Gram Negative Bacilli Infections-II

Gram‑negative bacilli infections include clinically significant pathogens that cause gastrointestinal, respiratory, urinary, and systemic diseases. This module covers key organisms, transmission, symptoms, and nursing management essential for exams and evidence‑based practice.

Introduction

Gram negative bacilli (GNB) constitute a diverse group of bacteria that are responsible for a wide range of infections in humans. These organisms are characterised by their cell wall structure, which includes an outer membrane containing lipopolysaccharide, contributing to their pathogenicity and resistance to several antibiotics. While fermentative GNB such as members of the Enterobacteriaceae family have been extensively studied, non-fermenting, fastidious, and miscellaneous Gram negative bacilli are increasingly recognised for their clinical significance, especially in hospital settings and among immunocompromised individuals.

The rise in healthcare-associated infections, the emergence of multidrug-resistant strains, and the expansion of at-risk populations have contributed to the growing importance of these organisms. Their identification, clinical implications, and management present unique challenges to clinicians and microbiologists.

Importance and Epidemiology

Non-fermenting and fastidious Gram negative bacilli are ubiquitous in nature, found in water, soil, and various animal reservoirs. In recent decades, their role as opportunistic pathogens has been accentuated by increased use of invasive medical devices, prolonged hospitalisation, and widespread antibiotic use.

These organisms are often implicated in nosocomial infections, including pneumonia, bacteraemia, urinary tract infections, and wound infections. Their detection requires specialised laboratory techniques, and their inherent or acquired resistance to multiple antimicrobials poses significant therapeutic dilemmas.

Classification of Gram Negative Bacilli

Gram negative bacilli are classified based on their metabolic and growth characteristics. For the purpose of this review, we focus on three groups beyond the classic fermenters:

1. Non-Fermenting Gram Negative Bacilli (NFGNB): These do not ferment glucose and include genera such as Pseudomonas, Acinetobacter, Stenotrophomonas, and Burkholderia.
2. Fastidious Gram-Negative Bacilli: These require specialised growth conditions and include Haemophilus, Bordetella, Brucella, Francisella, and Legionella.
3. Miscellaneous Gram Negative Bacilli: A heterogeneous group including Moraxella, Pasteurella, Capnocytophaga, Kingella, and others.

Non-Fermenting Gram Negative Bacilli (NFGNB)

Key Genera and Species

• Pseudomonas aeruginosa: The most clinically significant, known for its versatility and resistance.
• Acinetobacter baumannii: Notorious for causing outbreaks in intensive care units.
• Stenotrophomonas maltophilia: Emerging pathogen, particularly in immunocompromised patients.
• Burkholderia cepacia complex: Important in cystic fibrosis and chronic granulomatous disease.

Clinical Manifestations

NFGNB are responsible for a wide spectrum of infections, mainly in hospitalised and immunocompromised patients.

Pseudomonas aeruginosa:

• Pneumonia: Especially ventilator-associated pneumonia (VAP).
• Bacteraemia: Common in neutropenic and critically ill patients.
• Urinary Tract Infections: Often associated with catheter use.
• Wound and Burn Infections: Characterised by blue-green pus due to pyocyanin pigment.
• Otitis Externa (Swimmer’s Ear): Particularly in diabetics and swimmers.
• Endocarditis, osteomyelitis, and keratitis in specific risk groups.

Acinetobacter baumannii:

• Pneumonia: Frequently in ICU settings, often multidrug-resistant.
• Bacteraemia: Associated with high mortality rates.
• Wound Infections: Including post-surgical and traumatic wounds.
• Urinary Tract Infections: In patients with indwelling catheters.

Stenotrophomonas maltophilia:

• Respiratory Tract Infections: In patients with underlying lung disease.
• Bacteraemia and catheter-related infections.

Burkholderia cepacia complex:

• Chronic Lung Infections: Particularly in cystic fibrosis patients.
• Bacteraemia and opportunistic infections.

Laboratory Diagnosis

Diagnosis of NFGNB requires a systematic approach involving specimen collection, culture, and identification:

Specimen Collection:

• Appropriate specimens include sputum, blood, urine, wound swabs, and catheter tips, depending on the site of infection. Aseptic technique is essential to avoid contamination.

