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Branhamella catarrhalis in children and adults. A study of prevalence, time of colonisation, and association with upper and lower respiratory tract infections
Journal of Infection (1994) 29, 23-3I
B r a n h a m e l l a c a t a r r h a l i s in c h i l d r e n a n d adults. A s t u d y o f prevalence, time of colonisation, and association with upper a n d l o w e r r e s p i r a t o r y tract i n f e c t i o n s Tove Ejlertsen,* Ebbe Thisted,t Finn Ebbesen,T Bente Olesen~ and Jan Renneberg~ * Department of Clinical Microbiology and ~fDepartment of Pediatrics, Aalborg Hospital, Aalborg, ~ Department of Clinical Microbiology, Hvidovre Hospital, Hvidovre, ~Division of Preventive Microbiology, Statens Seruminstitut, Copenhagen, Denmark Accepted for publication 5 January 1994
Summary The colonisation rate of Branhamella catarrhalis in patients from o to 45 years of age was examined. Of 561 women admitted to hospital in labour, 6 (I %) carried B. catarrhalis in their throats but none carried the organism in their vaginas. None of 534 newborn babies became colonised at birth or during their 5 days' stay in hospital. Neither were lO2 neonates < I month of age in hospital colonised. The maximum colonisation rate during childhood was observed in children 1-48 months of age with 143 of 266 (54%) children colonised. Among children 4-15 years of age, four of 57 (7%) children with healthy respiratory tracts were colonised. Significantly more children with upper or lower respiratory tract infections (RTI) were colonised (68 %) than were children without such infections (36 %), (P < o.ooi). After recovery from RTI, the isolation rate in the R T I group fell to that of the non-RTI group. A seasonal variation in prevalence was not observed. Of all the strains of B. catarrhalis isolated, 84 % produced beta-lactamase.
Introduction Branhamella catarrhalis is a f r e q u e n t inhabitant of the oro- and n a s o p h a r y n x of children o--io years of age with a prevalence reported of approximately 25-35%.1-3 T h e time, m o d e and d u r a t i o n of colonisation as well as the m e c h a n i s m s of elimination o f B. catarrhalis have been poorly elucidated. One s t u d y has shown a high degree of acquisition and elimination of B. catarrhalis strains in the n a s o p h a r y n x of children w i t h o u t the influence of antibiotic treatment3 Sporadic observations have drawn attention to the role of B. catarrhalis in small children with b r o n c h o - p u l m o n a r y infections and tracheitis3 -s T h e organism has also been associated with atypical p s e u d o - c r o u p in preschool children 9 as well as cough and hoarseness in children and adults. 1' 10 T h u s , the presence of B. catarrhalis in respiratory tract infections ( R T I ) in children is certain b u t its role as a p r i m a r y p a t h o g e n remains controversial, * Address correspondence to : D r T. Ejlertsen, Dept. Clinical Microbiology, Aalborg Hospital, D K 9ooo Aalborg, Denmark. o163-4453/94/29oo23 +09 $08.00/0
© 1994 The British Society for the Study of Infection
24
T. EJLERTSEN E T A L .
In order to assess the importance of B. catarrhalis in tracheal secretions and throat cultures it may be helpful to know the expected incidence of colonisation with B. catarrhalis in each age group, and to what extent symptoms of respiratory tract infections (RTI) are associated with colonisation. T h e purpose of this study has been to investigate the transmission of B. catarrhalis from mother to child at birth, the incidence of colonisation during the first days of life, and the prevalence of B. catarrhalis throughout childhood. In children aged 1-47 months the relation to R T I has also been examined. Patients and m e t h o d s Patients
Healthy women admitted to the Maternity Department of Aalborg Hospital in labour and their newborn children were examined prospectively during a period of 4 months from September to December z989. Throat swabs and vaginal swabs were collected from the women before delivery. After birth, throat secretions were collected from their babies. On discharge from the hospital 5 days later, throat swabs were obtained from the babies. Children, aged o-47 months, admitted to the Paediatric Department of Aalborg Hospital, were examined prospectively during a period of I year from June z989 to May z99o. T h e y were divided into two groups according to their symptoms at the time of admission. Patients suffering from upper or lower R T I formed one group. R T I comprised catarrh, acute laryngitis, acute tracheitis, acute bronchiolitis and pneumonia. In the R T I group, tracheal secretions or a throat swab was collected on admission, and a throat swab was collected on discharge. During the period of 5 months from November I989 to March I99O, additional nasopharyngeal aspirates were collected and examined for the presence of respiratory syncytial virus (RSV) in this group. Patients not suffering from R T I (non-RTI), but in hospital for other reasons, formed another group. Throat swabs were obtained at the time of admission and of discharge in this group also. Antibiotic treatment of patients was noted in both groups. T h e last group consisted of outpatients aged 4-I5 years with healthy respiratory tracts when examined at Hvidovre Hospital, Copenhagen, during the period from July to December I992. All studies were approved by the local Ethics Committee, and the patients participated only after giving informed consent. A parent's informed consent was obtained in the case of each of the children. Microbiological methods
