Search Results (381)

Oropouche fever (OF), caused by Oropouche virus (OROV), has expanded beyond its Amazonian range into Minas Gerais (MG), Brazil, raising concern about transmission in extra-Amazonian Atlantic Forest landscapes. Critical gaps persist regarding Culicoides vector communities, anthropophily, and climate-sensitive transmission risk in these newly affected regions. We conducted targeted entomological surveys outbreak-driven by human OF cases, standardized across five MG communities using CDC light traps and Protected Human Attraction (PHA) to characterize Culicoides composition. Females of Culicoides underwent RT-qPCR for OROV (n = 819) and physiological assessment (n = 312). We developed an entomological alert framework that integrates blood-fed abundance, minimum infection rate (MIR) upper confidence bounds, and environmental drivers (i.e., mean temperature, relative humidity and precipitation) via generalized additive mixed models, which explained 68% of the variability in Culicoides abundance and the alert index across communities. We collected 1171 Culicoides individuals representing five species (C. leopoldoi, C. paraensis, C. pusillus, C. foxi, and C. limai). C. leopoldoi (79.1%) and C. paraensis (20.3%) were the predominant species; notably, C. paraensis is recognized as the primary vector of OROV in the Americas. C. paraensis was documented for the first time in all five outbreak areas and dominated PHA captures (90%), suggesting anthropophily. Although no specimens tested OROV-positive (consistent with expected field infection rates of 0.01–1%), MIR upper bounds reached 132/1000 in low-sample settings and humidity and temperature strongly modulated abundance. This operational baseline and alert index transform virologically negative, sparse surveillance data into prioritized targets for intensified sampling and vector control during early, low-prevalence phases, when containment of OROV’s extra-Amazonian spread is still achievable. Full article

Background/Objectives: Q fever, caused by Coxiella burnetii, is typically treated with doxycycline, but its efficacy is limited in chronic cases and may be poorly tolerated. Systemic ciprofloxacin shows limited activity for acute Q fever. However, inhaled liposomal formulations may provide therapeutic benefit. Methods: Two inhaled ciprofloxacin formulations (Lipoquin^® and Apulmiq^®) were evaluated in an A/J mouse model of Q fever and compared with intraperitoneal ciprofloxacin and oral doxycycline. Initially, pharmacokinetic studies were performed to determine an appropriate dosing regimen for the inhaled ciprofloxacin formulations. A separate cohort of mice were then infected with C. burnetii and treated once daily via nebulisation with Lipoquin or Apulmiq, initiated at 24, 48, or 72 h post-challenge. Clinical signs, weight change, splenomegaly, bacterial burden, and lung histopathology were evaluated. Results: Pharmacokinetic analysis confirmed sustained lung concentrations of inhaled ciprofloxacin, supporting once-daily dosing. Inhaled Lipoquin and Apulmiq significantly reduced clinical signs, weight loss, splenomegaly, and pulmonary bacterial burden compared to untreated controls and doxycycline-treated mice. Histopathology revealed decreased lung inflammation and lesion severity following inhalational dosing. Systemic ciprofloxacin slightly reduced splenic bacterial burden but was less effective in controlling pulmonary infection. Conclusions: Inhaled liposomal ciprofloxacin demonstrated superior protection and reduced respiratory manifestations of Q fever compared to doxycycline and systemic ciprofloxacin. These findings suggest inhaled formulations may represent a viable alternative for the treatment of Q fever pneumonia. Further studies are needed to evaluate clinical applicability and long-term outcomes. Full article

Objectives: Familial Mediterranean fever (FMF) is an autoinflammatory disease caused by MEFV mutations, leading to recurrent fever and inflammation. Dysregulation of innate and adaptive immunity, including altered expression of microRNAs and immune regulatory molecules, may contribute to disease heterogeneity. The role of CTLA-4, DTX1, and selected miRNAs in FMF pathogenesis remains unclear. Methods: We conducted a case–control study including 48 pediatric FMF patients and 36 age- and sex-matched healthy controls. Serum miR-204-3p and miR-223-3p levels were assessed via qRT-PCR. Plasma concentrations of pyrin, CTLA-4, and DTX1 were measured using ELISA. Clinical data and MEFV mutation types were analyzed in relation to biomarker levels. Results: There was no statistical significance between the groups in plasma CTLA-4 levels. Serum miR-204-3p, miR-223-3p, and plasma DTX1 levels were found to be significantly lower in FMF patients, while plasma pyrin levels (p < 0.05, in all) were significantly higher. CTLA-4 levels were positively correlated with pyrin and DTX1 levels (r = 0.602; p < 0.001; r = 0.740; p < 0.001, respectively). Conclusions: miR-204-3p and miR-223-3p may be associated with FMF pathogenesis. Increased levels of the pyrin protein, encoded by the MEFV gene, may have an important role in apoptotic and inflammatory signaling pathways. A decrease in DTX1 levels and a positive correlation between DTX1 and CTLA-4 suggest that subclinical inflammation may continue in attack-free periods in FMF patients. Full article