Cultural Characteristics:

• On MacConkey agar, most NFGNB form non-lactose fermenting colonies.
• Pseudomonas aeruginosa produces blue-green colonies with a fruity odour; may show metallic sheen.
• Acinetobacter forms smooth, opaque colonies; sometimes mistaken for Gram positive cocci due to variable Gram staining.
• Stenotrophomonas forms pale yellow colonies.

Biochemical Tests:

• Oxidase test: Pseudomonas is oxidase positive; Acinetobacter is oxidase negative.
• Motility: Pseudomonas is motile; Acinetobacter is non-motile.
• API 20NE, VITEK 2, and other automated systems are frequently used for identification.

Molecular Methods:

• PCR-based assays and MALDI-TOF MS are increasingly used for rapid and accurate identification.

Antimicrobial Susceptibility Testing:

• Disc diffusion, broth microdilution, or automated systems are used to determine susceptibility.
• Detection of resistance mechanisms (e.g., carbapenemases, metallo-beta-lactamases) is crucial for guiding therapy.

Fastidious Gram-Negative Bacilli

Key Genera and Species

• Haemophilus influenzae: Leading cause of respiratory tract infections, meningitis in children.
• Bordetella pertussis: Agent of whooping cough.
• Brucella species: Zoonotic pathogens causing brucellosis.
• Francisella tularensis: Causative agent of tularemia.
• Legionella pneumophila: Responsible for Legionnaires’ disease.

Clinical Manifestations

Haemophilus influenzae:

• Respiratory Tract Infections: Otitis media, sinusitis, epiglottitis, pneumonia.
• Meningitis: Especially in unvaccinated children.
• Sepsis: In infants and immunocompromised adults.

Bordetella pertussis:

• Pertussis (Whooping Cough): Characterised by paroxysmal cough, whoop, post-tussive vomiting.
• Complications: Pneumonia, seizures, encephalopathy.

Brucella species:

• Brucellosis: Undulating fever, sweats, malaise, hepatosplenomegaly, lymphadenopathy.
• Chronic infection: Arthritis, spondylitis, endocarditis.

Francisella tularensis:

• Tularemia: Ulceroglandular form is most common; also oculoglandular, pneumonic, typhoidal forms.

Legionella pneumophila:

• Legionnaires’ Disease: Severe atypical pneumonia, often with hyponatraemia, diarrhoea, confusion.
• Pontiac Fever: Milder, flu-like illness.

Laboratory Diagnosis

Fastidious GNB require enriched or selective media and specific laboratory techniques:

Specimen Collection:

• Nasopharyngeal swabs (for Bordetella), blood, CSF, sputum, tissue biopsies, depending on clinical syndrome.

Cultural Characteristics and Media:

• Haemophilus: Requires chocolate agar (factor V and X). Satellite phenomenon with Staphylococcus aureus.
• Bordetella: Bordet-Gengou or Regan-Lowe medium; slow-growing.
• Brucella: Grows on enriched media such as blood agar and Brucella agar; incubation up to 3 weeks may be needed.
• Francisella: Requires cysteine-enriched media (e.g., chocolate agar, buffered charcoal yeast extract).
• Legionella: Buffered charcoal yeast extract (BCYE) agar is essential.

Biochemical and Serological Tests:

• Haemophilus: Requirement for X (haemin) and V (NAD) factors, porphyrin test.
• Bordetella: Biochemical inactivity, agglutination with specific antisera.
• Brucella: Urease positive, oxidase positive, serological tests (Rose Bengal, SAT, ELISA).
• Francisella: Difficult to culture; serology and molecular methods preferred.
• Legionella: Urinary antigen test, direct fluorescent antibody (DFA), PCR.

Molecular Methods:

• PCR assays for rapid detection and identification, especially for Bordetella, Brucella, Francisella, and Legionella.