All specimens were sent to the local Department of Clinical Microbiology. Tracheal secretions were aspirated through a nasopharyngeal catheter and kept refrigerated for a maximum of 24 hours until they were cultured. Throat swabs were taken with a charcoal cotton-tipped applicator swabbed over the tonsils and palate. T h e tip of the applicator was stored in Stuart's transport medium and refrigerated until it was cultured. Tracheal secretions were centrifuged at z5oo × g for IO min. T h e sediment was recovered with a sterile cotton applicator and cultured on 5 % horse blood agar (streaked with
B. catarrhalis in children and adults
25
Staphylococcus aureus for detection of Haemophilus influenzae), 'chocolate' agar, and a Danish 'GC-chocolate' agar medium (selective for pathogenic Neisseria species) containing polymyxin B, a low concentration of lincomycin, mycostatin, and trimethoprim but without vancomycin (Statens Seruminstitut, Copenhagen, Denmark). This medium permitted all strains of B. catarrhalis observed on 'chocolate agar' and on horse blood agar to be recovered in pure growth without interference from the commensals of the pharynx other than Neisseria-species. Throat swabs were cultured on 5 % horse blood agar and on 'GC-chocolate' agar. All media were incubated aerobically with the addition of 5 % CO2 for 2 days. B. catarrhalis was identified according to colonial morphology, Gram-staining reaction, catalase and oxidase reactions, reduction of nitrate to nitrite as well as inability to produce acid from glucose, maltose, and lactose together with production of DNase. Isolates were tested for beta-lactamase activity by means of a chromogenic cephalosporin (Cefinase, BBL Microbiology Systems, U.S.A.). H. influenzae and Streptococcus pneumoniae were identified from tracheal secretions collected from the R T I group, 1-47 months of age, according to established methods. 11 RSV was identified by means of a direct immunofluorescence test (Imagen ® Respiratory Syncytial Virus, D A K O Diagnostics Ltd., U.K.) Statistical methods
A file was generated in M E D S T A T J ~X~was calculated from 2 x 2 contingency tables while Fisher's exact test (two-tailed) was used for small numbers. Results Characteristics of patients
A total of 1520 patients participated in the study (Table I). Of these, 561 were women admitted in labour with healthy respiratory tracts. T h e y formed the adult group aged z6-45 years. Of their babies, 534 formed the group of newborns examined at birth and of whom 499 were re-examined on discharge 5 days later. T h e neonatal group, aged < z month, consisted of zo2 babies, 9I without R T I and I z with R T I . Children aged 1-47 months formed a group of 266 patients, I I9 without R T I , 56 of whom were re-examined on discharge; and I47 with R T I , 88 of whom were re-examined on discharge. A further group was comprised of 57 outpatients aged 4-I5 years with healthy respiratory tracts. I s o l a t i o n rates o f B r a n h a m e l l a c a t a r r h a l i s
T h e isolation rates of B. catarrhalis were strongly age-dependent. Neonates < z month of age were not colonised with B. catarrhalis, regardless of whether or not they had a respiratory infection (Table I). Overall, z43/266 (54%) children I to 47 months of age were colonised. In this age group, a significant difference between the colonisation rate of those with and those without R T I was noticed. Children z-5 months of age were colonised in 58 % cases in the R T I - g r o u p and in 8 % cases in the n o n - R T I group. Children 6-47 months of age had a colonisation rate of 70 ~o in the R T I group and 43 % in the group
26
T. E J L E R T S E N
ET AL.
Table I Prevalence of B. catarrhalis in patients with and without respiratory tract infection at the time of admission to hospital and on discharge from hospital With RTI Admission Patients B. catarrhalis no. %
Age Neonates, < I month I n f a n t s , 1-47 m o n t h s C h i l d r e n , 4 - 1 5 years W o m e n , I 6 - 4 5 years
II I47
Patients no.
o 68
Discharge B. catarrhalis % n.e.
88
42
n.e. n.e.
n.e. n.e. Without RTI
Neonates, < r month I n f a n t s , 1-47 m o n t h s C h i l d r e n , 4 - I 5 years. W o m e n , I 6 - 4 5 years
625 I I9 57 56I
o 36 7 I
499 56
o 30 n.e. n.e.
n.e. = n o t e x a m i n e d .
Table II Isolation of B. catarrhalis in relation to age and respiratory tract infection (R TI) on admission to hospital With RTI
Age < I month I-- 5 m o n t h s
6-47 months 4 - 9 years IO--I5 years I 6 - 4 5 years
Patients no.
Without RTI
B. catarrhalis %
II 19
0 58
i28
70 n.e. n.e. n.e.
Patients no. 91
B. catarrhalis %
24
0 8
95 32 25 56I
43 9 4 I
n.e. = n o t examined.
without R T I (Table II). Children recovering from an R T I had a significant fall in colonisation rate at the time of discharge from hospital 4 days later, 42"I % on discharge versus 68"0 % on admission (P < o'ooI). Children without R T I had a colonisation rate on discharge similar to that on admission, 30"4 % versus 36"I %, (P = 0"29) (Table I). Patients, 4-15 years of age, without R T I were colonised in 7 % cases (4/57 children). Women without R T I , I6-45 years of age, were colonised in the throat in I'1% cases (6/56I women). None was colonised in the vagina. In children 1-47 months of age, comparison of simultaneously obtained tracheal secretions and throat swabs was possible in 2I of the patients with R T I (Table III). T h e finding of B. catarrhalis in tracheal secretions could be
B. catarrhalis in children and adults
27
Table I I I Isolation of B. catarrhalis from tracheal secretions and throat swabs
taken simultaneously at the time of admission of patients with respiratory tract infection Throat swab Growth of No growth of
Tracheal secretion
B. catarrhalis
B. catarrhalis
Total
I4 o
x 6
I5 6
14
7
2 I*
Growth of B. catarrhalis No growth of B . catarrhalis Total
* Agreement in 20 of 2I pairs of samples = 95.2 %.
