Clade 2.3.4.4 H5 chimeric cold-adapted attenuated influenza vaccines induced cross-reactive protection in mice and ferrets.

IMPORTANCE: Clade 2.3.4.4 H5Nx avian influenza viruses (AIVs) have circulated globally and caused substantial economic loss. Increasing numbers of humans have been infected with Clade 2.3.4.4 H5N6 AIVs in recent years. Only a few human influenza vaccines have been licensed to date. However, the licensed live attenuated influenza virus vaccine exhibited the potential of being recombinant with the wild-type influenza A virus (IAV). Therefore, we developed a chimeric cold-adapted attenuated influenza vaccine based on the Clade 2.3.4.4 H5 AIVs. These H5 vaccines demonstrate the advantage of being non-recombinant with circulated IAVs in the future influenza vaccine study. The findings of our current study reveal that these H5 vaccines can induce cross-reactive protective efficacy in mice and ferrets. Our H5 vaccines may provide a novel option for developing human-infected Clade 2.3.4.4 H5 AIV vaccines.

Improved structure stability and performance of a LiFeSO4F cathode material for lithium-ion batteries by magnesium substitution.

Tavorite LiFeSO4F with high Li-ion conductivity has been considered a promising alternative to LiFePO4. However, its poor cycle stability and low electronic conductivity limit the practical application of Tavorite LiFeSO4F. In the present study, we employ a solvothermal method to produce magnesium-substitution LiMgxFe1-xSO4F (x = 0, 0.02, 0.04) cathode materials in which the Mg substitutes the Fe(2) sites. The first-principles calculations demonstrate that Mg-substitution could reduce the bandgap of LiFeSO4F and increase its electronic conductivity to 2.5 × 10-11 S cm-1. Meanwhile, CI-NEB and BV calculations reveal that the diffusion energy barrier of lithium along the (100) direction after Mg substitution is lower than the pristine sample, and the electrochemical inactive Mg2+ could improve the structure stability. The results show that the Mg-substituted LiFeSO4F exhibits enhanced cycle stability and rate performance compared with the pristine LiFeSO4F, suggesting that the use of electrochemically inactive ion substitution may be critical for the development of high-performance LiFeSO4F cathode materials for lithium-ion batteries.

Humanized mice for investigating SARS-CoV-2 lung infection and associated human immune responses.

There is an urgent need for animal models of coronavirus disease 2019 to study immunopathogenesis and test therapeutic intervenes. In this study, we showed that NOD/SCID IL2rg-/- (NSG) mice engrafted with human lung (HL) tissue (NSG-L mice) could be infected efficiently by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and that live virus capable of infecting Vero cells was found in the HL grafts and multiple organs from infected NSG-L mice. RNA-Sequencing identified a series of differentially expressed genes, which are enriched in viral defense responses, chemotaxis, IFN stimulation and pulmonary fibrosis, between HL grafts from infected and control NSG-L mice. Furthermore, when infected with SARS-CoV-2, humanized mice with both human immune system (HIS) and autologous HL grafts (HISL mice) had bodyweight loss and hemorrhage and immune cell infiltration in HL grafts, which were not observed in immunodeficient NSG-L mice, indicating the development of anti-viral immune responses in these mice. In support of this possibility, the infected HISL mice showed bodyweight recovery and lack of detectable live virus at the later time. These results demonstrate that NSG-L and HISL mice are susceptible to SARS-CoV-2 infection, offering a useful in vivo model for studying SARS-CoV-2 infection and the associated immune response and immunopathology, and testing anti-SARS-CoV-2 therapies.

SARS-CoV-2 Virus Culture, Genomic and Subgenomic RNA Load, and Rapid Antigen Test in Experimentally Infected Syrian Hamsters.

