Weissella confusa N17 Derived from Loach (Misgurnus anguillicaudatus) Exhibits Promising for Further Applications in Loach Aquaculture.

The application of probiotics, in aquaculture, is becoming increasingly widespread and have had positive application effects. However, reports of loach-derived probiotics are quite limited. In this study, two representative strains of lactic acid bacteria with excellent traits, namely, Weissella confusa N17 and Lactobacillus saniviri N19, were screened from the intestine of healthy loaches. W. confusa N17 and L. saniviri N19 could inhibit different common various pathogenic bacteria, especially Aeromonas spp., and were sensitive to the most common antibiotics. The survival rate of the two strains exceeded 50% after 4 h of incubation in 10% loach bile. Moreover, the two strains showed significant tolerance to trypsin. Their autoaggregation capacity and hydrophobicity were greater than 30%. In addition, the aggregation ability of both strains was higher than 30% for both A. veronii TH0426 and A. hydrophila TPS. The two strains had a high biofilm-forming ability and strong adhesion to epithelioma papulosum cyprini (EPC) cells. Scanning electron microscopy results showed that the culture supernatants of the two strains had a significantly destructive effect on A. veronii TH0426 and A. hydrophila TPS. Overall, the traits of W. confusa N17 were better than those of L. saniviri N19. Genome sequencing and analysis demonstrated a lack of virulence factor-related or drug resistance-related genes in genome N17. The diet supplemented with the W. confusa N17 strain significantly improved the resistance of loaches to A. veronii infection, and the protection rate reached 57.1%. Therefore, W. confusa N17 exhibits promising for further applications in loach aquaculture.

Lc-pPG-612-OmpU-CTB: A promising oral vaccine for protecting Carassius auratus against Vibrio mimicus infection.

Vibrio mimicus (V. mimicus) is known to cause severe bacterial diseases with high mortality rates in fish, resulting in significant economic losses in the global aquaculture industry. Therefore, the objective of this study was to develop a safe and effective vaccine for protecting Carassius auratus (C. auratus) against V. mimicus infection. Recombinant Lactobacillus casei (L. casei) strains, Lc-pPG-612-OmpU and Lc-pPG-612-OmpU-CTB (surface-displayed), were constructed using a L. casei strain (ATCC 393) as an antigen delivery carrier and the cholera toxin B subunit (CTB) as an adjuvant. The two recombinant strains of L. casei were administered to C. auratus via oral immunization, and the protective efficacy of the oral vaccines was assessed. The results demonstrated that oral immunization with the two strains significantly increased the levels of nonspecific immune indicators in C. auratus, including alkaline phosphatase (AKP), lysozyme (LYS), acid phosphatase (ACP), complement 3 (C3), complement 4 (C4), lectin, and superoxide dismutase (SOD). Moreover, the experiment groups exhibited significant increases in specific immunoglobulin M (IgM) antibodies against OmpU, as well as the transcription of immune-related genes (ie., IL-1ß, TNF-α, IL-10, and TGF-ß), when compared to the control groups. Following infection of C. auratus with V. mimicus, the mortality rate of the recombinant L. casei-treated fish was observed to be lower compared to the control group. This finding suggests that recombinant L. casei demonstrates effective protection against V. mimicus infection in C. auratus. Furthermore, the addition of the immune adjuvant CTB was found to induce a more robust adaptive and innate immune response in C. auratus, resulting in reduced mortality after infection with V. mimicus.

Efficacy of a recombinant Lactobacillus plantarum Lp-pPG-Malt as an oral vaccine candidate against Aeromonas hydrophila infection in crucian carp.

