Antimicrobial resistance and population genomics of emerging multidrug-resistant Salmonella 4,[5],12:i:- in Guangdong, China.

Salmonella 4,[5],12:i:-, a monophasic variant of Salmonella Typhimurium, has emerged as a global cause of multidrug-resistant salmonellosis and has become endemic in many developing and developed countries, especially in China. Here, we have sequenced 352 clinical isolates in Guangdong, China, during 2009-2019 and performed a large-scale collection of Salmonella 4,[5],12:i:- with whole genome sequencing (WGS) data across the globe, to better understand the population structure, antimicrobial resistance (AMR) genomic characterization, and transmission routes of Salmonella 4,[5],12:i:- across Guangdong. Salmonella 4,[5],12:i:- strains showed broad genetic diversity; Guangdong isolates were found to be widely distributed among the global lineages. Of note, we identified the formation of a novel Guangdong clade (Bayesian analysis of population structure lineage 1 [BAPS1]) genetically diversified from the global isolates and likely emerged around 1990s. BAPS1 exhibits unique genomic features, including large pan-genome, decreased ciprofloxacin susceptibility due to mutation in gyrA and carriage of plasmid-mediated quinolone resistance (PMQR) genes, and the multidrug-resistant IncHI2 plasmid. Furthermore, high genetic similarity was found between strains collected from Guangdong, Europe, and North America, indicating the association with multiple introductions from overseas. These results suggested that global dissemination and local clonal expansion simultaneously occurred in Guangdong, China, and horizontally acquired resistance to first-line and last-line antimicrobials at local level, underlying emergences of extensive drug and pan-drug resistance. Our findings have increased the knowledge of global and local epidemics of Salmonella 4,[5],12:i:- in Guangdong, China, and provided a comprehensive baseline data set essential for future molecular surveillance.IMPORTANCESalmonella 4,[5],12:i:- has been regarded as the predominant pandemic serotype causing diarrheal diseases globally, while multidrug resistance (MDR) constitutes great public health concerns. This study provided a detailed and comprehensive genome-scale analysis of this important Salmonella serovar in the past decade in Guangdong, China. Our results revealed the complexity of two distinct transmission modes, namely global transmission and local expansion, circulating in Guangdong over a decade. Using phylogeography models, the origin of Salmonella 4,[5],12:i:- was predicted from two aspects, year and country, that is, Salmonella 4,[5],12:i:- emerged in 1983, and was introduced from the UK, and subsequently differentiated into the local endemic lineage circa 1991. Additionally, based on the pan-genome analysis, it was found that the gene accumulation rate in local endemic BAPS 1 lineage was higher than in other lineages, and the horizontal transmission of MDR IncHI2 plasmid associated with high resistance played a major role, which showed the potential threat to public health.

The receptor binding domain of SARS-CoV-2 Omicron subvariants targets Siglec-9 to decrease its immunogenicity by preventing macrophage phagocytosis.

The development of a vaccine specific to severe acute respiratory syndrome coronavirus 2 Omicron has been hampered due to its low immunogenicity. Here, using reverse mutagenesis, we found that a phenylalanine-to-serine mutation at position 375 (F375S) in the spike protein of Omicron to revert it to the sequence found in Delta and other ancestral strains significantly enhanced the immunogenicity of Omicron vaccines. Sequence FAPFFAF at position 371-377 in Omicron spike had a potent inhibitory effect on macrophage uptake of receptor-binding domain (RBD) nanoparticles or spike-pseudovirus particles containing this sequence. Omicron RBD enhanced binding to Siglec-9 on macrophages to impair phagocytosis and antigen presentation and promote immune evasion, which could be abrogated by the F375S mutation. A bivalent F375S Omicron RBD and Delta-RBD nanoparticle vaccine elicited potent and broad nAbs in mice, rabbits and rhesus macaques. Our research suggested that manipulation of the Siglec-9 pathway could be a promising approach to enhance vaccine response.

Polymorphisms of IFN signaling genes and FOXP4 influence the severity of COVID-19.

