A Salmonella fim Homologue in Citrobacter freundii Mediates Invasion In Vitro and Crossing of the Blood-Brain Barrier in the Rat Pup Model

Abstract

From the invasive Citrobacter freundii strain 3009, an invasion determinant was cloned, sequenced, and expressed. Sequence analysis of the determinant showed high homology with the fim determinant from Salmonella enterica serovar Typhimurium. The genes of the invasion determinant directed invasion of recombinant Escherichia coli K-12 strains into human epithelial cell lines of the bladder and gut as well as mannose-sensitive yeast agglutination and were termed fim Cf genes. Expression of the Fim Cf proteins was shown by 35 S labeling and/or Western blotting. In the infant rat model of experimental hematogenous meningitis, C. freundii strain 3009 and the in vitro invasive recombinant E. coli K-12 strain harboring the fim Cf determinant reached the cerebrospinal fluid, in contrast to the case for the control strain. The fim determinant was also necessary for efficient in vitro invasion by C. freundii, because a deletion mutant was strongly reduced in its invasion efficiency. The mutation could be complemented in trans by the corresponding genes. Invasion by C. freundii could be blocked only by D-mannose, GlcNAc, and chitin hydrolysate and not by other carbohydrates tested. In contrast, yeast agglutination was not affected by GlcNAc or chitin hydrolysate. This finding indicated mannose residues to be essential for both yeast agglutination and invasion, whereas GlcNAc (oligomer) residues of host cells are involved exclusively in invasion. These results showed the fim determinant of C. freundii to be responsible for D-mannose-and GlcNAc-dependent in vitro invasion without being assembled into pili and for crossing of the blood-brain barrier in the infant rat model.

Multiple Low-Dose Infusions of Human Umbilical Cord Blood Cells Improve Cognitive Impairments and Reduce Amyloid-β-Associated Neuropathology in Alzheimer Mice

“…Previous work in a mouse model of stroke has shown that HUCBC infusion results in significant reduction in the infarct volume as well as rescue of neurological deficits associated with decreased proinflammatory cytokine production [19]. Additionally, we previously found reduced parenchymal and vascular b-amyloid deposits in AD mouse model.…”

“…Immunohistochemical staining was performed using mouse monoclonal anti-human Ab [17][18][19][20][21][22][23][24] antibody (clone 4G8; Covance Research Products, Emeryville, CA) or rat anti-mouse CD45 antibody (AbD serotec, Raleigh, NC) in conjunction with the VectaStain Elite ABC kit (Vector Laboratories, Burlingame, CA) coupled with diaminobenzidine substrate. For all the staining, a set of sections without adding primary antibody were used as negative staining control.…”

“…To evaluate reductions of Ab plaques in HUCBC-infused mice brains, we performed immunohistochemistry using monoclonal Ab [17][18][19][20][21][22][23][24] antibody (4G8) staining. Mouse brain sections from both 6-and 10-month HUCBC-treated PSAPP mice were stained with 4G8 antibody at 12 and 16 months of age, respectively.…”

“…Recently, BM-derived cells, especially monocytes/macrophages (MO/MØ), have been shown to be able to cross the blood-brain barrier and differentiate into functional microglia [17][18][19][20][21].…”

Evolution of the Chaperone/Usher Assembly Pathway: Fimbrial Classification Goes Greek

“…The ␥ 1 -fimbrial clade corresponds to FUP cluster 1 (367) and consists of ushers present in several bacterial species, all belonging to a single family within the Gammaproteobacteria (the Enterobacteriaceae). Representatives are present in Citrobacter freundii (FimD) (133), E. coli (AufC, ECs0594, ECs2110, FimD, FmlC, FocD, SfaF, SfmD, YcbS, and YdeT) (17,33,45,56,73,128,177,202,203,271,274,297,340,352), Proteus mirabilis (AtfC) (213,214), and S. enterica serotype Typhimurium (BcfC and FimD) (218,337). A number of usher proteins are closely related to the ␥ 1 -fimbrial clade but are not connected by a node with a good bootstrapping value (Fig.…”