Tick-borne pathogens pose a significant threat to human health. In this study, a multiple droplet digital PCR (ddPCR) assay was developed to detect four tick-borne pathogens: Borrelia burgdorferi sensu lato (Bbsl), Coxiella burnetii (C. burnetii), spotted fever group Rickettsia (SFGR), and Borrelia miyamotoi (B. miyamotoi). Based on the singleplex ddPCR reaction system of Bbsl, the primer probes of the other three species were incorporated to develop a multiplex ddPCR reaction system. The annealing temperature and the final concentration of the primer probes were then optimized for multiplex ddPCR. The multiplex ddPCR assay was assessed for its sensitivity, specificity, repeatability, and ability to detect simulated and actual samples. The developed multiplex ddPCR approach enables the simultaneous detection of Bbsl, C. burnetii, SFGR, and B. miyamotoi. The positive target microtitre clusters are closely grouped and distinctly separated from each other, with the multiplex ddPCR assay demonstrating a dynamic range of five orders of magnitude. The limits of detection (LOD) for the multiplex ddPCR assay were 4 copies/20 µL for Bbsl, 3 copies/20 µL for C. burnetii, 3 copies/20 µL for SFGR, and 2 copies/20 µL for B. miyamotoi. The assay demonstrated high specificity, with no observed cross-reactivity against non-target pathogens. Performance was validated using both spiked samples and field-collected clinical specimens. In the evaluation of 30 ticks and 30 serum samples, the ddPCR method (in both singleplex and multiplex formats) achieved higher positive detection rates for all four target pathogens compared to quantitative real-time PCR (qPCR). In addition, the detection proportions of multiplex and singleplex ddPCR were consistent. Multiplex ddPCR can detect low DNA concentrations in samples and enables the absolute quantification of Bbsl, C. burnetii, SFGR, and B. miyamotoi, providing a novel detection approach for the clinical diagnosis of tick-borne diseases. Full article

Highly pathogenic avian influenza (HPAI) A/H5N1 has emerged as a cause of severe disease in domestic cats, but clinical data from field outbreaks remain limited. We retrospectively reviewed medical records, laboratory results, and ancillary examinations from 22 domestic cats with RT-qPCR-confirmed A/H5N1 infection diagnosed in Poland in June 2023. To the best of our knowledge, we report the first comprehensive retrospective case series from the 2023 Polish outbreak, combining 22 laboratory-confirmed cats with detailed clinical timelines and laboratory findings. For each cat, the temporal progression of clinical signs, hematology, serum biochemistry, and, when available, imaging findings were evaluated. Post-mortem examination data were not systematically available in this retrospective cohort. Notably, six of these cats were strictly indoor cats that received raw poultry meat as part of their diet. Disease onset was acute, with fever, lethargy, and anorexia rapidly progressing to severe dyspnea and neurological signs, including ataxia, seizures, and paraplegia; case fatality was 100%, with a typical interval of ≤3 days from first signs to death or euthanasia. Hematologic changes were dominated by thrombocytopenia, lymphopenia, and marked eosinopenia, consistent with a systemic inflammatory/stress leukogram. Biochemistry indicated marked tissue injury, with increased AST, LDH, and CK activities, whereas creatinine and urea remained largely within reference intervals, arguing against primary renal failure. Imaging supported the presence of interstitial to diffuse pneumonia. These data characterize the clinical and laboratory phenotypes of feline A/H5N1 infection and underscore its importance as a rapidly fatal respiratory and neurological disease with One Health implications. Full article

