Metatranscriptome of human lung microbial communities in a cohort of mechanically ventilated COVID-19 Omicron patients.

The Omicron variant of the severe acute respiratory syndrome coronavirus 2 (SARS­CoV­2) infected a substantial proportion of Chinese population, and understanding the factors underlying the severity of the disease and fatality is valuable for future prevention and clinical treatment. We recruited 64 patients with invasive ventilation for COVID-19 and performed metatranscriptomic sequencing to profile host transcriptomic profiles, plus viral, bacterial, and fungal content, as well as virulence factors and examined their relationships to 28-day mortality were examined. In addition, the bronchoalveolar lavage fluid (BALF) samples from invasive ventilated hospital/community-acquired pneumonia patients (HAP/CAP) sampled in 2019 were included for comparison. Genomic analysis revealed that all Omicron strains belong to BA.5 and BF.7 sub-lineages, with no difference in 28-day mortality between them. Compared to HAP/CAP cohort, invasive ventilated COVID-19 patients have distinct host transcriptomic and microbial signatures in the lower respiratory tract; and in the COVID-19 non-survivors, we found significantly lower gene expressions in pathways related viral processes and positive regulation of protein localization to plasma membrane, higher abundance of opportunistic pathogens including bacterial Alloprevotella, Caulobacter, Escherichia-Shigella, Ralstonia and fungal Aspergillus sydowii and Penicillium rubens. Correlational analysis further revealed significant associations between host immune responses and microbial compositions, besides synergy within viral, bacterial, and fungal pathogens. Our study presents the relationships of lower respiratory tract microbiome and transcriptome in invasive ventilated COVID-19 patients, providing the basis for future clinical treatment and reduction of fatality.

[miR-203 inhibits lung cancer cell metastasis by targeting fatty acid binding protein 4].

OBJECTIVE: To explore the role of fatty acid binding protein 4 (FABP4) in regulating lung cancer cell metastasis and identify miRNAs that target FABP4. METHODS: The expression of FABP4 in lung cancer cells with different metastatic potentials was detected using enzyme-linked immunosorbent assay (ELISA) and Western blotting. The effects of FABP4 knockdown or overexpression by shRNA or a recombinant lentivirus, respectively, on lung cancer cells metastasis were assessed. The miRNAs that targeted FABP4 were screened using target prediction algorithms and the results were verified with Q-PCR. RESULTS: FABP4 expression was significantly higher in lung cancer cell lines with high metastatic potentials (NL9980, H661, and 95C) than in those with low metastatic potentials (L9981, A549, and PC13) (P<0.05). FABP4 knockdown in NL9980 cells resulted in significantly inhibited metastasis of the cells (P<0.05), while FABP4 overexpression obviously promoted the metastasis of A549 cells (P<0.05). The expressions of miR-203, miR-361 and miR-539 were significantly higher in highly metastatic lung cancer cells than in the cells with low metastatic potentials (P<0.05). In NL9980 cells, FABP4 expression was most obviously suppressed by miR-203 (P<0.05), and target site mutational FABP4 overexpression significantly attenuated the inhibitory effect of miR-203 on NL9980 metastasis (P<0.05). CONCLUSION: FABP4 can promote lung cancer metastasis, and by targeting FABP4 to inhibit its expression, miR-203 can suppress the metastasis of lung cancer cells.

Synthesis of 13-ß-elemene ester derivatives and evaluation of their antioxidant activity in human umbilical vein endothelial cells.

In the present study, a series of 13-ß-elemene ester derivatives were designed and prepared, and their antioxidant activity was investigated in the H2O2-treated human umbilical vein endothelial cells (HUVECs). Among the test compounds, the dimer compounds 5v and 5w exhibited the most potent antioxidant activity with significant ROS suppression being observed. Both compounds markedly inhibited the H2O2-induced changes in various biochemical substances, such as superoxide dismutase (SOD), malonyldialdehyde (MDA), nitric oxide (NO), and lactic dehydrogenase (LDH), which were superior to that of the positive control vitamin E. Further more, they did not produce any obvious cytotoxicity, but increased the viability of HUVECs injured by H2O2 in a dose-dependent manner. Additionally, compound 5w, designed as a prodrug-like compound, showed improved stability relative to compound 4 in vitro.

Prevalence and characterization of plasmid-mediated blaESBL with their genetic environment in Escherichia coli and Klebsiella pneumoniae in patients with pneumonia.

