Schmallenberg virus (SBV), an emerging Culicoides‑borne arbovirus, is responsible for febrile illness and reduced milk production in adult ruminants and can induce congenital malformations in fetuses, representing a substantial concern for livestock health worldwide. Consequently, rapid, field‑adaptable diagnostic approaches are urgently needed. In this study, we developed a visual nucleic acid detection assay for SBV that combines reverse transcription recombinase‑aided amplification (RT‑RAA) with the CRISPR/Cas12a system, targeting conserved regions of the SBV S gene. Following systematic optimization of the reaction conditions, the assay was completed within 50 min, with a sensitivity of 8.6 copies/μL for the SBV-S plasmid and 8.6 × 101 copies/μL for the RNA transcripts. The assay exhibited high specificity, with no cross‑reactivity observed against a panel of relevant pathogens, including Seoul orthohantavirus (SEOV), infectious bovine rhinotracheitis virus (IBRV), Rift Valley fever virus (RVFV), Crimean‑Congo hemorrhagic fever virus (CCHFV), and bovine viral diarrhea virus (BVDV). When evaluated using simulated clinical samples, the sensitivity of the method was superior to that of conventional real-time fluorescent reverse transcription‒polymerase chain reaction (RT‑qPCR). Importantly, the entire reaction is performed in a single closed-tube format, significantly reducing the risk of cross-contamination and false-positive results. In summary, this method demonstrates favorable analytical performance for the detection of SBV-S plasmid-spiked nasal swab samples and holds promise for further clinical validation, although its diagnostic utility in authentic clinical specimens remains to be confirmed in future investigations.

Generating recombinant viruses lacking an essential gene for producing infective viral particles represents a promising approach to developing safer live viral vaccines. The envelope-spikes glycoprotein (G) of rabies virus (RABV) plays a critical role in mediating viral adsorption and entry into host cells, making it a key factor influencing its pathogenicity. Herein, we focused on constructing a replication-deficient rabies virus (RABV) with a specific G gene deletion (rSRV9-△G-eGFP). This engineered virus is incapable of forming plaques in cells unless supplemented externally with the G protein, confirming its status as a single-cycle virus producing non-infectious progeny. Despite this restriction, rSRV9-ΔG-eGFP efficiently initiated host immune activation following immunization. Notably, vaccination induced pronounced recruitment and activation of antigen-presenting cells, as well as enhanced T and B lymphocyte responses, highlighting a strong cellular immune component that distinguishes this platform from traditional inactivated rabies vaccines. Consistent with protective immunity, rSRV9-ΔG-eGFP elicited virus-neutralizing antibody titers exceeding the accepted protective threshold of 0.5 IU/mL and conferred protection against lethal RABV challenge. Together, these findings support rSRV9-ΔG-eGFP as a promising replication-deficient rabies vaccine candidate and suggest its potential utility as a viral vector platform for future vaccine development.

In 2024, Monkeypox virus (MPXV) clade I has triggered outbreaks in several countries world-wide. MPXV clade I demonstrates enhanced virulence and transmissibility, with a case fatality rate reaching 10%. In response, we have developed a one-pot detection assay specifically targeting MPXV clade I, combining recombinase polymerase amplification (RPA) and the CRISPR/Cas12a system. The assay can be completed within 40 min and achieved a 95% limit of detection (LOD95) of 27.16 copies/μL. No cross-reactivity was observed with MPXV clade II or other tested viral templates, including Vaccinia virus (Tiantan strain). A preliminary room-temperature evaluation showed that the assay retained detectable performance at 25°C, supporting its potential use in equipment-limited settings. The assay also showed good intra-assay and inter-assay repeatability for recombinant plasmid templates, with all coefficient of variation (CV) values below 10%. In simulated clinical samples, the RPA-CRISPR/Cas12a assay detected more low-concentration plasmid-spiked samples than quantitative polymerase chain reaction (qPCR). These results indicate that the established assay is specific, sensitive, repeatable, and easy to perform, providing a practical tool for field-based screening and decentralized detection of MPXV clade I.

