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This study aimed to understand the epidemiological characteristics of Salmonella in Tibetan pigs. We isolated, identified, and examined via antimicrobial susceptibility testing on Salmonella from Tibetan pigs breeder farms and slaughterhouses in Tibet, China. A genetic evolutionary tree was constructed on the basis of whole genome sequencing (WGS). A total of 81 Salmonella isolates were isolated from 987 samples. The main serovars were Salmonella Typhimurium and Salmonella London in Tibetan pigs. The isolated Salmonella Typhimurium isolates subjected to antimicrobial susceptibility testing showed varying degrees of resistance to ß-lactams, aminoglycosides, fluoroquinolones, sulfonamides, tetracyclines, and amphenicols. WGS analysis was performed on 20 Salmonella Typhimurium isolates in Tibet (n = 10), Jiangsu (n = 10), and 205 genome sequences downloaded from the Enterobase database to reveal their epidemiological and genetic characteristics. They were divided into two clusters based on core genome single-nucleotide polymorphisms: Cluster A with 112 isolates from Tibet and other regions in China and Cluster B with 113 isolates from Jiangsu and other regions. The isolates in Cluster A were further divided into two subclusters: A-1 with 40 isolates including Tibet and A-2 with 72 isolates from other regions. Virulence factors analysis revealed that all isolates from Tibet carried adeG, but this observation was not as common in Salmonella isolates from Jiangsu and other regions of China. Antibiotic resistance genes (ARGs) analysis showed that all isolates from Tibet carried blaTEM-55 and rmtB, which were absent in Salmonella isolates from Jiangsu and other regions of China. Genetic characteristic analysis and biofilm determination indicated that the biofilm formation capabilities of the isolates from Tibet were stronger than those of the isolates from Jiangsu and other regions of China. Our research revealed the epidemic patterns and genomic characteristics of Salmonella in Tibetan pigs and provided theoretical guidance for the prevention and control of local salmonellosis.
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Salmonellosis in poultry is detrimental to the advancement of the breeding industry and poses hazards to human health. Approximately 2,600 Salmonella varieties exist, among which S. Enteritidis, S. Pullorum, S. Typhimurium, and S. Infantis are prevalent serotypes in the poultry industry in recent years. They can also infect humans by contaminating poultry eggs and meat. Therefore, identifying these serotypes is crucial for successful preventive and control interventions. The White-Kauffmann-Le Minor scheme is time-consuming and requires expensive reagents. Whole-genome sequencing (WGS) and other molecular biology techniques require skilled technical staff. In comparison, the polymerase chain reaction (PCR) is more accurate, rapid, and inexpensive, thus proving suitable for widespread application in the poultry industry. Here, we selected 4 specific primers: lygD, mdh, ipaJ, and SIN_02055, which correspond to detecting S. Enteritidis, S. Typhimurium, S. Pullorum, and S. Infantis, respectively. They were integrated into a 1-step multiplex PCR method. We optimized the PCR method by utilizing specificity test results to determine the optimal annealing temperature (57°C). The PCR method exhibited excellent sensitivity for genomic DNA and bacterial cultures. We used the developed method to determine 157 clinical Salmonella isolates from various stages of the poultry production chain. The results aligned with serotype data generated via WGS analysis, demonstrating the method's excellent accuracy. In conclusion, this study developed a 1-step multiplex PCR method that simultaneously identifies S. Enteritidis, S. Typhimurium, S. Pullorum, and S. Infantis, allowing routine mass detection in the grass-root poultry industry.
