Mycoplasma pneumoniae – Habitat, Morphology, Pathogenesis, Treatment

What is Mycoplasma pneumoniae?

• Mycoplasma pneumoniae is a unique bacterium and one of the smallest self-replicating organisms. It belongs to the Mollicutes class, which is characterized by a lack of a cell wall. This absence of a cell wall makes it resistant to antibiotics that target cell wall synthesis, such as beta-lactams. M. pneumoniae is a known human pathogen that causes an atypical form of pneumonia, often referred to as “walking pneumonia,” which differs from typical bacterial pneumonia in its milder symptoms and slow onset.
• The discovery of Mycoplasma pneumoniae traces back to 1898, when Nocard and Roux isolated a microorganism believed to cause pneumonia in cattle. This organism shared similarities with pleuropneumonia-like organisms (PPLO), which were later found to be responsible for diseases in animals. However, its true nature remained unclear until 1944 when Monroe Eaton, using embryonated chicken eggs, identified the “Eaton agent,” an organism believed to cause primary atypical pneumonia in humans. At the time, researchers thought it might be a virus due to its cultivation method, which was common for viral pathogens. However, its susceptibility to broad-spectrum antibiotics raised doubts about its viral nature.
• In 1961, Robert Chanock and Leonard Hayflick revisited the Eaton agent’s classification. Hayflick, familiar with mycoplasmas, hypothesized that the Eaton agent might not be a virus but a mycoplasma. This theory was confirmed when Hayflick isolated the organism in a novel medium, establishing that the pathogen was indeed Mycoplasma pneumoniae. This marked the first time a mycoplasma was definitively identified as the cause of a human disease.
• M. pneumoniae’s pathogenic mechanisms involve its ability to adhere to the respiratory tract’s epithelial cells. It uses a specialized attachment organelle that enables it to bind to cells and disrupt their function. This attachment contributes to symptoms like persistent cough, bronchial inflammation, and even complications such as chronic obstructive pulmonary disease (COPD) and bronchial asthma. The bacterium also releases hydrogen peroxide, leading to further damage to the respiratory tissues. Additionally, M. pneumoniae produces a virulence factor known as the CARDS toxin, which plays a significant role in respiratory distress and inflammation.
• In terms of treatment, antibiotics like macrolides and tetracyclines are used to inhibit protein synthesis in M. pneumoniae. However, antibiotic resistance has been increasingly reported, especially in regions like Asia. Resistance often arises due to mutations in the 23S rRNA gene, which prevent the effective binding of macrolides. This has led to the need for alternative treatment strategies, highlighting the ongoing challenge in managing mycoplasma infections.
• From a biological perspective, M. pneumoniae is considered an obligate parasite, relying on the host’s cellular machinery for survival. It has a reduced genome with limited metabolic pathways, meaning it must scavenge resources from the host to sustain itself. This metabolic simplicity, coupled with its ability to mimic the host cell surface, contributes to the persistence of infections even after treatment, making M. pneumoniae a complex pathogen to manage in clinical settings.
• Taxonomically, M. pneumoniae belongs to the Mycoplasmataceae family and the Mycoplasmatales order. It shares a common evolutionary lineage with gram-positive bacteria, such as bacilli and streptococci, but has undergone significant genomic reduction, reflecting its parasitic lifestyle. Understanding the biology and pathology of M. pneumoniae is essential for developing more effective treatments and managing its increasing resistance to conventional antibiotics.

Domain:	Bacteria
Phylum:	Mycoplasmatota
Class:	Mollicutes
Order:	Mycoplasmatales
Family:	Mycoplasmataceae
Genus:	Mycoplasma
Species:	M. pneumoniae

Geographical Distribution and Habitat of Mycoplasma pneumoniae Infections

M. pneumoniae infections are found worldwide, with varying rates of occurrence depending on the region. This bacterium is a leading cause of community-acquired pneumonia, contributing to up to 20% of cases that require hospitalization.

Geographical Distribution

• M. pneumoniae infections occur in both epidemic and endemic forms, meaning they can cause localized outbreaks or be present continuously at low levels.
• The infection is well-documented in European countries and the United States, where it has a notable impact on respiratory health.
• Data from developing countries is limited, but seroprevalence studies suggest that M. pneumoniae may be endemic in many of these regions, indicating a consistent presence of the infection despite a lack of widespread reporting.