Severe fever with thrombocytopenia syndrome (SFTS) is a tick‑borne zoonosis with a case fatality rate of up to 16% and pandemic potential, yet no specific therapeutics or licensed vaccines are available. Subunit vaccines offer a safer alternative but typically suffer from poor immunogenicity as conventional adjuvants fail to coordinate effective antigen delivery with robust innate immune activation. Here, we develop a biomimetic mesoporous polydopamine (MPDA) nanoplatform that achieves high loading efficiency of the SFTS virus Gn antigen, enables lysosomal escape, and activates innate immunity. In a lethal SFTSV mouse model, MPDA-based vaccination provides 100% protection, MPDA-based vaccination provides 100% protection. Mechanistically, MPDA exhibits high antigen-loading efficiency, facilitates cytosolic delivery of Gn, enhances dendritic cell activation, and elicits substantially higher SFTSV-specific IgG and neutralizing antibody titers than the commercial aluminum hydroxide gel adjuvant (Alum). This work establishes MPDA as a multi-function delivery and adjuvant platform, offering a promising strategy for SFTSV subunit vaccine development.

The trans-cleavage activity of CRISPR/Cas systems has catalyzed significant progress in molecular diagnostics. Compared with traditional methods such as polymerase chain reaction (PCR) and its derivatives, CRISPR/Cas diagnostics are often credited with high specificity, portability, and visual readout. Among various CRISPR systems, CRISPR/Cas9, CRISPR/Cas12, and CRISPR/Cas13 have been extensively applied in pathogen detection owing to their distinct target-recognition and nucleic acid-cleavage mechanisms. In particular, Cas12- and Cas13-based systems exploit target-activated trans-cleavage activity for sensitive signal amplification, whereas Cas9-based diagnostic platforms generally rely on sequence-specific cis-cleavage. This review assesses the integrated CRISPR/Cas detection workflow from sample collection and processing through final result output, and systematically analyzes the intrinsic characteristics of Cas effector proteins with respect to target enrichment, reporter molecules, readout formats, sample background, and validation design. Based on a practical application-oriented framework, we analyzed the adaptability of various CRISPR/Cas systems in distinct scenarios, including point-of-care screening, quantitative laboratory testing, and multiplex pathogen identification. In addition, we highlight engineering innovations derived from mechanistic investigations of Cas9, Cas12, Cas13 and Class I CRISPR systems, discuss the specific diagnostic bottlenecks these effectors can resolve, and outline remaining challenges requiring further optimization prior to clinical translation.

Rabies virus (RABV) is a neurotropic virus that infects nearly all warm-blooded mammals and is almost invariably fatal once symptoms appear. Previous studies indicate that RABV uses its structural proteins to hijack host factors or modulate host signaling pathways to promote viral replication and pathogenicity. In this study, we observed upregulated expression of the host factor TGF β activated kinase 1 binding protein 2 (TAB2) in N2a cells following RABV infection. Loss and gain of function experiments confirmed that TAB2 overexpression enhances RABV replication, whereas TAB2 knockout suppresses it. We further demonstrated that TAB2 promotes RABV replication by activating the TGF β activated kinase 1 (TAK1)-p38 mitogen activated protein kinase (MAPK) signaling pathway. Co immunoprecipitation assays revealed an interaction between endogenous TAB2 and RABV M protein during infection. Further mapping showed that two TAB2 domains (aa 51-574 and aa 575-693) mediate this interaction; however, only the C terminal region (aa 575-693)-which contains ubiquitin modification sites and mediates TAK1 binding-is essential for the proviral function of TAB2. Mechanistically, early in RABV infection, M protein interacts with TAB2 and inhibits its K48 linked ubiquitination, leading to TAB2 stabilization and enhanced assembly of the TAB2-TAK1 complex. This process suppresses apoptosis via activation of the TAK1-p38/MAPK pathway and ultimately facilitates viral replication. These findings were corroborated in a TAB2 knockdown mouse infection model. Our study reveals a mechanism by which RABV M protein hijacks the host TAB2-TAK1-p38/MAPK signaling axis to inhibit apoptosis at early stage of infection, highlighting potential therapeutic targets for rabies intervention.