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The interplay among frustrated lattice geometry, non-trivial band topology and correlation yields rich quantum states of matter in kagome systems1,2. A series of recent members in this family, AV3Sb5 (A = K, Rb or Cs), exhibit a cascade of symmetry-breaking transitions3, involving the 3Q chiral charge ordering4-8, electronic nematicity9,10, roton pair density wave11 and superconductivity12. The nature of the superconducting order is yet to be resolved. Here we report an indication of dynamic superconducting domains with boundary supercurrents in intrinsic CsV3Sb5 flakes. The magnetic field-free superconducting diode effect is observed with polarity modulated by thermal histories, suggesting that there are dynamic superconducting order domains in a spontaneous time-reversal symmetry-breaking background. Strikingly, the critical current exhibits double-slit superconductivity interference patterns when subjected to an external magnetic field. The characteristics of the patterns are modulated by thermal cycling. These phenomena are proposed as a consequence of periodically modulated supercurrents flowing along certain domain boundaries constrained by fluxoid quantization. Our results imply a time-reversal symmetry-breaking superconducting order, opening a potential for exploring exotic physics, for example, Majorana zero modes, in this intriguing topological kagome system.
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African swine fever virus (ASFV) is a large double stranded DNA arbovirus that is highly contagious and seriously endangers domestic and wild pigs. In the past decade, African swine fever (ASF) has spread in many countries in the Caucasus, Russian Federation, Eastern Europe and Asia, causing significant losses to the pig industry. At present, there is a lack of effective vaccine and treatment for ASF. Therefore, the rapid and accurate detection is crucial for ASF prevention and control. In this study, we have developed a portable lateral flow strip (LFS) detection mediated by recombinase polymerase amplification (RPA) and CRISPR/LwCas13a, which is performed at 37 â and visualized by eyes without the need for complex instruments. This RPA-LwCas13a-LFS is based on the ASFV structural protein p17 gene (D117L), with a detection sensitivity up to 2 gene copies. This method is highly specific and has no cross reactivity to 7 other pig viruses. In the detection of two batches of 100 clinical samples, the p17 (D117L) RPA-LwCas13a-LFS had 100% coincidence with conventional quantitative PCR (qPCR). These findings demonstrate the potential of this simple, rapid, sensitive, and specific ASFV detection method for on-site ASFV detection.
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Virus de la Fiebre Porcina Africana , Fiebre Porcina Africana , Sistemas CRISPR-Cas , Animales , Fiebre Porcina Africana/virología , Fiebre Porcina Africana/diagnóstico , Virus de la Fiebre Porcina Africana/genética , Virus de la Fiebre Porcina Africana/aislamiento & purificación , Técnicas de Amplificación de Ácido Nucleico/métodos , Técnicas de Amplificación de Ácido Nucleico/veterinaria , Sensibilidad y Especificidad , Porcinos , Proteínas Estructurales Virales/análisis , Proteínas Estructurales Virales/genéticaRESUMEN
Exotic quantum phases and phase transition in the strongly interacting Dirac systems have attracted tremendous interests. On the other hand, non-Hermitian physics, usually associated with dissipation arising from the coupling to environment, emerges as a frontier of modern physics in recent years. In this Letter, we investigate the interplay between non-Hermitian physics and strong correlation in Dirac-fermion systems. We generalize the projector quantum Monte-Carlo (PQMC) algorithm to the non-Hermitian interacting fermionic systems. Employing PQMC simulation, we decipher the ground-state phase diagram of the honeycomb Hubbard model with spin resolved non-Hermitian asymmetric hopping processes. The antiferromagnetic (AFM) ordering induced by Hubbard interaction is enhanced by the non-Hermitian asymmetric hopping. Combining PQMC simulation and renormalization group analysis, we reveal that the quantum phase transition between Dirac semi-metal and AFM phases belongs to Hermitian chiral XY universality class, implying that a Hermitian Gross-Neveu transition is emergent at the quantum critical point although the model is non-Hermitian.
