Human Metapneumovirus and Respiratory Syncytial Virus Disease in ...

BRIEF REPORTS EPIDEMIOLOGIC STUDY OF CONJUNCTIVITISOTITIS SYNDROME Edouard Bingen, PhD,* Robert Cohen, MD,†‡ Nadejda Jourenkova, MD,§ and Pierre Gehanno, MD储 Abstract: We conducted a prospective cross-sectional epidemiologic study of conjunctivitis-otitis syndrome during two 3-week periods of consecutive winters. A representative sample of 184 pediatricians and ear, nose and throat specialists in the Paris area recruited 2901 children from 6 to 36 months of age presenting with acute otitis media, of whom 465 (16%) also had purulent conjunctivitis. In sampled children, the culture of the conjunctival exudate yielded bacterial pathogens in 419 patients: Haemophilus influenzae, 371 (89%); Streptococcus pneumoniae, 72 (17%); and Branhamella catarrhalis, 23 (5%). Pulsed field gel electrophoresis analysis of paired H. influenzae isolates from the conjunctival exudate and middle ear fluid of 21 patients showed identical strains in each patient. Key Words: conjunctivitis-otitis syndrome, Haemophilus influenzae Accepted for publication March 2, 2005. From *Service de Microbiologie, Hoˆpital Robert Debre´, Paris; †Pe´diatrie, Centre Hospitalier Inter Communal, Cre´teil; ‡ACTIV, Saint Maur; §ICARE, Massy; and 㛳Service d’ORL, Hoˆpital Bichat-Claude Bernard, Paris, France Supported by sanofi pasteur. Address for reprints: Dr Edouard Bingen, Service de Microbiologie, Hoˆpital Robert Debre´, 48 Boulevard Se´rurier, 75019 Paris, France. Fax 33 1 40 03 24 50; E-mail [email protected]. Copyright © 2005 by Lippincott Williams & Wilkins DOI: 10.1097/01.inf.0000172939.13159.3b

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he role of Haemophilus influenzae in acute otitis media (AOM) and purulent conjunctivitis was first recognized by Coffey1 nearly 40 years ago. Some years later, Bodor2,3 coined the term “conjunctivitis-otitis syndrome” (COS), and reported that H. influenzae was present in the conjunctival exudate in ⬃80% of cases. Those studies involved few patients, however, and little epidemiologic work has since been done on this syndrome. Increases in antibiotic resistance among H. influenzae and Streptococcus pneumoniae have complicated the treatment of AOM. In a large epidemiologic study, we examined the frequency of COS in French children with AOM and the distribution of H. influenzae, S. pneumoniae and Branhamella catarrhalis. PATIENTS AND METHODS This observational cross-sectional epidemiologic study was based on prospective patient enrollment during 2 consecutive winters (2001–2002 and 2002–2003) in the Paris region (population, ⬃10 million). In each study period, randomly selected pediatricians and ear, nose and throat (ENT) specialists were asked to participate. During a 3-week period, each physician was asked to register all children 6 –36 months of age with signs and symptoms of otitis media (fever, earache and irritability) associated with purulent otorrhea beginning ⬍24 hours previously, or opaque tympanic membrane plus either marked eardrum congestion or marked bulging, or moderate congestion and bulging.4 Among these children, COS was defined by the presence of purulent conjunctivitis. During two 3-week periods, children were enrolled in the study for bacteriologic samples after informing the parents of the nature of the study and obtaining their permission. Conjunctival samples were obtained for bacteriologic analysis, together with standardized his-

tory and physical findings. When otorrhea was present or tympanocentesis was required, samples of middle ear fluid (MEF) were obtained for bacteriologic analysis. The specimens were transported on selective transport medium at room temperature to a central bacteriology laboratory. All specimens (conjunctiva and MEF) were cultured aerobically on blood and chocolate agar in air-10% CO2. A further blood agar plate was incubated anaerobically. Bacteria were identified by standard methods. For the purposes of this study, only H. influenzae, S. pneumoniae, B. catarrhalis and Streptococcus pyogenes were considered potentially pathogenic; all other isolates were considered to be contaminants. H. influenzae and B. catarrhalis isolates were tested for ␤-lactamase production. The minimum inhibitory concentration (MIC) of amoxicillin for S. pneumoniae isolates was determined with the E-test technique, as recommended by the manufacturer, with the following cutpoints: susceptible (MIC ⱕ 0.5 mg/L); diminished susceptibility (MIC 1–2 mg/L); and resistant (MIC ⬎ 2 mg/L). H. influenzae isolates were subjected to capsular serotyping by the slide agglutination method with specific antisera (Phadebact; Boule Diagnostic, Huddinge, Sweden), and S. pneumoniae isolates were serogrouped with latex particles coated with specific antisera from the Copenhagen State Serum Institute. H. influenzae isolates cultured simultaneously from conjunctival exudates and MEF were subjected to pulsed field gel electrophoresis with SmaI restriction enzymes as described elsewhere,5 and the results were analyzed according to the method of Tenover et al.6 S. pneumoniae isolates cultured simultaneously from both sample types were subjected to ribotyping with the HindIII restriction enzyme as described elsewhere.7 Data were managed and analyzed with SAS software version 8 (SAS Institute, Cary, NC). The statistical analysis was descriptive, and the main results were the frequencies of distribution, which are reported with their 95% confidence intervals (95% CI). RESULTS Five hundred sixty randomly selected pediatricians and ENT specialists were invited to participate, and 184 (32.9%) accepted. AOM was diagnosed in 2901 infants, 465 of whom had COS (16%; 95% CI 15–19%). Of these, 57% were boys and 78% were younger than 21 months of age. H. influenzae serotype b immunization had been given to 81% of children and was complete for age in 94% of cases. Vaccination against pneumococci (heptavalent conjugate vaccine) had been given to 8% of children and was complete for age in 65% of these cases. Almost one-half of the children (47%) attended day care centers, and the remainder were kept at home by a parent or childminder. Conjunctival samples were obtained from all patients with COS (n ⫽ 465). A total of 152 MEF samples were obtained (72 tympanocentesis and 80 otorrhea). At least 1 bacterial pathogen was isolated from the conjunctival exudate of 419 patients and from MEF samples of 73 patients (Table 1). H. influenzae was the pathogen most frequently isolated in both types of sample. It accounted for 89% (95% CI 85–92%) of culture-positive conjunctival exudates (371 of 419) and for 66% (95% CI 55–77%) of culture-positive MEF samples (48 of 73). H. influenzae was the only pathogen identified in conjunctival exudates from 79% of patients. The results of bacterial cultures of MEF obtained by tympanocentesis and otorrhea were not substantially different. S. pyogenes was not found in our study. ␤-Lactamase production was observed in 218 (53%) of 429 H. influenzae isolates and in 26 (93%) of 28 B. catarrhalis isolates. Of the 39 S. pneumoniae strains isolated from MEF, 27 (69%) were susceptible to amoxicillin, whereas 11 (28%) had diminished susceptibility and 1 (3%) was resistant. All but 2 of the 429 H. influenzae isolates were nontypable. Serogroups covered by the heptavalent vaccine (4, 6, 9, 14, 18, 19

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TABLE 1. Conjunctival and Middle Ear Isolates Bacteria

Conjunctival Exudate

Middle Ear Fluid

No. of patients sampled No. of patients with pathogen-positive cultures

465 419

152 73

Haemophilus influenzae Alone Polymicrobial

371 (89%, CI 85–92%) 330 (79%) 41 (10%)*

48 (66%, CI 55–77%) 36 (49%) 12 (16%)†

Streptococcus pneumoniae Alone Polymicrobial

72 (17%, CI 14 –21%) 38 (9%) 34 (8%)*

33 (45%, CI 34 –57%) 23 (32%) 10 (14%)†

Branhamella catarrhalis Alone Polymicrobial

23 (5%, CI 3– 8%) 7 (2%) 16 (4%)*

5 (7%, CI 1–13%) 2 (3%) 3 (4%)†

*Twenty-eight patients with H. influenzae ⫹ S. pneumoniae, 10 patients with H. influenzae ⫹ B. catarrhalis, 3 patients with S. pneumoniae ⫹ B. catarrhalis, 3 patients with H. influenzae ⫹ S. pneumoniae ⫹ B. catarrhalis. † Nine patients with H. influenzae ⫹ S. pneumoniae, 2 patients with H. influenzae ⫹ B. catarrhalis, 1 patient with H. influenzae ⫹ S. pneumoniae ⫹ B. catarrhalis.

and 23) accounted for 83% of the 112 S. pneumoniae isolates. Pulsed field gel electrophoresis analysis of paired H. influenzae isolates from conjunctival and MEF samples showed that all 21 patients concerned harbored a single strain, whereas strains varied markedly among different patients. Ribotyping of paired S. pneumoniae isolates showed that they were identical in 8 patients and unrelated in 1 patient.

DISCUSSION Nontypable H. influenzae was the predominant pathogen isolated from both conjunctival and MEF samples of infants with COS. These data are consistent with the results of small studies reported by Bodor2,3 and French teams.8,9 S. pneumoniae was the second most frequent pathogen (respectively, 17 and 44% of isolates), also in keeping with previous work.2,3,8,9 The frequency of ␤-lactamase-producing H. influenzae strains varies from country to country10 but is very high in France. They accounted for ⬎50% of H. influenzae isolates in this study, suggesting that amoxicillin is no longer suitable for COS. French guidelines on AOM take into account both the association of COS with H. influenzae and the high frequency of ␤-lactamase-producing strains, recommending amoxicillin/clavulanate or oral third generation cephalosporins.11 COS was frequent in infants presenting with AOM (16%; 95% CI 15– 19%), confirming the results of one of our previous epidemiologic studies.12 The paired H. influenzae strains isolated from conjunctival and MEF samples of the same patient, and all but 1 of the paired S. pneumoniae isolates, were genetically identical by restriction fragment length polymorphism, and they differed from other strains. This confirms similar findings in 5 patients described by Bodor et al.3 Before the widespread implementation of pneumococcal conjugate vaccination, nontypable H. influenzae was the second bacterial cause of AOM, being responsible for one-third of bacteriologically documented cases. Because H. influenzae is involved in 70 – 89% of cases of COS2,3,8,9 and because conjunctivitis is associated with AOM in 16% of cases, one-third to one-half of children presenting with AOM are likely to be infected by H. influenzae. Palmu et al13 in Finland reported that 54% of children with AOM and H. influenzae-positive MEF culture had ocular symptoms at diagnosis. A relative increase in the frequency of AOM caused by nontypable H. influenzae and consequently a rise in COS are likely

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to occur as pneumococcal conjugate vaccination becomes more widespread.14

ACKNOWLEDGMENTS We thank Dr Ivan Brumpt (sanofi pasteur) for valuable contributions to the performance of the study and the preparation of the manuscript.

