Analysis of mycoplasma pneumoniae infection among children with respiratory tract infections in hospital in Chengdu from 2014 to 2020

Introduction

Respiratory tract infections (RTIs) are one of the most common diseases among children. RTIs are caused by a board range of pathogens, including viruses, bacterial pathogens, and atypical bacterial pathogens. Mycoplasma pneumoniae (M. pneumoniae) is a common bacterial pathogen that causes lower RTIs in children and young adults. It is one of the smallest prokaryotic cell types of microorganisms that lack cell walls, is highly polymorphic, and can pass the bacteria filter and grow on inanimate objects. In recent years, the incidence of M. pneumoniae has been increasing (1-3). Different areas in different cities have different pathogen spectrums and epidemiological features (3,4). To date, the data available about the prevalence of M. pneumoniae infection in pediatric patients in Chengdu, Sichuan Province, China, including during the period in which Coronavirus Disease 2019 (COVID-19) spread in China, have been fragmentary. This study sought to investigate the effects of M. pneumoniae in hospitalized children between January 2014 and December 2020. We present the following article in accordance with the STROBE reporting checklist (available at http://dx.doi.org/10.21037/tp-21-139).

Methods

Subject information

The data of 22,882 in-patients suffering from RTIs, who had been admitted to Chengdu Women’s and Children’s Hospital, Chengdu, China, were used in this study. Diagnoses were made in accordance with the World Health Organization’s criteria. Of the 22,882 patients, 13,600 children were male and 9,282 were female. Patients’ ages ranged from 1 month to 17 years old. Patients were classified into the following age subgroups based on different physiological stages: infants (1 month–1 year old); toddlers (>1–3 years old); preschoolers (>3–6 years old); school-age children (7–17 years old). Venous blood was drawn from every patient. Serum samples were separated and tested for the M. pneumoniae immunoglobulin M (IgM) and the antibody. A patient was diagnosed with M. pneumoniae infection, if M. pneumoniae IgM was positive or the antibody ≥1:160 (5). The demographic features of patients, clinical diagnoses and laboratory data were analyzed. Data provided by the Chengdu Meteorological Service were examined to determine if there was an association between gender, age, month, year, PM_2.5 (particulate matter in the air <2.5 µm), PM₁₀(particulate matter in the air <10 µm), temperature, humidity and cases of M. pneumoniae infection. All procedures performed in this study involving human participants were in accordance with the Declaration of Helsinki (as revised in 2013). The study was approved by institutional ethics board of Chengdu Women’s and Children’s Central Hospital, School of Medicine, University of Electronic Science and Technology of China [NO.: B2021(8)]. Individual consent for this retrospective study was waived.

Blood sampling

A sample of venous blood was drawn from each child. Each sample was centrifuged at 2,000 rpm for 10 min at 4 °C. The serum was separated and stored at –20 °C until it was assayed with the M. pneumoniae IgM, and the antibody test was undertaken.

M. pneumoniae IgM test: Indirect immunity and the antibody positive titer test

Each slide had 1 well containing M. pneumoniae antigens. Serum samples were diluted 1:1 with phosphate buffered saline (PBS), and then treated with anti-human immunoglobulin G (IgG) sorbent. The sorbent-treated diluted serum was incubated for 90 min at 37 °C with the slide wells. Each slide was washed twice with PBS before the fluorescent secondary IgM antibody was added to the well and incubated at 37 °C for 30 minutes. Each slide was washed twice with PBS and the greenish-yellow fluorescent signal was detected with a fluorescence microscope (Zeiss, Oberkochen, Germany). IgM antibodies against M. pneumoniae in serum were detected using a passive agglutination kid (Fujirebio, Japan) in accordance with the manufacturer’s instructions.

Statistical analysis

Categorical variables were presented as numbers with percentages and compared using a χ² test. A value of P<0.05 was considered statistically significant. The risk factors of M. pneumoniae infection were assessed using a binary logistic regression. Associations were quantified as odds ratios (ORs) and 95% confidence intervals (CIs). Analyses were carried out using the Statistical Package for the Social Sciences for Windows (SPSS, version 11.0).

Results

The positive rates of M. pneumoniae infection across different genders

Of the total of 22,882 samples tested, 4,213 samples (18.41%) were positive for M. pneumoniae infection. Of these, 2,104 (18.30%) males were positive, and 2,109 (22.72%) females were positive (χ²=198.078, P<0.01).

The positive rates of M. pneumoniae infection across different age groups

Overall, the positive percentages of M. pneumoniae infection were 4.24% in infants, 26.92% in toddlers, 33.98% in preschoolers, and 32.98% in school-age children. Notably, the positive percentage rate increased as the age of the patients increased (F=162.7532, P<0.01; see Table 1).

