M. pneumoniae is a species of bacteria without a cell wall that belongs to the class Mollicutes.1 Infection by M. pneumoniae spreads through respiratory droplets and the incubation period is of 2 to 4 weeks. The immunity generated by infection is temporary, and reinfection is frequent.1 Asymptomatic carriage status can persist for a few months, even in patients treated with antibiotics.1

M. pneumoniae has virulence factors which gives it a high affinity to respiratory epithelial cells, the capacity to cause direct damage, extracellular and intracellular survival and diverse interactions with the immune system. The pathogenesis of M. pneumoniae involves direct damage caused by the bacterium and indirect damage mediated by the host immune response and vasculitis or thrombosis.

M. pneumoniae follows a cyclic epidemic pattern that recurs every 3 to 5 years, with outbreaks usually occurring in summer or early autumn.2 During the COVID-19 pandemic, the measures implemented to control coronavirus disease were associated with a significant reduction in the incidence of M. pneumoniae infection.3 Contrary to the trends observed in other respiratory pathogens, the incidence of M. pneumoniae infection did not immediately surge back after the COVID containment measures were lifted,3 and instead an increasing trend was observed starting in January 2023, continuing through the second semester of 2023 in various countries in America, Europe, Oceania and Asia.2

China has reported a significant increase in new cases since June 2023, with a positive detection rate as high as 61% in the samples of children with respiratory illness.4 The current epidemic in China is characterised by an increased rate of macrolide resistance (MR), greater disease severity and an increased incidence in children aged less than 3 years compared to previous years.4

In Spain, accurate data are not available, as this is not a notifiable disease. However, since January 2024, paediatricians across the country have noticed an increase in cases, hospital admissions and extrapulmonary forms of disease, some of them severe. It is still unclear whether the latter increase is proportional to the overall increase in cases or due to the circulation of more virulent strains. Hospital-based registers have been set up to collect descriptive and epidemiological surveillance data on M. pneumoniae infections.

The most frequent presentation is respiratory infection including upper respiratory tract infection, tracheobronchitis and pneumonia. Extrapulmonary manifestations develop in approximately 1 in 4 patients (Table 2).5

The most frequent symptoms of pneumonia caused by M. pneumoniae are: fever (86–96%), persistent dry cough (85–96%), fatigue (78%), dyspnoea (67%), headache (11–48%) and sore throat (12%–47%), while rales are the most frequently detected sign on auscultation.5 These features are nonspecific and overlap with those of other pathogens. Table 2 presents the features associated with an increased or decreased probability of infection by M. pneumoniae, although none of them, together or combined, is sufficient to establish a certain diagnosis.2,5,6

Confirmation of a suspected case of M. pneumoniae infection requires direct detection of the pathogen or evidence of a compatible serologic response (Table 1B),2 as clinical, radiological and laboratory criteria are insufficient to make the diagnosis. At the same time, microbiological and serological criteria also have limitations and must always be interpreted taking into account the clinical presentation and context, as it is not always possible to differentiate between infection and asymptomatic carriage or coinfection with another pathogen.7

Infection by M. pneumoniae tends to be mild and many patients do not require hospitalization, but it may cause severe pneumonia or extrapulmonary manifestations.

The risk of severe pneumonia is greater in children with anatomical or functional asplenia (e.g., children with sickle-cell anaemia), Down syndrome or those who are immunocompromised (Table 1C).8

In addition, although the virulence determinants have only been partially elucidated, the severity of extrapulmonary disease may be determined in part by the individual strain and the toxin concentration it produces.8

Broadly speaking, it is reasonable to say that, as occurs in other types of infection, the characteristics and magnitude of the host immune response and immunocompetence can significantly affect the clinical picture of respiratory disease caused by M. pneumoniae and the wide array of possible extrapulmonary complications. For many years, research has been conducted on the role of the host immune response involving macrophages, mast cells, neutrophils and natural killer cells as well as T and B cells and humoral immune responses. Although the humoral response to infection does not provide full immunity against future infection, the importance of an intact antibody response in containing the disease cannot be overstated. Individuals with antibody deficiencies can develop severe and prolonged respiratory disease and are at increased risk of extrapulmonary complications such as meningitis or arthritis.9

At present, there are no laboratory tests that can be used in isolation for diagnosis of pulmonary infection by M. pneumoniae. Although there are some characteristic clinical and imaging features in children, it is not possible to differentiate community-acquired pneumonia caused by M. pneumoniae from pneumonia caused by other pathogens.2,10

On the other hand, the microbiological tests that are currently available generally offer information that is difficult to interpret.

The ideal specimens for aetiological diagnosis of pulmonary infection include bronchoalveolar lavage, pleural fluid, lung biopsy or tracheal aspiration samples, all techniques that are not used routinely. In real-world clinical practice, specimens tend to be collected from a site that is far from the primary site of infection (like the upper respiratory tract) or from tissues that offer indirect evidence of infection (serological tests), thus offering suboptimal results in terms of sensitivity and positive predictive value.11

Several studies demonstrate that a positive result in any of these tests does not necessarily entail active infection by M. pneumoniae, which severely compromises their positive predictive value.2,10,12 On the other hand, further complicating matters, a negative result in any one test does not rule out the involvement of M. pneumoniae with certainty.13

Due to all these challenges in the interpretation of results, it is strongly recommended that testing for aetiological diagnosis is reserved for patients in whom there is a strong clinical suspicion based on the characteristics discussed in the previous section. Combining tests to try to improve the precision of the aetiological diagnosis (paired serology testing + PCR) is not cost-effective in everyday clinical practice and is not recommended other than for research purposes.

