Abstract
In some Gram-negative bacteria, ompF encodes outer membrane protein F (OmpF), which is a cation-selective porin and is responsible for the passive transport of small molecules across the outer membrane. However, there are few reports about the functions of this gene in Cronobacter sakazakii. To investigate the role of ompF in detail, an ompF disruption strain (ΔompF) and a complementation strain (cpompF) were successfully obtained. We find that OmpF can affect the ability of biofilm formation in C. sakazakii. In addition, the variations in biofilm composition of C. sakazakii were examined using Raman spectroscopy analyses caused by knocking out ompF, and the result indicated that the levels of certain biofilm components, including lipopolysaccharide (LPS), were significantly decreased in the mutant (ΔompF). Then, SDS-PAGE was used to further analyze the LPS content, and the result showed that the LPS levels were significantly reduced in the absence of ompF. Therefore, we conclude that OmpF affects biofilm formation in C. sakazakii by reducing the amount of LPS. Furthermore, the ΔompF mutant showed decreased (2.7-fold) adhesion to and invasion of HCT-8 cells. In an antibiotic susceptibility analysis, the ΔompF mutant showed significantly smaller inhibition zones than the WT, indicating that OmpF had a positive effect on the influx of antibiotics into the cells. In summary, ompF plays a positive regulatory role in the biofilm formation and adhesion/invasion, which is achieved by regulating the amount of LPS, but is a negative regulator of antibiotic resistance in C. sakazakii.
Highlights
Cronobacter sakazakii is an opportunistic food-borne pathogen that is associated with outbreaks of life-threatening bacteremia, meningitis and necrotizing enterocolitis (NEC) in neonates and infants, with case fatality rates reported to be as high as 40–80% and survivors frequently left with severe neurological and developmental disorders [1,2]
The morphological characteristics of the wild_type, ∆ompF and cpompF strains were examined by SEM, and the results showed that the three strains exhibited similar morphologies (Figure 2b)
These results indicated that knocking out the ompF gene has no effect on the growth and morphology of C. sakazakii in
Summary
Cronobacter sakazakii is an opportunistic food-borne pathogen that is associated with outbreaks of life-threatening bacteremia, meningitis and necrotizing enterocolitis (NEC) in neonates and infants, with case fatality rates reported to be as high as 40–80% and survivors frequently left with severe neurological and developmental disorders [1,2]. While causing newborn disease, C. sakazaki infects adults. C. sakazakii CC4 and C. sakazakii ST12 have been recognized as specific pathovars associated with particular neonatal and adult infections [3,4]. C. sakazakii strains have the capacity to invade and translocate through the Caco-2 and human brain microvascular endothelial cell lines [8]. These organisms can form biofilms, which can adhere to substrate surfaces, survive in the presence of antibiotics and disinfectants and enhance the resistance of cells to environmental stress [9].
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