1 MINISTRY OF EDUCATION AND TRAINING MINISTRY OF HEALTH THE NATIONAL INSTITUTE OF HYGIENE AND EPIDEMIOLOGY PHAM THU HIEN THE STUDY OF EPIDEMIOLOGICAL CHARACTERISTICS, CLINICAL MANIFESTATIONS OF ATYPICAL PNEUMONIA CAUSED BY BACTERIA IN CHILDREN Science: Epidemiology Code: 62 72 01 17 SUMMARY OF THE DOCTORAL DISSERTATION HA NOI - 2014 The project was completed at the National Institute of Hygiene and Epidemiology The scientific advisors: Prof Dao Minh Tuan Prof Phan Le Thanh Huong Reviewer 1: Reviewer 2: Reviewer 3: The dissertation will be defended at the meeting hall of the National Institute of Hygiene and Epidemiology In… hours, …/… / 20… The dissertation is available at: The National Library The National Institute of Hygiene and Epidemiology LIST OF THE PUBLICATIONS BY THE AUTHORS RELATED TO THE DISSERTATION Pham Thu Hien, Dao Minh Tuan, Nguyen Phong Lan, Phan Le Thanh Huong (2011), "The role of Mycoplasma pneumoniae, Chlamydia pneumoniae, and Legionella pneumophyla in community -acquired pneumonia in children: preliminary results", Journal Journal preventive Medicine, Vol XXI, No ( 125 ), pp 62-69 Pham Thu Hien, Dao Minh Tuan, Nguyen Phong Lan, Phan Le Thanh Huong( 2012 ) , " Causes, clinical features , clinical manifestations of atypical pneumonia in children " , Journal of Medical Research , episode 80 , No A , pp 119124 Pham Thu Hien, Dao Minh Tuan, Nguyen Phong Lan, Phan Le Thanh Huong (2012), "Frequency , clinical characteristics , subclinical pneumonia caused by Mycoplasma pneumoniae, Chlamydia pneumoniae in children ", Journal Journal preventive Medicine, Vol XXII, No (133), tr 31 - 38 LIST OF ABBREVIATIONS ALT Alanine Aminotransferase AST BCYE Aspartate Aminotransferase buffered charcoal yeast extract C pneumoniae Chlamydia pneumoniae Chlamydia pneumoniae CRP Cs C protein reactive Protein C phản ứng Cộng ELISA IgG enzyme-linked immunosorbent assay Immunoglubulin G Kỹ thuật miễn dịch gắn men Immunoglubulin G IgM Immunoglubulin M Immunoglubulin M IL Interleukin Interleukin INF Tumor necrosis factor Yếu tố hoại tử u L pneumophila Legionella pneumophila Legionella pneumophila M pneumoniae Mycoplasma pneumoniae Mycoplasma pneumoniae PCR PPLO polymerase chain reaction Pleuropneumonia like organisms Phản ứng PCR Pleuropneumonia like organisms Real – time PCR S pneumoniae Real –time polymerase chain reaction Streptoccocus pneumoniae TNF Tumor necrosis factor Phản ứng Real – time PCR Streptoccocus pneumoniae Yếu tố hoại tử u VPĐH Viêm phổi điển hình VPKĐH (AP) Atypical pneumonia Viêm phổi khơng điển hình WHO World Health Organization Tổ chức y tế giới ABSTRACT OF THE DISSERTATION 1.Background Atypical pneumonia is frequent in developing countries However, the studies about these conditions in developing countries, including Vietnam are limited Forest (2007) reported that the incidence of atypical pneumonia in the community-acquired pneumonia was 22 % in the United States and 91% of those had been treated In Europe, the incidence of atypical pneumonia was 28%, the rate of treatment was 74% In Latin America, the incidence of atypical pneumonia was 21% and the rate of treatment was 57% In Asia / Africa, the incidence was 20%, the rate of treatment was 10% The diagnostic methods for atypical pneumonia include: bacterial culture in the special media, serology, and polymerase chain reaction method Polymerase chain reaction method (PCR) has helped confirmed and rapid diagnosis bacterial pathogens In Vietnam, PCR techniques are available in only few hospitals in central and major medical centers Most treatments have been done with empirical therapy that may result in increasing in antibiotic resistant, and prolonged treatment time Little is known about the epidemiology, clinical and laboratorial manifestations of atypical pneumonia in children, including the diagnostic techniques for bacterial causes of atypical pneumonia such as multiple primers PCR (multiplex-PCR) and enzyme-linked immunosorbent assay (ELISA), we conducted the study: "The study of epidemiological characteristics, clinical manifestations of atypical pneumonia caused by bacteria in children” The study objectives were: To describe the epidemiological characteristics, clinical