Rapid diagnostics and ceftazidime/avibactam for KPC-producing Klebsiella pneumoniae bloodstream infections: impact on mortality and role of combination therapy.

This study was aimed at investigating risk factors for mortality in patients suffering from KPC-producing Klebsiella pneumoniae (KPC-Kp) bloodstream infections (BSIs), evaluating the impact of rapid diagnostics and ceftazidime/avibactam use. This observational retrospective study (January 2017-May 2021) included all patients with a KPC-Kp BSI. Uni-multivariable analyses were carried out to evaluate the effect of clinical variables on both in-hospital death (IHD) and 30-day all-cause mortality, and the role of the combination of ceftazidime/avibactam plus polymyxin. One hundred and ninety-six patients met the study's inclusion criteria. Older age, having undergone renal replacement therapy during the 30 days preceding the KPC-Kp BSI onset, having an INCREMENT-CPE score ≥ 8, and having suffered from a superimposed and/or following KPC-Kp BSI treatment candidemia were found to be the main factors associated with both mortality rates. Among protective factors, the centrality of ceftazidime/avibactam in monotherapy (IHD: OR: 0.34; CI 95%: 0.11-1.00-30-day all-cause mortality: OR: 0.18; CI 95%: 0.04-0.77) or combination (IHD: OR: 0.51; CI 95%: 0.22-1.19-30-day all-cause mortality: OR: 0.62; CI 95%: 0.21-1.84) emerged and became even more evident once the effect of ceftazidime/avibactam plus polymyxin was removed. Rapid diagnostics may be useful to adopt more effective strategies for the treatment of KPC-Kp BSI patients and implement infection control measures, even if not associated with higher patient survival. Ceftazidime/avibactam, even when used alone, represents an important option against KPC-Kp, while combined use with polymyxin might not have altered its efficacy. Patient comorbidities, severity of BSI, and complications such as candidemia were confirmed to have a significant burden on survival.

Detection of Carbapenemase and CTX-M Encoding Genes Directly from Bronchoalveolar Lavage Using the CRE and ESBL ELITe MGB Assays: Toward Early and Optimal Antibiotic Therapy Management of Critically Ill Patients with Pneumonia.

The detection of carbapenemase extended-spectrum ß-lactamase (ESBL)-producing Enterobacterales (EB) has become a major issue among critically ill patients, especially due to their impact on appropriate antimicrobial therapy. This study aimed at evaluating the potential contribution of molecular assays to early optimization of empirical antibiotic therapy among critically ill patients with carbapenemase- and/or CTX-M-producing EB pneumonia. The CRE and ESBL ELITe MGB® assays were evaluated directly on 197 bronchoalveolar lavage (BAL) samples obtained from 120 patients. Molecular results were then compared to routine culture-based diagnostic results, and a retrospective analysis of the therapeutic antimicrobial management was performed. Among the 197 clinical specimens, blaKPC-like and blaCTX-M-like were detected in 20 (10.2%) and 12 (6.1%) specimens belonging to 15 and 11 patients, respectively. Positive predictive value (PPV) and negative predictive value (NPV) of the CRE ELITe MGB Kit were 85% [95% confidence interval [CI]: 64.9-94.6] and 100%, respectively. PPV and NPV of the ESBL ELITe MGB Kit were 75% [95% CI: 49.4-90.2] and 100%, respectively. Retrospective analysis of the therapeutic antimicrobial management at the time of BAL collection showed that in ∼50% of patients with carbapenemase- and CTX-M-producing EB pneumonia empirical antibiotic therapy could have been optimized at least 48-72 hr earlier if positive molecular data had been used. The CRE and ESBL ELITe MGB assays might be an interesting tool for expediting optimization of empirical antibiotic therapy in critically ill patients with pneumonia, depending on local epidemiology of antibiotic resistance, patient risk stratification for EB infection, and availability of an antimicrobial stewardship team.

Development of a competitive ELISA for the evaluation of sunflower pollen in honey samples.

We report the development of a rapid, specific, and sensitive enzyme-linked immunoassay (ELISA) for the evaluation of sunflower pollen in honey as a method alternative to melissopalynology, which is considered the standard technique for the evaluation of floral origin of honey. Two 33-36 kDa proteins, identified as characteristic of sunflower pollen, were isolated and used as coating antigens in the competitive ELISA. We verified its analytical performance by evaluating reproducibility, specificity, and exactitude in relation to melissopalynology. The competitive ELISA developed during this work is able to quantify sunflower pollen in honey, with a detection limit of 10%, showing linear response between 10 and 90%. The method afforded low cross reactivity with honey from other floral origin, thus evidencing an adequate selectivity. We also observed a significant correlation (r = 0.975; p < 0.001) when the proposed ELISA was referenced to melissopalynology. Hence, we conclude that the competitive ELISA constitutes a valuable and feasible alternative for authentication of sunflower honey. This work opens the possibility to develop similar assays for other pollen types.

Assessment of the floral origin of honey by SDS-page immunoblot techniques.

We report on the development of a novel alternative method for the assessment of floral origin in honey samples based on the study of honey proteins using immunoblot assays. The main goal of our work was to evaluate the use of honey proteins as chemical markers of the floral origin of honey. Considering that honeybee proteins should be common to all types of honey, we decided to verify the usefulness of pollen proteins as floral origin markers in honey. We used polyclonal anti-pollen antibodies raised in rabbits by repeated immunization of Sunflower (Elianthus annuus) and Eucalyptus (Eucalyptus sp.) pollen extracts. The IgG fraction was purified by immunoaffinity. These antibodies were verified with nitrocellulose blotted pollen and unifloral honey protein extracts. The antibodies anti-Sunflower pollen, bound to the 36 and 33 kDa proteins of Sunflower unifloral honey and to honey containing Sunflower pollen; and the antibodies anti-Eucalyptus sp. pollen bound to the 38 kDa proteins of Eucalyptus sp. unifloral honey in immunoblot assays. Satisfactory results were obtained in differentiating between the types of pollen analyzed and between Sunflower honey and Eucalyptus honey with less cross reactivity with other types of honey from different origin and also with good sensitivity in the detection. This immunoblot method opens an interesting field for the development of new antibodies from different plants, which could serve as an alternative or complementary method to the usual melissopalynological analysis to assess honey floral origin.