The duration of SARS-CoV-2 genomic RNA shedding is much longer than that of infectious SARS-CoV-2 in most COVID-19 patients. It is very important to determine the relationship between test results and infectivity for efficient isolation, contact tracing, and post-isolation. We characterized the duration of viable SARS-CoV-2, viral genomic and subgenomic RNA (gRNA and sgRNA), and rapid antigen test positivity in nasal washes, oropharyngeal swabs, and feces of experimentally infected Syrian hamsters. The duration of viral genomic RNA shedding is longer than that of viral subgenomic RNA, and far longer than those of rapid antigen test (RAgT) and viral culture positivity. The rapid antigen test results were strongly correlated with the viral culture results. The trend of subgenomic RNA is similar to that of genomic RNA, and furthermore, the subgenomic RNA load is highly correlated with the genomic RNA load. IMPORTANCE Our findings highlight the high correlation between rapid antigen test and virus culture results. The rapid antigen test would be an important supplement to real-time reverse transcription-RCR (RT-PCR) in early COVID-19 screening and in shortening the isolation period of COVID-19 patients. Because the subgenomic RNA load can be predicted from the genomic RNA load, measuring sgRNA does not add more benefit to determining infectivity than a threshold determined for gRNA based on viral culture.

Evidence for human infection with avian influenza A(H9N2) virus via environmental transmission inside live poultry market in Xiamen, China.

H9N2 avian influenza virus (AIV) has become prevalent in the live poultry market (LPM) worldwide, and environmental transmission mode is an important way for AIVs to infect human beings in the LPM. To find evidence of human infection with the influenza A(H9N2) virus via environmental contamination, we evaluated one human isolate and three environmental isolates inside LPMs in Xiamen, China. The phylogeny, transmissibility, and pathogenicity of the four isolates were sorted out systematically. As for the H9N2 virus, which evolved alongside the "Avian-Environment-Human" spreading chain in LPMs from the summer of 2019 to the summer of 2020, its overall efficiency of contact and aerosol transmissibility improved, which might contribute to the increasing probability of human infection. This study indicated that environmental exposure might act as an important source of human infection in LPMs.

Meningitis caused by oral anaerobes detected using mNGS tool: a case report and review of literature.

BACKGROUND: Bacterial meningitis is a central nervous system (CNS) infection disease of the meninges and brain parenchyma caused by the bacteria. Few cases of meningitis related to oral anaerobes have been reported in the literature. Here, we report a case of meningitis in a middle-aged woman, caused by oral anaerobes. CASE PRESENTATION: A 58-year-old woman was admitted to hospital with fever, headache for 21 days and left limb weakness for 2 days. The blood cell counts (11.73 × 109/L), neutrophil counts (9.22 × 109/L) and high-sensitivity C-reactive protein levels (> 5.00 mg/L) were elevated. The brain computerized tomography (CT) scanning indicated the new right thalamus infarct. The brain cranial-enhanced magnetic resonance imaging (MRI) showed the right lateral paraventricular and right thalamic infarct, and abnormal signal in occipital horns of bilateral lateral ventricles were increased. In addition, the brain enhanced nuclear magnetic resonance (NMR) scanning suggested that meninges were thickened and enhanced at the base of the brain, with meningitis changes. The neck CT angiography (CTA) revealed arteriosclerotic changes. The metagenomic next-generation sequencing (mNGS) revealed Eubacterium brachy, Porphyromonas gingivalis, Fusobacterium nucleatum and Torque teno virus in her cerebrospinal fluid (CSF). The patient was diagnosed with purulent meningitis caused by infection of oral anaerobes, and treated with mannitol, ceftriaxone and vancomycin. Her symptoms alleviated. Subsequently, she was transferred to the infectious department and treated with ceftriaxone plus metronidazole (anti-anaerobes) and mannitol (reduce intracranial pressure). Her symptoms improved and currently received rehabilitation treatment. CONCLUSION: We herein report a rare case involving meningitis caused by infection of oral anaerobes. The mNGS can accurately detect the pathogens of infectious diseases.

Real-Time Reverse Transcription Recombinase-Aided Amplification Assay for Rapid Amplification of the N Gene of SARS-CoV-2.