manifestations of atypical pneumonia caused by Mycoplasma pneumoniae, Chlamydia pneumoniae, and Legionella pneumophila in children who were treated at the National Hospital of Pediatrics in Hanoi, Vietnam from 07/2010 to 3/2012 To identify the factors associated with the severity of atypical pneumonia in children New contribution to the science - This project was the first study to identify the prevalence of atypical pneumonia caused by M pneumoniae, C pneumoniae and L pneumophila, co-infection rate of pneumonia in hospitalized children - The study has identified several factors associated with severity of atypical pneumonia Co-infection with bacteria and viruses was the related factors for severe atypical pneumonia - This study described the clinical manifestations and laboratorial characteristics of atypical pneumonia, atypical pneumonia co-infection in children Practical value of the subject - Evaluate the results of clinical manifestations and laboratory characteristics of atypical pneumonia to draw specific symptoms which suggesting early clinical diagnosis, help clinicians quickly optimal decision the choice of antibiotic therapy and have a more comprehensive view of the causes of pneumonia in children - The study's results are significant in establishing the pattern of microorganisms which cause respiratory infections in children, and to guide treatment and prevention strategies - Microbiological diagnostic techniques based on molecular biology (only in a few specialized laboratories) will be confirmed and efficient which can be replicated in the laboratory of clinical microbiology The structure of the dissertation The dissertation consists of 128 pages including: Background and objectives: pages; Literature review: 34 pages; Methods: 18 pages; Results: 34 pages; Discussion: 37 pages; Conclusion: pages, and recommendation: page There are 29 tables and 20 figures, 228 references including 22 in Vietnamese, 206 documents in foreign languages Chapter OVERVIEW 1.1 Introduction Atypical pneumonia: pneumonia caused by M pneumoniae has been recognized from the dairy failed to sulfonamides or penicillin used to treat pneumonia, help to distinguish pneumonia pathogens M pneumoniae pneumonia or pneumococcal (pneumococci) The failure to respond to antimicrobial therapy has been thought as "atypical" (atypical) This term, along with "Walking around pneumonia" is used broadly to refer to respiratory disease caused by M pneumoniae in humans Then other agents cause similar clinical picture was included in the group of atypical pneumonia as C pneumoniae, L pneumophila 1.2 Epidemiological characteristics of atypical pneumonia caused by Mycoplasma pneumoniae, and Legionella pneumophila Chlamydia pneumoniae 1.2.1 Epidemiology characteristics of pneumonia caused by Mycoplasma pneumoniae Disease appears in all countries, however studies about the disease mostly have been carried out in the United States, Europe and Japan In the U.S, infection caused by M pneumoniae accounts for 15-20% of all community-acquired pneumonia Especially in the summer time, M Pneumonia can reach up to 50% of all community-acquired pneumonia During 2010 to 2012, an outbreak of M pneumoniae infection happened in some Asian countries A multicenter study in Asia in 2005 found that M pneumoniae was an important cause of the community acquired pneumonia, which accounted for 22.3% Disease occurs in people of all ages, predominant in the age group from 5-9 years old Disease can occur throughout the year, and the peak during the period from late summer to early fall The bacteria can live everywhere in nature, transmitted from person to person via the respiratory tract The average Incubation period is weeks After suffering from this disease, an immune survived about years Immunology temporary and recurence 1.3.2 Epidemiology characteristics of Chlamydia pneumoniae pneumonia C pneumoniae infection distributes over the world A study from 10 different regions of the world showed a higher frequency in tropical populations In the U.S and many other countries, the seroprevalence of C pneumoniae infection was of 50% of total population 10 Estimated number of cases of