Efficient control of the fungal pathogens Colletotrichum gloeosporioides and Penicillium digitatum infecting citrus fruits by native soilborne Bacillus velezensis strains.

Destruction of citrus fruits by fungal pathogens during preharvest and postharvest stages can result in severe losses for the citrus industry. Antagonistic microorganisms used as biological agents to control citrus pathogens are considered alternatives to synthetic fungicides. In this study, we aimed to identify fungal pathogens causing dominant diseases on citrus fruits in a specialized citrus cultivation region of Vietnam and inspect soilborne Bacillus isolates with antifungal activity against these pathogens. Two fungal pathogens were characterized as Colletotrichum gloeosporioides and Penicillium digitatum based on morphological characteristics and ribosomal DNA internal transcribed spacer sequence analyses. Reinfection assays of orange fruits confirmed that C. gloeosporioides causes stem-end rot, and P. digitatum triggers green mold disease. By the heterologous expression of the green fluorescent protein (GFP) in C. gloeosporioides using Agrobacterium tumefaciens-mediated transformation, we could observe the fungal infection process of the citrus fruit stem-end rot caused by C. gloeosporioides for the first time. Furthermore, we isolated and selected two soilborne Bacillus strains with strong antagonistic activity for preventing the decay of citrus fruits by these pathogens. Molecular analyses of 16 S rRNA and gyrB genes showed that both isolates belong to B. velezensis. Antifungal activity assays indicated that bacterial culture suspensions could strongly inhibit C. gloeosporioides and P. digitatum, and shield orange fruits from the invasion of the pathogens. Our work provides a highly effective Bacillus-based preservative solution for combating the fungal pathogens C. gloeosporioides and P. digitatum to protect citrus fruits at the postharvest stages.

The contribution of hospital-acquired infections to the COVID-19 epidemic in England in the first half of 2020.

Background SARS-CoV-2 is known to transmit in hospital settings, but the contribution of infections acquired in hospitals to the epidemic at a national scale is unknown. Methods We used comprehensive national English datasets to determine the number of COVID-19 patients with identified hospital-acquired infections (with symptom onset >7 days after admission and before discharge) in acute English hospitals up to August 2020. As patients may leave the hospital prior to detection of infection or have rapid symptom onset, we combined measures of the length of stay and the incubation period distribution to estimate how many hospital-acquired infections may have been missed. We used simulations to estimate the total number (identified and unidentified) of symptomatic hospital-acquired infections, as well as infections due to onward community transmission from missed hospital-acquired infections, to 31 st July 2020. Results In our dataset of hospitalised COVID-19 patients in acute English hospitals with a recorded symptom onset date (n = 65,028), 7% were classified as hospital-acquired. We estimated that only 30% (range across weeks and 200 simulations: 20-41%) of symptomatic hospital-acquired infections would be identified, with up to 15% (mean, 95% range over 200 simulations: 14.1%-15.8%) of cases currently classified as community-acquired COVID-19 potentially linked to hospital transmission. We estimated that 26,600 (25,900 to 27,700) individuals acquired a symptomatic SARS-CoV-2 infection in an acute Trust in England before 31st July 2020, resulting in 15,900 (15,200-16,400) or 20.1% (19.2%-20.7%) of all identified hospitalised COVID-19 cases. Conclusions Transmission of SARS-CoV-2 to hospitalised patients likely caused approximately a fifth of identified cases of hospitalised COVID-19 in the "first wave" in England, but less than 1% of all infections in England. Using time to symptom onset from admission for inpatients as a detection method likely misses a substantial proportion (>60%) of hospital-acquired infections.

Rapid and efficient western blot assay by rotational cyclic draining and replenishing procedure.

Western blot is a principal technique for the detection of specific proteins in various biology disciplines. The antibody incubation is an imperative step in western blot. Antibody incubation by mild shaking on a rocker is generally used to facilitate mixing of antibodies. However, mild shaking is an inefficient process to remove antibody depletion layer on blotting membrane and requires hours of incubation time to achieve antibody binding to target proteins. We propose an alternative method of cyclic draining and replenishing (CDR) incubation of antibody solution using a rotational incubation chamber. The study demonstrated that rotational CDR incubation could shorten antibody incubation time with enhanced sensitivity. Moreover, rotational CDR incubation could achieve a stronger antibody binding with lower antibody concentration when compared with batch incubation. In addition, rotational CDR incubation significantly improved the detection of low abundance proteins. This simple modification in western blot procedure makes it rapid, sensitive, and cost-efficient.

Discovery of DNA dyes Hoechst 34580 and 33342 as good candidates for inhibiting amyloid beta formation: in silico and in vitro study.

Combining Lipinski's rule with the docking and steered molecular dynamics simulations and using the PubChem data base of about 1.4 million compounds, we have obtained DNA dyes Hoechst 34580 and Hoechst 33342 as top-leads for the Alzheimer's disease. The binding properties of these ligands to amyloid beta (Aß) fibril were thoroughly studied by in silico and in vitro experiments. Hoechst 34580 and Hoechst 33342 prefer to locate near hydrophobic regions with binding affinity mainly governed by the van der Waals interaction. By the Thioflavin T assay, it was found that the inhibition constant IC50 ≈ 0.86 and 0.68 µM for Hoechst 34580 and Hoechst 33342, respectively. This result qualitatively agrees with the binding free energy estimated using the molecular mechanic-Poisson Boltzmann surface area method and all-atom simulations with the AMBER-f99SB-ILDN force field and water model TIP3P. In addition, DNA dyes have the high capability to cross the blood brain barrier. Thus, both in silico and in vitro experiments have shown that Hoechst 34580 and 33342 are good candidates for treating the Alzheimer's disease by inhibiting Aß formation.