The global swine industry suffers persistent economic losses and health challenges due to major viral pathogens such as African swine fever virus (ASFV), porcine reproductive and respiratory syndrome virus (PRRSV), classical swine fever virus (CSFV), and porcine circovirus (PCV). Traditional diagnostic methods, including virus isolation, serology, and quantitative PCR (qPCR), are limited by time, equipment requirements, and field applicability. Recent advances in CRISPR-based diagnostics, particularly those leveraging the collateral cleavage activity of Cas12a and Cas13a, have enabled rapid, sensitive, and field-deployable nucleic acid detection. This review outlines the principles of CRISPR-Cas12a/13a systems, their integration with isothermal amplification techniques, and their application in detecting major swine viruses. Cas12a-based platforms (e.g., DETECTR) and Cas13a-based systems (e.g., SHERLOCK) achieve detection limits as low as single-copy/μL within 25–60 min at 37 °C, offering high specificity and compatibility with visual readouts. Applications include ASFV, PRRSV, CSFV, PCV, foot-and-mouth disease virus (FMDV), porcine rotavirus (PoRV), and porcine parvovirus 7 (PPV7). Despite significant advances, challenges remain, notably the reliance on nucleic acid extraction and the need for fully integrated “sample-in, result-out” systems. Ongoing innovations in extraction-free methods, lyophilized reagents, and multiplex detection will strengthen the role of CRISPR diagnostics in swine disease surveillance and control. From an application standpoint, the technology offers a low-capital, field-adaptable alternative to qPCR, with its value proposition rooted in early outbreak containment and loss prevention. Its adoption pathway is expected to vary across production systems—serving as a sentinel tool in intensive settings, a leapfrogging solution in rapidly intensifying regions, and through shared-service models in resource-limited contexts. However, translation to routine use still requires overcoming standardization hurdles, regulatory validation, and workflow integration. Full article

Candidatus Rickettsia colombiensis is a new candidate species of Rickettsiae spotted fever group that have been isolated only from ticks. The pathogenicity of Ca. R. colombiensis to human and animals is unknown. This study evaluated the pathogenic potential of Ca. R. colombiensis in Syrian hamsters and assessed the cross-reactivity between Ca. R. colombiensis and other Rickettsia in human and hamster sera. Shell vial technique was employed to isolate Ca. R. colombiensis. Subsequently, five male Syrian hamsters were inoculated intraperitoneally (IP) and five intradermally (ID) with 1 × 10⁶ Vero cells infected with Ca. R. colombiensis. One control hamster was used in each group. The health status was assessed daily, and necropsies were performed. Serum samples were tested by indirect immunofluorescence and tissues were processed by qPCR and histological stains. All Syrian hamsters remained healthy during the trial. No histopathological damages associated with rickettsial infection were observed. No Rickettsial DNA was detected in tissues. Syrian hamsters showed IgG antibody titers ranging from 1:64 to 1:1024. Control hamsters were negative. Regarding human sera, 56% (84/150) had IgG cross-reactivity antibodies against Ca. R. colombiensis. Subsequently, in a selected subset of 30 sera with moderate to high titers, all samples reacted with Ca. R. colombiensis antigen. Under specific conditions of this study, Ca. R. colombiensis did not behave as a highly virulent pathogen in the hamster model, although all infected Syrian hamsters developed IgG antibodies responses. Regarding cross-reactivity, it is possible to serologically diagnose rickettsial infection using Ca. R. colombiensis as an antigen. Full article

The flavivirus NS1 protein is a component of the viral replication complex and plays diverse, yet poorly understood, roles in the viral life cycle. To enable real-time visualization of the developing replication organelle and biochemical analysis of tagged NS1 and its interacting partners, we engineered a replication-competent yellow fever virus (YFV) replicon encoding a C-terminal fusion of NS1 with green fluorescent protein (NS1-GFP). The initial variant was non-viable in the absence of trans-complementation with wild-type NS1; however, viability was partially restored through the introduction of co-adaptive mutations in GFP (Q204R/A206V) and NS4A (M108L). Subsequent cell culture adaptation generated a 17-nucleotide frameshift within the NS1-GFP linker, resulting in a more flexible and less hydrophobic linker sequence. The optimized genome, in the form of a replicon, replicates in packaging cells that produce YFV structural proteins, as well as in naive BHK-21 cells. In the packaging cells, the adapted NS1-GFP replicon produces titers of infectious particles of approximately 10⁶ FFU/mL and is genetically stable over five passages. The expressed NS1-GFP fusion protein localizes to the endoplasmic reticulum and co-fractionates with detergent-resistant heavy membranes, a hallmark of flavivirus replication organelles. This NS1-GFP replicon provides a novel platform for studying NS1 functions and can be further adapted for proximity-labeling strategies aimed at identifying the still-unknown protease responsible for NS1-NS2A cleavage. Full article