Table IV Isolation of B. catarrhalis in paired admission and discharge samples
from patients with respiratory tract infections who did not receive antibiotic treatment Tracheal secretions on discharge Growth of No growth of
Tracheal secretions on admission
Growth of B . catarrhalis No growth of B. catarrhalis Total
B . catarrhalis
B. catarrhalis
23 2 25
I7 I3 30
Total 4o 15 55*
* Agreement in 36 of 55 pairs of samples = 65'5 %.
reproduced from the throat swab in I 4 / I 5 (93"3 %) cases. B. catarrhalis was not isolated from any throat swabs when the organism was not found in accompanying tracheal secretions. F r o m 72 patients with R T I , tracheal secretions on admission as well as a throat swab on discharge were obtained with a mean interval of 4 days. Excluding patients who received antibiotic treatment during their stay in hospital, admission and discharge samples were obtained from 55 patients (Table IV). In these pairs of samples, agreement was found in 65'5 %. Of 40 patients, I7 apparently lost their colonisation with B. catarrhalis without the influence of antibiotic treatment as they recovered from their R T I . A total of 60 patients without R T I gave throat swabs on admission and on discharge with a mean interval of 3"5 days. Excluding 15 patients who received antibiotic treatment during their stay in hospital, paired admission and discharge samples were obtained from 45 patients (Table V). Identical findings with regard to B. catarrhalis were noted in 40/45 (88"9%) paired samples. Streptococcus pneumoniae and Haemophilus influenzae also were sought in the tracheal secretions from the R T I group. B. catarrhalis was the most frequently isolated bacterium of the three with an isolation rate of 68 %. H. influenzae was isolated at a rate of 55"5 % while S. pneumoniae was isolated at a rate of 39"3 %. B. catarrhalis was isolated in pure growth from 22"6 % children with R T I .
T. EJLERTSEN ET AL.
28
Table V Isolation of B. catarrhalis from paired admission and discharge samples from patients without respiratory tract infections who did not receive antibiotic treatment Throat swabs on discharge Growth of No growth of B. catarrhalis B. catarrhalis Throat swabs on admission
Growth of B. catarrhalis No growth of B. catarrhalis Total
I4 I I5
4 26 3o
Total I8 27 45*
* Agreement in 40 of 45 pairs of samples = 88"9%. S e a s o n a l variation T h e isolation rate in relation to the time of year was investigated in the oneyear study (children 1-47 months of age). T h e problem was elucidated separately in those with and those without R T I . T h e isolation rate of B. catarrhalis in patients with R T I was 73 % in the winter and 67 % in the summer, while the isolation rate in those without R T I was 37 % in the winter and 4o % in the summer. Beta-lactamase production Beta-lactamase production was found in 84 % strains of B. catarrhalis. N o statistically significant difference was noted between the rates of betalactamase production in strains isolated from patients with R T I (88 %) and those without R T I (76 %), (P = 0.08). Relation to infection with respiratory syncytial virus Respiratory syncytial virus (RSV) was found in IO/57 patients with R T I . Since the prevalence of B. catarrhalis was 68 % in the R T I group, seven cases of infection with B. catarrhalis would be expected, and were found, within the RSV-positive group. T h e n u m b e r of RSV-positive cases, however, was too small to permit any evaluation of the correlation between R S V infection and colonisation with B. catarrhalis. T o find a I o % (or even 2 o % ) higher colonisation rate of B. catarrhalis in RSV-positive cases, with a type I error of 5 % and a type II error of 2o %, would require 328 (or 76) RSV-positive cases. Discussion This study demonstrated that the colonisation rate of B. catarrhalis correlates with age and respiratory tract infections. B. catarrhalis is not usually an inhabitant of the pharynx of adults and is rarely found in children < I month of age. In the age group of 1-47 months, children with healthy respiratory tracts were colonised with a prevalence of 36 %. This corresponds to the rate of 30-4 ° % reported by others.l-3 There was, however, a significantly increased colonisation rate in children with upper and lower R T I . In this study, the prevalence o f B . catarrhalis in children aged 1-47 months with R T I was found
B. catarrhalis in children and adults
29
to be 68 %. T h e relation to R T I has also been noted by Brorson et al. 1 who isolated B. catarrhalis from 66 % children with longstanding cough and from 28 % children with healthy respiratory tracts. Colonisation with B. catarrhalis is not static. M61stad et al. 4 found a high rate of acquisition and elimination of B. catarrhalis in children. Nearly IOO % children in their study group were colonised at various times during a period of I month, with a mean prevalence rate of 62 %. It may therefore be assumed that, in the age group of 1-47 months, children are frequently exposed to B. catarrhalis. Children with R T I may be prone to be colonised as long as their respiratory epithelium is damaged. T h e y thereby reveal a high colonisation rate during the period with infectious mucosal damage. Subsequently, they eliminate B. catarrhalis as they recover from their R T I , their colonisation rate returning to that of the n o n - R T I children. In our study, only patients without R T I in the older age groups were examined and found to have low colonisation rates. A high colonisation rate might also have been found in relation to R T I in these groups. Brorson et al? found a colonisation rate of 66 % in children aged x-9 years with R T I , and Schal~n et al. TM found B. catarrhalis in 55 % adults with longstanding cough. Likewise, S6derstr6m et al. 13 found a colonisation rate of I5 % in healthy children, 6-x3 years of age, 27 % in patients with non-bacterial R T I , and 37 % in patients with bacterial R T I . T h e low rate of 1% for pharyngeal carriage in adults with healthy respiratory tracts found in our study corresponds well with that of other reports, a°' 14 None of the women was colonised in the vagina. Furthermore, transmission from mother to child was not observed during birth or the first days of life. Seasonal variations have been noted in the prevalence of B. catarrhalis. '5 In the present study, seasonal variations were examined separately for those with and those without R T I . T h e R T I group had a constantly high colonisation rate throughout the year, while the n o n - R T I group had a constantly low colonisation rate throughout the year. T h e increased number of B. catarrhalis strains isolated by others in the winter may possibly be explained by an overrepresentation of samples collected in the winter when episodes of R T I are common. It has been suggested that preceding RSV or mycoplasmal infections play a role in the B. catarrhalis colonisation rate. 1° Studies of B. catarrhalis colonisation in relation to RS virus infections, however, have not been able to establish such a connection. T M Production of fl-lactamase was found in 84 % strains of B. catarrhalis. Such high prevalence of fl-lactamase production is a recent finding in D e n m a r k ? 7,18 Production of fl-lactamase by strains of B. catarrhalis was seen for the first time and simultaneously in I976-x977 in Europe and the U.S.A. Strains producing fl-lactamase then spread rapidly in the U.S.A. reaching a proportion of approximately 75 % in x98 I. In Europe, there was a delay of 4 years before the same increase was observed. Denmark and Sweden have been the two northern European countries with the longest delay of about 7 year s.17' 18 Now, however, all countries seem to have fl-lactamase production in 80-90 % strains of B. catarrhalis. In conclusion, B. catarrhalis does not belong to the normal flora of the respiratory tract of neonates. Isolation of B. catarrhalis in this age group should therefore command attention, since the organism may act as a virulent