BACKGROUND: The clinical manifestations of COVID-19 range from asymptomatic, mild to moderate, severe, and critical disease. Host genetic variants were recognized to affect the disease severity. However, the genetic landscape differs among various populations. Therefore, we explored the variants associated with COVID-19 severity in the Guangdong population. METHODS: A total of 314 subjects were selected, of which the severe and critical COVID-19 patients were defined as "cases", and the mild and moderate patients were defined as "control". Twenty-two variants in interferon-related genes and FOXP4 were genotyped using the MassARRAY technology platform. RESULTS: IFN signaling gene MX1 rs17000900 CA + AA genotype was correlated with a reduced risk of severe COVID-19 in males (P = 0.001, OR = 0.050, 95%CI = 0.008-0.316). The AT haplotype comprised of MX1 rs17000900 and rs2071430 was more likely to protect against COVID-19 severity (P = 6.3E-03). FOXP4 rs1886814 CC genotype (P = 0.001, OR = 3.747, 95%CI = 1.746-8.043) and rs2894439 GA + AA genotype (P = 0.001, OR = 5.703, 95% CI = 2.045-15.903) were correlated with increased risk of severe COVID-19. Haplotype CA comprised of rs1886814 and rs2894439 was found to be correlated with adverse outcomes (P = 7.0E-04). FOXP4 rs1886814 CC (P = 0.0004) and rs2894439 GA + AA carriers had higher neutralizing antibody titers (P = 0.0018). The CA + AA genotype of MX1 rs17000900 tended to be correlated with lower neutralizing antibody titers than CC genotype (P = 0.0663), but the difference was not statistically significant. CONCLUSION: Our study found a possible association between MX1 and FOXP4 polymorphisms and the severity of COVID-19. Distinguishing high-risk patients who develop severe COVID-19 will provide clues for early intervention and individual treatment strategies.

Potent Antibody Response Elicited by a Third Booster Dose of Inactivated COVID-19 Vaccine in Healthy Subjects.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccine has been used worldwide on a large scale because of its potent ability to contain the coronavirus disease 2019 (COVID-19) pandemic, and the antibody response induced by the vaccine needs to be elucidated. Thus, we conducted a prospective trial in healthy subjects to observe the antibody response after three doses of inactivated vaccines. Our results showed that neutralizing antibody (NAb) levels were significantly higher after the booster vaccination compared to the second, a 4.9-fold increase, with the peak occurring at 28 days. The NAb level could be maintained for a longer period after the third vaccination, with higher levels still observed after 3 months. We did not observe significantly higher levels of SARS-CoV-2 spike-specific immunoglobulin G (S-IgG) and immunoglobulin M (IgM) after the third vaccination compared with the second vaccination; this was especially true for SARS-CoV-2 spike-specific immunoglobulin M (S-IgM), which was barely expressed. Notably, those who did not undergo NAb seroconversion after two doses of the vaccine produced high and long-lasting NAb after the third vaccination, confirming that they were not completely unresponsive to the vaccine. The NAb titer in younger subjects (aged 20-40 years) rose 3.4-fold compared with older subjects (aged 40-60 years) after the second vaccination, but the difference was narrowed after the third vaccination (2.8-fold increase). In addition, the levels of antibodies in older men were 3.4-fold lower than those in the older women after the third vaccination. Overall, this study elucidates the dynamic change in antibodies after three doses of vaccination, which provides a reference for the improvement of vaccination strategies.

The instantly blocking-based fluorescent immunochromatographic assay for the detection of SARS-CoV-2 neutralizing antibody.

