C-Reactive Protein in the Diagnosis, Management, and Prognosis of Neonatal Necrotizing Enterocolitis
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《小儿科》
Pediatrics Obstetrics and Gynecology Radiology, University of Tennessee, Memphis, Tennessee
Department of Mathematical Sciences and Statistics, University of Memphis, Memphis, Tennessee
ABSTRACT
Objective. In this prospective, observational study, we determined whether serum C-reactive protein (CRP) correlated with necrotizing enterocolitis (NEC) stages II and III. We hypothesized that serial CRP measurement if used as an adjunct to abdominal radiographs would improve the identification of infants with NEC.
Methods. Serum CRP level was measured every 12 hours for 3 measurements and, when abnormal, once daily. When clinical signs persisted and the initial abdominal radiographs were abnormal, follow-up radiographs were obtained.
Results. Of 241 infants who were evaluated for gastrointestinal signs, 11 had ileus or benign pneumatosis intestinalis with persistently normal CRP; gastrointestinal manifestations resolved within 48 hours, antibiotics were discontinued in <48 hours, and feedings were restarted early without complications. Fifty-five infants had NEC stages II and III; all had abnormal CRP regardless of their blood culture results. In infants with stage II NEC, CRP returned to normal at a mean of 9 days except in those who developed complications such as stricture or abscess formation.
Conclusions. In infants with suspected NEC, normal serial CRP values would favor aborted antibiotic therapy and early resumption of feedings. CRP becomes abnormal in both stage II and stage III NEC. In infants with NEC, persistently elevated CRP after initiation of appropriate medical management suggests associated complications, which may require surgical intervention.
Key Words: necrotizing enterocolitis C-reactive protein newborn premature
Abbreviations: NEC, necrotizing enterocolitis GI, gastrointestinal CRP, C-reactive protein WBC, white blood cell
Necrotizing enterocolitis (NEC) is a common gastrointestinal (GI) disorder in premature infants and is associated with high morbidity and mortality.1 The initial clinical manifestations of NEC are often nonspecific and indistinguishable from neonatal septicemia and other forms of GI disorders.2 Usually, with progression, diagnosis may be established by abdominal radiographic findings, such as intestinal dilation, pneumatosis intestinalis, portal or hepatic vein gas, pneumoperitoneum, or ascites.3,4 However, some of these radiographic findings are not specific for NEC and may occur in other benign entities. For instance, many premature infants with intolerance to feeding show intestinal dilation and ileus.2 Also, infants with GI signs and radiographic findings of pneumatosis intestinalis might have other benign disorders, eg, neonatal pneumatosis coli,5 "benign necrotizing enterocolitis,"6 or presumed milk allergy.7,8 There is also an interobserver variability in interpretation of abdominal radiographs of infants with suspected NEC, predominantly overreading of films.9 Therefore, not all infants with intestinal dilation and/or pneumatosis intestinalis may be managed with prolonged parenteral nutrition while enteral feedings are withheld, as well as with a prolonged course of antibiotic therapy. It is important to have other objective tests to enable us to support the diagnosis and to manage these infants accordingly.
C-reactive protein (CRP) is an acute-phase reactant. CRP value increases in the serum in the presence of inflammation caused by infection or tissue injury.10 Elevated CRP has been reported in conditions such as septicemia, meningitis, meconium aspiration syndrome, and immunization and after surgical procedures.10–12 An elevated CRP level may not be evident until 12 to 24 hours after appearance of clinical signs because of its variable lag period.11,12 Also, CRP has a short half-life; with appropriate treatment, CRP level will rapidly decline.10 So as not to miss an elevated CRP value, frequent measurements are obtained during the early course of a disease.12 CRP abnormalities in NEC have been reported previously; however, these reports have involved relatively small cohorts of infants.12–14
We hypothesized that serial CRP measurement if used as an adjunct to abdominal radiographs would improve the identification of infants with NEC. Furthermore, we determined whether normal CRP values in the presence of GI manifestations would indicate no NEC. Therefore, clinical treatment of these infants could be based jointly on serial CRP measurements and abdominal radiographs but on neither factor alone. In this study, we assessed the relationship between serial CRP values and abdominal radiographs during the first 48 hours of abdominal signs in infants who are suspected of having NEC. Also, CRP responses were correlated with clinical course and treatment of infants with NEC.
METHODS
This study was reviewed and approved by The University of Tennessee Health Science Center Institutional Review Board and was conducted at the Regional Medical Center, Rout Center for Women and Newborns. Informed consent was waived because CRP determinations were being done routinely in our NICU. Moreover, interpretation of laboratory data and treatment of each infant were at the discretion of the attending physician and not dictated by this study. This observational study monitored infants who were evaluated by their attending physician for possible NEC because of their systemic and GI manifestations. GI manifestations included 2 or more of the following: increased gastric residuals or aspirates before feedings (>20% of feeding volume), bilious or bloody gastric aspirate or emesis, abdominal distention, abdominal tenderness, absent or decreased bowel sounds, abdominal wall erythema, and gross bloody stool. These early GI signs are commonly observed in infants with NEC.15
All patients had abdominal radiographs and CRP measurements. CRP level 1.0 mg/dL was considered normal.12 Other supporting laboratory determinations included blood culture and total white blood cell (WBC) count and differential. Normal WBC indices included total WBC count between 5000 and 25000/mm3, immature to total neutrophil ratio <0.17 or total band count <1500/mm3, based on the criteria by Manroe et al.16 Cerebrospinal fluid and urine cultures were obtained in selected cases. At completion of the initial work-up, antibiotic therapy (ampicillin or vancomycin and gentamicin) was initiated. Continuation or any change in antibiotic therapy was at the discretion of the attending physician, taking into consideration the infant’s clinical course, laboratory findings, and blood culture results. Infants received supportive care as appropriate: gastric decompression, volume expansion with fluid or blood products, inotropic or pressor support, and/or mechanical ventilation.