Table 1 The M. pneumoniae positive rates across different age groups
Full table

Annual incidence and seasonal incidence of M. pneumoniae infection

The positive rate of M. pneumoniae infection was significantly different among children of different ages (F=538.95, P<0.01). Notably, the incidence of M. pneumoniae infection in 2014 (13.14%), 2015 (10.47%), and 2020 (17.98%) was relatively low. However, the incidence of M. pneumoniae infection was significantly higher than the average rate in 2017 (26.59%) and 2019 (28.18%). Further, the incidence of M. pneumoniae infection was high from April to May, and September to October in 2016 (F=38.3378, P=0.0001), 2017 (F=110.8096, P<0.01), 2018 (F=21.6136, P=0.0275), and 2019 (F=35.6517, P=0.0002); however, no mensal preference was observed in 2014 (F=4.7018, P>0.05), 2015 (F=10.5878, P=0.4784), or 2020 (F=10.1404, P=0.5178; see Figure 1).

Figure 1 The seasonal incidence rates of M. pneumoniae from 2014 to 2020.

The association between M. pneumoniae infection and PM_2.5, PM₁₀, temperature, and humidity

There was no correlation between M. pneumoniae infection and temperature (R=0.1441, P>0.05) or humidity (R=−0.006, P>0.05; see Figures 2,3). However, there was a negative correlation between PM_2.5 (R=−0.293, P<0.01) and PM₁₀ (R=−0.285, P<0.01) (see Figures 4,5).

Figure 2 The correlation between a positive rate of M. pneumoniae infection and temperature.

Figure 3 The correlation between a positive rate of M. pneumoniae infection and humidity.

Figure 4 The correlation between the rate of M. pneumoniae infection and PM_2.5.

Figure 5 The correlation between the rate of M. pneumoniae infection and PM₁₀.

The constituent ratios of cases of M. pneumoniae infection

There was no difference between the constituent ratios of cases of M. pneumoniae infection between 2014 and 2019 (F=32.34, P>0.05). Bronchopneumonia, a common respiratory disease of M. pneumoniae infection, accounted for the highest proportion of cases, followed by the exacerbation of asthma and severe pneumonia. However, there was a difference in the constituent ratio of cases of M. pneumoniae infection in 2020 compared to other years (F=159.35, P<0.01). The incidence of severe pneumonia decreased significantly in 2020, and bronchopneumonia accounted for the highest proportion of cases, followed by acute bronchitis and the exacerbation of asthma (see Figure 6).

Figure 6 The constituent ratios of cases of M. pneumoniae infection.

Discussion

M. pneumoniae is a minimum microbe as bacteria and virus. It can cause pharyngitis, otitis, tracheobronchitis, or bronchopneumonia, and is transmitted by respiratory droplets. However, the positive rates of M. pneumoniae differ in different countries, different areas, across different ages, in different years and in different seasons. To provide more evidence for diagnoses and treatments, it is important to know the characteristics of M. pneumoniae in local areas.

The positive rate of M. pneumoniae was 18.41% for children in hospital for RTIs from 2014 to 2020 in Chengdu. This figure was lower than that of other areas in China (4). It was also lower than that of other countries, such as Japan and Peru (6,7). However, it was higher than that in Russia where M. pneumoniae was detected to be about 15.9% (8). Thus, geographical factors appear to contribute to apparent differences in M. pneumoniae infection. However, it should be noted that the different results could have been affected by the various research samples and the different detection methods used.

The distribution of M. pneumoniae across sex differed across reports (6). The present study found that the positive percentage of M. pneumoniae in male children was lower than that in female children. Similar results have been reported in other areas in China (9,10). However, the reasons for such differences between the sexes remain unexplained. Thus, more studies need to be conducted to examine this issue in the future.

The data showed a significant association among age groups. Age is an important factor that can affect pathogen distribution. We found that preschoolers and school-age children had high incidence rates of M. pneumoniae infection of 33.98% and 32.98%, respectively. In China, children go to daycare when they are about 3 years old and to primary school when they are about 7 years old. Compared to the spacious housing conditions of infants and toddlers, the preschoolers and school-aged children lived or studied in crowded places because of the large populations, and M. pneumoniae spread easily among children (see Table 1; P<0.05). A study in Indian also reported the positive rate of M. pneumoniae infection was 59.5% in children (11). However, in older children, the immune response of organisms improves, and the pathogen can produce an effective response. Thus, M. pneumoniae infection plays an important role in this age group. Future research should seek to examine the learning and living environments of preschoolers and school-age children as conditions allow.

The positive percentage of M. pneumoniae was different in different years from 2014 to 2020. Epidemics of M. pneumoniae infection occur at intervals of 3 to 7 years (12). There was a general upward trend in volatility in the last 7 years of our study. It appears that in 2017 and in 2019 in Chengdu, there was a M. pneumoniae infection epidemic (the rates for these years were 26.59% and 28.18%, respectively). Similarly, that there was an outbreak of M. pneumoniae in Finland in 2017 (13). Somewhat surprisingly, there was a lower incidence of M. pneumoniae infection in 2020 than other years, when COVID-19 spread in China. Further, there was no seasonal variation in the incidence of M. pneumoniae infection in 2020. The prevention and control of the COVID-19 epidemic began in January 2020. It appears that these prevention measures effectively controlled the infection rate of M. pneumoniae. M. pneumoniae infection can occur at any time of year, and were the most common from April to May and September to October in 2016, 2017, 2018, and 2019 in Chengdu (see Table 1; P<0.05); however, this was different in other areas (14,15). The present study demonstrated that there were seasonal variations in the incidence of M. pneumoniae infection. Interestingly, we also found that there was a relatively low rate of M. pneumoniae infection in winter vacation (February) and summer vacation (July to August) every year in Chengdu. It may be that the physical changes after vacations and crowded school environments play an important role in M. pneumoniae infection in children. It is important that doctors are aware of the character of sustainability and the aperiodic spreading of M. pneumoniae in local areas, and also regularly monitor M. pneumoniae to gain insights in how to avoid M. pneumonia infection at school.