Introduction: At present, there is an urgent need for the rapid and accurate detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) neutralizing antibodies (NAbs) to evaluate the ability of the human body to resist coronavirus disease 2019 (COVID-19) after infection or vaccination. The current gold standard for neutralizing antibody detection is the conventional virus neutralization test (cVNT), which requires live pathogens and biosafety level-3 (BSL-3) laboratories, making it difficult for this method to meet the requirements of large-scale routine detection. Therefore, this study established a time-resolved fluorescence-blocking lateral flow immunochromatographic assay (TRF-BLFIA) that enables accurate, rapid quantification of NAbs in subjects. Methods: This assay utilizes the characteristic that SARS-CoV-2 neutralizing antibody can specifically block the binding of the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein and angiotensin-converting enzyme 2 (ACE2) to rapidly detect the content of neutralizing antibody in COVID-19-infected patients and vaccine recipients. Results: When 356 samples of vaccine recipients were measured, the coincidence rate between this method and cVNT was 88.76%, which was higher than the coincidence rate of 76.97% between cVNT and a conventional chemiluminescence immunoassay detecting overall binding anti-Spike-IgG. More importantly, this assay does not need to be carried out in BSL-2 or 3 laboratories. Discussion: Therefore, this product can detect NAbs in COVID-19 patients and provide a reference for the prognosis and outcome of patients. Simultaneously, it can also be applied to large-scale detection to better meet the needs of neutralizing antibody detection after vaccination, making it an effective tool to evaluate the immunoprotective effect of COVID-19 vaccines.

SARS-CoV-2 ORF3a sensitizes cells to ferroptosis via Keap1-NRF2 axis.

Viral infection-induced cell death has long been considered as a double-edged sword in the inhibition or exacerbation of viral infections. Patients with severe Coronavirus Disease 2019 (COVID-19) are characterized by multiple organ dysfunction syndrome and cytokine storm, which may result from SARS-CoV-2-induced cell death. Previous studies have observed enhanced ROS level and signs of ferroptosis in SARS-CoV-2 infected cells or specimens of patients with COVID-19, but the exact mechanism is not clear yet. Here, we find SARS-CoV-2 ORF3a sensitizes cells to ferroptosis via Keap1-NRF2 axis. SARS-CoV-2 ORF3a promotes the degradation of NRF2 through recruiting Keap1, thereby attenuating cellular resistance to oxidative stress and facilitated cells to ferroptotic cell death. Our study uncovers that SARS-CoV-2 ORF3a functions as a positive regulator of ferroptosis, which might explain SARS-CoV-2-induced damage in multiple organs in COVID-19 patients and imply the potential of ferroptosis inhibition in COVID-19 treatment.

The genetic diversity, replication, and transmission of 2009 pandemic H1N1 viruses in China.

Background: The 2009 pandemic H1N1 influenza A virus (pdm09) continue to evolve, and few studies have systemically analyzed the evolution, replication, and transmission of pmd09 viruses in China. Methods: To better understand the evolution and pathogenicity of pdm09 viruses, we systematically analyzed viruses that were confirmed in 2009-2020 in China and characterized their replication and transmission ability. We extensively analyzed the evolution characteristics of pdm/09 in China over the past decades. The replication ability of 6B.1 and 6B.2 lineages on Madin-Darby canine kidney (MDCK) and human lung adenocarcinoma epithelial (A549) cells and their pathogenicity and transmission in guinea pigs were also compared. Results: In total, 3,038 pdm09 viruses belonged to clade 6B.1 (62% of all pdm09 viruses) and clade 6B.2 (4%). Clade 6B.1 pdm09 viruses are the predominant clade, with proportions of 54.1%, 78.9%, 57.2%, 58.6%, 61.7%, 76.3%, and 66.6% in the North, Northeast, East, Central, South, Southwest, and Northeast regions in China, respectively. The isolation proportion of clade 6B.1 pdm/09 viruses was 57.1%, 74.3%, 96.1%, 98.2%, 86.7%, and 78.5% in 2015-2020, respectively. A clear differentiation time point appeared in 2015 before which the evolution trend of pdm09 viruses in China was similar to that in North America but then showed a different trend after that point. To characterize pdm09 viruses in China after 2015, we further analyzed 33 pdm09 viruses isolated in Guangdong in 2016-2017, among which A/ Guangdong/33/2016 and A/Guangdong/184/2016 (184/2016) belonged to clade 6B.2, and the other 31 strains belonged to clade 6B.1. A/Guangdong/887/2017 (887/2017) and A/Guangdong/752/2017 (752/2017) (clade 6B.1), 184/2016 (clade 6B.2) and A/California/04/2009 (CA04) replicated efficiently in MDCK cells and A549 cells, as well as the turbinates of guinea pigs. 184/2016 and CA04 could transmit among guinea pigs through physical contact. Conclusion: Our findings provide novel insights into the evolution, pathogenicity, and transmission of pdm09 virus. The results show that enhancing surveillance of pdm09 viruses and timely evaluation of their virulence are essential.

