Loss of SigB in Listeria monocytogenes strains EGD-e and 10403S leads to hypersensitivity to hydrogen peroxide in stationary phase under aerobic conditions

SigB is the main stress gene regulator in L. monocytogenes affecting the expression of more than 150 genes and thus contributing in multiple stress resistance. Despite its clear role in most stresses, its role in oxidative stress is uncertain as results accompanying the loss of sigB range from hyperresistance to hypersensitivity. Previously, these differences have been attributed to strain variation. In this study, we show conclusively that in contrast to all other stresses, loss of sigB results in hyperresistance against H2O2 (more than 8 log CFU ml-1 compared to the wild type) in aerobically-grown stationary phase cultures of 10403S and EGD-e.. Furthermore, growth at 30°C resulted in higher resistance to oxidative stress than at 37°C. Oxidative stress resistance seemed to be higher with higher levels of oxygen. Under anaerobic conditions, loss of SigB in 10403S did not affect survival against H2O2 while in EGD-e it resulted in a sensitive phenotype. During exponential phase, minor differences occurred as expected due to the absence of sigB transcription. Catalase tests were performed under all conditions and stronger catalase results corresponded well with higher survival underpinning the important role of catalase in this phenotype. Furthermore, we assessed the catalase activity in protein lysates which corresponded with the catalase tests and survival. In addition, RT-PCR showed no differences in transcription between the wild type and the ΔsigB in various oxidative stress genes. Further investigation of the molecular mechanism behind this phenotype and its possible consequences for the overall phenotype of L. monocytogenes are underway

Loss of sigb in listeria monocytogenes strains egd-e and 10403s confers hyperresistance to hydrogen peroxide in stationary phase under aerobic conditions

SigB is the main stress gene regulator in Listeria monocytogenes affecting the expression of more than 150 genes and thus contributing to multiple-stress resistance. Despite its clear role in most stresses, its role in oxidative stress is uncertain, as results accompanying the loss of sigB range from hyperresistance to hypersensitivity. Previously, these differences have been attributed to strain variation. In this study, we show conclusively that unlike for all other stresses, loss of sigB results in hyperresistance to H2O2 (more than 8 log CFU ml(-1) compared to the wild type) in aerobically grown stationary-phase cultures of L. monocytogenes strains 10403S and EGD-e. Furthermore, growth at 30 degrees C resulted in higher resistance to oxidative stress than that at 37 degrees C. Oxidative stress resistance seemed to be higher with higher levels of oxygen. Under anaerobic conditions, the loss of SigB in 10403S did not affect survival against H2O2, while in EGD-e, it resulted in a sensitive phenotype. During exponential phase, minor differences occurred, and this result was expected due to the absence of sigB transcription. Catalase tests were performed under all conditions, and stronger catalase results corresponded well with a higher survival rate, underpinning the important role of catalase in this phenotype. Furthermore, we assessed the catalase activity in protein lysates, which corresponded with the catalase tests and survival. In addition, reverse transcription-PCR (RT-PCR) showed no differences in transcription between the wild type and the Delta sigB mutant in various oxidative stress genes. Further investigation of the molecular mechanism behind this phenotype and its possible consequences for the overall phenotype of L. monocytogenes are under way. IMPORTANCE SigB is the most important stress gene regulator in L. monocytogenes and other Gram-positive bacteria. Its increased expression during stationary phase results in resistance to multiple stresses. However, despite its important role in general stress resistance, its expression is detrimental for the cell in the presence of oxidative stress, as it promotes hypersensitivity against hydrogen peroxide. This peculiar phenotype is an important element of the physiology of L. monocytogenes, and it might help us explain the behavior of this organism in environments where oxidative stress is present

A novel genome editing platform for drug resistant Acinetobacter baumannii revealed an AdeR-unrelated tigecycline resistance mechanism

Infections with the Gram-negative coccobacillus Acinetobacter baumannii are a major threat in hospital settings. The progressing emergence of multidrug resistant clinical strains significantly reduces the treatment options for clinicians to fight A. baumannii infections. The current lack of robust methods to genetically manipulate drug resistant A. baumannii isolates impedes research on resistance and virulence mechanisms in clinically relevant strains. In this study we developed a highly efficient and versatile genome editing platform enabling the markerless modification of the genome of A. baumannii clinical and laboratory strains, regardless of their resistance profile.We applied this method for the deletion of AdeR, a transcription factor that regulates the expression of the AdeABC efflux pump in tigecycline resistant A. baumannii, to evaluate its function as a putative drug target. Loss of adeR reduced the MIC90 of tigecycline from 25 μg/ml in the parental strains to 3.1 μg/ml in the ΔadeR mutants indicating its importance in the drug resistant phenotype. However, 60% of the clinical isolates remained non-susceptible to tigecycline after adeR deletion. Evolution of artificial tigecycline resistance in two strains followed by whole genome sequencing revealed loss of function mutations in trm, suggesting its role in an alternative AdeABC-independent tigecycline resistance mechanism. This finding was strengthened by the confirmation of trm disruption in the majority of the tigecycline resistant clinical isolates. This study highlights the development and application of a powerful genome editing platform for A. baumannii enabling future research on drug resistance and virulence pathways in clinical relevant strains

Revista Temas Agrarios Volumen 26; Suplemento 1 de 2021

1st International and 2nd National Symposium of Agronomic Sciences: The rebirth of the scientific discussion space for the Colombian Agro.1 Simposio Intenacional y 2 Nacional de Ciencias Agronómicas: El renacer del espacio de discusión científica para el Agro colombiano