Cryo-EM structures of Smc5/6 in multiple states reveal its assembly and functional mechanisms.

Smc5/6 is a member of the eukaryotic structural maintenance of chromosomes (SMC) family of complexes with important roles in genome maintenance and viral restriction. However, limited structural understanding of Smc5/6 hinders the elucidation of its diverse functions. Here, we report cryo-EM structures of the budding yeast Smc5/6 complex in eight-subunit, six-subunit and five-subunit states. Structural maps throughout the entire length of these complexes reveal modularity and key elements in complex assembly. We show that the non-SMC element (Nse)2 subunit supports the overall shape of the complex and uses a wedge motif to aid the stability and function of the complex. The Nse6 subunit features a flexible hook region for attachment to the Smc5 and Smc6 arm regions, contributing to the DNA repair roles of the complex. Our results also suggest a structural basis for the opposite effects of the Nse1-3-4 and Nse5-6 subcomplexes in regulating Smc5/6 ATPase activity. Collectively, our integrated structural and functional data provide a framework for understanding Smc5/6 assembly and function.

Enhancing Tumor Immunotherapy by Multivalent Anti-PD-L1 Nanobody Assembled via Ferritin Nanocage.

Increasing immunotherapy response rate and durability can lead to significant improvements in cancer care. To address this challenge, a novel multivalent immune checkpoint therapeutic platform is constructed through site-specific ligation of anti-PD-L1 nanobody (Nb) on ferritin (Ftn) nanocage. Nb-Ftn blocks PD-1/PD-L1 interaction and downregulates PD-L1 levels via endocytosis-induced degradation. In addition, the cage structure of Ftn allows encapsulation of indocyanine green (ICG), an FDA-approved dye. Photothermal treatment with Nb-Ftn@ICG induces immunogenic death of tumor cells, which improves systemic immune response via maturation of dendritic cells and enhanced infiltration of T cells. Moreover, Nb-Ftn encapsulation significantly enhances cellular uptake, tumor accumulation and retention of ICG. In vivo assays showed that this nanoplatform ablates the primary tumor, suppresses abscopal tumors and inhibits tumor metastasis, leading to a prolonged survival rate. This work presents a novel strategy for improving cancer immunotherapy using multivalent nanobody-ferritin conjugates as immunological targeting and enhancing carriers.

A non-metal single atom nanozyme for cutting off the energy and reducing power of tumors.

Enzymes are considered safe and effective therapeutic tools for various diseases. With the increasing integration of biomedicine and nanotechnology, artificial nanozymes offer advanced controllability and functionality in medical design. However, several notable gaps, such as catalytic diversity, specificity and biosafety, still exist between nanozymes and their native counterparts. Here we report a non-metal single-selenium (Se)-atom nanozyme (SeSAE), which exhibits potent nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-mimetic activity. This novel single atom nanozyme provides a safe alternative to conventional metal-based catalysts and effectively cuts off the cellular energy and reduction equivalents through its distinctive catalytic function in tumors. In this study, we have demonstrated the substantial efficacy of SeSAE as an antitumor nanomedicine across diverse mouse models without discernible systemic adverse effects. The mechanism of the NADPH oxidase-like activity of the non-metal SeSAE was rationalized by density functional theory calculations. Furthermore, comprehensive elucidation of the biological functions, cell death pathways, and metabolic remodeling effects of the nanozyme was conducted, aiming to provide valuable insights into the development of single atom nanozymes with clinical translation potential.

A Tumor-Specific Cascade-Activating Smart Prodrug System for Enhanced Targeted Therapy.

Developing intelligently targeted drugs with low side effects is urgent for cancer treatment. Toward this goal, a tumor-specific cascade-activating smart prodrug system consisting of a G-quadruplex(G4)-modulated tumor-targeted DNA vehicle and a well-designed cellular stimuli-responsive ligand-drug conjugates (LDCs) is proposed. An original "donor-acceptor" binary fluorescent ligand, with ultrahigh affinity, brightness, and photostability, is engineered to tightly bind G4 structures and significantly improve the nuclease resistance of the DNA vehicle, which serves as a bridge contributing to the construction of the prodrug system, named ApG4/LDCs. Sodium nitroprusside and doxorubicin are loaded into ApG4/LDCs in one pot and generate nitric oxide and superoxide anion in response to cancer cellular environments, which in cascade generates peroxynitrite to cause DNA damage while promoting the self-monitored drug release to achieve enhanced targeted therapy. Such a cascade activation and self-reinforcement process is executed only when the prodrug system targets the tumor tissue followed by cell uptake, showing significant antitumor efficacy and greatly weakening the damage to normal tissues. Given the unique features, the innovative strategy for prodrug design may open a new door to precision disease treatment.

