Overexpression of high affinity K+ transporter from Nitraria sibirica enhanced salt tolerance of transgenic plants.

Nitraria sibirica Pall is a halophytic shrub growing in desert steppe zones. It exhibits extraordinary adaptability to saline-alkali soil, drought, and sand burial. In this study, the high-affinity K+ transporter NsHKT1 was identified and found to play a key role in salt tolerance in N. sibirica. NsHKT1 was used to improve salt tolerance in a poplar hybrid. The expression characteristics of NsHKT1 were analyzed by transforming Arabidopsis and poplar with the ß-glucuronidase (GUS) gene driven by the NsHKT1 promoter. The results showed that NsHKT1 expression was induced by various abiotic stresses and phytohormones. GUS expression was also detected in the reproductive organs of transgenic Arabidopsis, indicating its function in regulating plant reproductive growth. Transgenic 84 K poplar plants overexpressing NsHKT1 exhibited less damage, higher antioxidant capacity, higher chlorophyll and proline levels, and lower malondialdehyde content compared with non-transgenic plants under salt stress. These results are consistent with the salt tolerance results for transgenic Arabidopsis overexpressing NsHKT1, indicating that NsHKT1 plays a key role in salt tolerance in herbaceous and ligneous plants. Inductively coupled plasma-optical emission spectrometry showed a significantly lower leaf Na+ content in transgenic poplar than in the non-transgenic line, revealing that NsHKT1, as a member of HKT family subclass 1, was highly selective to Na+ and prevented shoot Na+ accumulation. Transcriptome analysis indicated that differentially expressed genes in transgenic poplars under salt stress were associated mainly with the isoflavonoid, cutin, suberine, wax, anthocyanin, flavonoid, and cyanoamino biosynthesis pathways, as well as the MAPK signaling pathway, indicating that NsHKT1 not only regulates ion homeostasis but also influences secondary metabolism and signal transaction in transgenic plants.

Synergistic effect of linezolid with fosfomycin against Staphylococcus aureus in vitro and in an experimental Galleria mellonella model.

BACKGROUND/PURPOSES: Treatment of Staphylococcus aureus infections is challenging owing to widespread multidrug resistance. There is now considerable interest in the potential of combination therapies. Although linezolid/fosfomycin combination appears to be a promising treatment option based on in vitro data, further preclinical work is needed. In this study, the Galleria mellonella system was employed to study the in vivo efficacy of this combination in order to determine whether it should be explored further for the treatment of S. aureus infections. METHODS: The antimicrobial activity of linezolid and fosfomycin alone and in combination was assessed versus four S. aureus. Synergy studies were performed using the microtitre plate chequerboard assay and time-kill methodology. The in vivo activity of linezolid/fosfomycin combination was assessed using a G. mellonella larvae model. RESULTS: The combination of linezolid and fosfomycin was synergistic and bacteriostatic against four tested strains. Treatment of G. mellonella larvae infected with lethal doses of S. aureus resulted in significantly enhanced survival rates when low-dose of combination has no significant differences with high-dose combination (P > 0.05), G. mellonella hemolymph burden of S. aureus suggest that combination therapy with rapid and sustained bacteriostatic activity compared monotherapy. CONCLUSION: This work indicated that linezolid combination with fosfomycin has synergistic effect against S. aureus in vitro and in an experimental G. mellonella model, and it suggests that high-dose of linezolid and fosfomycin may not necessary.

Pharmacodynamics Of Linezolid-Plus-Fosfomycin Against Vancomycin-Susceptible And -Resistant Enterococci In Vitro And In Vivo Of A Galleria mellonella Larval Infection Model.

OBJECTIVES: To explore the in vitro and in vivo antibacterial activity of linezolid/fosfomycin combination against vancomycin-susceptible and -resistant enterococci (VSE and VRE), and provide a theoretical basis for the treatment of VRE. METHODS: The checkerboard method and time-kill curve study were used to evaluate the efficacy of linezolid combined with fosfomycin against VSE and VRE. The transmission electron microscopy (TEM) was employed to observe the cell morphology of bacteria treated with each drug alone or in combination, which further elucidate the mechanism of action of antibiotic combination therapy. The Galleria mellonella infection model was constructed to demonstrate the in vivo efficacy of linezolid plus fosfomycin for VSE and VRE infection. RESULTS: The fractional inhibitory concentration index (FICI) values of all strains suggested that linezolid showed synergy or additivity in combination with fosfomycin against five of the six strains. Time-kill experiments demonstrated that the combination of linezolid-fosfomycin at 1×MIC or 2×MIC led to higher degree of bacterial killing without regrowth for all isolates tested than each monotherapy. TEM images showed that the combination treatment damaged the bacterial cell morphology more obviously than each drug alone. In the Galleria mellonella infection model, the enhanced survival rate of the combination treatment compared with linezolid monotherapy (P<0.05) was revealed. CONCLUSION: Our data manifested that the combination of linezolid and fosfomycin was a potential therapeutic regimen for VRE infection. The combination displayed excellent bacterial killing and inhibited amplification of fosfomycin-resistant subpopulations.

In vitro activity and post-antibiotic effects of linezolid in combination with fosfomycin against clinical isolates of Staphylococcus aureus.

OBJECTIVES: Linezolid combination therapy is recommended for the treatment of Staphylococcus aureus (S. aureus) infections. However, the optimal regimen of the combination therapy for S. aureus is unknown. The objective of this study was to investigate the antibacterial activity, post-antibiotic effect (PAE), and post-antibiotic subminimum inhibitory concentration (MIC) effect (PA-SME) of linezolid alone and in combination with fosfomycin against eleven clinical isolates of S. aureus. METHODS: The synergistic effects and antibacterial activity of linezolid and fosfomycin were assessed by checkerboard and time-kill assays. To determine the PAE and PA-SME, S. aureus strains in the logarithmic phase of growth were exposed for 1, 2, and 3 hours to the antibiotics, alone and in combination. Recovery periods of test strains were evaluated using viable counting after dilution. RESULTS: Synergistic effects were observed for eight strains and no antagonism was found with any combination. Moreover, linezolid combined with fosfomycin at 4x MIC showed the best synergistic antibacterial effect, and this effect was retained after 24 hours. In addition, both the antibiotics alone and in combination showed increased PAE and PA-SME values in a concentration- and time-dependent manner. CONCLUSION: Linezolid combined with fosfomycin exerted a good antibacterial effect against S. aureus, and the combinations have significant PAE and PA-SME.