REFERENCES 1. Coffey JD. Otitis media in the practice of pediatrics: bacteriological and clinical observations. Pediatrics. 1966;38:25–32. 2. Bodor FF. Conjunctivitis-otitis syndrome. Pediatrics. 1982;69:695– 698. 3. Bodor FF, Marchant CD, Shurin PA, et al. Bacterial etiology of conjunctivitis-otitis media syndrome. Pediatrics. 1985;76:26 –28. 4. Paradise JL. On classifying otitis media as suppurative or nonsuppurative, with a suggested clinical schema. J Pediatr. 1987;111:948 –951. 5. Gazagne L, Delmas C, Bingen E, et al. Molecular epidemiology of ampicillin-resistant non-␤-lactamase-producing Haemophilus influenzae. J Clin Microbiol. 1998;36:3629 –3635. 6. Tenover FC, Arbeit RD, Goering RV, et al. Interpreting chromosomal DNA restriction patterns produced by pulsed-field gel electrophoresis: criteria for bacterial strain typing. J Clin Microbiol. 1995;33:2233–2239. 7. Doit C, Loukil C, Geslin P, et al. Phenotypic and genetic diversity of invasive pneumococcal isolates recovered from French children. J Clin Microbiol. 2002;40:2994 –2998. 8. Cohen R, Danan C, Geslin P. Le syndrome otite-conjonctivite: a` propos de 81 observations. Med Mal Infect 1988;10(bis):553–557. 9. Cohen R, de la Rocque F, Bouhanna A, et al. Etude randomiseé ce´fatrizine versus ce´faclor dans le syndrome otite-conjontivite. Pathol Biol. 1990;38:517–520. 10. Felmingham D, Farrell DJ, Reinert RR, Morrissey I. Antibacterial resistance among children with community-acquired respiratory tract infections (PROTEKT 1999 –2000). J Infect. 2004;48:39 –55. 11. Agence Française de Sećurite´ Sanitaire des Produits de Sante´. Systemic antibiotic treatments in lower and upper respiratory tract infections: official French Guidelines. Clin Microbiol Infections 2003;9:1162–1178. 12. Cohen R, Levy C, Boucherat M, et al. Enqueˆte e´pide´miologique sur l’OMA en pratique pe´diatrique de ville. Me´decine Enfance 1996:27–31. 13. Palmu AA, Herva E, Savolainen H, Karma P, Makela PH, Kilpi TM. Association of clinical signs and symptoms with bacterial findings in acute otitis media. Clin Infect Dis. 2004;38:234 –242. 14. Black S, Hedrick J, Harrison C, et al. Community-wide vaccination with the heptavalent pneumococcal conjugate significantly alters the microbiology of acute otitis media. Pediatr Infect Dis J. 2004;23:829 – 833.

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CLINICAL PRESENTATION AND CHARACTERISTICS OF PHARYNGEAL ADENOVIRUS INFECTIONS Olga Dominguez, MD,* Pablo Rojo, MD,* Susana de las Heras, MD,* Dolores Folgueira, MD,† and Jesu´s Ruiz Contreras, MD* Abstract: We describe the clinical characteristics of 209 children younger than 15 years of age with positive pharyngeal cultures for adenovirus. The mean age of the children was 37 ⫾ 33 months, and the mean peak temperature was 39.2 ⫾ 0.76°C. On physical examination, tonsillitis was found for 88% of children; 52% of them had exudative tonsillitis. Forty-eight percent of the patients who had a white blood cell count performed had ⬎15,000 leukocytes per mm3, and 25% had ⬎20,000 leukocytes per mm3. C-reactive protein concentrations were ⬎7 mg/dL for 22.5% of the patients. Adenovirus pharyngeal infections in young children mimic severe bacterial infections. Key Words: adenovirus, pharyngeal infections, children Accepted for publication January 31, 2005. From the Departments of *Pediatrics and †Microbiology, Hospital Universitario Doce de Octubre, Madrid, Spain Reprints not available. DOI: 10.1097/01.inf.0000172942.96436.2d

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denovirus infections are common in pediatric patients.1 Adenovirus is distributed worldwide and is a well-known cause of respiratory tract infections and gastroenteritis in children.2 Transmission is through respiratory and fecal-oral routes. Cases occur year-round, but higher rates occur in spring and winter. Although most infections are self-limited and can be treated on an ambulatory basis, many children are hospitalized and receive unnecessary antibiotic therapy.3 The aim of this study was to describe the clinical and epidemiologic characteristics of children with pharyngeal adenovirus infections treated at a pediatric emergency department in Spain. METHODS This was a descriptive retrospective study carried out at the pediatric emergency department of Hospital Universitario Doce de Octubre, a tertiary teaching hospital in Madrid. We reviewed the records of children younger than 15 years of age for whom adenovirus was isolated from nasopharyngeal cultures between January 2000 and December 2003. Clinical samples taken at the discretion of the attending physician were inoculated (in a volume of 0.2 mL) onto monolayers of MRC-5 and 4 –549 cells in tubes, incubated at 37°C in stationary phase and scored daily for cytopathic effect for 7 days or until cytopathic effect developed. The following data were collected: age, gender, date of the visit at the emergency department, clinical manifestations, white blood cell (WBC) and differential counts and C-reactive protein (CRP) concentration. Statistical analyses were performed with Epi-Info 6.0; Student’s t test was used to compare continuous variables. RESULTS Epidemiologic and Demographic Data. We found 209 patients (85 girls and 124 boys) with positive adenovirus cultures. For 5 of them (2.3%), ⬎1 virus was isolated (3 respiratory syncytial virus, 1 herpes simplex virus and 1 parainfluenza type 3). The mean age of the children was 37 ⫾ 33 months (median, 28 months). Forty-eight (48.3%) were younger than 2 years of age, 38.7% were 2–5 years of age and 12.9% were older than 5 years of age. The incidence was © 2005 Lippincott Williams & Wilkins

Adenovirus Infections

high in winter and peaked in spring (April) but remained high well into the summer (Fig. 1). Seventeen percent of the patients were hospitalized. Only 4% of the children had first grade relatives with similar symptoms. Clinical Features. Almost all children (96%) presented with fever, with a mean peak temperature of 39.2 ⫾ 0.76°C, and 74% of the children had temperatures of ⬎39°C. Forty-seven children (22%) had upper respiratory tract symptoms such as rhinorrhea, cough or both. Diarrhea occurred for 34 patients (16%) and vomiting for 59 (28%). Tonsillitis was found for 184 children (88%), and 96 of them (52%) had exudative tonsillitis. Anterior cervical lymph node enlargement was present for 66 children (32%), conjunctivitis for 35 patients (17%) and rash for 26 patients (12%). Hepatomegaly was found for 17 children (8%) and splenomegaly for 10 (4%). The most common diagnoses were tonsillitis (48.3%), respiratory tract infection (23.9%) and fever without a focus (12.9%). Laboratory Data. Complete blood counts were performed for 81 children (39%). The mean WBC count was 14,999 ⫾ 6488 leukocytes per mm3 (range, 4210 – 44,400 leukocytes per mm3). Fortyeight percent of the patients who had a WBC count performed had ⬎15,000 leukocytes per mm3, and 25% had ⬎20,000 leukocytes per mm3. For children younger than 2 years of age, the mean WBC count was 18,884 leukocytes per mm3, which is statistically significantly higher (P ⬍ 0.05) than that for children older than 2 years of age. The mean differential count was 57% neutrophils, 29% lymphocytes and 11% monocytes. CRP concentrations were measured for 40 patients (mean, 4.4 ⫾ 3.5 mg/dL) and were ⬎7 mg/dL for 22.5% of them. Aspartate aminotransferase was elevated for 43% of the patients and alanine aminotransferase for 17% of them. Lactate dehydrogenase was not measured.

DISCUSSION This study focused on pharyngeal adenovirus infections confirmed by culture. The mean age of our patients was 37 months, and 87% of them were younger than 5 years of age. This is similar to a recent study that reported very few cases of school age children requiring hospital evaluation for adenovirus infection.3 Sixty percent of our patients were boys, close to the 2:1 ratio reported by Peled et al2 for adenovirus infections in hospitalized immunocompetent children. The seasonal distribution was a little different from that in previous studies4; although the incidence was high in the winter, the peak occurred in the spring and the incidence remained high during the summer. High grade fever in young children is of concern because of the risk of serious bacterial infection. In our study, 74% of the patients had peak temperatures of ⬎39°C and 29% had temperatures of ⬎40°C, reflecting the pyrogenicity of human adenoviruses.5 Leukocytosis and CRP concentration elevation, which are characteristic of severe bacterial infections and are often used to distinguish viral from bacterial origin, are also characteristic of adenoviral pharyngeal infections. In our study, the mean WBC count was high (14,999 leukocytes per mm3), in agreement with some studies.2,6 More importantly, 48% of the patients had ⬎15,000 leukocytes per mm3 and 24% had ⬎20,000 leukocytes per mm3, which might lead to unnecessary antibiotic therapy7 or chest radiographs.8 Appenzeller et al9 reported that adenovirus shows a clearly higher mean CRP level (4.9 mg/dL), compared with other viral respiratory tract infections (respiratory syncytial virus, 1.7 mg/dL; parainfluenza virus, 1 mg/dL; influenza virus, 2.3 mg/dL). This might be related to the high interleukin-6 values found in patients with adenoviral respiratory infections.10 For 22.5% of our patients, CRP concentrations were

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RESPIRATORY SYNCYTIAL VIRUS AND HUMAN METAPNEUMOVIRUS IN CHILDREN WITH ACUTE RESPIRATORY INFECTIONS IN YEMEN Najla Al-Sonboli, MTropPaed,*† Charles Anthony Hart, PhD,‡ Ahmed Al-Aeryani, MBBS,† Salem M. Banajeh, MBBS,† Nasher Al-Aghbari, MTropPaed,*† Winifred Dove, BSc,‡ and Luis Eduardo Cuevas, MTropMed* FIGURE 1. Seasonal pattern of pharyngeal adenovirus cases.

⬎7 mg/dL, a value that was previously used as a cutoff point to identify clinically undetectable, serious, bacterial infections.11 Our study has several limitations. First, this was a retrospective study; therefore, a protocol was not followed, and some children underwent laboratory analyses, whereas some did not. Also, pharyngeal cultures might have been performed for the patients who seemed to be more seriously affected, with higher temperatures or greater leukocytosis, for example. Second, typing of adenovirus isolates was not performed. The main conclusion of this study is that adenovirus pharyngeal infections in young children can mimic severe bacterial infections because of high fever associated with bacterial infection-like laboratory results. The development of rapid sensitive tests might help to decrease the high number of laboratory analyses and antibiotic prescriptions for these patients.

REFERENCES 1. Reina J, Ferres F, Gutie´rrez O, Ruiz de Gopegui E, Gonza´lez-Ca´rdenas M. Study of the clinical and epidemiological characteristics of respiratory infections caused by adenovirus in a pediatric population (1997– 2003) 关in Spanish兴. An Pediatr (Barc). 2004;61:137–142. 2. Peled N, Nakar C, Huberman H, et al. Adenovirus infection in hospitalized immunocompetent children. Clin Pediatr (Phila). 2004;43:223– 229. 3. Rocholl C, Gerber K, Daly J, et al. Adenoviral infections in children: the impact of rapid diagnosis. Pediatrics. 2004;113:e51– e56. 4. Irmen KE, Kelleher LL. Use of monoclonal antibodies for rapid diagnosis of respiratory viruses in a community hospital. Clin Diagn Lab Immunol. 2000;3:396 – 403. 5. Kato N. Pyrogenicity of human adenoviruses. J Gen Virol. 2000;81: 2611–2616. 6. Ruuskanen O, Mertsola J, Meurman O. Adenovirus infection in families. Arch Dis Child. 1988;63:1250 –1253. 7. Baraff LJ, Bass JW, Fleisher GR, et al. Practice guideline for the management of infants and children 0 to 36 months of age with fever without source: Agency for Health Care Policy and Research. Ann Emerg Med. 1993;22:1198 –1210. 8. Bachur R, Perry H, Harer MB. Occult pneumonias: empiric chest radiographs in febrile children with leukocytosis. Ann Emerg Med. 1999;33:166 –173. 9. Appenzeller C, Ammann RA, Duppenthaler A, et al. Serum C-reactive protein in children with adenovirus infection. Swiss Med Wkly. 2002; 132:345–350. 10. Kawasaki Y, Hosoya M, Katayose M, et al. Correlation between serum interleukin 6 and C-reactive protein concentrations in patients with adenoviral respiratory infection. Pediatr Infect Dis J. 2002;21:370 –374. 11. Pulliam PN, Attia MW, Cronan KM. C-reactive protein in febrile children 1 to 36 months of age with clinically undetectable serious bacterial infection. Pediatrics. 2001;108:1275–1279.

