Clinical features of severe Mycoplasma pneumoniae pneumonia with pulmonary complications in childhood: A retrospective study

Incidence of severe M. pneumoniae pneumonia (SMPP) reported in China has been increasing over the last decade. We aimed to evaluate the clinical features of pediatric SMPP with pulmonary complications, according to laboratory tests and chest radiographic resolution patterns.


| INTRODUCTION
Mycoplasma pneumoniae (MP) is a common respiratory pathogen of community-acquired pneumonia (CAP) in childhood.Epidemiological studies of Mycoplasma pneumoniae pneumonia (MPP) showed that 37.5% children with CAP were infected with MP. 1 MP infection was traditionally thought to be self-limited with a good outcome, but the incidence of severe Mycoplasma pneumoniae pneumonia (SMPP) cases has gradually increased in recent years, rising from 0.7% in 2006 to 35% in 2016. 1 Children with SMPP have a long hospitalization record, high medical expense and may have long-term complications such as bronchiectasis, bronchitis obliterans and bronchiolitis obliterans. 2,3There are no well-defined and unified diagnostic criteria for pediatric SMPP, and MPP patients either with extensive lung lesions spreading over more than two thirds of the chest area in radiographic image or developing intra-and extrapulmonary complications could be all considered as SMPP.Clearly, SMPP has been recognized to present diverse clinical phenotypes, including pulmonary complication subtype and nonpulmonary complication subtype, which might be associated with the occurrence of airway sequelae.
There are very few articles describing the relationship between pulmonary complications and prognosis in SMPP patients.Moynihan et al. 4 identified 30 confirmed M. pneumoniae infection cases in 11,526 pediatric intensive care unit (PICU) admissions, which included 22 pneumonia cases receiving invasive or noninvasive respiratory support with a median length of 2.0 days in PICU, but no further information of intra-pulmonary complications was reported in these SMPP patients.In Taiwan, Lee et al. 5 found 34 pediatric patients of SMPP, hospitalized in PICU between 2010 and 2019, and 22 of them presented with pleural effusion.The case of one SMPP patient with pleural effusion, showing rapid progression to acute respiratory distress syndrome and evolving into necrotizing pneumonia later, provided limited evidence of pulmonary complications as poor prognostic factor in SMPP.Other clinical studies found that MP genotypes might be useful to predict the progress of SMPP, or atopic individuals are more likely to suffer from SMPP for being more susceptible to extra-pulmonary complications. 6,7However, SMPP patients with or without pulmonary complications were not distinguished in these studies.
In the present study, we used serum biochemical indicators and airway secretion conditions obtained by bronchoscopy to describe the clinical features and prognosis in SMPP children with pulmonary complications, compared with nonpulmonary ones.(CT) features.The degree of secretion in the tracheobronchial tree was determined by using a standardized bronchoscopy scoring system according to the research of Chang et al. 9 This visual grading score graded the secretions in a scale from 1 to 6, based on distribution and amount of mucus in the airways.Trachea and lobar bronchi, including right main stem, right upper lobe, right bronchus intermedius, right middle lobe, right lower lobe, left main stem, left upper lobe, and left lower lobe, were scored.Subjects with grades 1 and 2 were considered to have no secretion, those with grade 3 had minimal secretion, and those with grades 4-6 had mild, moderate, and large secretion, respectively.

| Definition
Pulmonary complications were defined as the occurrence of lobar atelectasis, medium or large pleural effusion, and necrotizing pneumonia.Lobar atelectasis was referred to collapse or incomplete expansion of one or more lobes of lung observed by chest CT.Pleural effusion was confirmed by ultrasonographs; medium pleural effusion was defined as 300-500 mL fluid drained by thoracocentesis; and large pleura effusion, as ≥500 mL fluid drained by thoracocentesis.
Necrotizing pneumonia referred to MPP patients whose lung showed low attenuation area, with or without cavitation on postcontrast enhanced CT scan.Patients with high load of MP were referred to those whose sample exhibited MP DNA loads ≥10 6 copies/mL.Patients with near-complete resolution of chest x-ray or CT referred to those whose chest radiographic or CT image showed the absence of any abnormal findings as infiltrates, atelectasis, or pleural fluid, with only minimal residual changes left.Fever was defined as a temperature of 37.5°C or higher.The total duration of fever was calculated from the date of first symptom onset to the date of defervescence (defined as temperature <37.5°C for at least 24 h) during the hospital admission.