In the present study, we examined whether there was an association between M. pneumoniae infection and PM_2.5 (particulate matter in the air <2.5 µm) and PM₁₀ (particulate matter in the air <10 µm), temperature and humidity in Chengdu. Some cohort studies have identified consistent associations between ambient PM_2.5 and cardiorespiratory morbidity (16). Our study showed there was a negative correlation between the positive rate of M. pneumoniae infection and PM_2.5 and PM₁₀. Notably, there was serious air pollution in Chengdu in 2014 and 2015. This has been improved after some progress was made in environmental protection in 2020. Air pollution was found to increase the number RTI cases; however, it is not known whether the pathogeny of such cases was M. pneumoniae. The positive rate of M. pneumoniae infection was relatively low in 2014 and 2015. No association was found between M. pneumoniae infection and the average temperature and humidity of the climate, and it was different from other opinions (17). The temperature and humidity of the climate may affect the growth of M. pneumoniae; however, this study only analyzed the infection of M. pneumoniae, and did not examine the carriage of M. pneumoniae. It should be noted that high numbers of healthy children carry M. pneumoniae (18).

Of the children in-patients who were infected with M. pneumoniae between 2014 and 2019, cases of bronchopneumonia were the most common, followed by the acute exacerbation of asthma, and severe pneumonia. Some have conjectured that M. pneumoniae infection plays a key role in community-acquired pneumonia in children (19,20). However, other studies have shown that children with acute bronchitis have the highest positive rate of M. pneumoniae (21). These different results may have arisen as the data in this study was collected from in-patient children only and not outpatient children. Previous studies have shown that M. pneumoniae may play a role in the onset of asthma in predisposed children, and could trigger recurrent wheezing (22). The present study also confirmed that M. pneumoniae infection is frequently associated with exacerbations of asthma in children (23). It may be that children with M. pneumoniae infection with exacerbations of asthma have undergone empirical treatments before the exact pathogen had been identified. M. pneumoniae pneumonia is typically mild and self-limiting; however, severe M. pneumoniae pneumonia can cause severe pulmonary complications (e.g., obliterative bronchitis, bronchiectasis, and necrotizing pneumonia), and has increased in China (24). In the present study, we also found that cases of severe pneumonia are common in children in-patients who were infected with M. pneumoniae. Further, Lobar pneumonia increased in 2017, 2018, and 2019. However, the rate of severe pneumonia was significantly reduced in 2020. COIVD-19 may have weakened the virulence of M. pneumoniae. Future research should seek to confirm this in the future. However, in epidemic times, we should be aware that patients with M. pneumonia may be at risk of severe lung disease.

In conclusion, this retrospective analysis of M. pneumoniae in Chengdu from 2014 to 2020 revealed characteristics about M. pneumoniae in the local area. These results can help doctors to make clinical diagnoses, determine treatments, and prevent RTIs. By comparing the different characteristics of M. pneumoniae in 2020 to those of other years, we found that prevention strategies, such as health education, washing hands, and wearing masks, can control the epidemic of M. pneumoniae. This study had several limitations. First, it did not analyze the characteristics of outpatients with M. pneumoniae infection. Second, the tests conducted did not analyze patients with single M. pneumoniae infection or co-infections with other pathogenies.

Acknowledgments

Funding: None.

Footnote

Reporting Checklist: The authors have completed the STROBE reporting checklist. Available at http://dx.doi.org/10.21037/tp-21-139

Data Sharing Statement: Available at http://dx.doi.org/10.21037/tp-21-139

Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at http://dx.doi.org/10.21037/tp-21-139). The authors have no conflicts of interest to declare.

Ethical Statement: The authors are accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. All procedures performed in this study involving human participants were in accordance with the Declaration of Helsinki (as revised in 2013). The study was approved by institutional ethics board of Chengdu Women’s and Children’s Central Hospital, School of Medicine, University of Electronic Science and Technology of China (NO.: B2021[8]). Individual consent for this retrospective study was waived.

Open Access Statement: This is an Open Access article distributed in accordance with the Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International License (CC BY-NC-ND 4.0), which permits the non-commercial replication and distribution of the article with the strict proviso that no changes or edits are made and the original work is properly cited (including links to both the formal publication through the relevant DOI and the license). See: https://creativecommons.org/licenses/by-nc-nd/4.0/.

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(English Language Editor: L. Huleatt)