The study of Mycoplasma pneumonia infection among children with respiratory tract infection in hospital in Chengdu from 2014 to 2020

Aim: The hospitalized children with Mycoplasma pneumonia (M. pneumonia) infection caused by respiratory tract infection in Chengdu were studied and analysis of the epidemiological characteristics was carried out to provide a theoretical basis for clinical diagnosis and treatment. Method: 22882 hospitalized children with respiratory tract infections between January 2014 and December 2020 were collected M. pneumonia IgM antibody was detected by indirect immunouorescence method and passive agglutination method. Demographic characteristics, clinical diagnose and laboratory data of these children were analyzed.retrospectively.. Result : The 4213 specimens with M. pneumonia were tested positive, the total positive rate was18.41% (18.30% in male and 22.72% in female). Higher positive rates were found in female children,Look from the statistical analysis results, the consistency between the two sets of data is low (cid:0)(cid:0) 2 =198.078 (cid:0) P (cid:0) 0.01 (cid:0) . The results of different age patients with contrast different M. pneumonia infection degree were statistically signicant (cid:0) F=162.7532 (cid:0) P (cid:0) 0.01 (cid:0) ,there was higher M. pneumonia positive rate in Preschoolers and school-age children ,33.98% and 32.98%, respectively.The incidence rate of M. pneumonia in 2017 and 2019 was signicantly higher than average (cid:0) F=538.95, P (cid:0) 0.01)The difference of incidence rate of M. pneumonia was not signicant in different months in 2014, 2015 and 2020 (P>0.05). But the probability of M .pneumonia infection patients was much higher from April to May and September to October in2016,2017,2018 and 2019 (cid:0) P (cid:0) 0.05 (cid:0) . There was no correlation about M. pneumonia infection with temperature and humidity( P (cid:0) 0.05) (cid:0) there was negative correlation with PM 2.5 (cid:0) R=0.09362, P<0.01 (cid:0) and PM 10 . (cid:0) R=0.1185,


Background
Respiratory tract infection is one of common diseases among the children which could be caused by a board range of pathogens including viruses ,bacterial pathogens and atypical bacterial pathogens. Mycoplasma pneumonia is a common bacteral pathogen which is dominated by lower respiratory tract infections in children .M.pneumonia is one of the smallest prokaryotic cell type of microorganisms that lack of cell walls, highly polymorphic, and can pass the bacteria lter and grow on the inanimate medium.
In recent years, the incidence of M. pneumonia infection rate has an increasing trend [1][2][3] .
There is different pathogen spectrum and epidemiological feature in different areas in the city. Up to date,there are fragmentary data about prevalence of M. pneumonia infection pediatric patients in Chengdu, Sichuan Province, China, including during the time which 2019 coronavirus diseases  spreaded in China.The aim of this paper is mainly to understand the epidemiological distribution of M. pneumonia in hospitalized Children between January 2014and December 2020 Informed consent was obtained from a parent of patients. The retrosceptive studies was approved by the ethics committee of the Chengdu Women's and Children's Hospital ,SiChuan, China.

Subject information
The study was conducted on22882 inpatients suffering from respiratory tract infection in Chengdu Women's and Children's Hospital, Chengdu, China. Diagnosis followed World Health Organization Criteria. Among the patients, 13600 children were male and the rest were female. Aged from 1month to 17years old. Ages were classi ed into subgroups corresponding to different physiological stages.
infants group(1month 1 year old),toddlers group( 1 3 years old),preschoolers group( 3 6 years old),school-age children group (7 17 years old).Serums sample of every patient were separated from venous blood drawn and tested with pneumonia IgM and antibody. The case was de ned with M. pneumonia infection,which M. pneumonia IgM was positive and antibody ≥1:160. [4] Demographic features of patients,clinical diagnose and laboratory data were analyzed . To explore whether there was an association with gender, age, month, year,PM 2.5 ,PM 10 , temperature, humidity (data was kindly provided by Chengdu Meteorological Service and the aetiological organism,.all methods were carried out in accordance with relevant guidelines and regulations Blood sampling 22882 samples of venous blood were drawn from each child. The samples were centrifuged at 2000g for 10 min at 4℃. Serum was separated rstly and then stored at -20℃ until it was assayed with the pneumonia IgM and antibody test. M. Pneumonia IgM test (Indirect immunity)and antibody positive titer test .
First at 1:1 ratio in the sample with phosphate buffer salt (PBS), get after dilution of the sample, then drip in the samples of human immunoglobulin adsorbent, and the sample in 37 ℃ slide drilling stand for 90 minutes, to join in the rst samples of two kinds of uorescent auxiliary agent in PBS, observing the existence of IgM antibody in the samples, and placed at 37 ℃ environment, wait 30 minutes later, the two samples are recorded with a uorescent microscope imaging of imaging signal, ( Zeiss, Germany)..IgM antibodies against M. pneumonia in serum were detected using a passive agglutination kid (Fujirebio, Japan) based on the manufacturer's instructions.

Statistical analysis
Categorical variables were given as numbers with percentages and a value of p < 0.05 was considered to be statistically signi cant when compared with X 2 -test. Meanwhile, the risk factors of M. pneumonia infection were assessed by using binary logistic regression. Associations were quanti ed with odds ratios (OR) and their 95% con dence intervals (CI). Analyses were conducted using the Statistical Package for the Social Sciences for Windows (Spss, version11.0) The positive rates of M. pneumonia infection in different age groups As a whole to sum up, the positive percentage of M. pneumonia infection was 4.24% in infants, 26.92% in toddlers, 33.98% in preschoolers and 32.98% in school-age children. As listed in Table 1, the positive percentage increased with the age of the patients F=162.7532 P 0.01 (Table 1)  2014 to 2020 in Chengdu,It was lower than in other areas in China [5], It was also lower than other countries, such as in Japan ,in Peru . [6][7] but it was higher than in Russia where the M. pneumonia was detected about 15,9% [8] It was believed that geographical factor contributed to the apparent differences of M. pneumonia infections Of course, The different results were affected by various research samples and different detection methods .