| M. pneumoniae PCR assay
Extraction of DNA from NPA samples and detection of M. pneumoniae DNA by real-time fluorescence PCR assay (ABI 7500 Real-Time PCR System) in these samples were performed with one-step kit (Sansure Biotech Mycoplasma Pneumoniae DNA Fluorescence Diagnostic Kit).
The PCR reaction condition was: 45 amplification cycles of denaturation for 15 s at 94°C, annealing, elongation, and collection of fluorescence data for 30 s at 57°C.The minimum detection limit for MP was 400 copies/mL.

| Statistical analysis
Statistical analyses were performed using the Statistical Package for the Social Sciences (SPSS 22.0).For continuous variables, comparison of means was conducted using t test or Bonferroni test for post hoc analysis in ANOVA.For non-normal data, Mann-Whitney test was used.For categorical variables, the chi-square test, Fisher's exact test, or multiple chi-square test was used.Multiple regression analysis was performed to select the variables associated with intrapulmonary complication and radiographic resolution or complete absorption in chest CT scan.Probabilities of .05 or less were considered significant.

| Clinical and laboratory examination characteristics in 93 severe MPP patients
A total of 93 patients (45 males and 48 females) fulfilled the severe MPP diagnostic criteria between January 2016 and February 2019.
We compared age, total fever duration, and laboratory examinations in SMPP patients with and without pulmonary complications, and the data are shown in Table 2. SMPP patients with pulmonary complications were associated with older age, longer fever days, higher levels of CRP, PCT, D-dimer, LDH, and LDH to albumin ratio (LAR), but lower levels of albumin in serum, when compared to those with no pulmonary complications.Other variables (MP load in NPA, airway secretion conditions, and white blood cell count) showed no difference.One noticeable trend was that the number of patients on corticosteroid therapy was higher in the pulmonary complication group (9/63 vs. 0/30, p = .0537).

| Laboratory examination characteristics and risk indicators of SMPP with different pulmonary complications
Cases of single pulmonary complication were selected, and the data were analyzed and presented in Table 3.There were significant differences in the total duration of fever and levels of CRP, D-dimer, T A B L E 1 Descriptive analysis of demographic, laboratory, and radiographic findings of subjects.