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The newly discovered high-temperature superconductivity in La_{3}Ni_{2}O_{7} under pressure has attracted a great deal of attention. The essential ingredient characterizing the electronic properties is the bilayer NiO_{2} planes coupled by the interlayer bonding of 3d_{z^{2}} orbitals through the intermediate oxygen atoms. In the strong coupling limit, the low-energy physics is described by an intralayer antiferromagnetic spin-exchange interaction J_{â¥} between 3d_{x^{2}-y^{2}} orbitals and an interlayer one J_{â¥} between 3d_{z^{2}} orbitals. Taking into account Hund's rule on each site and integrating out the 3d_{z^{2}} spin degree of freedom, the system reduces to a single-orbital bilayer t-J model based on the 3d_{x^{2}-y^{2}} orbital. By employing the slave-boson approach, the self-consistent equations for the bonding and pairing order parameters are solved. Near the physically relevant 1/4-filling regime (doping δ=0.3â¼0.5), the interlayer coupling J_{â¥} tunes the conventional single-layer d-wave superconducting state to the s-wave one. A strong J_{â¥} could enhance the interlayer superconducting order, leading to a dramatically increased T_{c}. Interestingly, there could exist a finite regime in which an s+id state emerges.
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The inflammasome is a pivotal component of the innate immune system, acting as a multiprotein complex that plays an essential role in detecting and responding to microbial infections. Salmonella Enteritidis have evolved multiple mechanisms to regulate inflammasome activation and evade host immune system clearance. Through screening S. Enteritidis C50336ΔfliC transposon mutant library, we found that the insertion mutant of dinJ increased inflammasome activation. In this study, we demonstrated the genetic connection between the antitoxin DinJ and the toxin YafQ in S. Enteritidis, confirming their co-transcription. The deletion mutant ΔfliCΔdinJ increased cell death and IL-1ß secretion in J774A.1 cells. Western blotting analysis further showed elevated cleaved Caspase-1 product (p10 subunits) and IL-1ß secretion in cells infected with ΔfliCΔdinJ compared to cells infected with ΔfliC. DinJ was found to inhibit canonical inflammasome activation using primary bone marrow-derived macrophages (BMDMs) from Casp-/- C57BL/6 mice. Furthermore, DinJ specifically inhibited NLRP3 inflammasome activation, as demonstrated in BMDMs from Nlrp3-/- and Nlrc4-/- mice. Fluorescence resonance energy transfer (FRET) experiments confirmed the translocation of DinJ into host cells during infection. Finally, we revealed that DinJ could inhibit the secretion of IL-1ß and IL-18 in vivo, contributing to S. Enteritidis evading host immune clearance. In summary, our findings provide insights into the role of DinJ in modulating the inflammasome response during S. Enteritidis infection, highlighting its impact on inhibiting inflammasome activation and immune evasion.
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Antitoxinas , Inflamasomas , Animales , Ratones , Inflamasomas/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Salmonella enteritidis , Ratones Endogámicos C57BL , Macrófagos , Caspasa 1/genética , Interleucina-1beta/genética , Interleucina-1beta/metabolismoRESUMEN
Salmonella, which is widely distributed in nature, is an important zoonotic pathogen affecting humans, livestock, and other animals. Salmonella infection not only hinders the development of livestock and poultry-related industries but also poses a great threat to human health. In this study, we collected 1,537 samples including weak chicks, dead embryos, fecal samples and environmental samples from 2020 to 2023 (for a period of 1 to 2 months per year) to keep a long-term monitor the prevalence of Salmonella in an intensive laying hen farm, 105 Salmonella strains were isolated with an isolation rate of 6.83% (105/1,537). It revealed a significant decrease in prevalence rates of Salmonella over time (P < 0.001). Before 2020, the predominant serotype was S. Enteritidis. S. Kentucky was first detected in November 2020 and its proportion was gradually found to exceed that of S. Enteritidis since then. S. Kentucky isolates were distributed in various links of the four regions in the poultry farm. A total of 55 S. Kentucky strains, were assigned to ST198 based on whole genome sequencing. Among them, 54 strains were resistant to 12 to 16 antibiotics, indicating that they were extensively drug-resistant (XDR). Seventeen antimicrobial resistance genes were detected in 55 S. Kentucky isolates. For most of these isolates, antibiotic resistance phenotypes were concordant with their genotypes. All S. Kentucky strains isolated from this farm in 2020 to 2023 showed a high similarity based on their core-genome SNP-based phylogeny. The traceability analysis revealed that S. Kentucky was introduced to the farm through newly purchased flocks. The long-term existence of XDR S. Kentucky ST198 poses a substantial risk because of the multiage management and circulation of workers in this poultry farm. Thus, this study is the first to report extensively drug-resistant S. Kentucky ST198 detected in this intensive poultry farm in China.