BACKGROUND: The extended spectrum ß-lactamase (ESBL)-producing Escherichia coli (E. coli) and Klebsiella pneumoniae (K. pneumoniae) are the major pathogens causing pneumonia and have a significant impact on the clinical course. Limited data exist on molecular characterization of ESBL-producing E. coli and K. pneumoniae that cause pneumonia. The aim of this study was to investigate the comprehensive multilevel characteristics of E. coli and K. pneumoniae causing pneumonia in China for the first time. METHODS: E. coli (17) and K. pneumoniae (21) isolates responsible for pneumonia were isolated from 1270 specimens collected in a prospective multi-center study in eight teaching hospitals in China from June to December in 2007. The susceptibilities, ESBL confirmation, sequence typing, blaCTX-M and blaSHV genes, their genetic environment and plasmid Inc/rep types were determined. RESULTS: Sixteen E. coli (94.1%) and eleven K. pneumoniae (52.4%) isolates were ESBL producers. About 77.8% and 66.7% of them were resistance to ciprofloxacin and levofloxacin, and 100% were susceptible to imipenem. The most prevalent ESBL gene was CTX-M-14, followed by SHV-2, CTX-M-15, CTX-M-3, CTX-M-65, SHV-12, SHV-26 and SHV-28. SHV-1 and SHV-11 were also detected and coexisted with blaCTX-Ms in five strains, and three strains contained only SHV-1. All CTX-M-14 were detected ISEcp1 upstream and nine were found IS903 downstream and the majority of them (64.3%) were carried by IncF plasmids. All blaSHV were flanked by recF and deoR, located on IncF, IncN, IncX and IncH plasmids. Two SHV-2, one SHV-1 and the only SHV-28 were further preceded by IS26. Genes lacY and lacZ were detected at further upstream of two blaSHV-1. The K. pneumoniae carrying SHV-28 was susceptible to ß-lactams, and no mutations or deletions in gene or promoter sequences were identified to account for susceptibility. Multilocus sequence typing experiments showed the ESBL-producing strains were genetically diverse. CONCLUSIONS: The rate of occurrence of blaESBL in E. coli and K. pneumoniae causing pneumonia was high, and blaCTX-M-14 was dominant and probably mobilized by ISEcp1 mainly on IncF plasmids. Importantly, unexpressed blaESBL genes may occur in susceptible isolates and hence may have clinical implications.

Drug-resistant genes carried by Acinetobacter baumanii isolated from patients with lower respiratory tract infection.

BACKGROUND: Acinetobacter baumanii (A. baumanii ) remains an important microbial pathogen resulting in nosocomial acquired infections with significant morbidity and mortality. The mechanism by which nosocomial bacteria, like A. baumanii, attain multidrug resistance to antibiotics is of considerable interest. The aim in this study was to investigate the spread status of antibiotic resistance genes, such as multiple ß-lactamase genes and aminoglycoside-modifying enzyme genes, from A. baumanii strains isolated from patients with lower respiratory tract infections (LRTIs). METHODS: Two thousand six hundred and ninety-eight sputum or the bronchoalveolar lavage samples from inpatients with LRTIs were collected in 21 hospitals in the mainland of China from November 2007 to February 2009. All samples were routinely inoculated. The isolated bacterial strains and their susceptibility were analyzed via VITEK-2 expert system. Several kinds of antibiotic resistant genes were further differentiated via polymerase chain reaction and sequencing methods. RESULTS: Totally, 39 A. baumanii strains were isolated from 2698 sputum or bronchoalveolar lavage samples. There was not only a high resistant rate of the isolated A. baumanii strains to ampicillin and first- and second-generation cephalosporins (94.87%, 100% and 97.44%, respectively), but also to the third-generation cephalosporins (ceftriaxone at 92.31%, ceftazidine at 51.28%) and imipenem (43.59%) as well. The lowest antibiotic resistance rate of 20.51% was found to amikacin. The OXA-23 gene was identified in 17 strains of A. baumanii, and the AmpC gene in 23 strains. The TEM-1 gene was carried in 15 strains. PER-1 and SHV-2 genes were detected in two different strains. Aminoglycoside-modifying enzyme gene aac-3-Ia was found in 23 strains, and the aac-6'-Ib gene in 19 strains. aac-3-Ia and aac-6'-Ib genes hibernated in three A. baumanii strains that showed no drug-resistant phenotype. CONCLUSIONS: A. baumanii can carry multiple drug-resistant genes at the same time and result in multi-drug resistance. Aminoglycoside-modifying enzyme genes could be hibernating in aminoglycoside sensitive strains without expressing their phenotype.

[The analysis of "alpha" dominant mutation of hepatitis B virus in community-based Zhengding area, Hebei province].