Miscellaneous Gram Negative Bacilli

Examples and Relevance

• Moraxella catarrhalis: Common cause of otitis media, sinusitis, bronchitis, and exacerbations of COPD.
• Pasteurella multocida: Soft tissue infections following animal bites.
• Capnocytophaga canimorsus: Severe sepsis after dog bites, especially in asplenic individuals.
• Kingella kingae: Osteoarticular infections in children.
• Other genera: Eikenella, Cardiobacterium, Chromobacterium, etc.

Clinical Manifestations

Moraxella catarrhalis:

• Respiratory Tract Infections: Otitis media, sinusitis, bronchitis, pneumonia.

Pasteurella multocida:

• Cellulitis: Rapidly developing after animal bites or scratches.
• Systemic Infections: Rarely, can cause bacteraemia, endocarditis, meningitis.

Capnocytophaga canimorsus:

• Sepsis: Particularly in asplenic, alcoholic, or immunocompromised patients.
• Cellulitis and disseminated infections.

Kingella kingae:

• Osteoarticular Infections: Septic arthritis, osteomyelitis in young children.
• Bacteraemia: Less common.

Laboratory Diagnosis

Diagnosis of miscellaneous GNB relies on clinical suspicion and appropriate laboratory techniques:

Specimen Collection:

• Swabs from wounds, blood, synovial fluid, respiratory samples as indicated.

Cultural Methods:

• Moraxella: Grows well on blood and chocolate agar; oxidase positive, produces hockey puck colonies.
• Pasteurella: Grows on blood and chocolate agar, not on MacConkey. Characteristic musty odour.
• Capnocytophaga: Slow-growing, requires CO2 enrichment.
• Kingella: Beta-haemolytic, oxidase positive, may require enriched media.

Biochemical and Molecular Tests:

• Moraxella: Butyrate disk positive.
• Pasteurella: Indole positive, oxidase positive.
• MALDI-TOF MS and PCR for rapid species-level identification.

Distinguishing Features and Clinical Relevance

Laboratory Diagnosis

Specimen Collection

• Site-specific specimens (e.g., sputum, blood, urine, pus, CSF, synovial fluid).
• Strict aseptic technique to avoid contamination.
• Prompt transport to laboratory to preserve organism viability.

Culture Techniques

• Routine media: Blood agar, MacConkey agar, chocolate agar.
• Specialised media: BCYE for Legionella, Regan-Lowe for Bordetella, Brucella agar for Brucella.
• Enrichment with CO2 or specific nutrients for fastidious/miscellaneous GNB.

Biochemical Tests

• Oxidase, catalase, indole, urease, motility, and sugar utilisation tests.
• Commercial identification systems (API 20NE, VITEK, MALDI-TOF MS).

Molecular Methods

• PCR assays for rapid detection and identification (e.g., Bordetella, Brucella, Legionella).
• Multiplex PCR, real-time PCR, and sequencing for epidemiological typing.

Serological and Antigen Detection

• Urinary antigen tests (e.g., Legionella).
• Serological assays for Brucella, Francisella, and others.

Antimicrobial Susceptibility Testing

• Disc diffusion, broth microdilution, E-test.
• Detection of resistance genes (e.g., carbapenemases, ESBLs, metallo-beta-lactamases).

Clinical Manifestations: Overview

Common Infections and Organ Systems Affected

• Respiratory Tract: Pneumonia, bronchitis, sinusitis, otitis media (Pseudomonas, Haemophilus, Moraxella).
• Bloodstream: Bacteraemia, sepsis (Acinetobacter, Burkholderia, Capnocytophaga, Brucella).
• Central Nervous System: Meningitis, encephalitis (Haemophilus, Brucella, Pasteurella).
• Genitourinary Tract: UTIs (Pseudomonas, Acinetobacter).
• Soft Tissue and Bone: Cellulitis, osteomyelitis, septic arthritis (Pasteurella, Kingella, Burkholderia).
• Others: Endocarditis (Brucella, Capnocytophaga), chronic lung infections (Burkholderia).

Complications

• Septic shock and multi-organ failure (especially with multidrug-resistant NFGNB).
• Chronic and relapsing infections (Brucella, Burkholderia, Kingella).
• Neurological sequelae (pertussis, meningitis).
• Severe morbidity and mortality in immunocompromised and hospitalised patients.