Tick-borne encephalitis (TBE) is an important zoonotic viral disease transmitted by ticks. In recent decades, global climate change has increased human exposure to ticks, and mortality rate have gradually risen. Effective vaccines are essential for controlling TBE as specific antiviral treatment is unavailable. Vaccine candidates based on virus-like particles (VLPs) have previously been demonstrated to be effective in eliciting excellent immune responses against influenza virus and SARS-CoV-2. Here, we constructed TBE virus (TBEV) VLPs containing the envelope and membrane proteins derived from the Far Eastern TBEV strain (WH2012) using an insect cell-baculovirus expression system. Induction of immune responses was investigated in mice following intramuscular injection with the TBEV VLPs vaccine candidates formulated with a combination of poly(I:C) and Montanide ISA201VG adjuvants. Mice produced memory T-cells and serum-specific IgG antibodies that averaged up to 1:104.6 and remained at 1:104 (mean) at 24 wk after three immunizations. TBEV VLPs vaccine was able to provide long-term antibody protection against TBEV, making it a promising subunit vaccine candidate for this disease.

Conjugated polymers (CPs) exhibiting aggregation-induced emission (AIE) properties have found extensive applications in the biomedical sector due to their strong solid-state emission, enhanced light absorption capabilities, and effective production of reactive oxygen species (ROS). However, an explanation of the structure-function relationship was not provided. Herein, four CPs with AIE characteristics were synthesized by varying the composition and combination of donor-acceptor (D-A), among which PADAD' exhibited the highest ROS generation efficiency due to its D-A effect and aggregation-induced ROS generation ability. Furthermore, functionalized CPs were designed as a toolbox for biological applications using PADAD' as the skeleton. As a proof of concept, PADAD'-APT and PADAD'-GUA were synthesized by introducing target motifs designed to identify and photoinactivate tumor cells and bacteria, respectively. Thus, this platform holds great promise for biomedicine and provides ideas for the development of functionalized conjugated polymers.

The viruses of the genus Henipavirus within the family Paramyxoviridae are highly pathogenic and often associated with severe diseases in animals and humans (Basler 2012).For example,Nipah virus (NiV) and Hendra virus (HeV),two members of the genus Henipavirus,are known to infect humans and cause fatal disease (Field 2016;Singh et al.2019).

Dear Editor, The frequent emergence of life-threatening infectious diseases has posed a constant global threat in recent decades.

Rabies is a zoonotic infection with the potential to infect all mammals and poses a significant threat to mortality. Although enzyme-linked immunosorbent tests and real-time reverse transcription-quantitative polymerase chain reaction (RT-qPCR) have been established for rabies virus (RABV) detection, they require skilled staff. Here, we introduce a personal glucose meter (PGM)-based nucleic acid (NA-PGM) detection method to diagnose RABV. This method ensures sensitive and convenient RABV diagnosis through hybridization of reverse transcription-recombinase aided amplification (RT-RAA) amplicons with probes labeled with sucrose-converting enzymes, reaching a detection level as low as 6.3 copies/μL equivalent to 12.26 copies. NA-PGM allows for the differentiation of RABV from other closely related viruses. In addition, NA-PGM showed excellent performance on 65 clinical samples with a 100% accuracy rate compared with the widely adopted RT-qPCR method. Thus, our developed NA-PGM method stands out as sensitive, semiquantitative, and portable for RABV detection, showcasing promise as a versatile platform for a wide range of pathogens.

>Highlights The RT-RAA-VF assay developed for the NiV P gene can perform rapid detection of NiV within 20 min at 42°C with high specificity.This assay is capable of attaining sensitivity to a single copy of NiV RNA transcripts.This assay effectively avoids false positives caused by aerosol contamination with a sealed disposable nucleic acid visualization test paper device.