Aeromonas hydrophila (A. hydrophila), a gram-negative bacterium, causes serious diseases with various clinical symptoms in farm raised fish. Thus, different ways to prevent and control A. hydrophila infection need to be explored, including a vaccine. In this study, we evaluated the protective efficacy of an oral vaccine prepared from the A. hydrophila TPS maltoporin (Malt) with Lactobacillus plantarum (L. plantarum) against A. hydrophila infection in crucian carp (Carassius auratus). For the in vivo experiment, the oral vaccine was administered to crucian carp by feeding them fish diets containing Lp-pPG-Malt, Lp-pPG and PBS for 28 days. The enzyme-linked immunosorbent assay (ELISA), leukocyte phagocytosis assay and real-time quantitative polymerase chain reaction (RT-qPCR) were performed to measure the protective efficacy of the Lp-pPG-Malt. ELISA and leukocyte phagocytosis assay confirmed that Lp-pPG-Malt significantly enhanced the IgM level and nonspecific immune response of crucian carp compared with the control groups (Lp-pPG and PBS). The RT-qPCR results showed that the Lp-pPG-Malt increased the relative expression of immune-related genes (IL-10, IL-1ß, TNF-α, IFN-γ) of crucian carp in various tissues (liver, spleen, head kidney and hind intestine). Moreover, Lp-pPG-Malt significantly increased the relative percent survival of fish after intraperitoneal injection with A. hydrophila (55%) compared with the Lp-pPG and PBS groups (0%). These findings suggest that Lp-pPG-Malt can serve as an oral vaccine candidate for A. hydrophila infection and that Malt can be used as an effective antigen in crucian carp farming.

Oral vaccination with recombinant Lactobacillus casei with surface displayed OmpK fused to CTB as an adjuvant against Vibrio mimicus infection in Carassius auratus.

Vibrio mimicus (V. mimicus) is a pathogenic bacterium that causes diseases in humans and various aquatic animals. A particularly efficient way to provide protection against V. mimicus is through vaccination. However, there are few commercial vaccines against V. mimics, especially oral vaccines. In our study, two surface-display recombinant Lactobacillus casei (L. casei) Lc-pPG-OmpK and Lc-pPG-OmpK-CTB were constructed using L. casei ATCC393 as an antigen delivery vector, outer membrane protein K (OmpK) of V. mimicus as an antigen, and cholera toxin B subunit (CTB) as a molecular adjuvant; furthermore, the immunological effects of recombinant L.casei in Carassius auratus (C. auratus) were assessed. The results indicated that oral recombinant L.casei Lc-pPG-OmpK and Lc-pPG-OmpK-CTB stimulated higher levels of serum-specific immunoglobulin M (IgM) and increased the activity of acid phosphatase (ACP), alkaline phosphatase (AKP), superoxide dismutase (SOD), lysozyme (LYS), lectin, C3, and C4 in C. auratus, compared with control groups (Lc-pPG group and PBS group). Furthermore, the expression of interleukin-1ß (IL-1ß), interleukin-10 (IL-10), tumor necrosis factor-α (TNF-α), and transforming growth factor-ß (TGF-ß) in the liver, spleen, head kidney, hind intestine and gills of C. auratus was significantly increased, compared with that in the controls. These results demonstrated that the two recombinant L. casei strains could effectively trigger humoral and cellular immunity in C. auratus. In addition, two recombinant L.casei strains were able to survive and colonize the intestine of C. auratus. Importantly, after being challenged with V. mimicus, C. auratus fed Lc-pPG-OmpK and Lc-pPG-OmpK-CTB exhibited greater survival rates than the controls (52.08% and 58.33%, respectively). The data showed that recombinant L. casei could elicit a protective immunological response in C. auratus. The effect of the Lc-pPG-OmpK-CTB group was better than that of the Lc-pPG-OmpK group, and Lc-pPG-OmpK-CTB was found to be an effective candidate for oral vaccination.

Gut Microbiota and Metabolites May Play a Crucial Role in Sea Cucumber Apostichopus japonicus Aestivation.