Habitat

• M. pneumoniae is a strict human pathogen, meaning it only infects humans and does not have animal or environmental reservoirs.
• In infected individuals, the bacteria primarily reside in the upper respiratory tract, where they adhere to the mucosal surfaces.
• The bacteria live extracellularly, meaning they do not invade cells but instead remain on the surface of the mucosa, causing direct damage to epithelial cells.

Morphology of Mycoplasma pneumoniae

Mycoplasma pneumoniae, like other mycoplasmas, has unique morphological characteristics that distinguish it from other bacteria. Below are key features of its structure and behavior:

• Small Size: Mycoplasma pneumoniae is among the smallest self-replicating organisms, measuring only 150–250 nm in size.
• Lack of Cell Wall: It lacks a traditional cell wall, which is a defining feature of mycoplasmas. This absence contributes to its resistance to antibiotics that target cell wall synthesis, like penicillins and cephalosporins.
• Sterols in the Cell Membrane: The cell membrane of Mycoplasma pneumoniae contains sterols, a characteristic not seen in most other bacteria. This feature helps stabilize its membrane in the absence of a cell wall.
• Pleomorphism: Mycoplasma pneumoniae exhibits pleomorphism, meaning it can take various forms. It can appear as granular shapes or as filaments, which can vary in size and may form true branches. These variations contribute to its versatility in surviving in different environments.
• Absence of Flagella and Pili: Unlike many other bacteria, Mycoplasma pneumoniae does not possess flagella or pili, structures typically involved in motility and attachment. Despite this, it displays gliding motility on liquid-covered surfaces, which is believed to aid in its movement along respiratory tract cells.
• Binary Fission and Fragmentation: Mycoplasma pneumoniae reproduces through binary fission, although this process is often asynchronous. This results in the production of multinucleate fragments and chains of bead-like structures, adding to its complex morphology.
• Staining Characteristics: Mycoplasma pneumoniae does not stain well with the traditional Gram stain due to its lack of a cell wall. However, it can be effectively stained using Giemsa and Diene stains, which reveal its structure more clearly.
• Resistance to Certain Antibiotics: The lack of a cell wall and the presence of sterols in the membrane contribute to its resistance to common antibiotics like penicillins, cephalosporins, and vancomycin, which target cell wall synthesis.

Culture and Biochemical Reactions of Mycoplasma pneumoniae

Mycoplasma pneumoniae has distinct requirements for both culture and biochemical reactions, which play a significant role in its growth and identification.

• Culture Conditions:
  • Strict Aerobe: Unlike many mycoplasmas, M. pneumoniae is a strict aerobe and requires oxygen for growth.
  • Growth Temperature and pH: It grows best at a temperature of 37°C and a pH range of 7.3–7.8.
• Media for Culturing:
  • PPLO Broth: This medium is commonly used to isolate mycoplasmas. It contains 20% horse serum, 10% yeast extract, and glucose. The addition of phenol red helps to monitor pH changes.
    • Horse serum provides essential sterols like cholesterol and lipids, which mycoplasmas cannot synthesize themselves.
    • Antibiotics such as penicillin, ampicillin, polymyxin B, and amphotericin B are included to prevent contamination from bacteria and fungi.
  • PPLO Agar: When PPLO broth is solidified with agar, it allows for better observation of mycoplasma colonies. The colonies are small and typically exhibit a fried-egg appearance, characterized by:
    • A central opaque, granular area.
    • A surrounding translucent, peripheral zone.
  • Colony Characteristics: M. pneumoniae differs from other mycoplasmas in colony formation. It forms mulberry-shaped colonies, not the typical fried-egg appearance. These colonies lack the thin halo seen in other species.
  • Slow Growth: M. pneumoniae is particularly slow-growing, requiring 1 to 4 weeks for colonies to appear.
• Biochemical Reactions:
  • Carbohydrate Utilization: M. pneumoniae primarily utilizes glucose and other carbohydrates for energy. This is common among various mycoplasma species.
  • Chemo-organotrophic Metabolism: Mycoplasma species are chemo-organotrophs, meaning they rely on organic compounds for energy. The metabolism is largely fermentative, with glucose fermentation resulting in lactic acid production and a drop in pH.
  • Arginine Utilization: Some mycoplasma species, like M. salivarium and others, use arginine as their primary energy source. The breakdown of arginine produces ammonia, CO2, and adenosine triphosphate (ATP), which causes the pH of the medium to become alkaline.
  • Fermentation in Liquid Culture: For fermentation testing, liquid culture mediums are supplemented with glucose, arginine, and urea, with phenol red as an indicator.
    • Glucose-fermenting mycoplasmas turn the medium acidic due to lactic acid production.
    • Arginine-fermenting species result in an alkaline shift due to ammonia production.