Assessment of 2022 European LeukemiaNet risk classification system in real-world cohort from China.

BACKGROUND: The European LeukemiaNet (ELN) risk classification system for acute myeloid leukemia (AML) patients has been used worldwide. In 2022, the ELN risk classification system modified risk genes including CEBPA mutation status, myelodysplasia-related (MR) gene mutations and internal tandem duplications of FLT3 (FLT3-ITD). METHODS: We include newly diagnosed de novo AML patients at our center from January 2017 to December 2021, regardless of the further treatment received. Clinical data and date of survival were included. Survival analysis were performed using the Kaplan-Meier method, and the log-rank test was used to compare survival between different risk groups. RESULTS: We include 363 newly diagnosed de novo AML patients from 2017 to 2021 to assess the accuracy of the ELN risk classification system. Their survival results show that the ELN-2022 risk classification system is not superior to the ELN-2017 version; for patients with FLT3-ITD mutations but without FLT3 inhibitor treatment, their survival is similar to the ELN-2022 adverse risk group. The ELN-2022 risk classification system cannot accurately clarify ECOG performance status (PS) 2-4 patients, especially in the ELN-2022 favorable risk group. CONCLUSION: The ELN-2022 risk stratification system may not be appropriate for patients unable to receive intensive therapy or FLT3 inhibitor; more real-world data is needed to straify patients with worse ECOG PS and inferior intensive therapy.

A Leaking-Proof Theranostic Nanoplatform for Tumor-Targeted and Dual-Modality Imaging-Guided Photodynamic Therapy.

Objective: A protein-based leaking-proof theranostic nanoplatform for dual-modality imaging-guided tumor photodynamic therapy (PDT) has been designed. Impact Statement: A site-specific conjugation of chlorin e6 (Ce6) to ferrimagnetic ferritin (MFtn-Ce6) has been constructed to address the challenge of unexpected leakage that often occurs during small-molecule drug delivery. Introduction: PDT is one of the most promising approaches for tumor treatment, while a delivery system is typically required for hydrophobic photosensitizers. However, the nonspecific distribution and leakage of photosensitizers could lead to insufficient drug accumulation in tumor sites. Methods: An engineered ferritin was generated for site-specific conjugation of Ce6 to obtain a leaking-proof delivery system, and a ferrimagnetic core was biomineralized in the cavity of ferritin, resulting in a fluorescent ferrimagnetic ferritin nanoplatform (MFtn-Ce6). The distribution and tumor targeting of MFtn-Ce6 can be detected by magnetic resonance imaging (MRI) and fluorescence imaging (FLI). Results: MFtn-Ce6 showed effective dual-modality MRI and FLI. A prolonged in vivo circulation and increased tumor accumulation and retention of photosensitizer was observed. The time-dependent distribution of MFtn-Ce6 can be precisely tracked in real time to find the optimal time window for PDT treatment. The colocalization of ferritin and the iron oxide core confirms the high stability of the nanoplatform in vivo. The results showed that mice treated with MFtn-Ce6 exhibited marked tumor-suppressive activity after laser irradiation. Conclusion: The ferritin-based leaking-proof nanoplatform can be used for the efficient delivery of the photosensitizer to achieve an enhanced therapeutic effect. This method established a general approach for the dual-modality imaging-guided tumor delivery of PDT agents.

Patching sulfur vacancies: A versatile approach for achieving ultrasensitive gas sensors based on transition metal dichalcogenides.

