N6-Methyladenosine Positively Regulates Coxsackievirus B3 Replication.

Enteroviruses such as coxsackievirus B3 are identified as a common cause of viral myocarditis, but the potential mechanism of its replication and pathogenesis are largely unknown. The genomes of a variety of viruses contain N6-methyladenosine (m6A), which plays important roles in virus replication. Here, by using the online bioinformatics tools SRAMP and indirect immunofluorescence assay (IFA), we predict that the CVB3 genome contains m6A sites and found that CVB3 infection could alter the expression and cellular localization of m6A-related proteins. Moreover, we found that 3-deazaadenosine (3-DAA), an m6A modification inhibitor, significantly decreased CVB3 replication. We also observed that the m6A methyltransferases methyltransferase-like protein 3 (METTL3) and METTL14 play positive roles in CVB3 replication, whereas m6A demethylases fat mass and obesity-associated protein (FTO) or AlkB homolog 5 (ALKBH5) have opposite effects. Knockdown of the m6A binding proteins YTH domain family protein 1 (YTHDF1), YTHDF2 and YTHDF3 strikingly decreased CVB3 replication. Finally, the m6A site mutation in the CVB3 genome decreased the replication of CVB3 compared with that in the CVB3 wild-type (WT) strain. Taken together, our results demonstrated that CVB3 could exploit m6A modification to promote viral replication, which provides new insights into the mechanism of the interaction between CVB3 and the host.

Bronchoalveolar Lavage Fluid Metagenomic Second-Generation Sequencing Assists in Guiding the Treatment of Visceral Leishmaniasis: A Case Report.

Purpose: The incidence of visceral leishmaniasis (VL), a global infectious disease, has been on the rise in China's Hebei province. When patients achieve clinical cure, they often do not reach an etiological cure, which may lead to recurrence of the disease. Here, we report a case of visceral leishmaniasis with a negative blood smear and bone marrow cytology. Patients and Methods: A 65-year-old man and bronchoalveolar lavage fluid mNGS. Results: A 65-year-old man developed a chronic fever, anorexia, splenomegaly, and pancytopenia. The blood metagenomic second-generation sequencing (mNGS) revealed Leishmania sequence readings, which led to the diagnosis of VL. After sodium antimony gluconate treatment, the blood smear and bone marrow cytology revealed no Leishmania bodies. However, pancytopenia and respiratory failure did not fully subside, and cardiotoxic damage emerged. The bronchoalveolar lavage fluid (BALF) mNGS was performed to detect the pathogen. Through BALF mNGS, Leishmania sequence was still detectable. Therefore, after the ECG returned to normal, antimony sodium gluconate was administered as a next course of treatment. Conclusion: BALF mNGS may assist in evaluating the therapeutic efficacy of VL with respiratory failure, especially in patients with negative blood and bone marrow cytology.

Accurate detection of visceral leishmaniasis is essential for clinical diagnosis.It is uncommon to use alveolar lavage fluid mNGS in etiological diagnosis.Patient with negative bone marrow cytology may refer to alveolar lavage fluid mNGS.

Ferritin nanoparticle-based Nipah virus glycoprotein vaccines elicit potent protective immune responses in mice and hamsters.

Nipah virus (NiV) is a zoonotic paramyxovirus in the genus Henipavirus that is prevalent in Southeast Asia. NiV leads to severe respiratory disease and encephalitis in humans and animals, with a mortality rate of up to 75%. Despite the grave threat to public health and global biosecurity, no medical countermeasures are available for humans. Here, based on self-assembled ferritin nanoparticles (FeNPs), we successfully constructed two candidate FeNP vaccines by loading mammalian cells expressing NiV sG (residues 71-602, FeNP-sG) and Ghead (residues 182-602, FeNP-Ghead) onto E. coli-expressed FeNPs (FeNP-sG and FeNP-Ghead, respectively) through Spycatcher/Spytag technology. Compared with sG and Ghead alone, FeNP-sG and FeNP-Ghead elicited significant NiV specific neutralizing antibody levels and T-cell responses in mice, whereas the immune response in the FeNP-sG immunized group was greater than that in the FeNP-Ghead group. These results further demonstrate that sG possesses greater antigenicity than Ghead and that FeNPs can dramatically enhance immunogenicity. Furthermore, FeNP-sG provided 100% protection against NiV challenge in a hamster model when it was administered twice at a dose of 5 µg/per animal. Our study provides not only a promising candidate vaccine against NiV, but also a theoretical foundation for the design of a NiV immunogen for the development of novel strategies against NiV infection.