| Time and indicators of chest radiography resolution in SMPP patients
At least one chest x-ray or chest CT was performed on 63 (67.7%) patients who returned for their follow-up examinations, and complete or near-complete resolution was reported in 37 (58.7%)patients, including 22 patients with pulmonary complication.The incidence of SMPP in our study during the year from 2016 to early 2019 was 18.7%.In apparent contrast, Gao et al. 1 reported that the percentage of SMPP between 2015 and 2016 in North China was 35%, which was higher than ours.However, a state-wide multicenter retrospective study in Australia reported that the incidence of SMPP was only 0.3% in PICU admission in Queensland from 2008 to 2013. 4 Approximately 5% of SMPP hospitalized children required ICU admission in a children's hospital in Taipei from 2010 to 2019. 5 It is quite possible that the observed differences are due to geographical and climatic factors as well as the requirement of respiratory support.
We retrospectively analyzed the characteristics of 93 SMPP patients, including 30 patients with extensive pulmonary lesions and 63 patients with intrapulmonary complications, and no case of MPassociated myocarditis or encephalitis was found.Among the patients with pulmonary complications, 84.1% patients had single complication, and 49.1% of them presented pleural effusion.Similar to our results, 64.7% (22/34) of SMPP patients requiring PICU admission presented with pleura effusion in the Taipei study, 5 indicating a high occurrence of pleural effusion as pulmonary complication in SMPP.
Differences were found in clinical indicators between SMPP patients with and without pulmonary complications.Those with no intrapulmonary complications were mostly preschool children, while the ages of the intrapulmonary complications group were mostly over 6 years.
SMPP patients with atelectasis tend to be school-age children, which was a major difference from those with extensive lung lesions.Unlike the atelectasis group, in pleura effusion and necrotizing pneumonia group, fever time was significantly prolonged, and levels of CRP, D-dimer, and LDH, as well as MP load in BALF, were significantly higher than those without complications.Similarly, SMPP patients with pleural effusion in PICU were associated with higher CRP levels and longer fever duration. 5Differences in these parameters were seen in SMPP patients with pleura effusion or necrotizing pneumonia, but not in the atelectasis patients.The current evidence indicates that any inflammation resulting in atelectasis might be relatively mild in comparison to that causing pleural effusion and lung necrosis.
LDH is widely distributed in various tissues of the body, including the lung tissue, and serum LDH levels have long been used for the diagnosis and management of pulmonary infectious diseases as well as for outcome prediction. 10,11A previous study that quantified LDH, combined with ferritin, documented their roles as useful indicators for evaluating MPP conditions, 12 while the severity of MPP in this study was defined as hypoxia, dyspnea, extent of pleural effusion, and lung lesion in chest radiographic images; as mentioned earlier, patients with intrapulmonary complications were not included.Our study found that high LDH and low albumin levels were observed in SMPP patients with intrapulmonary complications, and both indicators were easy to obtain quickly.LAR was elevated in the pleural effusion or lung necrosis group and was in fact found to be one of the independent risk factors for moderate to large pleural effusion occurrence.Of note, LAR represents tissue damage, nutritional status, and systemic inflammatory response, and thus, could help the clinicians to fully assess the progression of severe MP infection.Coagulation abnormalities were common and persistent in CAP patients, especially related to D-dimer levels 13 that were significantly increased in the pleural effusion or lung necrosis group and were also an independent risk factor for pleural effusion and lung necrosis.These results indicate that a hypercoagulable state existed in these patients, which might not only contribute to microthrombus in pulmonary circulation, involved in pathogenesis of these two complications, but also alert us to the possibility of pulmonary thrombosis and lower venous thrombosis.5][16] Furthermore, asthmatic patients might have a different response after MP infection, as the IL-18 response was found to be significantly decreased in the asthmatic SMPP group compared to the non-severe group. 17Yet another study 18 showed that asthma patients were prone to be suffering from refractory Mycoplasma pneumoniae pneumonia (RMPP) indicating that the immune status of asthma patients may have a different pattern in the course of MPP.Since our study excluded asthma patients, further research will be needed to explore the indicators of SMPP with pulmonary complications in asthmatic children.