When abdominal radiographs were interpreted as being abnormal, radiographs were repeated every 6 hours for the first 48 hours and less frequently thereafter at the discretion of the attending physician. During the early phase of NEC, frequent radiographic examinations have been recommended.4,17 The abdominal radiographs were interpreted by the pediatric radiologist on-call. Both clinical manifestations and radiographic findings were used to establish the stage of NEC according to the modified Bell’s staging criteria.4 In modified Bell’s staging criteria,4 the radiographic findings included mild intestinal dilation and mild ileus that were associated with "NEC suspect," pneumatosis intestinalis with significant intestinal dilation and ileus with or without portal vein gas or ascites that were associated with NEC stage II, and additional findings of ascites and bowel perforation with advanced NEC stage III.
CRP was determined at the time of work-up and then repeated 12 and 24 hours later. When any CRP value was abnormal, determination was repeated once daily until a normal value was obtained. Serum concentrations of CRP were measured by rate immunonephelometry using automated instrumentation (Automated Immunochemistry System, Beckman Instrument, Inc, Fullerton, CA). The CRP assays were performed after a routine laboratory quality-control procedure for intra-assay variability was run.
Because the initial clinical manifestations of NEC are often nonspecific, all infants were reevaluated at 48 hours after disease onset to determine whether the infant’s clinical course was consistent with NEC, and treatment was tailored accordingly. By 48 hours, an infant would have shown either improvement or clinical deterioration. By such an approach, a more definitive diagnosis of NEC and its stage could be made. Diagnosis of benign pneumatosis intestinalis was made when an infant met the following criteria: (1) clinical signs and abdominal radiographic changes were resolved within 48 hours; (2) persistently normal WBC indices; (3) negative blood culture, or, when positive, it was positive for 1 of the commensals; (4) resumption of feeding within 48 hours without difficulty; and (5) discontinuation of antibiotics within 48 hours.
Statistical Analysis
The t test was used to compare means (continuous variables), whereas Fisher’s exact test or 2 test was used to determine statistical significance for categorical data from the 2 x 2 tables. Cross-tabulations were generated for NEC and CRP results for patients with abnormal GI manifestations and then for the subgroup of patients with positive abdominal radiographic findings, consistent with NEC stages II and III. False-positive predictive value and false-negative predictive value were generated from the 2 x 2 tables and determined on the basis of the infants’ final clinical outcome assessment at 48 hours after the onset of the GI manifestations. Fisher’s exact test was used to test the significance that CRP predicts NEC among all patients and a subgroup of patients. The 95% confidence interval of prediction accuracy using abdominal radiographs alone was estimated using 1000 bootstraps.18 Misclassification rates as to final clinical outcome assessment were also compared using an exact test.19 Final diagnosis or outcome was defined as definitive NEC (stages II and III) versus no definite NEC or ileus and benign pneumatosis ("NEC stage I").
RESULTS
During an 8-month study period in 1998, a total of 241 of 1126 infants who were admitted to the NICU required a work-up to rule out NEC. The most common GI manifestation was abdominal distention (84%). The presence of gross blood in the stool was observed in 55%; increased gastric residuals when attempting gavage feedings occurred in 44%. All infants who were evaluated for abdominal manifestations had abdominal radiographs and serial CRP determination per NICU protocol as described in Methods. Fifty percent of the infants were male, and 85% were black. Gestational age ranged from 25 to 34 weeks, and birth weight ranged from 609 to 1560 g. Postnatal age at the time of onset of abdominal signs ranged from 1 to 112 days; 30% of infants were <2 weeks of age.
Of these 241 infants, 175 (73%) had abdominal distention but with unremarkable abdominal radiographic findings and no progression on subsequent radiographs 12 hours later. Ninety-five of 175 infants had abnormal CRP values (Fig 1). Of these 95 infants, 60 had septicemia, 2 had meningitis, 1 had urinary tract infection, 10 had pneumonia, and 6 had meconium aspiration syndrome. The remaining 16 of 95 infants were presumed to have infection, because they were born with associated maternal complication suggestive of infection: fever, chorioamnionitis, urinary tract infection, or premature prolonged rupture of amniotic membranes. Although cultures were negative, these 16 infants were treated with antibiotics for 7 to 14 days per their attending physician. Of 175 infants, 80 had negative blood cultures and were assessed as having had nonspecific feeding intolerance; antibiotic therapy was aborted in these infants at 24 to 48 hours. These 80 infants also had persistently normal CRP.
The initial abdominal radiographs in the remaining 66 (27%) of 241 infants were interpreted as abnormal. These findings included intestinal dilation (n = 7), pneumatosis intestinalis (n = 56), and intestinal perforation and/or ascites (n = 3). Postnatal age of these 66 infants ranged from 8 to 34 days. At 48 hours after initial work-up, infants were reevaluated and final diagnosis was made (Fig 1). In 6 of 7 infants with intestinal dilation, radiographic findings resolved by 48 hours; all 6 infants had persistently normal CRP and WBC indices. Five of these 6 infants had blood cultures that were positive for commensals (Staphylococcus epidermidis in 4 infants and Streptococcus viridans in 1 infant) and were considered contaminated; antibiotics were administered for <48 hours (Table 1). Feedings were withheld in these infants for 48 hours and were tolerated when resumed. The remaining 1 infant with intestinal dilation had progression of radiographic findings to persistent pneumatosis intestinalis (stage II NEC).