pneumonia caused by C pneumoniae in the United States is 300,000 cases per year Globally, prevalence of pneumonia caused by C pneumoniae from 4337 patients was 8% in North America, 7% in Europe, 6% in Latin America and 5% in Asia Forest (2007) The disease affects both genders and all age groups Disease occurs throughout the year, and gets its peak during summer time Infectious reservoir is humans, and it is transmitted through respiratory secretions directly exposed to coughing, sneezing After suffering from this disease, patients have an immune temporarily and recurrent frequently Disease cycle every - years 1.3.3 Epidemiological characteristics of Legionella pneumophila pneumonia Legionella disease occurs worldwide The majority of cases disease been identified in tropical countries In the U.S., about 800018000 hospitalized cases every year In Europe, the prevalence of Legionella infection were 5,907 cases in 2007 and 5,960 cases in 2008 Most patients exposed to L pneumophila but no symptoms The risk increase in an older people Children after ages rare occurs pneumonia due to L pneumophila Legionella live everywhere, special in the aquatic environment, the disease transmitte through tiny droplets of water vapor Disease is not transmitted from person to person Incubation period -10 days Reinfection occurs in immunocompromised people 1.4 The clinical features, laboratory manifestations and treatment of pneumonia caused by M pneumoniae, C pneumoniae and L pneumophila 1.4.1 The clinical features of pneumonia caused by M pneumoniae, C pneumoniae, L pneumophila M pneumoniae, C pneumoniae is causative agents of pneumonia with the various degree of severity Majority of patients appear with mild illness and self-recover Few patients progress to severe condition, acute respiratory failure and death The common pulmonary manifestations are: eardrum inflammation, rash, urticaria, pleurisy, thrombocytopenia, meningitis, and mild anemia 17  Sample sizes for objective The purpose of the study is to describe a case series and combine with the analytic study to identify factors associated with severe atypical pneumonia cases, we used all cases diagnosed with atypical pneumonia (215 patients) that occurred during the study period (7/2010 - 3/2012), among them, 97 cases were clasified as severe atypical pneumonia 2.3.2 Sampling method:  Source of patients: Children from 12 months to 15 years, with an initial diagnosis of pneumonia, treatment in National Hospital of Pediatrics from the local different provinces  Sampling method fore objective 1: A convenient sampling technique, as a rule, chose one patient from patients ( k=3),according to data pneumonia in hospitalized patients in the preceding year divided by the total study minimum sample size was calculated) apply to case series research, prospective, longitudinal follow-up  Sampling method fore objective 2: a convenient sampling technique, take the whole 215 atypical pneumonia patients were selected by criteria subjects for the study objective From 215 cases with atypical pneumonia, we selected 97 cases with severe atypical pneuonia and 118 non- severe atypical pneumonia cases We conducted a comparative analysis to determine the factors associated with severe atypical pneumonia among patients who were treated at the National Hospital of Pediatrics The data collection was performed during the study, but the final analysis was carried out only at the end of the study when all clinical, liboratorial and microbiology data was collected for all patients 2.4 Study variables 2.4.1 Study variable for objective  Epidermiological data collection Demographic data: name, age, sex, location, education, information family Epidemiological factors: geography, season, habitat, level of income History: obstetrics, development, immunization, disease history  Clinical data collection: by interviewing parents and/or by physical examination 18  Laboratory data collection The laboratory tests included blood specimens for counting leukocyte (WBC), C-reactive protein (CRP), IL6 and for the detection of IgM, IgA, IgG, IgE antibody and IgM antibody against M pneumoniae, C pneumoniae and L pneumophila.Throat swabs/bronchial