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Abstract: A survey of 604 Yemeni children younger than 2 years with acute respiratory infections identified respiratory syncytial virus (RSV) in 244 (40%), human metapneumovirus (HMPV) in 41 (7%) and RSV/HMPV coinfection in 25 (4%) children. RSV infections occurred in younger children and were associated with more severe hypoxia than observed with HMPV. Both viruses are important causes of severe acute respiratory infection in Yemen. Key Words: acute respiratory infections, children, human metapneumovirus, respiratory syncytial virus, Yemen Accepted for publication February 8, 2005. From the *Liverpool School of Tropical Medicine, Liverpool, United Kingdom; the †Paediatrics Department, Sana’a University, Yemen; and the ‡Department of Medical Microbiology, University of Liverpool, Liverpool, United Kingdom Supported by the Islamic Development Bank as part of Dr Al-Sonboli’s PhD studies at the Liverpool School of Tropical Medicine. Reprints not available. DOI: 10.1097/01.inf.0000172937.80719.7f

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cute respiratory infections (ARI) remain the most important cause of death in children. Although the most frequent etiologic agent of ARI in young children is respiratory syncytial virus (RSV), human metapneumovirus (HMPV) has recently been identified as an important cause of ARI worldwide.1 HMPV was described in 1 small study from Israel,2 but there are no systematic data on its frequency and importance as a cause of ARI in the Middle East. We report here the epidemiology and clinical characteristics of children with ARI caused by RSV, HMPV or both viruses in children attending a tertiary hospital in Yemen.

MATERIALS AND METHODS Children younger than 2 years of age with a clinical diagnosis of ARI attending Alsabeen Hospital for Women and Children were enrolled from October 2002 to May 2003, after informed parental consent. Alsabeen is a reference hospital serving the population of Sana’a. All the patients attending the Accident and Emergency and outpatient clinics of the hospital between 8 AM and 1 PM with ARI were recruited consecutively at the time of consultation, independently of the severity of their illness. The clinical diagnosis of ARI at the time of enrollment was based on the presence of cough, tachypnea, chest indrawing or wheeze for ⬍7 days and followed the World Health Organization standard protocol for research on ALRI. Sixty-two percent of these patients were subsequently admitted to the hospital. In addition, all patients admitted to the wards during the night were recruited the next morning. Oxygen saturations (pO2) were measured by pulse oximetry (Nonin Medical, Inc., Plymouth, MN; model 8500) before the use of oxygen to classify the children as having mild/moderate (pO2 ⱖ 88%) or severe (pO2 ⬍ 88%) ARI, as suggested by Duke et al3 for research in high altitude locations. Nasopharyngeal aspirates were obtained by direct aspiration with sterile nasopharyngeal mucus extractors, mixed with 1 mL of phosphate-buffered saline and frozen at ⫺0°C until analysis. RSV and HMPV RNA were extracted by the Qiagen RNeasy extraction method (Qiagen, Crawley, United Kingdom). © 2005 Lippincott Williams & Wilkins


RSV and HMPV in Yemen

TABLE 1. Characteristics of Children with HMPV, RSV and RSV/HMPV Infections RSV (N ⫽ 241)

HMPV (N ⫽ 41)

RSV/HMPV Coinfection (N ⫽ 25)

Negative RSV and HMPV PCR (N ⫽ 294)

Median age (mo) Male

2 158 (66)*

6 29 (70)

3 18 (72)

4 189 (64)

History of Cough Tachypnea† Fever Blue lips† Wheeze† Difficulty waking Apnea Skin rash

241 (100) 216 (90) 178 (74) 177 (73) 102 (42) 23 (10) 15 (6) 11 (5)

40 (98) 24 (59) 33 (81) 23 (56) 6 (15) 2 (5) 0 (0) 1 (4)

25 (100) 22 (88) 19 (76) 19 (76) 14 (56) 3 (12) 2 (8) 1 (4)

294 (100) 183 (62) 198 (67) 134 (46) 72 (25) 17 (6) 8 (3) 10 (3)

Physical examination Crackles† Cyanosis† Chest indrawing† Wheeze† Temperature ⬎37.5°C Respiratory rate/min ⫾ SD† Hepatomegaly Mean pO2 ⫾ SD† pO2 ⬍ 88%

201 (83) 186 (77) 173 (72) 94 (39) 47 (19) 61 ⫾ 13 9 (4) 79 ⫾ 8% 210 (87)

21 (51) 17 (42) 17 (42) 9 (22) 11 (27) 50 ⫾ 15 3 (7) 84 ⫾ 8.2% 24 (59)

22 (88) 21 (84) 19 (76) 9 (36) 2 (8) 57 ⫾ 15 1 (4) 76% ⫾ 8.6% 22 (88)

156 (53) 137 (47) 117 (40) 63 (21) 67 (23) 54 ⫾ 14 18 (6) 83% ⫾ 9.6% 167 (59)

Variable

*Numbers in parentheses, percent. † P ⬍ 0.001.

HMPV and RSV genomes were detected by reverse transcriptionerase chain reaction amplification. To detect HMPV, we used primers amplifying the M gene between nucleotides 212 and 331 giving a 121-bp product, HMPVMF1 (5⬘-AAGTGAATGCATCAGCCCAAG-3⬘) and HMPVMR1 (5⬘-CACAGACTGTGAGTTTGTCAAA-3⬘). For the RSV genome, we used primers amplifying the N gene between nucleotides 858 and 1135 giving a 278-bp product. N1 primer (5⬘-GGAACAAGTTGTTGAGGTTTATGAATATGC-3⬘) and N2 primer (5⬘-CTTCTGCTGTCAAGTCTAGTACACTGTAGT-3⬘) were used.4 Ten percent of the HMPV positive by M gene amplification were confirmed by reverse transcription-polymerase chain reaction (PCR) amplification with L6 CAT GCC CAC TAT AAA AGG TCAG and L7 CAC CCC AGT CTT TCT TGA AA primers to amplify the polymerase gene.4 The DNA was purified by running it on a low melting temperature agarose (Nusieve GTG agarose; BioWhittaker Molecular Application). Selected PCR products were cloned into a TA cloning vector (Qiagen, pGEM-T; Promega, Crawley, United Kingdom) and transferred into Escherichia coli.4 After plasmid purification (Qiagen), 20 ␮L of purified DNA were sequenced to confirm identity. RSV PCR amplicons were subjected to restriction enzyme digestion without prior purification using HindIII, PstI, BglII, HaeIII and RsaI (Roche) and typed into NP genotypes on the basis of their digestion profiles.5 Ethical approval was obtained from the Research Ethics committees of the Liverpool School of Tropical Medicine and Alsabeen Hospital for Women and Children. RESULTS We recruited 604 children (395, 66% male) with a median age of 4 months as shown in Table 1. Of these, 179 (30%) children had pO2 ⱖ 88% and 425 had pO2 ⬍ 88%, reflecting the characteristics of patients referred to a tertiary hospital. Of these, 244 (40%) children had RSV, 41 (7%) had HMPV and 25 (4%) had both RSV and HMPV. Of the 241 children with RSV and clinical information, 210 (87%) had pO2 ⬍ 88%. In comparison, 24 (59%) of the 41 children with HMPV and 22 (88%) of 25 with RSV/HMPV coin© 2005 Lippincott Williams & Wilkins

fection had pO2 below this cutoff. In agreement with the pO2 concentrations, 186 (76%) children with RSV, 20 (80%) with RSV/HMPV and 19 (46%) with HMPV were hospitalized. confirming that RSV was associated with more severe disease (P ⬍ 0.01). Children with HMPV were also older than those with RSV or RSV/HMPV coinfections (P ⬍ 0.01); RSV was identified more frequently in children younger than 6 months of age, whereas HMPV occurred in older children. Children with RSV were more likely to have bronchospasm, crackles, cyanosis and chest indrawing, and the parents perceived wheezing and tachypnea more often than in children with HMPV. Children coinfected with RSV/HMPV, however, had symptoms resembling those of RSV. Younger children were also more likely to have hypoxia; this was more marked in children with RSV given that 90% of the children with RSV who were younger than 6 months of age had pO2 ⬍ 88% compared with 80% of those 6 –12 month old and 67% older than 1 year. In comparison, 62% of the children with HMPV younger than 6 months of age had pO2 ⬍ 88%, compared with 64% of those 6 –12 months old and 45% older than 1 year. Children with RSV/HMPV coinfection had a similar degree of hypoxia compared with those with RSV. ARIs increased from October and peaked in January, and RSV and HMPV infections coincided during the study period although HMPV peaked 1 month later than RSV. Group A RSV were identified in 171 (82%) and group B in 37 (18%) of 208 positive nasopharyngeal aspirates. Children with severe ARI seemed to have a higher prevalence of group B, but this was not statistically significant. Ninety-six (46%) RSV were NP2 genotype, 75 (36%) NP4, 25 (12%) NP3 and 1 (5%) NP1. NP1 and NP2 were identified in all 8 months of the study but NP1 and NP3 were absent during April and May. Six children with RSV and 1 with HMPV died (P ⬎ 0.05). DISCUSSION Studies across the world have reported that HMPV is responsible for 0 –24% of ARI,6 although during epidemics it can represent up to 50% of cases. This is the first report of HMPV from the Arabian Peninsula and confirms its importance as a

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cause of ARI in Yemen. Our findings also confirm that RSV is the most frequent pathogen causing severe ARI in young children and that coinfection with both viruses often occurs. Although we studied only 8 months of the year, RSV and HMPV had marked seasonal variations with an RSV peak in December and January and an HMPV peak in February and March. This agrees with reports of HMPV having a peak in the first months of the year in the Northern hemisphere and during April and May in the Southern hemisphere.6 RSV most frequently affected patients younger than 6 months of age, whereas HMPV affected slightly older children. This is similar to the age distribution of the patients included in original reports from the Netherlands1 and elsewhere7 and might be one of the reasons why infection with HMPV results in less severe clinical symptoms. Although it was not possible to differentiate the 2 infections solely on clinical findings, wheezing was less frequent in children with HMPV. This is consistent with studies from Hong Kong and Japan8,9 but different to Canada where in a small study 12 children with HMPV had more wheezing (83%) that those with RSV (65%).10 The milder presentation of HMPV could be explained in part by the older age of the children, because they would have larger respiratory airways and fewer maternally derived antibodies. Consistent with this hypothesis is the fact that younger children with HMPV infection had a higher frequency of hypoxia than older children. It has also been suggested that RSV/HMPV coinfection results in increased episode severity.4 Although a higher proportion of our children with dual infections had severe hypoxia that children with single viral infections, the number of coinfected children was too small for a meaningful statistical analysis. Information on the relative frequencies of group A and B RSV in the Middle East and Africa is scarce. Group A RSV was predominant in our study and in Jordan, group A RSV predominated during 3 consecutive years.11 This is consistent with the worldwide predominance of group A, although group B has been reported as the main pathogen in some years. Our data confirm that HMPV is a significant pathogen in children younger than 2 years of age in the Middle East, and further studies are necessary to quantify its burden of disease and delineate its pathogenicity mechanisms to inform potential treatment and prevention strategies. REFERENCES 1. van den Hoogen BG, de Jong JC, Groen J, et al. A newly discovered human pneumovirus isolated from young children with respiratory tract disease. Nat Med. 2001;7:719 –724. 2. Wolf DG, Zakay-Rones Z, Fadeela A, Greenberg D, Dagan R. High seroprevalence of human metapneumovirus among young children in Israel. J Infect Dis. 2003;188:1865–1867. 3. Duke T, Blaschke AJ, Sialis S, Bonkowsky JL. Hypoxaemia in acute respiratory and non-respiratory illnesses in neonates and children in a developing country. Arch Dis Child. 2002;86:108 –112. 4. Greensill J, McNamara PS, Dove W, Flanagan B, Smyth RL, Hart CA. Human metapneumovirus in severe respiratory syncytial virus bronchiolitis. Emerg Infect Dis. 2003;9:372–375. 5. Cane PA, Pringle CR. Molecular epidemiology of respiratory syncytial virus: rapid identification of subgroup A lineages. J Virol Methods. 1992;40:297–306. 6. Cuevas LE, Nasser AM, Dove W, Gurgel RQ, Greensill J, Hart CA. Human metapneumovirus and respiratory syncytial virus, Brazil. Emerg Infect Dis. 2003;9:1626 –1628. 7. Ebihara T, Endo R, Kikuta H, Ishiguro N, Ishiko H, Kobayashi K. Comparison of the seroprevalence of human metapneumovirus and human respiratory syncytial virus. J Med Virol. 2004;72:304 – 306.

