Coronaviruses in children, Greece.To the Editor: Two recently detected human coronaviruses (HCoVs), NL63 and HKU HKU University of Hong Kong HKU Hogeschool voor de Kunsten Utrecht (Utrecht School of The Arts, The Netherlands) HKU Hot Key Users 1, increased the number of coronaviruses known to infect humans to 5 (1-3). HCoV-229E and HCoV-NL63 belong to antigenic group 1, HCoV-OC43 and HCoVHKU1 belong to antigenic group 2, and severe acute respiratory syndrome Severe Acute Respiratory Syndrome (SARS) Definition Severe acute respiratory syndrome (SARS) is the first emergent and highly transmissible viral disease to appear during the twenty-first century. (SARS)-associated coronavirus coronavirus /co·ro·na·vi·rus/ (ko-ro´nah-vi?rus) any virus belonging to the family Coronaviridae. Coronavirus /Co·ro·na·vi·rus/ (ko-ro´nah-vi?rus (SARS-CoV) is most closely related to group 2 coronaviruses. In 2005, an optimized pancoronavirus reverse transcription-PCR assay was used to explore the incidence of HCoV-NL63 infection in children in Belgium who had a diagnosis of respiratory tract infection Noun 1. respiratory tract infection - any infection of the respiratory tract respiratory infection infection - the pathological state resulting from the invasion of the body by pathogenic microorganisms (4). We report the results of an epidemiologic study epidemiologic study A study that compares 2 groups of people who are alike except for one factor, such as exposure to a chemical or the presence of a health effect; the investigators try to determine if any factor is associated with the health effect that used a universal coronavirus RT-PCR RT-PCR reverse transcriptase-polymerase chain reaction. See PCR1. assay to detect coronaviruses among children in Greece with acute respiratory tract infections. We tested throat swab specimens obtained from children hospitalized in Greece during June 2003 through May 2004 (200 children 2 months to 14 years of age, mean 4.09 years) and during December 2005 through March 2006 (44 children 1.6-8.5 years of age, mean 5.05). Specimens were obtained the first day of each child's hospitalization, and all specimens were included in the study, regardless whether other respiratory microorganisms were detected. The 25-[micro]L reaction contained 200 [micro]M dNTPs, 0.2 [micro]M primer PC2S2 (equimolar e·qui·mo·lar adj. Chemistry Having an equal number of moles. mixture of 5'TTATGGGTTGGGATTATC-3' and 5'-TGATGGGATGGGACTATC-3'), 0.8 [micro]M primer PC2As1 (5'-TCATCAGAAAGAATCATCA-3'), 1 [micro]L of enzyme mix from the QIAGEN OneStep RT-PCR Kit (QIAGEN GmbH, Hilden, Germany), and 5 [micro]L of RNA RNA: see nucleic acid. RNA in full ribonucleic acid One of the two main types of nucleic acid (the other being DNA), which functions in cellular protein synthesis in all living cells and replaces DNA as the carrier of genetic . The initial 30-min reverse transcription reverse transcription n. The process by which DNA is synthesized from an RNA template. step at 48[degrees]C was followed by 10 cycles of 20 sec at 94[degrees]C, 30 sec at 62[degrees]C with a decrease of 1[degrees]C per cycle, 40 sec at 72[degrees]C; 40 cycles of 20 sec at 94[degrees]C, 30 sec at 52[degrees]C, 40 sec at 72[degrees]C; and a final extension step at 72[degrees]C for 10 min. To determine the sensitivity after optimization, we tested quantified RNA in vitro in vitro /in vi·tro/ (in ve´tro) [L.] within a glass; observable in a test tube; in an artificial environment. in vi·tro adj. In an artificial environment outside a living organism. transcripts that included the natural primer binding sites of the respective coronavirus genomes. Sensitivities for SARS-CoV, HCoVOC43, HCoV-229E, and HCoV-NL63 were 61.0, 800.0, 8.2, and 82.3 nominal RNA copies per assay, respectively. A separate test was not done for HCoV-HKU1 because it had the same primer binding sites as HCoV-OC43. A phylogenetic tree phylogenetic tree Diagram showing the evolutionary interrelations of a group of organisms that usually originated from a shared ancestral form. The ancestor is in the tree trunk; organisms that have arisen from it are placed at the ends of tree branches. based on a 400-bp genome fragment of the polymerase gene was constructed (online Appendix Figure, available from www.cdc. gov/EID/13/6/947-appG.htm). Of 200 samples collected in 2003-2004, 5 (2.5%) were positive for coronaviruses (2 each for HCoV-NL63 and HCoV-229E and 1 for HCoVOC43), and of 44 samples collected in 2005-2006, 2 (4.5%) were positive for coronaviruses (1 for HCoV-229E and 1 for HCoV-OC43) (GenBank accession nos. EF103180-EF103184, EF394298, and EF394299). CoVHKU1 was not detected. The amplified genome