Tubercle Tuber& (1991) 72. NO-192 Q Longman Group UK L.td 1991
Adenosine deaminase levels in cerebrospinal fluid in tuberculosis and bacterial meningitis R. K. CHAWLA, R. K. SETH, B. RAJ and A. S. SAINI Department of Chest Diseases and Biochemistry Medical College, Rohtak. India
Summary - Adenosine deaminase activity was measured in cerebrospinal fluid of patients with confirmed tuberculous and bacterial meningitis. The values were compared with those of control subjects without meningitis. A statistically significant increase in the level of this enzyme was noted in the two types of meningitis, but no definite demarcation in the levels was observed between the two types. Therefore increases in adenosine deaminase activity may not be of such diagnostic significance as reported elsewhere.
Introduction
confirmed by clinical, radiological (chest X-ray) and microbiological (sputum positive for acid-fast bacilli by Ziehl-Neelsen staining and/or culture) criteria.
Over the past decade various biochemical tests on cerebrospinal fluid (CSF) have been reported to help in the early diagnosis of tuberculous meningitis (IBM). These include the determination of lactate concentration [l, 21, lactate dehydrogenase activity [3,4] and the bromide partition test [5,6]. These tests, however, are of little value in culture negative and low cell count meningitis. It has recently been reported that assay of adenosine deaminase (ADA: EC 3.5.4.4) activity in CSF enables TBM to be distinguished from bacterial meningitis [6-8]. This study was therefore undertaken to evaluate the status of ADA assay in the diagnosis of tuberculous meningitis.
Bacterial meningitis. This group included 27 patients with a mean age of 30 months. Only those patients from whom bacteria were grown on culture of CSF were included in this study. The causative organism was Streptococcus pneumoniae in 8 cases and Staphylococcus aureus in the other 19 cases.
Patients and methods Patients All CSF specimens were taken by lumbar puncture under aseptic conditions. Blood-strained traumatic taps were excluded from the present study as human erythrocytes have a high ADA content [Q]. Patients were divided into the following groups: Tuberculous meningitis. This group comprised 12 patients of a mean age of 33 years with tuberculosis Correspondence Rohtak-124M)l.
to: Dr R.K. Seth, 28/9-J, Medical Enclave, Hatyana. India.
Controls. The control group contained 36 individuals; 14 were in age group of 1-15 years (mean age 3.5 years) who were investigated for convulsions. These individuals did not show any evidence of central nervous system disease. The remaining 22 controls were adults in the age range of 30-47 years (mean age 35.2 years) investigated for various reasons. Their subsequent conrse remained uneventful. It has previously been reported that there were no age or sex differences in ADA levels in normal individuals aged between 2 and 79 years (mean 19.4 f 11.7) 181. From the ethical point of view, samples of CSF were only taken when there was a need to rule out any organic disease including secondary causes of epilepsy or convulsions. The control CSF samples
190
191
ADENOSEVE DEAMINASE LEVELS IN CSF
Table Adenosine deaminase activity (u/I) in the CSF of the different groups of patients and controls Groyp Control Children Adults Total Tuberculous meningitis Bacterial meningitis
Number
Mean fsD
Minimwn
Maimw?l
14 22 36 12
1.06 f 0.68 1.86 f 1.02 1.55 It 0.98 10.45 f 4.50
0.2 0.3 0.2 4.8
2.2 3.1 3.1 19.2
27
11.22 * 4.59
4.9
20.2
Statistical significance (Student’s t-test): A vs B p < 0.001; A vs C p < 0.001; B vs C p > 0.05)
were those that were found to be normal biochemically (with regard to the protein, sugar and chloride levels), cytologically and microbiologically. Neither the meningitis patients nor controls had received antibiotic therapy before lumbar puncture. Assay of ADA activity Assay of ADA activity was carried out on all the CSF samples at 37’C by the method of Giusti [9]. ADA catalyses the following reaction: Adenosine + Hz0 - - -4nosine
+ NH3
The release of ammonia was determined colorimetritally after the development of an intense blue colour with hypochlorite and phenol in alkaline solution. Alkaline hypochlorite was prepared from commercial bleaching powder and sodium hydroxide. All other reagents used were of analytical grade. Adenosine was obtained from Sigma Chemical Co,, USA.