Polyvalent mpox mRNA vaccines elicit robust immune responses and confer potent protection against vaccinia virus.

The 2022 mpox outbreak led the World Health Organization (WHO) to declare it a public health emergency of international concern (PHEIC). There is a need to develop more effective and safer mpox virus (MPXV)-specific vaccines in response to the mpox epidemic. The mRNA vaccine is a promising platform to protect against MPXV infection. In this study, we construct two bivalent MPXV mRNA vaccines, designated LBA (B6R-A29L) and LAM (A35R-M1R), and a quadrivalent mRNA vaccine, LBAAM (B6R-A35R-A29L-M1R). The immunogenicity and protective efficacy of these vaccines alone or in combination were evaluated in a lethal mouse model. All mRNA vaccine candidates could elicit potential antigen-specific humoral and cellular immune responses and provide protection against vaccinia virus (VACV) infection. The protective effect of the combination of two bivalent mRNA vaccines and the quadrivalent vaccine was superior to that of the individual bivalent mRNA vaccine. Our study provides valuable insights for the development of more efficient and safer mRNA vaccines against mpox.

Circular RNA vaccines against monkeypox virus provide potent protection against vaccinia virus infection in mice.

Since the outbreak of monkeypox (mpox) in 2022, widespread concern has been placed on imposing an urgent demand for specific vaccines that offer safer and more effective protection. Using an efficient and scalable circular RNA (circRNA) platform, we constructed four circRNA vaccines that could induce robust neutralizing antibodies as well as T cell responses by expressing different surface proteins of mpox virus (MPXV), resulting in potent protection against vaccinia virus (VACV) in mice. Strikingly, the combination of the four circular RNA vaccines demonstrated the best protection against VACV challenge among all the tested vaccines. Our study provides a favorable approach for developing MPXV-specific vaccines by using a circular mRNA platform and opens up novel avenues for future vaccine research.

A potential bivalent mRNA vaccine candidate protects against both RSV and SARS-CoV-2 infections.

As the world continues to confront severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), respiratory syncytial virus (RSV) is also causing severe respiratory illness in millions of infants, elderly individuals, and immunocompromised people globally. Exacerbating the situation is the fact that co-infection with multiple viruses is occurring, something which has greatly increased the clinical severity of the infections. Thus, our team developed a bivalent vaccine that delivered mRNAs encoding SARS-CoV-2 Omicron spike (S) and RSV fusion (F) proteins simultaneously, SF-LNP, which induced S and F protein-specific binding antibodies and cellular immune responses in BALB/c mice. Moreover, SF-LNP immunization effectively protected BALB/c mice from RSV infection and hamsters from SARS-CoV-2 Omicron infection. Notably, our study pointed out the antigenic competition problem of bivalent vaccines and provided a solution. Overall, our results demonstrated the potential of preventing two infectious diseases with a single vaccine and provided a paradigm for the subsequent design of multivalent vaccines.

Ebola virus VP35 perturbs type I interferon signaling to facilitate viral replication.