COVID-19 was officially declared a global pandemic disease on 11 March 2020, with severe implications for healthcare systems, economic activity, and human life worldwide. Fast and sensitive amplification of the severe acute respiratory syndrome virus 2 (SARS-CoV-2) nucleic acids is critical for controlling the spread of this disease. Here, a real-time reverse transcription recombinase-aided amplification (RT-RAA) assay, targeting conserved positions in the nucleocapsid protein gene (N gene) of SARS-CoV-2, was successfully established for SARS-CoV-2. The assay was specific to SARS-CoV-2, and there was no cross-reaction with other important viruses. The sensitivity of the real-time RT-RAA assay was 142 copies per reaction at 95% probability. Furthermore, 100% concordance between the real-time RT-RAA and RT-qPCR assays was achieved after testing 72 clinical specimens. Further linear regression analysis indicated a significant correlation between the real-time RT-RAA and RT-qPCR assays with an R2 value of 0.8149 (p < 0.0001). In addition, the amplicons of the real-time RT-RAA assay could be directly visualized by a portable blue light instrument, making it suitable for the rapid amplification of SARS-CoV-2 in resource-limited settings. Therefore, the real-time RT-RAA method allows the specific, sensitive, simple, rapid, and reliable detection of SARS-CoV-2.

SARS-CoV-2 Aerosol Exhaled by Experimentally Infected Cynomolgus Monkeys.

We analyzed size of severe acute respiratory coronavirus 2 (SARS-CoV-2) aerosol particles shed by experimentally infected cynomolgus monkeys. Most exhaled particles were small, and virus was mainly released early during infection. By postinfection day 6, no virus was detected in breath, but air in the isolator contained large quantities of aerosolized virus.

Photoacoustic-fluorescence microendoscopy in vivo.

A miniature endoscope capable of imaging multiple tissue contrasts in high resolution is highly attractive, because it can provide complementary and detailed tissue information of internal organs. Here we present a photoacoustic (PA)-fluorescence (FL) endoscope for optical-resolution PA microscopy (PAM) and FL microscopy (FLM). The endoscope with a diameter of 2.8 mm achieves high lateral resolutions of 5.5 and 6.3 µm for PAM and FLM modes, respectively. In vivo imaging of zebrafish larvae and a mouse ear is conducted, and high-quality images are obtained. Additionally, in vivo endoscopic imaging of a rat rectum is demonstrated, showing the endoscopic imaging capability of our endoscope. By providing dual contrasts with high resolution, the endoscope may open up new opportunities for clinical endoscopic imaging applications.

Genetic diversity and evolutionary dynamics of Ebola virus in Sierra Leone.

A novel Ebola virus (EBOV) first identified in March 2014 has infected more than 25,000 people in West Africa, resulting in more than 10,000 deaths. Preliminary analyses of genome sequences of 81 EBOV collected from March to June 2014 from Guinea and Sierra Leone suggest that the 2014 EBOV originated from an independent transmission event from its natural reservoir followed by sustained human-to-human infections. It has been reported that the EBOV genome variation might have an effect on the efficacy of sequence-based virus detection and candidate therapeutics. However, only limited viral information has been available since July 2014, when the outbreak entered a rapid growth phase. Here we describe 175 full-length EBOV genome sequences from five severely stricken districts in Sierra Leone from 28 September to 11 November 2014. We found that the 2014 EBOV has become more phylogenetically and genetically diverse from July to November 2014, characterized by the emergence of multiple novel lineages. The substitution rate for the 2014 EBOV was estimated to be 1.23 × 10(-3) substitutions per site per year (95% highest posterior density interval, 1.04 × 10(-3) to 1.41 × 10(-3) substitutions per site per year), approximating to that observed between previous EBOV outbreaks. The sharp increase in genetic diversity of the 2014 EBOV warrants extensive EBOV surveillance in Sierra Leone, Guinea and Liberia to better understand the viral evolution and transmission dynamics of the ongoing outbreak. These data will facilitate the international efforts to develop vaccines and therapeutics.

