chronic meningitis is a potentially treatable condition if diagnosed early....

1. CHAIR PERSONS : Dr . Nagarathna . S
Additional professor of Neuro
Micro Biology
Dr . Netravathi . M
Associate professor of Neurology
PRESENTER : Dr .Ashok Vardhan
Reddy. T
LABORATORY DIAGNOSIS OF
CHRONIC
MENINGITIS

2. What is Chronic Meningitis
Case vignettes
Clinical scenario
Possibilities
Basic Investigation workup
Detailed investigations of common diseases
Conclusion
OUTLINE

3. What is Chronic meningitis ?
 Chronic meningitis is defined as predominantly
leptomeningeal disease with clinical and CSF
inflammatory changes persisting for 4 weeks or
longer.
 Notifiable disease in many countries.
 Multiple etiologies.
 Even with detailed evaluation, 1/3rd of the cases
remain inconclusive. Jerzy Hilderbrand & Marc Hilderbrand : Infections of
Central Nervous system .
Mandell, Douglas and Bennet: Principles and practice of
infectious diseases,8th edition

4. Case vignettes

5. Case 1
 48 year old gentleman
 Vomiting and headache -1 month
 Diplopia, drooping of both eyelids -1 week
 On examination-
Bilateral ptosis present
Extra ocular movement- full

8. Blood investigations
 Hb-12g/dl
 Platelet count-4 lakh /mm3
 WBC -8000/mcl
 TSH -0.3 mc Iu/ml
 Vit.B12 -275pg/ml
 LFT ,RFT ,Serum electrolytes- normal
 ANA Profile, ANCA –negative.
 ESR -23 mm after1 hr

9. CSF
 Appearance-clear
 Cells- 45
 100 % lymphocytes
 India ink–Negative
 Cytospin- Small and large lymphocytes,
reactive monocytoid cells and, polymorphs, in
the background of few RBCs and several
degenerated cells, Suggestive of chronic
inflammation.
 VDRL –Non reactive
 Anti Mycobacterial antibodies by ELISA-Positive
 Culture for Mycobacteria -Negative
TB MENINGITIS

10. Case 2
 38 yr old gentleman presented with c/o fever,
headache, vomiting for 3 weeks.
 Recently diagnosed as HIV positive.
 On examination :
Visual acuity 6/12 both eyes
Papilledema present- Bilateral
No other focal deficits

13. Investigations
 Hb-12.2 g/dl
 TC-8000/mm3
 Platelet count -1.36L/mcl
 RBC count -5 Millions/mm3
 LFT ,RFT ,Serum electrolytes- Normal
 Blood culture, stool culture-Normal commensals

14. CSF analysis
 Appearance–Clear
 Nil cells
 Budding yeast like cells present on India Ink
staining

15.  Cryptococcal capsular polysaccharide
antigen test by latex agglutination –Positive
 CSF Culture- Cryptococcus.neoformans
 ELISA for anti Cysticercal and
anti Mycobacterial antibodies-Negative
CRYPTOCOCCAL MENINGITIS

16. Case 3
 26 yr old gentleman presented with tingling and
numbness of
Both lower limbs for 6 months
Upper limbs for 5 months
Imbalance while walking for 3 months
Urinary and fecal incontinence for 1 month

17. On Examination
 Cranial nerves-Normal
 Power -5/5 in all 4 limbs
 Sensory –Touch ,pain decreased by 50 % in both
lower limbs below knees .
 Plantar reflex –Bilateral extensor
 DTR s Brisk in lower limbs
2 + in upper limbs
 Gait - Spastic ataxic

20. Blood Investigations
 CBC,LFT , RFT, Serum electrolytes –Normal .
 HIV –Non reactive
 RA,ANA –Negative
 Serum VDRL-Non reactive
 Serum ACE levels -35.3 u/l

21. CSF
 Clear
 20 cells
 Lymphocytes -20
 Protein-45mg/dl
 Sugar-126mg/dl; Chloride-130 mEq/l
 Cytospin -Lymphocytes and degenerated cells.
No abnormal cells
 India ink-Negative
 Cryptococcal antigen test-Negative
 Anti -Mycobacterial antibodies-Negative

22. Serum agglutination test for Brucella abortus -
1:320
Serum agglutination test for Brucella melitensis-
Negative
Blood culture for Brucella- Positive
PCR -Positive

23. Case 4
 51 yr old agriculturist
 Known HIV positive for last 1 yr. Took ART for 6
months and then stopped.
 H/O fever and loose stools for last 1 month.
 For past 15 days he developed involuntary
movements of left lower limb and occasional
movement of upper limb. Movements are
predominantly proximal and disappear during
sleep.

24.  On Examination-
Involuntary movements of left lower limb-
Flexion,
extension at hip and knee, rotation at ankle.
Power in left lower limb –Cannot be tested
Other limbs -5/5.

27. Investigations
 Hb- 9.2g/dl
 TC- 6000/mcl
 RBS- 86mg/dl
 Serum lipid profile- normal
 LFT ,RFT, Serum electrolytes- normal
 HIV-Positive
 VDRL –Non reactive

28. CSF
 Clear
 4 cells
 All lymphocytes
 Proteins-46mg/dl
 Sugar -40mg/dl
 India ink –Negative
 Mycobacteria- culture, ELISA- Negative
 VDRL –Negative
 No bacterial growth on culture

29.  CSF-Anti toxoplasma antibody –negative
 Serum anti Toxoplasma antibody (1:32)-
positive
TOXOPLASMA MENINGITIS

30. Case 5
 44yr old gentleman
 Intermittent ,mild headache for past 5 yrs
 Behavioral disturbances -3 yrs
 Speech disturbance- 2 yrs
 Memory disturbance -1 yr
 On examination-
 MMSE- 20/30
 Motor system, sensory system – normal

