Comprehensive Microbiology Case Study: Investigation of a Nosocomial Infection Outbreak

Background:

An outbreak of nosocomial (hospital-acquired) infections has been reported in a major urban hospital. Several patients in the Intensive Care Unit (ICU) have developed symptoms of sepsis, and initial investigations suggest a common bacterial source. This case study outlines the steps taken to identify the causative agent, determine its source, and implement control measures.

Case Presentation:

Patient Symptoms:

• High fever
• Chills
• Elevated heart rate
• Hypotension
• Elevated white blood cell count

Initial Steps:

Patient Sample Collection:

• Blood cultures from all symptomatic patients
• Swabs from wound sites and central venous catheter insertion points
• Urine and sputum samples if applicable

Laboratory Techniques and Results:

1. Blood Culture and Gram Staining:

• Procedure: Inoculated blood culture bottles were incubated at 37°C. Growth was monitored, and Gram staining was performed.
• Results: Gram-positive cocci in clusters were observed in all positive cultures.

1. Culturing on Selective Media:

• Procedure: Blood samples were streaked on Blood Agar (BA), MacConkey Agar (MAC), and Mannitol Salt Agar (MSA) plates and incubated at 37°C for 24-48 hours.
• Results: Yellow colonies grew on MSA, indicating the presence of Staphylococcus aureus. No growth on MAC, confirming the organism is not Gram-negative. Beta-hemolytic colonies on BA.

1. Biochemical Testing:

• Procedure: Coagulase test, catalase test, and other relevant biochemical tests were performed on isolated colonies.
• Results: Positive coagulase and catalase tests confirmed the presence of Staphylococcus aureus.

1. Molecular Techniques:

• Procedure: DNA was extracted from isolated colonies. PCR targeting the mecA gene and pulsed-field gel electrophoresis (PFGE) for strain typing were performed.
• Results: PCR confirmed the presence of the mecA gene, indicating methicillin resistance (MRSA). PFGE showed identical banding patterns among patient isolates, indicating a common source.

1. Antibiotic Sensitivity Testing:

• Procedure: The Kirby-Bauer disk diffusion method was used on Mueller-Hinton Agar.
• Results: The isolates were resistant to methicillin, oxacillin, and several other antibiotics but susceptible to vancomycin and linezolid.

Environmental Sampling:

• Procedure: Swabs from potential reservoirs (sinks, ventilators, hospital staff hands, and equipment) were cultured and identified using the same techniques as patient samples.
• Results: MRSA was isolated from the ventilators used in the ICU, matching the strain from Case Study Questions for Students:

1. What are the primary characteristics of Methicillin-resistant Staphylococcus aureus (MRSA) that make it a significant pathogen in nosocomial infections?
2. Explain the significance of using selective media such as Blood Agar, MacConkey Agar, and Mannitol Salt Agar in isolating and identifying the pathogen.
3. Describe the role of Gram staining in the identification process of the bacteria isolated from the patients. Why is this step crucial?
4. How does the presence of the mecA gene detected by PCR confirm methicillin resistance in Staphylococcus aureus?
5. Discuss the importance of antibiotic sensitivity testing in the management of bacterial infections, specifically in the context of this outbreak.