneumococci

H influenzae

42



Diagnosis of Viral Meningitis

The virus are to be

isolated from CSF Presence of Viral

antibodies by pairedsampling of serum

In few viral infections thevirus can be isolated from

Throat swabsSpecimens of feces

43



Tuberculosis Meningitis -Diagnosis

CSF should betested for presenceof Acid fast bacilliby simple ZiehlNeelsen method

The deposit of theconcentrate can beinoculated ontoLowensteinJensen’s Medium

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Ziehl- Neelsen ProcedureMake a smear. Air Dry. Heat Fix.

2. Flood smear with Carbol Fuchsin stain Carbol Fuchsin is a lipid soluble, phenolic compound, which is able to penetrate

the cell wall

3. Cover flooded smear with filter paper4. Steam for 10 minutes. Add more Carbol Fuchsin stain as needed5. Cool slide6. Rinse with DI water7. Flood slide with acid alcohol (leave 15 seconds). The acid alcohol

contains 3% HCl and 95% ethanol, or you

can declorase with 20% H2 S04 The waxy cell wall then prevents the stain from being removed by the acid alcohol

(decolorizer) once it has penetrated the cell wall. The acid alcohol decolorizer willremove the stain from all other cells.

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Ziehl- Neelsen Procedure (continued)47

8. Tilt slide 45 degrees over the sink and add acid alcoholdrop wise (drop by drop) until the red color stopsstreaming from the smear

9. Rinse with DI water

10. Add Loeffler’s Methylene Blue stain (counter stain). Thisstain adds blue color to non-acid fast cells!! LeaveLoeffler’s Blue stain on smear for 1 minute

11. Rinse slide. Blot dry.

12. Use oil immersion objective to view.

1 2 3



Ziehl-Neelsenstain

4 5 6

7

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Viruses - Meningitis

The following viruses cancause Aseptic meningitis

1 Echovirus

2 Coxsackie3 Herpes virus

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Tele contact is crucial in serious patients

When the patient is

serious, write a

Tele contact

number whichcan help in prompt

delivery of results

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The Programme is created by Dr.T.V.Rao

MD for ‘e’ learning for Young ResidentDoctors in the Developing World

[email protected]

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DIAGNOSIS OF MENINGITIS SKILL BASED DIAGNOSIS OF MENINGITIS