| Abstract|| |
Context: Chlamydophila pneumoniae (C. pneumoniae) is an emerging infectious agent with a spectrum of clinical manifestations including lower and upper respiratory tract infections. Aims: To investigate the role of C. pneumoniae in community-acquired lower respiratory tract infections (LRTIs) in children using serological tests. Settings and Design: Two hundred children, age 2 months to 12 years, hospitalized for community-acquired LRTIs were investigated for C. pneumoniae etiology. Materials and Methods: We investigated 200 children hospitalized for community-acquired LRTIs, using ELISA for detecting anti-C. pneumoniae IgM and IgG antibodies. The demographic, clinical and radiological findings for C. pneumoniae antibody positive and C. pneumoniae antibody negative cases were compared. Statistical Analysis Used: Data analysis was performed by Chi-square test and Fisher's exact tests using Epi Info (2002). Results: Clinical and radiological findings in both the groups were comparable. Serological evidence of C. pneumoniae infection was observed in 12 (6%) patients; specific IgM antibodies were detected in 11 (91.67%; specific IgG antibodies in 1 (8.33%) patients, while 4-fold rise in C. pneumoniae IgG antibody titers were noted in none of the patients. Conclusions: C. pneumoniae has a role in community-acquired LRTIs, even in children aged < 5 years. Serological detection using ELISA would enable pediatricians in better management of C. pneumoniae infections.
Keywords: Community acquired pneumonia, Chlamydophila pneumoniae, respiratory tract infections
|How to cite this article:|
Kumar S, Saigal SR, Sethi GR. Detection of IgM and IgG antibodies to Chlamydophila pneumoniae in pediatric community-acquired lower respiratory tract infections. Indian J Pathol Microbiol 2011;54:782-5
|How to cite this URL:|
Kumar S, Saigal SR, Sethi GR. Detection of IgM and IgG antibodies to Chlamydophila pneumoniae in pediatric community-acquired lower respiratory tract infections. Indian J Pathol Microbiol [serial online] 2011 [cited 2019 Jun 16];54:782-5. Available from: http://www.ijpmonline.org/text.asp?2011/54/4/782/91501
| Introduction|| |
Lower respiratory tract infections (LRTIs) are a common cause of morbidity and mortality among young children, an overwhelming majority occurring in developing countries. The causative organisms of LRTIs are believed to be mainly respiratory viruses, Streptococcus pneumoniae and Haemophilus influenzae in young children and S. pneumoniae and Mycoplasma pneumoniae in older children. 
Respiratory infection with C. pneumoniae occurs worldwide and in all age groups.Few data are available with regard to C. pneumoniae in pediatric age groups. Some studies have recently shown that M. pneumoniae and C. pneumoniae seem to play a more significant role than previously thought as causes of LRTIs in children of all ages. 
Originally within the genus Chlamydia four species were recognized: C. picorum, C. psittaci, C. pneumoniae, and C. trachomatis and all except C. picorum have been associated with human disease. Later on, the taxonomy of family Chlamydiaceae was revised in 1999 on the basis of genomic studies of these organisms including analysis of 16S and 23S rRNA gene sequences and the family has been divided into two genera, Chlamydia and Chlamydophila. Chlamydia trachomatis was retained in the genus Chlamydia, but C. psittaci and C. pneumoniae were transferred to the new genus, Chlamydophila0.
Since it is difficult to detect C. pneumoniae in clinical practice, specific etiological diagnosis is established only in a minority of cases. The detection of antibodies in paired serum samples has been considered the standard laboratory diagnostic method, but PCR has recently been found to be useful to rapidly detect this pathogen in respiratory secretions. 
The aim of this study was to investigate the role of C. pneumoniae in community-acquired LRTIs in Indian children, using C. pneumoniae ELISA.
| Materials and Methods|| |
This was a prospective, single-center study designed to evaluate the incidence of C. pneumoniae infection in hospitalized children with community-acquired LRTIs and the sampling strategy and sample size estimation methodology adopted was based on prevailing estimated data on the prevalence of this infection and statistical considerations.
