Risk factors associated with the occurrence of multiresistant Staphylococcus spp. isolated from bovine subclinical mastitis in northern Brazil

Staphylococcus bacteria are often associated with subclinical bovine mastitis. This study aimed to identify multiresistant Staphylococcus spp. associated with subclinical mastitis and the associated risk factors. Twenty-three dairy farms with a history of decrease in milk production, located in the lower Acre region, Brazil, were selected. An epidemiological questionnaire was provided in all farms. All animals were examined using the California Mastitis Test (CMT) and their milk samples were collected for bacterial culture. After isolation and identification, the disk diffusion antimicrobial susceptibility test was performed against nine classes of antimicrobials. Of the 339 cows examined using the CMT, 108 had mastitis. A total of 229 milk samples were collected from individual teats. MALDI-TOF MS found isolates belonging to eight species of Staphylococcus, in 101 of these samples. S. chromogenes (58.4%) demonstrated strongest resistance to the nine classes of antimicrobial active principles. Nineteen isolates with multidrug resistance phenotypic profile were identified. This phenotypic expression indicates wide circulation of resistant genes in this species. The presence of multidrug resistance in Staphylococcus spp. in this study was correlated with lack of water for cleaning the corral, which is a preventive factor, minimizing the transmission and persistence of pathogens in the farms.

Anlotinib Reverses Multidrug Resistance (MDR) in Osteosarcoma by Inhibiting P-Glycoprotein (PGP1) Function In Vitro and In Vivo

Overexpression of the multidrug resistance (MDR)-related protein P-glycoprotein (PGP1), which actively extrudes chemotherapeutic agents from cells and significantly decreases the efficacy of chemotherapy, is viewed as a major obstacle in osteosarcoma chemotherapy. Anlotinib, a novel tyrosine kinase inhibitor (TKI), has good anti-tumor effects in a variety of solid tumors. However, there are few studies on the mechanism of anlotinib reversing chemotherapy resistance in osteosarcoma. In this study, cellular assays were performed in vitro and in vivo to evaluate the MDR reversal effects of anlotinib on multidrug-resistant osteosarcoma cell lines. Drug efflux and intracellular drug accumulation were measured by flow cytometry. The vanadate-sensitive ATPase activity of PGP1 was measured in the presence of a range of anlotinib concentrations. The protein expression level of ABCB1 was detected by Western blotting and immunofluorescence analysis. Our results showed that anlotinib significantly increased the sensitivity of KHOSR2 and U2OSR2 cells (which overexpress PGP1) to chemotherapeutic agents in vitro and in a KHOSR2 xenograft nude mouse model in vivo. Mechanistically, anlotinib increases the intracellular accumulation of PGP1 substrates by inhibiting the efflux function of PGP1 in multidrug-resistant cell lines. Furthermore, anlotinib stimulated the ATPase activity of PGP1 but affected neither the protein expression level nor the localization of PGP1. In animal studies, anlotinib in combination with doxorubicin (DOX) significantly decreased the tumor growth rate and the tumor size in the KHOSR2 xenograft nude mouse model. Overall, our findings suggest that anlotinib may be useful for circumventing MDR to other conventional antineoplastic drugs.

Plethora of Resistance Genes in Carbapenem-Resistant Gram-Negative Bacteria in Greece: No End to a Continuous Genetic Evolution

Carbapenem-resistant Gram-negative bacteria are a public health threat that requires urgent action. The fact that these pathogens commonly also harbor resistance mechanisms for several other antimicrobial classes further reduces patient treatment options. The present study aimed to provide information regarding the multidrug resistance genetic background of carbapenem-resistant Gram-negative bacteria in Central Greece. Strains from a tertiary care hospital, collected during routine practice, were characterized using a DNA microarray-based assay. Various different resistance determinants for carbapenems, other beta-lactams, aminoglycosides, quinolones, trimethoprim, sulfonamides and macrolides were detected among isolates of the same sequence type. Eighteen different multidrug resistance genomic profiles were identified among the twenty-four K. pneumoniae ST258, seven different profiles among the eight K. pneumoniae ST11, four profiles among the six A. baumannii ST409 and two among the three K. oxytoca. This report describes the multidrug resistance genomic background of carbapenem-resistant Gram-negative bacteria from a tertiary care hospital in Central Greece, providing evidence of their continuous genetic evolution.

Crystal structures of bacterial Small Multidrug Resistance transporter EmrE in complex with structurally diverse substrates

Proteins from the bacterial small multidrug resistance (SMR) family are proton-coupled exporters of diverse antiseptics and antimicrobials, including polyaromatic cations and quaternary ammonium compounds. The transport mechanism of the Escherichia coli transporter, EmrE, has been studied extensively, but a lack of high-resolution structural information has impeded a structural description of its molecular mechanism. Here we apply a novel approach, multipurpose crystallization chaperones, to solve several structures of EmrE, including a 2.9 Å structure at low pH without substrate. We report five additional structures in complex with structurally diverse transported substrates, including quaternary phosphonium, quaternary ammonium, and planar polyaromatic compounds. These structures show that binding site tryptophan and glutamate residues adopt different rotamers to conform to disparate structures without requiring major rearrangements of the backbone structure. Structural and functional comparison to Gdx-Clo, an SMR protein that transports a much narrower spectrum of substrates, suggests that in EmrE, a relatively sparse hydrogen bond network among binding site residues permits increased sidechain flexibility.

LINC00883 Promotes Drug Resistance of Glioma Through a microRNA-136/NEK1-Dependent Mechanism

ObjectiveAccumulating evidence has highlighted the roles of long noncoding RNAs (lncRNAs) as competing endogenous RNAs (ceRNAs) of microRNAs (miRNAs) through their binding sites in the progression of glioma. Hereby, we aim to explore the role of LINC00883 as a regulator of miR-136 and its target, NIMA-related kinase 1 (NEK1), thus, its involvement in the drug resistance of glioma cells.Methods and ResultsMechanistic investigations by dual-luciferase reporter, RNA pull-down, and RNA-binding protein immunoprecipitation (RIP) assays indicated that LINC00883 bound to miR-136, thereby blocking miR-136-induced downregulation of NEK1. Through gain-of-function experiments in U251 cells that presented a high drug resistance, we found that ectopic expression of LINC00883 resulted in increased MRP (encoding multidrug resistance-associated protein), limited cell apoptosis, and increased proliferation. Expectedly, depleting LINC00883 yielded tumor-suppressive and anti-chemoresistance effects on U251 cells by increasing miR-136 and inhibiting NEK1. Next, drug-resistant glioma cell line SOWZ1, drug-sensitive glioma cell line SOWZ2, and drug-resistant glioma cell line SOWZ2-BCNU (SOWZ2 cultured in BCNU) were applied to validate the roles of LINC00883 in the regulation of multidrug resistance. LINC00883 knockdown suppressed the viability of SWOZ1, SWOZ2, and SWOZ2-BCNU cells.ConclusionIn conclusion, LINC00883 knockdown reduces drug resistance in glioma. Hence, our study provides a future strategy to prevent drug resistance-induced therapeutic failure in glioma.