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The presence of the meningococcus in the nasopharynx of normal individuals, and the bactericidal property of blood against the meningococcus
T H E P R E S E N C E OF T I t E MENINGOCOCCUS IN T H E NASOPHARYNX OF NORMAL INDIVIDUALS, AND T t I E B A C T E R I C I D A L P R O P E R T Y OF BLOOD AGAINST T H E MENINGOCOCCUS NELLES
SILVERTHORNE,
M.B.
TORONTO, ONTARIO
epidemics of meningoeoccic meningitis many authors have D URING reported high contact carrier rates. However, the presence of carriers has also been found during interepidemic periods. I n 1931 Cameron 1 demonstrated four carriers in a group of twenty-nine persons swabbed, all of whom were in the same building. None of these individuals had been in contact with eases of meningitis. One o~ these individnals was still found to be a carrier one year-later. In 1934 Rake 2 discovered carriers of the meningococcus in healthy individuals. Dudley and Brennan s in 1935 found a non-contact carrier rate as high as 60 per cent in a naval and military population. As these observations are of practical importance to the pediatrician, during the past two years we have studied the carrier rate in a noncontact population. As it is also relevant to assess the importance of certain immune reactions in the blood of individuals who may be hatboring strains of the meningoeoecus, we have investigated t he baetericidal property of the blood of normal individuals and of noncontaet carriers against various strains of meningococci. TECHNIC
West tube swabs were taken of the posterior nasopharynx, planted, and spread on dextrose blood agar plates. The plates were incubated at 37 ~ C. for twenty-four to forty-eight hours, and colonies were picked and examined microscopically. Cultures of these colonies were planted in carbohydrate media containing dextrose, maltose, and saccharose. A suspension of each culture was tested with polyvalent antimeningoeoceus serum. All strains isolated were found to be gram-negative diplococei and produced acid in dextrose and maltose but not in saceharose. They agglutinated with polyvalent antimeningococcus serum. RESULTS
Previous to the present study we had isolated meningococci from 16 carriers in a group of 125 medical students. During the last two F r o m the w a r d s and l a b o r a t o r i e s of the H o s p i t a l for Sick Children, the D e p a r t m e n t of P a e d i a t r i c s and the C o n n a u g h t L a b o r a t o r i e s , U n i v e r s i t y of Toronto. l~ before the F o r t y - E i g h t h A n n u a l Me e t i ng of t he A m e r i c a n P e d i a t r i c Society in ]olnt session w i t h the C a n a d i a n Society for the S t udy of D i s e a s e s of Children, a t Bolton Landing, l'~. Y., J u n e 11-13, 1936. 328
SILVERTtIORNE:
i~(ENINGOCOCCUS 1N N O R M A L
NASOPHARYNX
329
years of this study, 63 healthy adults mainly physicians and technicians were swabbed at approximately monthly intervals. In all, 1,227 swabs were taken over the two-year period. No less than 243 were positive for the meningoeoccus. The percentage of the total positive swabs taken was 19.8. The monthly percentage of positive individuals varied between 16 and 28, with an average of 20. For the two-year period, of the 63 healthy adults tested, no less than 26, or 41 per cent, showed the presence of the ~neningoeoceus in the nasopharynx at one time or another. In these 26 individuals, 11 were shown to be harboring the meningococcus fairly persistently. Thirteen carried the meningococeus intermittently, and in 2 instances cultures of the meningoeoecus were obtained on only one occasion. It was also noted that there was either a complete absence or a marked reduction in the colonies of meningocoeci when swabs were taken during an acute upper respiratory infection in any given individual. These findings are in general agreement with those reported by Rake? In 1934 Silverthorne and Fraser ~ demonstrated that samples of blood from many adults contained bactericidal power to a recently isolated strain of the meningococeus. Other adults and infants failed to manifest this property in their blood. These authors :further showed that two infants with meningitis possessed relatively little or no bactericidal power in samples of their blood at the onset of the disease, but developed it after recovery. It was also shown that strains of meningoeoeci vary