Cell Wall Components and Antigenic Structure of Mycoplasma pneumoniae

Mycoplasma pneumoniae lacks a typical cell wall, but its membrane still carries several critical components that play a central role in its ability to cause disease. These components include glycolipids and proteins that contribute to its antigenic structure and interactions with the host immune system.

• Membrane Glycolipids and Antigenic Cross-Reactivity
  • Glycolipids in the membrane are major antigenic determinants of M. pneumoniae.
  • These glycolipids have been shown to cross-react with human tissues and other bacteria, particularly human neurons.
  • Cross-reactivity occurs because the glycolipids share similar antigenic structures with those found in neurons.
  • Complement fixation tests are used to identify these glycolipid antigens in the laboratory.
  • The antibodies produced against M. pneumoniae glycolipids can attack the human brain, leading to potential neurological damage.
  • This cell damage could explain the neurological symptoms observed in some cases of M. pneumoniae infections, such as confusion or encephalitis.
• Surface Proteins of M. pneumoniae and their Role
  • Two major surface proteins on M. pneumoniae include P1 adhesion protein and another unnamed surface protein.
  • The P1 protein is essential for the bacteria to attach to cell structures in the human respiratory tract.
  • These surface proteins are identified using enzyme-linked immunosorbent assay (ELISA), a common diagnostic technique.
• P1 Protein and Autoimmune Responses
  • The P1 protein triggers the body to produce antibodies not only against itself but also against antigenic determinants present on red blood cells (RBCs).
  • This leads to autoimmune reactions, where the immune system mistakenly attacks the body’s own cells.
  • The reaction can result in erythrocyte lysis (destruction of red blood cells), contributing to anemia or other related issues.

Virulence Factors of Mycoplasma pneumoniae

Mycoplasma pneumoniae uses several mechanisms to establish infection and cause disease. One of the primary factors contributing to its virulence is its ability to attach to host cells, and this process is largely driven by its adhesion protein P1.

• Adhesion Protein P1
  • P1 protein is the key virulence factor of M. pneumoniae.
  • This membrane-associated protein helps the bacteria attach to epithelial cells in the human respiratory tract.
  • P1 protein binds specifically to sialated glycoprotein receptors, which are located at the base of cilia on the epithelial surface.
  • The same receptors that P1 attaches to are also present on erythrocytes (red blood cells), enabling a dual interaction with both the respiratory epithelial cells and the bloodstream.
  • By binding to these receptors, P1 enables the bacteria to remain firmly attached to the host, allowing for prolonged infection.
• Autoimmune Reactions Triggered by P1
  • Antibodies produced against P1 protein can cross-react with red blood cells (RBCs), leading to their agglutination.
  • This interaction causes an autoimmune response, as the immune system attacks the body’s own RBCs, which can result in further complications like hemolysis (destruction of red blood cells).
• Localized Infections with Limited Systemic Spread
  • While M. pneumoniae can cause significant damage at the site of infection in the respiratory tract, it typically does not invade the bloodstream, preventing widespread systemic manifestations.
  • This characteristic helps limit the infection to the lungs and respiratory system but still allows for severe respiratory illness.

Pathogenesis of Mycoplasma pneumoniae Infections

Mycoplasma pneumoniae causes respiratory infections primarily through its interaction with the host’s epithelial cells. The bacteria rely on their ability to attach to the host cells and disrupt normal respiratory function, leading to the disease’s hallmark symptoms.

• Initial Attachment and Cell Damage
  • The bacteria first attach to the epithelial cells in the upper respiratory tract.
  • After attachment, M. pneumoniae damages cilia and then destroys the ciliated epithelial cells.
  • The loss of these cells impairs normal function of the upper respiratory tract, making it less effective at filtering and clearing pathogens.
  • As a result, the lower respiratory tract becomes vulnerable to secondary infections and irritation by microbes.
• Infection and Persistent Cough
  • The destruction of epithelial cells creates mechanical irritation in the respiratory system.
  • This irritation leads to the development of a persistent cough, which is a common symptom in patients with M. pneumoniae infections.
• Immune Response and Inflammatory Reaction
  • M. pneumoniae acts as a superantigen, triggering an exaggerated immune response.
  • The bacteria cause the migration of inflammatory cells to the site of infection, producing key cytokines such as tumor necrosis factor-alpha (TNF-α), interleukin-1 (IL-1), and interleukin-6 (IL-6).
  • These cytokines help in the clearance of the bacteria and contribute to the inflammatory response.
• Limited Invasion and Systemic Effects
  • M. pneumoniae generally does not invade the bloodstream, which helps prevent widespread systemic infection.
  • In most cases, the infection remains localized in the respiratory tract, without causing systemic manifestations.
• Rare Cases of Invasion
  • Although rare, M. pneumoniae can penetrate the submucosa, particularly in cases of immunosuppression or after certain medical procedures (like instrumentation).
  • When the bacteria invade the bloodstream in these rare instances, they can lead to infections in other organs of the body.