Of 56 infants with initial pneumatosis intestinalis, findings resolved by 48 hours in 5 infants. These infants had persistently normal CRP values, and their WBC indices were recurrently normal. No additional abdominal complication was observed during their hospitalization, regardless of the short-term antibiotic therapy and early resumption of feeding. Forty-eight of 56 infants had persistent pneumatosis intestinalis and at 48 hours had definitive diagnosis of stage II NEC. Of these 48 infants, 29 (60%) had positive blood cultures; the most common organism was Staphylococcus epidermidis (n = 9) followed by Escherichia coli (n = 7; Table 1). Twenty-eight (58%) of 48 infants with stage II NEC had abnormal CRP at first determination. On subsequent determination at 12 hours, 39 (81%) of 48 had abnormal CRP and 100% by 24 hours (Fig 2). The sensitivity of CRP in stage II NEC increased with serial determination, regardless of whether the blood culture results were positive or negative. Peak CRP level of 9.1 ± 5.1 mg/dL was reached at 1.7 ± 1.5 days from onset of the disease, irrespective of blood culture results. Abnormal CRP values in infants with stage II NEC returned to normal at a mean of 9 days from the disease onset, except in 7 infants, 4 of whom required exploratory surgery for stricture formation and 3 for intra-abdominal abscesses and peritonitis. The remaining 4 of 56 infants with pneumatosis intestinalis developed intestinal perforation by 48 hours of disease onset (stage III NEC). All infants with stage III NEC (n = 7) had abnormal initial CRP, except for 1 infant in whom CRP became abnormal on subsequent determination. Peak CRP value was 7.3 ± 6.4 mg/dL, demonstrated at 0.9 ± 0.4 days of onset of GI manifestations. No follow-up CRP was done in 7 infants who were transferred to the Children’s Hospital for surgery; 4 of 7 survived, and 3 died. Five (71%) of the 7 infants with stage III NEC had positive blood cultures, the most common organism being Enterobacter aerogenes (Table 1). The incidence of positive blood cultures did not differ between those with NEC stage II and NEC stage III (2 = 1.05, P = .31). Mean peak CRP values did not differ between those with NEC stage II and stage III. The time to reach the peak CRP value, however, was shorter with NEC stage III compared with stage II (0.9 ± 0.4 days vs 1.7 ± 1.5 days; t = 3.0, P < .005).
We also compared the abdominal radiographic findings along with CRP in the diagnosis of NEC stages II and III. The 95 infants with abnormal CRP and established diagnoses (septicemia, meningitis, urinary tract infection, pneumonia, meconium aspiration syndrome, or presumptive infection) were not included in the analysis because these conditions are known to be associated with increased CRP.12,20 Similarly, 80 infants who had a diagnosis of nonspecific feeding intolerance and normal abdominal radiographs and normal CRP values were excluded. When abdominal radiographic findings in the first 48 hours were used alone, we had a true-positive rate of 92% (55 of 60) for diagnosis of NEC stages II and III and a false-positive rate of 8% (5 of 60), with 95% confidence interval of 1.5% to 15.2%. However, when both serial CRP values and abdominal radiographs were considered together, specificity increased from 55% (6 of 11) to 100%, and no patients were misclassified, which is significantly better than the 8% misclassification rate when CRP was not used (P = .004). In the subgroup of 60 infants who had radiographic findings consistent with NEC stages II or III, 5 were assessed to have no NEC stage II (false positive) but were correctly classified as not having NEC with addition of CRP (P < .001 by Fisher’s exact test).
DISCUSSION
Our present experience with a large number of infants demonstrates the usefulness of CRP in discriminating NEC stage II from the benign form of pneumatosis intestinalis, NEC suspect, or spuriously suggestive GI conditions. Few investigators have evaluated the use of CRP in infants with NEC.12–14,21 Philip et al13 reported increased CRP in 12 (80%) of 15 infants with classical NEC at initial evaluation, but the exact time interval between CRP determination and onset of clinical signs was not reported. Isaacs et al14 reported that 10 (83%) of 12 infants with definite diagnosis of NEC had an abnormal CRP at the time of onset; by 48 hours, 11 of 12 infants had abnormal values. We also reported previously on abnormal CRP values within 24 hours of onset of GI signs in infants with NEC stages II and III.12 There are, however, infants who present with GI signs and abnormal radiographs suggestive of NEC stage II but CRP values remain persistently normal. In these instances, nonspecific feeding intolerance, ileus, and benign pneumatosis intestinalis are considered, and the normal CRP values are consistent with a mild course or with a mistaken diagnosis (overreading film).
NEC suspect, or NEC stage I, is virtually a clinical nonentity most often caused by feeding intolerance, ileus, or other nonspecific GI disease.5–8 The physical findings are usually transient; feedings can often be restarted with relative safety. In the present study, infants with NEC suspect ("NEC stage I") followed a benign, uncomplicated course.