31.  zz

32. Investigations
 Blood investigations-normal
 CSF-
Clear
6 cells -5 neutrophils ,1 lymphocyte
India ink –Negative
CSF- Chloride/glucose/protein-
124mEq/l /67mg/dl /75mg/dl
Culture for Mycobacteria and Fungus-
Negative

33. Serum VDRL-Reactive
CSF VDRL-Reactive
NEURO SYPHILIS

34. Case 6
 40 yr old lady
 Headache for 3 months
 Diplopia on looking towards right ,drooping of
right eye lid for 20 days.
 On examination –
Right eye lid ptosis
Right lateral rectus palsy
Motor ,sensory system
normal

36. Investigations
 Hb-14g/dl
 RBC count – 5 millions/mm3
 WBC count- 7,000/ mcl
 Platelets 2.1 lakh / mcl
 LFT ,RFT ,Serum electrolytes-normal
 Serum VDRL-Negative

38. CSF
 Opening pressure -14 cm H2O
 Appearance-clear
 Cell count-3/mm³
 Differential count –All lymphocytes
 Glucose- 68mg/dl, protein-25mg/dl, chloride-
123mEq/l
 India ink -Negative
 Cryptococcal antigen test – Negative
 Anti -Mycobacterial anti bodies- Negative
 VDRL – Negative

39. Serum ACE -124.6 u/l

40. When to suspect Chronic meningitis
• Most common presentation –Headache,
insidious
onset increasing over 4 weeks.
• Associated symptoms -Fever, vomiting, abnormal
mental
state.
• Visual symptoms -Diplopia, blurring of vision.
• Seizures.
• Signs-Normal sensorium, coma, nuchal rigidity,
papilledema, multiple cranial nerve palsies,
cerebrovascular accident .
• Weakness of limbs, back ache, burning
paraesthesias occur if spinal nerve roots areJoseph R . Zunt et al .Review article.Continuum.,2012,American Academy of Neurology; Karen L Ross, Allan R Tunkel : Hand

41. Causes of Chronic meningitis
Jerzy Hilderbrand &Marc Hilderbrand , Scheld, Whitley , Marra : Infections of
INFECTIVE NON INFECTIVE
1. Bacterial
2. Viral
3. Fungal
4. Parasitic
5. Endocarditis
6. Para meningeal
Infection
1. Uveo meningitic
syndromes
2. Hypertrophic
patchy
meningitis
3. Drugs
4. Malignant
meningitis
5. Miscellaneous

42. INFECTIVE
BACTERIAL
Partially treated
pyogenic
meningitis
Tuberculosis
Syphilis
Lyme disease
Others –
Leptospira
Brucellosis
Listeriosis
Mycoplasma
pneumoniae
Nocardia
Actinomyces
VIRAL
MENIN
GITIS
HIV
Ebstein
Barr virus
Echo
virus
FUNGAL
MENINGITI
S
Cryptococco
sis
Blastomycosi
s
Histoplasmosi
s
Coccidiomyco
sis
Aspergillosis
Mucor
mycosis
PARASITE
S
Toxoplasm
a
Acanthame
ba
Angiostron
gylus
cantonensi
s
Cysticercos
is
OTHERS:
Infective
endocarditis
Para
meningeal
infection due
to epidural
abscess
sinusitis
Mandell, Douglas and Bennett’s Principles and practice of Infectious D
edition a

43. Non Infective causes
UVEO
MENINGITIC
SYNDROMES
Sarcoidosis
Behcet’s disease
Wegener”s
granulomatosis
Vogt –Koyanagi
Harada
syndrome
Sjogren ‘s
syndrome
SLE
Other
vasculitidess
DRUGS
Non steroidal anti
inflammatory
drugs
Anti microbial
agents
I V Immuno
globulins
Immuno
suppressants
Allopurinol
Vaccination
Intrathecal agents
MALIGNAN
T
MENINGITI
S
Carcinoma
Lymphoma
Leukemia
MISCELLAN
EOUS
Hereditary
auto
inflammatory
periodic fever
syndromes
Cholesterol
emboliation
syndrome
Fabry’s
disease
Subarachnoid
hemorrhage
Migraine
IDIOPA
THIC
Hypertr
ophic
pachy
meningi
tis
Lionel Ginsberg ,Desmond Kidd :Review of chronic and recurrent meningitis. Pra

44. Investigations
Imaging –CT ,MRI Brain
CSF Analysis
• Ancillary tests -Blood investigations, Chest X ray
USG abdomen, Mantoux test, Meningeal biopsy.
Lionel Ginsberg ,Desmond Kidd :Review of chronic and recurrent meningitis. Pract Neu

45.  Non enhanced CT scan-Normal in >50 % of
patients.
 Hyper dense lepto meninges with diffuse/nodular
Contrast enhancement.
 Hyper densities around basal cisterns.
 Hydrocephalus.
 Diagnosis of complications- Subdural effusion ,
Epidural empyema, Dural sinus thrombosis,
Infarcts, Cerebritis, cerebral abscess.
 CT Angiogram – Vasospasm / infarction
CT Brain
Karen l ross,Allan R Tunkel Hand book of clinical Neurology
.Imaging in CNS Infections.

46.  .

49. MRI
 Exudate appears isointense in T1 sequence.
 Obliterated cisterns, distension of sub
arachnoid space with widening of
interhemispheric fissure.
 T1 contrast (Gd): Leptomeningeal thickening and
enhancement is highly sensitive for meningitis
(sensitivity>90%).
 Cranial nerve enhancement in subarachnoid
cisterns.
 Intra dural extra medullary enhancing nodules.
Black DF et al :MR Imaging of Central Nervous System Infections .AJNR Am J
Neuroradiol.31 (8):1493-7,2010.