Treatment and Management

General Principles

• Empiric therapy should be guided by local epidemiology and susceptibility patterns.
• Definitive therapy based on culture and susceptibility results.
• Combination therapy may be necessary for multidrug-resistant organisms.
• Removal of infected devices (e.g., catheters) when feasible.

Antimicrobial Resistance

• High rates of resistance among NFGNB, particularly Acinetobacter and Pseudomonas (carbapenem resistance, ESBLs, aminoglycoside resistance).
• Emerging resistance in fastidious and miscellaneous GNB (macrolide resistance in Bordetella, beta-lactamase production in Moraxella).
• Antimicrobial stewardship and infection control are critical to limit spread.

Specific Considerations

• Pseudomonas: Piperacillin-tazobactam, ceftazidime, carbapenems, aminoglycosides, colistin (in resistant cases).
• Acinetobacter: Carbapenems, sulbactam, colistin, tigecycline (depending on susceptibility).
• Stenotrophomonas: Trimethoprim-sulfamethoxazole preferred.
• Haemophilus: Amoxicillin-clavulanate, cephalosporins, macrolides (depending on resistance).
• Bordetella: Macrolides (azithromycin, clarithromycin).
• Brucella: Doxycycline with rifampicin or streptomycin.
• Legionella: Macrolides or fluoroquinolones.
• Pasteurella: Penicillin, amoxicillin-clavulanate.
• Moraxella: Augmentin, cephalosporins, macrolides.

Prevention and Control

Infection Control Measures

• Strict hand hygiene and use of personal protective equipment in healthcare settings.
• Environmental cleaning and disinfection to prevent outbreaks (especially for Acinetobacter, Pseudomonas, Legionella).
• Surveillance and prompt isolation of infected/colonised patients.
• Vaccination (e.g., Haemophilus influenzae type b, pertussis) to reduce incidence of vaccine-preventable infections.
• Safe handling of animals and animal products (Brucella, Pasteurella).

Public Health Implications

• Reporting of notifiable diseases (e.g., brucellosis, pertussis, legionellosis).
• Education of healthcare workers and the public regarding risk factors and prevention.
• Antimicrobial stewardship programmes to limit emergence and spread of resistance.

Conclusion

Non-fermenting, fastidious, and miscellaneous Gram negative bacilli are increasingly recognised as significant pathogens in both community and healthcare settings. Their clinical manifestations are diverse, ranging from mild respiratory tract infections to severe, life-threatening sepsis. Laboratory diagnosis relies on a combination of conventional and advanced techniques, emphasising the need for clinical suspicion and close liaison between clinicians and microbiologists.

The rising tide of antimicrobial resistance among these organisms underscores the importance of judicious antibiotic use, robust infection control measures, and ongoing research into novel diagnostics and therapeutics. Vaccination and public health interventions remain vital in reducing the burden of certain fastidious GNB. Future directions include the development of rapid molecular diagnostics, new antimicrobial agents, and enhanced surveillance to stay ahead of these adaptable and often elusive pathogens.

REFERENCES

1. Apurba S Sastry, Essential Applied Microbiology for Nurses including Infection Control and Safety, First Edition 2022, Jaypee Publishers, ISBN: 978-9354659386
2. Joanne Willey, Prescott’s Microbiology, 11th Edition, 2019, Innox Publishers, ASIN- B0FM8CVYL4.
3. Anju Dhir, Textbook of Applied Microbiology including Infection Control and Safety, 2nd Edition, December 2022, CBS Publishers and Distributors, ISBN: 978-9390619450
4. Gerard J. Tortora, Microbiology: An Introduction 13th Edition, 2019, Published by Pearson, ISBN: 978-0134688640 
5. Durrant RJ, Doig AK, Buxton RL, Fenn JP. Microbiology Education in Nursing Practice. J Microbiol Biol Educ. 2017 Sep 1;18(2):18.2.43. https://pmc.ncbi.nlm.nih.gov/articles/PMC5577971/

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