Dynamic changes in polioviruses identified by environmental surveillance in Guangzhou, 2009-2021.

Polio cases can be missed by acute flaccid paralysis (AFP) case surveillance alone, emphasizing the importance of environmental surveillance (ES). In this study, to investigate the serotype distribution and epidemiological trends of poliovirus (PV), we characterized PV isolated from domestic sewage in Guangzhou City, Guangdong Province, China from 2009 to 2021. A total of 624 sewage samples were collected from the Liede Sewage Treatment Plant, and the positive rates of PV and non-polio enteroviruses were 66.67% (416/624) and 78.37% (489/624), respectively. After sewage sample treatment, each sewage sample was inoculated in six replicate tubes of three cell lines, and 3370 viruses were isolated during the 13-year surveillance period. Among these, 1086 isolates were identified as PV, including type 1 PV (21.36%), type 2 PV (29.19%), and type 3 PV (49.48%). Based on VP1 sequences, 1057 strains were identified as Sabin-like, 21 strains were high-mutant vaccines, and eight strains were vaccine-derived poliovirus (VDPV). The numbers and serotypes of PV isolates in sewage were influenced by the vaccine switch strategy. After type 2 OPV was removed from the trivalent oral PV (OPV) vaccine and a bivalent OPV (bOPV) was adopted in May 2016, the last type 2 PV strain was isolated from sewage, with no detection thereafter. Type 3 PV isolates increased significantly and became the dominant serotype. Before and after the second vaccine switch in January 2020, that is, from the first dose of IPV and second-fourth doses of bOPV to the first two doses of IPV and third-fourth doses of bOPV, there was also a statistical difference in PV positivity rates in sewage samples. Seven type 2 VDPVs and one type 3 VDPV were identified in sewage samples in 2009-2021, and phylogenetic analysis indicated that all VDPVs isolated from ES in Guangdong are newly discovered VDPVs, different from VDPV previously discovered in China, and were classified as ambiguous VDPV. It is noteworthy that no VDPV cases were reported in AFP case surveillance in the same period. In conclusion, continued PV ES in Guangzhou since April 2008 has been a useful supplement to AFP case surveillance, providing an important basis for evaluating the effectiveness of vaccine immunization strategies. ES improves early detection, prevention, and control; accordingly, this strategy can curb the circulation of VDPVs and provide a strong laboratory basis for maintaining a polio-free status.

Somatically hypermutated antibodies isolated from SARS-CoV-2 Delta infected patients cross-neutralize heterologous variants.

SARS-CoV-2 Omicron variants feature highly mutated spike proteins with extraordinary abilities in evading antibodies isolated earlier in the pandemic. Investigation of memory B cells from patients primarily with breakthrough infections with the Delta variant enables isolation of a number of neutralizing antibodies cross-reactive to heterologous variants of concern (VOCs) including Omicron variants (BA.1-BA.4). Structural studies identify altered complementarity determining region (CDR) amino acids and highly unusual heavy chain CDR2 insertions respectively in two representative cross-neutralizing antibodies-YB9-258 and YB13-292. These features are putatively introduced by somatic hypermutation and they are heavily involved in epitope recognition to broaden neutralization breadth. Previously, insertions/deletions were rarely reported for antiviral antibodies except for those induced by HIV-1 chronic infections. These data provide molecular mechanisms for cross-neutralization of heterologous SARS-CoV-2 variants by antibodies isolated from Delta variant infected patients with implications for future vaccination strategy.