in their ability to thrive in a sample of blood containing no bactericidal power when measured against a " v i r u l e n t " spinal fluid strain. A ' ' v i r u l e n t " strain is one that is lethal to mice in the more diluted suspensions in the mouse-mucin test. 7 These findings furnish one with two facts, first, that strains of-meningoeoeei vary in their power to " i n v a d e " a nonbacterieidal sample of human blood and, second, that individuals possess varying amounts of the bactericidal principle in their blood against virulent meningoeoeei. In 1935 we 5 found that the bactericidM principle was preseIlt in serum or plasma and depended on the presettce of complement for its action. We also demonstrated the specificity of the bactericidal power of blood against the meningoeoeeus and were able to produce this property in the blood of guinea pigs by vaccination with recently isolated virulent eerebrospinal fluid strains. Recently, we 6 have explored and extended some of these observations. The relation of the bactericidal power of blood to the virulence of the microorganism has been studied. Using as a control test a previously known nonbaeterieidal sample of blood, and a virulent recently isolated cerebrospinal fluid strain of the meningocoeeus, we demonstrated that samples of blood from five carriers were bacterieidM to this virulent strain and to each of the five carrier strains. One might suppose from these findings that carriers possess bactericidal property in their Mood which has been developed due to the continued presence of the menin-
330
TIIE JOURNAL
OF PFA)IATRICS
gococcus in their nasopharynx. Most of the samples of blood f r o m carriers which were tested possessed this p r o p e r t y to certain virulent spinal fluid strains. W e also demonstrated that the power of meningococci to invade a nonbactericidal sample of blood was, in general, parallel to the virulence of strains as judged on the basis of the mousemucin test of Miller. B y the use of the mouse-mucin test and the bactericidal test, eleven noneontact carrier strains were examined. Three of these proved to be as virulent as any of the recently isolated spinal fluid strains, and the remaining eight strains proved to be avirulent. I n later studies, strains both f r o m spinal fluid and noncontact carriers were f o u n d to be in a " m i d p o s i t i o n " in respect of virulence, and one spinal fluid strain was avirulent as j u d g e d by the results of these tests. This latter finding m a y be due to the lack of the rapid subeulturing on suitable media of this strain before being received. Samples of blood obtained from two of the three carriers of virulent nasopharyngeal strains had marked bactericidal power to their respective strains; the third was not tested, CONCLUSIONS
1. A total of 1,227 swabs were t a k e n f r o m 63 normal healthy adults over a two-year period; 19.8 per cent of these were positive for the meningococcus. 2. Sixty-three individuals were swabbed at a p p r o x i m a t e l y monthly intervals. On these monthly examinations the n u m b e r of positive individuals varied f r o m 16 to 28 per cent. No less than 41 per cent of the group of 63 were positive at one time or another during the twoy e a r period. Eleven individuals were persistent, 13 intermittent, and 2 transient carriers. 3. Eleven noncontact carrier strains were examined by the bactericidal and mouse-muein tests; 3 of these strains w e r e found to be virulent, and 8 were f o u n d to be avirulent. 4. Carriers of the meningoeoecus possess bactericidal power in their blood to their respective strains whether these strains are virulent or aviru]ent. Also, samples of blood f r o m carriers possess bactericidal power to certain virulent cerebrospinal fluid strains. 5. The determination of the virulence of strains of meningoeocei by means of the mouse-muein and bactericidal tests m a y serve as a useful means of selecting strains for the p r e p a r a t i o n of therapeutic serum. REFERENCES
]. 2. 3. 4. 5. 6. 7.
Cameron~ D.W." Tr. Roy Soc. Canad,, Sec. V. 9: 97, ]931. l~ake, G.: J. Exper. Med. 59: 553, ]934. Dudley, S. F., and Brennan, J.R.: J, Ityg. 34: 525, 1935. Silverthorne, N., and Fraser, D. T.: Brit. J. Exper. Path. 15: 362, 1934. Silverthorne, N., and Fraser, D.T.: J. Immunol. ~9: 6, 1935. Silverthorne, N., and Fraser, D.T.: (In press.) lV[iller, C.P.: Science 78: 340, 1933.