Influence of Storage Conditions on SARS-CoV-2 Nucleic Acid Detection in Throat Swabs.

The detection of SARS-CoV-2 infection is the premise of quarantine. In many countries or areas, samples need to be shipped or inactivated before SARS-CoV-2 testing. In this study, we checked the influence of sample storage conditions on SARS-CoV-2 nucleic acid testing results, including sample inactivation time, storage temperature, and storage time. All of these conditions caused an increase in the cycle threshold values of the nucleic acid tests and led to the misclassification of at least 10.2% of positive cases as negative or suspected. The results highlight the importance of immediate testing of samples for SARS-CoV-2 nucleic acid detection.

Aerosol and Surface Distribution of Severe Acute Respiratory Syndrome Coronavirus 2 in Hospital Wards, Wuhan, China, 2020.

To determine distribution of severe acute respiratory syndrome coronavirus 2 in hospital wards in Wuhan, China, we tested air and surface samples. Contamination was greater in intensive care units than general wards. Virus was widely distributed on floors, computer mice, trash cans, and sickbed handrails and was detected in air ≈4 m from patients.

Transmission and prevention of SARS-CoV-2.

The coronavirus disease 2019 (COVID-19), caused by a novel virus of the ß-coronavirus genus (SARS-CoV-2), has been spreading globally. As of July 2020, there have been more than 17 million cases worldwide. Determining multiple transmission routes of SARS-CoV-2 is critical to improving safety practices for the public and stemming the spread of SARS-CoV-2 effectively. This article mainly focuses on published studies on the transmission routes of SARS-CoV-2 including contact transmission, droplet transmission, aerosol transmission and fecal-oral transmission, as well as related research approaches, such as epidemiological investigations, environmental sampling in hospitals and laboratories and animal models. We also provide four specific recommendations for the prevention and control of SARS-CoV-2 that may help reduce the risk of SARS-CoV-2 infection under different environmental conditions. First, social distancing, rational use of face masks and respirators, eye protection, and hand disinfection for medical staff and the general public deserve further attention and promotion. Second, aerodynamic characteristics, such as size distribution, release regularity, aerosol diffusion, survival and decline, infectious dose and spread distance, still require further investigation in order to identify the transmissibility of COVID-19. Third, background monitoring of the distribution of pathogenic microorganisms and environmental disinfection in crowded public places, such as railway stations, schools, hospitals and other densely populated areas, can give early warning of outbreaks and curb the transmission routes of SARS-CoV-2 in those high-risk areas. Forth, establishing novel predictive models can help us to not only assess transmission and impacts in communities, but also better implement corresponding emergency response measures.

Identification of cellular microRNA miR-188-3p with broad-spectrum anti-influenza A virus activity.

BACKGROUND: Influenza A virus (IAV) continues to pose serious threats to public health. The current prophylaxis and therapeutic interventions for IAV requires frequent changes due to the continuous antigenic drift and antigenic shift of IAV. Emerging evidence indicates that the host microRNAs (miRNAs) play critical roles in intricate host-pathogen interaction networks. Cellular miRNAs may directly target virus to inhibit its infection and be developed as potential anti-virus drugs. METHODS: In this study, we established a broad-spectrum anti-IAV miRNA screening method using miRanda software. The screened miRNAs were further verified by luciferase assay, viral protein expression assay and virus replication assay. RESULTS: Five cellular miRNAs (miR-188-3p, miR-345-5p, miR-3183, miR-15-3p and miR-769-3p), targeting 99.96, 95.31, 92.9, 94.58 and 97.24% of human IAV strains recorded in NCBI, respectively, were chosen for further experimental verification. Finally, we found that miR-188-3p downregulated PB2 expression at both mRNA and protein levels by directly targeted the predicted sites on PB2 and effectively inhibited the replication of IAV (H1N1, H5N6 and H7N9) in A549 cells. CONCLUSIONS: This is the first report screening cellular miRNAs that broad-spectrum inhibiting IAV infection. These findings suggested that cellular miR-188-3p could be used for RNAi-mediated anti-IAV therapeutic strategies.