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8. Peiris JS, Tang WH, Chan KH, et al. Children with respiratory disease associated with metapneumovirus in Hong Kong. Emerg Infect Dis. 2003;9:628 – 633. 9. Ebihara T, Endo R, Kikuta H, et al. Human metapneumovirus infection in Japanese children. J Clin Microbiol. 2004;42:126 –132. 10. Boivin G, De Serres G, Cote S, et al. Human metapneumovirus infections in hospitalized children. Emerg Infect Dis. 2003;9:634 – 640. 11. Bdour S. Respiratory syncytial virus subgroup A in hospitalized children in Zarqa, Jordan. Ann Trop Paediatr. 2001;21:253–261.

RISK FACTORS FOR MORTALITY IN CHILDREN WITH CANDIDEMIA Theoklis E. Zaoutis, MD,*‡§ Susan E. Coffin, MD, MPH,*‡ Jaclyn H. Chu, MHS,* Kateri Heydon, MS,* Huaqing Zhao, MA,† H. Mollie Greves, MD,储 and Thomas J. Walsh, MD¶ Abstract: We performed a retrospective cohort study of hospitalized children with positive blood cultures for Candida species. Independent risk factors for mortality by multivariable analysis were location in the pediatric intensive care unit at the time of infection (hazard ratio, 6.3; 95% confidence interval, 1.6 –24.3) and the presence of an arterial catheter (hazard ratio, 2.4; 95% confidence interval, 1.1–5.8). Our findings help identify a group of pediatric patients that should be targeted for future interventions to prevent and treat candidemia. Key Words: pediatric, candidemia, fungal infections, invasive candidiasis, mortality Accepted for publication March 2, 2005. From the *Divisions of Infectious Diseases and †Biostatistics and Epidemiology, The Children’s Hospital of Philadelphia, and the ‡Department of Pediatrics and the §Center for Clinical Epidemiology and Biostatistics, Center for Education and Research on Therapeutics, University of Pennsylvania School of Medicine, Philadelphia, PA; the 㛳Department of Pediatrics, Children’s Hospital and Regional Medical Center, University of Washington School of Medicine, Seattle, WA; and the ¶Immunocompromised Host Section, Pediatric Oncology Branch, National Cancer Institute, Bethesda, MD Supported in part by Center for Education and Research on Therapeutics grant U-18-HS10399 from the Agency of Healthcare Research and Quality. Presented in part at the Pediatric Academic Societies Meeting, 2003, Seattle, WA, and at the 13th Annual Meeting of the Society for Healthcare Epidemiology, April 5– 8, 2003, Arlington, VA. Address for reprints: Theoklis E. Zaoutis, MD, Division of Infectious Diseases, The Children’s Hospital of Philadelphia, 34th and Civic Center Boulevard, Philadelphia, PA 19104. Fax 215-590-2025; E-mail [email protected]. DOI: 10.1097/01.inf.0000172938.76561.8e

C

andidemia is a frequent and potentially life-threatening complication experienced by many chronically or critically ill children.1 Infections caused by Candida are the most common invasive fungal infections in hospitalized patients, and these organisms have been identified as the fourth most common cause of nosocomial bloodstream infection in the United States.2 Candidemia has also been associated with significant morbidity and mortality. Mortality rates in children with candidemia are from 19 –26% to as high as 43–54% among infants.3– 8 Candidemia is often associated with signs and symptoms of sepsis and fungal infections possess the second highest case fatality rate (13%) among all causes of sepsis in children.9 To date, most studies of risk factors for mortality in patients with candidemia have focused on adult populations. However, risk factors for mortality identified in adults may not be relevant in © 2005 Lippincott Williams & Wilkins


pediatric patients. For example, hospitalized children with candidemia often have unique underlying diagnoses and comorbidities that might result in a different set of predictors of mortality than in adults.6 This study was undertaken to determine the risk factors for mortality in a large cohort of hospitalized children with candidemia. METHODS This retrospective cohort study was conducted at a large tertiary care children’s hospital. All study patients were identified through a systematic record review of the clinical microbiology laboratory. All patients who had an episode of candidemia (defined as a positive blood culture for any Candida species obtained from a peripheral vessel or vascular catheter) from January 1, 1998 through December 31, 2001 were eligible for inclusion in the study. If a patient experienced multiple episodes of infection during the study period, only the first episode was included in the analysis of risk factors for mortality, because the risk of death could change with subsequent episodes of candidemia. Data on demographic and clinical characteristics, presence of comorbid conditions, antifungal treatment and infecting Candida species were collected on all patients. To assess a possible association between total placement duration and disposition of an intravascular catheter during infection and the risk of death, detailed information on exposure to central venous catheters (CVCs) during infection was collected. A catheter was considered replaced if the patient was still candidemic (positive blood cultures for Candida) on the day of new catheter placement. The presence of arterial catheters was also assessed. The primary outcome was defined as in-hospital death (from any cause) within 1 month (30 days) of the initial positive culture for Candida. Univariate analyses were conducted to determine the association between potential risk factors and in-hospital death among children with candidemia. Categoric variables were compared by Fisher’s exact test, and continuous variables were compared by the Wilcoxon rank sum test. All variables with a P value of ⬍0.10 on univariate Cox analysis were considered for inclusion in the multivariable model, as well as other variables determined a priori to be key factors for predicting mortality among patients with candidemia (eg, cancer, neutropenia, prematurity, total parenteral nutrition, CVC use, antifungal treatment, transplantation). We assessed central venous catheter exposure during infection and antifungal use as timedependent variables, because these exposures can change over the course of observation, respectively. A 2-tailed P value of ⬍0.05 was considered statistically significant. Statistical calculations were performed with the computing packages Stata version 8.0 (Stata Corp., College Station, TX) and SAS version 9.1 (SAS Corp., Cary, NC). RESULTS We identified a total of 172 eligible patients during the study period. Eleven patients (6%) had 2 or more episodes of infection. Of the 172 patients, complete records were available for review for 168 (98%) of the patients. The microbiology data for this cohort have been previously reported.10 In summary, Candida albicans accounted for 43% of the cases of candidemia followed by Candida parapsilosis (27%), Candida glabrata (10%), Candida tropicalis (10%), Candida krusei (3%) and Candida lusitaniae (2%). Table 1 summarizes the patient demographics and the univariate analysis of risk factors for mortality. Overall 29 (17%) children died within 1 month of the first positive culture of Candida. The types of antifungal therapy administered after the onset of candidemia are reported in Table 1. Twelve patients were not treated with any antifungal therapy after the first positive Candida © 2005 Lippincott Williams & Wilkins

Fatal Candidemia

culture was reported, for reasons including death before receipt of culture results, clinician determination of a catheter-related infection that resolved after a line was pulled or unexplained resolution of infection by subsequent negative culture without initiation of antifungal treatment. The multivariable Cox proportional hazard model revealed that patients located in the pediatric intensive care unit (PICU) at the time of infection were at higher risk for death of when compared with patients located outside the PICU at the time of infection 关hazard ratio (HR); 6.3, 95% confidence interval (CI), 1.6 –24.3兴. The overall 30-day mortality rate for patients in the PICU was 37% compared with 10% for patients not located in the PICU. The other independent predictor for death in children with candidemia was the presence of an arterial line (HR 2.4; 95% CI 1.1–5.8). Neutropenia approached but did not reach statistical significance (HR 2.4; 95% CI 0.09 – 6.2). In multivariate analysis, we found that time-dependent exposure to catheters and antifungal therapies during infection were not significantly associated with mortality.

DISCUSSION In our cohort of 168 children with candidemia, we found that significant independent risk factors for death were location in the PICU and the presence of an arterial catheter at the time of infection. Both of these variables are likely strong markers for severity of illness. Thirty-seven percent of children with candidemia in the PICU died compared with 10% in all other areas including the neonatal intensive care unit. No association was seen between the time with a central venous catheter while blood cultures remained positive for Candida or duration of antifungal use and death. The mortality rate in children with candidemia has been reported to range between 17–24%.3– 8 Few studies have reported PICU-specific rates of mortality rates. One study of PICU patients from Spain reported a crude mortality rate of 22%.7 The variability in rates may reflect different definitions of mortality (ie, 30-day, all-cause, in-hospital). Multiple studies have reported risk factors for mortality in adult patients with candidemia. However, the outcomes of children with candidemia have not been extensively studied. Pappas et al,6 in a prospective cohort study that included 144 children, identified neutropenia and endotracheal intubation as independent risk factors for mortality. In our study, patients with neutropenia had a higher HR for death, but this did not reach statistical significance. Because endotracheal intubation occurs only in an intensive care unit, our finding of an association between presence in the PICU and an increased risk of death is probably comparable with the findings of Pappas et al.6 Furthermore we found no association between duration of CVC use while patients were candidemic and increased mortality. However, our sample size might have not been sufficient to detect a small difference if one existed. Amphotericin B was the drug of choice for the treatment of candidemia in children. This can reflect a more conservative approach to the management of children with candidemia as well as the lack of clinical trials of azoles and other newer antifungal agents in children. Although fluconazole was available during the study period, newer triazole antifungals such as voriconazole and echinocandins (caspofungin) were not available. These and other newer antifungal agents can change the treatment strategies used to treat candidemia in children in the future. Previous studies have provided evidence supporting the recommendation that all candidemic patients should receive antifungal therapy.6,11 Our analysis revealed that almost all children with candidemia who did not receive antifungal therapy survived. However, this finding is limited by the small number of patients in this group and should not be considered as evidence for not treating children with candidemia.

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TABLE 1. Risk Factors for Mortality in Children With Candidemia Variable Demographics Age (yr), median Gender Female Hospital characteristics ICU Non-ICU PICU NICU LOS preceding candidemia (d), median Total LOS (d), median Clinical characteristics Cancer Leukemia and lymphoma Solid organ History of transplant Bone marrow Solid organ Neutropenia† Prematurity Prior surgery‡ Immunosuppression§ Total parenteral nutrition† Antifungal prophylaxis (fluconazole) Characteristics of infection Infecting Candida spp. Candida albicans Other Candida spp. Days culture-positive, median CVCs CVC burden† No CVC 1 CVC 2 or more CVCs Catheter removed Catheter rewired Catheter replaced Arterial catheter present† Antifungal treatment None Amphotericin B formulation only Amphotericin B formulation followed by fluconazole Fluconazole only Fluconazole followed by amphotericin B formulation Amphotericin B formulation ⫹ 5FC Amphotericin B formulation ⫹ 5FC ⫹ fluconazole Other antifungal therapy

Alive (N ⫽ 139)

Died (N ⫽ 29)

3.1 (0.7–14.4)*

4.8 (0.2–14.0)*

Total (N ⫽ 168) 3.6 (0.7–14.4)*

66

17

83 (49%)

76 40 23 12 (2, 19) 38 (18, 79)

3 23 3 18 (9, 32) 29 (13, 46)

79 (47%) 63 (38%) 26 (15%) 13 (3–27) 35 (18 – 69)

P

0.92 0.31

⬍0.001

0.02 0.05

0.69 16 17

5 3

21 (13%) 20 (12%)

9 5 17 29 44 37 88 9

6 0 9 7 10 8 24 8

15 (9%) 5 (3%) 26 (15%) 36 (21%) 54 (32%) 45 (27%) 112 (67%) 17 (10%)

58 81 1 (1, 3)

14 15 1 (1, 5)

0.05 0.02 0.80 0.82 0.99 0.05 ⬍0.01 0.54 72 (43%) 96 (57%) 1 (1–3)

0.60

0.05 11 119 9 76 6 23 21

1 22 6 11 2 7 18

12 (7%) 141 (84%) 15 (9%) 87 (87%) 8 (5%) 30 (18%) 39 (23%)

9 84 19 2 3 13 9 0

3 17 2 3 0 0 1 3

12 (7%) 101 (60%) 21 (13%) 5 (3%) 3 (2%) 13 (8%) 10 (6%) 3 (2%)

0.11 0.62 0.42 ⬍0.001

*Numbers in parentheses, interquartile range, unless otherwise noted. † At time of infection (absolute neutrophil count, ⬍500 mm3). ‡ Within 30 days before infection. § Use of corticosteroids, calcineurin inhibitors, or chemotherapeutic agents in the 2 weeks preceding infection. ICU indicates intensive care unit; LOS, length of stay; 5FC, 5-fluorocytosine.

