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Cronobacter species are linked with life-treating diseases in neonates and show strong tolerances to environmental stress. However, the information about factors involved in oxidative tolerance in Cronobacter remains elusive. Here, factors involved in oxidative tolerance in C. malonaticus were identified using a transposon mutagenesis. Eight mutants were successfully screened based on a comparison of the growth of strains from mutant library (n = 215) and wild type (WT) strain under 1.0 mM H2O2. Mutating sites including thioredoxin 2, glutaredoxin 3, pantothenate kinase, serine/threonine protein kinase, pyruvate kinase, phospholipase A, ferrous iron transport protein A, and alanine racemase 2 were successfully identified by arbitrary PCR and sequencing alignment. Furthermore, the comparison about quantity and structure of biofilms formation among eight mutants and WT was determined using crystal violet staining (CVS), scanning electron microscopy (SEM), and confocal laser scanning microscopy (CLSM). Results showed that the biofilms of eight mutants significantly decreased within 48 h compared to that of WT, suggesting that mutating genes play important roles in biofilm formation under oxidative stress. The findings provide valuable information for deeply understanding molecular mechanism about oxidative tolerance of C. malonaticus.

Cronobacter species are linked with life-treating diseases in neonates and show strong tolerances to environmental stress. However, the information about factors involved in oxidative tolerance in Cronobacter remains elusive. Here, factors involved in oxidative tolerance in C. malonaticus were identified using a transposon mutagenesis. Eight mutants were successfully screened based on a comparison of the growth of strains from mutant library (n = 215) and wild type (WT) strain under 1.0 mM H2O2. Mutating sites including thioredoxin 2, glutaredoxin 3, pantothenate kinase, serine/threonine protein kinase, pyruvate kinase, phospholipase A, ferrous iron transport protein A, and alanine racemase 2 were successfully identified by arbitrary PCR and sequencing alignment. Furthermore, the comparison about quantity and structure of biofilms formation among eight mutants and WT was determined using crystal violet staining (CVS), scanning electron microscopy (SEM), and confocal laser scanning microscopy (CLSM). Results showed that the biofilms of eight mutants significantly decreased within 48 h compared to that of WT, suggesting that mutating genes play important roles in biofilm formation under oxidative stress. The findings provide valuable information for deeply understanding molecular mechanism about oxidative tolerance of C. malonaticus.

Microbial metal reduction is a process by which microorganisms are able to deliver electrons to a solid phase that is external to the cell, this reduction plays an important role in bioremediation approaches. The gram-negative bacterium Shewanella oneidensis is the focus of this work; it is known that when oxygen becomes limiting, the expression of the outer membrane cytochrome MtrC and OmcA (needed for extracellular electron transfer) increases, however, our understanding of how this organism is able to senses the change in oxygen concentration into the environment remains unclear. The aim of this project, therefore, is to identify the redox sensor in S. oneidensis. The approach used is to screen a large number of mutants, which consists on a strain (KM001) containing mtrC-lacZ fusion in which the transposon Himar is inserted into the genome, and to find the ones with reduced b-galactosidase activity (compared to KM001) under anoxic conditions. For each mutant which meets the criteria, the transposon insertion site will be identified using arbitrary PCR, while batch cultures under anoxic and under aerobic conditions will be performed to confirm reduced activity. No relevant results were recorded relating to genes influencing the expression of MtrC, however, some interesting behaviours were detected on genes encoding for proteins AroE and AroB, which seems to be important for both aerobic and anaerobic growth.