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Pollos , Farmacorresistencia Bacteriana Múltiple , Enfermedades de las Aves de Corral , Salmonelosis Animal , Salmonella enterica , Secuenciación Completa del Genoma , Animales , Enfermedades de las Aves de Corral/microbiología , Enfermedades de las Aves de Corral/epidemiología , Salmonelosis Animal/epidemiología , Salmonelosis Animal/microbiología , China/epidemiología , Prevalencia , Salmonella enterica/genética , Salmonella enterica/efectos de los fármacos , Salmonella enterica/aislamiento & purificación , Secuenciación Completa del Genoma/veterinaria , Femenino , Serogrupo , Antibacterianos/farmacologíaRESUMEN
Introduction: Apicomplexan AP2 family of proteins (ApiAP2) are transcription factors (TFs) that regulate parasite growth and development, but little is known about the ApiAP2 TFs in Eimeria spp. ENH_00027130 sequence is predicted to encode a Eimeria necatrix ApiAP2 protein (EnApiAP2). Methods: The cDNAs encoding full-length and truncated EnApiAP2 protein were cloned and sequenced, respectively. Then, the two cDNAs were cloned into the pET28a(+) expression vector and expressed expressed in Escherichia coli BL21. The mouse polyclonal antibody (pAb) and monoclonal antibody (mAb) against recombinant EnApiAP2 (rEnApiAP2) and EnApiAP2tr (rEnApiAP2tr) were prepared and used to localize the native EnApiAP2 protein in E. necatrix, respectively. Finally, the recombinant pEGFP-C1-ΔNLS-EnApiAP2s (knockout of a nuclear localization sequence, NLS) and pEGFP-C1-EnApiAP2 plasmid were constructed and transfected into DF-1 cells, respectively, to further observe subcellular localization of EnApiAP2 protein. Results: The EnApiAP2 gene had a size of 5019 bp and encoded 1672 amino acids, containing a conserved AP2 domain with a secondary structure consisting of an α-helix and three antiparallel ß-strands. The rEnApiAP2 and rEnApiAP2tr were predominantly expressed in the form of inclusion bodies, and could be recognized by the 6×His tag mAb and the serum of convalescent chickens after infection with E. necatrix, respectively. The native EnApiAP2 protein was detected in sporozoites (SZ) and second generation merozoites (MZ-2) extracts, with a size of approximately 210 kDa. A quantitative real-time PCR (qPCR) analysis showed that the transcription level of EnApiAP2 was significantly higher in SZ than in MZ-2, third generation merozoites (MZ-3) and gametocytes (P<0.01). EnApiAP2 protein was localized in the nuclei of SZ, MZ-2 and MZ-3 of E. necatrix. The protein of EnApiAP2 was localized in the nucleus of the DF-1 cells, whereas the ΔNLS-EnApiAP2 was expressed in the cytoplasm, which further confirmed that EnApiAP2 is nucleoprotein. Discussion: EnApiAP2 protein encoded by ENH_00027130 sequence was localized in the nucleus of E. necatrix parasites, and relied on the NLS for migration to DF-1 cell nucleus. The function of EnApiAP2 need further study.
