Nanoarchitectures for efficient IgE cross-linking on effector cells to study amoxicillin allergy.

BACKGROUND: Amoxicillin (AX) is nowadays the ß-lactam that more frequently induces immediate allergic reactions. Nevertheless, diagnosis of AX allergy is occasionally challenging due to risky in vivo tests and non-optimal sensitivity of in vitro tests. AX requires protein haptenation to form multivalent conjugates with increased size to be immunogenic. Knowing adduct structural features for promoting effector cell activation would help to improve in vitro tests. We aimed to identify the optimal structural requirement in specific cellular degranulation to AX using well-precised nanoarchitectures of different lengths. METHOD: We constructed eight Bidendron Antigens (BiAns) based on polyethylene glycol (PEG) linkers of different lengths (600-12,000 Da), end-coupled with polyamidoamine dendrons that were terminally multi-functionalized with amoxicilloyl (AXO). In vitro IgE recognition was studied by competitive radioallergosorbent test (RAST) and antibody-nanoarchitecture complexes by transmission electron microscopy (TEM). Their allergenic activity was evaluated using bone marrow-derived mast cells (MCs) passively sensitized with mouse monoclonal IgE against AX and humanized RBL-2H3 cells sensitized with polyclonal antibodies from sera of AX-allergic patients. RESULTS: All BiAns were recognized by AX-sIgE. Dose-dependent activation responses were observed in both cellular assays, only with longer structures, containing spacers in the range of PEG 6000-12,000 Da. Consistently, greater proportion of immunocomplexes and number of antibodies per complex for longer BiAns were visualized by TEM. CONCLUSIONS: BiAns are valuable platforms to study the mechanism of effector cell activation. These nanomolecular tools have demonstrated the importance of the adduct size to promote effector cell activation in AX allergy, which will impact for improving in vitro diagnostics.

Dendrimeric Antigens for Drug Allergy Diagnosis: A New Approach for Basophil Activation Tests.

Dendrimeric Antigens (DeAns) consist of dendrimers decorated with multiple units of drug antigenic determinants. These conjugates have been shown to be a powerful tool for diagnosing penicillin allergy using in vitro immunoassays, in which they are recognized by specific IgE from allergic patients. Here we propose a new diagnostic approach using DeAns in cellular tests, in which recognition occurs through IgE bound to the basophil surface. Both IgE molecular recognition and subsequent cell activation may be influenced by the tridimensional architecture and size of the immunogens. Structural features of benzylpenicilloyl-DeAn and amoxicilloyl-DeAn (G2 and G4 PAMAM) were studied by diffusion Nuclear Magnetic Resonance (NMR) experiments and are discussed in relation to molecular dynamics simulation (MDS) observations. IgE recognition was clinically evaluated using the basophil activation test (BAT) for allergic patients and tolerant subjects. Diffusion NMR experiments, MDS and cellular studies provide evidence that the size of the DeAn, its antigen composition and tridimensional distribution play key roles in IgE-antigen recognition at the effector cell surface. These results indicate that the fourth generation DeAns induce a higher level of basophil activation in allergic patients. This approach can be considered as a potential complementary diagnostic method for evaluating penicillin allergy.

Pseudomonas aeruginosa isolates and their antimicrobial susceptibility pattern among catheterized patients at Jimma University Teaching Hospital, Jimma, Ethiopia.

BACKGROUND: Pseudomonas aeruginosa is among the most common bacterial pathogens with wide spread distribution in health care settings. Despite advances in medical and surgical care and introduction of wide variety of antimicrobial agents, Pseudomonas aeruginosa continues to cause life threatening infection. Thus, this study aims to isolate and determine antimicrobial susceptibility patterns of Pseudomonas aeruginosa from catheterized patients with urinary tract infection. RESULT: A cross-sectional study was conducted from January to May, 2013. Urine specimens of 73 catheterized patients who developed urinary tract infection after catheterization were collected from sampling port of the catheter. The urine samples were inoculated on MaConckey and blood agar plates, and incubated at 37 °C for 24 h. The isolates were identified by conventional microbiological tests. Antimicrobial susceptibility pattern was determined by modified Kirby-Bauer disk diffusion method. From a total of 73 urine samples collected P. aeruginosa was isolated from 36 (49.32%) catheterized patients; 17 (23.29%) males and 19 (26.03%) females. While all P. aeruginosa isolates were found to be susceptible to Norfloxacin and Ciprofloxacin most isolates were also susceptible to Gentamicin (86.12%). CONCLUSION: The result shows higher prevalence of P. aeruginosa isolates among catheterized patients and the isolates were susceptible to the antimicrobials studied. All P. aeruginosa isolates were susceptible to Ciprofloxacin and Norfloxacin with some of the isolates shown resistance to Gentamicin. While the susceptibility of the isolates to the two fluoroquinolones is a good news for the prescribers their future rational prescription and use should be the main focus.