The evolution of coronaviruses, such as SARS-CoV-2, makes broad-spectrum coronavirus preventional or therapeutical strategies highly sought after. Here we report a human angiotensin-converting enzyme 2 (ACE2)-targeting monoclonal antibody, 3E8, blocked the S1-subunits and pseudo-typed virus constructs from multiple coronaviruses including SARS-CoV-2, SARS-CoV-2 mutant variants (SARS-CoV-2-D614G, B.1.1.7, B.1.351, B.1.617.1 and P.1), SARS-CoV and HCoV-NL63, without markedly affecting the physiological activities of ACE2 or causing severe toxicity in ACE2 "knock-in" mice. 3E8 also blocked live SARS-CoV-2 infection in vitro and in a prophylactic mouse model of COVID-19. Cryo-EM and "alanine walk" studies revealed the key binding residues on ACE2 interacting with the CDR3 domain of 3E8 heavy chain. Although full evaluation of safety in non-human primates is necessary before clinical development of 3E8, we provided a potentially potent and "broad-spectrum" management strategy against all coronaviruses that utilize ACE2 as entry receptors and disclosed an anti-coronavirus epitope on human ACE2.
Objective To study the species and amount of bacteria in sputum of patients with ventilator associated pneumonia (VAP) by using 16S rDNA sequencing analysis,and to explore the new method for etiologic diagnosis of VAP.Methods Bronchoalveolar lavage sputum samples were collected from 31 patients with VAP.Bacterial DNA of the samples were extracted and identified by polymerase chain reaction (PCR).At the same time,sputum specimens were processed for routine bacterial culture.The high flux sequencing experiment was conducted on PCR positive samples with 16S rDNA macro genome sequencing technology,and sequencing results were analyzed using bioinformatics,then the results between the sequencing and bacteria culture were compared.Results ① 550 bp of specific DNA sequences were amplified in sputum specimens from 27 cases of the 31 patients with VAP,and they were used for sequencing analysis.103 856 sequences were obtained from those sputum specimens using 16S rDNA sequencing,yielding approximately 39 Mb of raw data.Tag sequencing was able to inform genus level in all 27 samples.② Alpha-diversity analysis showed that sputum samples of patients with VAP had significantly higher variability and richness in bacterial species (Shannon index values 1.20,Simpson index values 0.48).Rarefaction curve analysis showed that there were more species that were not detected by sequencing from some VAP sputum samples.③ Analysis of 27 sputum samples with VAP by using 16S rDNA sequences yielded four phyla:namely Acitinobacteria,Bacteroidetes,Firmicutes,Proteobacteda.With genus as a classification,it was found that the dominant species included Streptococcus 88.9％ (24/27),Limnohabitans 77.8％ (21/27),Acinetobacter 70.4％ (19/27),Sphingomonas 63.0％ (17/27),Prevotella 63.0％ (17/27),Klebsiella 55.6％ (15/27),Pseudomonas 55.6％ (15/27),Aquabacterium 55.6％ (15/27),and Corynebacterium 55.6％ (15/27).④ Pyrophosphate sequencing discovered that Prevotella,Limnohabitans,Aquabacterium,Sphingomonas might not be detected by routine bacteria culture.Among seven species which were identified by both methods,pyrophosphate sequencing yielded higher positive rate than that of ordinary bacteria culture [Streptococcus:88.9％ (24/27) vs.18.5％ (5/27),KlebsieHa:55.6％ (15/27) vs.18.5％ (5/2 7),Acinetobacter:70.4％ (19/27) vs.37.0％ (10/27),Corynebacterium:55.6％ (15/27) vs.7.4％ (2/27),P＜0.05 or P＜0.01].Sequencing positive rate was found to increase positive rate for culture of Pseudomonas [55.6％ (15/27) vs.25.9％ (7/27),P=0.050].No significant differences were observed between sequencing and ordinary bacteria culture for detection Staphylococcus [7.4％ (2/27) vs.11.1％ (3/27)] and Neisseria bacteria genera [18.5％ (5/27) vs.3.7％(1/27),both P＞0.05].Conclusions 16S rDNA sequencing analysis confirmed that pathogenic bacteria in sputum of VAP were complicated with multiple drug resistant strains.Compared with routine bacterial culture,pyrophosphate sequencing had higher positive rate in detecting pathogens.16S rDNA gene sequencing technology may become a new method for etiological diagnosis of VAP.
