Discover a New Antibiotic
Looking for Antibiotic-Resistant Bacteria

Christine L.Case
This was published in Strategies for Success Fall 1998.
















S. aureus are gram-positive bacteria that cause pneumonia and infections of the bloodstream, skin, soft tissues, and bone; this pathogen frequently causes community-acquired infections and is the most common cause of nosocomial infections. In the preantibiotic era, S. aureus infections were a common cause of death. Although the availability of penicillin in the 1940s offered an important advance in the treatment of infection, susceptibility of S. aureus was short-lived.

Resistance was first recognized in 1944 and was due to production of penicillinase; by the late 1950s, approximately 50% of strains were resistant to penicillin. These strains were associated with wide-spread disease outbreaks before the development of semisynthetic penicillinase-resistant agents, such as methicillin, in 1960. Resistance to methicillin was reported as early as 1961 in England. In the United States, the proportion of methicillin-resistant S. aureus (MRSA) isolates increased from 2% in 1975 to 35% in 1996. Since the 1980s, vancomycin has been the last uniformly effective antimicrobic available for treating serious S. aureus infections. In July 1997, intermediate-resistant S. aureus (VISA)-associated peritonitis was diagnosed in a patient who was being treated with long-term ambulatory peritoneal dialysis. The patient had been treated with intravenous vancomycin for repeated episodes of MRSA-associated peritonitis. Of six S. aureus isolates from the patient one demonstrated intermediate resistance to vancomycin. As a part of the investigation, cultures were obtained from the hands and nares of the patient's household contacts, hospital roommates, and health-care providers. Although S. aureus was isolated from 13 of 51 hand cultures and eight of 51 nares cultures, none of these cultures were positive for VISA.

The emergence of VISA in the United States suggests that S. aureus strains with full resistance to vancomycin may eventually emerge. Such resistance could result in serious clinical and public health consequences. Infections caused by less virulent coagulase-negative staphylococci (e.g., S. haemolyticus and S. epidermidis) with reduced susceptibility to vancomycin have been previously recognized.

European studies have provided compelling evidence that vancomycin-resistant enterococci, now a frequent cause of nosocomial infections, come from the (veterinary) use of antibiotics in animal feed.

MMWR 46(27):624-626, 7/11/97; MMWR 46(33):765-766, 8/22/97; MMWR 46(35):813-815, 9/5/97.


The Experiment




















Isolate a bacterium from your skin, your house, or your pet and test its susceptibility to antimicrobics to compare resistance to penicillin, methicillin, and vancomycin or to the four generations of cephalosporins. Any bacterial isolate can be used but antibiotics will have to be selected according to the gram-reaction of the isolate. Enterococci can be isolated from feces and perianal skin by swabbing with a moist swab and inoculating SF broth. An easy isolation procedure for staphylococci is to:

1) Moisten a sterile swab with sterile water and swab any surface.

2) Swab one-half of a mannitol salt agar plate. Discard the swab in disinfectant.

3) Using a sterile loop, streak back and forth perpendicular to the swabbed area a few times.

4) Incubate the plate inverted at 35°C for 24-48 hr.

Collect epidemiologic data including the source of bacterium and information about the host (e.g., Species, age, prior antibiotic exposure, clinical work experience, and for pets, whether it is an indoor or outdoor animal.

Select one isolated colony and transfer it to a tube of nutrient broth. Incubate the broth at 35°C for 24-48 hr. S. aureus are mannitol-fermenting, coagulase-positive.

Swab in three directions to ensure complete coverage of the agar. Using sterilized forceps, place antibiotic disks on the surface of the agar. Incubate the plate inverted at 35°C for 24-48 hr.

Keep the Petri plate closed and measure the zones of inhibition from the underside of the plate. (All cultures should be autoclaved before disposal.) Zones of inhibition can be interpreted from data in the antibiotic-disk package insert.

Conclusions and Discussion

1. Which antibiotic could be used to treat an infection caused by the organism you tested?

2. Combine class results to determine if there is a relationship between antibiotic-use and resistance.

3. What is the method of action of the antibiotic(s) tested?

4. Explain the mechanisms of resistance to the antibiotic(s) tested.

5. How can the spread of resistant strains be prevented?