51. MRI
FLAIR sequences:
 Pre contrast FLAIR can be normal.
 FLAIR images are more sensitive compared
to post contrast T1WI for leptomeningeal
enhancement.
 T1WI are more specific compared to FLAIR
for parenchymal enhancement.
 MR Angiography- Arterial narrowing and
occlusion.
. Black DF et al :MR Imaging of Central Nervous System Infections .AJNR Am J Neuroradi

52. T1 T2 FLAIR CONTRAST MP
RAGE

53. Clues for Diagnosis
 Basal cisterns enhancement
 Dura and arachnoid enhancement
 Infarctions especially in basal ganglia and internal capsule
 Progressive hydrocephalus
 Involvement of cranial nerves (Commonly II, III, IV, VII)
- Highly suggestive of
TUBERCULOUS etiology.
 Hyper intense meninges on pre contrast FLAIR is the most specific sign
 Triad of hydrocephalus, infarction, non contrast basal
enhancement are 89% sensitive and 100 % specific for TBM.
Leiguarda et al 1988 , Wilson & Castillo 199 9, Morgado & Ruvio 2005,Bernaerts A et al : Tuberculosis of Central Nervous

56. Clues for Diagnosis
 Dilated perivascular spaces
 Basal ganglion pseudo cysts with soap bubble
appearance
 T1WI & FLAIR –Hypo intensity, not enhancing on
contrast
 T2WI hyper intensity
 Dense enhancement of choroid plexus in lateral and
4th ventricle.
-suggestive of Cryptococcal
meningitis.
 Normal brain imaging - 44% of HIV positive cases
versus 13% of non HIV infected cases(Lee et al 2011)Eric F Greif et al 2011.Luthra G et al :Comparative evaluation of fungal , tubercular, pyogenic infections with MR imaging . AJNR Am J

58.  Non contrast CT - Multiple hypodense lesions in
basal ganglia, thalami and cortico medullary
junction.
 T1Hypo intense, smooth ring enhancement to
contrast.
 FLAIR Images-Target sign
 T2 weighted imaging- Hypo intense core- Hyper
intense intermediate zone -Hypo intense rim.
 Pathognomonic -Asymmetric target sign.
 Sensitivity -30 %
 Specificity >90%
-Cerebral
toxoplasmosis
Clues for Diagnosis
Ashley et al .Central nervous

60. Clues for Diagnosis
 Multiple small (2-8 mm) sub cortical and peri
ventricular white matter hyper intensities on
T2 FLAIR images predominant in frontal area.
 Uniform enhancement on T1 contrast around
Facial nerve.
 Multi-focal hyper intense lesions on T2WI with
patchy cord and linear nerve root enhancement .
-Lyme Neuro
Borreliosis
 The reported rate of abnormal MRI findings in
LNB varies from 17% to 43%.Hildenbrand P et al :Lyme Neuro Borreliosis :manifestations of rapidly emerging zoonosis AJNR Am J
87, 2009
Ashley et al Review article –CNS infections

62. Clues for Diagnosis
 Diffuse/Nodular leptomeningeal
enhancement-40%
(Predominant supra sellar and frontal basal meninges)
 Dural enhancement
-30%
 Enhancement in hypothalamus/pituitary stalk
-18%
 Cranial nerve thickening and
enhancement(VII &II are commonly involved )
-NeuroLury KM et al :Neuro Sarcoidosis –Review of imaging findings .Semin
Roentgenol.39(4).495-504,2004
Zunt et al :chronic and sub acute meningitis.continuum .Review

64. Clues for Diagnosis
 Lesions in :
Meso diencephalic junction and
ponto bulbar
area
Bilateral thalamus and basal ganglia
Cerebellar peduncles
Internal capsule
 T1 Hypo intense,T2 Hyper intense with patchy
contrast enhancement . DWI –Iso to hyper intense
lesions.
-Neuro Behcet’s
disease
Naci Kocer et al .CNS Involvement in Neuro Behcet
syndrome :An MR study . AJNR.2001

66. Blood tests
 Complete blood count.
 ESR, C-reactive protein.
 Blood cultures(Bacterial meningitis, Brucellosis,
Cryptococcosis), urine (leptospirosis, Cryptococcosis).
 Serology-HIV, Syphilis, Borreliosis, Brucellosis.
 Auto antibodies: Anti nuclear factor, rheumatoid
factor, extractable nuclear antigens, ANCA
(vasculitis).
 LFT , RFT, Serum electrolytes (TB Meningitis,
Leptospirosis)
 ACE levels(sarcoidosis).

67. CSF Analysis
 Indications, contra indications
 Collection, transport and storage
 Macroscopic examination
 Biochemistry -protein ,sugar, chloride.
 Cytological examination
 Chronic meningitis workup

68. CSF
 Indications-
Any suspected case of acute or chronic
meningitis
vasculitis, demyelinating diseases.
Therapeutic- Idiopathic intra cranial
hypertension
 Contra indications -
Midline shift
Coagulopathy, platelet count <50
000/mcl,
INR>1.4
Intra cranial space occupying lesion

69. AMOUNT
Fishman , Greenlee ,Carroll 1997.
TEST CSF Required
Cell count &differential 0.5-1 ml
Glucose &protein 0.5 ml
Bacterial culture 3ml
Mycobacterial &fungal culture, smear for
AFB, India ink
12 ml
PCR for mycobacterium 0.1-1ml
Viral culture &PCR 1-2 ml
Cryptococcal antigen 0.5 ml
VDRL 0.5 ml
Oligo clonal bands 2 ml
Cytospin 0.5ml

70. CSF Amount
 10-15 ml is required for culture of mycobacteria,
fungus, parasite.( Tenny et al .,1982 as cited in Gray &Fedorko,1992)
 Specimen must be stored at 37ºC until
microscopy and bacterial cultures are
performed(Kaste.,1990 cited in Gray &Federko ,1992.WHO recommendations)
 Sample must be analyzed with in 1 hr. If delay is
expected, one bottle should contain plain CSF for
making smears to identify organism. The other bottle
is transported after adding few drops of glucose
broth and used for culture of organism(EFNS Task force
recommendations)
EFNS Task force ,F Deisenhammer et al ,European jo
Neurology ,2006

