1. The Plot Thickens

2. Patient History
A 69 year old male presented to the emergency department (ED) with fever, altered mental status, and hiccups.
Past medical history
Seizures
Brain cyst removed 2010
Brain shunt placed 2011
History of kidney stones and recurrent urinary tract infections
Empiric treatment started - vancomycin, cefepime, and metronidazole.
Patient’s condition continued to decline. Cefepime was changed to ceftriaxone for possible pneumonia; patient continued on vancomycin.
Blood cultures were drawn.

3. Why are blood cultures (BCx) obtained?
A healthy human’s blood should be sterile
The BCx is the most critical test to determine if a patient has bacteremia (bacterial infection of the bloodstream)
The first 2 sets of blood cultures (from two different “needle sticks”) are collected before antimicrobial treatment is started
Depending on patient’s condition, additional blood cultures may be obtained
Add blood culture insert my Gram stain of GPCC.
Blood draw
10 ml blood/bottle
1 set=1 aerobic + 1 anaerobic bottle
Place bottles in incubator/shaker; automatically monitors for growth
When “positive”:
Gram stain
Subculture for ID & susceptibility

4. How is antimicrobial susceptibility determined?
Disk diffusion (DD) and Broth Microdilution (BMD) minimal inhibitory concentration (MIC) tests are used for antimicrobial susceptibility testing
BMD is done in a 96-well microtiter plate with varying concentrations of several antimicrobial agents
MIC = the lowest concentration of antimicrobial agent showing no growth/turbidity (red circles)
DD is done by spreading bacteria on an agar plate (Muller-Hinton agar) and adding antimicrobial disks
After incubation, the diameters of zones of inhibition around each antimicrobial disk are measured
MIC or zone measurements are interpreted as susceptible, intermediate, or resistant based on predefined criteria

5. Patient’s microbiology workup
Day 1:
Urine and respiratory cultures: Negative
Blood cultures: 2 sets positive
Gram-positive cocci in clusters
MRSA susceptible to vancomycin MIC = 2 µg/ml
Day 3:
Blood Culture: 2 sets positive again with MRSA; vancomycin MIC = 2 µg/ml
Day 6:
Blood Culture: 1 set positive with MRSA; vancomycin MIC increased to 4 µg/ml (interpreted as “intermediate” and this MRSA is a VISA!)
Medical technologist recognized the importance of detecting vancomycin-intermediate Staphylococcus aureus (VISA) (MIC 4-8 µg/ml) and called the physician and infection control

6. Overview of Vancomycin
Glycopeptide first released in 1958
Drug of choice for severe infections caused by MRSA
Most MRSA do not develop vancomycin resistance even when serially passaged on media containing vancomycin
Vancomycin-resistant Staphylococcus aureus (VRSA; (MIC >8 µg/ml) are very uncommon
First VRSA case was reported in US in 2002
Only 16 VRSA cases reported worldwide to date

7. Definitions: VSSA, VISA, VRSA
Classification
S. aureus
MIC (µg/mL)
Susceptible (VSSA) ≤2
Intermediate (VISA)
4-8
Resistant (VRSA)
≥16
Detection of VISA is challenging because:
Colonies may grow slower
Colony morphology may be unlike typical S. aureus
CDC recommends using an algorithm, depending on the method used to detect VISA

8. Detection of VISA

9. Tell-tale Signs of VISA From the Bench
VISA is typically isolated from patients who have been on extended vancomycin therapy (weeks)
The vancomycin MIC of this patient’s initial MRSA isolate was 2 µg/ml susceptible
The vancomycin MIC of the third MRSA was 4 µg/ml (VISA)
It is likely selective pressure from vancomycin therapy contributed to emergence of VISA
Patient’s cultures grew both VSSA and VISA
VISA: Pinpoint 48 h; atypical S. aureus appearance; patient on extended vancomycin therapy
MRSA (not VISA): typical S. aureus appearance

10. Treatment Options for VISA
Rifampin + fusidic acid: Combination therapy (not FDA-approved for use in US).
Linezolid (Oxazolidinones): resistance is uncommon
Tigecycline: option for select cases

11. Back to our patient…
Vancomycin was stopped and patient was started on linezolid
Three subsequent blood cultures were all negative
Fever dropped and the patient’s mental status cleared
Patient was discharged 7 days later

12. Max T. Wu, MS, PhD
Dr. Max Wu is an active duty officer in the US Army, attending the CPEP clinical microbiology fellowship at UCLA. Prior to his direct-commissioning into the Army, Dr. Wu held an assistant professor position in Louisiana Tech University, LA. In his Army career, MAJ(P) Wu has successfully contributed to the development of vaccines for group B meningococcal meningitis and shigellosis, and established the first US DoD’s modern microbiology reference lab in sub-Saharan Africa. His research interests include antibiotic resistance mechanisms, detection of MDR organisms in combat wound infections, and vaccine development for diarrheal diseases.