PEDIATRICS Vol. 107 No. 2 February 2001, pp. 274-279

Early Diagnosis of Cystic Fibrosis in the Newborn Period and Risk of Pseudomonas aeruginosa Acquisition in the First 10 Years of Life: A Registry-Based Longitudinal Study

Sophia S. Wang, PhD^*, , Stacey C. FitzSimmons, PhD^§, Leslie A. O'Leary, PhD, Michael J. Rock, MD, Marta L. Gwinn, MD, MPH, and Muin J. Khoury, MD, PhD

From the ^* Epidemic Intelligence Service, Division of Applied Public Health Training, Epidemiology Program Office, Centers for Disease Control and Prevention, Atlanta, Georgia, and  Office of Genetics and Disease Prevention, National Center for Environmental Health, Centers for Disease Control and Prevention, Atlanta, Georgia, and ^§ Cystic Fibrosis Foundation, Bethesda, Maryland and  University of Wisconsin Hospital and Clinics, Madison, Wisconsin.

	ABSTRACT

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Objective.  Controlled clinical trial data have suggested that identifying asymptomatic cystic fibrosis (CF) patients through newborn screening improves health outcomes of affected children in the first decade of life. However, it is unclear whether these improvements also include a reduction in risk for bronchial infection, the major determinant of CF morbidity. The authors therefore investigated the association between early CF diagnosis and acquisition of Pseudomonas aeruginosa, the major bronchial pathogen, in the first decade of life.

Methodology.  Longitudinal data on 3625 CF patients diagnosed between 1982 and 1990 and before 36 months of age were ascertained from the National Cystic Fibrosis Patient Registry. We compared P aeruginosa acquisition in the first 10 years of life among 4 groups: EAD (early asymptomatic diagnosis)<6 weeks, by pre/neonatal screening, genotype, family history (n = 157); ESD (early symptomatic diagnosis) (n = 227); LAD (late asymptomatic diagnosis)6 weeks to 36 months (n = 161); and LSD (late symptomatic diagnosis) (n = 3080). P aeruginosa acquisition was determined from yearly sputum and/or bronchoscopy cultures. Children whose CF diagnoses followed meconium ileus or whose cultures were obtained only from nasal samples were excluded from the study.

Results.  Kaplan Meier analyses for P aeruginosa acquisition were conducted for each diagnostic group. Regression models were used to generate adjusted relative hazards with EAD as the referent group. Relative hazards were 0.9 (95% confidence interval [CI]: 0.7-1.2) for ESD, 0.8 (95% CI: 0.6-1.2) for LAD, and 1.0 (95% CI: 0.7-1.2) for LSD. The risk of acquiring P aeruginosa was therefore not significantly different between children diagnosed early, late, asymptomatically, or symptomatically.

Conclusions.  These data suggest that, despite improvements in other health outcomes from newborn screening for CF, early asymptomatic diagnosis of CF does not affect P aeruginosa acquisition.  Key words:  cystic fibrosis, Pseudomonas aeruginosa, neonatal screening, Kaplan Meier, epidemiology.

Cystic fibrosis (CF) is a common, serious genetic disorder affecting an estimated 30 000 individuals in the United States. The mean age at diagnosis of CF is 6 months. Most of the 1000 CF cases diagnosed annually in the United States are diagnosed by the time the patient is 3 years old.¹ The major cause of morbidity and mortality in CF patients is from respiratory infections, which lead to a subsequent decline in pulmonary function. Although antibiotics may be given as prophylaxis, they are more often administered orally or intravenously to control acute episodes of infection.

By the end of the first decade of life, Pseudomonas aeruginosa is the predominant bacterial pathogen colonizing the lower respiratory tract in CF patients.^2,3,4 Although it is not the first pathogen to colonize the lungs of CF patients,⁵ it is the major pathogen contributing to pulmonary disease and decline in pulmonary function.^6-11 Studies have demonstrated significant associations between P aeruginosa colonization and impaired pulmonary function, pulmonary deterioration, and mortality in CF patients.^12,13 Once acquired, P aeruginosa is difficult to eradicate,¹⁴ often persisting until pulmonary failure and death.^5,6,8,11

Respiratory failure in CF patients is a consequence of lung damage resulting from intense inflammatory responses.^3,10,11,15 Therefore, although P aeruginosa colonization is regarded as a leading contributor to death in CF patients in combination with lung inflammation and lung obstruction,^6,16 it is the resulting pulmonary insufficiency and respiratory failure that ultimately lead to morbidity and mortality in CF patients.^5,8

