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
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SUMMARY OF THE DOCTORAL DISSERTATION
HA NOI - 2014
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The project was completed at the
National Institute of Hygiene and Epidemiology
The scientific advisors:
1. Prof. Dao Minh Tuan
2. 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….
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The dissertation is available at:
1. The National Library
2. The National Institute of Hygiene and Epidemiology
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men
IgG Immunoglubulin G 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 polymerase chain reaction Phản ứng PCR
PPLO Pleuropneumonia like
organisms
Pleuropneumonia like
organisms
Real – time PCR Real –time polymerase
chain reaction
Phản ứng Real – time
PCR
S. pneumoniae Streptoccocus pneumoniae Streptoccocus
pneumoniae
TNF Tumor necrosis factor Yếu tố hoại tử u
VPĐH Viêm phổi điển hình
VPKĐH (AP) Atypical pneumonia Viêm phổi không điển
hình
WHO World Health Organization Tổ chức y tế thế giới
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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

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.
3. 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.
4. The structure of the dissertation
The dissertation consists of 128 pages including: Background and
objectives: 2 pages; Literature review: 34 pages; Methods: 18 pages;
Results: 34 pages; Discussion: 37 pages; Conclusion: 2 pages, and
recommendation: 1 page. There are 29 tables and 20 figures, 228
references including 22 in Vietnamese, 206 documents in foreign
languages.
Chapter 1. OVERVIEW
1.1. Introduction
Atypical pneumonia: pneumonia caused by M. pneumoniae has

is 3 weeks. After suffering from this disease, an immune survived about
4 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 sero-
prevalence of C. pneumoniae infection was of 50% of total population.
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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 4 - 8 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 8000-
18000 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 4 rare occurs
pneumonia due to L. pneumophila.
Legionella live everywhere, special in the aquatic environment, the

phosphate).
Extrapulmonary manifestations of Legionella can present with the
damage in spleen, liver, kidney, heart, bone and bone marrow, joints,
inguinal lymph nodes, nervous and digestive tract.
1.4.2. Laboratory manifestations of pneumonia caused by M.
pneumoniae, C. pneumoniae and L. pneumophila
Chest X-ray (CXR):
Radiographic manifestations of atypical pneumonia can be
extremely variable and can mimic with a wide variety of lung diseases.
The inflammatory response causes interstitial mononuclear cell
inflammation that may be manifested radiographically as diffuse,
reticular infiltrates of bronchopneumonia in the perihilar regions or
lower lobes, usually with a unilateral distribution, and hilar adenopathy.
Bilateral involvement may occur in about 20% of cases.
Bacteriological tests
- Blood culture: L. pneumophila can be isolated from blood
culture with low sensitivity.
- Gram stain: L. pneumophila start gram paler color when dyed. M.
pneumoniae results because bacteria do not have cell walls so they do not
color when dyed.
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Respiratory secretions culture: by using a special medium (PPLO
broth environmental bacterium M. pneumoniae culture, environment
chick embryo cells or mice, Hella 229 and cultured Hep 2 find C.
pneumoniae; BCYE environment - Buffered Charcoal Yeast Extract Agar
detect L. pneumophila culture). L. pneumohila usually grows after 3-5
days, M. pneumoniae usually results after 7-21 days later.
Serological methods: the methods are: complement fixation technique
(Complement Fixation - CF), immunofluorescence technique (Immuno
Fluorescence Assay - IFA), Enzyme-linked immunosorbent technique

pneumophila causes severe disease in adults, it occurs rarely in
children under 4 years of age.
1.5.1.2. In Vietnam
Some research interest in disease incidence and clinical features of
pneumonia caused by M. pneumonia among hospitalized children in some
provinces of Vietnam only. Molecular biology techniques are deployed in
some centers, large hospitals nearly.
1.5.2. Studies of severe atypical pneumonia and related factors
1.5.2.1. Etiological bacteria
Pneumonia caused by L.pneumophila that disease is second,
followed by pneumococcal pneumonia requiring intensive treatment.
For people with normal immune systems, the mortality rate is usually in
the range of 10-15%.
1.5.2.2. Coinfection status
Co-infection status was considered as aggravating factors in
community-acquired pneumonia in adults has been demonstrated by
Gutiérrez: pleural effusion, atelectasis, septic shock, hypoxemia
requiring mechanical ventilation, death in patients with pneumonia due
to coinfected patients higher than agent patients (OR = 2.84, 95% CI
1.24 to 6.54, p = 0,02).
1.5.2.3. Accompanying diseases
Studies in adults show that with diseases such as asthma, chronic
obstructive pulmonary disease, malignancies, cardiovascular, diabetes,
immunosuppression are factors that increase the severity of the disease.
1.5.2.4. Specific treatment late
Specific treatment late is emphasized associated with significant
mortality in adults suffer from pneumonia caused by L. pneumophila.
According to Gacouin A., duration of illness before admission to the
ICU for more than 5 days (OR 7:46, 95% CI 1.17 to 47.6) were risk
factors for mortality of L . pneumophila pneumonia.