Transition metal dichalcogenides (TMDCs) garner significant attention for their potential to create high-performance gas sensors. Despite their favorable properties such as tunable bandgap, high carrier mobility, and large surface-to-volume ratio, the performance of TMDCs devices is compromised by sulfur vacancies, which reduce carrier mobility. To mitigate this issue, we propose a simple and universal approach for patching sulfur vacancies, wherein thiol groups are inserted to repair sulfur vacancies. The sulfur vacancy patching (SVP) approach is applied to fabricate a MoS2-based gas sensor using mechanical exfoliation and all-dry transfer methods, and the resulting 4-nitrothiophenol (4NTP) repaired molybdenum disulfide (4NTP-MoS2) is prepared via a sample solution process. Our results show that 4NTP-MoS2 exhibits higher response (increased by 200 %) to ppb-level NO2 with shorter response/recovery times (61/82 s) and better selectivity at 25 °C compared to pristine MoS2. Notably, the limit of detection (LOD) toward NO2 of 4NTP-MoS2 is 10 ppb. Kelvin probe force microscopy (KPFM) and density functional theory (DFT) reveal that the improved gas sensing performance is mainly attributed to the 4NTP-induced n-doping effect on MoS2 and the corresponding increment of surface absorption energy to NO2. Additionally, our 4NTP-induced SVP approach is universal for enhancing gas sensing properties of other TMDCs, such as MoSe2, WS2, and WSe2.

An AND-gate bioluminescent probe for precise tumor imaging.

Sensitivity and specificity are two indispensable requirements to ensure diagnostic accuracy. Dual-locked probes with "AND-gate" logic theory have emerged as a powerful tool to enhance imaging specificity, avoid "false positive" results, and realize correlation analysis. In addition, bioluminescence imaging (BLI) is an excitation-free optical modality with high sensitivity and low background and can thus be combined with a dual-locked strategy for precise disease imaging. Here, we developed a novel AND-gate bioluminescent probe, FK-Luc-BH, which is capable of responding to two different tumor biomarkers (cathepsin L and ClO-). The good specificity of FK-Luc-BH was proven, as an obvious BL signal could only be observed in the solution containing both cathepsin L (CTSL) and ClO-. 4T1-fLuc cells and tumors treated with FK-Luc-BH exhibited significantly higher BL signals than those treated with unresponsive control compound Ac-Luc-EA or cotreated with FK-Luc-BH and a ClO- scavenger/cathepsin inhibitor, demonstrating the ability of FK-Luc-BH to precisely recognize tumors in which CTSL and ClO- coexist.

Gas-Mediated Tumor Energy Remodeling for Sensitizing Mild Photothermal Therapy.

The metabolic reprogramming of tumors requires high levels of adenosine triphosphate (ATP) to maintain therapeutic resistance, posing a major challenge for photothermal therapy (PTT). Although raising the temperature helps in tumor ablation, it frequently leads to severe side effects. Therefore, improving the therapeutic response and promoting healing are critical considerations in the development of PTT. Here, we proposed a gas-mediated energy remodeling strategy to improve mild PTT efficacy while minimizing side effects. In the proof-of-concept study, a Food and Drug Administration (FDA)-approved drug-based hydrogen sulfide (H2 S) donor was developed to provide a sustained supply of H2 S to tumor sites, serving as an adjuvant to PTT. This approach proved to be highly effective in disrupting the mitochondrial respiratory chain, inhibiting ATP generation, and reducing the overexpression of heat shock protein 90 (HSP90), which ultimately amplified the therapeutic outcome. With the ability to reverse tumor thermotolerance, this strategy delivered a greatly potent antitumor response, achieving complete tumor ablation in a single treatment while minimizing harm to healthy tissues. Thus, it holds great promise to be a universal solution for overcoming the limitations of PTT and may serve as a valuable paradigm for the future clinical translation of photothermal nanoagents.

High temporal waveform fidelity stimulated Brillouin scattering phase conjugate mirror using Novec-7500.

A high temporal waveform fidelity stimulated Brillouin scattering phase conjugate mirror (SBS-PCM) with high energy efficiency, based on a novel medium, Novec-7500, is proposed and practically achieved in this study. A theoretical analysis reveals that the temporal-domain waveform distortion is caused by the inherent pulse duration compression effect of the SBS, and this undesirable phenomenon can be significantly suppressed by decreasing the compression coefficient (CC afterwards), which is defined as the gain coefficient divided by the phonon lifetime, which coefficient and is identified as the key parameter for high waveform-fidelity in SBS-PCM. The feasibility of this approach was demonstrated experimentally, in which a reflected pulse with waveform symmetry equals to the pump and an average pulse duration of 0.974 τp (τp is the duration of pump) with an energy efficiency of over 90% was achieved using Novec-7500.