SILVERTHORNE,
M.B.
TORONTO, ONTARIO
epidemics of meningoeoccic meningitis many authors have D URING reported high contact carrier rates. However, the presence of carriers has also been found during interepidemic periods. I n 1931 Cameron 1 demonstrated four carriers in a group of twenty-nine persons swabbed, all of whom were in the same building. None of these individuals had been in contact with eases of meningitis. One o~ these individnals was still found to be a carrier one year-later. In 1934 Rake 2 discovered carriers of the meningococcus in healthy individuals. Dudley and Brennan s in 1935 found a non-contact carrier rate as high as 60 per cent in a naval and military population. As these observations are of practical importance to the pediatrician, during the past two years we have studied the carrier rate in a noncontact population. As it is also relevant to assess the importance of certain immune reactions in the blood of individuals who may be hatboring strains of the meningoeoecus, we have investigated t he baetericidal property of the blood of normal individuals and of noncontaet carriers against various strains of meningococci. TECHNIC
West tube swabs were taken of the posterior nasopharynx, planted, and spread on dextrose blood agar plates. The plates were incubated at 37 ~ C. for twenty-four to forty-eight hours, and colonies were picked and examined microscopically. Cultures of these colonies were planted in carbohydrate media containing dextrose, maltose, and saccharose. A suspension of each culture was tested with polyvalent antimeningoeoceus serum. All strains isolated were found to be gram-negative diplococei and produced acid in dextrose and maltose but not in saceharose. They agglutinated with polyvalent antimeningococcus serum. RESULTS
Previous to the present study we had isolated meningococci from 16 carriers in a group of 125 medical students. During the last two F r o m the w a r d s and l a b o r a t o r i e s of the H o s p i t a l for Sick Children, the D e p a r t m e n t of P a e d i a t r i c s and the C o n n a u g h t L a b o r a t o r i e s , U n i v e r s i t y of Toronto. l~ before the F o r t y - E i g h t h A n n u a l Me e t i ng of t he A m e r i c a n P e d i a t r i c Society in ]olnt session w i t h the C a n a d i a n Society for the S t udy of D i s e a s e s of Children, a t Bolton Landing, l'~. Y., J u n e 11-13, 1936. 328
SILVERTtIORNE:
i~(ENINGOCOCCUS 1N N O R M A L
NASOPHARYNX
329
years of this study, 63 healthy adults mainly physicians and technicians were swabbed at approximately monthly intervals. In all, 1,227 swabs were taken over the two-year period. No less than 243 were positive for the meningoeoccus. The percentage of the total positive swabs taken was 19.8. The monthly percentage of positive individuals varied between 16 and 28, with an average of 20. For the two-year period, of the 63 healthy adults tested, no less than 26, or 41 per cent, showed the presence of the ~neningoeoceus in the nasopharynx at one time or another. In these 26 individuals, 11 were shown to be harboring the meningococcus fairly persistently. Thirteen carried the meningococeus intermittently, and in 2 instances cultures of the meningoeoecus were obtained on only one occasion. It was also noted that there was either a complete absence or a marked reduction in the colonies of meningocoeci when swabs were taken during an acute upper respiratory infection in any given individual. These findings are in general agreement with those reported by Rake? In 1934 Silverthorne and Fraser ~ demonstrated that samples of blood from many adults contained bactericidal power to a recently isolated strain of the meningococeus. Other adults and infants failed to manifest this property in their blood. These authors :further showed that two infants with meningitis possessed relatively little or no bactericidal power in samples of their blood at the onset of the disease, but developed it after recovery. It was also shown that strains of meningoeoeci vary in their ability to thrive in a sample of blood containing no bactericidal power when measured against a " v i r u l e n t " spinal fluid strain. A ' ' v i r u l e n t " strain is one that