Previous studies reported that the chest radiographic resolution of 90.3% of RMPP patients occurred in 12 weeks. 19Our study showed that 86.6% of patients without intrapulmonary complications had imaging resolution at 4 weeks, and 72.7% of patients with resolution at 12 weeks.Our results also suggest that the high-level LDH in patients without complication may indicate delayed resolution, which is consistent with the study by Huang et al. 19 D-dimer is also involved in the severity and prognosis of CAP, 20,21 and likewise, we found high levels of D-dimer to be an independent risk factor for delayed resolution in patients with intrapulmonary complications, which suggests that the benefits of early use of anticoagulant therapy need to be evaluated.The bronchoscopic secretion (BS) scoring system was first used to quantify the extent of airway secretions to identify wet coughs. 22Moderate to high airway secretion, as evaluated by this system, was found in over 70% of patients, suggesting that the invasion pathway of MP in our SMPP patients was likely through the respiratory tract and not via the bloodstream.There was no significant correlation between BS grade and the presence of intrapulmonary complications, but patients with complications and lower BS grade showed a tendency of delayed resolution.We speculate that mycoplasma causes damage from distal parenchyma to proximal airways as the invasion progresses.One pleural effusion patient presented minimal airway secretion in the course of 2 weeks, and found mucus plug in the course of 6 weeks, followed by bronchi obliterans in lobe bronchus in the 8th week, suggesting that the local inflammatory injury gradually progressed from the distal alveolar to the large proximal airway and that the recovery was slow and delayed.
Intervention of bronchoscopy, therefore, may be beneficial for discharged patients with delayed resolution, especially those with pulmonary complications in the acute phase of MPP.
The advantage of this study is that all three markers studied, namely LDH, D-dimer, and LAR, can be easily detected, quantified, There were 892 patients with CAP, hospitalized between January 2016 and February 2019 in the Division of Respiratory Medicine at Children's Hospital of Chongqing Medical University, who were diagnosed with MPP.The MPP diagnosis in our study is as follows: acute respiratory infection symptoms (fever, cough, or wheezing), physical examination and chest imaging with infiltrates, and laboratory tests in nasopharyngeal aspiration (NPA) or bronchoalveolar lavage fluid (BALF) samples to confirm MP infection.MPP patients that satisfied the criteria of severe CAP according to the "Guidelines for management of community-acquired pneumonia in children" published by the Society of Pediatrics, Chinese Medical Association 8 were included.The following four characteristics were estimated as criteria of SMPP: (1) tachycardia (judgment criteria: <1 year old, respiratory rate >50 times/min; 1-5 years old, respiratory rate ≥40 times/min; >5 years old, respiratory rate >30 times/min), accompanied with three concave signs and cyanosis, (2) hypoxemia (pulse oxygen saturation is less than 0.92 in condition of air inhalation) (3) lung lesions over more than 2/3 area in chest radiographic image, (4) pulmonary complications such as atelectasis, pleural effusion or necrosis (also called as necrotizing pneumonia).The following patients were excluded from the study: (1) those with conditions of bronchopulmonary dysplasia, congenital heart disease, asthma, and malnutrition (17 patients); and (2) those with other pathogens detected in NPA or BALF (379 patients).In the end, 496 MPP patients were the only MP detected in respiratory samples without any other pathogens.Ninety-three patients (18.7%, 93/496) were diagnosed as SMPP and were enrolled in this study.All observations followed the relevant guidelines and regulations of the Children's Hospital of Chongqing Medical University.The study was approved by the Institutional Review Board, Children's Hospital of Chongqing Medical University.Medical records of all subjects were retrospectively reviewed.Collected data included clinical presentations, NAP and BALF detection of MP load by polymerase chain reaction (PCR), serum biochemical examination of all indicators, bronchoscopy record, and chest radiographic/computed tomography and calculated for a quick evaluation of disease severity and prognosis, combined with clinical manifestations.Nevertheless, some limitations of this study can be noted.First, it is a retrospective study, and we had relatively low amounts of follow-up data after patient discharge.Overall, 32.3% of subjects did not come back for follow-up once after discharge.Second, macrolide-resistant MP was not detected, and hence was not studied.There were four patients with delayed clearance of chest image and they presented moderate mucus in the lumen of affected segment bronchus by follow-up bronchoscopy test.In such cases, macrolide-resistant MP might be associated with the delayed chest image clearance.5| CONCLUSIONSThe clinical features of SMPP were different between the pulmonary complication subtype and the non-complication subtype.Older age, longer duration of fever, higher levels of CRP, LDH, D-dimer, and LAR, and longer time of radiographic resolution were observed in patients with pulmonary complication.LAR and D-dimer might serve as useful predictors of medium or large pleural effusion in SMPP patients; D- dimer might also be an indicator of lung necrosis and delayed radiographic resolution in SMPP patients with intrapulmonary complications.Further studies should shed light on the distribution of macrolide-resistant MP in pulmonary complications and its influence on the chest radiographic resolution.
Laboratory findings in SMPP patients with different pulmonary complications.

Table 4
Abnormality in the last known chest image examination of SMPP patients failed to follow up.