Transboundary and Emerging Diseases最新文献

Pentatrichomonas hominis (P. hominis) and Tritrichomonas foetus (T. foetus) are prevalent intestinal protozoa. P. hominis is associated with chronic diarrhea in humans and animals, whereas T. foetus causes gastrointestinal disease in companion animals and reproductive-tract infection in cattle. Rapid and accurate identification of these infections at the point-of-care (POC) is crucial for the diagnosis and effective management of zoonotic diseases. In this study, we developed two novel recombinase polymerase amplification (RPA) assays coupled with CRISPR/Cas12a detection. The dual-species assay, using a lateral-flow format, targeted species-specific regions of the 18S rRNA gene of P. hominis and T. foetus, and under ideal conditions, delivered visual results within 40 min for a single sample at 37°C. P. hominis-specific assay: To differentiate P. hominis in mixed infections with T. foetus, a second assay targeted the highly conserved Spo11-1 gene of P. hominis. Optimal crRNA-412 and RPA primers were selected for maximal Cas12a cleavage efficiency. Analytical sensitivity and specificity were compared with conventional nested polymerase chain reaction (PCR) and Sanger sequencing. The results showed that The dual-species assay detected as few as 50 DNA copies/µL of either parasite with no cross-reactivity to Giardia lamblia, Cystoisospora canis, Cryptosporidium spp., Toxoplasma gondii, Toxocara canis, and Toxascaris leonina. Among 70 fecal samples of companion animal (48 dogs and 22 cats), 14 (29.2%) dogs tested positive for P. hominis, and eight cats (36.4%) tested positive for T. foetus by nested PCR. Due to financial and logistical constraints, we selected a smaller subset for subsequent analysis with the RPA-CRISPR/Cas12a lateral-flow strip (LFS) assay, which showed 100% diagnostic concordance with PCR. The Spo11-1 assay achieved a limit of detection of 20 DNA copies/µL and specifically recognized P. hominis among a panel that included seven non-target protozoa and helminths. Validation on 10 additional canine and feline samples (four positives and six negatives) showed complete agreement with nested-PCR results. In conclusion, this CRISPR–based diagnostic approach significantly enhances the efficiency and accuracy of Trichomonads detection, offering a practical, cost-effective solution particularly suitable for veterinary and potentially human healthcare diagnostics in resource-limited settings.

Pseudorabies virus (PRV), the causative agent of Aujeszky’s disease, leads to great economic losses on swine production worldwide. Receptor of activated protein kinase C 1 (RACK1) is initially known as a receptor for protein kinase C, and recent studies indicate that RACK1 can also play critical roles in various virus infections. However, the role of RACK1 during PRV infection has not yet been determined. In this investigation, we observed a strong positive correlation between the expression levels of RACK1, interferon-β (IFN-β), and the IFN-stimulated gene 15 (ISG15) and ISG20 in PRV-infected porcine kidney-15 (PK-15) cells at 24 h postinfection. Further experiments revealed that RACK1 exerted an inhibitory effect on PRV replication and enhanced the activation of the Type I IFN (IFN-I) signaling pathway. Interestingly, RACK1 was found to facilitate stimulator of IFN genes (STING)–dependent phosphorylation of IFN regulatory factor 3 (IRF3). More specifically, RACK1 could interact with STING and then promote aggregation of STING around the Golgi apparatus. Taken together, these findings demonstrated that RACK1 could associate with STING to promote IFN-I activation and inhibit PRV infection. These results will provide new data on host factors that limit PRV infection, and facilitate our understanding of IFN-I-mediated antiviral responses during PRV infection.

Lumpy skin disease (LSD) is a disease of cattle and other ruminants caused by the LSD Virus (LSDV). LSD infections are characterized by high morbidity, low mortality, and significant economic impact. Initially endemic to Africa only, LSD has spread to the Middle East, Europe, and Asia in the past decade. In 2023, LSD posed a significant threat to Bhutan’s livestock population, with outbreaks reported across 20 Dzongkhags (districts) and 192 Gewogs (groups of villages). This study investigated the epidemiology and molecular aspects of the outbreak. A total of 19,907 animals (16,728 cattle and 3179 yaks) were affected, with 2888 deaths recorded, leading to an apparent case fatality rate (CFR) of 9.92% in cattle and 38.66% in yaks. Molecular characterization of the LSDV-positive samples from cattle and yaks based on LSDV-differentiating genes (i.e., RPO30, GPCR, EEV, and B22R) revealed 100% similarity among the samples, clustering them with LSDV field isolates from Sudan, India, and China in Clade 1.2. Further whole-genome sequence characterization of a representative sample (LSDV_Bhutan_03) from cattle skin scrapings and phylogenetic network analysis clustered the virus with Neethling Warmbaths (NW)-like LSDVs (Clade 1.2.2). Within the NW-like clade, the Bhutan LSDV was closely related to recent isolates from cattle and buffalo in India and yaks in China. These data highlight the importance of LSDV surveillance in both domestic and wild bovines to identify spillover incidences, understand the extent of disease spread, and strengthen control measures.