is lethal to mice in the more diluted suspensions in the mouse-mucin test. 7 These findings furnish one with two facts, first, that strains of-meningoeoeei vary in their power to " i n v a d e " a nonbacterieidal sample of human blood and, second, that individuals possess varying amounts of the bactericidal principle in their blood against virulent meningoeoeei. In 1935 we 5 found that the bactericidM principle was preseIlt in serum or plasma and depended on the presettce of complement for its action. We also demonstrated the specificity of the bactericidal power of blood against the meningoeoeeus and were able to produce this property in the blood of guinea pigs by vaccination with recently isolated virulent eerebrospinal fluid strains. Recently, we 6 have explored and extended some of these observations. The relation of the bactericidal power of blood to the virulence of the microorganism has been studied. Using as a control test a previously known nonbaeterieidal sample of blood, and a virulent recently isolated cerebrospinal fluid strain of the meningocoeeus, we demonstrated that samples of blood from five carriers were bacterieidM to this virulent strain and to each of the five carrier strains. One might suppose from these findings that carriers possess bactericidal property in their Mood which has been developed due to the continued presence of the menin-
330
TIIE JOURNAL
OF PFA)IATRICS
gococcus in their nasopharynx. Most of the samples of blood f r o m carriers which were tested possessed this p r o p e r t y to certain virulent spinal fluid strains. W e also demonstrated that the power of meningococci to invade a nonbactericidal sample of blood was, in general, parallel to the virulence of strains as judged on the basis of the mousemucin test of Miller. B y the use of the mouse-mucin test and the bactericidal test, eleven noneontact carrier strains were examined. Three of these proved to be as virulent as any of the recently isolated spinal fluid strains, and the remaining eight strains proved to be avirulent. I n later studies, strains both f r o m spinal fluid and noncontact carriers were f o u n d to be in a " m i d p o s i t i o n " in respect of virulence, and one spinal fluid strain was avirulent as j u d g e d by the results of these tests. This latter finding m a y be due to the lack of the rapid subeulturing on suitable media of this strain before being received. Samples of blood obtained from two of the three carriers of virulent nasopharyngeal strains had marked bactericidal power to their respective strains; the third was not tested, CONCLUSIONS
1. A total of 1,227 swabs were t a k e n f r o m 63 normal healthy adults over a two-year period; 19.8 per cent of these were positive for the meningococcus. 2. Sixty-three individuals were swabbed at a p p r o x i m a t e l y monthly intervals. On these monthly examinations the n u m b e r of positive individuals varied f r o m 16 to 28 per cent. No less than 41 per cent of the group of 63 were positive at one time or another during the twoy e a r period. Eleven individuals were persistent, 13 intermittent, and 2 transient carriers. 3. Eleven noncontact carrier strains were examined by the bactericidal and mouse-muein tests; 3 of these strains w e r e found to be virulent, and 8 were f o u n d to be avirulent. 4. Carriers of the meningoeoecus possess bactericidal power in their blood to their respective strains whether these strains are virulent or aviru]ent. Also, samples of blood f r o m carriers possess bactericidal power to certain virulent cerebrospinal fluid strains. 5. The determination of the virulence of strains of meningoeocei by means of the mouse-muein and bactericidal tests m a y serve as a useful means of selecting strains for the p r e p a r a t i o n of therapeutic serum. REFERENCES
]. 2. 3. 4. 5. 6. 7.
Cameron~ D.W." Tr. Roy Soc. Canad,, Sec. V. 9: 97, ]931. l~ake, G.: J. Exper. Med. 59: 553, ]934. Dudley, S. F., and Brennan, J.R.: J, Ityg. 34: 525, 1935. Silverthorne, N., and Fraser, D. T.: Brit. J. Exper. Path. 15: 362, 1934. Silverthorne, N., and Fraser, D.T.: J. Immunol. ~9: 6, 1935. Silverthorne, N., and Fraser, D.T.: (In press.) lV[iller, C.P.: Science 78: 340, 1933.