Design, synthesis and immunological evaluation of monophosphoryl lipid A derivatives as adjuvants for a RBD-hFc based SARS-CoV-2 vaccine.

Toll-like receptor 4 (TLR4) is a reliable target for the development of vaccine adjuvants. To identify novel TLR4 ligands with improved immunological properties for use as adjuvants for a RBD-hFc based SARS-CoV-2 vaccine, herein, natural E. coli monophosphoryl lipid A (MPLA) and nine of its derivatives were designed and synthesized. Immunological evaluation showed that compounds 1, 3, 5 and 7 exhibited comparative or better adjuvant activity than clinically used Al adjuvants, and are expected to be a promising platform for the development of new adjuvants used for a RBD-hFc based SARS-CoV-2 vaccine. Preliminary structure-activity relationship analysis of the MPLA derivatives showed that the replacement of the functional groups at the C-1, C-4' or C-6' position of E. coli MPLA has an effect on its biological activity. In addition, we found that the combination of MPLA and Al was feasible for immunotherapy and could further enhance immune responses, providing a new direction toward the immunological enhancement of RBD-hFc based SARS-CoV-2 vaccines.

Carriage and Transmission of mcr-1 in Salmonella Typhimurium and Its Monophasic 1,4,[5],12:i:- Variants from Diarrheal Outpatients: a 10-Year Genomic Epidemiology in Guangdong, Southern China.

The banning of colistin as a feed additive for food-producing animals in mainland China in 2017 caused the decline in the prevalence of Escherichia coli-mobilized colistin resistance (mcr-1) in China. Salmonella Typhimurium and its monophasic 1,4,[5],12:i:- variants are also the main species associated with the spread of mcr-1; however, the evidence of the prevalence and transmission of mcr-1 among Salmonella is lacking. Herein, the 5,354 Salmonella isolates recovered from fecal samples of diarrheal patients in Guangdong, Southern China, from 2009 to 2019 were screened for colistin resistance and mcr-1, and mcr-1-positive isolates were characterized based on whole-genome sequencing (WGS) data. Relatively high prevalence rates of colistin resistance and mcr-1 (4.05%/4.50%) were identified, and more importantly, the prevalence trends of colistin-resistant and mcr-1-positive Salmonella isolates had a similar dynamic profile, i.e., both were first detected in 2012 and rapidly increased during 2013 to 2016, followed by a sharp decrease since 2017. WGS and phylogenetic analysis indicate that, whether before or after the ban, the persistence and cross-hospital transmission of mcr-1 are primarily determined by IncHI2 plasmids with similar backbones and sequence type 34 (ST34) Salmonella in specific clades that are associated with a high prevalence of IncHI2 plasmids and clinically important antimicrobial resistance genes, including blaCTX-M-14-fosA3-oqxAB-floR genotypes. Our work reveals the difference in the prevalence rate of mcr-1 in clinical Salmonella before and after the Chinese colistin ban, whereas mcr-1 transmission was closely linked to multidrug-resistant IncHI2 plasmid and ST34 Salmonella across diverse hospitals over 10 years. Continued surveillance is required to explore the factors related to a sharp decrease in mcr-1 after the recent ban and determine whether the ban has affected the carriage of mcr-1 in Salmonella circulating in the health care system. IMPORTANCE Colistin is one of the last-line antibiotics for the clinical treatment of Enterobacteriaceae. However, the emergence of the mobilized colistin resistance (mcr-1) gene has spread throughout the entire human health system and largely threatens the usage of colistin in the clinical setting. In this study, we investigated the existence of mcr-1 in clinical Salmonella from a 10-year continuous surveillance and genomic study. Overall, the colistin resistance rate and mcr-1 carriage of Salmonella in tertiary hospitals in Guangdong (2009 to 2019) were relatively high and, importantly, rapidly increased from 2013 to 2016 and significantly decreased after the Chinese colistin withdrawal. However, before or after the ban, the MDR IncHI2 plasmid with a similar backbone and ST34 Salmonella were the main vectors involved in the spread of mcr-1. Interestingly, these Chinese mcr-1-carrying Salmonella obtain phylogenetically and phylogeographically distinct patterns compared with those from other continents and are frequently associated with clinically important ARGs including the extended-spectrum ß-lactamases. Our data confirmed that the national stewardship intervention seems to be successful in blocking antibiotic resistance determinants and that continued surveillance of colistin resistance in clinical settings, farm animals, and related products is necessary.