Patterns and trends of alcohol consumption in rural and urban areas of China: findings from the China Kadoorie Biobank.

BACKGROUND: In China, alcohol consumption has increased significantly in recent decades. Little evidence exists, however, about temporal trends in levels and patterns of alcohol consumption and associated factors in adult populations. METHODS: In 2004-08, the China Kadoorie Biobank recruited ~ 512,000 adults (41% men, mean age 52 years [SD 10.7]) from 10 (5 urban, 5 rural) geographically diverse regions across China, with ~ 25,000 randomly selected participants resurveyed in 2013-14. The self-reported prevalence and patterns (e.g., amount, beverage type, heavy drinking episodes) of alcohol drinking at baseline and resurvey were compared and related to socio-demographic, health and other factors. RESULTS: At baseline, 33% of men drank alcohol at least weekly (i.e., current regular), compared to only 2% of women. In men, current regular drinking was more common in urban (38%) than in rural (29%) areas at baseline. Among men, the proportion of current regular drinkers slightly decreased at resurvey (33% baseline vs. 29% resurvey), while the proportion of ex-regular drinkers slightly increased (4% vs. 6%), particularly among older men, with more than half of ex-regular drinkers stopping for health reasons. Among current regular drinkers, the proportion engaging in heavy episodic drinking (i.e., > 60 g/session) increased (30% baseline vs. 35% resurvey) in both rural (29% vs. 33%) and urban (31% vs. 36%) areas, particularly among younger men born in the 1970s (41% vs. 47%). Alcohol intake involved primarily spirits, at both baseline and resurvey. Those engaging in heavy drinking episodes tended to have multiple other health-related risk factors (e.g., regular smoking, low fruit intake, low physical activity and hypertension). CONCLUSIONS: Among Chinese men, the proportion of drinkers engaging in harmful drinking behaviours increased in the past decade, particularly among younger men. Harmful drinking patterns tended to cluster with other unhealthy lifestyles and health-related risk factors.

Miniature fiber-optic high-intensity focused ultrasound device using a candle soot nanoparticles-polydimethylsiloxane composites-coated photoacoustic lens.

We present a miniature fiber-optic ultrasound transmitter for generating high-intensity focused ultrasound (HIFU) based on photoacoustic transduction. The HIFU device consists of a fiber and a photoacoustic lens. We develop a simple fabrication procedure for making the photoacoustic lens, which is coated with candle soot nanoparticles-polydimethylsiloxane composites. The fiber is used to deliver pulsed laser for photoacoustic excitation, which facilitates the use of the HIFU device by eliminating the need of free-space optical alignment. The HIFU device (6.5 mm in diameter) produces focused acoustic pressures up to >30 MPa in peak positive with a tight -6-dB focal volume of ~100 µm and ~500 µm in the lateral and axial directions, respectively. Acoustic cavitation induced by the HIFU device is demonstrated. The miniature HIFU device facilitates handheld operation. It holds promise for clinical applications in intraoperative high-precision HIFU therapy. It can even be used for intracavitary therapy with further miniaturization.

Miniature probe for in vivo optical- and acoustic-resolution photoacoustic microscopy.

We present a miniature probe capable of both optical-resolution (OR) and acoustic-resolution (AR) photoacoustic microscopy. A gradient-index-lens fiber and a multimode fiber are used to deliver light for OR and AR illumination, respectively. The probe achieves lateral resolution of 3.1 µm for OR mode and 46-249 µm (at depth of 1.2-4.3 mm) for AR mode, respectively. The size of the probe attains 3.7 mm in diameter, which can be used for endoscopic applications. In vivo imaging of several different parts of a mouse demonstrates the excellent imaging ability of the probe.

A Rapid Screen for Host-Encoded miRNAs with Inhibitory Effects against Ebola Virus Using a Transcription- and Replication-Competent Virus-Like Particle System.