A total of 200 patients were enrolled comprising of newly clinically diagnosed cases of acute LRTI, admitted to the pediatrics units of Lok Nayak Hospital and Maulana Azad Medical College, New Delhi, India. Protocols employed in this study were approved by the Ethical committee of Maulana Azad Medical College and associated hospitals, New Delhi, India, and the work undertaken conformed to the provisions of the Declaration of Helsinki in 1995 (as revised in Edinburgh 2000).
Selection of Cases
Previously healthy male and female children who had been hospitalized for signs and symptoms of community-acquired LRTIs and fulfilling the following criteria were included in this study: children aged 2 months to 12 years; presence of cough and fever with breathlessness of less than 30 days duration; increased respiratory rate (with or without features of respiratory distress) on examination; presence of signs of consolidation or bronchopneumonia with or without wheeze on auscultation; radiological findings suggestive of consolidation; bronchopneumonia or interstitial infiltrates with or without hyperinflation.
A newly diagnosed case of acute LRTI comprises of a child with less than 4 weeks history of LRTI symptoms and is within one week of admission in the pediatric ward.
Hospital acquired pneumonia, i.e. pneumonia that developed 72 h after hospitalization or within 7 days of discharge.
Enrollment and evaluation of patients
Written informed consent was taken from the parents or legal guardian of infants (aged 2 months to 1 year) and older children (aged 13 months to 12 years) before enrolling them in the study and collecting clinical specimens, in conformation with standard bio-ethical norms as per provisions of the Declaration of Helsinki in 1995 (as revised in Edinburgh 2000). Detailed history and clinical examination was performed on admission, and chest X-rays were conducted in all children, the details of which were noted in a pre-designed performa.
One to two milliliter blood specimens were collected from children on enrollment (acute phase) for Hb, TLC, DLC, and ELISA for IgM and IgG antibodies against C. pneumoniae. The children's wards/attendants were given written instructions to report back with their child after 4 weeks for obtainment of a second serum sample (convalescent phase) to investigate for convalescent stage antibodies to C. pneumoniae0.
ELISA for IgM/IgG antibody to Chlamydophila pneumoniae
Commercially available ELISA (enzyme-linked immunosorbent assay) technique-based kit (CALBIOTECH INC,CA,91978, USA) was employed for detecting IgM and IgG antibodies in sera of patients. Tests were performed and results interpreted according to manufacturer's instructions. The cut-off value was calculated by multiplying the mean optical value of the calibrator with the calibration factor (mentioned on the calibrator vial in kit). The antibody index of the test sample was determined by dividing the optical density value of each sample by the cut off value.
Commercially available M. pneumoniae ELISA-based kit (Novatec Immunodiagnostica GmbHD, Germany) was used for detection of IgM antibodies to M. pneumoniae. Serological tests were performed and interpreted according to the manufacturer's instructions.
Data analysis was performed using Epi Info 2002. Chi-square test and Fisher's exact test were applied to determine any difference between the proportions. A P value less than 0.05 was considered as statistically significant.
| Results|| |
There were 127 (63.5%) males and 73 (36.5%) females in the 200 cases investigated. The presence of C. pneumoniae antibody was higher in 10 (7.87%) males than in 2 (2.74%) females, though this difference was statistically non-significant (P value 0.22) [Table 1]. In all, 129 (64.5%) children were aged 0 to 1 years (Group 1), 47 (23.5%) aged 2 to 5 years (Group 2), while 24(12%) children were of age 6-12 years (group 3).The difference in the prevalence of C. pneumoniae antibodies (in diagnostic titer) among cases in the three age groups (Group 1 vs. Group 2 vs. Group 3) was borderline significant (P=0.05) [Table 2].
|Table 1: Sex distribution in 200 children with lower respiratory tract infections|
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Clinical Profile in Chlamydophila pneumoniae Antibody Positive Cases
Two (16.67%) cases had a history of LRTIs; tachpnea was documented in 9 (75%) cases; cough and coryza in 12 (100%) cases; fever in 8 (66.67%), while 4 (33.33%) cases were afebrile. Five (41.67%) cases documented audible/auscutable wheezing and 3 (25%) crepitations [Table 3].
