New antibiotics that may offer additional and much needed treatment options will only be used in hospital systems if the clinical microbiology laboratory can provide timely antimicrobial susceptibility testing data. Historically there has been a time lag in the availability of susceptibility testing methods either at reference laboratories (long delay to results), incorporation in automated commercial systems (4 years), or simpler methods like disk diffusion and gradient strips that can be performed manually on an as needed basis. I was excited to learn that delafloxacin and meropenem/vaborbactam disk diffusion and gradient strip methods finally became available. Fantastic. However, before we can introduce those methods in the clinical laboratory, we need to validate performance of these methods per CLIA '88 regulations and good laboratory practice. This requires either comparing the new methods to a reference standard (broth microdilution -- need antibiotic powder, and a lot of set up time) or a set of strains that has been previously characterized by a reference dilution method and has a good representation of susceptible and resistant isolates. Those are serious roadblocks. By chance, I happened to give a talk at the Northeast Branch of the American Society of Microbiology and a someone from Melinta Pharmaceuticals happened to be there, and that someone referred me to our local Key Account Manager who offerred a solution "on request". Specifically, Melinta or other pharmaceutical companies are not allowed to approach me and tell me about a solution, but if I inquire independently and ask for a previously characterized set of bacterial isolates, they are allowed to tell me that in fact they have a series of previously characterized isolates for both drugs available for validation. These isolates are provided free of charge from Laboratory Specialist, Inc, Sent by FEDEX with first class documentation, data for broth microdilution performed circa ten times on each isolates with modal MICs and MIC distribution - wow, that is awesome. Methods validated beautifully. It just seems odd to me that such a valuable resource, providing clinical laboratories the ability to robustly evaluate AST methods for newly marketed drugs, needs to remain on a need to know basis. Clinical labs, please take note of this available resource. The same proved to be true for plazomicin. Achaogen uses the same Laboratory Specialists, Inc. to provide 30 characterized isolates on request; however, the FEDEX chargers are not absorbed. The FDA-CDC Biobank also now has a set of isolates characterized for plazomicin susceptibility and I see now also delafloxacin., I suspect the exact same set of organisms. The existing rule restricting such company-clinical laboratory communication about these important resources should be relaxed! We need facile access to validation strains sets to bring new testing into our laboratories in a timely fashion, to facilitate the availability of new antibiotics when they are most needed, and to help support profitability of antibiotic development by pharmaceutical companies.
JoVE manuscript accepted on use of Inkjet Printing to perform antimicrobial susceptibility testing for single and drug combinations and follow up time-kill study methodology
Postdoctoral fellow, Thea Brennan-Krohn, recently had a manuscript accepted in the Journal of Visualized Experimentation, aka JoVE. The title of the manuscript and link to the abstract are "Antimicrobial Synergy Testing by the Inkjet Printer-Assisted Automated Checkerboard Array and the Manual Time-Kill Method." We have been fielding a lot of questions over the past two years about implementation of inkjet printing antimicrobial susceptibility testing technology and thought it would be useful to share a video of the technique as well as classic time-kill analysis to analyze antimicrobial synergy. We are excited to learn that the CDC has decided to implement the technology in the near future in their Antimicrobial Resistance Laboratory Network (ARLN), initially to test, the combination of ceftazidime-avibactam and aztreonam for activity against multidrug-resistant Gram-negatives.
KP Smith Speaking tomorrow at Harvard Antimicrobial Resistance Laboratory Network as well as Broad Institute collaborator, Alejandro Pironti
BIDMC news release on Thea's antimicrobial synergy paper:
"Bacteria—especially Gram-negative strains—are becoming increasingly resistant to current antibiotic drugs, and the development of new classes of antibiotics has slowed. Faced with these challenges, investigators are studying the potential of combination therapy, in which two or more drugs are used together to increase or restore the efficacy of both drugs against a resistant bacterial pathogen. Now new research indicates that such synergy may work even when bacteria become resistant to colistin, which is considered a treatment agent of last resort.