OBJECTIVE: To determine the "alpha"dominant mutation of hepatitis B virus (HBV) in community-based Zhengding. Analysis the role of the newborn hepatitis B vaccination on the mutation. METHODS: Based on the national surveillance of hepatitis B, 11,478 people's sera were collected and tested by SPRIA with kits. Collect people's sera with positive HBsAg and amplify the S gene. Sequencing and clastwaling them with the standard sequences. RESULTS: Overall, HBV DNA was successfully amplified and sequenced in 434 of 443 samples. 6.7% samples mutated in HBV "alpha" dominant region. The difference between the mutation ratio of the two loops of HBV "alpha" dominant between the people born before and after the year 1986 has no significance. CONCLUSION: There were HBV "alpha" dominant mutant virus in the local area with a low infection rate in the population born after the year 1986. It could not explain the newborn hepatitis B vaccination can induce the prevalence of the "alpha" dominant mutate HBV.

Potential cost-effectiveness of a rotavirus immunization program in rural China.

BACKGROUND: To assess the incidence and economic burden of rotavirus diarrhea and the potential cost-effectiveness of a rotavirus immunization program in rural Zhengding County in Hebei Province, China. METHODS: Population-based surveillance was conducted during the peak season for diarrhea among children who were <5 years of age in Zhengding County from 14 October 2004 through 19 January 2005. The cost of illness was measured from the perspectives of both patient and society. A decision-analytic model was applied to the cost-effectiveness analysis using real data derived from surveillance and from a cost-of-illness study. RESULTS: During the surveillance period, 500 episodes of diarrhea were registered. Of these 500 episodes, 125 (25%) occurred in patients who were positive for rotavirus. Of these 125 episodes, 63 (50%) occurred in patients who were hospitalized. The overall incidence rate of rotavirus infection was 61.4 cases per 1000 children per year during the 14-week epidemic season. For a Chinese cohort of 5000 newborns, a universal rotavirus immunization program would prevent 1764 cases of rotavirus diarrhea, averting 882 hospitalizations of patients

[The immunological effects of three doses of a live attenuated hepatitis A vaccine (H2 strain) in 8 years].

OBJECTIVE: To observe the immunological effects of three doses of H2 strain live attenuated hepatitis A vaccine 8 years after the administration and to compare with that of one dose of the vaccine. METHODS: In a country area, 110 children of 1 to 7 years old susceptible to HAV were screened and administered with one dose of the vaccine, as group B; Group A were 42 children from one of the villages and administered with 3 doses of the vaccine according to 0, 2, 6 month schedule. Blood samples were taken for the children 1, 2, 6, 7, 8, 12, 24, 36 and 96 months after the administrations respectively and detected for anti-HAV antibody. RESULTS: For group B, the sero conversion rate of anti-HAV and GMC reached peak at 92.2% and 126.2 mIU/ml respectively, and then, began to drop with time; For group A, after 2 dose of the vaccine, the sero-conversion rate reached 100%, and the GMC reached peak of 2 739 mIU/ml one month after the third dose at 7 months. So that, group A has a better short-term immunological effects than that of group B. During 36 through 96 months, the anti-HAV positive rate in group B was 75%-71% and 80-89 mIU/ml respectively, and comparatively in group A were 100% and 918.2-480.6 mIU/ml respectively. The differences between group A and B were significantly important. CONCLUSION: A 3-dose schedule administration of H2 strain live attenuated hepatitis A vaccine has better immunological effects than 1-dose schedule in 8years and further observations are needed.

Long-term immunogenicity after single and booster dose of a live attenuated hepatitis A vaccine: results from 8-year follow-up.

Live, attenuated hepatitis A vaccines are used widely in China but there is uncertainty regarding the persistence of vaccine-induced anti-HAV antibodies after single dose and booster dose administrated at month 12. A large scale clinical trial to evaluate the live, attenuated hepatitis A vaccine was conducted in Hebei province between 1996 and 1999. Five years after the trials, children in single dose and booster dose groups were bled and followed. Seventy two percent (61/85) of children who received a single trial dose had detectable anti-HAV antibodies for 96 months (GMC at 96 months: 89.0 mIU/mL). In the booster group 98% (48/49) children remained anti-HAV positive with GMC of 262.8 mIU/mL at month 96. The reinjection with live attenuated HAV vaccine can elicit a booster effect. Results from single dose group seems not to support the need for booster doses of live attenuated hepatitis A vaccine in immunocompetent individuals regarding the persisting anti-HAV and anamnestic response of a single dose vaccine. Continued monitoring of anti-HAV antibodies is needed for a rational hepatitis A immunization strategy in China.