T. EJLERTSEN E T AL.
30
p a t h o g e n in n e o n a t e s . 6-s I n c h i l d r e n 1 - 4 7 m o n t h s o f age, B . catarrhalis b e l o n g s to t h e n o r m a l flora w i t h a p r e v a l e n c e o f a p p r o x i m a t e l y 4 0 % in h e a l t h y c h i l d r e n a n d 70 % in c h i l d r e n w i t h r e s p i r a t o r y t r a c t i n f e c t i o n s . T h e h i g h c o l o n i s a t i o n rate in c h i l d r e n w i t h R T I m a k e s it difficult to d e t e r m i n e w h e t h e r B . catarrhalis has a p a t h o g e n i c role in t h e i n i t i a t i o n or in the m a i n t e n a n c e o f t h e i n f e c t i o n . T h e p r o b l e m b e c o m e s e v e n m o r e c o m p l i c a t e d b y t h e fact t h a t B. catarrhalis is v e r y o f t e n a c c o m p a n i e d b y H . influenzae o r S. pneumoniae o r b o t h . I n g e n e r a l , B . catarrhalis is n o t h e l d to b e a v i r u l e n t p r i m a r y p a t h o g e n in this age g r o u p . F u r t h e r m o r e , t h e o r g a n i s m does n o t s e e m to b e l o n g to t h e n o r m a l flora o f c h i l d r e n > 4 y e a r s o f age o r o f a d u l t s w i t h h e a l t h y r e s p i r a t o r y tracts. (This work was supported by grants from N o r t h Jutland Research Council, Aalborg, County Christmas Lottery as well as the August Petersen and Thorkild Petersen Grant. We thank the staff of the Maternity D e p a r t m e n t of Aalborg Hospital for their enthusiastic assistance in collecting the relevant specimens from women admitted to their department in labour. D r Brita Bruun is thanked for a critical review of this work.) References
i. Brorson J-E, Malmvall B-E. Branhamella catarrhalis and other bacteria in the nasopharynx of children with longstanding cough. ScandJ Infect Dis 1981; 13: 111-113. 2. Ingvarsson L, Lundgren K, Ursing J. The bacterial flora in the nasopharynx in healthy children. Acta Otolaryngol 1982; Suppl. 386: 94-96. 3. Vaneechoutte M, Verschraegen G, Claeys G, Weise B, Abele AMVD. Respiratory tract carrier rates of Moraxella (Branhamella) catarrhalis in adults and children and interpretation of the isolation of M. catarrhalis from sputum. J Clin Microbiol 199o; 28: 2674-2680. 4. M61stad S, Eliasson I, Hovelius B, Kamme C, Schal6n C. fl-lactamase production in the upper respiratory tract flora in relation to antibiotic consumption: A study in children attending day nurseries. ScandJ Infect Dis I988; zo: 3z9-334. 5. Gunnarsson M, Grip L. Atypical croup and Branhamella catarrhalis. Lancet I98z; ii: i56. 6. Ohlsson A, Bailey T. Neonatal pneumonia caused by Branhamella catarrhalis. Scand ff Infect Dis I985; x7: 225-228. 7. Haddad J, Faou AL, Simeoni U, Messer J. Hospital-acquired bronchopulmonary infection in premature infants due to Branhamella catarrhalis. J Hosp Inf I986; 7: 3oI-3O2. 8. Berg AB, Bartley DL. Pneumonia associated with Branhamella catarrhalis in infants. Pediatr Infect Dis J I987; 6: 569-573. 9. Barren/is M-L, Beran M, Blomberg J, Ejnell H, Jeppsson P-H. Etiological aspects on pseudocroup. Acta Otolaryngol I982; Suppl. 386: 206-208. IO. Schal6n L, Christensen K, Kamme C, Mi6rner H, Pettersson K-I, Schal6n C. High isolation rate of Branhamella catarrhalis from the nasopharynx in adults with acute laryngitis. Scand Infect Dis I98O; I2: 277-280. I I. Balows A, Hausler WJ Jr, Herrmann KL, Isenberg HD, Shadomy HJ (eds). Manual of Clinical Microbiology. 5th ed. American Society for Microbiology. Washington, D.C., 199I. I2. Wulff HR, S chlichting P. Manual for M E D - S TAT. Guide to MED-STAT version 2. I. Copenhagen I988: The ASTRA Group A/S. I3. S6derstr6m M, Hovelius B, Prellner K, Schal6n C. Quantification of nasopharyngeal bacteria for diagnosis of respiratory tract infections in children. Scand J Infect Dis I99O; 22 : 333-337. I4. Verghese A, Berro E, Berro J, Franzus BW. Pulmonary clearance and phagocytic cell response in a murine model of Branhamella catarrhalis infection, ff Infect Dis I99O; I62: I 189-I I92.