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Eimeria , Enfermedades de las Aves de Corral , Animales , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Pollos/genética , ADN Complementario/genética , Eimeria/genética , Eimeria/metabolismo , Proteínas de Microfilamentos/metabolismo , Proteínas Nucleares/metabolismo , Enfermedades de las Aves de Corral/parasitología , Esporozoítos/metabolismoRESUMEN
IMPORTANCE: The development of safe and effective vaccines is needed to control the transmission of coronavirus disease 2019 (COVID-19). Synthetic DNA vaccines represent a promising platform in response to such outbreaks. Here, DNA vaccine candidates were developed using an optimized antibiotic-resistance gene-free asd-pVAX1 vector. An optimized flagellin (FliC) adjuvant was designed by fusion expression to increase the immunogenicity of the S1 antigen. S1 and S1-FliCΔD2D3 proteins were strongly expressed in mammalian cells. The FliCΔD2D3-adjuvanted DNA vaccine induced Th1/Th2-mixed immune responses and high titers of neutralizing antibodies. This study provides crucial information regarding the selection of a safer DNA vector and adjuvant for vaccine development. Our FliCΔD2D3-adjuvanted S1 DNA vaccine is more potent at inducing both humoral and cellular immune responses than S1 alone. This finding provides a new idea for the development of novel DNA vaccines against COVID-19 and could be further applied for the development of other vaccines.
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COVID-19 , Vacunas de ADN , Humanos , Animales , Ratones , Salmonella typhimurium/metabolismo , Vacunas de ADN/genética , Vacunas de ADN/metabolismo , SARS-CoV-2 , Flagelina/genética , Flagelina/metabolismo , Vacunas contra la COVID-19 , COVID-19/prevención & control , Adyuvantes Inmunológicos/metabolismo , Adyuvantes Inmunológicos/farmacología , Anticuerpos Neutralizantes/metabolismo , Anticuerpos Antivirales/metabolismo , Inmunogenicidad Vacunal , MamíferosRESUMEN
Eimeria necatrix is a high pathogenic pathogen, which seriously endangers the poultry industry. The surface antigens (SAGs) of Apicomplexa are a kind of membrane protein anchored on the surface of the parasites through its carboxyl terminal glycosylphosphatidylinositol (GPI) structure. However, little is known about GPI-linked surface proteins in E. necatrix. In the present work, the E. necatrix sag gene (Ensag-CAP) was amplified and cloned for expression of the recombinant protein (rEnSAG-CAP). The full length Ensag-CAP gene was 813 bp, coding 270 amino acids with a predicated molecular weight of 28.86 kDa and contained a CAP domain with four sequence motifs CAP1, CAP2, CAP3 and CAP4. The rEnSAG-CAP was about 32 kDa and mainly expressed in a soluble form. Western blot analysis indicated that the rEnSAG-CAP could be recognized by anti-rEnSAG-CAP monoclonal antibody (anti-rEnSAG-CAP McAb) and the convalescent serum of chicken infected with E. necatrix. Native protein of EnSAG-CAP was detected in second-generation merozoites (MZ-2) using anti-rEnSAG-CAP polyclonal antibody (anti-rEnSAG-CAP pAb). The findings from the indirect immunofluorescence assay and enzyme digestion utilizing Bacillus cereus phosphoinositide-specific phospholipase C (PI-PLC) revealed that EnSAG-CAP predominantly localized at the surfaces of SZ and MZ-2 via a GPI anchor. It was observed that EnSAG-CAP can be cleaved from MZ-2 by PI-PLC. Real-time quantitative PCR (qPCR) analysis showed that transcript levels of Ensag-CAP in MZ-2 was significantly higher than that in SZ (P < 0.05). The anti-rEnSAG-CAP McAb in vitro could significantly inhibit the sporozoite invasion into MDBK cells (P < 0.01), which suggests that the protein might participate in sporozoite invasion into MDBK cells. rEnSAG-CAP afforded an immune protection against E. necatrix. The ACI value was 164.99 in the chickens immunized with 200 µg rEnSAG-CAP. Chickens immunized with rEnSAG-CAP had a significantly higher antigen-specific serum IgY response (P < 0.0001). The data indicates that EnSAG-CAP could serve as a potential candidate antigen for the development of a recombinant coccidiosis vaccine.