As one of the deadliest viruses, Ebola virus (EBOV) causes lethal hemorrhagic fevers in humans and nonhuman primates. The suppression of innate immunity leads to robust systemic virus replication of EBOV, leading to enhanced transmission. However, the mechanism of EBOV-host interaction is not fully understood. Here, we identified multiple dysregulated genes in early stage of EBOV infection through transcriptomic analysis, which are highly clustered to Jak-STAT signaling. EBOV VP35 and VP30 were found to inhibit type I interferon (IFN) signaling. Moreover, exogenous expression of VP35 blocks the phosphorylation of endogenous STAT1, and suppresses nuclear translocation of STAT1. Using serial truncated mutations of VP35, N-terminal 1-220 amino acid residues of VP35 were identified to be essential for blocking on type I IFN signaling. Remarkably, VP35 of EBOV suppresses type I IFN signaling more efficiently than those of Bundibugyo virus (BDBV) and Marburg virus (MARV), resulting in stable replication to facilitate the pathogenesis. Altogether, this study enriches understanding on EBOV evasion of innate immune response, and provides insights into the interplay between filoviruses and host.

Duration of humoral immunity from smallpox vaccination and its cross-reaction with Mpox virus.

The ongoing pandemic caused by mpox virus (MPXV) has become an international public health emergency that poses a significant threat to global health. The vaccinia virus Tiantan strain (VTT) was used to vaccinate against smallpox in China 42 years ago. It is urgent to assess the level of immunity to smallpox in individuals vaccinated 43 or more years ago and evaluate their immunological susceptibility to MPXV. Here, we recruited 294 volunteers and detected the level of residual humoral immunity, including the vaccinia-specific IgG level and neutralizing antibody titer, and the cross-antibodies of MPXV A29L, B6R, A35R, and M1R. Our results showed that the humoral immunity from the smallpox vaccine in the population still remains, and VTT-specific NAb levels wane with age. The majority of the population pre-1981 who should be immunized with VTT still maintains certain levels of MPXV-specific antibodies, in particular, targeting A35R and B6R antigens. Furthermore, we separately analyzed the correlations between the OD450 values of VTT-specific IgG and A35R-specific IgG, B6R-specific IgG, and A29L-specific IgG with plasma samples diluted 1:40, showing a linear correlation (p < 0.0001). Our findings suggest that most Chinese populations still maintain VTT-specific IgG antibodies for 42 or more years after smallpox vaccination and could provide some level of protection against MPXV.

Analysis of multidrug-resistant determinants of clinically isolated Acinetobacter baumannii CYZ via whole genome sequencing.

Acinetobacter baumannii is a multidrug-resistant coccobacillus responsible for severe nosocomial infectious diseases. This study mainly focuses on investigating the antimicrobial resistance features of a clinically isolated strain (A. baumannii CYZ) using the PacBio Sequel II sequencing platform. The chromosomal size of A. baumannii CYZ is 3,960,760 bp, which contains a total of 3803 genes with a G + C content of 39.06%. Functional analysis performed using the Clusters of Orthologous Groups of Proteins (COGs), Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) databases, as well as the Comprehensive Antibiotic Resistance Database (CARD) revealed a complicated set of antimicrobial resistance determinants in the genome of A. baumannii CYZ, which were mainly classified into multidrug efflux pumps and transport systems, ß-lactamase relative and penicillin-binding proteins, aminoglycoside modification enzymes, alternation of antibiotic target sites, lipopolysaccharide relative, and other mechanisms. A total of 35 antibiotics were tested for the antimicrobial susceptibility of A. baumannii CYZ, and the organism exhibited a stronger antimicrobial resistance ability. The phylogenetic relationship indicated that A. baumannii CYZ has high homology with A. baumannii ATCC 17978; however, the former also exhibited its specific genome characteristics. Our research results give insight into the genetic antimicrobial-resistant features of A. baumannii CYZ as well as provide a genetic basis for the further study of the phenotype.

HBV precore G1896A mutation promotes growth of hepatocellular carcinoma cells by activating ERK/MAPK pathway.