Viral diseases in chickens, such as avian influenza, Newcastle disease, and infectious bronchitis, cause significant economic losses to the global chicken industry, while the cross-species transmission risks of avian influenza viruses (AIVs) pose potential threats to public health security. Conventional inactivated vaccines and live attenuated vaccines exhibit limitations in terms of protective coverage, immunization duration, and safety profiles, necessitating the development of novel vaccine strategies. In recent years, the application of nanotechnology has been bringing new opportunities for chicken vaccine development. As innovative vaccine carriers, nanoparticles demonstrate unique advantages through their size effects, surface modifiability, and antigen-loading capacity, enabling precise regulation of antigen delivery efficiency and enhance innate immune responses via activation of pattern recognition receptors. This review summarizes recent advancements in nanoparticle-based vaccines for chickens, with particular emphasis on nanocarrier design principles, immunological mechanisms, and protective efficacy. The potential of these systems to improve immune responses and extend protective duration is systematically analyzed, with future research priorities outlined to guide the development of next-generation chicken vaccines.

Far Eastern (FE) tick-borne encephalitis virus (TBEV) was thought to be the only endemic subtype in northeastern China. However, in 2024, the Siberian (Sib) subtype of TBEV was found in ticks. This study investigates Sib-TBEV infection in tick-bitten patients in the region. A total of 2573 tick-bitten patients, including 506 inpatients and 2067 outpatients, were enrolled between April and September in both 2023 and 2024 and were tested for Sib-TBEV using real-time quantitative polymerase chain reaction (RT-qPCR). Seven cases were confirmed, with four among inpatients and three among outpatients. All four inpatients exhibited headaches and had hospital stays ranging from 5 to 21 days. Two inpatients with longer hospital stays showed additional symptoms: one experienced dizziness and fatigue, while the other exhibited liver injury and neurological symptoms, including facial paralysis, dysarthria, and meningeal signs. All patients recovered and were discharged without sequelae. Seven Sib-TBEV complete genomes were amplified and classified into the Sib-TBEV Zausaev and Vasilchenko lineages. Bayesian analysis estimated that the most recent common ancestor (tMRCA) of these strains dates to a period between 1898 and 1983, with possible migration pathways from Irkutsk or Zabaykalsky to the Daxing’an Mountains (DXAM) in northeastern China. We identified Sib-TBEV-infected patients in northeastern China and characterized their epidemiological and clinical features. Sib-TBEV may have been circulating in northeastern China for decades, underscoring the need for TBEV surveillance in ticks, animals, and tick-bitten patients.

Although several biosecurity and control measures are currently in place to mitigate the African swine fever (ASF) epidemic, vaccination is being explored as a potential long-term strategy. However, standardized guidelines for evaluating the safety and efficacy of ASF vaccines are not yet fully established. Understanding infection dynamics in wild boar is crucial, as they play a key role in the spread and persistence of the virus. This work aims to provide comprehensive information on viral distribution through immunohistochemical analysis (p72) with histopathologic assessment in wild boar. The study design comprises animals: (i) intramuscular infected with the high-virulence genotype II isolate Arm07 (highly virulent isolate [HVI]; n = 6); (ii) orally vaccinated with the low virulence isolate Lv17/WB/Rie1-ΔCD (low virulent isolate [LVI]; n = 6); and (iii) orally vaccinated with Lv17/WB/Rie1-ΔCD, either with a single dose (LVI-HVI1; n = 6) or repeated doses (LVI-HVI2; n = 6), followed by intramuscular challenge with Arm07. Clinical monitoring, viral load quantification in blood and tissues via real-time quantitative PCR, and virus viability in tissue cultures using peripheral blood mononuclear cells were performed. HVI animals had hemorrhagic and inflammatory lesions, along with generalized lymphoid depletion, correlated with widespread viral dissemination. LVI animals rarely showed mild lymphoid depletion of the lymph nodes; minimal immunostaining was observed in macrophages of the tonsils and lymph nodes, typically restricted to the oral entry point. A few LVI–HVI1 cases had infected resident sinus macrophages related to necrotic lesions at tonsils and lymph nodes, preventing the virus from disseminating to vital organs. No viral immunostaining or associated histopathologic lesions were observed in LVI–HVI2 animals, indicating that revaccination enhances safety against virulent challenges. Observed changes following vaccination do not reflect chronic infection but rather a transient one, followed by lymphoid system recovery. Immunohistochemical and histological evaluation has proven valuable in advancing our understanding of ASF pathogenesis in wild boar, contributing to improved vaccination safety and disease management strategies.