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Coccidiosis , Eimeria , Enfermedades de las Aves de Corral , Animales , Eimeria/fisiología , Pollos/parasitología , Coccidiosis/prevención & control , Coccidiosis/veterinaria , Proteínas Recombinantes/genética , Esporozoítos , Vacunas Sintéticas , Enfermedades de las Aves de Corral/parasitologíaRESUMEN
BACKGROUND: COVID-19 caused mild to severe infections in humans. The long-term epidemic environment harms people's mental health. To explore the impact of the epidemic on people's mental and psychological conditions, we surveyed in Wenzhou. METHODS: We collected the data of people who visited the First Affiliated Hospital of Wenzhou Medical University for five types of mental and psychological diseases from January 2018 to December 2021. Then, taking December 2019 as the cut-off point, the 48-month data were divided into the pre-epidemic group and the dur-epidemic group. Based on the above data, statistical analysis was done. RESULTS: From 2018 to 2021, the number of initial diagnoses, the number of disease visits, and drug consumption for these five types of mental and psychological diseases were all on the rise. Compared with the number of disease visits for all disorders in both psychiatry and neurology departments, it was found that the growth rate of these five diseases was higher than the growth rate of all disorders. We found that the number of disease visits, drug consumption, and scale scores after the COVID-19 outbreak were significantly different from those before the outbreak (P < 0.05). And the number of disease visits positively correlated with drug consumption (P < 0.0001, r = 0.9503), which verified the stability of the data. CONCLUSION: The epidemic environment has had a long-term and negative impact on people's mental and psychological conditions. Therefore, whether or not the epidemic is receding, we still need to be concerned about the impact of COVID-19 on mental and psychological health.
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COVID-19 , Trastornos Mentales , Psiquiatría , Humanos , Pandemias , COVID-19/epidemiología , Trastornos Mentales/epidemiología , Salud MentalAsunto(s)
Infecciones por Klebsiella , Klebsiella pneumoniae , Porcinos , Animales , Klebsiella pneumoniae/genética , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , beta-Lactamasas , China , Infecciones por Klebsiella/veterinaria , Infecciones por Klebsiella/tratamiento farmacológico , Pruebas de Sensibilidad MicrobianaRESUMEN
Most cases of chicken salmonellosis are caused by Salmonella enterica serovar Gallinarum biovars Gallinarum and Pullorum, which lead to a significant morbidity and fatality rate. Although the conventional Kaufmann-White scheme is the reliable method for the serotyping of Salmonella, it does not distinguish between closely related biotypes like S. Pullorum and S. Gallinarum. Herein, we conducted a single one-step multiplex PCR assay that can identify and distinguish between S. Pullorum and S. Gallinarum in an accurate manner. This PCR method was based on three genes, including torT for S. Pullorum identification, I137_14430 for S. Gallinarum identification, and stn as the genus-level reference gene for Salmonella. By comparing S. Pullorum to S. Gallinarum and other serovars of Salmonella, in silico study revealed that only the former has a deletion of 126 bp-region in the carboxyl terminus of torT. The I137_14430 gene does not exist in S. Gallinarum. However, it is present in all other Salmonella serotypes. The multiplex PCR approach utilizes unique sets of primers that are intended to specifically target these three different genes. The established PCR method was capable of distinguishing between the biovars Pullorum and Gallinarum from the 29 distinct Salmonella serotypes as well as the 50 distinct pathogens that are not Salmonella, showing excellent specificity and exclusivity. The minimal amount of bacterial cells required for PCR detection was 100 CFU, while the lowest level of genomic DNA required was 27.5 pg/µL for both S. Pullorum and S. Gallinarum. After being implemented on the clinical Salmonella isolates collected from a poultry farm, the PCR test was capable of distinguishing the two biovars Pullorum and Gallinarum from the other Salmonella strains. The findings of the PCR assay were in line with those of the traditional serotyping and biochemical identification methods. This new multiplex PCR could be used as a novel tool to reinforce the clinical diagnosis and differentiation of S. Pullorum and S. Gallinarum, particularly in high-throughput screening situations, providing the opportunity for early screening of infections and, as a result, more effective management of the illness among flocks.