region is one of the most conserved regions of the coronavirus genome. However, sequences for HCoV-NL63 strains isolated in Greece are genetically closer to the sequence for a strain (AY567487) isolated in Amsterdam in 2003 (1) than to a strain (AY518894) from a specimen collected in Rotterdam in 1988 (2) (0.6% vs. 1.1% nucleotide divergence). Sequences for HCoV-229E and HCoV-OC43 strains isolated in Greece differ from sequences for strains isolated elsewhere by 0.5%-1.7%. The HCoV-NL63--positive specimens in our study were obtained from a 9- and a 14-month-old child during winter 2003-2004; no cases were identified during 2005-2006. Specimens positive for HCoV-229E and HCoV-OC43 were detected during both study periods (Table). HCoV-OC43 affected children with a mean age of 3.1 years (median, 1.4 years), and HCoV-229E affected children with a mean (and median) age of 5.5 years. However, no general conclusions can be drawn from these data because number of cases is too few. None of the patients in Greece had an underlying disease, and all recovered completely. Patients infected with HCoV-229E had been hospitalized for upper respiratory tract infections upper respiratory tract infection URI Infectious disease A nonspecific term used to describe acute infections involving the nose, paranasal sinuses, pharynx, and larynx, the prototypic URI is the common cold; flu/influenza is a systemic illness involving the URT , and those with HCoV-OC43 had lower respiratory tract infections; all cases were mild. Both children infected with HCoV-NL63 had symptoms of lower respiratory tract infections: 1 child had severe pneumonia and was hospitalized for 12 days, while the other had a mild course of bronchiolitis Bronchiolitis Definition Bronchiolitis is an acute viral infection of the small air passages of the lungs called the bronchioles. Description Bronchiolitis is extremely common. . HCoV-NL63 was first identified in Amsterdam, the Netherlands, by van der Hoek et al. (1) from a nasopharyngeal nasopharyngeal pertaining to the nasal and pharyngeal cavities. nasopharyngeal meatus see nasopharyngeal meatus. nasopharyngeal spasm see reverse sneeze. specimen obtained in 2003 from a 7-month-old child with bronchiolitis, conjunctivitis conjunctivitis (kənjəngtəvī`təs), inflammation or infection of the mucosal membrane that covers the eyeball and lines the eyelid, usually acute, caused by a virus or, less often, by a bacillus, an allergic reaction, or an , and fever. One month later, Fouchier et al. (2) reported the characterization of the same virus isolated from a specimen collected in 1988. The specimen had been obtained from an 8-month-old child with pneumonia in Rotterdam, the Netherlands. Later, HCoV-NL63 was detected in 2.5% of bronchiolitis patients <2 years of age in Japan (5) and in most children hospitalized with bronchiolitis in Australia and Canada (6,7). Coinfection with HCoVs and other respiratory viruses is frequently observed and is associated with severe clinical syndromes, especially in infants and young children (6,8). Coinfection was observed in 3 of the 7 HCoV-positive patients in our study. The 3 patients were infected with HCoV-OC43 or HCoV-229E; coinfection with respiratory syncytial virus respiratory syncytial virus (sĭnsĭsh`əl): see cold, common. was found in 2 patients, and coinfection with Mycoplasma pneumoniae Mycoplasma pneu·mo·ni·ae n. A microorganism causing primary atypical pneumonia in humans. was found in 1 patient. It was not possible to determine the role of the HCoVs in these coinfections. In addition, because coronaviruses can be detected even 3 weeks after an acute episode, some cases of coinfection might represent former rather than current HCoV infection (9). In conclusion, we detected 3 types of HCoVs in Greece: 229E, OC43, and NL63. This finding provides initial insight into the epidemiologic features of coronaviruses in Greece. Further studies are needed to find the exact clinical effect of these HCoVs in humans and to elucidate the epidemiology of coronaviruses worldwide. This work was supported by the Greek State Scholarships Foundation (IKY IKY I Know You IKY I Kiss You ) and the Deutscher Akademischer Austauschdienst (DAAD DAAD Deutscher Akademischer Austauschdienst (German Academic Exchange Service) ) (program IKYDA 2004). References (1.) van der Hoek L, Pyrc K, Jebbink MF, Vermeulen-Oost W, Berkhout RJ, Wolthers KC, et al. Identification of a human coronavirus. Nat Med. 2004; 10:368-71. (2.) Fouchier RA, Hartwig NG, Bestebroer TM, Niemeyer B, de Jong De Jong is the most common Dutch surname. Many people bear this name, including many important historical figures. Some of these people are mentioned below. De Jong may mean:
