Knowledge, Attitudes, and Beliefs Regarding Antimicrobial Therapy and Resistance Among Physicians in Alexandria University Teaching Hospitals and the Associated Prescription Habits.

Irresponsible prescription of antimicrobials (AMs) is the driving factor for the growing antimicrobial resistance (AMR) crisis. In this study, we assessed the knowledge, attitudes, perceptions, and beliefs regarding AMs and AMR together with the prescription habits of physicians in three University hospitals in Alexandria, Egypt. A 40-question survey was used. Physicians were stratified into residents and practicing staff members, and further into various departments. Clinical pharmacists at the University main hospital were included for comparative purposes. A total of 319 questionnaires were completed (response rate = 91.4%). Participants demonstrated fair average knowledge about AMs (4.71 ± 1.29 out of 7), with no significant difference between residents and staff members, whereas clinical pharmacists scored significantly higher on knowledge questions (p < 0.005). Participants showed poor awareness regarding local AMR patterns of Klebsiella pneumoniae and Pseudomonas aeruginosa (13% and 23%, respectively). AMR was perceived as a global (95%), national (97%), and local (85%) problem. High confidence regarding use of AMs was noticed with significantly higher levels among staff members (70.3% vs. 86.7%, p < 0.05). Most participants agreed that the patients' demands (78.5%) and socioeconomic statuses (76.3%) do influence their choices. The most significant knowledge deficit was regarding dosage adjustment in renal patients, and the survey highlighted poor engagement in educational activities, limited awareness of local resistance patterns, and neglect in explaining the side-effects to patients. Patients' demands and socioeconomic statuses were also shown to influence the physicians' decisions.

Binding of the Fap2 protein of Fusobacterium nucleatum to human inhibitory receptor TIGIT protects tumors from immune cell attack.

Bacteria, such as Fusobacterium nucleatum, are present in the tumor microenvironment. However, the immunological consequences of intra-tumoral bacteria remain unclear. Here, we have shown that natural killer (NK) cell killing of various tumors is inhibited in the presence of various F. nucleatum strains. Our data support that this F. nucleatum-mediated inhibition is mediated by human, but not by mouse TIGIT, an inhibitory receptor present on all human NK cells and on various T cells. Using a library of F. nucleatum mutants, we found that the Fap2 protein of F. nucleatum directly interacted with TIGIT, leading to the inhibition of NK cell cytotoxicity. We have further demonstrated that tumor-infiltrating lymphocytes expressed TIGIT and that T cell activities were also inhibited by F. nucleatum via Fap2. Our results identify a bacterium-dependent, tumor-immune evasion mechanism in which tumors exploit the Fap2 protein of F. nucleatum to inhibit immune cell activity via TIGIT.

Identification and characterization of fusolisin, the Fusobacterium nucleatum autotransporter serine protease.

Fusobacterium nucleatum is an oral anaerobe associated with periodontal disease, adverse pregnancy outcomes and colorectal carcinoma. A serine endopeptidase of 61-65 kDa capable of damaging host tissue and of inactivating immune effectors was detected previously in F. nucleatum. Here we describe the identification of this serine protease, named fusolisin, in three oral F. nucleatum sub-species. Gel zymogram revealed fusobacterial proteolytic activity with molecular masses ranging from 55-101 kDa. All of the detected proteases were inhibited by the serine protease inhibitor PMSF. analysis revealed that all of the detected proteases are encoded by genes encoding an open reading frame (ORF) with a calculated mass of approximately 115 kDa. Bioinformatics analysis of the identified ORFs demonstrated that they consist of three domains characteristic of autotransporters of the type Va secretion system. Our results suggest that the F. nucleatum fusolisins are derived from a precursor of approximately 115 kDa. After crossing the cytoplasmic membrane and cleavage of the leader sequence, the C-terminal autotransporter domain of the remaining 96-113 kDa protein is embedded in the outer membrane and delivers the N-terminal S8 serine protease passenger domain to the outer cell surface. In most strains the N-terminal catalytic 55-65 kDa domain self cleaves and liberates itself from the autotransporter domain after its transfer across the outer cell membrane. In F. nucleatum ATCC 25586 this autocatalytic activity is less efficient resulting in a full length membrane-anchored serine protease. The mature serine protease was found to cleave after Thr, Gly, Ala and Leu residues at the P1 position. Growth of F. nucleatum in complex medium was inhibited when serine protease inhibitors were used. Additional experiments are needed to determine whether fusolisin might be used as a target for controlling fusobacterial infections.