Modeling and characterization of high-power single frequency free-space Brillouin lasers.

Free-space Brillouin lasers (BLs) are capable of generating high-power, narrow-linewidth laser outputs at specific wavelengths. Although there have been impressive experimental demonstrations of these lasers, there is an absence of a corresponding theory that describes the dynamic processes that occur within them. This paper presents a time-independent analytical model that describes the generation of the first-order Stokes field within free-space BLs. This model is based on the cavity resonance enhancement theory and coupled wave equations that govern the processes of stimulated Brillouin scattering (SBS). This model is validated using an experimental diamond BL to numerically simulate the influence of the cavity design parameters on the SBS threshold, pump enhancement characteristics, and power of the generated Stokes field. Specifically, the model is used to determine the SBS cavity coupler reflectance to yield the maximum Stokes field output power and efficiency, which is also a function of the pump power and other cavity design parameters. This analysis shows that the appropriate choice of Brillouin cavity coupler reflectance maximizes the Stokes field output power for a given pump power. Furthermore, the onset of higher-order Stokes fields that are undesirable in the context of single-frequency laser operation were inhibited. This study aids in understanding the relationship between the cavity parameters and resultant laser characteristics for the design and optimization of laser systems.

Boosting Ferroptosis Therapy with Iridium Single-Atom Nanocatalyst in Ultralow Metal Content.

Nanocatalysts are promising tumor therapeutics due to their ability to induce reactive oxygen species in the tumor microenvironment. Although increasing metal loading can improve catalytic activity, the quandary of high metal content versus potential systemic biotoxicity remains challenging. Here, a fully exposed active site strategy by site-specific anchoring of single iridium (Ir) atoms on the outer surface of a nitrogen-doped carbon composite (Ir single-atom catalyst (SAC)) is reported to achieve remarkable catalytic performance at ultralow metal content (≈0.11%). The Ir SAC exhibits prominent dual enzymatic activities to mimic peroxidase and glutathione peroxidase, which catalyzes the conversion of endogenous H2 O2 into •OH in the acidic TME and depletes glutathione (GSH) simultaneously. With an advanced support of GSH-trapping platinum(IV) and encapsulation with a red-blood-cell membrane, this nanocatalytic agent (Pt@IrSAC/RBC) causes intense lipid peroxidation that boosts tumor cell ferroptosis. The Pt@IrSAC/RBC demonstrates superior therapeutic efficacy in a mouse triple-negative mammary carcinoma model, resulting in complete tumor ablation in a single treatment session with negligible side effects. These outcomes may provide valuable insights into the design of nanocatalysts with high performance and biosafety for biomedical applications.

Protein encapsulation of nanocatalysts: A feasible approach to facilitate catalytic theranostics.

Enzyme-mimicking nanocatalysts, also termed nanozymes, have attracted much attention in recent years. They are considered potential alternatives to natural enzymes due to their multiple catalytic activities and high stability. However, concerns regarding the colloidal stability, catalytic specificity, efficiency and biosafety of nanomaterials in biomedical applications still need to be addressed. Proteins are biodegradable macromolecules that exhibit superior biocompatibility and inherent bioactivities; hence, the protein modification of nanocatalysts is expected to improve their bioavailability to match clinical needs. The diversity of amino acid residues in proteins provides abundant functional groups for the conjugation or encapsulation of nanocatalysts. Moreover, protein encapsulation can not only improve the overall performance of nanocatalysts in biological systems, but also bestow materials with new features, such as targeting and retention in pathological sites. This review aims to report the recent developments and perspectives of protein-encapsulated catalysts in their functional improvements, modification methods and applications in biomedicine.

Photothermal Enhanced and Tumor Microenvironment Responsive Nanozyme for Amplified Cascade Enzyme Catalytic Therapy.