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Poly (triazine imide) (PTI/Li+ Cl- ), one of the crystalline versions of polymeric carbon nitrides, holds great promise for photocatalytic overall water splitting. In principle, the photocatalytic activity of PTI/Li+ Cl- is closely related to the morphology, which could be reasonably tailored by the modulation of the polycondensation process. Herein, we demonstrate that the hexagonal prisms of PTI/Li+ Cl- could be converted to hexagonal nanosheets by adjusting the binary eutectic salts from LiCl/KCl or NaCl/LiCl to ternary LiCl/KCl/NaCl. Results reveal that the extension of in-plane conjugation is preferred, when the polymerisation was performed in the presence of ternary eutectic salts. The hexagonal nanosheets bears longer lifetimes of charge carriers than that of hexagonal prisms due to lower intensity of structure defects and shorter hopping distance of charge carriers along the stacking direction of triazine nanosheets. The optimized hexagonal nanosheets exhibits a record apparent quantum yield value of 25 % (λ=365â nm) for solar hydrogen production by one-step excitation overall water splitting.
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Salmonella Enteritidis is an important intracellular pathogen, which can cause gastroenteritis in humans and animals and threaten life and health. S. Enteritidis proliferates in host macrophages to establish systemic infection. In this study, we evaluated the effects of Salmonella pathogenicity island-1 (SPI-1) and SPI-2 to S. Enteritidis virulence in vitro and in vivo, as well as the host inflammatory pathways affected by SPI-1 and SPI-2. Our results show that S. Enteritidis SPI-1 and SPI-2 contributed to bacterial invasion and proliferation in RAW264.7 macrophages, and induced cytotoxicity and cellular apoptosis of these cells. S. Enteritidis infection induced multiple inflammatory responses, including mitogen-activated protein kinase (ERK-mediated) and Janus kinase-signal transducer and activator of transcript (STAT) (STAT2-mediated) pathways. Both SPI-1 and SPI-2 were necessary to induce robust inflammatory responses and ERK/STAT2 phosphorylation in macrophages. In a mouse infection model, both SPIs, especially SPI-2, resulted in significant production of inflammatory cytokines and various interferon-stimulated genes in the liver and spleen. Activation of the ERK- and STAT2-mediated cytokine storm was largely affected by SPI-2. S. Enteritidis ΔSPI-1-infected mice displayed moderate histopathological damage and drastically reduced bacterial loads in tissues, whereas only slight damage and no bacteria were observed in ΔSPI-2- and ΔSPI-1/SPI-2-infected mice. A survival assay showed that ΔSPI-1 mutant mice maintained a medium level of virulence, while SPI-2 plays a decisive role in bacterial virulence. Collectively, our findings indicate that both SPIs, especially SPI-2, profoundly contributed to S. Enteritidis intracellular localization and virulence by activating multiple inflammatory pathways.
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Islas Genómicas , Salmonella enteritidis , Animales , Humanos , Ratones , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Proliferación Celular , Modelos Animales de Enfermedad , Salmonella enteritidis/genética , VirulenciaRESUMEN
Listeria monocytogenes (Lm) is a deadly foodborne pathogen that comprises 14 serotypes, among which, serotype 4b Lm is the primary cause of listeriosis outbreaks in humans and animals. Here, we evaluated the safety, immunogenicity, and protective efficacy of a serotype 4b vaccine candidate Lm NTSNΔactA/plcB/orfX in sheep. The infection dynamics, clinical features, and pathological observation verified that the triple genes deletion strain has adequate safety for sheep. Moreover, NTSNΔactA/plcB/orfX significantly stimulated humoral immune response and provided 78% immune protection to sheep against lethal wild-type strain challenge. Notably, the attenuated vaccine candidate could differentiate infected and vaccinated animals (DIVA) via serology determination of the antibody against listeriolysin O (LLO, encoded by hly) and phosphatidylinositol-specific phospholipase C (PI-PLC, encoded by plcB). These data suggest that the serotype 4b vaccine candidate has high efficacy, safety, and DIVA characteristics, and may be used to prevent Lm infection in sheep. Our study provides a theoretical basis for its future application in livestock and poultry breeding.