The findings are especially promising because recent evidence indicates the potential for rapid worldwide spread of colistin resistance. “For an infected patient, if the multidrug-resistant Gram-negative bacterial pathogen is resistant to colistin, then there is a big problem,” said senior author James Kirby, MD, Director of the Clinical Microbiology Laboratory at BIDMC.
In their Antimicrobial Agents and Chemotherapy study, Kirby and his colleagues Thea Brennan-Krohn, MD and Alejandro Pironti, PhD screened 19 different antibiotics for synergy with colistin. The team discovered several combinations where synergy was present and infections with resistant pathogens could potentially be treated with the combination therapy.
Of particular interest, colistin demonstrated high rates of synergy with linezolid, fusidic acid, and clindamycin, which are protein synthesis inhibitor antibiotics that individually have no activity against Gram-negative bacteria. “It was remarkable to see two drugs, each of which is inactive on its own against these bacteria, inhibiting them in combination,” notes Brennan-Krohn. “These findings suggest that colistin retains sub-lethal activity against colistin-resistant bacteria, which may enable drugs like linezolid to reach their targets.”
“Faced with highly resistant pathogens, clinicians often currently treat with multiple antibiotics without knowing the benefit the combinations may provide,” said Kirby. “This study now provides some scientific underpinning for these choices and direction for future investigation.” He added that combination therapy may also allow clinicians to use lower effective doses of colistin and other drugs, which would help avoid toxicities associated with the medications as well as slow the development of antibiotic resistance.
This work was funded in part with Federal funds from the National Institute of Allergy and Infectious Diseases, the National Institutes of Health, and the Department of Health and Human Services."
"A Little Change Can Make a Lot of MIC Difference: the Inoculum Effect and Antibiotic Susceptibility Testing"
Thanks to American Society of Microbiology science writer, Julie Wolf, for highlighting our inoculum effect manuscript in the ASM mBiosphere Blog, "A Little Change Can Make a Lot of MIC Difference: the Inoculum Effect and Antibiotic Susceptibility Testing"
Now live on the Antimicrobial Agents and Chemotherapy Webpage: The Inoculum Effect in the Era of Multidrug Resistance: Minor Differences in Inoculum Have Dramatic Effect on MIC Determination.
Our inoculum effect manuscript in the journal Antimicrobial Agents and Chemotherapy highlighted in BIDMC news release.
"The Inoculum Effect in the Era of Multidrug Resistance: Minor Differences in Inoculum Have Dramatic Effect on Minimal Inhibitory Concentration Determination."
The manuscript describes use of D300-based inkjet printing technology to investigate the inoculum effect with a resolution not previously possible. The inoculum effect is the general observation that the minimal inhibitor concentration (in other words level of resistance) of an organism to an antibiotic increases when a higher density of organisms is tested. This is effect is especially prominent for beta-lactam antiibiotics. It is of potential clinical concern during some types of infections when the organism burden is high. Here we explored whether subtle differences in inoculum within the range allowed by current standards can effect the susceptibility testing results that clinical laboratories obtain and provide to clinicians. Our findings for organisms with certain types of multidrug-resistance and very important classes of antibiotics was that these small allowable differences in inoculum could change the MIC determinations and the determination of whether organisms were susceptible or resistant to the antibiotics tested.
KP Smith wins an "Outstanding Abstract Award" otherwise known as the OAA, for his experimental work on the inoculum effect.
The award description on the ASM Microbe 2018 awards website: "Sponsored by ASM and determined by the ASM Microbe Program Committee, these awards highlight outstanding abstracts presented by students, residents, or medical/clinical fellows. All abstracts submitted by the deadline will be considered for these awards. Awards will include a cash prize of $200."
The laboratory has been fortunate to participate in Harvard Catalyst Reactor Program through the "Big Ideas, Small Features” Pilot Grant Award. A description of the program was recently published last week in Harvard Medical School news titled "Catalyzing Research Innovation."
A description of the supported work can be found at: "Reactor Program Awards Eight New Pilot Grants: Supports researchers with novel solutions to major clinical challenges."
Kirby Lab Blog