ISG15, the first ubiquitin-like molecule identified two decades ago, is encoded by interferon stimulated gene 15 ( ISG 15), where its robust expression can be induced by viral infections or interferon treatments. ISG 15 conjugate to other proteins as the ubiquitin and was found to be involved in innate immune response. However, the functions of ISG15 modification remained unclear. We cloned bovine ISG15(bisg15) into a prokaryotic expression vector pET28a( + ) with a His-tag to generate a soluble form of bISG15 fusion protein, and purified with Ni-NTA Sepharose chromatography. The purified protein was concentrated and used to immune Balb/c mice to raise the antiserum, which could specifically recognize bISG15 expressed in eukaryotic cells by Western blot analysis. The concentrated bISG15 protein and its antiserum were then used to establish an in vitro bISG15 modification system. Our studies have demonstrated that cellular proteins could be conjugated to bISG15 with this system.

Objective To analyze the pathogenic features and risk factors of hospital-acquired pneumonia in patients with acute spontaneous intracerebral hemorrhage (sICH) in intensive care unit (ICU).Methods The clinical data of 110 patients with sICH admitted in ICU during January 2015 and February 2017 were collected.Patients were divided into hospital-acquired pneumonia group (HAP group,n =66) and non-HAP group (n =44).Multivariate Logistic regression was used to study the risk factors of HAP,and pathogen distribution and drug susceptibility were analyzed.Results Multivariate Logistic regression demonstrated that long-term mechanical ventilation (OR =1.028,95％ CI 1.012-1.044,P ＜ 0.01),lower score of glasgow coma scale (GCS) (OR =1.550,95％ CI 1.148-2.093,P ＜ 0.01),prolonged hospital stay (OR =1.131,95％ CI 1.046-1.224,P ＜0.01) and underlying diseases more than two forms (OR =9.793,95％ CI 1.012-1.044,P ＜ 0.01) were the independent risk factors of HAP,while high plasma albumin level was protective factor for HAP (OR =0.897,95％ CI O.811-0.992,P ＜ 0.05).One hundred and eighty-three bacterial strains were isolated from 66 patients,the top 4 pathogens were Acinetobacter baumannii (28.96％,53/183),Klebsiella pneumonia (15.85％,29/183),Pseudomonas aeruginosa (13.11％,24/183) and Staphylococcus aureus (12.02％,22/183).Acinetobacter baumannii,Klebsiella pneumoniae and Pseudomonas aeruginosa were highly resistant to the majority of antibiotics,some of which even reached 100％.Staphylococcus aureus showed high resistance to macrolides,fluoroquinolones and β-lactam antibiotics.Conclusions There is high incidence of HAP in patients with sICH,and the pathogenic bacteria are mainly gram-negative bacteria.Effective prevention and treatment measures should be taken to reduce the incidence of HAP for patients with sICH in ICU.