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Table 1. Selection of Tn-seq identified genes required for adherence of M. catarrhalis BBH18.

CSV
Figure 2. Adhesion of directed gene deletion mutants to respiratory tract epithelial cells.

Adherence of aroA, ecnAB , lgt1, and the putative lipoprotein MCR_1483 to both Detroit 562 (A) and A549 (B) cells was significantly attenuated compared to wild-type (WT). Adherence levels are expressed relative to WT (n ≥ 3) and shown as means and SEM. Statistical difference was determined with a Mann-Whitney test with * P < 0.05, ** P < 0.01, and *** P < 0.001.

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Loss of the putative outer membrane lipoprotein gene MCR_1483 and aroA resulted in the most severe attenuation of adherence, to 30-39% and 34-39% of wild-type, respectively. MCR_1483 encodes a small 48 amino acid putative lipoprotein sharing 58% identity with the LppL protein of Pseudomonas sp. GM102. In M. catarrhalis BBH18 MCR_1483 is part of a putative operon together with lysA, dapF , and xerC , whereas the representative gene cluster in Pseudomonas sp. contains two additional uncharacterized ORFs [ Cheap Sale Recommend Cheap Sale Discounts Proenza Schouler Woman Printed Silktwill Dress Black Size 8 Proenza Schouler Lowest Price Cheap Price In China u7l7OwwFu
]. In Pseudomonas fluorescens WCS365 this operon, more specifically xerC encoding a site specific recombinase, is required for root colonization [ Sleeveless Top floral b by VIDA VIDA Cheap Inexpensive Best Prices Free Shipping Fashionable 39LjeI
]. Others postulated that in Pseudomonas sp . LppL is involved in regulation of the lysine biosynthesis gene lysA [ 29 ]. The attenuated adhesion of the aroA mutant is intriguing, especially since a direct role of this predicted cytoplasmic protein in cellular adhesion is unlikely. The aroA gene encodes a 3-phosphoshikimate 1-carboxyvinyltransferase that is involved in aromatic amino acid metabolism and generation of chorismate-derived metabolites [ Womens Divia Dress Strenesse Cheap Sale Explore Explore Online L3qCoLx9aa
]. Deletion of aro genes leads to attenuated virulence in many bacterial pathogens and often results in defective cell envelope biosynthesis [ 32 ]. It could be possible that a defective outer membrane function in M. catarrhalis BBH18Δ aroA caused in incorrect localization of “true” adhesins, thereby leading to attenuated adhesion. Interestingly, both MCR_1483 and aroA were also found to be required for complement resistance of M. catarrhalis (De Vries et al., unpublished data) and thus appear to fulfill or affect an essential virulence mechanism. More subtle effects on adherence efficiency were observed upon deletion of ecnAB and lgt1 , 59-78% and 64-80% of wild-type, respectively. In E. coli , ecnAB is reported to direct the biosynthesis of two small outer membrane lipoproteins that regulate programmed cell death [ How Much Sale Online Discounts Sale Online Mens Tramper Jeans Mustang Cheap Sale Store UGRoN
]. Deletion of lgt1 , which encodes an α-(1,2) glucosyltransferase involved in LOS biosynthesis [ Womens Straight Jeans GStar Order Free Shipping 2018 Unisex Quality Free Shipping Sale Supply Shopping LJlq1
], is expected to result in a truncated LOS structure. An altered LOS structure may lead to decreased membrane stability and altered surface display of adhesins as has been shown for a LOS-deficient mutant of strain O35E [ 34 ].

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