71. APPEARANCE

72. Normal
Viral meningitis
Cryptococcal meningitis
TB meningitis
Clear

76. Biochemistry
 CSF glucose : 45-60 mg/dl
 <45 mg/dl is considered as low
 Decrease in glucose-Bacterial
>Tubercular>Fungal
 Normal in aseptic meningitis.
 CSF/serum glucose <0.4 is helpful in
distinguishing between bacterial and aseptic
meningitis with sensitivity and specificity of 91 %
and 96%.(Genton & Berger 1990).
Bonadio et al ,1992,lab diagnosis of chronic meningitis. R
article

77. Biochemistry
 Protein levels 15-45 mg/dl
 >55 mg/dl -Abnormal
 Bacterial, fungal ,tubercular(100 -500 mg/dl)
meningitis.
Normal
mg/dl
Acute
Bacterial
Tubercular Fungal Viral syphilis
15-45 200-300 100-300 50-100 50-100 50-100
Bonadio et al ,1992,lab diagnosis of chronic meningitis. Review article
Nather C Bahr et al :Methods of rapid diagnosis fr the etiology of meningitis in adults.Review article.Biomarkers in medicine.

78. CSF Chloride (116 - 127mEq/l)
 Chloride decreases in both pyogenic and tubercular
meningitis. More so in tubercular.
Chloride Content of the Cerebrospinal Fluid by H. W. GIERSON, M.D., and G. J. OWENS. M.D., Los Angeles.California
medicine.1961
CSF Chloride
mEq/l
PYOGENIC MENINGITIS TUBERCULAR
MENINGITIS
<113 19% 60%
<108 7% 41%

79. Cell count and differential
count

80. CELL COUNT
CELL COUNT CAUSE EXCEPTIONS
1000-2000 ↑N Bacterial Meningitis 10% -↑L
0-1000 (223) ↑L Tubercular Meningitis 20%-↑N
<20 in 52%; >100 in 20 % Fungal meningitis
10-100 ↑L Viral meningitis
10-50 ↑E Parasitic meningitis
S . Nagarathna , H. B. Veenakumari , A . Chandramuki ,NIMHANS.Laboratory diagnosi
Meningitis. 2012
TRUE WBC in CSF= WBC in CSF ⼀WBC in Blood Ⅹ RBC in CSF/RBC in Blood

81. Chronic meningitis workup
 India ink
 Cryptococcal polysaccharide antigen test
 ELISA for anti mycobacterial and anti Cysticercal
antibodies
 AFB Culture
 CSF VDRL

82. Detailed investigations of common
diseases

83. Partially treated pyogenic
meningitis
 Clinical profile:
High fever, signs of meningeal irritation with
history of receiving antibiotics recently, undergone
cranial surgery, head injury.
 CSF –Increased proteins, decreased glucose,
CSF/Blood glucose< 0.2 and / or Pleocytosis
with predominance of PMN cells.

84. Differentiating TBM form PTPM

85. TB MENINGITIS
CSF PARAMETER MOST COMMON EXCEPTIONS
CELL COUNT 0-1000/mcl, L↑ Normal -6-10%
N↑20-27%
GLUCOSE <45 mg/dl Normal-27-30%
Immuno Compromsed
State-
29-31%
MEAN PROTEIN 224 mg/dl Normal -6-10%
Immuno Compromised
State-
40-45%
Verdon et al 1996,Gracia –Monoco et al 199

86. CSF Ziehl –Neelsen staining
 3-4 serial samples of 10 -15ml and spinning for
30 minutes is required.
 Positive in 25 % of adults and 3 % children with
TBM.
 Modification by treating with Triton prior to ZN
staining increases the yield.
Davis 1993,Hooker 2003,Chrutensen 2011;EFNS Task force
Recommendations2011

87. CSF culture for Mycobacteria
 Lowenstein Jensen Medium
 BACTEC 460 TB System
 MGIT 320 /640/960

88. Lowenstein Jensen Medium
 Time consuming method
 10-15 ml of CSF inoculated in to the medium
 Kept at 37 ºC plus 5% CO2 for 8 weeks
 Reading taken once in a week
 Positivity rate -40-85 % in adults
35-85 % in children
Gracia Monco et al ,1999.

89. BACTEC 460 TB
 Automated Radiometric assay
 BACTEC12 B medium with PANTA and C14 are
used
 Drug susceptibility test done with in 10-14 days
 Total 6 weeks required to declare negative growth
 Advantages-Speed of the results –faster than LJ
medium
Higher recovery rate
Drug sensitivity testing
 Disadvantages-
Radio labelled products are used
High cost

90. BACTEC
Takes less time, Completes DST.
Almost 90% pick up rate if combined with conventional culture.

91. MGIT
 Modified middle Brooks 7H9 broth with
fluorescent growth indicator embedded in silicon
at the bottom of tube.
 Low oxygen content stimulates the fluorescence
 PANTA –Polymyxin B, Azlocillin, Nalidixic acid,
Trimethoprim, Amphotericin B.
 Para Nitro Benzoic acid test is done to
differentiate Mycobacteria from Non
tuberculous(Resistant ) organisms.
Rajeev Thakur et al ,Dept.of Micro
Biology ,IHBAS,India

92. .