exudates were used for detection of M pneumoniae, C pneumoniae and L pneumophila specific DNA by multiplex PCR In addition, RT-PCR was applied to determine the presence of co – infections involving other viral respiratory pathogens such as Adenovirus, Respiratory Syncytial Virus (RSV), Rhinovirus, Influenza A & B(RNA extraction using Qiamp Viral RNA Mini kit, RT-PCR using Kit SuperSckip III One- Step Kit [Invitrogen] 2.4.2 Study variable for objective Factors associated with severe atypical pneumonia: time from onset to admission, antibiotic use before admission, signs, clinical symptoms, WBC, CPR, IL6, IgA, IgM, IgG, IGE antibody, coinfected status with severity of disease 2.5 Data analysis Statistical analysis was performed using Stata.10, SPSS.13, Epidata 3.1 Chapter RESULTS 3.1 Epidemiological characteristics of Atypical bacterial pneumonia in children 3.1.1 General epidemiological characteristics of atypical pneumonia Table 3.1 The rate of common pneumonia Type of pneumonia The number Rate% of Typical pneumonia caused by bacteria Viral pneumonia Pneumonia caused by typical bacteria coinfection with virus Atypical bacterial pneumonia Pneumonia with unknown etiology Total 82 80 14 11.35 11.08 1.93 215 331 722 29.8 45.84 100 Table 3.1 shows the overall incidence of atypical pneumonia was 29.8% of the total pneumonia 19 Table 3.2 Classification of atypical pneumonia Classification of atypical pneumonia Atypical simple pneumonia: or M pneumoniae, or C pneumoniae, or L pneumophila Atypical Atypical pneumonia coinfection: by pneumonia M pneumoniae and C pneumoniae or M pneumoniae and L pneumophila or C pneumoniae and L pneumophila Atypical pneumonia + typical bacterial The number of 144 38 19 pneumoni a Outside group Total 67 10 Coinfected pneumonia atypical Atypical pneumonia + viral pneumonia Atypicalpneumonia + typical Rate % pneumonia + viral pneumonia 215 17.67 8.84 1.86 100 Table 3.2 shows the co-infection rate was 33%, which co-infected with typical pneumonia and viral pneumonia accounted for 28.37% Table 3.3 classify of pneumonia cases by agent bacterial atypical pneumonia (data not shown here ): it found that M pneumoniae was the most predominant among community acquired pneumonia in 26.3%; C pneumoniae and L.pneumophila detected with low rate (3.7%, 1.8%) 20 Chart 3.1 Age distribution of typical pneumonia and atypical pneumonia Chart 3.1 shows that the proportion of children aged greater than years old with atypical pneumonia was 23.3% But among 215 patients with atypical pneumonia The incidence of children aged higher in the group under years old The age variables were significantly difference (p < 0.001) There was no difference in gender distribution among children with atypical pneumonia (data not shown) Spring Summer Auturm Winter Chart 3.3 Seasonal distribution of atypical pneumonia Atypical pneumonia occurs throughout all seasons, more frequent during spring-summer seasons than Auturm - Winter season There was statistically significant difference with p = 0.003 (χ test) 21 Table 3.11 Factors associated with co-infection Characteristics Age Under years >2 years - years old > years - 10 years > 10 years old Adjusted OR 0.79 0.46 0.43 95% CI 0.39 0.19 0.08 1.58 1.12 2.48 p 0.50 0.09 0.35 Gender Female 1.81 0.94 3.48 0.07 Male Family economic conditions Non-poor households 1.37 0.54 3.47 0.51 Poor households Method of birth Vaginal delivery 2.12 1.05 4.30 0.037 Caesareans Asthma Yes 0.73 0.29 1.84 0.50 No Nutritional status Normal 1.82 0.77 4.29 0.17 Wasting 0.83 0.38 1.81 0.63 Overweight and obesity Test fit the Hosmer-Lemeshow test pattern n = 215, p = 0.8619 Table 3:11 shows the relationship between co – infected status and each of variables, including age, gender, family economic conditions, method of birth, asthma disease, nutritional status Except for Caesarean section, the other variables were not significantly associated with co - infection (OR = 2.12, p = 0.037) 22 3.1 Table The functional symptoms of hospitalized atypical pneumonia patients (data not presented here): signs dyspnea in coinfected atypical pneumonia outside group higher statistical significance compared with atypical pneumonia in group (p