Occurrence of shigellosis in the young and elderly in rural China: results of a 12-month population-based surveillance study.

In 2002, population- and treatment center-based surveillance was used to study the disease burden of shigellosis in rural Hebei Province in the People's Republic of China. A total of 10,105 children with diarrhea or dysentery were enrolled. Infants were treated most frequently for diarrhea (1,388/1,000/year) followed by children < or = 5 years old (618/1,000/year). Shigellosis was treated most often in children 3-4 years old (32/1,000/year) and people > 60 years of age (7/1,000/year). Fifty-six percent (184 of 331) Shigella isolates were detected in patients who had non-bloody diarrhea. Shigella flexneri was identified in 93% of 306 isolates. The most common S. flexneri serotypes were 1a (34%), X (33%), and 2a (28%). More than 90% of the Shigella isolates were resistant to cotrimoxazole and nalidixic acid, but remained susceptible to ciprofloxacin, norfloxacin, and gentamicin. Widespread resistance to antibiotics adds urgency to the development and use of vaccines to control shigellosis.

Incidence of diarrhea caused by rotavirus infections in rural Zhengding, China: prospective, population-based surveillance.

Rotavirus is the pathogen most commonly associated with severe gastroenteritis in young children in the People's Republic of China, yet there are few population-based data on the incidence of rotavirus infection. The present study investigated the burden of rotavirus diarrhea and rotavirus infections in rural China, according to age. Population-based surveillance was used to study the incidence of rotavirus infection among children <5 years of age in 4 townships of Zhengding County, Hebei Province, China. The total population in the catchment area in 2002 was 75,630 individuals, including 2997 children aged <5 years. Stool samples were obtained and were tested for rotavirus antigen by use of an enzyme-linked immunosorbent assay. During 2002, a total of 2010 cases of diarrhea were detected among children <5 years of age. The incidence of treated cases of diarrhea was 671 cases/1000 children/year for children <5 years of age, and it was highest for children <12 months of age (1467 cases/1000 children/year). The estimated incidence of rotavirus infection was 151 cases/1000 children/year for children <5 years of age. The highest incidence of rotavirus infection was among children aged 1-2 years (340 cases/1000 children/year). Widespread immunization of children against rotavirus before 6 months of age should be considered for the control of rotavirus diarrhea.

[Construction of the mutants of rice nonspecific lipid transfer protein and expression comparison in two kinds of thioredoxin fusion expression vectors].

Five structural important residues of rice nonspecific lipid transfer protein LTP110 were mutated by site-directed mutagenesis. Sequence results showed that they were all mutated successfully. After trying various E. coli expression systems, thioredoxin fusion expression system was found to be a proper system to express wild type and mutant LTP110. cDNA sequences encoding wild type LTP110 and the mutants Y17A, P72L, R46A, D43A, C50A were cloned into two kinds of thioredoxin fusion expression vectors. The expression results were compared. In pTrxFus/GI724 expression system, wild type LTP110 and the mutants Y17A, P72L, R46A could be expressed at low level while D43A and C50A could not be expressed normally; in pET32a(+)/BL21 (DE3) trxB- expression system, wild type LTP110 and all mutant proteins could be expressed very well and the levels were higher than that in pTrxFus/GI724 system. LTP110 fusion protein expressed in pET32a(+) vector was purified and its activity was checked by fluorescence labeled fatty acid. Results indicated that the recombinant LTP110 fusion protein has lipid binding activity. This work provides good basis for the further study.

[Expression, purification and function of rice nonspecific lipid transfer protein].

Plant nonspecific lipid transfer protein(nsLTP) is a class of protein which has in vitro lipid transferring activity between biomembranes. In order to study the antimicrobial function of rice nonspecific lipid transfer protein, a gene LTP110 encoding rice nsLTP was cloned into ThioFusion expression vector pET32a (+) and expressed in host strain Bl21(DE3)trxB-. After induction by IPTG at 30 degrees C for 5 h, the fusion protein thio-LTP110 was in large amount produced. The expressed protein was purified by Ni2(+)-chelating Sepharose fast flow column, then digested by enterokinase. By passing through nickel affinity column again, the cleavage product, LTP110, was obtained. CD spectrum scanning from 185 nm to 250 nm showed that the recombinant protein LTP110 had similar secondary structure with the nsLTP purified from rice etiolated seedlings. Activity determination by fluorescent lipid P-96 showed that it had lipid binding activity. Microbial inhibition test results revealed that LTP110 deterred germination of the spores of rice pathogen P. oryzae, showing it might be involved in plant microbial resistance function. Therefore, it has the potential to be used in plant transgene engineering to improve plant resistance.