Several potential limitations exist in our study. We defined 30-day mortality as the outcome of interest. Although this defined end point is unambiguous, it does not differentiate according to the cause of death so that some deaths may not have been directly attributable to candidemia. Autopsies were not performed on most of diseased patients. In addition, efforts to designate outcomes as “attributable” to a specific infection are often very subjective and inconsistently applied.12 Therefore our identified associations do not necessarily imply biologic causality. However, our results may indicate a secondary effect of candidemia in worsening the health of critically ill pediatric patients that subsequently may increase the risk of death. Additionally we did not assess birth weight or

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antifungal resistance as predictors of mortality, which may have resulted in residual confounding. Finally our study was conducted in a large academic tertiary care medical children’s hospital, and the results might thus not be generalizable to other institutions. REFERENCES 1. Hostetter MK. New insights into candidal infections. Adv Pediatr. 1996;43:209 –230. 2. Edmond MB, Wallace SE, McClish DK, Pfaller MA, Jones RN, Wenzel RP. Nosocomial bloodstream infections in United States hospitals: a three-year analysis. Clin Infect Dis. 1999;29:239 –244.



3. Saiman L, Ludington E, Pfaller M, et al. Risk factors for candidemia in neonatal intensive care unit patients: the National Epidemiology of Mycosis Survey study group. Pediatr Infect Dis J. 2000;19:319 –324. 4. Stamos JK, Rowley AH. Candidemia in a pediatric population. Clin Infect Dis. 1995;20:571–575. 5. MacDonald L, Baker C, Chenoweth C. Risk factors for candidemia in a children’s hospital. Clin Infect Dis. 1998;26:642– 645. 6. Pappas PG, Rex JH, Lee J, et al. A prospective observational study of candidemia: epidemiology, therapy, and influences on mortality in hospitalized adult and pediatric patients. Clin Infect Dis. 2003;37:634 – 643. 7. Rodriguez-Nunez A. Incidence and mortality of proven invasive Candida infections in pediatric intensive care patients. Infect Control Hosp Epidemiol. 2001;22:477– 478. 8. Turner RB, Donowitz LG, Hendley JO. Consequences of candidemia for pediatric patients. Am J Dis Child. 1985;139:178 –180. 9. Watson RS, Carcillo JA, Linde-Zwirble WT, Clermont G, Lidicker J, Angus DC. The epidemiology of severe sepsis in children in the United States. Am J Respir Crit Care Med. 2003;167:695–701. 10. Zaoutis TE, Greves HM, Lautenbach E, Bilker WB, Coffin SE. Risk factors for disseminated candidiasis in children with candidemia. Pediatr Infect Dis J. 2004;23:635– 641. 11. Rex JH, Walsh TJ, Sobel JD, et al. Practice guidelines for the treatment of candidiasis: Infectious Diseases Society of America. Clin Infect Dis. 2000;30:662– 678. 12. Cosgrove SE, Carmeli Y. The impact of antimicrobial resistance on health and economic outcomes. Clin Infect Dis. 2003;36:1433–1437.

FREQUENT HAEMOPHILUS INFLUENZAE TYPE B COLONIZATION IN RURAL THAILAND Sonja J. Olsen, PhD,* Surang Dejsirilert, MSc,† Leelawadee Sangsuk, MSc,† Supamit Chunsutiwat, MD, MPH,‡ and Scott F. Dowell, MD, MPH* Abstract: In Asia, there is uncertainty regarding whether the burden of Haemophilus influenzae type b (Hib) disease is great enough to warrant vaccination. In this nasopharyngeal carriage study in rural Thailand, 7% of children younger than 5 years of age carried Hib, a prevalence similar to that found in countries with high rates of Hib disease before vaccine introduction. Key Words: Haemophilus influenzae, carriage, resistance, Asia Accepted for publication March 2, 2005. From the *International Emerging Infections Program, Centers for Disease Control and Prevention-Thai Ministry of Public Health Collaboration, Nonthaburi, Thailand; and †National Institute of Health and ‡Department of Disease Control, Ministry of Public Health, Nonthaburi, Thailand Reprints not available. DOI: 10.1097/01.inf.0000172940.77549.94

S

ince the late 1980s, the use of conjugate vaccines against Haemophilus influenzae type b (Hib) has resulted in a 98% decline in the incidence of Hib disease in young children in the United States; several other wealthy countries have seen similar results.1,2 Although it is well recognized that Hib can cause bacterial meningitis and pneumonia in Asian children,3,4 there is uncertainty about the true burden of Hib disease and whether it is sufficient to warrant routine vaccination, given limited health care resources. What are needed are studies with well defined populations that measure the burden of invasive disease with the highest quality microbiologic techniques. Carriage of Hib bacteria in the nasopharynx may be helpful in assessing the possible burden of Hib disease in a country; high rates of colonization often, although not invariably, correlate with high rates of invasive disease. There are several high risk groups, such as Alaskan natives and some African populations, that have the highest © 2005 Lippincott Williams & Wilkins

Hib Colonization

rates of invasive Hib disease and corresponding high carriage rates ranging from 7% to 15%.5– 8 Before vaccination, the prevalence of colonization in Alaskan natives was ⬃2 times higher than the rates in the U.S. general population, and rates of disease were 5 times higher.9 Hib disease and carriage rates in the United States and Europe in the prevaccination era were lower, with carriage rates in the range of 3– 6%.10 –12 In contrast, studies in Asian nations typically found very low invasive disease incidence and carriage rates of ⬃0 –1%.13–15 Here we present results from a Hib carriage study conducted in rural Thailand that suggest that Hib is widely circulating and colonization is relatively common. METHODS As part of a study of influenza and other respiratory pathogens, patients with influenza-like illness at 5 hospital outpatient departments were enrolled in Sa Kaeo Province, Thailand. Influenza-like illness was defined by using the WHO criteria of acute fever of ⬎38°C within the past 3 days and either cough or sore throat in the absence of another diagnosis.16 After informed written consent was obtained from the patient or guardian, trained nurses administered a brief questionnaire to the patient or guardian, placed a calcium alginate swab in one nostril to the nasopharynx and rotated the swab once, to collect epithelial tissue and to absorb secretions. The specimen was immediately placed in skim milk-tryptone-glucose-glycerin bacterial transport medium and was stored at 2– 8°C for ⬍48 hours until transport to the provincial hospital for storage at ⫺70°C. Specimens were transported once weekly, on dry ice, to the Thai National Institute of Health laboratory in Bangkok for bacterial isolation. Each nasopharyngeal swab in skim milk-tryptone-glucose-glycerin medium was plated onto chocolate agar with X and V factors and 0.3 mg/mL bacitracin. Serotyping was performed with latex agglutination assays, using a Pastorex meningitis/Hib kit (Bio-Rad Laboratories, Marnes-la-Coquette, France) and standard methods. Antimicrobial susceptibility testing was performed with broth microdilution assays, and the 2003 National Committee for Clinical Laboratory Standards criteria were used for interpretation. ␤-Lactamase production was detected with nitrocefin disks (Cephinase disks; BBL Microbiology, Becton Dickinson). Quality assurance included periodic monitoring for percentage Streptococcus pneumoniae isolation, with retraining of nurses when the percentage dropped below 25%. In addition, there was regular testing of chocolate agar batches with standard stock strains. Data were analyzed with SPSS version 12.0 (SPSS, Chicago, IL). The study was approved by the institutional review board at the CDC and the ethics review committee at the Thai Ministry of Public Health. RESULTS Prevalence of H. influenzae. Between August 15, 2003, and August 14, 2004, 1050 outpatients with influenza-like illness were enrolled; 541 (52%) were male. Four hundred ninety-two (47%) were younger than 5 years of age, 415 (40%) were 5–17 years of age and 143 (14%) were ⱖ18 years of age. At no time during the year did the frequency of S. pneumoniae isolation drop below 25%. H. influenzae was isolated in 383 (36%) cases, ie, 36 (9%) cases in infants younger than 1 year of age, 58 (15%) in children 12–23 months of age, 124 (32%) in children 24 –59 months of age, 147 (38%) in children 5–17 years of age and 18 (5%) in adults ⱖ18 years of age. Hib was isolated in 67 cases, for an overall colonization prevalence of 6.4%. Hib carriage rates were highest in young children, ie, 4 (6%) cases in infants younger than 1 year of age, 7 (10%) in children 12–23 months of age, 24 (36%) in children 24 –59 months of age, 27 (7%) in children 5–17 years of age and 5 (4%) in adults ⱖ18 of age. Hib colonization by month was as follows: 4% in August, 35% in

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FIGURE 1. Hib carriage in young children. ■ indicates data from the current study; 䊐, healthy populations; u, sick or mixed populations. Reference numbers are given in parentheses after the country names. September and October, 30% in November, 40% in December, 14% in January, 5% in February, 30% in March, 0% in April, 9% in May, 7% in June and 18% in July. Figure 1 compares the Hib colonization prevalence in children from published studies in the prevaccination era.5,7,8,10 –15,17–35 All studies except 7 used nasopharyngeal specimens; the 7 that used throat swabs detected carriage across a range of prevalence from 0.6% to 6.8%. Risk Factors. The times between illness onset and date of specimen collection were similar for persons with and without Hib (2.6 days versus 2.4 days; P ⫽ 0.4). The presence of a household smoker 关51% for colonized versus 51% for noncolonized; odds ratio (OR) 1.0; 95% confidence interval (CI) 0.6 –1.6兴, a chronic health condition (6% for colonized versus 7% for noncolonized; OR 0.8; 95% CI 0.3–2.4) or having another ill household member (25% for colonized versus 26% for noncolonized; OR 1.0; 95% CI 0.6 –1.7) was not associated with Hib carriage. There was no difference in the median number of people sleeping in the household ⱖ15 nights per month (4 for colonized versus 4 for noncolonized; P ⫽ 0.4) or the number of people younger than 5 years of age living in the house (1 versus 1; P ⫽ 0.8). Among children younger than 5 years of age, 40% of persons with Hib colonization and 38% of persons without colonization reported taking antibiotics in the past 3 months (P ⫽ 0.8). Antimicrobial Susceptibility. Of the 383 H. influenzae isolates, 103 (27%) produced ␤-lactamase. An additional 2 (1%) were ␤-lactamase negative and ampicillin resistant. Two hundred-twenty (57%) isolates were not susceptible to trimethoprim-sulfamethoxazole, 57 (15%) to chloramphenicol and none to cefuroxime, ceftriaxone or cefotaxime. Of the 67 Hib isolates, 22 (33%) produced ␤-lactamase and none were ␤-lactamase negative and ampicillin resistant. Thirtyfour (51%) isolates were not susceptible to trimethoprim-sulfamethoxazole, 11 (16%) to chloramphenicol and none to cefuroxime, ceftriaxone or cefotaxime.