93. MGIT

94. Study conducted in a tertiary care hospital in India comparing various
diagnostic tests for TBM .
Showing 27 .4 % culture positivity by MGIT.
4 samples yielded positive only in LJ medium probably due to some
growth factors in egg based media.
So it is recommended to use conventional solid media along with MGIT .
Samples (n = 164) CSF AFB LJ MEDIUM MGIT
7.9 % 10.9% 27.4
Rajeev thakur et al ,IHBAS ,Newdelhi .2010 ,Journal of La

95. BACTEC 460TB vs MGIT 960.
95
•The average time required for
completion of the test was 2.5 days
shorter with MGIT
•BACTEC MGIT 960 - suitable
replacement for the radiometric method
of antimicrobial susceptibility testing of
M. tuberculosis.

96. CSF ELISA
 Total anti Mycobacterial antibodies –MTSE
(Mycobacterium tuberculosis Soluble Extract )
 Specific antibody to a defined antigen –LAM
,38KDa,A60, 30KDa,1,6 k Da.
 Secretory Mycobacterial antigens –ESAT 6,MPT
63,MPT 64,MPT 70 ,14 kDa,19
kDa,38kDa,85B,ORF9,ORF10 a , culture filtrate
antigens.
 Sensitivity and specificity varies from 70- 87 %.
Akepati 1986,chandramukhi 1989,akepati et al 2002

97. Role of NAA in Tubercular
meningitis
 Sensitivity - 50% and a specificity - 100%
 Retrospective study from India(MP)found a LAMP(Loop
mediated isothermal Amplification assay) assay
performed on CSF was 88% sensitive and 80% specific
for TBM diagnosis
 The sensitivity of CSF microscopy and culture falls
rapidly after the start of treatment
Thwaites GE, Caws M, Chau TT, Dung NT, Campbell JI, Phu NH et al. 42(3):996 e 1002.

98. Role of NAA in Tubercular
meningitis
 Mycobacterial DNA may remain detectable within the
CSF until 1
month after the start of treatment.
 The Xpert MTB/RIF assay performed in a single
cartridge, provides
concurrent detection of M. tuberculosis and the genetic
mutations
which detect rifampicin resistance within around 120
min.
Thwaites GE, Caws M, Chau TT, Dung NT, Campbell JI, Phu NH et al. Clin Microbiol 2004;
42(3):996 e 1002.

99.  Pulmonary kochs -Sensitivity of nearly 100% for
smear-positive sputum and around 70% for smear-
negative and specificity of 100%

Thwaites GE, Caws M, Chau TT, Dung NT, Campbell JI, Phu NH et al. Clin MicroBiology 42(3):996 e 1002.

100. Recommendations
NAA in CNS tuberculosis
 A commercial NAA assay to be performed on
CSF for all forms of suspected CNS tuberculosis.
 Negative test does not rule out tuberculosis.
 NAA tests are more useful than conventional
bacteriology after the start of anti-tuberculosis
treatment.
 NAA assays that detect the rifampicin
resistance genotypes should be requested when
the risk of drug resistant tuberculosis is high.
Thwaites GE, Caws M, Chau TT, Dung NT, Campbell JI, Phu NH et al. 42(3):99

101. Tuberculous meningitis
Alternative cerebrospinal fluid diagnostic markers
 A real-time quantitative (q) PCR and an ELISA to detect a
panel of M. tuberculosis antigens (GlcB, HspX, MPT51,
Ag85B and PstS1)
 The sensitivity and specificity of both assays
approached 100% in the 29 patients with culture-confirmed
TBM
 In probable /possible TBM, with a sensitivity of 98% and
specificity of 98% for qPCR, and 92–95% sensitivity and 93–
96% specificity for GlcB, HspX and MPT51 antigen ELISAs.

102. At NIMHANS

103. Tuberculous Meningitis: Tuberculin test
 10 -20% of patients with CNS tuberculosis have a
positive test.
 Rates for children vary between 30 - 65%
 Varies with age, vaccination with BCG, nutritional
status, HIV infection, and technique of administration.
Thwaites GE, Caws M, Chau TT, Dung NT, Campbell JI, Phu NH et al. Clin Microbiol 2004;

104. Tuberculous meningitis: Adenosine
deaminase
 Adenosine de Aminase (ADA) is raised in the
CSF of patients with TBM.
 High CSF ADA activity -Lymphomas,
Malaria, Brucellosis and pyogenic
meningitis.
Thwaites GE, Caws M, Chau TT, Dung NT, Campbell JI, Phu NH et al. Clin Microbiol
2004; 42(3):996 e 1002.

106. Tuberculous meningitis
Interferon Gamma Release Assay
 QuantiFERON-TB gold and T-SPOT.TB
 Better than Tuberculin test for diagnosing latent
tuberculosis.
 Sensitivity - 58% and specificity - 94% which
adds to the diagnostic yield
 CSF IGRA has major limitations.
 1)Relatively large CSF volumes (5–10 ml) are required
 2) Indeterminate results are common (10–80%
reported).
 3)The test lacks diagnostic sensitivity.
 4) CSF lymphocytes die rapidly when stimulated with M.
tuberculosis specific antigens ex vivo and the test fails.Vidhate MR, Singh MK, Garg RK, et al. Diagnostic and prognostic value of Mycobacterium tuberculosis complex specific interferon gamma
release assay in patients with tuberculous meningitis. J Infect 2011; 62:400–403.

108. Recommendations
TST,ADA,IGRA
 CSF adenosine deaminase activity not
recommended as a routine diagnostic test for
CNS tuberculosis .
 TST &IGRAs are only licensed for the
diagnosis of latent tuberculosis and cannot
be recommended for the diagnosis of active
CNS disease .

