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We aimed to obtain the recombinant aminopeptidase encoded by Listeria monocytogenes (L. monocytogenes) gene lmo1711, and characterized the enzyme. First, the amino acid sequences of Lmo1711 from L. monocytogenes EGD-e and its homologues in other microbial species were aligned and the putative active sites were analyzed. The putative model of Lmo1711 was constructed through the SWISS-MODEL Workspace. Then, the plasmid pET30a-Lmo1711 was constructed and transformed into E. coli for expression of the recombinant Lmo1711. The his-tagged soluble protein was purified using the nickel-chelated affinity column chromatography. With the amino acid-p-nitroaniline as the substrate, Lmo1711 hydrolyzed the substrate to free p-nitroaniline monomers, whose absorbance measured at 405 nm reflected the aminopeptidase activity. The specificity of Lmo1711 to substrates was then examined by changing various substrates, and the effect of metal ions on the catalytic efficiency of this enzyme was further determined. Based on the bioinformatics data, Lmo1711 is a member of the M29 family aminopeptidases, containing a highly conserved catalytic motif (Glu-Glu-His-Tyr-His-Asp) with typical structure arrangements of the peptidase family. The recombinant Lmo1711 with a size of about 49.3 kDa exhibited aminopeptidase activity and had a selectivity to the substrates, with the highest degree of affinity for leucine-p-nitroaniline. Interestingly, the enzymatic activity of Lmo1711 can be activated by Cd²⁺, Zn²⁺, and is strongly stimulated by Co²⁺. We here, for the first time demonstrate that L. monocytogenes lmo1711 encodes a cobalt-activated aminopeptidase of M29 family.

Objective: To investigate the curative effect of lateral position ventilation combined with vibration sputum drainage on the patients with acute respiratory distress syndrome (ARDS)., Methods: A prospective randomized controlled trial was conducted. The patients with ARDS undergoing mechanical ventilation (MV) admitted to intensive care unit (ICU) of Linyi Central Hospital from January 2013 to June 2017 were enrolled, and they were divided into simple ventilation group and combined treatment group according to random number table. The patients in both groups received etiological treatment, protective ventilation strategy, sensitive antibiotics for anti-infection, and calefacient and humidifying treatment. The patients in the simple ventilation group received bilateral discontinuous alternative lateral position ventilation [pressure controlled ventilation (PCV), tidal volume (VT) ≤ 6 mL/kg, the inhaled oxygen concentration (FiO 2 ) and positive end expiratory pressure (PEEP) was adjusted to maintain the airway plateau pressure (Pplat) ≤ 30 cmH 2 O (1 cmH 2 O = 0.098 kPa)], and those in the combined treatment group received lateral position ventilation combined with vibration sputum drainage, twice a day, 15 minutes each time. The parameters of respiratory function and inflammation as well as excretion of sputum before and after treatment were compared between the two groups, and the complication and prognosis were recorded., Results: A total of 200 patients with ARDS were included, 4 patients were excluded because of severe pneumothorax, massive hemorrhage of the digestive tract, or elevated intracranial pressure, so 196 patients were finally enrolled in the analysis. There were 98 patients in the simple ventilation group and the combined treatment group, respectively. There were no significant differences in parameters of respiratory function and inflammation as well as excretion of sputum before treatment between the two groups. After treatment, the above parameters in both groups were improved, and the effect of combined treatment was more significant. Compared with the simple ventilation group, white blood cell count (WBC), procalcitonin (PCT), C-reactive protein (CRP) at 24 hours of treatment in the combined treatment group were significantly decreased [WBC (×10 9 /L): 9.1±1.6 vs. 11.8±3.6, PCT (μg/L): 14.5±2.4 vs. 22.7±3.2, CRP (mg/L): 32.2±6.3 vs. 67.2±7.2, all P < 0.01], the quantity of sputum excretion was significantly increased (mL: 49.3±12.5 vs. 36.8±11.8, P < 0.01); 72 hours after treatment, the oxygenation index (PaO 2 /FiO 2 ) in the combined treatment group was significantly increased [mmHg (1 mmHg = 0.133 kPa): 278±28 vs. 238±39, P < 0.01], and PEEP, FiO 2 , airway resistance (Raw) were significantly lowered [PEEP (cmH 2 O): 5±2 vs. 7±3, FiO 2 : 0.40±0.11 vs. 0.49±0.12, Raw (cmH 2 O): 12.8±0.7 vs. 16.2±0.8, all P < 0.01]. There was no significant difference in static lung compliance (Cst) or Pplat between the two groups. Compared with the simple ventilation group, the duration of MV (days: 5.5±3.0 vs. 8.6±2.1), the length of ICU stay (days: 7.5±5.4 vs. 11.3±4.6) and the extraction time of artificial airway (days: 6.6±2.8 vs. 9.4±3.3) in the combined treatment group were significantly shortened (all P < 0.01). However, there was no significant difference in the 28-day mortality rate [4.1% (4/98) vs. 3.1% (3/98)], the rate of tracheotomy [2.0% (2/98) vs. 3.1% (3/98)] or the incidence of I-II skin pressure sore [1.0% (1/98) vs. 2.0% (2/98)] between simple ventilation group and the combined treatment group (all P > 0.05). No artificial airway disappearance, pneumothorax, grade III or IV pressure sore was found in the two groups., Conclusions: Compared with lateral position ventilation alone, the treatment of combined with vibration sputum drainage is more effective for improving the respiratory function of patients with ARDS, reducing infection action, shortening the duration of MV and the length of ICU stay, and improve the recovery, promote the rehabitation of patients.

Fumonisin B1 (FB1) is a carcinogenic mycotoxin found in commodities such as corn and corn-originated products. An aptamer-based method for detection of FB1 was developed using the fluorescent dye PicoGreen, which can recognize and bind double-stranded DNA. A peak fluorescence of PicoGreen was obtained in 15 min in the presence of FB1 aptamer, which formed a double-stranded hybridizer DNA with its complementary strand. The excitation and emission wavelengths for PicoGreen detection were 480 nm and 520 nm, respectively. The sensitivity of this aptamer/PicoGreen-based method was 0.1 μg/L. This method showed a good linearity for FB1 concentration ranging from 0.1 to 1 μg/L. The entire detection procedure for FB1 could be completed within 40 min. No cross reactions were observed with any other mycotoxins against aflatoxin B1, ochratoxin A, citrinin and zearalenone, demonstrating high specificity towards FB1 aptamer. Agreement between commercial, antibody-based enzyme-linked immunosorbent assay (ELISA) kit and aptamer method was excellent with a kappa value of 0.857. Taken together, this aptamer/PicoGreen-based method is more cost-effective, time-saving and useful than ELISA for detection of FB1.