Feline calicivirus (FCV) is a significant pathogen in cats, with sporadic outbreaks of infections with virulent systemic (VS-FCV) strains causing significant health problems. Nineteen FCV strains were isolated and identified in China from 2021 to 2022. The nucleotide and amino acid phylogenetic analysis of the VP1 gene showed that 16 strains were GI genotype and three strains were GII genotype. HBDL2 strain was further found to be in the same clade as the reported VS-FCV SH/2014 strain, with 87.1% nucleotide sequence identity and 93.0% amino acid sequence identity. However, given that phylogenetic and homology analysis alone is insufficient to predict virulence, the pathogenic potential of HBDL2 was subsequently assessed through experimental infection in cats. The results revealed that HBDL2 was able to cause systemic clinical signs and caused severe tracheal and lung damage, with a similar characterization as the VS-FCV strain. Serum neutralization assays confirmed that HBDL2 elicited broad-spectrum neutralizing antibodies in cats against multiple FCV strains, including diverse GI and GII genotypes. Notably, neutralizing antibody titers against the VS-FCV strains were elevated. Additionally, we established an infectious clone of HBDL2 as a critical technical tool. These findings indicate that the HBDL2 strain holds promise for use in vaccine development against FCV infection.

Porcine circovirus type 2 (PCV2) is a major swine pathogen causing significant economic losses in the swine industry worldwide. Continual monitoring of genetic and antigenic diversity is essential for the early detection of emerging variants. This study investigates the evolutionary dynamics and genetic variation of PCV2 circulating in Thailand and across Asia from 2007 to 2024, using integrated phylogenetic and phylodynamic analyses. Analysis of 2739 PCV2 genomes, after excluding 0.99% recombinant strains, revealed four main groups circulating in Asia, with PCV2d as the predominant genotype. PCV2d has diversified into several distinct clades, including a recently identified variant with the ¹³³HDAM¹³⁶ amino acid motif, which likely originated from earlier variants, the ¹³³ANAL¹³⁶ and ¹³³ATAL¹³⁶ motifs. Recombination analysis detected intragenotypic recombination events within PCV2d strains circulating in Thailand, emphasizing the role of genetic recombination in driving the evolutionary changes of the virus. Phylodynamic analysis demonstrated significant fluctuations in the virus population size, correlating with changes in genotype dominance over time. Selective pressure analysis identified positively selected sites in the capsid protein (codons 63, 131, 134, 169, and 190), indicating ongoing adaptation under host immune pressures. Structural modeling and epitope analyses revealed mutations affecting antigenic sites and immune recognition, suggesting concerns for vaccine efficacy. This integrated approach enhances our understanding of PCV2 evolution, informing strategies for effective vaccine development and disease control.

In October 2019, the first cases of African swine fever (ASF) were confirmed in wild boar in the Republic of Korea (ROK). Since then, ASF has continued to spread throughout the country, particularly among wild boar populations, despite intensive efforts to contain the outbreak. Our objective was to assess the results of current ASF surveillance in wild boars and identify various risk factors related to ASF outbreak in ROK. Between September 2019 and June 2023, a total of 122,078 wild boar samples were submitted, with 3135 tested as positive to ASF infection. Among them, 90.6% (2839/3135) of ASF-positive cases were detected from carcasses. Within the carcass samples, ASF prevalence showed an increase from 5.98% (95% confidence interval [CI] 4.47–7.81) to 37.15% (95% CI 34.12–40.27) through years 2019 and 2023, respectively. Geographically, from the demilitarized zone (DMZ), the region where ASF was first detected in wild boars, the disease has spread toward the east and south direction across the Baekdu mountain range, which is a continuous forest habitat. The movement of the ASF outbreak region between years has been relatively constant for the last 45 months, except for between 2020 and 2021 due to rapid spread during the year 2021. From the ASF-specific antibody assay, only two wild boars were seropositive out of 17,275 serum samples. This is the first study presenting results and patterns of wild boar surveillance and ASF virus prevalence changes in the ROK during 2019–2023. The study showed that output of surveillance and control effort, as well as the epidemiologic course of ASF in ROK is potentially associated with various spatial and anthropogenic factors, with subsets of factors more linked with specific circumstances of ROK. A better understanding regarding the past and current status of the disease will contribute to improving control measures required for preventing further spread of the ASF virus in the Korean peninsula.