The use of flow cytometry to detect expression of subunits encoded by 11 <i>Salmonella enterica</i> serotype Typhimurium fimbrial operons

Humphries, Andrea D.; Raffatellu, Manuela; Winter, Sebastian; Weening, Eric H.; Kingsley, Robert A.; Droleskey, Robert E.; Zhang, Shuping; Figueiredo, Josely F.; Khare, Sangeeta; Nunes, Jairo; Adams, L. Garry; Tsolis, Renée M.; Bäumler, Andreas J.

doi:10.1046/j.1365-2958.2003.03507.x

Cited by 150 publications

(184 citation statements)

References 49 publications

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“…Flow cytometry was performed essentially as described (38), labeling FimH on LS76, LS60, LS68, LS154, LS141, and LSpBB using primary antibodies against FimH and FITC-conjugated secondary antibodies (see also SI Text).…”

Section: Methodsmentioning

confidence: 99%

Adaptive mutations in the signal peptide of the type 1 fimbrial adhesin of uropathogenic Escherichia coli

Ronald¹,

Yakovenko

Yazvenko³

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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Signal peptides (SPs) are critical for protein transport across cellular membranes, have a highly conserved structure, and are cleaved from the mature protein upon translocation. Here, we report that naturally occurring mutations in the SP of the adhesive, tipassociated subunit of type 1 fimbriae (FimH) are positively selected in uropathogenic Escherichia coli. On the one hand, these mutations have a detrimental effect, with reduced FimH transport across the inner membrane, fewer FimH and fimbriae expressed on the bacterial surface, and decreased bacterial adhesion under flow conditions. On the other hand, the fimbriae expressed by the mutants are significantly longer on average, with many fimbriae able to stretch to >20 m in length. More surprisingly, the SP mutant bacteria display an increased ability to resist detachment from the surface upon a switch from high to low flow. This functional effect of longer fimbriae highlights the importance of the nonadhesive fimbrial rod for adhesive function. Also, whereas bacterial adhesion to bladder epithelial cells was preserved in most mutants, binding to and killing by human neutrophils was decreased, providing an additional reason the SP mutations are relatively common among uropathogenic strains. Thus, this study demonstrates how mutations in an SP, while decreasing transport function and not affecting the final structure of the translocated protein, can lead to functional gains of the extracellular organelles that incorporate the protein and overall adaptive changes in the organism's fitness.bacterial pathogenesis ͉ fimbrial morphology ͉ protein transport ͉ shear force

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Section: Methodsmentioning

confidence: 99%

Adaptive mutations in the signal peptide of the type 1 fimbrial adhesin of uropathogenic Escherichia coli

Ronald¹,

Yakovenko

Yazvenko³

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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“…With this in mind, caution is required when extrapolating findings from in vitro models to target hosts. This is reinforced by the finding that only one of 11 S. Typhimurium fimbrial subunits was expressed during culture in vitro, compared with nine of 11 in bovine ileal loops (Humphries et al 2003).…”

Section: Molecular Basis Of Intestinal Colonization Of Food-producingmentioning

confidence: 99%

Molecular insights into farm animal and zoonoticSalmonellainfections

Stevens

Humphrey

Maskell

2009

Phil. Trans. R. Soc. B

145

102

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Salmonella enterica is a facultative intracellular pathogen of worldwide importance. Infections may present in a variety of ways, from asymptomatic colonization to inflammatory diarrhoea or typhoid fever depending on serovar-and host-specific factors. Human diarrhoeal infections are frequently acquired via the food chain and farm environment by virtue of the ability of selected non-typhoidal serovars to colonize the intestines of food-producing animals and contaminate the avian reproductive tract and egg. Colonization of reservoir hosts often occurs in the absence of clinical symptoms; however, some S. enterica serovars threaten animal health owing to their ability to cause acute enteritis or translocate from the intestines to other organs causing fever, septicaemia and abortion. Despite the availability of complete genome sequences of isolates representing several serovars, the molecular mechanisms underlying Salmonella colonization, pathogenesis and transmission in reservoir hosts remain ill-defined. Here we review current knowledge of the bacterial factors influencing colonization of food-producing animals by Salmonella and the basis of host range, differential virulence and zoonotic potential.

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“…Flow cytometry was used to detect the production of ECP by all E. coli strains studied, as described (42). Briefly, bacteria grown overnight in DMEM were incubated with anti-ECP antibodies (1:1,000) followed by goat anti-rabbit IgG Alexa fluor conjugate (Invitrogen).…”

mentioning

confidence: 99%

Commensal and pathogenic Escherichia coli use a common pilus adherence factor for epithelial cell colonization

Rendón

Saldaña

Erdem

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

262

302

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Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is a food-borne pathogen that causes hemorrhagic colitis and the hemolytic uremic syndrome. Colonization of the human gut mucosa and production of potent Shiga toxins are critical virulence traits of EHEC. Although EHEC O157:H7 contains numerous putative pili operons, their role in the colonization of the natural bovine or accidental human hosts remains largely unknown. We have identified in EHEC an adherence factor, herein called E. coli common pilus (ECP), composed of a 21-kDa pilin subunit whose amino acid sequence corresponds to the product of the yagZ (renamed ecpA) gene present in all E. coli genomes sequenced to date. ECP production was demonstrated in 121 (71.6%) of a total of 169 ecpA ؉ strains representing intestinal and extraintestinal pathogenic as well as normal flora E. coli. High-resolution ultrastructural and immunofluorescence studies demonstrated the presence of abundant peritrichous fibrillar structures emanating from the bacterial surface forming physical bridges between bacteria adhering to cultured epithelial cells. Isogenic ecpA mutants of EHEC O157:H7 or fecal commensal E. coli showed significant reduction in adherence to cultured epithelial cells. Our data suggest that ECP production is a common feature of E. coli colonizing the human gut or other host tissues. ECP is a pilus of EHEC O157:H7 with a potential role in host epithelial cell colonization and may represent a mechanism of adherence of both pathogenic and commensal E. coli.pili ͉ enterohemorrhagic Escherichia coli ͉ normal flora

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The use of flow cytometry to detect expression of subunits encoded by 11 Salmonella enterica serotype Typhimurium fimbrial operons

Cited by 150 publications

References 49 publications

Adaptive mutations in the signal peptide of the type 1 fimbrial adhesin of uropathogenic Escherichia coli

Adaptive mutations in the signal peptide of the type 1 fimbrial adhesin of uropathogenic Escherichia coli

Molecular insights into farm animal and zoonoticSalmonellainfections

Commensal and pathogenic Escherichia coli use a common pilus adherence factor for epithelial cell colonization

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