Chronic hepatitis B virus (HBV) infection is one of the leading causes of hepatocellular carcinoma (HCC). The HBV genome is prone to mutate and several variants are closely related to the malignant transformation of liver disease. G1896A mutation (G to A mutation at nucleotide 1896) is one of the most frequently observed mutations in the precore region of HBV, which prevents HBeAg expression and is strongly associated with HCC. However, the mechanisms by which this mutation causes HCC are unclear. Here, we explored the function and molecular mechanisms of the G1896A mutation during HBV-associated HCC. G1896A mutation remarkably enhanced the HBV replication in vitro. Moreover, it increased tumor formation and inhibited apoptosis of hepatoma cells, and decreased the sensitivity of HCC to sorafenib. Mechanistically, the G1896A mutation could activate ERK/MAPK pathway to enhanced sorafenib resistance in HCC cells and augmented cell survival and growth. Collectively, our study demonstrates for the first time that the G1896A mutation has a dual regulatory role in exacerbating HCC severity and sheds some light on the treatment of G1896A mutation-associated HCC patients.

Ferrets: A powerful model of SARS-CoV-2.

The rapid spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in recent years not only caused a global pandemic but resulted in enormous social, economic, and health burdens worldwide. Despite considerable efforts to combat coronavirus disease 2019 (COVID-19), various SARS-CoV-2 variants have emerged, and their underlying mechanisms of pathogenicity remain largely unknown. Furthermore, effective therapeutic drugs are still under development. Thus, an ideal animal model is crucial for studying the pathogenesis of COVID-19 and for the preclinical evaluation of vaccines and antivirals against SARS-CoV-2 and variant infections. Currently, several animal models, including mice, hamsters, ferrets, and non-human primates (NHPs), have been established to study COVID-19. Among them, ferrets are naturally susceptible to SARS-CoV-2 infection and are considered suitable for COVID-19 study. Here, we summarize recent developments and application of SARS-CoV-2 ferret models in studies on pathogenesis, therapeutic agents, and vaccines, and provide a perspective on the role of these models in preventing COVID-19 spread.

The life cycle of Hyalomma scupense (Acari: Ixodidae) under laboratory conditions.

The life cycle of Hyalomma scupense on rabbit hosts was investigated under laboratory conditions. Hy. scupense exhibited one- and two-host life cycles of 163.2 and 161.4 days, respectively. The incubation of eggs required an average period of 52 days, which was the longest period among the four developmental stages. The average time for pre-feeding of larvae was 3.5 days. It took 20 days for larvae to become engorged nymphs and 52.3 days to become engorged females. The duration of the pre-feeding, feeding, pre-oviposition, and oviposition stages of female adults was 2.3, 13.5, 27.5, and 27.9 days, respectively. The average weight of engorged females was 390.0 mg (ranging from 129.3 mg to 828.6 mg), which was 28.95 times the weight of unfed females. There was a positive relationship between the weight and the number of eggs laid by engorged females (r = 0.927). The reproductive efficiency index (REI) was 8.63.

Monkeypox virus: a re-emergent threat to humans.

Human monkeypox (MPX) is a rare zoonotic infection characterized by smallpox-like signs and symptoms. It is caused by monkeypox virus (MPXV), a double stranded DNA virus belonging to the genus Orthopoxvirus. MPX was first identified in 1970 and mostly prevailed in the rural rainforests of Central and West Africa in the past. Outside Africa, MPX was reported in the United Kingdom, the USA, Israel, and Singapore. In 2022, the resurgence of MPX in Europe and elsewhere posed a potential threat to humans. MPXV was transmitted by the animals-human or human-human pathway, and the symptoms of MPXV infection are similar to that of smallpox, but in a milder form and with lower mortality (1%-10%). Although the smallpox vaccination has been shown to provide 85% protection against MPXV infection, and two anti-smallpox virus drugs have been approved to treat MPXV, there are still no specific vaccines and drugs against MPXV infection. Therefore it is urgent to take active measures including the adoption of novel anti-MPXV strategies to control the spread of MPXV and prevent MPX epidemic. In this review, we summarize the biological features, epidemiology, pathogenicity, laboratory diagnosis, and prevention and treatment strategies on MPXV. This review provides the basic knowledge for prevention and control of future outbreaks of this emerging infection.

Clinical Value of Serum Neuron-Specific Enolase Combined with Serum S100B Protein in the Diagnosis of Systemic Lupus Erythematosus.