109. Diagnosis of TB Meningitis
TEST SENSITIVITY (%) SPECIFICITY(%)
ZN Staining 3-25
LJ Medium 30-40
MGIT 35-50
ELISA (Anti Mycobacterial
antibodies)
70-87 >80
LAMP Assay 88 80
Gene
Xpert/MTB/RIF(Sputum)
>95 >95
Quantiferon TB-Gold 60 94
RT PCR 100 >95

110. Cryptococcal meningitis
 There are about 50 species of Cryptococcus
 All are Non fermentors , assimilate inositol ,
produce urease.
 C.neoformans is pathogenic to humans .
 Rarely C.laurentii , C.albidus are pathogenic (Horowit
et al 1993 )
 Specific type of Glucuron xylo mannan is
present in the capsule.
 Inability to grow at 37ºcelsius indicates non
pathogenic species.

111.  POOR PROGNOSTIC factors :
Low glucose levels
Antigenic titre >1:1024
High CSF Lactate
Hydrocephalus
High opening pressure >
30cm H20
Altered sensorium
Joseph R Zunt et al ,Continuum ,American academy of Ne

112. India ink test

113. Cryptococcal antigen test

114. SDA showing glistening mucoid colonies of
Cryptococcus

115. Other tests for Cryptococcus
Urease positive Gram positive

116. Species IdentificationC.neoformans in Caffeic acid
ferric
citrate test agar
C.gattii in canavanine glycine
Bromo thymol blue

117. Sl.
No
INDIA INK
Sensitivity/Sp
ecificity
CRYPTOCOCCAL
ANTIGEN (LAT)
Sensitivity/Specifi
city
CULTURE
Sensitivity/Specif
icity
Authors
1 >90 % in HIV
positive ,>50%
in HIV negative
>90%/>90% 100 % Satish chandra et al
Dept.of Neurology
NIMHANS ,Bangalore
2007
2 ------------ 100 %/100% 90%/90% Kelly et al ,Dept .of
Neurology ,Geisinger
Medical Center ,USA
2014
3 50 % /100 % 100%/100% 80% RMS Dominic et al
,KMC Mangalore
2010
4 93.5/100% 100%/86.5% 95.7%/100% DC Saha et al ,Dept.of
Micro Biology ,AIIMS
2009

118. M kingston et al UK National guidelines on management of Syphilis ,international journal of STD &AIDS
,vol 19,2008
CLINICAL
SYNDROME
TIMING AFTER
INFECTION
SIGNS & SYMPTOMS
Asymptomatic Early Abnormal CSF can be present in 30 %
of primary and secondary syphilis.
Does not progress in majority of
patients.
Meningovascular 2-7 yrs Focal arteritis, Meningitis, emotional
lability, insomnia.
General paresis 10-20 yrs Decline in memory ,emotional lability,
psychosis, dementia.
Tabes dorsalis 15-25yrs Paraesthesia, sensory ataxia,
Charcot’s joints, optic atrophy, Argyll
Robertson pupil.
Cardiovascular 10-30 yrs Aortitis, heart failure, aneurysms.
Gummatous 1-46 yrs (average -
15 yrs)
Destructive Gummas most commonly
over bones and skin
Neuro syphilis

119. CSF Serology
 Indications :
-Any stage of disease with clinical evidence
of
neurological, ocular involvement .
-Tertiary syphilis (Gummatous,
Cardiovascular )
 Should not be macroscopically/microscopically
contaminated with blood (gives false positive result )
 WBC Count >5/mm3 with lymphocytic predominance
 Protein levels are usually normal
 Tests
1. VDRL
2. TPHA
3. TPHA IndexM Kingston et al UK National guidelines on management of Syphilis 2008 .international journal

120. CSF VDRL
 Quantitative VDRL is useful
 Sensitivity -50 % ( 10 % for asymptomatic cases
to 90 % for symptomatic cases).
 CSF and serum VDRL are performed
simultaneously .

121.  Serum VDRL titer of 1:8 is considered as positive
M Janier et al :2014 European guidelines on management of Syphilis ,Southampton ,UK.European centre for
disease prevention and control.
False negative False positive (0.2-0.8%)
Early syphilis Acute Chronic
Secondary syphilis Post Immunisation Auto immune diseases
Recent MI Injection drug users
Infections (Malaria, Hepatitis,
Chicken pox, measles)
HIV, Leprosy, Malignancy.
Pregnancy

122. Negative VDRL in serum -100 %sensitivity in excluding CSF
abnormalities.
VDRL titer >1:32 is associated with high chances of CSF
positivity
M Kingston et al UK National guidelines on management of Syphilis 2008 .international journal o

123. TPHA-Treponema pallidum
Haemagglutination Assay
 Highly sensitive (>95%)
 Lacks specificity (can be positive due to transfer of
immuno globulins from serum to CSF if blood brain
barrier is breached).
 Significant titer is >1:320
 TPHA Index = CSF TPHA /Albumin quotient.
{Albumin Quotient =CSF Albumin/serum
albuminⅩ1000}
 Highly sensitive and specific (>95%)
 TPHA index>70 TPHA titer >320 are most reliable in
supporting the diagnosis of Neuro syphilis
M Kingston et al : UK National guidelines on management of syphilis 2008 ,International

124. Neuro Brucellosis
 Neurologic involvement occurs in approximately
10% of cases.
 Categories:
1. Acute meningitis or meningoencephalitis.
2. chronic peripheral form (Radiculo
neuropathy),
and chronic central nervous system infection
(Meningoencephalitis, Myelitis, Cerebellar
involvement,
Cranial nerve palsies).