A recent clinical trial in Wisconsin demonstrated improvements in nutritional status, measured by height and weight, in children identified with CF through newborn screening, compared with children identified by standard means (usually with the onset of symptoms).^17-19 These findings suggested that newborn screening may lead to improved long-term health outcomes in children with CF. Better nutrition made possible by early diagnosis and treatment might also enable children with CF to delay acquisition of P aeruginosa, and remain free from infection with this ubiquitous organism as persons without CF are able to do. However, the same trial recently tested the hypothesis that P aeruginosa would be accelerated with early diagnosis attributable to increased interaction with older CF patients and increased exposure.¹⁴ The trial did not find that early diagnosis led to a delay or acceleration in P aeruginosa acquisition,^14,19 but because of its limited sample size, it may not have had the power to detect a statistical difference. As P aeruginosa pathogenesis causes substantial morbidity and mortality in CF patients, any effect on P aeruginosa acquisition in children with CF would be significant.

Using data from CF patients in the national Cystic Fibrosis Foundation Patient Registry, we attempted to determine whether early asymptomatic diagnosis of CF is associated with acquisition of P aeruginosa in the first 10 years of life. The purpose of newborn screening is not only to identify newborns with CF, but also to diagnose CF before clinical symptoms present. In this study, early asymptomatic diagnosis (diagnosis before 6 weeks of age) is used to approximate a diagnosis from newborn screening.

	METHODS

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Study Population

The study population for this longitudinal study included approximately 4000 children with CF who were diagnosed between 1982 and 1990, registered in the Cystic Fibrosis Foundation (CFF) Patient Registry, and seen at 1 of the 111 accredited CF care centers in the United States and followed for up to 10 years. The CFF supports and accredits CF care centers nationwide. These centers provide a national network of specialized care for persons with CF and offer comprehensive diagnosis and treatment, as well as participation in clinical trials of experimental therapies. The CFF has sponsored the patient registry since 1966,²¹ requiring all CFF-accredited care centers to complete standardized questionnaires for all patients seen in their centers.²¹ The 1998 Annual Report of the CFF National Patient Registry reported 21 044 individuals with CF enrolled in the registry or approximately 91% of the estimated 23 000 CF patients under care in the United States.¹ Our analysis included children diagnosed with CF within 36 months of age. Only children with annual P aeruginosa cultures obtained by bronchoscopy or from sputum samples were included. Children who presented with meconium ileus, or whose P aeruginosa cultures were obtained only from nasal samples, were excluded.

CF Diagnosis

Diagnosis with CF was confirmed at CF care centers by sweat test, or by DNA analysis. We grouped children into 4 categories: early asymptomatic diagnosis (EAD), early symptomatic diagnosis (ESD), late asymptomatic diagnosis (LAD) and late symptomatic diagnosis (LSD). Early diagnosis was defined as diagnosis before 6 weeks of age, and late diagnosis was defined as diagnosis between 6 weeks and 36 months of age. We selected 6 weeks as the threshold for early diagnosis because it is considered standard for newborn screening programs. Asymptomatic diagnosis is defined as diagnosis by family history, genotype, prenatal diagnosis (chorionic villus sampling, amniocentesis), or neonatal screening. Symptomatic diagnosis is defined as diagnosis because of clinical presentation with acute or persistent respiratory symptoms, failure to thrive or malnutrition, steatorrhea, abnormal stools, malabsorption, electrolyte imbalance, nasal polyps or sinus disease, rectal prolapse, or liver disease. In addition to diagnostic information, the National CF Patient Registry includes demographic, health care indicators, microbiology, lung function, anthropometric, and health status information. Methods for collection of this data are described elsewhere.²¹ Characteristics specifically of interest for this study included P aeruginosa cultures, method of diagnosis, age at diagnosis, race, gender, acute exacerbation, pancreatic status, year of birth, and state of birth.

Statistical Methods

Univariate and stratified analyses were conducted for the cohort at baseline, which was defined as a CF patient's first visit to a CF care center. Statistical significance was set at P < .05, two-sided. All analyses were performed with SAS 6.12 for Windows (SAS Institute, Cary, NC). There was sufficient power (>80%) to detect decreases or increases in risk for P aeruginosa acquisition of at least 0.8 or 1.2.

Colonization rates for P aeruginosa by diagnostic category (EAD, ESD, LAD, LSD) were assessed longitudinally, from time of diagnosis with CF to 10 years of age. Colonization of P aeruginosa was defined by a positive culture from sputum or bronchoscopy. It should be noted that between 1982 and 1990, only 85 cultures by bronchoscopy were obtained to identify P aeruginosa acquisition; the remainder and therefore vast majority of cultures included in this study to measure colonization and acquisition were ascertained from sputum. The proportion of patients colonized with P aeruginosa was calculated by age; crude and adjusted odds ratios (ORs) were calculated at baseline and at 1, 6, and 10 years of age. Adjusted ORs were calculated by multivariate logistic regression, adjusting for race, gender, acute exacerbation, pancreatic insufficiency, year of birth, and state of birth (dichotomized into states universally screening for CF and states who are not). All covariates were dichotomized in the model except year of birth, which was categorized into tertiles (1979-1984, 1985-1987, 1988-1990). Year of birth was originally divided into quartiles as seen in Table 1; however, because of the small number of individuals born between 1979 and 1981, they were combined with individuals born between 1982 and 1984. Acute exacerbation was defined as hospitalization or home-based course of intravenous antibiotic therapy for respiratory illness. Adjusted ORs with 95% confidence intervals (CIs) for P aeruginosa colonization are reported for 1, 6, and 10 years of age.