Table 4 summarises the available diagnostic tests.

Customarily, the development of specific IgM or an increase in the IgG titre by at least 4-fold in 2 samples collected at least 2 to 4 weeks apart is diagnostic of acute infection by M. pneumoniae.

This diagnostic criterion only allows retrospective diagnosis and has little impact on care delivery. IgM levels can remain elevated for months after the initial infection and can also, on the contrary, increase only mildly in future reinfections in older children or adults, who may respond only with a transient increase in IgG levels.

A positive IgM result in the first week from onset of symptoms is unlikely.10 The seroprevalence of IgM in the healthy paediatric population can be as high as 40% and some studies have found IgG, IgM and IgA antibodies in healthy children with M. pneumoniae carriage, so the validity and sensitivity of these tests are questionable.10. In addition, serologic tests for detection of IgM can yield false positives on account of: (1) cross-reactivity with other antibodies, (2) the presence of substances that can interfere with the technique, or (3) the limitations of the technique used.

A positive result in molecular diagnostic tests indicates the presence of genetic material of the bacterium in the sample, but there is strong evidence that this does not necessarily prove a causative role in the ongoing infection.2,10,12 In well-selected patients in whom samples are collected correctly, the specificity is greater than 90%, and gradually declines from the first week from onset of symptoms.

The frequency of positive PCR detection in nasopharyngeal samples in the general population can be as high as 45% to 56%10 on account of either asymptomatic carriage or the prolonged presence of the pathogen in the oropharynx after active infection, which may persist for several months.

The near universal availability of PCR techniques allows rapid testing for detection of multiple respiratory pathogens, but the results have to be interpreted correctly to determine whether the detected pathogen is the causative agent.14

Testing for identification of M. pneumoniae should be individualised and only performed when the pre-test probability is high and test results are likely to change the management of the patient (Table 3B).11

In general, performance of diagnostic tests is not recommended in the outpatient management of pneumonia in healthy children in whom empirical therapy tends to be effective, even if it does not include antibiotics with activity against M. pneumoniae.11 The reported positivity rate of PCR testing for M. pneumoniae in respiratory specimens of children with community-acquired pneumonia varies widely in the scientific literature (4%–39%), and a positive result does not allow differentiation between active infection and asymptomatic carrier status.12 The prevalence of M. pneumonia carriage in the upper respiratory tract varies considerably in different years and seasons, and is higher after an epidemic season (given the cyclic pattern of M. pneumoniae). In patients appropriately selected based on clinical suspicion, the positive predictive value and especially the negative predictive value of testing increase and testing results contribute significantly to increasing the appropriate use of antibiotherapy.15

Despite the limitations noted above, PCR testing of respiratory specimens is the quickest method currently available (1–2 h) and is sensitive and specific compared to other microbiological tests, although the value ranges have not been clearly established.16 In the most recent meta-analysis on the usefulness of point-of-care and rapid tests for aetiological diagnosis in paediatric respiratory infections, out of the 57 included studies, only one focused specifically on M. pneumoniae, and it found a substantial increase in appropriate antibiotic prescribing with the use of the test.16 In Spain, molecular tests for detection of MR are not widely available.

Ordering tests for microbiological diagnosis is indicated in the circumstances detailed in Table 3B.2,17,18 The technique to be used will be selected based on availability, cost, the characteristics of the patient and the clinical status (taking into account how quickly results need to be available).

In addition to respiratory infection, neurologic, cutaneous, cardiac, renal, haematological, immunological, muscular, osteoarticular and gastrointestinal involvement in M. pneumonia infection have been described.18 Testing for detection of M. pneumoniae in these systems is not performed routinely, and extrapulmonary testing is most frequently performed in cerebrospinal fluid samples. Although not conducted on a routine basis, testing for detection of the bacterium can be performed in specimens from sites with clinical manifestations resulting from direct damage, but is not indicated in the case of immune-mediated manifestations (Table 5).

It is important to take into account that in many instances, these techniques have not been validated for use in non-respiratory specimens, so their diagnostic value must be interpreted with caution. In light of this, it is more reliable to assess whether the mechanism of damage is direct or indirect based on the form of disease, and the detection of genetic material of M. pneumoniae and the detection by PCR of genetic material of M. pneumoniae in a non-respiratory specimen would simply support the diagnosis, and would not be used to select or modify treatment.19

The recommendations and indications of imaging tests in patients with suspected infection by M. pneumoniae are the same as those that apply to any case of community-acquired pneumonia. In Spain, the gold standard of diagnosis is the plain chest radiograph. A chest radiograph is also indicated if the course of disease differs from what is expected, if complications are suspected or if there is diagnostic uncertainty. The chest radiograph is no more useful for diagnosis than it is for any other cause of pneumonia, and its role in differential diagnosis remains unclear.2

When performed by adequately trained paediatricians, the point-of-care lung ultrasound is an adequate screening technique, as it does not involve exposure to radiation and its sensibility is equal or even superior to that of the chest radiograph if performed 48 to 72 hours from the onset of fever, and is also the imaging test of choice for diagnosis of parapneumonic pleural effusion.