respiratory disorder, respiratory illness adult respiratory distress syndrome, ARDS, wet lung, white lung - acute lung injury characterized by coughing and rales; inflammation of the in humans. Proc Natl Acad Sci U S A. 2004;101:6212-6. (3.) Woo PC, Lau SK, Chu CM, Chan KH, Tsoi HW, Huang Y, et al. Characterization and complete genome sequence of a novel coronavirus, coronavirus HKUI, from patients with pneumonia. J Virol. 2005;79:884-95. (4.) Moes E, Vijgen L, Keyaerts E, Zlateva K, Li S. A novel pancoronavirus RT-PCR assay: frequent detection of human coronavirus NL63 in children hospitalized with respiratory tract infections in Belgium. BMC (BMC Software, Inc., Houston, TX, www.bmc.com) A leading supplier of software that supports and improves the availability, performance, and recovery of applications in complex computing environments. Infect Dis. 2005;5:6. (5.) Ebihara T, Endo R, Ma X, Ishiguro N, Kikuta H. Detection of human coronavirus NL63 in young children with bronchiolitis. J Med Virol. 2005;75:463-5. (6.) Arden KE, Nissen MD, Sloots TP, Mackay IM. New human coronavirus, HCoV-NL63, associated with severe lower respiratory tract Noun 1. lower respiratory tract - the bronchi and lungs lung - either of two saclike respiratory organs in the chest of vertebrates; serves to remove carbon dioxide and provide oxygen to the blood disease in Australia. J Med Virol. 2005;75:455-62. (7.) Bastien N, Robinson JL, Tse A, Lee BE, Hart L, Li Y. Human coronavirus NL-63 infections in children: a l-year study. J Clin Microbiol. 2005;43:4567-73. (8.) Gerna G, Campanini G, Rovida F, Percivalle E, Sarasini A, Marchi A, et al. Genetic variability Introduction Genetic Variability
(9.) Kaiser L, Regamey L, Roiha H, Deffernez C, Frey U. Human coronavirus NL63 associated with lower respiratory tract symptoms in early life. Pediatr infect Dis J. 2005;24:1015-7. Anna Papa, * Evangelia Papadimitriou, * Luciano Kleber de Souza De Souza or D'Souza is a common Portuguese family name. Although it is still quite common outside Portugal -- especially in Brazil and India --, Souza is the old spelling of present-day Sousa. Luna, ([dagger]) Motassim Al Masri, * Efimia Souliou, * Maria Eboriadou, ([double dagger]) Antonis Antoniadis, * and Christian Drosten ([dagger]) * Aristotle University of Thessaloniki The Aristotle University of Thessaloniki (often referred to in English as Aristotelian University), named after the philosopher Aristotle, is the largest university of Greece. Its campus covers 429 metric acres close to the center of the city of Thessaloniki. , Thessaloniki, Greece; ([dagger]) Berhard Nocht Institute for Tropical Medicine tropical medicine, study, diagnosis, treatment, and prevention of certain diseases prevalent in the tropics. The warmth and humidity of the tropics and the often unsanitary conditions under which so many people in those areas live contribute to the development and , Hamburg, Germany; and ([double dagger]) University General AHEPA AHEPA American Hellenic Educational Progressive Association Hospital, Thessaloniki, Greece Address for correspondence: Anna Papa, Department of Microbiology, School of Medicine, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece; email: annap@ med.auth.gr
Table. Epidemiologic and laboratory data for patients with
coronavirus infection, Greece *
Specimen
no., HCoV Sample
strain Age, sex date Symptoms
10/03, 229E 3 y, F Jun 3, Fever
2003 (39
[degrees]C),
cough,
pharyngitis
16/03, 229E 8 y, M Jun 14, Fever
2003 (41
[degrees]C),
headache,
rhinitis,
sinusitis
109/03, NL63 14 mo, F Nov 30, Fever
2003 (39
[degrees]C),
cough, severe
pneumonia
173/04, NL63 10 mo, M Feb 10, Fever
2004 (38.5
[degrees]C),
cough,
rhinitis,
tachypnea,
bronchiolitis
185/04, OC43 17 mo, F Feb 25, Pharyngitis,
2004 rhinitis,
respiratory
distress,
bronchiolitis
12A/O6, OC43 6 mo, F Jan 11, Fever
2006 (38.8
[degrees]C),
cough,
tachypnea,
bronchiolitis
14A/06, 229E 7.5 y, M Feb 13, Fever
2006 (40.5
[degrees]C),
couqh,
rhinitis
Specimen WBC
no., HCoV (cells/ ESR
strain [mm.sup.3]) Granulocytes, % (mm/h)
10/03, 229E 10,400 87 40
16/03, 229E 18,900 86.4 30
109/03, NL63 18,700 44.0 85
173/04, NL63 7,100 57.9 55
185/04, OC43 10,100 63.2 30
12A/O6, OC43 19,950 80.3 35
14A/06, 229E 20,600 83.1 98
Specimen
no., HCoV Days in
strain hospital Coinfection
10/03, 229E 3 RSV
16/03, 229E 4 ND
109/03, NL63 12 ND
173/04, NL63 3 ND
185/04, OC43 2 ND
12A/O6, OC43 6 RSV
14A/06, 229E 4 Mycoplasma
pneumoniae
* HCoV, human coronavirus; WBC, white blood cell count; ESR,
erythrocyte sedimentation rate; RSV, respiratory syncytial virus;
ND, not detected.
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