MicroRNAs (miRNAs) may become efficient antiviral agents against the Ebola virus (EBOV) targeting viral genomic RNAs or transcripts. We previously conducted a genome-wide search for differentially expressed miRNAs during viral replication and transcription. In this study, we established a rapid screen for miRNAs with inhibitory effects against EBOV using a tetracistronic transcription- and replication-competent virus-like particle (trVLP) system. This system uses a minigenome comprising an EBOV leader region, luciferase reporter, VP40, GP, VP24, EBOV trailer region, and three noncoding regions from the EBOV genome and can be used to model the life cycle of EBOV under biosafety level (BSL) 2 conditions. Informatic analysis was performed to select up-regulated miRNAs targeting the coding regions of the minigenome with the highest binding energy to perform inhibitory effect screening. Among these miRNAs, miR-150-3p had the most significant inhibitory effect. Reverse transcription polymerase chain reaction (RT-PCR), Western blot, and double fluorescence reporter experiments demonstrated that miR-150-3p inhibited the reproduction of trVLPs via the regulation of GP and VP40 expression by directly targeting the coding regions of GP and VP40. This novel, rapid, and convenient screening method will efficiently facilitate the exploration of miRNAs against EBOV under BSL-2 conditions.

Miniature all-optical probe for large synthetic aperture photoacoustic-ultrasound imaging.

A miniature all-optical probe for high-resolution photoacoustic (PA)-ultrasound (US) imaging using a large synthetic aperture is developed. The probe consists of three optical fibers for PA excitation, US generation, and detection of acoustic waves, respectively. The fiber for PA excitation has a large numerical aperture (NA) for wide-angle laser illumination. On the other hand, the fiber with a carbon black-polydimethylsiloxane composite coated on the end face of the optical fiber is used for wide-angle US transmission through laser-US conversion. Both the excited PA and backscattered US signals are detected by a fiber-tip Fabry-Perot cavity for wide-angle acoustic detection. The probe outer diameter is only ~2 mm. The synergy of the three optical fibers makes a large-NA synthetic aperture focusing technique for high-resolution PA and US imaging possible. High PA lateral resolutions of 104-154 µm and high US lateral resolutions of 64-112 µm over a depth range of > 4 mm are obtained. Compared with other existing miniature PA-US probes, to our knowledge, our probe achieves by far the best performance in terms of lateral resolutions and imaging depth range. The constructed probe has potential for endoscopic and intravascular imaging applications that require PA and US contrasts with high resolutions over a large depth range.

Photoacoustic microscopy in vivo using synthetic-aperture focusing technique combined with three-dimensional deconvolution.

Acoustic-resolution photoacoustic microscopy (ARPAM) plays an important role in studying the microcirculation system of biological tissues with deep penetration. High lateral resolution of ARPAM is achieved by using a high numerical aperture acoustic transducer. The deteriorated lateral resolution in the out-of-focus region can be alleviated by synthetic aperture focusing technique (SAFT). Previously, we reported a three-dimensional (3D) deconvolution ARPAM to improve both lateral and axial resolutions in the focus region. In this study, we present our extension of resolution enhancement to the out-of-focus region based on two-dimensional SAFT combined with the 3D deconvolution (SAFT+Deconv). In both the focus and out-of-focus regions, depth-independent lateral resolution provided by SAFT, together with inherently depth-independent axial resolution, ensures a depth-independent point spread function for 3D deconvolution algorithm. Imaging of 10 µm polymer beads shows that SAFT+Deconv ARPAM improves the -6 dB lateral resolutions from 65-700 µm to 20-29 µm, and the -6 dB axial resolutions from 35-42 µm to 12-19 µm in an extended depth of focus (DOF) of ∼2 mm. The signal-to-noise ratio is also increased by 6-30 dB. The resolution enhancement in three dimensions is validated by in vivo imaging of a mouse's dorsal subcutaneous microvasculature. Our results suggest that SAFT+Deconv ARPAM may allow fine spatial resolution with deep penetration and extended DOF for biomedical photoacoustic applications.