Five of 6 infants who had radiographic findings of intestinal dilation/ileus and normal CRP values also had positive blood culture. On review of their records, the blood cultures seem to have been contaminated: (1) the organisms were contaminating commensals, (2) they had normal WBC indices, and (3) they improved clinically despite early discontinuation of antibiotic therapy, ie, less than a 2-day course of treatment. We do not suggest that all blood cultures that grow Streptococcus viridans or Staphylococcus epidermidis should be considered as contaminated. Eleven infants with NEC stages II and III also had positive blood cultures with these contaminating commensals. As was suggested earlier, to discern contamination from pathogen, serial CRP determination should be a helpful tool.12,22,23
Infants who receive a diagnosis of having NEC with or without concomitant positive blood culture commonly receive antibiotic therapy for 10 to 14 days and have no enteral feedings for at least the same period.4,14 Radiographic findings of pneumatosis intestinalis is often erroneously considered by some clinicians as a confirmatory sign of stage II NEC. However, pneumatosis intestinalis may be confused with intestinal gas mixed with stool or blood.4 It can be indistinguishable from "neonatal pneumatosis coli," a benign GI disorder.5 Also, interobserver variability exists in interpreting the abdominal radiographs with more tendencies for overreading films.5,24 A small number of our patients who initially received a diagnosis of stage II NEC because of pneumatosis intestinalis did not demonstrate a rise in their CRP values. The clinical and radiographic findings resolved within 48 hours, and their reassessment on the basis of the clinical course was consistent with benign pneumatosis. Also, the absence of CRP response in these patients was compatible with their benign clinical course.
All infants with confirmed NEC and abnormal CRP values also had abnormal clinical and radiographic finding for a protracted period. Abnormal CRP was first observed at initiation of work-up in stage III NEC and also in infants who eventually required surgery for intestinal perforation. In stage II NEC, CRP increased within the first 24 hours of GI manifestations. This rise in CRP is presumably a response to tissue inflammation and/or necrosis.10,12,20,25 Mean CRP values were similar in stages II and III NEC, regardless of whether an associated positive blood culture was present. In these infants, antibiotic therapy was continued for 10 to 14 days regardless of blood culture results.
Our results indicate the importance of serial CRP determination. Had we obtained only a single early CRP, we would have missed the abnormal elevations in 42% of infants with stage II NEC. Also, a 12-hour interval between CRP determinations in the first 24 hours of work-up will allow for closer monitoring of patients. Our experience also indicates that once an abnormal CRP value is demonstrated, daily determinations of CRP are helpful in assessing the clinical course. Persistence of high levels of CRP suggests ongoing disease and/or complications that may require surgical intervention. Our experience is consistent with the reports of other investigators.13,14,26 Normalization of CRP values is associated with subsequent improvement in clinical status and a potential to resume oral feedings.
Our data, however, should be interpreted with caution because CRP can be elevated in non-NEC disorders such as septicemia or meningitis, conditions that may present with GI manifestation. On the basis of our experience, we recommend that serial CRP determinations be included with abdominal radiographs for diagnostic evaluation of NEC. Once the diagnosis of NEC is established, daily determination of CRP is valuable in the recognition of a subsiding disease or a worsening process that may require surgery. However, the lack of abnormal CRP response would favor aborted antibiotic therapy and early resumption of oral feedings.
ACKNOWLEDGMENTS
We thank Marion Haynes for editorial assistance.
FOOTNOTES
Accepted Feb 2, 2005.
No conflict of interest declared.
Dr Bada’s current address is University of Kentucky, Division of Neonatology, 800 Rose St, MS-473, Lexington, KY 40536-0298.
Dr Yang’s current address is St Jude Children’s Research Hospital, 332 North Lauderdale, Memphis, TN 38105.
REFERENCES
Stoll B. Epidemiology of necrotizing enterocolitis. Clin Perinatol. 1994;21 :205 –218
Kliegman RM, Fanaroff AA. Necrotizing enterocolitis. N Engl J Med. 1984;310 :1093 –1103
Bell MJ, Ternberg JL, Feigin RD, et al. Neonatal necrotizing enterocolitis. Therapeutic decisions based upon clinical staging. Ann Surg. 1978;187 :1 –7
Walsh MC, Kliegman RM. Necrotizing enterocolitis: treatment based on staging criteria. Pediatr Clin North Am. 1986;33 :179 –201
Leonidas JC, Hall RT. Neonatal pneumatosis coli: a mild form of neonatal necrotizing enterocolitis. J Pediatr. 1976;89 :456 –459
Richmond JA, Mikity V. Benign form of necrotizing enterocolitis. AJR Radium Ther Nucl Med. 1975;123 :301 –306
Aziz EM. Neonatal pneumatosis intestinalis associated with milk intolerance. Am J Dis Child. 1983;125 :560 –563
Powell GK. Enterocolitis in low birth weight infants associated with milk and soy protein intolerance. J Pediatr. 1976;88 :840 –844
Mata AG, Rosengart M. Interobserver variability in the radiographic diagnosis of necrotizing enterocolitis. Pediatrics. 1980;66 :68 –71
Pepys MB. C-reactive protein fifty years on. Lancet. 1981;1 :653 –656