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I5. Boyle FM. Georghion PR, Tilse MH, McCormack JG. Branhamella (Moraxella) catarrhalis: Pathogenic significance in respiratory infections. Med J Austr I99I; I54: 592-596 • I6. Korppi M, Leinonen M, Koskela M, Mfikelfi PH, Launiala K. Bacterial coinfection in children hospitalized with respiratory syncyteal virus infections. Pediatr Infect Dis J I989; 8 : 687-692. 17. Ejlertsen T, Schonheyder CH, Thisted E. fl-lactamase production in Branhamella catarrhalis isolated from lower respiratory tract secretions in Danish children: An increasing problem. Infection I99I; I9:328-33 O. I8. Christensen J J, Keiding J, Bruun B. Antimicrobial susceptibility and beta-lactamase characterization of Branhamella catarrhalis strains from i983/i984 and I988. A P M I S I99O; 98: Io39--IO44.
B r a n h a m e l l a c a t a r r h a l i s in c h i l d r e n a n d adults. A s t u d y o f prevalence, time of colonisation, and association with upper a n d l o w e r r e s p i r a t o r y tract i n f e c t i o n s Tove Ejlertsen,* Ebbe Thisted,t Finn Ebbesen,T Bente Olesen~ and Jan Renneberg~ * Department of Clinical Microbiology and ~fDepartment of Pediatrics, Aalborg Hospital, Aalborg, ~ Department of Clinical Microbiology, Hvidovre Hospital, Hvidovre, ~Division of Preventive Microbiology, Statens Seruminstitut, Copenhagen, Denmark Accepted for publication 5 January 1994
Summary The colonisation rate of Branhamella catarrhalis in patients from o to 45 years of age was examined. Of 561 women admitted to hospital in labour, 6 (I %) carried B. catarrhalis in their throats but none carried the organism in their vaginas. None of 534 newborn babies became colonised at birth or during their 5 days' stay in hospital. Neither were lO2 neonates < I month of age in hospital colonised. The maximum colonisation rate during childhood was observed in children 1-48 months of age with 143 of 266 (54%) children colonised. Among children 4-15 years of age, four of 57 (7%) children with healthy respiratory tracts were colonised. Significantly more children with upper or lower respiratory tract infections (RTI) were colonised (68 %) than were children without such infections (36 %), (P < o.ooi). After recovery from RTI, the isolation rate in the R T I group fell to that of the non-RTI group. A seasonal variation in prevalence was not observed. Of all the strains of B. catarrhalis isolated, 84 % produced beta-lactamase.
Introduction Branhamella catarrhalis is a f r e q u e n t inhabitant of the oro- and n a s o p h a r y n x of children o--io years of age with a prevalence reported of approximately 25-35%.1-3 T h e time, m o d e and d u r a t i o n of colonisation as well as the m e c h a n i s m s of elimination o f B. catarrhalis have been poorly elucidated. One s t u d y has shown a high degree of acquisition and elimination of B. catarrhalis strains in the n a s o p h a r y n x of children w i t h o u t the influence of antibiotic treatment3 Sporadic observations have drawn attention to the role of B. catarrhalis in small children with b r o n c h o - p u l m o n a r y infections and tracheitis3 -s T h e organism has also been associated with atypical p s e u d o - c r o u p in preschool children 9 as well as cough and hoarseness in children and adults. 1' 10 T h u s , the presence of B. catarrhalis in respiratory tract infections ( R T I ) in children is certain b u t its role as a p r i m a r y p a t h o g e n remains controversial, * Address correspondence to : D r T. Ejlertsen, Dept. Clinical Microbiology, Aalborg Hospital, D K 9ooo Aalborg, Denmark. o163-4453/94/29oo23 +09 $08.00/0