referred to “ atypical bacterial pneumonia”.
- Patients with atypical pneumonia due to at least one of the three
studied bacteria: M. pneumoniae, C. pneumoniae và L. pneumophila,
were diagnosed severe atypical pneumonia.
2.2. Inclusion criteria
2.
- Pneumonia were diagnosed by using the WHO's criteria:
cough, fever, tachypnea, infiltration on chest radiograph.
2.2.2. Atypical bacterial pneumonia case
Patients were diagnosed with pneumonia
-Three bacteria M. pneumoniae, C. pneumoniae and L.
pneumophila were confirmed by Multiplex PCR in bronchial secretions
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or nasopharyngeal or throat swab positive for the three studied
bacterias, or ELISA: double the serum samples were positive for one of
three studied bacterias.
2.2.3. Severe atypical pneumonia case.
Children were diagnosed with severe atypical pneumonia entitled to
classify pneumonia and WHO standards and the Association of
Pediatric Infection of the America.
- Patients with atypical pneumonia due to at least one of the
three studied bacteria.
- The severity of pneumonia was determined by the criteria for
severe pneumonia of the American Association of Pediatric Infection .
a. One or more major signs: required mechanical ventilation; sepsis
b. Or at least two of the following signs: tachypnea, apnea,
consciousness disorders; hypotension; pleural effusion, SpO
2
<90%
with room air and Pao

−
where n is the minimum sample size, Z (1 - α / 2) is the coefficient of
reliability, corresponding to 95% confidence level we have Z
(1 - α / 2)
= 1.96. p
dependence incidence of pneumonia by M. pneumoniae, C.
pneumoniae, L. pneumophila, estimated in prospective studies on the
incidence of atypical pneumonia in hospital (in this study the rate of p
= 18% = 0.18). q = 1-p = 1 18 = 0.82; p.ε accuracy desired sample, choose
ε = 0.16. A required minimum sample size was 718 patients. We did
enrolled 722 patients for this study.
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 Sample sizes for objective 2
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
2 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.

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 2
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 3. 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
of
Rate%
Typical pneumonia caused by bacteria 82 11.35
Viral pneumonia 80 11.08
Pneumonia caused by typical bacteria co-
infection with virus
14 1.93
Atypical bacterial pneumonia 215 29.8
Pneumonia with unknown etiology 331 45.84
Total 722 100

38 17.67
Atypical pneumonia + viral
pneumonia
19
8.84
Atypicalpneumonia + typical
pneumonia + viral pneumonia
4
1.86
Total 215 100
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%).
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Chart 3.1. Age distribution of typical pneumonia and atypical
pneumonia
Chart 3.1 shows that the proportion of children aged greater than 5 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 5 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)

Chart 3.3. Seasonal distribution of atypical pneumonia
Atypical pneumonia occurs throughout all seasons, more frequent

Caesareans
2.12 1.05 4.30 0.037
Asthma
Yes
1 - - -
No
0.73 0.29 1.84 0.50
Nutritional status
Normal
1 - - -
Wasting
1.82 0.77 4.29 0.17
Overweight and obesity
0.83 0.38 1.81 0.63
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).
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3.1 Table 2. 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 <0,05).
Chart 3.7. Physical symptoms in the lungs of atypical pneumonia
patients
Examination finding moisture and crackles among coinfected
atypical pneumonia outside group higher statistical significance
compared than atypical pneumonia in group (p <0.05) (Figure 3.7).

%
n = 8
Rate
%
Fever 122 94, 57 6 85, 71 7 87, 5
Cough 129 100 7 100 8 100
Headache 37 28.68 3 42.86 2 25
Chest Pain 25 19.38 2 28.57 2 25
Wheeze 84 65.12 4 57.14 4 50
Hoarseness 42 32.56 4 57.14 3 37.5
Table 3:19. Physical symptoms of atypical simple pneumonia
Clinical
Characteristics
Atypical pneumonia
by
M. pneumoniae
Atypical
pneumonia by
L. neumophila
Atypical
pneumonia by
C. pneumoniae
n = 129
Rate
%
n = 7
Rate
%
n = 8
Rate

Congenital malformation of respiratory system
No - - - -
Yes - - - -
Mental development
Normal 1 - - -
Retardation 4.02 1.25 12.91 0.02
Motor development
Normal 1 - - -
Delayed 5.93 1.64 21.48 0.01
Having asthma
No 1 - - -
Yes 1.58 0.73 3.39 0.24
Immunization
Enough 1 - - -
Not enough 5.21 1.08 25.16 0.04
Nutritional status
Normal 1 - - -
Wasting 1.25 0.59 2.68 0.55
Overweight and obesity 0.99 0.51 1.91 0.97
Table 3:23. through univariate analysis the likelihood of severe
atypical pneumonia includes: history of mental retardation (OR = 4.02,
p = 0.02), children with a history of delayed motor development (OR =
5.93, p = 0. 01); Children with a history of inadequate vaccination (OR =
5.21, p = 0.04).
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Table 3.24. Association between etiologies and severe atypical
pneumonia
Factors OR 95% CI p
Etiologic bacteria
M. pneumoniae