Objective: To investigate the clinical value of serum neuron-specific enolase (NSE) combined with serum S100B protein in the diagnosis of systemic lupus erythematosus (SLE). Methods: Sixty patients with SLE treated in our hospital from January 2019 to April 2021 were enrolled as the study group. According to the degree of activity, the study group was assigned into three groups: mild activity group (n = 20), moderate activity group (n = 20), and severe activity group (n = 20). A total of 60 healthy people who underwent physical examination in our hospital in the same period were enrolled as the control group. The NSE and serum S100B protein were detected in the two groups, and the correlation between serum nerve-specific enolase and serum S100B protein and the clinical value in the diagnosis of SLE were analyzed. Results: First of all, we compared the general data of the two groups. There was no significant difference in sex, age, marital status, and education level, and no significant difference was exhibited (p > 0.05). There was no significant difference in sex, age, marital status, and education level among mild activity group, moderate activity group, and severe activity group, and no significant difference in data was exhibited (p > 0.05). Secondly, we compared the levels of serum S100B protein and NSE. The levels of serum S100B protein and NSE in the study group were higher compared to the control group (p < 0.05). The levels of serum S100B protein and NSE in patients with different activity levels of SLE were compared. The levels of serum S100B protein and NSE in mild activity group < moderate activity group < severe activity group were significantly different (p < 0.05). Correlation analysis between serum S100B, NSE levels, and SLE activity indicated that serum S100B and NSE levels were positively correlated with SLE activity. With the increase of SLE activity, serum S100B and NSE levels gradually increased, and the data difference was statistically significant (r = 0.855, 0.844, p < 0.05). Finally, we established the logistic prediction model, take the probability of generating prediction as the analysis index, and draw the ROC curve to evaluate the diagnostic value of different combinations to SLE. The highest AUC and sensitivity of the two indexes in the diagnosis of SLE were 0.773 and 0.836, respectively. The levels of serum S100B protein and NSE have a certain value in the diagnosis of SLE, while the combined diagnosis is of higher value, sensitivity, and specificity in the diagnosis of SLE. Conclusion: Serum S100B protein and NSE are very sensitive indexes to judge the damage of central nervous system. However, due to the small number of cases in this study, there were as many as 19 kinds of NPSLE classification, so the relationship between serum S100B protein, NSE levels, and various NPSLE and their exact application value in diagnosing the disease and judging the prognosis needs to be confirmed by expanding the number of cases.

Long-term asymptomatic SARS-CoV-2 infection associated with deficiency on multiple immune cells.

The immune responses and the function of immune cells among asymptomatic severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection cases, especially in immuno-compromised individuals, remain largely unknown. Here we present a case of asymptomatic SARS-CoV-2 infection that lasted for at least 67 days. The patient has administrated Thymalfasin as 1.6 mg per dose every other day from Day 45 to 70, plus 200 mg per dose Arbidol antiviral therapy three doses per day from Day 48 to 57. Throughout the infection, no anti-SARS-CoV-2 specific IgM or IgG antibodies were detected. Instead, the patient showed either a low percentage or an absolute number of non-classical monocytes, dendritic cells (DCs), CD4+ T cells, and regulatory T cells (Tregs), which may account for the clinical feature and absence of antibody response. This case may shed new light on the outbreak management related to control/prevention, treatment, and vaccination of SARS-CoV-2 and other virus infections in immunocompromised individuals.

Hold Breath: Autonomic Neural Regulation of Innate Immunity to Defend Against SARS-CoV-2 Infection.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a novel member of the genus of betacoronavirus, which caused a pandemic of coronavirus disease 2019 (COVID-19) worldwide. The innate immune system plays a critical role in eliminating the virus, which induces inflammatory cytokine and chemokine secretion, produces different interferons, and activates the adaptive immune system. Interactions between the autonomic nervous system and innate immunity release neurotransmitters or neuropeptides to balance the excess secretion of inflammatory cytokines, control the inflammation, and restore the host homeostasis. However, more neuro-immune mechanisms to defend against viral infection should be elucidated. Here, we mainly review and provide our understanding and viewpoint on the interaction between respiratory viral proteins and host cell receptors, innate immune responses to respiratory viral infection, and the autonomic neural regulation of the innate immune system to control respiratory viruses caused by lungs and airways inflammation.