© 1994 The British Society for the Study of Infection
24
T. EJLERTSEN E T A L .
In order to assess the importance of B. catarrhalis in tracheal secretions and throat cultures it may be helpful to know the expected incidence of colonisation with B. catarrhalis in each age group, and to what extent symptoms of respiratory tract infections (RTI) are associated with colonisation. T h e purpose of this study has been to investigate the transmission of B. catarrhalis from mother to child at birth, the incidence of colonisation during the first days of life, and the prevalence of B. catarrhalis throughout childhood. In children aged 1-47 months the relation to R T I has also been examined. Patients and m e t h o d s Patients
Healthy women admitted to the Maternity Department of Aalborg Hospital in labour and their newborn children were examined prospectively during a period of 4 months from September to December z989. Throat swabs and vaginal swabs were collected from the women before delivery. After birth, throat secretions were collected from their babies. On discharge from the hospital 5 days later, throat swabs were obtained from the babies. Children, aged o-47 months, admitted to the Paediatric Department of Aalborg Hospital, were examined prospectively during a period of I year from June z989 to May z99o. T h e y were divided into two groups according to their symptoms at the time of admission. Patients suffering from upper or lower R T I formed one group. R T I comprised catarrh, acute laryngitis, acute tracheitis, acute bronchiolitis and pneumonia. In the R T I group, tracheal secretions or a throat swab was collected on admission, and a throat swab was collected on discharge. During the period of 5 months from November I989 to March I99O, additional nasopharyngeal aspirates were collected and examined for the presence of respiratory syncytial virus (RSV) in this group. Patients not suffering from R T I (non-RTI), but in hospital for other reasons, formed another group. Throat swabs were obtained at the time of admission and of discharge in this group also. Antibiotic treatment of patients was noted in both groups. T h e last group consisted of outpatients aged 4-I5 years with healthy respiratory tracts when examined at Hvidovre Hospital, Copenhagen, during the period from July to December I992. All studies were approved by the local Ethics Committee, and the patients participated only after giving informed consent. A parent's informed consent was obtained in the case of each of the children. Microbiological methods
All specimens were sent to the local Department of Clinical Microbiology. Tracheal secretions were aspirated through a nasopharyngeal catheter and kept refrigerated for a maximum of 24 hours until they were cultured. Throat swabs were taken with a charcoal cotton-tipped applicator swabbed over the tonsils and palate. T h e tip of the applicator was stored in Stuart's transport medium and refrigerated until it was cultured. Tracheal secretions were centrifuged at z5oo × g for IO min. T h e sediment was recovered with a sterile cotton applicator and cultured on 5 % horse blood agar (streaked with
B. catarrhalis in children and adults
25
Staphylococcus aureus for detection of Haemophilus influenzae), 'chocolate' agar, and a Danish 'GC-chocolate' agar medium (selective for pathogenic Neisseria species) containing polymyxin B, a low concentration of lincomycin, mycostatin, and trimethoprim but without vancomycin (Statens Seruminstitut, Copenhagen, Denmark). This medium permitted all strains of B. catarrhalis observed on 'chocolate agar' and on horse blood agar to be recovered in pure growth without interference from the commensals of the pharynx other than Neisseria-species. Throat swabs were cultured on 5 % horse blood agar and on 'GC-chocolate' agar. All media were incubated aerobically with the addition of 5 % CO2 for 2 days. B. catarrhalis was identified according to colonial morphology, Gram-staining reaction, catalase and oxidase reactions, reduction of nitrate to nitrite as well as inability to produce acid from glucose, maltose, and lactose together with production of DNase. Isolates were tested for beta-lactamase activity by means of a chromogenic cephalosporin (Cefinase, BBL Microbiology Systems, U.S.A.). H. influenzae and Streptococcus pneumoniae were identified from tracheal secretions collected from the R T I group, 1-47 months of age, according to established methods. 11 RSV was identified by means of a direct immunofluorescence test (Imagen ® Respiratory Syncytial Virus, D A K O Diagnostics Ltd., U.K.) Statistical methods
A file was generated in M E D S T A T J ~X~was calculated from 2 x 2 contingency tables while Fisher's exact test (two-tailed) was used for small numbers. Results Characteristics of patients
A total of 1520 patients participated in the study (Table I). Of these, 561 were women admitted in labour with healthy respiratory tracts. T h e y formed the adult group aged z6-45 years. Of their babies, 534 formed the group of newborns examined at birth and of whom 499 were re-examined on discharge 5 days later. T h e neonatal group, aged < z month, consisted of zo2 babies, 9I without R T I and I z with R T I . Children aged 1-47 months formed a group of 266 patients, I I9 without R T I , 56 of whom were re-examined on discharge; and I47 with R T I , 88 of whom were re-examined on discharge. A further group was comprised of 57 outpatients aged 4-I5 years with healthy respiratory tracts. I s o l a t i o n rates o f B r a n h a m e l l a c a t a r r h a l i s
T h e isolation rates of B. catarrhalis were strongly age-dependent. Neonates < z month of age were not colonised with B. catarrhalis, regardless of whether or not they had a respiratory infection (Table I). Overall, z43/266 (54%) children I to 47 months of age were colonised. In this age group, a significant difference between the colonisation rate of those with and those without R T I was noticed. Children z-5 months of age were colonised in 58 % cases in the R T I - g r o u p and in 8 % cases in the n o n - R T I group. Children 6-47 months of age had a colonisation rate of 70 ~o in the R T I group and 43 % in the group
26
T. E J L E R T S E N
ET AL.
Table I Prevalence of B. catarrhalis in patients with and without respiratory tract infection at the time of admission to hospital and on discharge from hospital With RTI Admission Patients B. catarrhalis no. %
Age Neonates, < I month I n f a n t s , 1-47 m o n t h s C h i l d r e n , 4 - 1 5 years W o m e n , I 6 - 4 5 years
II I47
Patients no.
o 68
Discharge B. catarrhalis % n.e.
88
42
n.e. n.e.
n.e. n.e. Without RTI
Neonates, < r month I n f a n t s , 1-47 m o n t h s C h i l d r e n , 4 - I 5 years. W o m e n , I 6 - 4 5 years
625 I I9 57 56I
o 36 7 I
499 56
o 30 n.e. n.e.
n.e. = n o t e x a m i n e d .
Table II Isolation of B. catarrhalis in relation to age and respiratory tract infection (R TI) on admission to hospital With RTI
Age < I month I-- 5 m o n t h s
6-47 months 4 - 9 years IO--I5 years I 6 - 4 5 years
Patients no.
Without RTI
B. catarrhalis %
II 19
0 58
i28
70 n.e. n.e. n.e.
