Fatty acids are commonly present in foods and animal blood and tissues, and can easily assimilate in the membranes of essentially all types of cells. We are examining the impact of fatty acid assimilation on opportunist pathogen bacteria, especially how they might change the pathogen life style and sensitivities to antibiotics.
Bacterial pathogens are usually tested for antibiotic or stress sensitivities when grown in laboratory medium and conditions. Our results indicate that bacterial expression and behavior is significantly altered by exogenous fatty acids (e.g., in blood, organs, or foods). This project aims to re-examine the factors that might inhibit growth of bacterial pathogens, this time in the presence of exogenous fatty acids. In this way, we expect to find physiologically relevant approaches to inhibit or slow pathogen growth.
Basic approaches include determinations of cell fatty acid composition, molecular biology/genetic screens and cell culture assays, genome sequencing, qRTPCR, physiological tests. We added screening of hospital strains while developing the project.
Results thus far establish the general capacity of Gram-positive opportunist pathogens of the streptococcus, enterococcus and staphylococcus genera to assimilate exogenous fatty acids and bypass the action of FASII-targeting antibiotics. They show that bacterial expression is reprogrammed when grown in fatty-acid-rich environments such as serum.
Bacteria isolated from hospitals have properties similar to those shown for laboratory strains.
Our results, showing that exogenous fatty acids trigger bacterial reprogramming, reveal a novel behavioral response of bacteria under physiologically relevant conditions. These results should enrich computational biology approaches aiming to accurately predict cell functions based on high throughput studies and the literature. Our studies may provide the right conditions for identifying functional bacterial inhibitors.
The development of new antimicrobial targets to overcome multi-drug-resistant bacteria has led to numerous candidate targets and drugs in recent years. The fatty acid synthesis (FASII) pathway, which is required to produce membrane fatty acids (FA), has stimulated interest as a promising target for drug development as evidenced by a multitude of publications in top-line journals. FASII pathway inhibitors were isolated from Streptomyces by extensive screenings, and gave rise to novel drugs (e.g;, platencin and platencimycin) plus synthetic derivatives, which were developed in the context of drug discovery programs designed for commercial use.
Our recent results questioned the validity of FASII pathway as antibacterial target for major low GC% Gram-positive bacterial species (streptococci, enterococci, and staphylococci) (Brinster et al, Nature 2009). We discovered that these opportunist pathogens scavenge FA, which are abundant in blood (serum contains 3g FA/liter) to constitute their membranes. In-depth studies in Streptococcus agalactiae (Group B streptococcus; GBS) and Staphylococcus aureus showed that serum FA fully overcame drug inhibition of de novo FA biosynthesis. Importantly, deletion of several genes of the GBS FASII pathway did not perturb growth when FA were present in the medium, and mutants were as virulent as the parental strain. Moreover, when grown in serum, six tested major Gram-positive pathogens escaped killing by FASII-targeted antibiotics. These results gave strong evidence that FASII pathway is not an appropriate target for antimicrobial development, particularly in treating septicemic infections.
The above published findings solicited support from scientists that had come to similar conclusions in studies of parasites. However, they also raised opposition from companies that were developing FASII-targeting drugs aimed at eliminating S. aureus. To assuage any doubts concerning the applicability of our findings, we performed detailed experiments with S. aureus that validate the generality of our findings. We showed that while FASII-targeted drugs reach their targets, they do not stop staphylococcal growth in serum; moreover, we successfully deleted the gene encoding FabI, the main S. aureus candidate target of FASII inhibitors (Brinster et al, Nature Commentary in press). These findings raise major questions on a large field of research and costly investment on antimicrobial drug development based on targeting the FASII biosynthesis pathway.
We hypothesize that the inherent property of bacteria to modify their membrane composition according to environmental FA availability has far-reaching consequences on bacterial behavior in vitro and during infection. This project aims to study the impact of blood FAs on the in vitro and in vivo life styles of major Gram-positive opportunist pathogens using two main models. 1) S. agalactiae, a major cause of septicemia in newborns and an emerging cause of infection in immuno-compromised adults. 2) S.aureus, a main cause of nosocomial-associated infections, which is exacerbated by the widespread emergence of multiresistance to antibiotics. Some studies will be validated on major human streptococcal pathogens S. pyogenes and S. pneumoniae, which cause a wide range of pathologies, and Enterococcus faecalis, a major agent of nosocomial infections.
Our goals are to determine:
1) the mechanisms and localization of FA assimilation,
2) the impact of FA incorporation in natural isolates and fab mutants on genome expression and bacterial sensitivity to antibiotics and in vitro and in vivo stress,
3) the physical changes due to FA incorporation in bacterial membranes, and consequences on membrane fluidity, biofilms, adherence to host cells, and in vivo localization,
4) the impact of saturated FA (SFA)-rich versus unsaturated FA (UFA)-rich diet on bacterial infectivity.
This project aims to provide leads on strategies for treatment of infections.
Madame Alexandra GRUSS (INSTITUT NATIONAL DE LA RECHERCHE AGRONOMIQUE - CENTRE DE RECHERCHE DE JOUY-EN-JOSAS) – firstname.lastname@example.org
The author of this summary is the project coordinator, who is responsible for the content of this summary. The ANR declines any responsibility as for its contents.
IP INSTITUT PASTEUR
INRA INSTITUT NATIONAL DE LA RECHERCHE AGRONOMIQUE - CENTRE DE RECHERCHE DE JOUY-EN-JOSAS
INSERM INSTITUT NATIONAL DE LA SANTE ET DE LA RECHERCHE MEDICALE - DELEGATION DE PARIS V
Help of the ANR 450,000 euros
Beginning and duration of the scientific project: - 36 Months