tuberculous and bacterial meningitis. The ADA is distributed in various mammalian tissues including cerebral cortex and lymphoid tissues [ 10, 111. It has been reported previously that ADA cannot pass the bloodbrain barrier in typhoid fever [12]. The increase in ADA levels in CSF of patients with the two types of meningitis could therefore be a result of partial damage to the blood-brain barrier, allowing the entry of ADA from plasma or cerebral tissue into the CSF. Malan et al. [6] have also reported an increase in the activity of ADA in both bacterial and tuberculous meningitis. Nevertheless, they claimed that levels of this enzyme in CSF are useful in distinguishing TBM from bacterial meningitis. A value higher than 6u/l character&d tuberculous meningitis while values lower than 6u/l were found in bacterial meningitis. In contrast to the findings of Malan et al [6], we found that the mean values and range of ADA in CSF were virtually identical in tuberculous and bacterial meningitis. We therefore conclude that the assay of ADA levels in CSF is of little help in the early diagnosis of tuberculous meningitis.
Results References Results of the assay of ADA activity are shown in the Table. No significant difference was observed in the activity of ADA of neonates (with convulsions) and adult controls (> 0.05). The mean ADA levels were found to be less than 2 u/l in all the controls. A statistically significant increase in the activity of this enzyme was found in CSF of patients with TBM and bacterial meningitis compared to the normal controls (p c 0.001) but the values of ADA in the two groups of meningitis patients were almost identical with a very large overlap (Table). Discussion It is apparent from the results of this study that the level of ADA in CSF is considerably elevated in both
1. Brook I. Bricknell KS, Overmff GD, Fiiegold SM. Measurement of lactic acid in cerebrospinal fluid of patients with infections of the central nervous system. J Infecf Dis 1978; 137: 384-390. 2. Lauwers S. Lactic acid concentration in cerehspinal fluid and differential diagnosis of meningitis. Lancer 1978; ii: 163. 3. Wroblewski F, Dacker B, Wroblewski R. The clinical implications of spinal fluid lactic dehydrogenase activity. New Engl J Med 1958; 258: 635. 4. Khanna SK, Gupa DK. Khanna F’.Value of lactic dehydrogenase in cer&rc~~pinal fluid of tuberculous meningitis patients. J Indian Med Ass 1977; 68: 4-6. 5. Wiggelinkhuizen J. Mann M. The radioactive bromide partition test in the diagnosis of tuberculous meningitis in children. J Pediatr 1980; 97: 843-847. 6. Malan C. Donald PR. Golden M. Taljaard JJE Adenosine deaminase levels in cerebrospinal fluid in the diagnosis of tuberculous meningitis. .I Trop Med Hyg 1984; 87: 334.
192 7. MulLx-Beissenhintz van W. Keller H. Die Bestimmungder Dtsch Md Wochendu l%a; Adenosinedeamiiasein se-. 91: 159-168. 8. Goldberg DM. &mm adenosinedcamii in the differential diagnosisofjaundice Br Meci J 1965; 1: 353-355. 9. Giusti G. Abosine dcamiiase.In: BergmeyerHU. ed.Methods k Enzymic anatys~ 2nd edn. Wieahea~:V&g Chemie, 1974:pp 1092-1099.
c!HAwLAANo-
10. Haukiewicz J. bsniak h-f, Adenosine deambse in cerel1 bro&aIaI¶uiadL~ia 1972;43: 385-395. w&rw5wxadalosmedmmiMse act& and its shcklar distrii. Bkhm J 1974;144: 3741. 12. GalantiB. Nonliello S. RussoM, Flomtino F. InmmaI lymphoqtc adamsinehaminase ia qphoid fever.Sand J Itfed Dir 1981; 13: 4750.