Clinical Syndromes of Mycoplasma pneumoniae Infections

M. pneumoniae is primarily known for causing respiratory infections in humans. While most cases are mild and self-limiting, the bacteria can lead to various clinical manifestations that affect different parts of the respiratory tract and even lead to extrapulmonary complications.

Respiratory Infections

• Upper Respiratory Tract Infections
  • Typically cause mild symptoms like low-grade fever, malaise, and headache.
  • Patients experience a nonproductive cough that usually appears 2-3 weeks after exposure.
  • Over time, the cough may become productive, with sputum that can range from small amounts to mucopurulent or even blood-tinged in more severe cases.
• Lower Respiratory Tract Infections
  • Includes tracheobronchitis and bronchopneumonia.
  • The infection begins in the bronchi, leading to infiltration of bronchial epithelial cells by lymphocytes and plasma cells.
• Primary Atypical Pneumonia (Walking Pneumonia)
  • Incubation period typically lasts 2-3 weeks.
  • Patients often appear not as ill, making the illness commonly referred to as walking pneumonia.
  • The pharynx becomes edematous, but no cervical adenopathy is present.
  • Chest X-rays show patchy bronchopneumonia, which often does not correlate with the mild physical findings, creating a disparity between symptoms and radiological evidence.
  • The condition is mostly self-limiting, but pleural effusion can occur in 5-20% of patients.

Extrapulmonary Infections

• Cardiac Abnormalities
  • Myocarditis and pericarditis are the most frequently reported extrapulmonary manifestations of M. pneumoniae infection.
• Neurological Abnormalities
  • Infections may also result in various neurological issues, although these are less common.
• Otitis Media
  • Middle ear infections can also occur as a result of the infection, especially in younger individuals.
• Erythema Multiforme and Stevens–Johnson Syndrome
  • Skin reactions like erythema multiforme, including Stevens–Johnson syndrome, are reported in some patients.
• High-Risk Populations
  • M. pneumoniae infections tend to be more severe in:
    • Children with immunosuppressive diseases.
    • Individuals with sickle cell anemia or functional asplenia.
    • Children with Down syndrome.
• Subclinical Infections
  • Around 20% of adults may experience subclinical infections, which are often undiagnosed due to the absence of severe symptoms.

Reservoir, Host, and Transmission of Mycoplasma pneumoniae Infection

M. pneumoniae primarily infects humans, and humans are the main host for this bacterium. The infection is often passed from one individual to another through close contact and aerosolized droplets.

Reservoir and Host

• Humans act as the primary reservoir for M. pneumoniae. People with active infections are the key sources of transmission.
• Children aged 9-10 years and young adults are most likely to carry and spread the infection.
• The infection is also increasingly seen in older adults, particularly those over the age of 65, where it contributes to about 15% of cases of community-acquired pneumonia.
• M. pneumoniae is the second most common cause of pneumonia in older adults, following Streptococcus pneumoniae.

Transmission

• Transmission occurs when infected individuals release aerosolized droplets through coughing, sneezing, or talking. These droplets are inhaled by others, leading to new infections.
• Close living conditions, such as those found in college dormitories or military barracks, increase the likelihood of transmission due to frequent, close contact.
• The infection is more common among groups who live in close quarters, where respiratory droplets can spread easily.

Laboratory Diagnosis of Mycoplasma pneumoniae Infection

Diagnosing Mycoplasma pneumoniae infection in the lab isn’t always straightforward, and the process typically takes weeks. Treatment is often started based on clinical signs before test results are available. Here’s how the process generally goes.

Specimen Collection

• Respiratory specimens like throat washings, bronchial washings, and expectorated sputum are typically collected.
• Tracheal washings are more effective than sputum since many patients have a dry, nonproductive cough.
• Once collected, specimens must be transported to the lab immediately.
• If there’s a delay, transport media such as SP4 can be used to prevent the specimens from drying out.
• If transportation is delayed further, specimens can be stored at -70°C.