Single-dose rAAV5-based vaccine provides long-term protective immunity against SARS-CoV-2 and its variants.

The COVID-19 pandemic, caused by the SARS-CoV-2 virus and its variants, has posed unprecedented challenges worldwide. Existing vaccines have limited effectiveness against SARS-CoV-2 variants. Therefore, novel vaccines to match mutated viral lineages by providing long-term protective immunity are urgently needed. We designed a recombinant adeno-associated virus 5 (rAAV5)-based vaccine (rAAV-COVID-19) by using the SARS-CoV-2 spike protein receptor binding domain (RBD-plus) sequence with both single-stranded (ssAAV5) and self-complementary (scAAV5) delivery vectors and found that it provides excellent protection from SARS-CoV-2 infection. A single-dose vaccination in mice induced a robust immune response; induced neutralizing antibody (NA) titers were maintained at a peak level of over 1:1024 more than a year post-injection and were accompanied by functional T-cell responses. Importantly, both ssAAV- and scAAV-based RBD-plus vaccines produced high levels of serum NAs against the circulating SARS-CoV-2 variants, including Alpha, Beta, Gamma and Delta. A SARS-CoV-2 virus challenge showed that the ssAAV5-RBD-plus vaccine protected both young and old mice from SARS-CoV-2 infection in the upper and lower respiratory tracts. Whole genome sequencing demonstrated that AAV vector DNA sequences were not found in the genomes of vaccinated mice one year after vaccination, demonstrating vaccine safety. These results suggest that the rAAV5-based vaccine is safe and effective against SARS-CoV-2 and several variants as it provides long-term protective immunity. This novel vaccine has a significant potential for development into a human prophylactic vaccination to help end the global pandemic.

Progress of the COVID-19: Persistence, Effectiveness, and Immune Escape of the Neutralizing Antibody in Convalescent Serum.

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), a new coronavirus causing Coronavirus Disease 2019 (COVID-19), is a major topic of global human health concern. The Delta and Omicron variants have caused alarming responses worldwide due to their high transmission rates and a number of mutations. During a one-year follow-up (from June 2020 to June 2021), we included 114 patients with SARS-CoV-2 infection to study the long-term dynamics and the correlative factors of neutralizing antibodies (NAbs) in convalescent patients. The blood samples were collected at two detection time points (at 6 and 12 months after discharge). We evaluated the NAbs response of discharged patients by performing a micro-neutralization assay using a SARS-CoV-2 wild type. In addition, a total of 62 serum samples from discharged COVID-19 patients with Alpha, Beta, Delta, and Omicron variants of infection were enrolled to perform cross-neutralization tests using the original SARS-CoV-2 strain and VOCs variants (including Alpha, Beta, Gamma, Delta, and Omicron variants) and to assess the ability of NAbs against the SARS-CoV-2 variants. NAbs seroconversion occurred in 91.46% of patients (n = 82) in the first timepoint and in 89.29% of patients (n = 84) in the second detection point, and three kinds of NAbs kinetics curves were perceived. The NAbs levels in young patients had higher values than those in elder patients. The kinetics of disease duration was accompanied by an opposite trend in NAbs levels. Despite a declining NAbs response, NAbs activity was still detectable in a substantial proportion of recovered patients one year after discharge. Compared to the wild strain, the Omicron strain could lead to a 23.44-, 3.42-, 8.03-, and 2.57-fold reduction in neutralization capacity in "SAlpha", "SBeta", "SDelta", and "SOmicron", respectively, and the NAbs levels against the Omicron strain were significantly lower than those of the Beta and Delta variants. Remarkably, the NAbs activity of convalescent serum with Omicron strain infection was most obviously detectable against six SARS-CoV-2 strains in our study. The role of the vaccination history in NAbs levels further confirmed the previous study that reported vaccine-induced NAbs as the convincing protection mechanism against SARS-CoV-2. In conclusion, our findings highlighted the dynamics of the long-term immune responses after the disappearance of symptoms and revealed that NAbs levels varied among all types of convalescent patients with COVID-19 and that NAbs remained detectable for one year, which is reassuring in terms of protection against reinfection. Moreover, a moderate correlation between the duration of disease and Nabs titers was observed, whereas age was negatively correlated with Nabs titers. On the other hand, compared with other VOCs, the Omicron variant was able to escape the defenses of the immune system more significantly, and the convalescent serum infected with the Omicron variant played a critical part in protection against different SARS-CoV-2 variants. Recovery serum from individuals vaccinated with inactivated vaccine preceding infection with the Omicron strain had a high efficacy against the original strain and the VOCs variants, whereas the convalescent serum of persons vaccinated by inactivated vaccine prior to infection with the Delta variant was only potent against the wild-type strain.