Peltola H, Jaakkola M. C-reactive protein in early detection of bacteremic versus viral infections in immunocompetent compromised children. J Pediatr. 1988;113 :641 –646
Pourcyrous M, Bada HS, Korones SB, Baselski V, Wong SP. Significance of serial C-reactive protein responses in neonatal infection and other disorders. Pediatrics. 1993;92 :431 –435
Philip AGS, Sann L, Bienvenu F. Acute phase proteins in necrotizing enterocolitis. Acta Paediatr Scand. 1986;75 :1032 –1033
Isaacs D, North J, Lindsell D, Wilkinson AR. Serum acute phase reactants in necrotizing enterocolitis. Acta Paediatr Scand. 1987;76 :923 –927
Grosfeld JL, Cheu H, Schlatter M, West KW, Rescorla FJ. Changing trends in necrotizing enterocolitis. Ann Surg. 1991;214 :300 –307
Manroe BL, Weinberg AG, Rosenfeld CR, Browne R. The neonatal blood count in health and disease, I: reference values for neutrophilic cells. J Pediatr. 1979:95; 89 –98
Morrison SC, Jacobson JM. The radiology of necrotizing enterocolitis. Clin Perinatol. 1994;21 :347 –363
Efron B, Tibshirani R. An Introduction to the Bootstrap. New York, NY: Chapman and Hall/CRC; 1998
Hollander M, Wolfe DA. Nonparametric Statistical Influence. NewYork, NY: John Wiley and Son; 1973
Pourcyrous M, Bada HS, Korones SB, et al. Acute phase reactants in neonatal bacterial infection. J Perinatol. 1991;11 :319 –325
Schober PH, Nassiri J. Risk factors and severity indices in necrotizing enterocolitis. Acta Paediatr Suppl. 1994;396 :49 –52
Schmidt BK, Kirpalani HM, Corey M. Coagulase-negative staphylococci as true pathogens in newborn infants: a cohort study. Pediatr Infect Dis J. 1987;6 :1026 –1031
McCartney AC, McGovern T, Cobb S, Gemmell CG. The measurement of C-reactive protein and immune complexes in endocarditis caused by coagulase-negative staphylococci. J Infect. 1987;15 :213 –219
Rehan VK, Seshia MM, Johnston B, Reed M, Wilmot D, Cook V. Observer variability in interpretation of abdominal radiographs of infants with suspected necrotizing enterocolitis. Clin Pediatr. 1999;38 :637 –643
Harris MC, Costarino AT, Sullivan JS, et al. Cytokine elevations in critically ill infants with sepsis and necrotizing enterocolitis. J Pediatr. 1994;124 :105 –111
Costin BS, Singleton EB. Bowel stenosis as a late complication of acute necrotizing enterocolitis. Radiology. 1978;128 :435 –43(Massroor Pourcyrous, MD, )
Department of Mathematical Sciences and Statistics, University of Memphis, Memphis, Tennessee
ABSTRACT
Objective. In this prospective, observational study, we determined whether serum C-reactive protein (CRP) correlated with necrotizing enterocolitis (NEC) stages II and III. We hypothesized that serial CRP measurement if used as an adjunct to abdominal radiographs would improve the identification of infants with NEC.
Methods. Serum CRP level was measured every 12 hours for 3 measurements and, when abnormal, once daily. When clinical signs persisted and the initial abdominal radiographs were abnormal, follow-up radiographs were obtained.
Results. Of 241 infants who were evaluated for gastrointestinal signs, 11 had ileus or benign pneumatosis intestinalis with persistently normal CRP; gastrointestinal manifestations resolved within 48 hours, antibiotics were discontinued in <48 hours, and feedings were restarted early without complications. Fifty-five infants had NEC stages II and III; all had abnormal CRP regardless of their blood culture results. In infants with stage II NEC, CRP returned to normal at a mean of 9 days except in those who developed complications such as stricture or abscess formation.
Conclusions. In infants with suspected NEC, normal serial CRP values would favor aborted antibiotic therapy and early resumption of feedings. CRP becomes abnormal in both stage II and stage III NEC. In infants with NEC, persistently elevated CRP after initiation of appropriate medical management suggests associated complications, which may require surgical intervention.
Key Words: necrotizing enterocolitis C-reactive protein newborn premature
Abbreviations: NEC, necrotizing enterocolitis GI, gastrointestinal CRP, C-reactive protein WBC, white blood cell
Necrotizing enterocolitis (NEC) is a common gastrointestinal (GI) disorder in premature infants and is associated with high morbidity and mortality.1 The initial clinical manifestations of NEC are often nonspecific and indistinguishable from neonatal septicemia and other forms of GI disorders.2 Usually, with progression, diagnosis may be established by abdominal radiographic findings, such as intestinal dilation, pneumatosis intestinalis, portal or hepatic vein gas, pneumoperitoneum, or ascites.3,4 However, some of these radiographic findings are not specific for NEC and may occur in other benign entities. For instance, many premature infants with intolerance to feeding show intestinal dilation and ileus.2 Also, infants with GI signs and radiographic findings of pneumatosis intestinalis might have other benign disorders, eg, neonatal pneumatosis coli,5 "benign necrotizing enterocolitis,"6 or presumed milk allergy.7,8 There is also an interobserver variability in interpretation of abdominal radiographs of infants with suspected NEC, predominantly overreading of films.9 Therefore, not all infants with intestinal dilation and/or pneumatosis intestinalis may be managed with prolonged parenteral nutrition while enteral feedings are withheld, as well as with a prolonged course of antibiotic therapy. It is important to have other objective tests to enable us to support the diagnosis and to manage these infants accordingly.
