Preparation and Evaluation of a New Lipopolysaccharide-based Conjugate as a Vaccine Candidate for Brucellosis

Objectives: Development of an efficacious vaccine against brucellosis has been a challenge for scientists for many years. At present, there is no licensed vaccine against human brucellosis. To overcome this problem, currently, antigenic determinants of Brucella cell wall such as Lipopolysaccharide (LPS) are considered as potential candidates to develop subunit vaccines. Methods: In this study, Brucella abortus LPS was used for conjugation to Neisseria meningitidis serogroup B outer membrane vesicle (OMV) as carrier protein using carbodiimide and adipic acid-mediated coupling and linking, respectively. Groups of eight BALB/c mice were injected subcutaneously with 10μg LPS alone, combined LPS+OMV and conjugated LPS-OMV on 0 days, 14 days, 28 days and 42 days. Anti-LPS IgG was measured in serum. Results: The yield of LPS to OMV in LPS-OMV conjugate was 46.55, on the basis of carbohydrate content. The ratio for LPS to OMV was 4.07. The LPS-OMV conjugate was the most immunogenic compound that stimulated following the first injection with increased IgG titer of ~5-fold and ~1.3-fold higher than that produced against LPS and LPS in noncovalent complex to OMV (LPS+OMV), respectively. The highest anti-LPS IgG titer was detected 2 weeks after the third injection (Day 42) of LPS-OMV conjugate. The conjugated compound elicited higher titers of IgG than LPS+OMV, that showed a 100-120-fold rise of anti-LPS IgG in mice. Conclusion: These results indicate that our conjugated LPS-OMV can be used as a brucellosis vaccine, but further investigation is required. © 2014

The association of surface adhesin genes and the biofilm formation among Klebsiella oxytoca clinical isolates

Bacterial adhesins mediate the attachment and biofilm production leading to the persistence of colonized strains. The aim of this study was evaluation of the association of surface adhesin genes with the biofilm formation among Klebsiella oxytoca isolates. Among 50 isolates of K. oxytoca from patients with antibiotic-associated diarrhoea, the susceptibility test, MIC (according to CLSI 2016) and phenotypic biofilm formation (with microtitre tissue-plate assay) were performed. The presence of adhesins was investigated using PCR. Thirty-three (66%) isolates produced moderate-level biofilms, but none of them exhibited strong biofilm formation. The presence of adhesins was as follows: fimA, 60% (n = 30), mrkA, 42% (n = 21), matB, 96% (n = 48) and pilQ, 92% (n = 46). The biofilm formation was related to the presence of fimA (odds ratio (OR) 0.8571, 95% CI 1.733–6.267, p <0.0001), mrkA (OR 0.2462, 95% CI 2.723–4.622, p 0.001), matB (OR 0.4521, 95% CI 1.353–5.332, p 0.008) and pilQ (OR 0.1481, 95% CI 1.691–6.117, p <0.0001). The npsB toxin-encoding gene was detected among 46 (92%) isolates. Resistance to non-β-lactam antibiotics was significantly associated with the presence of adhesin-encoding genes. The presence of adhesins and the capsular encoding gene was significantly associated with biofilm formation among K. oxytoca isolates. The presence of surface adhesin-encoding genes was significantly associated with the biofilm formation and also with resistance to non-β-lactam antibiotics among K. oxytoca clinical isolates. In addition, biofilm production was not significantly associated with β-lactam resistance among the isolates. Keywords: Adhesins, antibiotic resistance, biofilms, Klebsiella oxytoc