Nipah virus (NiV) is an emerging bat-borne zoonotic virus that can be transmitted to humans and other animals through infected bats or contaminated foods. The disease is highly lethal in humans (40%-75%) and has the potential for human-to-human transmission. Currently, there are no approved treatments or vaccines for NiV infection in humans or animals. Consequently, there is a pressing need for a highly sensitive, precise, and visually detectable assay to enable early intervention and mitigate the transmission of NiV infection. Here, we report a single-copy sensitive, field-deployable, one-pot visual reverse transcription-recombinase polymerase amplification (RT-RPA)-clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR associate system (Cas)12 for the detection of NiV. The assay works by targeting the N gene of NiV, and the results are directly visible to the naked eye. The assay has demonstrated the ability to detect as few as 5.5 copies/μl of positive plasmids or 5.5 × 101 copies/μl of RNA transcripts when reacted at constant temperature for 40 min. It showed high specificity for NiV and had no cross-reaction with other pathogens, including rabies virus (RABV), Japanese encephalitis virus (JEV), herpes simplex virus type 1 (HSV-1), Hendra virus (HeV), and Streptococcus suis (S. suis), that can cause clinical symptoms similar to those of NiV infection. Moreover, this assay had a 100% coincidence rate with the reverse transcription quantitative polymerase chain reaction (RT-qPCR) method recommended by the World Organization for Animal Health (WOAH) for the detection of simulated clinical samples, indicating that it has great potential as an ultrasensitive, simple, and portable novel assay for the onsite diagnosis of NiV infection.

Rabies, which is caused by the rabies virus (RABV), is a neurological disease of mammals, including humans. The disease is untreatable, as drugs and antibodies are prevented from entering brain tissue by the blood-brain barrier (BBB). However, it has been reported that BBB permeability can be enhanced in mice infected with a laboratory-attenuated strain of RABV, which is not the case in those infected with wild-type RABV. Moreover, it is not the RABV infection directly that enhances BBB, but rather the inflammatory molecules that are induced by the virus. Understanding the mechanism underlying the RABV-induced production of chemokines and cytokines is crucial for the prevention and management of rabies. In this study, we found that mice infected with the RABV challenge strain CVS11 showed significant inflammatory infiltration in the later stages of infection. At the cellular level, we confirmed that the NF-κB signaling pathway was activated following RABV infection, leading to significant upregulation of the expression of downstream chemokines and inflammatory factors (CCL2, CCL5, CXCL10, TNF-α, and IL-6). Surprisingly, the replication of RABV was upregulated when the NF-κB pathway was inhibited, and the levels of its downstream chemokines and inflammatory factors were reduced following treatment with the inhibitor JSH-23. This suggests that RABV infection activates the NF-κB pathway, which is then able to negatively regulate the replication of RABV. This pathway is therefore a potential target for drugs and other therapies in the treatment of rabies.

Feline viral rhinotracheitis (FVR) is caused by the feline herpesvirus-1 (FHV-1), which commonly results in upper respiratory symptoms, and can result in death in the kittens and weak cats. Rabies is an infectious disease with zoonotic characteristics highly relevant to public health and also poses a serious threat to cats. Vaccines are the most effective method to control the spread of both FHV-1 and RABV and have the advantage that they produce long-term specific immune responses. In this study, we constructed a bivalent vaccine against FHV-1 and rabies virus (RABV) simultaneously. The vaccine was constructed by cloning FHV-1 gB into a RABV based vector, and the recombinant RABV (SRV9-FHV-gB) expressing the FHV-1 gB protein was rescued. The growth characteristics of SRV9-FHV-gB were analyzed on NA and BSR cells. To assess the immunogenicity of the vaccine, mice and cats were immunized with SRV9-FHV-gB supplemented with Gel02 adjuvant. The SRV9-FHV-gB exhibited the same growth characteristics as the parent virus SRV9 in both BSR cells and NA cells. The safety of SRV9-FHV-gB was evaluated using 5-day-old and 14-day-old suckling mice. The results showed that mice infected with the SRV9-FHV-gB survived for longer than those in the SRV9 group. Mice immunized with inactivated SRV9-FHV-gB produced high titers of specific antibodies against FHV-1 and neutralizing antibodies against RABV. Cats that received three immunizations with SRV9-FHV-gB also produced neutralizing antibodies against both FHV-1 and RABV. This study represents the first time that a bivalent vaccine targeting FHV-1 and RABV has been constructed, laying the foundations and providing inspiration for the development of other multivalent vaccines.

