No abstract

This study evaluated the bioactivity of 3D-printed β-tricalcium phosphate (β-TCP) scaffolds or hydroxyapatite (HA) scaffolds associated with polydioxanone (PDO) membrane (Plenum® Guide) for guided bone regeneration in rats. Fifty-four rats were divided into three groups (n = 18 animals): autogenous bone + PDO membrane (Auto/PG); 3D-printed β-TCP + PDO membrane (TCP/PG); and 3D-printed HA + PDO membrane (HA/PG). A surgical defect in the parietal bone was made and filled with the respective scaffolds and PDO membrane. The animals were euthanized 7, 30, and 60 days after the surgical procedure for micro-CT, histomorphometric, and immunolabeling analyses. Micro-CT showed an increase in trabecular thickness and a decrease in trabecular separation, even with similar bone volume percentages between TCP/PG and HA/PG vs. Auto/PG. Histometric analysis showed increased bone formation at 30 days in the groups compared to 7 days postoperatively. Immunolabeling analysis showed an increase in proteins related to bone formation at 30 days, and both groups showed a similar immunolabeling pattern. This study concludes that 3D-printed scaffolds associated with PDO membrane (Plenum® Guide) present similar results to autogenous bone for bone regeneration.

bThe extraintestinal pathogen termed avian pathogenic Escherichia coli (APEC) is known to cause colibacillosis in chickens. The molecular basis of APEC pathogenesis is not fully elucidated yet. In this work, we deleted a component of the Yad gene cluster (yadC) in order to understand the role of Yad in the pathogenicity of the APEC strain SCI-07. In vitro, the transcription level of yadC was upregulated at 41°C and downregulated at 22°C. The yadC expression in vivo was more pronounced in lungs than in spleen, suggesting a role in the early steps of the infection. Chicks infected with the wild-type and mutant strains presented, respectively, 80% and 50% mortality rates. The ⌬yadC strain presented a slightly decreased ability to adhere to HeLa cells with or without the D-mannose analog compared with the wild type. Real-time PCR (RT-PCR) assays showed that fimH was downregulated (P < 0.05) and csgA and ecpA were slightly upregulated in the mutant strain, showing that yadC modulates expression of other fimbriae. Bacterial internalization studies showed that the ⌬yadC strain had a lower number of intracellular bacteria recovered from Hep-2 cells and HD11 cells than the wild-type strain (P < 0.05). Motility assays in soft agar demonstrated that the ⌬yadC strain was less motile than the wild type (P < 0.01). Curiously, flagellum-associated genes were not dramatically downregulated in the ⌬yadC strain. Taken together, the results show that the fimbrial adhesin Yad contributes to the pathogenicity and modulates different biological characteristics of the APEC strain SCI-07. Avian pathogenic Escherichia coli (APEC) strains cause a variety of extraintestinal infections in poultry collectively known as colibacillosis (1, 2). Although the complete mechanisms of APEC pathogenicity are not fully elucidated, it is believed that colibacillosis starts with colonization of the host's upper respiratory tract, with the bacterium expressing one or more colonization factors known as adhesins (3). The infection can subsequently spread into the lungs and other inner organs, leading to a fatal septicemia (1,4,5). Among the known APEC adhesins, those more studied are the type 1 fimbriae, P fimbriae, and curli fimbriae, respectively (6).Besides their roles in the adhesion and subsequent colonization of environmental surfaces, those fimbriae are assumed to be essential for the establishment of a host-parasite relationship and further disease progression (7). Both type 1 and curli fimbriae play important roles in the initial bacterial colonization of the respiratory epithelium (1, 8), while P fimbriae (9) are important for later stages of infection (1).In addition to the fimbriae mentioned above, E. coli possesses several other fimbrial operons (i.e., Yad, Ycb, Ybg, Yfc, Yra, Sfm, Ygi, and Yeh) that display sequence and organizational homologies to type 1 fimbria and could contribute to E. coli's ability to adhere to and colonize the host epithelia (10, 11).The adhesion operon Yad is composed of seven genes, yadN (major subunit), ecpD (usher...

BackgroundAvian pathogenic Escherichia coli strains cause extraintestinal diseases in birds, leading to substantial economic losses to the poultry industry worldwide. Bacteria that invade cells can overcome the host humoral immune response, resulting in a higher pathogenicity potential. Invasins are members of a large family of outer membrane proteins that allow pathogen invasion into host cells by interacting with specific receptors on the cell surface.ResultsAn in silico analysis of the genome of a septicemic APEC strain (SEPT362) demonstrated the presence of a putative invasin homologous to the ychO gene from E. coli str. K-12 substr. MG1655. In vitro and in vivo assays comparing a mutant strain carrying a null mutation of this gene, a complemented strain, and its counterpart wild-type strain showed that ychO plays a role in the pathogenicity of APEC strain SEPT362. In vitro assays demonstrated that the mutant strain exhibited significant decreases in bacterial adhesiveness and invasiveness in chicken cells and biofilm formation. In vivo assay indicated a decrease in pathogenicity of the mutant strain. Moreover, transcriptome analysis demonstrated that the ychO deletion affected the expression of 426 genes. Among the altered genes, 93.66 % were downregulated in the mutant, including membrane proteins and metabolism genes.ConclusionThe results led us to propose that gene ychO contributes to the pathogenicity of APEC strain SEPT362 influencing, in a pleiotropic manner, many biological characteristics, such as adhesion and invasion of in vitro cultured cells, biofilm formation and motility, which could be due to the possible membrane location of this protein. All of these results suggest that the absence of gene ychO would influence the virulence of the APEC strain herein studied.

Avian Pathogenic Escherichia coli is responsible for significant economic losses in the poultry industry by causing a range of systemic or localized diseases collectively termed colibacillosis. The virulence mechanisms of these strains that are pathogenic in poultry and possibly pathogenic in humans have not yet been fully elucidated. This work was developed to study if over-expressed genes in a microarray assay could be potentially involved in the pathogenicity of an Avian Pathogenic Escherichia coli strain isolated from a swollen head syndrome case. For this study, five over-expressed genes were selected for the construction of null mutants [flgE (flagellar hook), tyrR (transcriptional regulator), potF (putrescine transporter), yehD (putative adhesin) and bfr (bacterioferritin)]. The constructed mutants were evaluated for their capacity for the adhesion and invasion of in vitro cultured cells, their motility capacity, and their pathogenic potential in one-day-old chickens compared with the wild-type strain (WT). The Δbfr strain showed a decreased adhesion capacity on avian fibroblasts compared with WT, in the presence and absence of alpha-D-mannopyranoside, and the ΔpotF strain showed decreased adhesion only in the absence of alpha-D-mannopyranoside. The ΔtyrR mutant had a reduced ability to invade Hep-2 cells. No mutant showed changes in invading CEC-32 cells. The mutants ΔflgE and ΔtyrR showed a decreased ability to survive in HD-11 cells. The motility of the mutant strains Δbfr, ΔyehD and ΔpotF was increased, while the ΔtyrR mutant showed reduction, and the ΔflgE became non-motile. No mutant strain caused the same mortality of the WT in one-day-old chickens, showing attenuation to different degrees.