Escherichia coliK1 aslAContributes to Invasion of Brain Microvascular Endothelial Cells In Vitro and In Vivo

ABSTRACTNeonatal Escherichia colimeningitis remains a devastating disease, with unacceptably high morbidity and mortality despite advances in supportive care measures and bactericidal antibiotics. To further our ability to improve the outcome of affected neonates, a better understanding of the pathogenesis of the disease is necessary. To identify potential bacterial genes which contribute toE. coliinvasion of the blood-brain barrier, a cerebrospinal fluid isolate of E. coliK1 was mutagenized with TnphoA. TnphoAmutant 27A-6 was found to have a significantly decreased ability to invade brain microvascular endothelial cells compared to the wild type. In vivo, 32% of the animals infected with mutant 27A-6 developed meningitis, compared to 82% of those infected with the parent strain, despite similar levels of bacteremia. The DNA flanking the TnphoAinsertion in 27A-6 was cloned and sequenced and determined to be homologous toE. coliK-12 aslA(arylsulfatase-like gene). The deduced amino acid sequence of the E. coliK1aslAgene product shows homology to a well-characterized arylsulfatase family of enzymes found in eukaryotes, as well as prokaryotes. Two additional aslAmutants were constructed by targeted gene disruption and internal gene deletion. Both of these mutants demonstrated decreased invasion phenotypes, similar to that of TnphoAmutant 27A-6. Complementation of the decreased-invasion phenotypes of these mutants was achieved whenaslAwas supplied in trans. This is the first demonstration that this locus contributes to invasion of the blood-brain barrier by E. coliK1.