Adenosine deaminase levels in cerebrospinal fluid in tuberculosis and bacterial meningitis R. K. CHAWLA, R. K. SETH, B. RAJ and A. S. SAINI Department of Chest Diseases and Biochemistry Medical College, Rohtak. India
Summary - Adenosine deaminase activity was measured in cerebrospinal fluid of patients with confirmed tuberculous and bacterial meningitis. The values were compared with those of control subjects without meningitis. A statistically significant increase in the level of this enzyme was noted in the two types of meningitis, but no definite demarcation in the levels was observed between the two types. Therefore increases in adenosine deaminase activity may not be of such diagnostic significance as reported elsewhere.
Introduction
confirmed by clinical, radiological (chest X-ray) and microbiological (sputum positive for acid-fast bacilli by Ziehl-Neelsen staining and/or culture) criteria.
Over the past decade various biochemical tests on cerebrospinal fluid (CSF) have been reported to help in the early diagnosis of tuberculous meningitis (IBM). These include the determination of lactate concentration [l, 21, lactate dehydrogenase activity [3,4] and the bromide partition test [5,6]. These tests, however, are of little value in culture negative and low cell count meningitis. It has recently been reported that assay of adenosine deaminase (ADA: EC 3.5.4.4) activity in CSF enables TBM to be distinguished from bacterial meningitis [6-8]. This study was therefore undertaken to evaluate the status of ADA assay in the diagnosis of tuberculous meningitis.
Bacterial meningitis. This group included 27 patients with a mean age of 30 months. Only those patients from whom bacteria were grown on culture of CSF were included in this study. The causative organism was Streptococcus pneumoniae in 8 cases and Staphylococcus aureus in the other 19 cases.
Patients and methods Patients All CSF specimens were taken by lumbar puncture under aseptic conditions. Blood-strained traumatic taps were excluded from the present study as human erythrocytes have a high ADA content [Q]. Patients were divided into the following groups: Tuberculous meningitis. This group comprised 12 patients of a mean age of 33 years with tuberculosis Correspondence Rohtak-124M)l.
to: Dr R.K. Seth, 28/9-J, Medical Enclave, Hatyana. India.
Controls. The control group contained 36 individuals; 14 were in age group of 1-15 years (mean age 3.5 years) who were investigated for convulsions. These individuals did not show any evidence of central nervous system disease. The remaining 22 controls were adults in the age range of 30-47 years (mean age 35.2 years) investigated for various reasons. Their subsequent conrse remained uneventful. It has previously been reported that there were no age or sex differences in ADA levels in normal individuals aged between 2 and 79 years (mean 19.4 f 11.7) 181. From the ethical point of view, samples of CSF were only taken when there was a need to rule out any organic disease including secondary causes of epilepsy or convulsions. The control CSF samples
190
191
ADENOSEVE DEAMINASE LEVELS IN CSF
Table Adenosine deaminase activity (u/I) in the CSF of the different groups of patients and controls Groyp Control Children Adults Total Tuberculous meningitis Bacterial meningitis
Number
Mean fsD
Minimwn
Maimw?l
14 22 36 12
1.06 f 0.68 1.86 f 1.02 1.55 It 0.98 10.45 f 4.50
0.2 0.3 0.2 4.8
2.2 3.1 3.1 19.2
27
11.22 * 4.59
4.9
20.2
Statistical significance (Student’s t-test): A vs B p < 0.001; A vs C p < 0.001; B vs C p > 0.05)
were those that were found to be normal biochemically (with regard to the protein, sugar and chloride levels), cytologically and microbiologically. Neither the meningitis patients nor controls had received antibiotic therapy before lumbar puncture. Assay of ADA activity Assay of ADA activity was carried out on all the CSF samples at 37’C by the method of Giusti [9]. ADA catalyses the following reaction: Adenosine + Hz0 - - -4nosine
+ NH3
The release of ammonia was determined colorimetritally after the development of an intense blue colour with hypochlorite and phenol in alkaline solution. Alkaline hypochlorite was prepared from commercial bleaching powder and sodium hydroxide. All other reagents used were of analytical grade. Adenosine was obtained from Sigma Chemical Co,, USA.