Oropouche virus (OROV) has emerged as a significant arboviral pathogen in South America, responsible for recurrent outbreaks of febrile illness. In the Loreto region of Peru, more than 600 cases were reported in 2024, markedly exceeding expected incidence rates. We conducted a retrospective observational study using clinical–epidemiological records of all RT-qPCR-confirmed cases of Oropouche fever from the Regional Health Directorate of Loreto between October 2024 and March 2025. A total of 100 confirmed cases were identified. The most frequent symptoms were fever (88%), headache (78%), and myalgia (72%). No atypical or neurological presentations were reported. No severe cases or deaths occurred. Eight patients required hospitalization, mainly due to severe abdominal pain, persistent vomiting, arthralgia, and pregnancy. Six pregnant women were identified; three experienced pregnancy complications, though no fetal malformations or miscarriages were observed. This outbreak represents a new OROV epidemic in the region, with fewer cases than in 2024 and predominantly mild clinical courses. Although outcomes were generally favorable, the occurrence of complications in pregnant women underscores the importance of continued molecular surveillance and targeted public health interventions. Full article

Pathogens such as Toxoplasma gondii, lentiviruses (e.g., CAE), Hypoderma spp., Neospora caninum, Mycoplasma spp., and pestiviruses are important for goat farming in Lithuania; however, data on their prevalence remain limited. To address this gap, a multi-pathogen study was conducted between 2021 and 2024 using selected ELISA kits (ID.vet, Innovative Diagnostics, France). A total of 380 blood samples were collected from 30 goat herds across different regions of Lithuania; the sample size varied depending on the pathogen. Serum samples were tested for antibodies, and seroprevalence was calculated for each pathogen. The highest seroprevalence was detected for T. gondii (38.9%, 143/368) and CAE virus (19.5%, 74/380). Antibodies to Mycoplasma spp. (0.3%, 1/368), Hypoderma spp. (3.8%, 7/184), and N. caninum (0.5%, 2/368) were detected only sporadically, while no antibodies to Border disease virus or Q fever were identified. Mixed infections were found in 7.6% of samples. Chi-square analysis showed that co-infections with toxoplasmosis and CAE occurred more frequently than expected (χ² = 19.05, p < 0.001). Herd size was significantly associated only with CAE seroprevalence (χ² = 7.913, df = 1, p < 0.05). Overall, toxoplasmosis and CAE were identified as the most epidemiologically relevant infections in the Lithuanian goat population. Full article

This study integrates data on the prevalence, infection dynamics and risks associated with African swine fever virus (ASFV) outbreaks in Croatian wild boar during 2023–2024. Although the overall ASFV DNA prevalence in Croatia was 0.24%, the highest prevalence (2.29% in 2023 and 4.69% in 2024) was recorded in Vukovar-Srijem County. Genetic typing identified ASFV genotype II, subgroup 19, consistent with strains isolated from domestic pigs in Croatia and circulating in neighboring countries. Anti-ASFV specific antibodies were detected in 10.34% of wild boar tested in counties with previously reported DNA findings. In Vukovar-Srijem County, 4.60% of wild boar were positive for both, ASFV DNA and antibodies, suggesting ongoing virus infection, whereas the proportion of boar positive only for antibodies was 5.75%, indicating survival of acute infection. Statistical analysis revealed an increase in ASFV DNA detection from 2023 to 2024 (p = 0.043), with a higher prevalence in carcasses than in hunted animals (p = 0.001), highlighting the need for passive monitoring. While gender showed no statistical significance, a higher infection rate was observed in older animals (p = 0.001). The identified course of infection involved spillover events between domestic pigs and wild boar, with a significant anthropogenic influence. Full article

Background: Coxiella burnetii is a common zoonotic pathogen that can lead not only to acute or chronic Q fever but also to post-infectious syndromes, where autonomic nervous system (ANS) dysfunction has been suggested as a contributing mechanism. This study aimed to assess autonomic function in patients presenting with polymorphic symptoms, dysautonomia, or ME/CFS who had serological evidence of acute infection with Coxiella burnetii. Methods: A total of 156 participants were evaluated, including 100 seropositive patients and 56 matched controls. All subjects underwent standardized cardiovascular reflex tests (CART), beat-to-beat analysis of heart rate and blood pressure with baroreflex indices, 24 h Holter ECG with HRV assessment, and, in the Coxiella group, head-up tilt testing (HUTT). Results: A significantly higher prevalence of autonomic dysfunction was observed in the Coxiella group, predominantly affecting parasympathetic regulation, with abnormal CART scores, reduced LF power and baroreflex effectiveness, and a high rate of positive HUTT findings characterized by extreme blood pressure variability. Although long-term HRV measures did not differ significantly between groups, short-term indices consistently indicated ANS impairment. Conclusions: These findings suggest that Coxiella burnetii infection may trigger persistent autonomic dysfunction, potentially contributing to the development of ME/CFS and syncope in affected individuals. Further longitudinal studies are needed to clarify pathophysiological mechanisms and clinical implications. Full article