Human Fc-Conjugated Receptor Binding Domain-Based Recombinant Subunit Vaccines with Short Linker Induce Potent Neutralizing Antibodies against Multiple SARS-CoV-2 Variants.

The coronavirus disease-19 (COVID-19) pandemic has been ongoing since December 2019, with more than 6.3 million deaths reported globally as of August 2022. Despite the success of several SARS-CoV-2 vaccines, the rise in variants, some of which are resistant to the effects of vaccination, highlights the need for a so-called pan-coronavirus (universal) vaccine. Here, we performed an immunogenicity comparison of prototype vaccines containing spike protein receptor-binding domain (RBD) residues 319-541, or spike protein regions S1, S2 and S fused to a histidine-tagged or human IgG1 Fc (hFC) fragment with either a longer (six residues) or shorter (three residues) linker. While all recombinant protein vaccines developed were effective in eliciting humoral immunity, the RBD-hFc vaccine was able to generate a potent neutralizing antibody response as well as a cellular immune response. We then compared the effects of recombinant protein length and linker size on immunogenicity in vivo. We found that a longer recombinant RBD protein (residues 319-583; RBD-Plus-hFc) containing a small alanine linker (AAA) was able to trigger long-lasting, high-titer neutralizing antibodies in mice. Finally, we evaluated cross-neutralization of wild-type and mutant RBD-Plus-hFc vaccines against wild-type, Alpha, Beta, Delta and Omicron SARS-CoV-2 variants. Significantly, at the same antigen dose, wild-type RBD-Plus-hFc immune sera induced broadly neutralizing antibodies against wild-type, Alpha, Beta, Delta and Omicron variants. Taken together, our findings provide valuable information for the continued development of recombinant protein-based SARS-CoV-2 vaccines and a basic foundation for booster vaccinations to avoid reinfection with SARS-CoV-2 variants.

Phylogenomic analysis of Salmonella Indiana ST17, an emerging MDR clonal group in China.

OBJECTIVES: To reconstruct the genomic epidemiology and evolution of MDR Salmonella Indiana in China. METHODS: A total of 108 Salmonella Indiana strains were collected from humans and livestock in China. All isolates were subjected to WGS and antimicrobial susceptibility testing. Phylogenetic relationships and evolutionary analyses were conducted using WGS data from this study and the NCBI database. RESULTS: Almost all 108 Salmonella Indiana strains displayed the MDR phenotype. Importantly, 84 isolates possessed concurrent resistance to ciprofloxacin and cefotaxime. WGS analysis revealed that class 1 integrons on the chromosome and IncHI2 plasmids were the key vectors responsible for multiple antibiotic resistance gene (ARG) [including ESBL and plasmid-mediated quinolone resistance (PMQR) genes] transmission among Salmonella Indiana. The 108 Salmonella Indiana dataset displayed a relatively large core genome and ST17 was the predominant ST. Moreover, the global ST17 Salmonella Indiana strains could be divided into five distinct lineages, each of which was significantly associated with a geographical distribution. Genomic analysis revealed multiple antimicrobial resistance determinants and QRDR mutations in Chinese lineages, which almost did not occur in other global lineages. Using molecular clock analysis, we hypothesized that ST17 isolates have existed since 1956 and underwent a major population expansion from the 1980s to the 2000s and the genetic diversity started to decrease around 2011, probably due to geographical barriers, antimicrobial selective pressure and MDR, favouring the establishment of this prevalent multiple antibiotic-resistant lineage and local epidemics. CONCLUSIONS: This study revealed that adaptation to antimicrobial pressure was possibly pivotal in the recent evolutionary trajectory for the clonal spread of ST17 Salmonella Indiana in China.