Microscopy

• Microscopy isn’t much help for diagnosing M. pneumoniae. Since the bacteria lack a cell wall, they stain poorly and don’t show up well under a microscope.

Direct Antigen Detection

• The antigen capture immunoassay is used for direct detection of M. pneumoniae in sputum, with high sensitivity and specificity.
• Tests like direct immunofluorescence, counter-current immunoelectrophoresis, and immunoblotting can detect M. pneumoniae antigens in specimens.

Culture

• Culturing M. pneumoniae can confirm the diagnosis but takes time—3 to 4 weeks due to the bacteria’s slow growth.
• For culturing, specimens are inoculated into mycoplasma medium like PPLO agar, supplemented with nutrients and antibiotics to suppress unwanted bacterial and fungal growth.
• Growth is slow and requires incubation at 37°C in an atmosphere of 95% N2 and 5% CO2.
• Colonies appear as mulberry-shaped clusters, and their growth is typically slow and non-distinctive without special stains or tests.

Colony Identification

• Color change in the agar is one way to identify M. pneumoniae colonies. The pH indicator in the medium will turn from red to yellow as glucose fermentation occurs, lowering the pH.
• Diene stain is another useful technique. Colonies stained with Diene stain appear granular with a fried-egg appearance. The center stains dark blue, and the outer edge light blue.
• Hemadsorption test: M. pneumoniae colonies will adsorb guinea pig erythrocytes when incubated with them. This test confirms identification when observed under the microscope.
• Tetrazolium reduction test: M. pneumoniae reduces tetrazolium to form a red-colored compound, indicating the presence of the bacteria.

Serodiagnosis

• Diagnosing via serology looks for specific antibodies in the patient’s serum.
• Complement fixation test can indicate a recent infection if there’s a rise in antibody titer, typically after 7-10 days of infection. The antibody titer peaks around 4-6 weeks.
• IgM ELISA is a frequently used method. It detects IgM antibodies in serum with high sensitivity (97%) and specificity (99%). Results can often be obtained within 30 minutes.

Nonspecific Serological Tests

• Streptococcus MG agglutination test uses heat-killed Streptococcus MG as an antigen. If the serum produces an agglutination at 1:20 titer or higher, it suggests M. pneumoniae infection.
• Cold agglutination test detects antibodies that agglutinate human O group erythrocytes at low temperatures (4°C). The titer of 1:32 or more is indicative of M. pneumoniae infection. However, this test can be nonspecific since similar agglutinins appear in other infections, like rubella, infectious mononucleosis, and others.

Treatment of Mycoplasma pneumoniae Infection

When it comes to treating Mycoplasma pneumoniae infections, antibiotics aren’t always needed for upper respiratory issues. But when it comes to pneumonia caused by this bacteria, antibiotics make a difference.

Here’s a closer look at how treatment works:

• Antibiotics aren’t always necessary for upper respiratory infections but are helpful when it comes to pneumonia. They help to:
  • Reduce the illness duration.
  • Lower the number of Mycoplasma bacteria in clinical specimens.
  • Alleviate symptoms.
  • Support faster recovery.
  • Promote pneumonia resolution.
• Pneumonia caused by M. pneumoniae is generally self-limiting. In most cases, it’s not life-threatening, but symptoms can still be bothersome.
• Effective antibiotics for treating M. pneumoniae include:
  • Tetracyclines: These are effective against M. pneumoniae and also work against many other mycoplasmas and chlamydiae, which are common causes of nongonococcal urethritis.
  • Erythromycin: Also works by inhibiting protein synthesis in mycoplasma bacteria.
• What to avoid: M. pneumoniae is resistant to antibiotics that target the cell wall, such as:
  • Penicillins
  • Cephalosporins Since Mycoplasma bacteria lack cell walls, these antibiotics won’t work.

Prevention and Control of Mycoplasma pneumoniae Infection

Preventing the spread of Mycoplasma pneumoniae and controlling infections requires careful measures, as no vaccine currently exists for this pathogen. Focusing on isolation and appropriate use of antibiotics is key to reducing transmission.

• Isolation of infected individuals:
  • The best way to stop the spread of M. pneumoniae is isolating infected patients.
  • This prevents the bacteria from reaching healthy individuals and keeps outbreaks contained.
• Antibiotic prophylaxis:
  • Using tetracyclines or erythromycin as preventive measures can help control the spread, especially in close-contact settings.
  • These antibiotics are effective at preventing infection in those who have been exposed.
• No vaccine available:
  • At this point, there is no vaccine for Mycoplasma infections, which means prevention relies heavily on isolation and antibiotics.