Patients no. 91
B. catarrhalis %
24
0 8
95 32 25 56I
43 9 4 I
n.e. = n o t examined.
without R T I (Table II). Children recovering from an R T I had a significant fall in colonisation rate at the time of discharge from hospital 4 days later, 42"I % on discharge versus 68"0 % on admission (P < o'ooI). Children without R T I had a colonisation rate on discharge similar to that on admission, 30"4 % versus 36"I %, (P = 0"29) (Table I). Patients, 4-15 years of age, without R T I were colonised in 7 % cases (4/57 children). Women without R T I , I6-45 years of age, were colonised in the throat in I'1% cases (6/56I women). None was colonised in the vagina. In children 1-47 months of age, comparison of simultaneously obtained tracheal secretions and throat swabs was possible in 2I of the patients with R T I (Table III). T h e finding of B. catarrhalis in tracheal secretions could be
B. catarrhalis in children and adults
27
Table I I I Isolation of B. catarrhalis from tracheal secretions and throat swabs
taken simultaneously at the time of admission of patients with respiratory tract infection Throat swab Growth of No growth of
Tracheal secretion
B. catarrhalis
B. catarrhalis
Total
I4 o
x 6
I5 6
14
7
2 I*
Growth of B. catarrhalis No growth of B . catarrhalis Total
* Agreement in 20 of 2I pairs of samples = 95.2 %.
Table IV Isolation of B. catarrhalis in paired admission and discharge samples
from patients with respiratory tract infections who did not receive antibiotic treatment Tracheal secretions on discharge Growth of No growth of
Tracheal secretions on admission
Growth of B . catarrhalis No growth of B. catarrhalis Total
B . catarrhalis
B. catarrhalis
23 2 25
I7 I3 30
Total 4o 15 55*
* Agreement in 36 of 55 pairs of samples = 65'5 %.
reproduced from the throat swab in I 4 / I 5 (93"3 %) cases. B. catarrhalis was not isolated from any throat swabs when the organism was not found in accompanying tracheal secretions. F r o m 72 patients with R T I , tracheal secretions on admission as well as a throat swab on discharge were obtained with a mean interval of 4 days. Excluding patients who received antibiotic treatment during their stay in hospital, admission and discharge samples were obtained from 55 patients (Table IV). In these pairs of samples, agreement was found in 65'5 %. Of 40 patients, I7 apparently lost their colonisation with B. catarrhalis without the influence of antibiotic treatment as they recovered from their R T I . A total of 60 patients without R T I gave throat swabs on admission and on discharge with a mean interval of 3"5 days. Excluding 15 patients who received antibiotic treatment during their stay in hospital, paired admission and discharge samples were obtained from 45 patients (Table V). Identical findings with regard to B. catarrhalis were noted in 40/45 (88"9%) paired samples. Streptococcus pneumoniae and Haemophilus influenzae also were sought in the tracheal secretions from the R T I group. B. catarrhalis was the most frequently isolated bacterium of the three with an isolation rate of 68 %. H. influenzae was isolated at a rate of 55"5 % while S. pneumoniae was isolated at a rate of 39"3 %. B. catarrhalis was isolated in pure growth from 22"6 % children with R T I .
T. EJLERTSEN ET AL.
28
Table V Isolation of B. catarrhalis from paired admission and discharge samples from patients without respiratory tract infections who did not receive antibiotic treatment Throat swabs on discharge Growth of No growth of B. catarrhalis B. catarrhalis Throat swabs on admission
Growth of B. catarrhalis No growth of B. catarrhalis Total
I4 I I5
4 26 3o
Total I8 27 45*
* Agreement in 40 of 45 pairs of samples = 88"9%. S e a s o n a l variation T h e isolation rate in relation to the time of year was investigated in the oneyear study (children 1-47 months of age). T h e problem was elucidated separately in those with and those without R T I . T h e isolation rate of B. catarrhalis in patients with R T I was 73 % in the winter and 67 % in the summer, while the isolation rate in those without R T I was 37 % in the winter and 4o % in the summer. Beta-lactamase production Beta-lactamase production was found in 84 % strains of B. catarrhalis. N o statistically significant difference was noted between the rates of betalactamase production in strains isolated from patients with R T I (88 %) and those without R T I (76 %), (P = 0.08). Relation to infection with respiratory syncytial virus Respiratory syncytial virus (RSV) was found in IO/57 patients with R T I . Since the prevalence of B. catarrhalis was 68 % in the R T I group, seven cases of infection with B. catarrhalis would be expected, and were found, within the RSV-positive group. T h e n u m b e r of RSV-positive cases, however, was too small to permit any evaluation of the correlation between R S V infection and colonisation with B. catarrhalis. T o find a I o % (or even 2 o % ) higher colonisation rate of B. catarrhalis in RSV-positive cases, with a type I error of 5 % and a type II error of 2o %, would require 328 (or 76) RSV-positive cases. Discussion This study demonstrated that the colonisation rate of B. catarrhalis correlates with age and respiratory tract infections. B. catarrhalis is not usually an inhabitant of the pharynx of adults and is rarely found in children < I month of age. In the age group of 1-47 months, children with healthy respiratory tracts were colonised with a prevalence of 36 %. This corresponds to the rate of 30-4 ° % reported by others.l-3 There was, however, a significantly increased colonisation rate in children with upper and lower R T I . In this study, the prevalence o f B . catarrhalis in children aged 1-47 months with R T I was found
B. catarrhalis in children and adults
29