Etiology of acute febrile illnesses in Southern China: Findings from a two-year sentinel surveillance project, 2017-2019.

BACKGROUND: Southern China is at risk for arborvirus disease transmission, including Zika virus and dengue. Patients often present to clinical care with non-specific acute febrile illnesses (AFI). To better describe the etiology of AFI, we implemented a two-year AFI surveillance project at five sentinel hospitals in Yunnan and Guangdong Provinces. METHODS: Between June 2017 and August 2019, we enrolled patients between 2 and 65 years of age presenting at one sentinel hospital in Mengla County, Yunnan, and four in Jiangmen City, Guangdong, with symptoms of AFI (acute onset of fever ≥ 37.5°C within the past 7 days) without respiratory symptoms or diarrhea. Demographic, epidemiologic, and clinical information was obtained and entered into a web-based AFI surveillance database. A custom TaqMan Array card (TAC) was used to test patients' whole blood specimens for 27 different pathogens using real-time polymerase chain reaction assays. RESULTS: During the two-year project period, 836 patients were enrolled; 443 patients from Mengla County and 393 patients from Jiangmen City. The median age was 33 years [range: 2-65], and most were hospitalized [641, 77%]. Of 796 patients with valid TAC results, 341 (43%) were positive for at least one of the 10 unique pathogens detected. This included 205 (26%) patients positive for dengue virus, 60 (8%) for Orientia tsutsugamushi, and 42 (5%) for Coxiella burnetii. Ten patients (1%) in Jiangmen City tested positive for malaria, 8 of whom reported recent travel outside of China. TAC results were negative for 455 (57%) patients. None of the patients had a positive TAC detection for Zika virus. CONCLUSIONS: The project detected variability in the etiology of AFI in Southern China and highlighted the importance of differential diagnosis. Dengue, O. tsutsugamushi, and C. burnetii were the most frequently identified pathogens among enrolled AFI patients. As a non-notifiable disease, the frequent detection of C. burnetii is noteworthy and warrants additional investigation. The project provided a framework for routine surveillance for persons presenting with AFI.

Broad and durable antibody response after vaccination with inactivated SARS-CoV-2 in individuals with a history of 2003 SARS-CoV infection.

In vaccinees who were infected with SARS-CoV in 2003, we observed greater antibody responses against spike and nucleoprotein of both SARS-CoV-2 and SARS-CoV after a single dosage of inactivated SARS-CoV-2 vaccine. After receiving the second vaccination, antibodies against RBD of SARS-CoV-2 Wuhan, Beta, Delta, and recently emerged Omicron are significantly higher in SARS-CoV experienced vaccinees than in SARS-CoV naïve vaccinees. Neutralizing activities measured by authentic viruses and pseudoviruses of SARS-CoV, SARS-CoV-2 Wuhan, Beta, and Delta are greater in SARS-CoV experienced vaccinees. In contrast, only weak neutralizing activities against SARS-CoV-2 and variants were detected in SARS-CoV naïve vaccinees. By 6 months after the second vaccination, neutralizing activities were maintained at a relatively higher level in SARS-CoV experienced vaccinees but were undetectable in SARS-CoV naïve vaccinees. These findings suggested a great possibility of developing a universal vaccine by heterologous vaccination using spike antigens from different SARS-related coronaviruses.