Nanocatalysts, a class of nanomaterials with intrinsic enzyme-like activities, have been widely investigated for cancer catalytic therapy in recent years. However, precise construction of nanocatalysts with excellent enzyme catalytic activity and biosafety for tumor therapy still remains challenging. Here, a biodegradable nanocatalyst, PEGylated Cux Mny Sz (PCMS), is reported that can promote cascade catalytic reactions in tumor microenvironment (TME) while confining off-target side effects on normal tissues. PCMS not only catalyzes the cascade conversion of endogenous hydrogen peroxide (H2 O2 ) to oxygen (O2 ) via catalase-like activity and then to superoxide radical (·O2 - ) via oxidase-like activity in the TME, but also effectively depletes intracellular glutathione via glutathione oxidase-like activity. The cascade catalytic reactions, by taking advantage of high H2 O2 level in tumor cells, result in an enhanced enzyme catalytic effect in generation of ·O2 - . More importantly, PCMS exhibits prominent photothermal effect under NIR-II 1064 nm laser irradiation that can further enhance chemodynamic therapy (CDT) efficacy in tumors. In addition, the biodegradation in TME and excellent photothermal effect of PCMS are beneficial to magnetic resonance imaging, photoacoustic imaging and infrared thermal imaging, resulting in tracing the fate of PCMS in vivo. This study provides a new tool for rational design of TME-responsive nanocatalysts with high biocompatibility for tumor catalytic therapy.

A new isolate of Streptomyces lateritius (Z1-26) with antibacterial activity against fish pathogens and immune enhancement effects on crucian carp (Carassius auratus).

The Streptomyces lateritius Z1-26 was isolated from soil samples which showed broad-spectrum antibacterial activity against a broad range of fish pathogens. The In Vivo Imaging System (IVIS) monitored that strain Z1-26 could survive and colonize in the gills and abdomen of crucian carp. The effects of dietary supplementation with strain Z1-26 were evaluated with respect to the growth performance, antioxidant capacity, and immune response of crucian carp. The results showed that the Z1-26-fed fish had a significantly higher growth rate than the fish fed the control diet. The immune and antioxidant parameters revealed that the non-specific immune indicators (AKP, SOD, and LZM) of the serum, the expression of immune-related genes (IgM, C3, and LZM), and antioxidant-related genes (Nrf2 and Keap1) of the immune organs were significantly increased, whereas the expression of pro-inflammatory factors (IL-1ß, IL-8, and TNF-α) of the immune organs was significantly down-regulated in crucian carp fed strain Z1-26 compared with fish fed a control diet. Moreover, fish fed with Z1-26 supplemented diets showed a significantly improved survival rate after Aeromonas hydrophila infection. In addition, the whole genome analysis showed that strain Z1-26 possesses 28 gene clusters, including 6 polyketide synthetase (PKS), 4 non-ribosomal peptide-synthetase (NRPS), 1 bacteriocin, and 1 lantipeptide. In summary, these results indicated that strain Z1-26 could improve the growth performance and disease resistance in crucian carp, and has the potential to be developed as a candidate probiotics for the control of bacterial diseases in aquaculture.

Screening of new Paenibacillus polymyxa S3 and its disease resistance of grass carp (Ctenopharyngodon idellus).

A new strain of Paenibacillus polymyxa S3 with antagonistic effects on 11 major fish pathogens (especially Aeromonas hydrophila), but had no toxicity to grass carp, was screened from the sediment of fishponds. In vivo colonization studies showed that strain S3 could be colonized and distributed in the gill and abdomen of the grass carp. Bioassay results showed that the weight growth rate of grass carp in the strain S3 oral group (16.01%) and strain S3 immersion group (16.44%) was significantly higher than those of the control group (8.61%). At the same time, the activities of ACP, AKP, CAT and GSH-Px in the serum of grass carp in oral and immersion groups were significantly higher than those of the control group. In addition, the treatment with strain S3 could significantly upregulate the expression of the antioxidant-related genes and immune-related genes Keap1, Nrf2, C3, LZM, IgM, TLR-4 and MyD-88 in grass carp tissues. The challenge test showed that strain S3 treatment significantly increased the survival rate of grass carp infected with Aeromonas hydrophila. Whole genome sequencing analysis showed that strain S3 had 16 active metabolite gene clusters, indicating that it had abundant gene resources, which provided important support for its development for fish microecological preparations. In summary, a new strain of Paenibacillus polymyxa S3 with antibacterial activity against a variety of fish pathogens was screened in this study and its probiotic function was evaluated, proving its potential value in fisheries.

Narrow laser-linewidth measurement using short delay self-heterodyne interferometry.