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DISCUSSION In this study in rural Thailand, the prevalence of Hib carriage was relatively high, especially in young children. Our findings of high carriage rates are consistent with a study conducted in Bangkok in 1979, which found that 9 –17% of children in an orphanage were colonized with Hib after several cases of invasive disease were recognized.19 High rates of colonization are often found in populations with a correspondingly high incidence of Hib disease. Despite these moderately high carriage rates, estimates of the incidence of Hib disease in children younger than 5 years of age in Thailand, based on data from the national passive surveillance system, are low, ranging from 0.92 to 5.86 cases per 100,000 for Hib meningitis and from 8.83 to 80.0 cases per 100,000 for Hib pneumonia.36 These rates are 1.3– 4.5 times lower than prevaccination rates in the United States and Canada.37 A recent population-based study in northern Thailand was able to document an annual incidence of Hib meningitis of 3.8 cases per 100,000; however, the authors speculated that this might be a minimal estimate, for the same reasons that have limited other studies, including laboratory limitations and pretreatment with antibiotics.38 Although carriage and incidence of invasive disease are correlated, there may be differences in the types of strains from invasive and noninvasive sites that make inferences from colonization to invasive disease difficult. For example, one study found that more virulent strains were unable to colonize the nasopharynx.39 Although our data suggest that the bacteria are circulating widely, studies with well defined populations that measure the burden of invasive disease with the highest quality microbiologic techniques are needed. One feature of our study that merits comment is the enrollment of children with influenza-like illness, as opposed to healthy © 2005 Lippincott Williams & Wilkins


children. Although the prevalence of Hib carriage has been higher in some ill populations than in the corresponding healthy ones, the prevalence of Hib colonization in healthy and sick children does not appear to vary widely.40 We emphasize that caution should be used when extrapolating between studies, given the different methods and populations studied. Because carriage is directly related to exposure, other factors, such as household crowding or the number of children in a household, may influence Hib colonization rates,40 although we did not document this in rural Thailand. The Hib vaccine has been remarkably successful in reducing Hib disease. One reason is that childhood Hib vaccination has resulted in decreased infection rates in both vaccinated and unvaccinated children, through a herd effect.37 Vaccination either delays or prevents nasopharyngeal carriage, which is the ecologic niche for the bacteria. Remaining pockets of disease are largely attributed to undervaccination.41 Similar to results of the other study conducted in Thailand, there was moderate resistance to trimethoprim-sulfamethoxazole and ampicillin.19 Resistance to chloramphenicol was low (11%), and there was no resistance to the second-generation cephalosporin cefuroxime or the third-generation cephalosporins ceftriaxone and cefotaxime, which are commonly used to treat serious cases of invasive Hib disease. Although this study suggests that Haemophilus is widely circulating in rural Thailand, the question of whether the incidence of disease is high enough to warrant use of the vaccine remains. On the basis of estimates from existing data, the Thai Ministry of Public Health has not yet recommended adoption of routine infant vaccination in the public sector basic immunization program. However, our findings are provocative. Recent developments in vaccine production, such as synthetically produced Hib vaccine,42 may make less costly vaccination possible in the near future.

REFERENCES 1. Bisgard KM, Kao A, Leake J, Strebel PM, Perkins BA, Wharton M. Haemophilus influenzae invasive disease in the United States, 1994 – 1995: near disappearance of a vaccine-preventable childhood disease. Emerg Infect Dis. 1998;4:229 –237. 2. Peltola H. Worldwide Haemophilus influenzae type b disease at the beginning of the 21st century: global analysis of the disease burden 25 years after the use of the polysaccharide vaccine and a decade after the advent of conjugates. Clin Microbiol Rev. 2000;13:302–317. 3. Peltola H. Spectrum and burden of severe Haemophilus influenzae type b diseases in Asia. Bull World Health Organ. 1999;77:878 – 887. 4. Lolekha S, Cooksley G, Chan V, et al. A review of Hib epidemiology in Asia. Southeast Asian J Trop Med Public Health. 2000;31:650 – 657. 5. Hall DB, Lum MK, Knutson LR, Heyward WL, Ward JI. Pharyngeal carriage and acquisition of anticapsular antibody to Haemophilus influenzae type b in a high-risk population in southwestern Alaska. Am J Epidemiol. 1987;126:1190 –1197. 6. Das BK, Arora NK, Mathur P, et al. Nasopharyngeal carriage of Haemophilus influenzae. Indian J Pediatr. 2002;69:775–777. 7. Adegbola RA, Mulholland EK, Secka O, Jaffar S, Greenwood BM. Vaccination with a Haemophilus influenzae type b conjugate vaccine reduces oropharyngeal carriage of H. influenzae type b among Gambian children. J Infect Dis. 1998;177:1758 –1761. 8. Hussey GD, Coetzee G, Hitchcock J, van Schalkwyk E, van Wyk H, Kibel M. Carriage of Haemophilus influenzae in Cape Town children. S Afr Med J. 1994;84:135–137. 9. Millar EV, O’Brien KL, Levine OS, Kvamme S, Reid R, Santosham M. Toward elimination of Haemophilus influenzae type b carriage and disease among high-risk American Indian children. Am J Public Health. 2000;90:1550 –1554. 10. Michaels RH, Poziviak CS, Stonebraker FE, Norden CW. Factors affecting pharyngeal Haemophilus influenzae type b colonization rates in children. J Clin Microbiol. 1976;4:413– 417.


Hib Colonization

11. Takala AK, Eskola J, Leinonen M, et al. Reduction of oropharyngeal carriage of Haemophilus influenzae type b (Hib) in children immunized with an Hib conjugate vaccine. J Infect Dis. 1991;164:982–986. 12. Barbour ML, Mayon-White RT, Coles C, Crook DW, Moxon ER. The impact of conjugate vaccine on carriage of Haemophilus influenzae type b. J Infect Dis. 1995;171:93–98. 13. Hu YY, Yu SJ, Liu G, Gao W, Yang YH. Antimicrobial susceptibility of Haemophilus influenzae among children in Beijing, China, 1999 – 2000. Acta Paediatr. 2002;91:136 –140. 14. Sung RY, Ling JM, Fung SM, et al. Carriage of Haemophilus influenzae and Streptococcus pneumoniae in healthy Chinese and Vietnamese children in Hong Kong. Acta Paediatr. 1995;84:1262–1267. 15. Kuroki H, Ishikawa N, Uehara S, Himi K, Sonobe T, Niimi H. Nasopharyngeal colonization with Haemophilus influenzae type b among infants and children in Japan. Acta Paediatr Jpn. 1997;39:541– 545. 16. World Health Organization. WHO Recommended Surveillance Standards. Geneva, Switzerland: World Health Organization; 1999. 17. Homoe P, Prag J, Farholt S, et al. High rate of nasopharyngeal carriage of potential pathogens among children in Greenland: results of a clinical survey of middle-ear disease. Clin Infect Dis. 1996;23:1081–1090. 18. Manary MJ, Lehmann D, Michael A, et al. Antigenuria in healthy Papua New Guinean children with nasal Haemophilus influenzae type b carriage. Ann Trop Paediatr. 1993;13:385–389. 19. Simasathien S, Duangmani C, Echeverria P. Haemophilus influenzae type b resistant to ampicillin and chloramphenicol in an orphanage in Thailand. Lancet. 1980;2:1214 –1217. 20. Gomez E, Moore A, Sanchez J, et al. The epidemiology of Haemophilus influenzae type b carriage among infants and young children in Santo Domingo, Dominican Republic. Pediatr Infect Dis J. 1998;17:782–786. 21. Huebner RE, Wasas A, Mushi A, Mazhani L, Klugman K. Nasopharyngeal carriage and antimicrobial resistance in isolates of Streptococcus pneumoniae and Haemophilus influenzae type b in children under 5 years of age in Botswana. Int J Infect Dis. 1998;3:18 –25. 22. Bakir M, Yagci A, Ulger N, et al. Pharyngeal colonization with Haemophilus influenzae type b among healthy Turkish infants and children. Pediatr Int. 2002;44:381–386. 23. Naaber P, Tamm E, Putsepp A, Koljalg S, Maimets M. Nasopharyngeal carriage and antibacterial susceptibility of Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis in Estonian children. Clin Microbiol Infect. 2000;6:675– 677. 24. Ward JI, Margolis HS, Lum MK, Fraser DW, Bender TR, Anderson P. Haemophilus influenzae disease in Alaskan Eskimos: characteristics of a population with an unusual incidence of invasive disease. Lancet. 1981;1:1281–1285. 25. Turk DC. Naso-pharyngeal carriage of Haemophilus influenza type b. J Hyg (Lond). 1963;61:247–256. 26. Gessner BD, Sutanto A, Steinhoff M, et al. A population-based survey of Haemophilus influenzae type b nasopharyngeal carriage prevalence in Lombok Island, Indonesia. Pediatr Infect Dis J. 1998;17(suppl):S179 – S182. 27. Mpairwe Y. Observations on the nasopharyngeal carriage of Haemophilus influenzae type b in children in Kampala, Uganda. J Hyg (Lond). 1970;68:337–341. 28. Akcakaya N, Camcioglu Y, Belbek S, Eskazan G, Cokugras H. Haemophilus influenzae type b colonization in children in a hospitalbased day care center. Eur J Epidemiol. 2001;17:313–316. 29. Kilian M, Heine-Jensen J, Bulow P. Haemophilus in the upper respiratory tract of children: a bacteriological, serological and clinical investigation. Acta Pathol Microbiol Scand 关B兴 Microbiol Immunol. 1972;80: 571–578. 30. Scheifele DW, Fussell SJ. Ampicillin-resistant Haemophilus influenzae colonizing ambulatory children. Am J Dis Child. 1981;135:406 – 409. 31. Levine OS, Liu G, Garman RL, Dowell SF, Yu S, Yang YH. Haemophilus influenzae type b and Streptococcus pneumoniae as causes of pneumonia among children in Beijing, China. Emerg Infect Dis. 2000;6:165–170. 32. Lerman SJ, Kucera JC, Brunken JM. Nasopharyngeal carriage of antibiotic-resistant Haemophilus influenzae in healthy children. Pediatrics. 1979;64:287–291.

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33. Howard AJ, Dunkin KT, Millar GW. Nasopharyngeal carriage and antibiotic resistance of Haemophilus influenzae in healthy children. Epidemiol Infect. 1988;100:193–203. 34. Granoff DM, Gilsdorf J, Gessert C, Basden M. Haemophilus influenzae type b disease in a day care center: eradication of carrier state by rifampin. Pediatrics. 1979;63:397– 401. 35. Neto AS, Lavado P, Flores P, et al. Risk factors for the nasopharyngeal carriage of respiratory pathogens by Portuguese children: phenotype and antimicrobial susceptibility of Haemophilus influenzae and Streptococcus pneumoniae. Microb Drug Resist. 2003;9:99 –108. 36. Tangkanakul W, Tharmaphornpilas P. Epidemiology of Haemophilus influenzae type b in Thailand. J Health Sci. 1998;7:571–581. 37. Wenger JD. Epidemiology of Haemophilus influenzae type b disease and impact of Haemophilus influenzae type b conjugate vaccines in the United States and Canada. Pediatr Infect Dis J. 1998;17(suppl):S132– S136. 38. Rerks-Ngarm S, Treleaven SC, Chunsuttiwat S, et al. Prospective population-based incidence of Haemophilus influenzae type b meningitis in Thailand. Vaccine. 2004;22:975–983. 39. Weiser JN. Relationship between colony morphology and the life cycle of Haemophilus influenzae: the contribution of lipopolysaccharide phase variation to pathogenesis. J Infect Dis. 1993;168:672– 680. 40. Barbour ML. Conjugate vaccines and the carriage of Haemophilus influenzae type b. Emerg Infect Dis. 1996;2:176 –182. 41. Singleton R, Bulkow LR, Levine OS, Butler JC, Hennessy TW, Parkinson A. Experience with the prevention of invasive Haemophilus influenzae type b disease by vaccination in Alaska: the impact of persistent oropharyngeal carriage. J Pediatr. 2000;137:313–320. 42. Verez-Bencomo V, Fernandez-Santana V, Hardy E, et al. A synthetic conjugate polysaccharide vaccine against Haemophilus influenzae type b. Science. 2004;305:522–525.

