Bacteria of the genus Enterobacter are amongst essentially the most dangerous bacteria related to hospital infections worldwide. A few of their representatives are highly proof against commonly used antibiotics, in order that the reserve antibiotic colistin is increasingly used as a final resort therapy option. To avoid unnecessary reliance on colistin and risk increasing resistance, bacteria are tested for sensitivity or resistance to colistin before recommending treatment. Nonetheless, commonly used tests for Enterobacter aren’t reliable. In a comprehensive study with broad participation inside the German Center for Infection Research (DZIF), scientists have now resolved this problem. Based on these findings, they developed a straightforward, sensitive and robust test for the genus Enterobacter, which now enables targeted antibiotic therapy for the assorted Enterobacter species.

The currently used microbiological tests for colistin resistance and other antibiotic resistances didn’t allow accurate conclusions regarding the spread of resistance in several Enterobacter species. This was partly since the taxonomic classification of clinical Enterobacter isolates was imprecise, and partly since the error rate in determining resistance was high. The massive-scale DZIF study has now achieved a breakthrough and clarified the relationships between the various Enterobacter species in addition to optimized resistance testing. To this end, researchers on the Institute of Medical Microbiology of Justus Liebig University Giessen (Deep-iAMR project), along with DZIF scientists on the Research Center Borstel Leibniz Lung Center, analyzed Enterobacter isolates collected at German university hospitals over a period of three years.

Using genome-based taxonomic studies, Enterobacter xiangfangensis was found to be essentially the most continuously occurring species in German hospitals: From an information pool of over 3246 isolates worldwide-; representing a group from over twenty years-;this species accounted for 68.7 percent of all Enterobacter detected.

Determination of antibiotic resistance profiles using phenotypic assays advisable by the European Committee on Antimicrobial Susceptibility Testing (EUCAST) yielded mixed results with regard to colistin resistance.

“It turned out that many isolates were either not or barely resistant in these tests, though the bacteria carried all of the genes vital for the expression of colistin resistance,” explains the study’s first creator, Dr. Swapnil Doijad, of Friedrich Schiller University, Jena, Germany. A result that raised questions. The researchers obtained an initial answer by further investigating the isolates wherein resistance was not clearly detectable using mass spectrometry.

“Depending on the actual Enterobacter species, we detected low levels of modified lipid A, the anchor structure of lipopolysaccharides (LPS)-;a vital component of the bacterial membrane and required for colistin resistance-;even from bacteria grown in colistin-free medium,” comments the co-first creator of the study, Dr. Nicolas Gisch of the Research Center Borstel Leibniz Lung Center. “These modifications of lipid A seem like depending on the bacterial species and are inherent, meaning their expression is embedded in a more complex regulation and never alone triggered by colistin,” he adds.

The result suggests that there’s species-dependent variation within the heteroresistance seen in Enterobacter: In routine test systems, the bacteria are sometimes resistant, sometimes not.”

Dr. Can Imirzalioglu, co-author of the study and Acting Director for Clinical Microbiology and Diagnostics on the Institute of Medical Microbiology of Justus Liebig University Giessen

Using further sophisticated methods, the authors were in a position to elucidate the phenomenon of heteroresistance within the genus Enterobacter. “Our analyses revealed that these bacteria have a sensor on their surface that responds to the pH value, i.e., the acidity within the environment, and regulates accordingly, either up or down, the genes required for the expression of colistin resistance,” explains Prof. Trinad Chakraborty, senior creator and former director of the Institute of Medical Microbiology at Justus Liebig University Giessen. Genetic variations and interactions on this sensing pathway for environmental pH led to species-dependent differences regarding the extent of colistin resistance in the assorted Enterobacter species in conventional test systems.

Based on these findings, the researchers developed a straightforward recent assay that eliminates heteroresistance effects and allows unambiguous and reliable determination of the true levels of colistin resistance for any isolate. The assay will avoid therapeutic failures when recommending the antibiotic, thereby paving the way in which for a targeted and economical treatment of Enterobacter species with this reserve antibiotic worldwide. A result that has come about not least through translational collaborations inside the DZIF.

Source:

German Center for Infection Research

Journal reference:

Doijad, S.P., et al. (2023) Resolving colistin resistance and heteroresistance in Enterobacter species. Nature Communications. doi.org/10.1038/s41467-022-35717-0.