125. Clinical profile (According to a multinational study on
Neuro brucellosis)
 Headache >4 weeks occurs in >50 % of cases.
 Non specific symptoms- malaise, sweating
,weight loss in >50 % cases.
 Cranial nerve involvement, mostly affecting the
sixth
and eighth cranial nerves -20%
 Poly neuropathy, radiculopathy, paraplegia -10%
 Brain abscess- 4%
 Associated with hepatomegaly, splenomegaly in
30 %cases.
.
 Mandell,Douglas,Bennet’s principles and practice of infectious diseases,8

126. Neuro Brucellosis
Clinical profile :
 Chronic meningitis -33.3 %
 Infective CVT- 25 %
 Demyelination -16.6%
 Myelo radiculopathy -8.33%
Nagarathna S, Rajeshwari S, Veenakumari HB, Netravathi M, Padmashree BS, Sayani M, Triveni K, Satishchandra
P.NIMHANS.NeuroBrucellosis-a clinical masquerade in Indian scenario;Int J Bio Med Res.2015 ;6(3):5143-5147

127. Lab diagnosis
 Specimens: Blood, Bone marrow, CSF .
Tests- Rose Bengal Test
Tube Agglutination test
ELISA
PCR
Culture

128. Rose Bengal slide agglutination
test:
 Rapid screening test
 Antigen-Brucella abortus suspension at pH 3.6-
3.7
 Agglutination indicates concentration of
antibodies >25 IU/ml

129. Mohd Zakaria Hussein et al .Evaluation of Rose Bengal Test ,Standard Tube Agglutination test and Nested PCR for diagnosis of Human

130.  Sensitivity 57.5-68.6% in serum samples
 False negatives are common in early stages
of disease.
 It is the 1st screening investigation when we
suspect brucellosis.
Mohd Zakaria Hussein et al .Evaluation of Rose Bengal Test ,Standard Tube Agglutination test and Nested PCR for diagnosis

131. Standard tube agglutination
 Titer of 1/160 is considered as positive
 Superior sensitivity (62.5%) compared to
RBT
 Recognizes both Ig G and IgM
 False negative results occur in 1st 3 months of
illness.
Mohd Zakaria Hussein et al .Evaluation of Rose Bengal Test ,Standard Tube Agglutination test and Nested PCR for diagnosis of Human
Brucellosis. Egyptian journal of medical microbiology , April 2006

132. 
ELISA IgM anti-
Brucella
1/100 dilution -serum
1/5 dilution -CSF
Sensitivity & specificity for serum
>90%

133. PCR
 Target sequence 223 bp in a gene encoding a 31 kDa
immunogenic outer membrane protein (bcsp 31)
 Sensitivity 70%
 Advantages:
Rapid assay ,provides results in less than 24 hrs
Risk of infection to laboratory personnel is reduced
The sample can be stored at (-20º.C )until
processing
Can be used for evaluation of treatment regimen
 Disadvantages:
Costly procedure.
Highly equipped laboratory is needed.
Mohd Zakaria Hussein et al .Evaluation of Rose Bengal Test ,Standard Tube Agglutination test and Nested PCR for diagnosis
of Human Brucellosis. Egyptian journal of medical microbiology ,April 2006

134. Lane 1 and Lane 8 is 100bp ladder.
Lane 2 and Lane 7 is positive control
Lane 3, 4 and 5 are positive samples.
Lane 6 is negative control.

135. Culture
 Castaneda method
 1week
 As the duration of illness increases, chance of
isolation decreases
 Sensitivity : Bone marrow(85-90%) >Blood
culture(62-70%)
 Bone marrow, blood, lymph nodes, liver biopsy
increases the sensitivity .
 CSF Culture positivity in neuro brucellosis is
13 -20%.
P Yagupiny et al J cl Microbio .1999-
37(11);3437-3442

136. Toxoplasma meningitis
 When to suspect ?
 HIV patients with CD4 < 200 /mcl
 Clinical features:
Headache -83.3%
Altered sensorium-40%
Hemi paresis-37.5%
Fever -30.8%
Cranial nerve palsy-18%
Seizures-15 %
Ganiem et al (2013):Cerebral Toxoplasmosis mimicking subacute meningitis in HIV patients ;a coho
Indonesia.PLOS.Neglected tropical diseases

137. Lab Diagnosis
 CSF -Clear
 Lymphocytic / eosinophilic pleocytosis,
elevated protein
 Toxoplasma.gondii IgG on CSF and Sera :
sensitivity, specificity
>90%
 Sensitivity of PCR (B22/B23 primer set)-98%.
 Specificity -92-94%
C.Baliu et al :Toxoplasma encephalitis associated with meningitis .Review.Journal of infectious di
,Spain 2014

139. Acanthameba
 Primary amebic encephalitis (acute)
 Granulomatous Amoebic Encephalitis(chronic)
 Suppressed immune function.
 Symptoms:
Mental status abnormalities(86%)
Seizures (66%)
Fever, headache, and hemiparesis (53%)
Meningismus(40%)
Visual disturbances (26%)
Ataxia (20%)
.
Mandell, Douglas and Bennet’s principles and practice of infectious diseases,8th e

140. Blood Tests
 Serum
Indirect immuno fluorescense antibody titer –
1:256 to 1:1024 – Favors
diagnosis
<1:80 –Exposure
Bruno et al :Diagnosis of infections caused by pathogenic free living Amoebae .Review article.Hindawi Publish

141. CSF
 Elevated WBC count with lymphocytic
predominance.
 Elevated protein
 Slightly decreased glucose
 CSF Wet mount preparation
 Culture-Non nutrient Agar plates covered by
E.coli and Enterobacter aerogenes
 PCR for Acanthameba 18S ribosomal RNA gene
Bruno et al :Diagnosis of infections caused by pathogenic free living Amoebae .Review article.Hindawi Publishin

142. Aspergillus -Meningitis
 Cerebral aspergillosis constitutes 10-20% of
invasive disease.
 Aspergillus fumigatus -90% cases.
 Chronic basilar meningitis
 Angio Invasion
 Cerebral vasculitis
 Granulomatous inflammation

143. Diagnosis
 CT –Head ,para nasal sinuses, thorax.
 MRI Brain –Multiple abscesses, meningeal enhancement,
infarction, hemorrhage.
 Fungal culture-
Maxillary sinus aspirate through antral
puncture.
Induced sputum
Bronchoscopy guided biopsy
 Blood and BAL –PCR assays-86-100%
 CSF-
ELISA for Galactomannan & Beta D Glucan
-sensitivity -70-80%
Latex agglutination for Galactomannan
-sensitivity -70-80%