C-reactive protein (CRP) is an acute-phase reactant. CRP value increases in the serum in the presence of inflammation caused by infection or tissue injury.10 Elevated CRP has been reported in conditions such as septicemia, meningitis, meconium aspiration syndrome, and immunization and after surgical procedures.10–12 An elevated CRP level may not be evident until 12 to 24 hours after appearance of clinical signs because of its variable lag period.11,12 Also, CRP has a short half-life; with appropriate treatment, CRP level will rapidly decline.10 So as not to miss an elevated CRP value, frequent measurements are obtained during the early course of a disease.12 CRP abnormalities in NEC have been reported previously; however, these reports have involved relatively small cohorts of infants.12–14
We hypothesized that serial CRP measurement if used as an adjunct to abdominal radiographs would improve the identification of infants with NEC. Furthermore, we determined whether normal CRP values in the presence of GI manifestations would indicate no NEC. Therefore, clinical treatment of these infants could be based jointly on serial CRP measurements and abdominal radiographs but on neither factor alone. In this study, we assessed the relationship between serial CRP values and abdominal radiographs during the first 48 hours of abdominal signs in infants who are suspected of having NEC. Also, CRP responses were correlated with clinical course and treatment of infants with NEC.
METHODS
This study was reviewed and approved by The University of Tennessee Health Science Center Institutional Review Board and was conducted at the Regional Medical Center, Rout Center for Women and Newborns. Informed consent was waived because CRP determinations were being done routinely in our NICU. Moreover, interpretation of laboratory data and treatment of each infant were at the discretion of the attending physician and not dictated by this study. This observational study monitored infants who were evaluated by their attending physician for possible NEC because of their systemic and GI manifestations. GI manifestations included 2 or more of the following: increased gastric residuals or aspirates before feedings (>20% of feeding volume), bilious or bloody gastric aspirate or emesis, abdominal distention, abdominal tenderness, absent or decreased bowel sounds, abdominal wall erythema, and gross bloody stool. These early GI signs are commonly observed in infants with NEC.15
All patients had abdominal radiographs and CRP measurements. CRP level 1.0 mg/dL was considered normal.12 Other supporting laboratory determinations included blood culture and total white blood cell (WBC) count and differential. Normal WBC indices included total WBC count between 5000 and 25000/mm3, immature to total neutrophil ratio <0.17 or total band count <1500/mm3, based on the criteria by Manroe et al.16 Cerebrospinal fluid and urine cultures were obtained in selected cases. At completion of the initial work-up, antibiotic therapy (ampicillin or vancomycin and gentamicin) was initiated. Continuation or any change in antibiotic therapy was at the discretion of the attending physician, taking into consideration the infant’s clinical course, laboratory findings, and blood culture results. Infants received supportive care as appropriate: gastric decompression, volume expansion with fluid or blood products, inotropic or pressor support, and/or mechanical ventilation.
When abdominal radiographs were interpreted as being abnormal, radiographs were repeated every 6 hours for the first 48 hours and less frequently thereafter at the discretion of the attending physician. During the early phase of NEC, frequent radiographic examinations have been recommended.4,17 The abdominal radiographs were interpreted by the pediatric radiologist on-call. Both clinical manifestations and radiographic findings were used to establish the stage of NEC according to the modified Bell’s staging criteria.4 In modified Bell’s staging criteria,4 the radiographic findings included mild intestinal dilation and mild ileus that were associated with "NEC suspect," pneumatosis intestinalis with significant intestinal dilation and ileus with or without portal vein gas or ascites that were associated with NEC stage II, and additional findings of ascites and bowel perforation with advanced NEC stage III.
CRP was determined at the time of work-up and then repeated 12 and 24 hours later. When any CRP value was abnormal, determination was repeated once daily until a normal value was obtained. Serum concentrations of CRP were measured by rate immunonephelometry using automated instrumentation (Automated Immunochemistry System, Beckman Instrument, Inc, Fullerton, CA). The CRP assays were performed after a routine laboratory quality-control procedure for intra-assay variability was run.
Because the initial clinical manifestations of NEC are often nonspecific, all infants were reevaluated at 48 hours after disease onset to determine whether the infant’s clinical course was consistent with NEC, and treatment was tailored accordingly. By 48 hours, an infant would have shown either improvement or clinical deterioration. By such an approach, a more definitive diagnosis of NEC and its stage could be made. Diagnosis of benign pneumatosis intestinalis was made when an infant met the following criteria: (1) clinical signs and abdominal radiographic changes were resolved within 48 hours; (2) persistently normal WBC indices; (3) negative blood culture, or, when positive, it was positive for 1 of the commensals; (4) resumption of feeding within 48 hours without difficulty; and (5) discontinuation of antibiotics within 48 hours.
Statistical Analysis
The t test was used to compare means (continuous variables), whereas Fisher’s exact test or 2 test was used to determine statistical significance for categorical data from the 2 x 2 tables. Cross-tabulations were generated for NEC and CRP results for patients with abnormal GI manifestations and then for the subgroup of patients with positive abdominal radiographic findings, consistent with NEC stages II and III. False-positive predictive value and false-negative predictive value were generated from the 2 x 2 tables and determined on the basis of the infants’ final clinical outcome assessment at 48 hours after the onset of the GI manifestations. Fisher’s exact test was used to test the significance that CRP predicts NEC among all patients and a subgroup of patients. The 95% confidence interval of prediction accuracy using abdominal radiographs alone was estimated using 1000 bootstraps.18 Misclassification rates as to final clinical outcome assessment were also compared using an exact test.19 Final diagnosis or outcome was defined as definitive NEC (stages II and III) versus no definite NEC or ileus and benign pneumatosis ("NEC stage I").