Background/Objectives: The detection of anti-Coxiella antibodies using serological methods is essential for identifying exposed ruminants and preventing this important zoonotic disease in livestock. In recent years, numerous attempts have been made to increase diagnostic performance as well as simplify the production of serological assays. Commercially available tests often use whole-cell antigens, which can decrease specificity and require high-level biosafety facilities for manufacturing. The aim of this work was to produce three Coxiella burnetii (C. burnetii) antigens in recombinant form and assess them for the detection of anti-Coxiella antibodies in ruminants. Methods: Three recombinant C. burnetii antigens (Com-1, MceB, AdaA) were selected among immunodominant antigens and produced in a heterologous system (Escherichia coli). Following purification, the proteins were utilized to coat ELISA plates and evaluated for seroreactivity against sera from both negative and positive cattle. Results: Com-1 demonstrated the greatest agreement with the commercial test, albeit moderate. MceB exhibited nonspecific reactivity against a large number of sera, while the AdaA showed reactivity against only a few positive sera. Conclusions: Our findings are consistent with previous research, indicating that utilizing a single antigen to identify exposed animals is unfeasible with current knowledge, most likely due to the complex immunological response following C. burnetii infection in cattle. Consequently, it is critical to continue testing and identifying immunoreactive antigens in order to further investigate them and, potentially, select the most appropriate. Full article

Background: Individual differences in immune responses to African swine fever virus (ASFV), whether induced by vaccination or natural infection, may be linked to genetic variation in the genes involved in antigen presentation. Methods: A total of nine pigs from the 112-population were selected for RNA-seq analysis. To pinpoint key transcription factors (TFs) regulating gene expression in the lymph nodes, weighted Kendall’s Tau rank correlation analysis was performed to link the TF binding potential with the extent of differential expression of target genes. Results: CD8⁺ T cells expressing a specific epitope of the ASFV p72 protein (ACD8⁺) accounted for 41% of the total CD8⁺ T cells in peripheral blood. A total of 2062 transcripts were identified as differentially expressed across the nine pigs (q-value < 1 × 10⁻⁸). Differential expression levels of the target genes for MECP2, ETS1, ZBTB33, ELK4, and E2F4 were significantly correlated with their TF binding potential (p < 0.05). Six SNPs were identified in the promoter region of ELK4. Analysis of the 112-pig population revealed that SNPs at S.-404A>G and S.-668C>T loci were significantly associated with ACD8⁺ levels (q-value < 0.01). Individuals with the AA genotype at S.-404A>G had significantly higher ACD8⁺ counts compared to those with AG and GG genotypes (q-value < 0.05). At the S.-668C>T locus, ACD8⁺ levels were highest in the CC genotype, followed by CT and TT genotypes, with CC showing notably higher ACD8⁺ counts (q-value < 0.05). Notably, the S.-404A>G site overlaps with potential binding sites for TFs FOXA2, GATAs, and TRPS1, while the S.-668C>T site lies within the binding regions for NR1H3, RARA, VDR, and NR1I3. Conclusion: These mutations may disrupt TFs binding to the ELK4 promoter, potentially reducing ELK4 expression and impairing antigen processing and presentation. Full article

Goats are important reservoirs of Coxiella burnetii, the causative agent of Q fever in humans. Monitoring infection in goats is therefore essential for effective human disease prevention. However, epidemiological data on C. burnetii infection in Thai goats remain limited, particularly in the central and western regions, where goat farming has expanded. This study aimed to determine the seroprevalence of C. burnetii infection in goats raised in these two regions. A total of 947 serum samples from 101 herds were collected and tested using a commercial enzyme-linked immunosorbent assay (ELISA). Individual- and herd-level seroprevalence were calculated. Pearson’s chi-square test was used to compare infection proportions between regions (central vs. western) and herd types (meat vs. dairy), with p < 0.05 considered statistically significant. Overall, individual- and herd-level seroprevalence were 22.39% and 58.42%, respectively. Individual seroprevalence in the central and western regions was 25.16% and 19.71%, while herd-level seroprevalence was 71.15% and 44.90%, respectively. The proportions of infection at both levels were significantly higher in the central region (p = 0.044 and p = 0.007). In contrast, no significant differences were observed between meat and dairy herds: individual-level seroprevalence was 22.81% and 25.25% (p = 0.586), and herd-level seroprevalence was 56.98% and 60.00% (p = 0.855), respectively. These findings highlighted the urgent need to strengthen control measures for C. burnetii infection in goats in central and western Thailand to protect human, animal, and environmental health. Full article