The data were also analyzed with Kaplan Meier and Cox proportional hazards regression models, with age as the time scale and time to P aeruginosa acquisition (in years) as the outcome. P aeruginosa acquisition for each CF patient was defined as the first documented positive culture of P aeruginosa by sputum or bronchoscopy. Relative hazards were adjusted for race, gender, acute exacerbation, pancreatic status, year of birth, and state of birth. All covariates were categorized as in the logistic regression models. Adjusted relative hazards with 95% CIs for P aeruginosa acquisition are reported.

	RESULTS

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Table 1 displays demographic and clinical characteristics of all children included in the study at their baseline visit. The median ages of diagnoses were 3.6 weeks for EAD, 4.2 weeks for ESD, 2.8 months for LAD, and 6 months for LSD. There were no significant differences among the 4 diagnostic groups in gender, ethnicity, or F508 mutation status. However, there were significant differences among the groups in race, pancreatic status, height and weight, year of birth, P aeruginosa colonization, and state of birth (Table 1). Despite significant differences in height and weight among the 4 diagnostic groups at baseline, they were not associated with P aeruginosa acquisition and their inclusion of height and weight in multivariate models did not affect the results; they were therefore not included in the final multivariate model.

Logistic regression models were used to determine significant differences in colonization with P aeruginosa between the 4 diagnostic groups while controlling for confounders. Logistic regression models controlling for race, gender, acute exacerbation, pancreatic status, year of birth, and state of birth demonstrated no differences between the 4 diagnostic groups with regards to P aeruginosa colonization at 1, 6, or 10 years of age (Table 2). Individuals with at least 1 reported acute exacerbation were consistently more likely to be colonized with P aeruginosa.

To assess whether early diagnosis was associated with the acquisition of P aeruginosa, we performed a Kaplan Meier analysis for P aeruginosa acquisition (defined as the first visit in which a culture is positive for P aeruginosa) for the 4 diagnostic categories by age (Fig 1). A Cox proportional hazard model adjusting for race, gender, acute exacerbation, pancreatic status, year of birth, and state of birth did not yield significant differences between relative hazards for P aeruginosa acquisition in ESD, LAD, or LSD groups compared with the EAD group (Table 3). Significantly increased risk of P aeruginosa acquisition was associated with at least 1 acute exacerbation (OR 1.6; 95% CI: 1.5-1.8), being born in a later cohort (1988 and beyond) (OR 1.4; 95% CI: 1.3-1.4), and being born in a state not universally screening for CF (OR 1.5; 95% CI: 1.2-1.9).

View larger version (69K): [in this window] [in a new window]   Fig. 1.   Kaplan Meier analysis of Pseudomonas acquisition in CF patients measured through sputum and/or bronchoscopy, from ages 1 through 10 years, by diagnosis (excluding patients with meconium ileus).

	DISCUSSION

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As P aeruginosa is the major organism contributing to respiratory tract infection and associated morbidity and mortality in persons with CF, the goal of this study was to determine whether early diagnosis of CF by newborn screening delays P aeruginosa acquisition, or accelerates acquisition because of increased interaction with older CF children. This observational study of patients from the CFF Patient Registry did not find significant differences between the 4 diagnostic groups (EAD, ESD, LAD, and LSD) in P aeruginosa acquisition.

Use of the CFF Patient Registry provided many benefits to this study. This large registry offered sufficient sample size and power to examine whether P aeruginosa acquisition was delayed in children who were diagnosed with CF asymptomatically and before 6 weeks of age. Despite the exclusion criteria imposed, over 3600 patients were included in the analyses. Furthermore, longitudinal analysis was possible to assess P aeruginosa acquisition with adjustment for possible confounding factors. All multivariate analyses were adjusted for year of birth because of concern that improving treatments over time could lead to a cohort effect. State of birth was also adjusted for in the multivariate analysis, dichotomizing between states with newborn screening programs for CF (Wisconsin, Wyoming, Colorado) and the remaining states to control for any differences in treatments in the newborn populations in these states. It is possible that children diagnosed in states with newborn screening programs for CF may have had differences in treatment, possibly affecting P aeruginosa acquisition or colonization, which is the outcome of interest.