RESULTS
During an 8-month study period in 1998, a total of 241 of 1126 infants who were admitted to the NICU required a work-up to rule out NEC. The most common GI manifestation was abdominal distention (84%). The presence of gross blood in the stool was observed in 55%; increased gastric residuals when attempting gavage feedings occurred in 44%. All infants who were evaluated for abdominal manifestations had abdominal radiographs and serial CRP determination per NICU protocol as described in Methods. Fifty percent of the infants were male, and 85% were black. Gestational age ranged from 25 to 34 weeks, and birth weight ranged from 609 to 1560 g. Postnatal age at the time of onset of abdominal signs ranged from 1 to 112 days; 30% of infants were <2 weeks of age.
Of these 241 infants, 175 (73%) had abdominal distention but with unremarkable abdominal radiographic findings and no progression on subsequent radiographs 12 hours later. Ninety-five of 175 infants had abnormal CRP values (Fig 1). Of these 95 infants, 60 had septicemia, 2 had meningitis, 1 had urinary tract infection, 10 had pneumonia, and 6 had meconium aspiration syndrome. The remaining 16 of 95 infants were presumed to have infection, because they were born with associated maternal complication suggestive of infection: fever, chorioamnionitis, urinary tract infection, or premature prolonged rupture of amniotic membranes. Although cultures were negative, these 16 infants were treated with antibiotics for 7 to 14 days per their attending physician. Of 175 infants, 80 had negative blood cultures and were assessed as having had nonspecific feeding intolerance; antibiotic therapy was aborted in these infants at 24 to 48 hours. These 80 infants also had persistently normal CRP.
The initial abdominal radiographs in the remaining 66 (27%) of 241 infants were interpreted as abnormal. These findings included intestinal dilation (n = 7), pneumatosis intestinalis (n = 56), and intestinal perforation and/or ascites (n = 3). Postnatal age of these 66 infants ranged from 8 to 34 days. At 48 hours after initial work-up, infants were reevaluated and final diagnosis was made (Fig 1). In 6 of 7 infants with intestinal dilation, radiographic findings resolved by 48 hours; all 6 infants had persistently normal CRP and WBC indices. Five of these 6 infants had blood cultures that were positive for commensals (Staphylococcus epidermidis in 4 infants and Streptococcus viridans in 1 infant) and were considered contaminated; antibiotics were administered for <48 hours (Table 1). Feedings were withheld in these infants for 48 hours and were tolerated when resumed. The remaining 1 infant with intestinal dilation had progression of radiographic findings to persistent pneumatosis intestinalis (stage II NEC).
Of 56 infants with initial pneumatosis intestinalis, findings resolved by 48 hours in 5 infants. These infants had persistently normal CRP values, and their WBC indices were recurrently normal. No additional abdominal complication was observed during their hospitalization, regardless of the short-term antibiotic therapy and early resumption of feeding. Forty-eight of 56 infants had persistent pneumatosis intestinalis and at 48 hours had definitive diagnosis of stage II NEC. Of these 48 infants, 29 (60%) had positive blood cultures; the most common organism was Staphylococcus epidermidis (n = 9) followed by Escherichia coli (n = 7; Table 1). Twenty-eight (58%) of 48 infants with stage II NEC had abnormal CRP at first determination. On subsequent determination at 12 hours, 39 (81%) of 48 had abnormal CRP and 100% by 24 hours (Fig 2). The sensitivity of CRP in stage II NEC increased with serial determination, regardless of whether the blood culture results were positive or negative. Peak CRP level of 9.1 ± 5.1 mg/dL was reached at 1.7 ± 1.5 days from onset of the disease, irrespective of blood culture results. Abnormal CRP values in infants with stage II NEC returned to normal at a mean of 9 days from the disease onset, except in 7 infants, 4 of whom required exploratory surgery for stricture formation and 3 for intra-abdominal abscesses and peritonitis. The remaining 4 of 56 infants with pneumatosis intestinalis developed intestinal perforation by 48 hours of disease onset (stage III NEC). All infants with stage III NEC (n = 7) had abnormal initial CRP, except for 1 infant in whom CRP became abnormal on subsequent determination. Peak CRP value was 7.3 ± 6.4 mg/dL, demonstrated at 0.9 ± 0.4 days of onset of GI manifestations. No follow-up CRP was done in 7 infants who were transferred to the Children’s Hospital for surgery; 4 of 7 survived, and 3 died. Five (71%) of the 7 infants with stage III NEC had positive blood cultures, the most common organism being Enterobacter aerogenes (Table 1). The incidence of positive blood cultures did not differ between those with NEC stage II and NEC stage III (2 = 1.05, P = .31). Mean peak CRP values did not differ between those with NEC stage II and stage III. The time to reach the peak CRP value, however, was shorter with NEC stage III compared with stage II (0.9 ± 0.4 days vs 1.7 ± 1.5 days; t = 3.0, P < .005).
We also compared the abdominal radiographic findings along with CRP in the diagnosis of NEC stages II and III. The 95 infants with abnormal CRP and established diagnoses (septicemia, meningitis, urinary tract infection, pneumonia, meconium aspiration syndrome, or presumptive infection) were not included in the analysis because these conditions are known to be associated with increased CRP.12,20 Similarly, 80 infants who had a diagnosis of nonspecific feeding intolerance and normal abdominal radiographs and normal CRP values were excluded. When abdominal radiographic findings in the first 48 hours were used alone, we had a true-positive rate of 92% (55 of 60) for diagnosis of NEC stages II and III and a false-positive rate of 8% (5 of 60), with 95% confidence interval of 1.5% to 15.2%. However, when both serial CRP values and abdominal radiographs were considered together, specificity increased from 55% (6 of 11) to 100%, and no patients were misclassified, which is significantly better than the 8% misclassification rate when CRP was not used (P = .004). In the subgroup of 60 infants who had radiographic findings consistent with NEC stages II or III, 5 were assessed to have no NEC stage II (false positive) but were correctly classified as not having NEC with addition of CRP (P < .001 by Fisher’s exact test).