The rapid emergence of Severe Acute Respiratory Syndrome Coronavirus type 2 (SARS-CoV-2) variants, coupled with severe immune evasion and imprinting, has jeopardized the vaccine efficacy, necessitating urgent development of broad protective vaccines. Here, we propose a strategy employing recombinant rabies viruses (RABV) to create a universal SARS-CoV-2 vaccine expressing heterologous tandem receptor-binding domain (RBD) trimer from the SARS-CoV-2 Prototype, Delta, and Omicron strains (SRV-PDO). The results of mouse immunization indicated that SRV-PDO effectively induced cellular and humoral immune responses, and demonstrated higher immunogenicity and broader SARS-CoV-2 neutralization compared to the recombinant RABVs that only expressed RBD monomers. Moreover, SRV-PDO exhibited full protection against SARS-CoV-2 in the challenge assay. This study demonstrates that recombinant RABV expressing tandem RBD-heterotrimer as a multivalent immunogen could elicit a broad-spectrum immune response and potent protection against SARS-CoV-2, making it a promising candidate for future human or veterinary vaccines and offering a novel perspective in other vaccine design.

Crimean-Congo hemorrhagic fever（CCHF） is a zoonotic disease caused by the CCHF virus（CCHFV）, which is primarily transmitted by ticks（Lorenzo Juanes et al. 2023）. It is an emerging disease that occurs sporadically in Africa, Asia, and Europe, with a high morbidity and mortality rate, as high as 30% in humans（Ceylan et al. 2013）. CCHFV, belonging to genus Nairovirus,

Rabies is a zoonotic neurological disease caused by the rabies virus (RABV) that is fatal to humans and animals. While several post-infection treatment have been suggested, developing more efficient and innovative antiviral methods are necessary due to the limitations of current therapeutic approaches. To address this challenge, a strategy combining photodynamic therapy and immunotherapy, using a photosensitizer (TPA-Py-PhMe) with high type I and type II reactive oxygen species (ROS) generation ability is proposed. This approach can inactivate the RABV by killing the virus directly and activating the immune response. At the cellular level, TPA-Py-PhMe can reduce the virus titer under preinfection prophylaxis and postinfection treatment, with its antiviral effect mainly dependent on ROS and pro-inflammatory factors. Intriguingly, when mice are injected with TPA-Py-PhMe and exposed to white light irradiation at three days post-infection, the onset of disease is delayed, and survival rates improved to some extent. Overall, this study shows that photodynamic therapy and immunotherapy open new avenues for future antiviral research.

Background Rabies, caused by the rabies virus (RABV), is an ancient and neglected zoonotic disease posing a large public health threat to humans and animals in developing countries. Immunization of animals with a rabies vaccine is the most effective way to control the epidemic and the occurrence of the disease in humans. Therefore, the development of cost-effective and efficient rabies vaccines is urgently needed. The activation of dendritic cells (DCs) is known to play an important role in improving the host immune response induced by rabies vaccines. Methodology/Principal findings In this study, we constructed a recombinant virus, rCVS11-MAB2560, based on the reverse genetic system of the RABV CVS11 strain. The MAB2560 protein (a DC-targeting molecular) was chimeric expressed on the surface of the viral particles to help target and activate the DCs when this virus was used as inactivated vaccine. Our results demonstrated that inactivated rCVS11-MAB2560 was able to promote the recruitment and/or proliferation of DC cells, T cells and B cells in mice, and induce good immune memory after two immunizations. Moreover, the inactivated recombinant virus rCVS11-MAB2560 could produce higher levels of virus-neutralizing antibodies (VNAs) in both mice and dogs more quickly than rCVS11 post immunization. Conclusions/Significance In summary, the recombinant virus rCVS11-MAB2560 chimeric-expressing the molecular adjuvant MAB2560 can stimulate high levels of humoral and cellular immune responses in vivo and can be used as an effective inactivated rabies vaccine candidate.