to be 68 %. T h e relation to R T I has also been noted by Brorson et al. 1 who isolated B. catarrhalis from 66 % children with longstanding cough and from 28 % children with healthy respiratory tracts. Colonisation with B. catarrhalis is not static. M61stad et al. 4 found a high rate of acquisition and elimination of B. catarrhalis in children. Nearly IOO % children in their study group were colonised at various times during a period of I month, with a mean prevalence rate of 62 %. It may therefore be assumed that, in the age group of 1-47 months, children are frequently exposed to B. catarrhalis. Children with R T I may be prone to be colonised as long as their respiratory epithelium is damaged. T h e y thereby reveal a high colonisation rate during the period with infectious mucosal damage. Subsequently, they eliminate B. catarrhalis as they recover from their R T I , their colonisation rate returning to that of the n o n - R T I children. In our study, only patients without R T I in the older age groups were examined and found to have low colonisation rates. A high colonisation rate might also have been found in relation to R T I in these groups. Brorson et al? found a colonisation rate of 66 % in children aged x-9 years with R T I , and Schal~n et al. TM found B. catarrhalis in 55 % adults with longstanding cough. Likewise, S6derstr6m et al. 13 found a colonisation rate of I5 % in healthy children, 6-x3 years of age, 27 % in patients with non-bacterial R T I , and 37 % in patients with bacterial R T I . T h e low rate of 1% for pharyngeal carriage in adults with healthy respiratory tracts found in our study corresponds well with that of other reports, a°' 14 None of the women was colonised in the vagina. Furthermore, transmission from mother to child was not observed during birth or the first days of life. Seasonal variations have been noted in the prevalence of B. catarrhalis. '5 In the present study, seasonal variations were examined separately for those with and those without R T I . T h e R T I group had a constantly high colonisation rate throughout the year, while the n o n - R T I group had a constantly low colonisation rate throughout the year. T h e increased number of B. catarrhalis strains isolated by others in the winter may possibly be explained by an overrepresentation of samples collected in the winter when episodes of R T I are common. It has been suggested that preceding RSV or mycoplasmal infections play a role in the B. catarrhalis colonisation rate. 1° Studies of B. catarrhalis colonisation in relation to RS virus infections, however, have not been able to establish such a connection. T M Production of fl-lactamase was found in 84 % strains of B. catarrhalis. Such high prevalence of fl-lactamase production is a recent finding in D e n m a r k ? 7,18 Production of fl-lactamase by strains of B. catarrhalis was seen for the first time and simultaneously in I976-x977 in Europe and the U.S.A. Strains producing fl-lactamase then spread rapidly in the U.S.A. reaching a proportion of approximately 75 % in x98 I. In Europe, there was a delay of 4 years before the same increase was observed. Denmark and Sweden have been the two northern European countries with the longest delay of about 7 year s.17' 18 Now, however, all countries seem to have fl-lactamase production in 80-90 % strains of B. catarrhalis. In conclusion, B. catarrhalis does not belong to the normal flora of the respiratory tract of neonates. Isolation of B. catarrhalis in this age group should therefore command attention, since the organism may act as a virulent
T. EJLERTSEN E T AL.
30
p a t h o g e n in n e o n a t e s . 6-s I n c h i l d r e n 1 - 4 7 m o n t h s o f age, B . catarrhalis b e l o n g s to t h e n o r m a l flora w i t h a p r e v a l e n c e o f a p p r o x i m a t e l y 4 0 % in h e a l t h y c h i l d r e n a n d 70 % in c h i l d r e n w i t h r e s p i r a t o r y t r a c t i n f e c t i o n s . T h e h i g h c o l o n i s a t i o n rate in c h i l d r e n w i t h R T I m a k e s it difficult to d e t e r m i n e w h e t h e r B . catarrhalis has a p a t h o g e n i c role in t h e i n i t i a t i o n or in the m a i n t e n a n c e o f t h e i n f e c t i o n . T h e p r o b l e m b e c o m e s e v e n m o r e c o m p l i c a t e d b y t h e fact t h a t B. catarrhalis is v e r y o f t e n a c c o m p a n i e d b y H . influenzae o r S. pneumoniae o r b o t h . I n g e n e r a l , B . catarrhalis is n o t h e l d to b e a v i r u l e n t p r i m a r y p a t h o g e n in this age g r o u p . F u r t h e r m o r e , t h e o r g a n i s m does n o t s e e m to b e l o n g to t h e n o r m a l flora o f c h i l d r e n > 4 y e a r s o f age o r o f a d u l t s w i t h h e a l t h y r e s p i r a t o r y tracts. (This work was supported by grants from N o r t h Jutland Research Council, Aalborg, County Christmas Lottery as well as the August Petersen and Thorkild Petersen Grant. We thank the staff of the Maternity D e p a r t m e n t of Aalborg Hospital for their enthusiastic assistance in collecting the relevant specimens from women admitted to their department in labour. D r Brita Bruun is thanked for a critical review of this work.) References
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I5. Boyle FM. Georghion PR, Tilse MH, McCormack JG. Branhamella (Moraxella) catarrhalis: Pathogenic significance in respiratory infections. Med J Austr I99I; I54: 592-596 • I6. Korppi M, Leinonen M, Koskela M, Mfikelfi PH, Launiala K. Bacterial coinfection in children hospitalized with respiratory syncyteal virus infections. Pediatr Infect Dis J I989; 8 : 687-692. 17. Ejlertsen T, Schonheyder CH, Thisted E. fl-lactamase production in Branhamella catarrhalis isolated from lower respiratory tract secretions in Danish children: An increasing problem. Infection I99I; I9:328-33 O. I8. Christensen J J, Keiding J, Bruun B. Antimicrobial susceptibility and beta-lactamase characterization of Branhamella catarrhalis strains from i983/i984 and I988. A P M I S I99O; 98: Io39--IO44.