tuberculous and bacterial meningitis. The ADA is distributed in various mammalian tissues including cerebral cortex and lymphoid tissues [ 10, 111. It has been reported previously that ADA cannot pass the bloodbrain barrier in typhoid fever [12]. The increase in ADA levels in CSF of patients with the two types of meningitis could therefore be a result of partial damage to the blood-brain barrier, allowing the entry of ADA from plasma or cerebral tissue into the CSF. Malan et al. [6] have also reported an increase in the activity of ADA in both bacterial and tuberculous meningitis. Nevertheless, they claimed that levels of this enzyme in CSF are useful in distinguishing TBM from bacterial meningitis. A value higher than 6u/l character&d tuberculous meningitis while values lower than 6u/l were found in bacterial meningitis. In contrast to the findings of Malan et al [6], we found that the mean values and range of ADA in CSF were virtually identical in tuberculous and bacterial meningitis. We therefore conclude that the assay of ADA levels in CSF is of little help in the early diagnosis of tuberculous meningitis.
Results References Results of the assay of ADA activity are shown in the Table. No significant difference was observed in the activity of ADA of neonates (with convulsions) and adult controls (> 0.05). The mean ADA levels were found to be less than 2 u/l in all the controls. A statistically significant increase in the activity of this enzyme was found in CSF of patients with TBM and bacterial meningitis compared to the normal controls (p c 0.001) but the values of ADA in the two groups of meningitis patients were almost identical with a very large overlap (Table). Discussion It is apparent from the results of this study that the level of ADA in CSF is considerably elevated in both
1. Brook I. Bricknell KS, Overmff GD, Fiiegold SM. Measurement of lactic acid in cerebrospinal fluid of patients with infections of the central nervous system. J Infecf Dis 1978; 137: 384-390. 2. Lauwers S. Lactic acid concentration in cerehspinal fluid and differential diagnosis of meningitis. Lancer 1978; ii: 163. 3. Wroblewski F, Dacker B, Wroblewski R. The clinical implications of spinal fluid lactic dehydrogenase activity. New Engl J Med 1958; 258: 635. 4. Khanna SK, Gupa DK. Khanna F’.Value of lactic dehydrogenase in cer&rc~~pinal fluid of tuberculous meningitis patients. J Indian Med Ass 1977; 68: 4-6. 5. Wiggelinkhuizen J. Mann M. The radioactive bromide partition test in the diagnosis of tuberculous meningitis in children. J Pediatr 1980; 97: 843-847. 6. Malan C. Donald PR. Golden M. Taljaard JJE Adenosine deaminase levels in cerebrospinal fluid in the diagnosis of tuberculous meningitis. .I Trop Med Hyg 1984; 87: 334.
192 7. MulLx-Beissenhintz van W. Keller H. Die Bestimmungder Dtsch Md Wochendu l%a; Adenosinedeamiiasein se-. 91: 159-168. 8. Goldberg DM. &mm adenosinedcamii in the differential diagnosisofjaundice Br Meci J 1965; 1: 353-355. 9. Giusti G. Abosine dcamiiase.In: BergmeyerHU. ed.Methods k Enzymic anatys~ 2nd edn. Wieahea~:V&g Chemie, 1974:pp 1092-1099.
c!HAwLAANo-
10. Haukiewicz J. bsniak h-f, Adenosine deambse in cerel1 bro&aIaI¶uiadL~ia 1972;43: 385-395. w&rw5wxadalosmedmmiMse act& and its shcklar distrii. Bkhm J 1974;144: 3741. 12. GalantiB. Nonliello S. RussoM, Flomtino F. InmmaI lymphoqtc adamsinehaminase ia qphoid fever.Sand J Itfed Dir 1981; 13: 4750.