Some authors suggest the use of imaging tests to support the diagnosis in patients with reactive infectious mucocutaneous eruption (RIME) preceded by respiratory symptoms.6

The role of antibiotherapy in the management of M. pneumoniae infections is not well established.20–22 In Spain, most of these infections are self-limiting and mild.23,24

Although some studies conducted in epidemiological contexts different from ours suggest that complications, extrapulmonary manifestations or severe or prolonged disease can develop if diagnosis is delayed or an ineffective antibiotic agent is prescribed,23,25–27 several studies, including a Cochrane review, have concluded that the evidence on the efficacy of antibiotherapy for treatment of respiratory infections by M. pneumoniae is insufficient.22–24

Recent observational studies have found no differences in the course and outcomes of respiratory tract infection by M. pneumoniae in patients with community-acquired pneumonia who received empirical treatment with beta-lactam antibiotics including vs excluding a macrolide, supporting the hypothesis that routine initiation of treatment is not indicated.22,28

Given the lack of scientific evidence in support or against antibiotherapy, it seems reasonable to apply the benefit of the doubt criterion and prescribe antibiotherapy in patients with severe illness that could be caused by M. pneumoniae. In most cases, a watchful waiting approach can be considered, even in patients with community-acquired pneumonia, in the absence of warning signs, as long as it is possible to guarantee close outpatient follow-up.2,29,30 In such cases, the decision whether to perform tests for aetiological diagnosis should be made based on the statements regarding their indication and interpretation presented in Table 3B.

A key factor is the age of the patient. Young children are less likely to have infection by M. pneumoniae compared to school-aged children. However, the incidence of symptomatic infection has increased significantly in infants and preschool-aged children in Asia and Europe, although the impact in terms of the clinical picture is not yet clear.31 In children under 3–5 years, complicated disease with development of pneumonia is infrequent if additional aetiological agents are not concurrently involved.25,26 In other age groups, it is estimated that 10% to 40% may develop pneumonia.25

Considering what has been discussed thus far, prescribing of targeted antibiotherapy against M. pneumoniae is indicated when there is a high clinical suspicion of this aetiology or microbiological tests evince the presence of the bacterium and the patient has risk factors for severe disease (Table 1C) or presents with severe respiratory or extrapulmonary manifestations. Pneumonia with hypoxaemia requiring supplemental oxygen or ventilatory support, encephalitis and certain severe mucosal or cutaneous manifestations are considered indications for antibiotic treatment.2,32,33

In outpatient care settings (primary and emergency care), where molecular tests may not be immediately available, the epidemiological context, age and disease severity must be considered to identify patients with a high probability of infection by M. pneumoniae requiring empirical antibiotherapy (Table 7).2,26

Table 6C presents the different treatment options with the recommended doses and duration of treatment. Azithromycin and clarithromycin are equivalent in terms of efficacy, antimicrobial spectrum, safety profile (especially regarding QT prolongation) and antimicrobial drug resistance mechanisms. Both interact with multiple drugs (they inhibit cytochrome P450 enzymes).

Traditionally, azithromycin has been considered the first-line treatment due to the ease of use and excellent tolerability (gastrointestinal intolerance is reported more frequently with the use of clarithromycin). However, there is growing concern about the induction of antimicrobial resistance due to its long half-life, which results in a prolonged persistence of the drug in the body.2 Thus, from an antimicrobial stewardship standpoint, the use of clarithromycin is prefer, as it reduces the duration of antimicrobial exposure.

In patients with central nervous system involvement, since the evidence on the efficacy of macrolides is insufficient,34 doxycycline is considered the treatment of choice. In real-world clinical practice, macrolides are widely used with the aim of reducing the bacterial load in the body to avoid indirect immune-mediated pathophysiological mechanisms. However, considering the severity of extrapulmonary presentations involving the central nervous system and the possibility that some neurologic manifestations may result from direct damage, tetracyclines are a more judicious option, especially since their use is no longer considered contraindicated in children younger than 8 years as long as the treatment duration is less than 3 weeks.35

Fluoroquinolones are the antimicrobials of choice if a broader spectrum is required to cover other pathogens in empirical antibiotherapy, in immunocompromised patients and if macrolide resistance is suspected.

Macrolide resistance has been increasing significantly, especially in Asia.36,37 In Europe, the reported rates of MR range from 1% to 25%,38 with an approximate prevalence of 8% in Spain.39 Testing for MR is not performed in Spain outside of research projects. Few studies offer data on the clinical impact of MR, which could increase the length of stay or the incidence of complications.40 Since tests for MR are not available in clinical practice, antibiotic selection must be guided by local data on drug resistance, disease severity or the patient’s response to treatment with macrolides.