144. Mucor Mycosis
 Occurs predominantly in immunocompromised
host
 HIV, DKA, IV Drug abusers, malignancies.
 Most common-Rhino cerebral mucor mycosis
 CNS Infections occurs in 30 % of Mucor
mycosis cases.
 Periorbital pain, facial numbness, conjunctival
suffusion, blindness followed by proptosis and
vision loss, ophthalmoplegia, Cavernous sinus
thrombosis and chronic meningitis.
Spellberg et al .Recent advances in management of Mucor mycosis.From bench to bedside. Clin Infect Dis

145. Lab diagnosis
 CSF –clear
 Cell count increases –predominantly lymphocytes
 Elevated protein, decreased glucose.
 CSF –Culture positivity <50 % of cases.
 Other samples- Sputum, Broncho alveolar
Lavage, Biopsy of the lesion
 Histopathology- Broad ribbon like non septate
hyphae.
 PCR is under investigation for diagnosis
Spellberg et al .Recent advances in management of Mucor mycosis.From bench to bedside.
Clin Infect Dis.48:1743,2009

146. Neuro Sarcoidosis
 Clinical profile:
 Neurological involvement -25 % cases
 Cranial neuropathies in 50 % -Facial nerve>
optic nerve> vestibulo cochlear nerve.
 Chronic meningitis-26% of patients
 Longitudinally extensive myelitis (>3
segments)
 Sub acute axonal polyneuropathy/GBS
 Diabetes insipidus
Chad Hoyle et al :Neuro sarcoidosis:Clinical review of a disorder with challenging inpatien

147. Lab diagnosis –Neuro
Sarcoidosis
 Serum ACE elevated -50 % cases with Neuro sarcoidosis
 CSF—Elevated protein(>200mg/dl) with positive Oligoclonal
bands, lymphocytic pleocytosis (10-200 cells /mcl).
 Sensitivity of CSF ACE 24-55%
 High CSF ACE levels- Infections
- carcinoma
Chad Hoyle et al :Neuro sarcoidosis:Clinical review of a disorder with challenging inpatient presentations .The

148. Neuro Sarcoidosis
 Other useful CSF markers-CSF lysosome,
Beta2 microglobulin, Tcell lymphocyte ratios-
under trials
 Chest X ray is found to be more sensitive
than CSF ACE levels. Sensitivity -55-60 %
 Ga scanning evaluating uptake in lung and
parotid regions –sensitivity -50-65%
 Chad Hoyle et al :Neuro sarcoidosis:Clinical review of a disorder with challenging inpatient presentations .The

149. Neuro Sarcoidosis
Other investigations:
 Lymph node /skin/trans bronchial lung biopsy
 Muscle /conjunctival/bone marrow biopsy if no
obvious systemic abnormality is detected.
 Meningeal biopsy if enhancing in MRI and
accessible
Sensitivity
-30-50%
Chad Hoyle et al :Neuro sarcoidosis:Clinical review of a disorder with challenging inpatient presentations .The Ne

150. Tests done at NIMHANS
Yield from January 2015 till August 2015
SUSPECTED
DISEASE
TESTS YIELD
TB Meningitis ZN staining
CULTURE-LJ Medium, MGIT 320
ELISA –Anti Mycobacterial Antibodies
PCR, Gene Xpert (sputum), Line probe
assay(sputum)
Culture positive-
165/884 (18.55%)
12.3%-SIRE Sensitive
6.22%-Resistant -At
least 1 drug
0.03 % -MDR
Neuro
Syphilis
Serum and CSF -VDRL
Serum TPHA
40 /3000 CSF
(1.11%)
Neuro
Brucellosis
Serum and CSF-Rose Bengal Test
ELISA for antibodies
Culture
PCR
PCR- 6/22
4-IgM, 1-IgG, 1-Both

151. Tests done at NIMHANS
SUSPECTED DIAGNOSIS TEST YIELD
Cryptococcal Meningitis India Ink
Latex Agglutination Test for Antigen
Fungal culture
II positive in
50 out of 60
culture
positive
cases.
( 83.3%)
Aspergillus and Mucor
Meningitis
Fungal culture _
Cysticercal Meningitis ELISA Anti cysticercal antibodies 8/10 Cases
(80%)
Toxoplasma Meningitis Latex Agglutination Test for Antigen,
ELISA for antibodies
_
Acanthameba Meningitis Wet mount preparation, Culture _

152. CONCLUSIONS
Chronic meningitis is defined as meningeal inflammation persisting
for at least 4 weeks.
The presentation may be non-specific and diagnosis of underlying
cause may be difficult. Cause may not be found in up to 1/3rd of
patients.
Tuberculosis is the commonest identifiable cause world wide. Febrile
patient with cranial nerve palsies, CSF lymphocytosis and low CSF
glucose is highly likely to have Tuberculosis.
In HIV patients with CD4 < 100 presenting with chronic headache,
most common cause is Tubercular, followed by Cryptococcal
meningitis.

153. CONCLUSIONS
In any patient with syphilis, CSF examination should be
performed in those with neurological symptoms and signs.
If infection and malignancy are reasonably excluded, a trial of
corticosteroids may be given.
In case of sarcoidosis, opportunistic infections must be
excluded before attributing it as the cause.
Meningeal biopsy is the diagnostic modality helpful when no
clue is found after extensive investigations.

154. PETER
PERLMANN KARY MULLISE
L
I
S
A
P
C
R
Thought is the
original source
of all wealth, all
success, all
material gain,
all great
discoveries and
inventions, and
of all
achievements.
Claude M.
BristolTHANK
YOU