|Table 3: Clinical and laboratory data on 12 children with lower respiratory tract infections with positive Chlamydophila pneumoniae findings|
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Documentation of hyperinflation, infiltration, consolidation, bronchopneumonia, and normal chest X-ray was numerically comparable and differences statistically insignificant in C. pneumoniae antibody positive and antibody negative categories [Table 4].
|Table 4: Radiological findings in children with lower respiratory tract infections|
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Serological evidence of C. pneumoniae infection was observed in 12 (6%) patients; specific IgM antibodies (serum dilution 1:100) in acute phase were detected in 11 (91.67%) patients; specific IgG antibodies in both acute and paired sera in one (8.33%) child though this case failed to document significant (four-fold) rise in antibody titer. All samples were negative for both IgM and IgG antibodies. One (8.33%) specimen documented the presence of anti-M. pneumoniae IgM antibodies in diagnostic titer concurrently with IgM antibodies to C. pneumoniae.
| Discussion|| |
Laboratory tests to detect Chlamydophila infections include isolation in cell culture, direct detection of Chlamydia antigen by direct fluorescent antibody (DFA), enzyme-linked immunosorbant assay (ELISA), by DNA-RNA probes, and polymerase chain reaction (PCR). Micro immunofluorescence is the test of choice, though no technique is ideal. Commercial ELISA's are concordant with microimmunofluorescence, have high throughput, objective endpoints, and are technically accessible, though problems with sensitivity and specificity have persisted. The present study investigated the role of C. pneumoniae community-acquired LRTIs in Indian children by employing the enzyme-linked immunosorbent assay technique.
In this study, differential presence of C. pneumoniae antibodies within male and female children was statistically insignificant (P=0.022). A vast majority of 176 (88%) cases investigated were aged less than 5 years, and in all 129 (64.50%) were aged less than 1 year. Within the three age groups investigated in children, relationship between age group and the presence of C. pneumoniae antibodies (in diagnostic titer) was statistically borderline significant (P=0.05). A relatively higher rate of infection in children below 5 years of age is in minor contrast to western and Indian reports where prevalence was highest in the school going age group.  C. pneumoniae antibodies have been documented in 30-45% school going children and adolescents with acute respiratory disease. 
Wheezing was observed in significant (41.67%) C. pneumoniae antibody-positive cases. C. pneumoniae infection has been known to trigger acute episodes of wheezing in asthma  as otherwise, we could not demonstrate any useful association to predict C. pneumoniae from clinical characteristics, which is in line with other data casting doubt on the sensitivity and specificity of clinical and laboratory features in predicting the etiology of community-acquired pneumonia in children, emphasizing the need for effective laboratory diagnosis. Radiological features in C. pneumoniae antibody positive and antibody negative cases were comparable and no feature was significantly associated with positive C. pneumoniae antibody responses. Literature reports, however, reveal that classic C. pneumoniae respiratory tract infection shows unilateral infiltration.  In a recent study,  which compared radiological features in patients with pneumonia due to M. pneumoniae, C. pneumoniae, and cases where etiology was unknown, a higher and statistically significant proportion of C. pneumoniae positive cases showed bilateral patchy features and also infiltration in comparison to the category of unknown etiology.
Overall, 12 (6%) children documented serological evidence of C. pneumoniae infection, spreading across age groups in children, which falls in line with western reports ,,, Indian studies are scattered, conducted on small populations, and have employed different methods of diagnosis of this infection. ,, One (8.33%) C. pneumoniae antibody positive case simultaneously documented IgM antibodies to M. pneumoniae suggesting probable co-infection. M. pneumoniae and C. pneumoniae co-infections are not rare but their clinical implications are unclear.
In conclusion, our data underlines that C. pneumoniae plays a role in Indian children with community-acquired LRTIs even in children aged less than 5 years.
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Director Professor, Department of Microbiology, Maulana Azad Medical College, New Delhi - 110 002
Source of Support: None, Conflict of Interest: None
[Table 1], [Table 2], [Table 3], [Table 4]