ACANTHOSIS NIGRICANS AS AN ADVERSE EFFECT OF HIGHLY ACTIVE ANTIRETROVIRAL THERAPY IN AN ADOLESCENT GIRL WITH HUMAN IMMUNODEFICIENCY VIRUS INFECTION Antonio Mur, PhD, MD,* Veronica Seidel, MD,* Marıá Ańgeles Lo´pez-Vı´lchez, MD,* Mariona Bonet, MD,* and Montserrat Gilaberte, MD† Abstract: We report an 11-year-old girl with acanthosis nigricans that appeared after 4 years of treatment with didanosine, stavudine and amprenavir. Laboratory studies showed hyperglycemia, hyperinsulinemia and hypertriglyceridemia. Withdrawal of amprenavir resulted in disappearance of acanthosis nigricans and improvement of metabolic abnormalities. Key Words: human immunodeficiency virus infection, antiretroviral therapy, acanthosis nigricans Accepted for publication January 27, 2005. From the Services of *Pediatrics and †Dermatology, Hospital del Mar, Universitat Auto`noma de Barcelona, Barcelona, Spain Reprints not available. DOI: 10.1097/01.inf.0000172936.20623.fd

H

ighly active antiretroviral therapy is associated with numerous adverse effects, of which metabolic abnormalities and endocrine changes are of particular interest in the human immunodeficiency virus (HIV)-infected pediatric population. Some of the metabolic disorders reported for children include the lipodystrophy syndrome,1,2 frequently associated with hyperlipidemia, hyperglycemia and sometimes also insulin resistance3; xanthomas,4 as a cutaneous manifestation of hyperlipidemia; and fatal lactic acidosis5 among children exposed to antiretroviral agents for the prevention of vertical transmission.

742

FIGURE 1. Acanthosis nigricans, with hyperpigmentation and roughening of the right axilla. We report a case of acanthosis nigricans attributed to antiretroviral therapy for an HIV-infected girl. This skin disorder has been documented for adults receiving antiretroviral therapy, but we have found no previously described cases of acanthosis nigricans in a child under the same circumstances.6 CASE REPORT The patient was an 11-year-old girl with HIV infection acquired through vertical transmission. The diagnosis of HIV infection was made at 6 months of age, when the patient had a 3-week febrile episode with liver enlargement and multiple lymph nodes evident in the physical examination. At the time of diagnosis, the Centers for Disease Control and Prevention classification was A1. Initial treatment included zidovudine therapy and monthly administration of intravenous ␥-globulin. At 4 years of age, lamivudine was added to the previous treatment; at 6 years of age, treatment with didanosine, stavudine and amprenavir was started because of failure of previous therapy. Two years later, at 8 years of age, changes in the patient’s body composition were observed, with abdominal adiposity and peripheral fat wasting as signs of lipodystrophy syndrome. The patient’s body weight was in the 75th percentile, height in the 50th–75th percentile and body mass index in the 85th percentile (17.75 kg/m2). After 4 years of treatment with the combination of didanosine, stavudine and amprenavir, at 11 years of age, hyperpigmentation and roughening of the skin in the axillae (Fig. 1), neck and periumbilical region were observed. The physical examination showed fat redistribution and Tanner scale results of S2–S3, P2 and A1. Laboratory studies showed hyperglycemia (140 mg/dL), hyperinsulinemia (113 ␮U/mL) and hypertriglyceridemia (297 mg/dL), with normocholesterolemia (181 mg/dL). Results of thyroid and hypothalamic-pituitary-adrenal axis studies were normal. Metabolic abnormalities were attributed to the protease inhibitor amprenavir, and this drug was switched to efavirenz, a nonnucleoside reverse transcription inhibitor. The metabolic abnormalities slowly improved, and acanthosis nigricans completely disappeared within 6 months. The viral load always remained undetectable. DISCUSSION Acanthosis nigricans is a skin lesion characterized by hyperpigmentation localized in large skinfolds such as the axillae, the © 2005 Lippincott Williams & Wilkins


groins and the neck. It appears predominantly in situations of peripheral insulin resistance, such as endocrine diseases, obesity and some malignant tumors.7 It is used as a skin marker for insulin resistance, although this association has been called into question in recent endocrinologic studies.8 The pathophysiologic mechanism of this skin alteration consists of direct or indirect activation of the insulin-like growth factor-1 receptor of fibroblasts and keratinocytes by elevated concentrations of circulating insulin, with subsequent proliferation of these epidermal cells. The skin alterations consist of hyperkeratosic and hyperpigmented plaques with velvet texture and brown coloration. The metabolic changes induced by antiretroviral therapy can lead to a state of insulin resistance,1,2 which may result in acanthosis nigricans, as shown in the present case.

REFERENCES 1. Leonard EG, McComsey GA. Metabolic complications of antiretroviral therapy in children. Pediatr Infect Dis J. 2003;22:77– 84. 2. Brambilla P, Bricalli D, Sala N, et al. Highly active antiretroviral-treated HIV-infected children show fat distribution changes even in absence of lipodystrophy. AIDS. 2001;18:2415–2422. 3. Amaya RA, Kozinetz CA, McBeans A, Schwarzwald H, Kline M. Lipodystrophy syndrome in human immunodeficiency virus-infected children. Pediatr Infect Dis J. 2002;21:405– 410. 4. Babl FE, Regan AM, Pelton SI. Xanthomas and hyperlipidemia in a human immunodeficiency virus-infected child receiving highly active antiretroviral therapy. Pediatr Infect Dis J. 2002;21:259 –260. 5. Rey C, Prieto S, Medina A, Pe´rez C, Concha A, Meneńdez S. Fatal lactic acidosis during antiretroviral therapy. Pediatr Crit Care Med. 2003;4: 485– 487. 6. Mellor-Pita S, Yebra-Bango M, Alfaro-Martıńez J, Sua´rez E. Acanthosis nigricans: a new manifestation of insulin resistance in patients receiving treatment with protease inhibitors. Clin Infect Dis. 2002;34:716 –717. 7. Torley D, Bellus GA, Munro CS. Genes, growth factors and acanthosis nigricans. Br J Dermatol. 2002;147:1096 –1101. 8. Nguyen TT, Keil MF, Russell DL, et al. Relationship of acanthosis nigricans to hyperinsulinemia and insulin sensitivity in overweight African American and white children. J Pediatr. 2001;138:474 – 480.

ADULT-TYPE PULMONARY TUBERCULOSIS IN CHILDREN 10-14 YEARS OF AGE Ben J. Marais, MRCP (Paed UK), FCP (Paed SA), MMed Paed, Robert P. Gie, FCP (Paed SA), Anneke H. Hesseling, MB. ChB, MSc, and Nulda Beyers, FCP (Paed SA), PhD Abstract: We report 8 children (10 –14 years of age) who were diagnosed with tuberculosis at their local primary health care clinic from July to September 2004, after routine sputum testing was extended to all children older than 10 years of age with suspected tuberculosis. This case series emphasizes that older children develop adult-type cavitating disease, which can be diagnosed by sputum smear microscopy, in contrast to younger children for whom smear microscopy has very little diagnostic value. Key Words: adolescent children, adult-type pulmonary tuberculosis Accepted for publication January 31, 2005. From the Department of Paediatrics and Child Health, Faculty of Health Sciences, Stellenbosch University, Tygerberg, South Africa Address for reprints: Dr B. J. Marais, Department of Paediatrics and Child Health, Faculty of Health Sciences, Stellenbosch University, P.O. Box 19063, Tygerberg, 7505, South Africa. Fax 27-21-8389138; E-mail [email protected]. DOI: 10.1097/01.inf.0000173305.04212.09


Pulmonary Tuberculosis in Children

B

ecause of the rising burden of tuberculosis worldwide, the World Health Organization took the unprecedented step of declaring a global tuberculosis emergency in 1993. Children constitute a significant proportion of the tuberculosis caseload, up to 40% in high burden settings, and experience considerable tuberculosis-related morbidity and mortality.1,2 Childhood tuberculosis is regarded as a paucibacillary disease that is difficult to diagnose and that poses little risk of Mycobacterium tuberculosis transmission. However, children older than 10 years of age develop a different spectrum of disease compared with younger children, with the majority developing adult-type cavitating disease that is sputum smear-positive.3,4 This emphasizes the importance of correct disease classification in childhood tuberculosis, both for prognosis and estimation of the transmission risk.5 We report 8 children (10 –14 years of age) who were diagnosed with tuberculosis at their local primary health care clinic in Cape Town, South Africa during a period of 3 months (July–September 2004); after routine sputum testing was extended to all children older than 10 years of age with suspected tuberculosis. CASE SERIES During the 3-month period, 92 cases of tuberculosis were recorded in total. Children (younger than 15 years of age) constituted 22.8% (21 of 92) of the total disease burden. All sputum smear-positive disease occurred in those 10 years of age or older. In total, 65 (70.6%) cases were sputum smear-positive, of whom 7(10.8%) were children 10 –14 years of age. One child in this age group, with a large pleural effusion and blistering tuberculin skin test, were sputum smear-negative. The male-female ratio was 2:6, and none of these children was infected with human immunodeficiency virus. The majority (6 of 8, 75.0%) reported known contact with a sputum smear-positive source case in the preceding 6 –18 months (Table 1). All of the children were still at school. Table 1 describes the clinical presentation and disease characteristics of the 8 cases. Two children reported erythema nodosum (probably indicative of primary M. tuberculosis infection) in the preceding year. Two children had a tuberculous pleural effusion, of whom 1 had additional parenchymal cavitation and was sputum smear-positive. Both presented with intense, localized pleuritic chest pain, accompanied by fever. Most children without pleural effusion (4 of 6, 66.7%) reported vague and poorly localized chest pain. The children with sputum smear-positive disease had high organism loads as reflected by their sputum smear grading. Complete symptom resolution occurred within 2 months of treatment in 7 (87.5%) cases, using the standard initial four-drug treatment of isoniazid, rifampin, pyrazinamide and ethambutol in a combined fixed dose tablet. Treatment adherence during the intensive phase was excellent (⬎90% of prescribed doses in all cases). Sputum smear conversion was documented in all 7 sputum smear-positive cases after 2 months of treatment. At this time, only 1 child reported incomplete symptom resolution, although she did show significant symptomatic and radiologic improvement. This was attributed to the extent of disease at diagnosis, and her treatment was continued with the standard short course (6 months) treatment regimen. DISCUSSION Despite the methodologic limitations, this case series highlights a few clinical observations with important public health implications. It illustrates that tuberculosis in children 10 –14 years of age frequently presents like adult tuberculosis,6 and that the majority of these children can be diagnosed with routine sputum smear microscopy at the primary health care level. It is well-known that adolescent girls are at higher risk to develop tuberculosis after recent primary infection than are boys.3,4 Erythema nodosum may represent a marker of recent primary M. tuberculosis infection, identifying those with a particularly high risk of progression to tuberculosis in the coming months.7

743

Asymptomatic Pleural effusion No None 12 8

M 14 7

F 13 6

F

No

No

Yes 12 5

F

10 11 3 4

F M

13 2

F

⫺

⫹⫹⫹

Asymptomatic

Marked improvement, 2/12 sm⫺

Adult-type with cavitation and pleural effusion Adult-type with cavitation ⫹⫹⫹

Asymptomatic, 2/12 sm⫺ Adult-type with cavitation ⫹⫹⫹

Asymptomatic, 2/12 sm⫺ Asymptomatic, 2/12 sm⫺ Adult-type with cavitation Adult-type with cavitation

Cough (4 wk), night sweats (3 wk), fatigue (3 wk) Cough (8 wk), chest pain (2 wk), fever (2 wk), night sweats (6 wk), fatigue (6 wk) Cough (5 wk), chest pain (1 wk), fatigue (4 wk), weight loss (4 wk) Cough (8 wk), chest pain (1 wk), fever (1 wk), fatigue (4 wk), weight loss (4 wk) Cough (6 wk), night sweats (2 wk), fatigue (4 wk), weight loss (4 wk) Cough (1 wk), chest pain (2 wk), fever (2 wk), fatigue (2 wk) Yes No

No

⫹⫹ ⫹⫹⫹

Asymptomatic, 2/12 sm⫺ Adult-type with cavitation ⫹⫹⫹

Asymptomatic, 2/12 sm⫺ Adult-type with cavitation ⫹⫹

Cough (⬎12 wk), chest pain (1 wk), night sweats (8 wk), fatigue (8 wk), weight loss ( 8 wk) Cough (4 wk), night sweats (2 wk), fatigue (2 wk), weight loss (4 wk) No

HH: mother (12–18/12) HH: mother and father (6 –12/12) Uncle (6 –12/12) HH: brother (6 –12/12) Grandmother (6 –12/12) HH: uncle (12–18/12) None 14 1

F

Response to First 2/12 of Chemotherapy Radiologic Disease Classification Sputum Smear* Symptoms at Presentation: Cough, Chest Pain, Fever, Night Sweats, Fatigue, Weight Loss 关Uninterrupted Duration (wk)兴 Recent Erythema Nodosum TB Contact HH (Time Since) Sex Age (yr) Case

TABLE 1. Disease Characteristics in Children 10 –14 Years of Age Diagnosed With Tuberculosis

744

*Sputum smear microscopy grading according to the number of AFB observed .⫺ indicates no AFB in at least 100 fields; ⫹⫹, 1–10 AFB/field in at least 50 fields; ⫹⫹⫹, ⬎10 AFB/field in at least 20 fields. TB contact indicates contact with a sputum smear-positive tuberculosis source case; HH, contact inside the household; (time since), time since known contact with a sputum smear-positive source case in months; Recent erythema nodosum, probable primary Mycobacterium tuberculosis infection in the preceding 12 months; AFB, acid-fast bacilli; 2/12 sm⫺, sputum smear-negative after 2 months of treatment.


