Mathematical Modelling Highlights the Potential for Genetic Manipulation as an Adjuvant to Counter Efflux-Mediated MDR in Salmonella.

Bacteria have developed resistance to antibiotics by various mechanisms, notable amongst these is the use of permeation barriers and the expulsion of antibiotics via efflux pumps. The resistance-nodulation-division (RND) family of efflux pumps is found in Gram-negative bacteria and a major contributor to multidrug resistance (MDR). In particular, Salmonella encodes five RND efflux pump systems: AcrAB, AcrAD, AcrEF, MdsAB and MdtAB which have different substrate ranges including many antibiotics. We produce a spatial partial differential equation (PDE) model governing the diffusion and efflux of antibiotic in Salmonella, via these RND efflux pumps. Using parameter fitting techniques on experimental data, we are able to establish the behaviour of multiple wild-type and efflux mutant Salmonella strains, which enables us to produce efflux profiles for each individual efflux pump system. By combining the model with a gene regulatory network (GRN) model of efflux regulation, we simulate how the bacteria respond to their environment. Finally, performing a parameter sensitivity analysis, we look into various different targets to inhibit the efflux pumps. The model provides an in silico framework with which to test these potential adjuvants to counter MDR.

Time dependent asymptotic analysis of the gene regulatory network of the AcrAB-TolC efflux pump system in gram-negative bacteria.

Efflux pumps are a mechanism of intrinsic and evolved resistance in bacteria. If an efflux pump can expel an antibiotic so that its concentration within the cell is below a killing threshold the bacteria are resistant to the antibiotic. Efflux pumps may be specific or they may pump various different substances. This is why many efflux pumps confer multi drug resistance (MDR). In particular over expression of the AcrAB-TolC efflux pump system confers MDR in both Salmonella and Escherichia coli. We consider the complex gene regulation network that controls expression of genes central to controlling the efflux associated genes acrAB and acrEF in Salmonella. We present the first mathematical model of this gene regulatory network in the form of a system of ordinary differential equations. Using a time dependent asymptotic analysis, we examine in detail the behaviour of the efflux system on various different timescales. Asymptotic approximations of the steady states provide an analytical comparison of targets for efflux inhibition.

Robust Phagocyte Recruitment Controls the Opportunistic Fungal Pathogen Mucor circinelloides in Innate Granulomas In Vivo.

Mucormycosis is an emerging fungal infection with extremely high mortality rates in patients with defects in their innate immune response, specifically in functions mediated through phagocytes. However, we currently have a limited understanding of the molecular and cellular interactions between these innate immune effectors and mucormycete spores during the early immune response. Here, the early events of innate immune recruitment in response to infection by Mucor circinelloides spores are modeled by a combined in silico modeling approach and real-time in vivo microscopy. Phagocytes are rapidly recruited to the site of infection in a zebrafish larval model of mucormycosis. This robust early recruitment protects from disease onset in vivoIn silico analysis identified that protection is dependent on the number of phagocytes at the infection site, but not the speed of recruitment. The mathematical model highlights the role of proinflammatory signals for phagocyte recruitment and the importance of inhibition of spore germination for protection from active fungal disease. These in silico data are supported by an in vivo lack of fungal spore killing and lack of reactive oxygen burst, which together result in latent fungal infection. During this latent stage of infection, spores are controlled in innate granulomas in vivo Disease can be reactivated by immunosuppression. Together, these data represent the first in vivo real-time analysis of innate granuloma formation during the early stages of a fungal infection. The results highlight a potential latent stage during mucormycosis that should urgently be considered for clinical management of patients.IMPORTANCE Mucormycosis is a dramatic fungal infection frequently leading to the death of patients. We know little about the immune response to the fungus causing this infection, although evidence points toward defects in early immune events after infection. Here, we dissect this early immune response to infectious fungal spores. We show that specialized white blood cells (phagocytes) rapidly respond to these spores and accumulate around the fungus. However, we demonstrate that the mechanisms that enable phagocytes to kill the fungus fail, allowing for survival of spores. Instead a cluster of phagocytes resembling an early granuloma is formed around spores to control the latent infection. This study is the first detailed analysis of early granuloma formation during a fungal infection highlighting a latent stage that needs to be considered for clinical management of patients.