This study found that children who were born more recently in a state without newborn screening for CF and who had at least 1 acute exacerbation were at significantly increased risk for P aeruginosa acquisition. The association between recent births and P aeruginosa acquisition may be attributable to the increasing ability of the clinics to obtain cultures from newborns who normally do not have the ability to produce sputum for culture. The association between children from states without universal screening for CF and P aeruginosa acquisition is unclear because universal screening was initiated midway through the inclusion period (1982 to 1990). The association between pancreatic status (insufficiency) and P aeruginosa acquisition is plausible and may be serving as a proxy for F508 mutational status, which has been demonstrated to be involved in bacterial colonization. Lastly, the association between acute exacerbation and P aeruginosa is also a plausible one. Children reporting >1 acute exacerbations at the time of their annual examinations are not only more susceptible to infection because of their respiratory complications, but they may also have increased exposure to organisms from their hospitalization, leading to subsequent detection of colonization at their next CF clinic visit. However, it is unclear whether acute exacerbations preceded P aeruginosa colonization or whether colonization led to the documented hospitalization or home-based course of intravenous antibiotic therapy for respiratory illness.

There are several issues that must be considered when interpreting these findings. First, this was an observational study and there were unmeasured factors that cannot be controlled for as they might be in a controlled clinical trial such as access to care. Other biases include possible selection bias; for example, although the registry includes most CF patients under care in the United States, those who are not enrolled in the clinics may be older in age and healthier in general.²¹ Excluding persons who remain asymptomatic for >3 years may also have biased our analysis to include a sicker cohort. Nevertheless, it is important to note these biases would be minimal because the majority of CF patients are diagnosed by 3 years of age. In clinical practice, the healthiest nonsputum- producing patients may be less likely to have cultures, again biasing the cohort to sicker patients and higher P aeruginosa rates. This bias would apply to the entire cohort, equally to each diagnostic category; however, the sicker referent group would make differences between the groups more difficult to identify.²¹

There is no standard method for detecting P aeruginosa in lower airways in this registry; although potential for misclassification was reduced by excluding cultures from nasal samples. The majority of cultures were obtained from sputum with a minimal number of cultures obtained by bronchoscopy. Methods for ascertaining culture were limited to culture obtained by sputum, from throat/nasal swab, or by bronchoscopy. Therefore, for this study, culture from sputum was the best available method for identifying lower respiratory infection. Furthermore, not every patient was cultured for P aeruginosa in any given year. Therefore, logistic regression models at specific age groups do not include all eligible children in the population. Estimates of risk calculated longitudinally are therefore more reliable than estimates from cross-sectional data, as all children were included for this analysis. In this analysis, P aeruginosa acquisition was defined as the first positive culture documented in the registry. As cultures were not necessarily attained every year, there is potential for delayed identification of acquisition, possibly resulting in a more conservative estimate of risk for the overall time period.

The Wisconsin clinical trial compared P aeruginosa acquisition of patients diagnosed through neonatal screening and treated in early infancy with those identified by standard diagnostic methods. Although they found no differences in acquisition between the diagnostic categories, there were differences when stratified by the type of CF care center. A significant difference was observed with earlier acquisition of P aeruginosa in an urban CF care center where there was more opportunity for social interactions between younger CF patients and older CF patients.¹⁴ Subsequent analyses further identified the use of aerosols (aerosolized saline, bronchodilators, antibiotics, or DNase) to be the significant risk factor for P aeruginosa acquisition.²²

Despite the limitations discussed, the CFF Patient Registry cohort offers unique opportunities to longitudinally investigate P aeruginosa acquisition in CF patients. The results presented in this study are consistent with those observed in the Wisconsin Clinical Trial, where acquisition for P aeruginosa was not associated with early diagnosis of CF. Policy decisions regarding newborn screening for CF should include P aeruginosa acquisition among the health outcomes to be considered.

	FOOTNOTES

Dr FitzSimmons is now with the Pharmaceutical Research and Manufacturers of America.

Dr Wang is now with the National Institutes of Health.

Received for publication Jan 7, 2000; accepted May 22, 2000.

Reprint requests to (S.S.W.) Division of Cancer Epidemiology and Genetics, National Cancer Institute, NIH 6120 Executive Blvd, Rockville, MD 20854. E-mail: wangso{at}mail.nih.gov

	ABBREVIATIONS

CF, cystic fibrosis; CFF, Cystic Fibrosis Foundation; EAD, early asymptomatic diagnosis; ESD, early symptomatic diagnosis; LAD, late asymptomatic diagnosis; LSD, late symptomatic diagnosis; CI, confidence interval; OR, odds ratio.

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