DISCUSSION
Our present experience with a large number of infants demonstrates the usefulness of CRP in discriminating NEC stage II from the benign form of pneumatosis intestinalis, NEC suspect, or spuriously suggestive GI conditions. Few investigators have evaluated the use of CRP in infants with NEC.12–14,21 Philip et al13 reported increased CRP in 12 (80%) of 15 infants with classical NEC at initial evaluation, but the exact time interval between CRP determination and onset of clinical signs was not reported. Isaacs et al14 reported that 10 (83%) of 12 infants with definite diagnosis of NEC had an abnormal CRP at the time of onset; by 48 hours, 11 of 12 infants had abnormal values. We also reported previously on abnormal CRP values within 24 hours of onset of GI signs in infants with NEC stages II and III.12 There are, however, infants who present with GI signs and abnormal radiographs suggestive of NEC stage II but CRP values remain persistently normal. In these instances, nonspecific feeding intolerance, ileus, and benign pneumatosis intestinalis are considered, and the normal CRP values are consistent with a mild course or with a mistaken diagnosis (overreading film).
NEC suspect, or NEC stage I, is virtually a clinical nonentity most often caused by feeding intolerance, ileus, or other nonspecific GI disease.5–8 The physical findings are usually transient; feedings can often be restarted with relative safety. In the present study, infants with NEC suspect ("NEC stage I") followed a benign, uncomplicated course.
Five of 6 infants who had radiographic findings of intestinal dilation/ileus and normal CRP values also had positive blood culture. On review of their records, the blood cultures seem to have been contaminated: (1) the organisms were contaminating commensals, (2) they had normal WBC indices, and (3) they improved clinically despite early discontinuation of antibiotic therapy, ie, less than a 2-day course of treatment. We do not suggest that all blood cultures that grow Streptococcus viridans or Staphylococcus epidermidis should be considered as contaminated. Eleven infants with NEC stages II and III also had positive blood cultures with these contaminating commensals. As was suggested earlier, to discern contamination from pathogen, serial CRP determination should be a helpful tool.12,22,23
Infants who receive a diagnosis of having NEC with or without concomitant positive blood culture commonly receive antibiotic therapy for 10 to 14 days and have no enteral feedings for at least the same period.4,14 Radiographic findings of pneumatosis intestinalis is often erroneously considered by some clinicians as a confirmatory sign of stage II NEC. However, pneumatosis intestinalis may be confused with intestinal gas mixed with stool or blood.4 It can be indistinguishable from "neonatal pneumatosis coli," a benign GI disorder.5 Also, interobserver variability exists in interpreting the abdominal radiographs with more tendencies for overreading films.5,24 A small number of our patients who initially received a diagnosis of stage II NEC because of pneumatosis intestinalis did not demonstrate a rise in their CRP values. The clinical and radiographic findings resolved within 48 hours, and their reassessment on the basis of the clinical course was consistent with benign pneumatosis. Also, the absence of CRP response in these patients was compatible with their benign clinical course.
All infants with confirmed NEC and abnormal CRP values also had abnormal clinical and radiographic finding for a protracted period. Abnormal CRP was first observed at initiation of work-up in stage III NEC and also in infants who eventually required surgery for intestinal perforation. In stage II NEC, CRP increased within the first 24 hours of GI manifestations. This rise in CRP is presumably a response to tissue inflammation and/or necrosis.10,12,20,25 Mean CRP values were similar in stages II and III NEC, regardless of whether an associated positive blood culture was present. In these infants, antibiotic therapy was continued for 10 to 14 days regardless of blood culture results.
Our results indicate the importance of serial CRP determination. Had we obtained only a single early CRP, we would have missed the abnormal elevations in 42% of infants with stage II NEC. Also, a 12-hour interval between CRP determinations in the first 24 hours of work-up will allow for closer monitoring of patients. Our experience also indicates that once an abnormal CRP value is demonstrated, daily determinations of CRP are helpful in assessing the clinical course. Persistence of high levels of CRP suggests ongoing disease and/or complications that may require surgical intervention. Our experience is consistent with the reports of other investigators.13,14,26 Normalization of CRP values is associated with subsequent improvement in clinical status and a potential to resume oral feedings.
Our data, however, should be interpreted with caution because CRP can be elevated in non-NEC disorders such as septicemia or meningitis, conditions that may present with GI manifestation. On the basis of our experience, we recommend that serial CRP determinations be included with abdominal radiographs for diagnostic evaluation of NEC. Once the diagnosis of NEC is established, daily determination of CRP is valuable in the recognition of a subsiding disease or a worsening process that may require surgery. However, the lack of abnormal CRP response would favor aborted antibiotic therapy and early resumption of oral feedings.
ACKNOWLEDGMENTS
We thank Marion Haynes for editorial assistance.
FOOTNOTES
Accepted Feb 2, 2005.
No conflict of interest declared.
Dr Bada’s current address is University of Kentucky, Division of Neonatology, 800 Rose St, MS-473, Lexington, KY 40536-0298.
Dr Yang’s current address is St Jude Children’s Research Hospital, 332 North Lauderdale, Memphis, TN 38105.
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