Delayed self-heterodyne interferometry is a commonly used technique for the measurement of laser linewidth. It typically requires the use of a very long delay fiber when measuring narrow linewidth (especially linewidths in the kHz-range) lasers. The use of long fibers can result in system losses and the introduction of 1/f noise that causes spectral line broadening. In this paper, we present a calculation method for processing the output of a delayed self-heterodyne setup using a short length of delay fiber, to determine laser linewidth. The method makes use of pairs of data points (corresponding to adjacent maxima and/or minima) in the signal generated from the self-heterodyne setup to determine the laser linewidth. Here, the power ratio or amplitude difference of the signal at these data points is of importance. One of the key benefits of this method is that it avoids 1/f noise which would otherwise be introduced into the measurement through the application of long fibers. The experimental results highlight that the method has a high calculation accuracy. Furthermore, the capacity for the method to utilize different pairs of data points in the self-heterodyne output to determine the laser linewidth, imparts a high degree of flexibility and usability to the technique when applied to real-world measurements.

22.5-W narrow-linewidth diamond Brillouin laser at 1064†nm.

Stimulated Brillouin scattering (SBS), with its advantages of low quantum defect and narrow gain bandwidth, has recently enabled an exciting path toward narrow-linewidth and low-noise lasers. Whereas almost all work to date has been in guided-wave configurations, adaptation to unguided Brillouin lasers (BLs) offers a greater capacity for power scaling, cascaded Stokes control, and greater flexibility for expanding wavelength range. Here, we report a diamond Brillouin laser (DBL) employing doubly resonant technology at 1064 nm. Brillouin output power of 22.5 W with a linewidth of 46.9 kHz is achieved. The background noise from the pump amplified spontaneous emission (ASE) is suppressed by 35 dB. The work represents a significant step toward realizing Brillouin oscillators that simultaneously have high power (tens-of-watts+) and kHz-linewidths.

The Influence of Noise Floor on the Measurement of Laser Linewidth Using Short-Delay-Length Self-Heterodyne/Homodyne Techniques.

Delayed self-heterodyne/homodyne measurements based on an unbalanced interferometer are the most used methods for measuring the linewidth of narrow-linewidth lasers. They typically require the service of a delay of six times (or greater) than the laser coherence time to guarantee the Lorentzian characteristics of the beat notes. Otherwise, the beat notes are displayed as a coherent envelope. The linewidth cannot be directly determined from the coherence envelope. However, measuring narrow linewidths using traditional methods introduces significant errors due to the 1/f frequency noise. Here, a short fiber-based linewidth measurement scheme was proposed, and the influence of the noise floor on the measurement of the laser linewidth using this scheme was studied theoretically and experimentally. The results showed that this solution and calibration process is capable of significantly improving the measurement accuracy of narrow linewidth.

Aeromonas veronii infection remarkably increases expression of lysozymes in grass carp (Ctenopharyngodon idellus) and injection of lysozyme expression cassette along with QCDC adjuvant significantly upregulates immune factors and decreases cumulative mortality.

Aeromonas veronii AvX005 is a pathogenic bacterium with high toxicity to grass carp (Ctenopharyngodon idellus). The expression levels of g-type (goose-type lysozyme, Lys-g) and c-type lysozyme (chicken-type lysozyme, Lys-c) in the spleen of grass carp infected with AvX005 were significantly increased by approximately 4.5 times and 27 times, respectively. The recombinant proteins rLys-g and rLys-c produced in a recombinant expression system of Escherichia coli showed significant antibacterial activity against the pathogenic bacteria AvX005. A challenge test was conducted after rLys-g and rLys-c were expressed in grass carp L8824 liver cells, and compared with the survival rate of the control cells (46.3%), the survival rate of the experimental cells (77.6% for rLys-g and 68.6% for rLys-c) was significantly increased. Grass carp were infected with AvX005 on the second day after delivering pcDNA3.1-lys-g and pcDNA-lys-c with the Quil A/cholesterol/DDA/Carbopol (QCDC) adjuvant, and both pcDNA3.1-lys-g and pcDNA-lys-c provided 70% relative protection for grass carp. The activity of lysozyme and alkaline phosphatase in the serum of grass carp was significantly increased after injection of DNA. The expression of the immune factors IgM, C3 and IL8 in the kidney was upregulated to varying degrees for pcDNA3.1-lys-g and immune factors C3 and IgM was upregulated for pcDNA-lys-c. The results indicated that pcDNA3.1-lys-g and pcDNA-lys-c may be used as immunostimulants to protect grass carp from the pathogenic bacterium AvX005.