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Listeria monocytogenes , Listeriosis , Humanos , Animales , Ovinos , Listeria monocytogenes/genética , Listeriosis/prevención & control , Listeriosis/veterinaria , Serogrupo , Vacunas Atenuadas , Anticuerpos , Proteínas Hemolisinas/genéticaRESUMEN
Inflammasome activation is an essential innate immune defense mechanism against Salmonella infections. Salmonella has developed multiple strategies to avoid or delay inflammasome activation, which may be required for long-term bacterial persistence. However, the mechanisms by which Salmonella evades host immune defenses are still not well understood. In this study, Salmonella Enteritidis (SE) random insertion transposon library was screened to identify the key factors that affect the inflammasome activation. The type I secretion system (T1SS) protein SiiD was demonstrated to repress the NLRP3 inflammasome activation during SE infection and was the first to reveal the antagonistic role of T1SS in the inflammasome pathway. SiiD was translocated into host cells and localized in the membrane fraction in a T1SS-dependent and partially T3SS-1-dependent way during SE infection. Subsequently, SiiD was demonstrated to significantly suppress the generation of mitochondrial reactive oxygen species (mtROS), thus repressing ASC oligomerization to form pyroptosomes, and impairing the NLRP3 dependent Caspase-1 activation and IL-1ß secretion. Importantly, SiiD-deficient SE induced stronger gut inflammation in mice and displayed NLRP3-dependent attenuation of the virulence. SiiD-mediated inhibition of NLRP3 inflammasome activation significantly contributed to SE colonization in the infected mice. This study links bacterial T1SS regulation of mtROS-ASC signaling to NLRP3 inflammasome activation and reveals the essential role of T1SS in evading host immune responses.
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Inflamasomas , Proteína con Dominio Pirina 3 de la Familia NLR , Animales , Ratones , Proteína con Dominio Pirina 3 de la Familia NLR/genética , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Salmonella enteritidis , Sistemas de Secreción Tipo I , Transducción de Señal , Caspasa 1/metabolismo , Interleucina-1beta/metabolismoRESUMEN
The flagellin (FliC) of Salmonella typhimurium is a potential vaccine adjuvant as it can activate innate immunity and promote acquired immune responses. Macrophages are an important component of the innate immune system. The mechanism of flagellin's adjuvant activity has been shown to be related to its ability to activate macrophages. However, few studies have comprehensively investigated the effects of Salmonella flagellin in macrophages using transcriptome sequencing. In this study, RNA-Seq was used to analyze the expression patterns of RAW264.7 macrophages induced by FliC to identify novel transcriptomic signatures in macrophages. A total of 2204 differentially expressed genes were found in the FliC-treated group compared with the control. Gene ontology and KEGG pathway analyses identified the top significantly regulated functional classification and canonical pathways, which were mainly related to immune responses and regulation. Inflammatory cytokines (IL-6, IL-1ß, TNF-α, etc.) and chemokines (CXCL2, CXCL10, CCL2, etc.) were highly expressed in RAW264.7 cells following stimulation. Notably, flagellin significantly increased the expression of interferon (IFN)-ß. In addition, previously unidentified IFN regulatory factors (IRFs) and IFN-stimulated genes (ISGs) were also significantly upregulated. The results of RNA-Seq were verified, and furthermore, we demonstrated that flagellin increased the expression of IFN-ß and IFN-related genes (IRFs and ISGs) in bone marrow-derived dendritic cells and macrophages. These results suggested that Salmonella flagellin can activate IFN-ß-related immune responses in macrophages, which provides new insight into the immune mechanisms of flagellin adjuvant.