Regulation of Antiviral Immune Response by N 6-Methyladenosine of mRNA.

Host innate and adaptive immune responses play a vital role in clearing infected viruses. Meanwhile, viruses also evolve a series of mechanisms to weaken the host immune responses and evade immune defense. Recently, N 6-methyladenosine (m6A), the most prevalent mRNA modification, has been revealed to regulate multiple steps of RNA metabolism, such as mRNA splicing, localization, stabilization, and translation, thus participating in many biological phenomena, including viral infection. In the process of virus-host interaction, the m6A modification that presents on the virus RNA impedes capture by the pattern recognition receptors, and the m6A modification appearing on the host immune-related molecules regulate interferon response, immune cell differentiation, inflammatory cytokine production, and other immune responses induced by viral infection. This review summarizes the research advances about the regulatory role of m6A modification in the innate and adaptive immune responses during viral infections.

Characteristic antimicrobial resistance of clinically isolated Stenotrophomonas maltophilia CYZ via complete genome sequence.

OBJECTIVES: Stenotrophomonas maltophilia is an important multidrug-resistant pathogen that is associated with various serious nosocomial infections. In our study, we investigated the antimicrobial resistance traits of clinical S. maltophilia strain CYZ isolated from the sputum of an immunocompromised patient. METHODS: The whole genome sequence of S. maltophilia CYZ was investigated using a PacBio RS II system. The functions of all the predicted genes were annotated by the COG, GO and KEGG databases. Several types of antibiotics were selected to test the antimicrobial susceptibility, and a phylogenetic tree was constructed based on 16S rRNA gene sequence. RESULTS: The genome of S. maltophilia CYZ has a length of 4,517,685 bp and contains 4077 predicted genes, with an average G + C content of 66.65%. Functional genomic analysis via the annotations of the COG and GO databases revealed that the isolate exhibited specific means to resist antibiotics. The annotated genes involved in flagella, pili or fimbriae, biofilm formation, polysaccharide and cyclic di-GMP may contribute to promote the ability of antimicrobial resistance. This strain showed susceptibility to levofloxacin, trimethoprim/sulfamethoxazole and minocycline according to antimicrobial susceptibility testing. The phylogenetic relationship indicated that S. maltophilia CYZ was closely related to S. maltophilia strains isolated from the nosocomial environment. CONCLUSIONS: The current results give a better understanding of the genetic characteristics of antimicrobial resistance in S. maltophilia CYZ and provide a genetic basis for further study of the phenotype.

Legionella feeleii: pneumonia or Pontiac fever? Bacterial virulence traits and host immune response.

Gram-negative bacterium Legionella is able to proliferate intracellularly in mammalian host cells and amoeba, which became known in 1976 since they caused a large outbreak of pneumonia. It had been reported that different strains of Legionella pneumophila, Legionella micdadei, Legionella longbeachae, and Legionella feeleii caused human respiratory diseases, which were known as Pontiac fever or Legionnaires' disease. However, the differences of the virulence traits among the strains of the single species and the pathogenesis of the two diseases that were due to the bacterial virulence factors had not been well elucidated. L. feeleii is an important pathogenic organism in Legionellae, which attracted attention due to cause an outbreak of Pontiac fever in 1981 in Canada. In published researches, it has been found that L. feeleii serogroup 2 (ATCC 35849, LfLD) possess mono-polar flagellum, and L. feeleii serogroup 1 (ATCC 35072, WRLf) could secrete some exopolysaccharide (EPS) materials to the surrounding. Although the virulence of the L. feeleii strain was evidenced that could be promoted, the EPS might be dispensable for the bacteria that caused Pontiac fever. Based on the current knowledge, we focused on bacterial infection in human and murine host cells, intracellular growth, cytopathogenicity, stimulatory capacity of cytokines secretion, and pathogenic effects of the EPS of L. feeleii in this review.