Matsuzawa et al

It is difficult to establish the exact time of M. tuberculosis infection, but it is striking that 75% of children had known contact with a sputum smear-positive source case in the preceding 6 –18 months. This correlates with the natural history of disease, which indicates that adolescent children are at high risk to develop adulttype tuberculosis within 1–2 years after primary infection.3,7 Therefore all contacts of sputum-smear positive source cases, especially adolescent girls, should be informed about their risk to develop tuberculosis, even if their initial screening tests are negative. Persistent, nonremitting symptoms should be reported without delay, and tuberculosis should be excluded. Delayed diagnosis poses a significant transmission risk to the community, especially fellow pupils8 and household members. ACKNOWLEDGMENTS We thank Sister L. E. Mpambaniso and staff nurse G. N. Summers for their diligence in identifying the patients and assisting with data collection; the patients for their kind assistance; and the local health authorities for permission to report the findings. REFERENCES 1. van Rie A, Beyers N, Gie RP, et al. Childhood tuberculosis in an urban population in South Africa: burden and risk factor. Arch Dis Child. 1999;80:433– 437. 2. Donald PR. Childhood tuberculosis: the hidden epidemic. Int J Tuberc Lung Dis. 2004;8:627– 629. 3. Marais BJ, Gie RP, Schaaf HS, et al. The natural history of childhood intra-thoracic tuberculosis: a critical review of the literature from the pre-chemotherapy era. Int J Tuberc Lung Dis. 2004;8:392– 402. 4. Davies PDB. The natural history of tuberculosis in children. Tubercle. 42(suppl):1– 40, 1961. 5. Marais BJ, Gie RP, Starke JR, et al. A proposed radiological classification of childhood intra-thoracic tuberculosis. Pediatr Radiol. 2004;33:886–894. 6. Weber HC, Beyers N, Gie RP, et al. The clinical and radiological features of tuberculosis in adolescents. Ann Trop Paediatr. 2000;20:5–10. 7. Wallgren A. The time-table of tuberculosis. Tubercle. 1948;29:245–251. 8. Curtis AB, Ridzon R, Vogel R, et al. Extensive transmission of Mycobacterium tuberculosis from a child. N Engl J Med.1999;341:1491–1495.

CONGENITAL CANDIDA GLABRATA INFECTION WITHOUT SPECIFIC NODULES ON THE PLACENTA AND UMBILICAL CORD Sachie Matsuzawa, MD,* Makiko Ohyama, MD,* Motoyoshi Kawataki, MD,* Yasufumi Itani, MD,* Sakae Hashimoto, MD,† Michiko Yamanaka, MD,† Rieko Ijiri, MD,‡ and Yukichi Tanaka, MD‡ Abstract: Two extremely premature infants died as a result of congenital Candida glabrata infection, and their placentas and umbilical cords were free of macroscopic Candida nodules. Because non-Candida albicans Candida infections are less likely to produce necrotic foci, we should not exclude Candida infections in the absence of macroscopic nodules on the placenta and umbilical cord. Key Words: Candida glabrata, Candida nodules, placenta, pneumonia Accepted for publication March 2, 2005. From the Departments of *Neonatology, †Obstetrics and ‡Pathology, Kanagawa Children’s Medical Center, Yokohama City, Japan Address for reprints: Makiko Ohyama, MD, Department of Neonatology, Kanagawa Children’s Medical Center, 2-138-4 Mutsukawa, Minami-ku, Yokohama City, 232-8555, Japan. Fax ⫹81-45-711-2351; E-mail [email protected]. DOI: 10.1097/01.inf.0000173611.59475.30



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eonatal Candida glabrata infection has been reported in infants with large birth weights and have been reported to be less severe than those caused by Candida albicans.1 In congenital Candida infections, the placenta is characterized by yellow-white nodules on the umbilical cord.2 We treated 2 neonates who developed fatal congenital C. glabrata infections and whose placentas and umbilical cords were free of macroscopic Candida nodules. In addition, we retrospectively reviewed the records for 8 infants with congenital Candida infections who were treated in Kanagawa Children’s Medical Center between 1992 and 2002. CASE REPORTS Case 1. A 34-year-old multiparous woman delivered twin infants at 26 weeks of gestation by cesarean section. This pregnancy was established after in vitro fertilization and embryo transfer and was complicated by recurrent episodes of vaginal bleeding. The patient had a history of C. glabrata vaginitis that responded to topical therapy. She was given ritodrine hydrochloride for treatment of preterm labor, as well as 3 courses of intravenous antibiotics for 3 weeks for treatment of a suspected intrauterine infection. The firstborn infant (twin A), who was near the uterine orifice, weighed 915 g, with Apgar scores of 5/9. Although twin A required mechanical ventilation because of pulmonary surfactant deficiency and was treated for congenital heart failure, he never showed any sign of congenital infection and survived. The second infant (twin B), who weighed 808 g, developed severe asphyxia, with Apgar scores of 1/1. Twin B died as a result of ventilatory failure, despite vigorous resuscitation, 13 hours after birth. Cultures taken from the nasopharynx of twin B showed C. glabrata, but a blood culture was sterile. The placenta was diamniotic and dichorionic. The fetal surface of the twin A placenta was dark brown. Microscopically, the chorionic plate for twin A showed stage 3 subacute chorioamnionitis, as well as diffuse chorioamniotic hemosiderosis.3 However, neither the free membrane nor the umbilical cord was inflamed, and Candida hyphae were not identified. The placenta for twin B, which was yellow-white, showed stage 3 acute chorioamnionitis. The umbilical cord was macroscopically discolored, without characteristic nodules. Microscopically, severe subacute funisitis, with diffuse necrosis of the amnion, was revealed. Grocott stain revealed a few foci of microspores suggesting Candida on the amnion of the chorionic plate. No fungus was identified on the umbilical cord. The autopsy demonstrated necrotizing pneumonia with scattered foci of microspores. Other organs did not show any inflammatory reaction. Case 2. A 29-year-old woman was admitted to our hospital because of threatened abortion at 25 weeks of gestation. She had gestational diabetes mellitus but had not received any treatment. She delivered an 808-g female infant by cesarean section. The infant had asphyxia, with Apgar scores of 1/1. The infant died as a result of pneumonia complicated by persistent pulmonary hypertension of the newborn on day 5, despite intensive therapy. Cultures from her pharynx and umbilicus yielded negative results. A culture of lung tissue obtained at autopsy yielded C. glabrata. The fetal surface of the placenta was macroscopically turbid. Microscopically, the chorionic plate showed stage 3 chorioamnionitis. The umbilical cord was yellow-tan, but overt Candida nodules were not recognized. Microscopically necrotizing funisitis was revealed. The amnion was necrotic, and Grocott stain demonstrated scattered microspores suggesting Candida. The autopsy demonstrated necrotizing bronchitis with hemorrhage. Grocott stain of the lesion revealed foci of microspores suggesting Candida species.


Congenital Candida glabrata Infection

Retrospective Review. The placentas were examined in all of the 10 cases of congenital Candida infections. Macroscopically, 5 of the 10 placentas showed overt Candida nodules. The infants with these 5 Candida nodule-positive placentas developed postnatal C. albicans infections. The remaining 5 placentas did not show overt nodules. Causative organisms were C. albicans in 3 cases and C. glabrata in 2 cases (present cases). DISCUSSION Candida infections of the placenta are characterized by yellow-white nodules, mostly on the umbilical cord.2 Candida nodules are commonly seen in C. albicans infections but are rarely found in non-C. albicans infection, including C. glabrata infection.2 In a report of 32 placentas with Candida funisitis, as many as 13 placentas (41%) did not show Candida nodules on the umbilical cords.4 That report did not provide speciation of the Candida. The present report suggests that the absence of nodules is more frequent in non-C. albicans species infections than in C. albicans infections. Candida nodules are not necessarily seen even in C. albicans infections. C. glabrata is known to grow without forming pseudohyphae, which are thought to be responsible for tissue invasion.5 In our series, we found scattered microspores on the necrotic surface of the Wharton’s jelly, but they did not form pseudohyphae or microabscesses. It is possible that this finding could account for the absence of nodule formation by C. glabrata. Mortality rates of C. glabrata fungemia in adults have been reported to be higher than those of C. albicans infections in patients with advanced age or underlying malignancy.6 In neonates, nosocomial C. glabrata infections showed variable outcomes.1,7 However, a few reported cases of congenital C. glabrata infections resulted in neonatal death; all were accompanied by fungemia.8,9 We think that mortality rates for C. glabrata infections are variable and death attributable to fungal pneumonia is possible irrespective of fungemia. Macroscopically apparent Candida nodules are quite helpful for treating acutely ill infants. It is important, however, that the absence of such nodules should not lead clinicians to exclude Candida. REFERENCES 1. Fairchild KD, Tomkoria S, Sharp EC, et al. Neonatal Candida glabrata sepsis: clinical and laboratory features compared with other Candida species. Pediatr Infect Dis J. 2002;21:39 – 43. 2. Benirschke K, Kaufmann P. Infectious disease. In: Benirschke K, Kaufmann P, eds. Pathology of the Human Placenta. 4th Ed. New York, NY: Springer-Verlag; 2000:591– 684. 3. Ohyama M, Itani Y, Yamanaka M, et al. Maternal, neonatal, and placental features associated with diffuse chorioamniotic hemosiderosis, with special reference to neonatal morbidity and mortality. Pediatrics. 2004;113:800–805. 4. Qureshi F, Jacques SM, Benton RW, et al. Candida funisitis: a clinico-pathologic study of 32 cases. Pediatr Dev Pathol. 1998;1: 118 –124. 5. Fidel PL, Vazquez JA, Sobel JD. Candida glabrata: review of epidemiology, pathogenesis, and clinical disease with comparison to C. albicans. Clin Microbiol Rev. 1999;12:80 –96. 6. Safdar A, Bannister TW, Safdar Z. The predictors of outcome in immunocompetent patients with hematogenous candidiasis. Int J Infect Dis. 2004;8:180 –186. 7. Reich JD, Huddleston K, Jorgensen D, et al. Neonatal Torulopsis glabrata fungemia. South Med J. 1997;90:246 –248. 8. Quirke P, Hwang W, Validen CC. Congenital Torulopsis glabrata infection in man. Am J Clin Pathol. 1980;73:144 –152. 9. Sander CH, Martin JN, Rogers AL, et al. Perinatal infection with Torulopsis glabrata: a case associated with maternal sickle cell anemia. Obstet Gynecol. 1983;61(suppl):21S–24S.

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