Results
The gender distribution in M. pneumonia had different reports [6] , which showed the positive percentage of M. pneumonia in male children was lower than that of female children.The similar result was reported in other areas in China [9] [10] . but the reasons for differences in prevalence according to sex are unexplained now,The more studies should be con rmed in the future.
Our data showed a signi cant association among age groups. Age is an important factor that can affect pathogen distribution,We found the preschoolers and school-age children had the higher incidence rate of M. pneumonia infections which reached 33.98% and32.98% . In China, children start live or study in crowed because of large population at age of three ,such as in kindergarten,in primary school ,so the M. pneumonia is most easily spread among preschoolers and school-age children .(chart1 P 0.05) ..A study conducted in India which has the same large population also reported the positive rate of M. pneumonia infection was 59.5% in Children [11] .At the same time, the immune response ability of organism is improved, and the pathogen can be produced effective response in older children.So the M. pneumonia infections played an important role in this age group .Therefor,It should pay more attention to improve the learning and living environment of preshoolers and school-age children as conditions allow.
There were different positive percentage of M. pneumonia in different years from 2014 to 2020. It was reported the epidemics of M. pneumonia infections occurred at intervals of 3 to 7 years [12] There was a general upward trend in volatility in recent seven years in our study. It seemed the M. pneumonia infection epidemic time was 2017and 2019 in Chengdu , respectively 26.59%and 28.18%.it was the similar report that there was a outbreak of M. pneumonia in Finland in 2017 [13] . To our surprise, there was a lower  [14] [ 15] .Though,Our study demonstrated that there was seasonal variation in the incidence of M. pneumonia infection. Interestingly,we also found that there was a relatively lower rate of M. pneumonia infection in winter vacation (February)and summer vacation(July,August) every year in Chengdu. We Suspicious the physical changes after the vacation, and the crowded environment in school also play a important role in M. pneumonia infection in children.So it is important for doctors not only to know the character of sustainability and aperiodic spreading about M. pneumonia in local area , to do regular monitoring of M. pneumonia forever,but also to enhance the education how to avoid the M. pneumonia infection in school.
In this study ,We uniquely explored whether there was an association between M. pneumonia infection and PM 2.5 ( The ne particulate matter in the air<2.5um) PM 10 ( The ne particulate matter in the air<10um) , temperature and humidity in Chengdu . Some cohort studies had identi ed consistent associations between ambient PM 2.5 and cardiorespiratory morbidity [16] .Our study showed there was negative correlation between the positive rate of M. pneumonia infection and PM 2.5 ,PM 10 . In fact ,There was serious air pollution in Chengdu in 2014and 2015. It had been improvement after making some progress in environmental protection to 2020. Air pollution increased the number respiratory tract infection cases ,but the pathogeny may be not the M. pneumonia,so the positive rate of M. pneumonia infection was relatively low in 2014 and 2015. There was no associated between M. pneumonia infection with the average temperature and humidity of the climate,it was also different from other opinion [17], The temperature and humidity of the climate maybe affect the growth of M.pneumonia ,but during the study ,we only analyzed the infection of M.pneumonia,not carriage of M. pneumonia .After all,there was high rates of healthy children carry M.
In the hospitalized children who were infected with M. pneumonia between 2014 and 2019, The cases of bronchopneumonia was the most common,followed by acute exacerbation of asthma and severe pneumonia. it was the similar opinion that the M.pneumonia infection played a key role in CAP (community-acquried pneumonia) in children [19] [20]. .But other studies showed the highest M. pneumonia positive rate for children with acute bronchitis [21]. The different results were may be affected by the object being investigated who came from the inpatient children only, not outpatient children in our study . Previous studies showed M. pneumonia may act a part in the onset of asthma in predisposed children and could cause recurrent wheezing [22] . our study also con rmed that the infection with M. pneumonia was frequently correlated with exacerbation of asthma in Children [23] , so it maybe suggested empirical treatments to M. pneumonia infection in the children with exacerbation of Asthma, before knowing the exact pathogen.Though the M. pneumonia pneumonia (MPP) is typically mild and self-limiting, but severe M. pneumonia pneumonia (SMPP) , including severe pulmonary complications( such as obliterative bronchitis,bronchiectasis and necrotizing pneumonia (NP)) have increased in China [24]. In our study ,we also found the cases of severe pneumonia was common too in the children inpatients who were infected with M. pneumonia,what's more ,Lobar pneumonia increased in 2017,2018 and 2019. But the the rate of severe pneumonia had signi cantly reduced in 2020. We suspected whether the COIVD-19 maybe weaken the virulence of M. pneumonia, The more studies should be con rmed in the future. However, in the M. pneumonia, epidemic time,we should aware of the possibility of severe lung disease.
In conclusions, through retrospective analysis of the M. pneumonia results in Chengdu from 2014 to 2020,we know the characteristic about M. pneumonia in local area. It can help doctors to make clinical diagnosis and treatments and prevention of respiratory tract infection reasonably. Comparing the different characteristic of M. pneumonia in 2020 with in other years ,we found the prevention strategies such as health education,washing hands,wearing mask can also control the epidemic of M. pneumonia.But there were several limitations to our study .First ,this study do not analyze the characteristic of outpatients of M. pneumonia infection. Second,our testing did not include analyzing the patients with the single M. pneumonia infection or co-infection with other pathogen.