The constant increase in temperatures under global warming has led to a prolonged aestivation period for Apostichopus japonicus, resulting in considerable losses in production and economic benefits. However, the specific mechanism of aestivation has not been fully elucidated. In this study, we first tried to illustrate the biological mechanisms of aestivation from the perspective of the gut microbiota and metabolites. Significant differences were found in the gut microbiota of aestivating adult A. japonicus (AAJSD group) compared with nonaestivating adult A. japonicus (AAJRT group) and young A. japonicus (YAJRT and YAJSD groups) based on 16S rRNA gene high-throughput sequencing analysis. The abundances of Desulfobacterota, Myxococcota, Bdellovibrionota, and Firmicutes (4 phyla) in the AAJSD group significantly increased. Moreover, the levels of Pseudoalteromonas, Fusibacter, Labilibacter, Litorilituus, Flammeovirga, Polaribacter, Ferrimonas, PB19, and Blfdi19 genera were significantly higher in the AAJSD group than in the other three groups. Further analysis of the LDA effect size showed that species with significant variation in abundance in the AAJSD group, including the phylum Firmicutes and the genera Litorilituus, Fusibacter, and Abilibacter, might be important biomarkers for aestivating adult A. japonicus. In addition, the results of metabolomics analysis showed that there were three distinct metabolic pathways, namely biosynthesis of secondary metabolites, tryptophan metabolism, and sesquiterpenoid and triterpenoid biosynthesis in the AAJSD group compared with the other three groups. Notably, 5-hydroxytryptophan was significantly upregulated in the AAJSD group in the tryptophan metabolism pathway. Moreover, the genera Labilibacter, Litorilituus, Ferrimonas, Flammeovirga, Blfdi19, Fusibacter, Pseudoalteromonas, and PB19 with high abundance in the gut of aestivating adult A. japonicus were positively correlated with the metabolite 5-HTP. These findings suggest that there may be potential biological associations among the gut microbiota, metabolites, and aestivation in A. japonicus. This work may provide a new perspective for further understanding the aestivation mechanism of A. japonicus.

A systematic review, meta-analysis and meta-regression of the global prevalence of foodborne Vibrio spp. infection in fishes: A persistent public health concern.

Human vibriosis, caused by pathogenic Vibrio spp., such as Vibrio parahaemolyticus, Vibrio cholerae and Vibrio vulnificus, has been increasing worldwide, mediated by increasing consumption of seafood. The present study was conducted to examine the global prevalence of V. vulnificus, V. parahaemolyticus and V. cholerae in fishes. We searched PubMed, Web of Science, Scopus, and CNKI for peer-reviewed articles and dissertations prior to December 31, 2021. A total of 24,831 articles were retrieved, and 82 articles contained 61 fish families were included. The global pooled prevalence of V. cholerae, V. parahaemolyticus and V. vulnificus in fishes was 9.56 % (95 % CI: 2.12-20.92), 24.77 % (95 % CI: 17.40-32.93) and 5.29 % (95 % CI: 0.38-13.61), respectively. Subgroup and meta-regression analyses showed that study-level covariates, including temperature, country, continent, origin and detection methods partly explained the between-study heterogeneity. These heterogeneities were underpinned by differences of the three Vibrio spp. in fishes at geographical and climatic scales. These results reveal a high global prevalence of pathogenic Vibrio spp. in fishes and highlight the need for implementation of more effective prevention and control measures to reduce food-borne infection in humans.

Critical roles of VipB protein on virulence and oxidative stress tolerance in Aeromonas veronii.

Aeromonas veronii is a zoonotic pathogen capable of causing sepsis and ulceration in freshwater fish. Recently, reports of numerous cases indicate a marked increase in pathogenicity. Nonetheless, little is known about the pathogenesis of A. veronii infections. In this study, an in-frame mutant of the A. veronii vipB gene was generated to investigate its biological function. Deletion of the vipB gene resulted in a significant 204.71-fold decrease in the LD50 of A. veronii against zebrafish and a 2-fold and 4-fold reduction in the toxicity to EPC cells at 1 h and 2 h of infection, respectively. The virulence-related genes of the mutant ΔvipB all showed significantly reduced expression levels compared to the wild strain. In addition, the motility of the mutant ΔvipB decreased significantly, the adhesion ability to EPC cells was 3.25-fold lower than that of the parental strain, and the oxidative stress tolerance was 2.31-fold lower than that of TH0426 strain. In contrast, the biofilm formation amount of ΔvipB strain increased by 1.65-fold at both 12 h and 24 h. Our findings suggest that the vipB gene is associated with flagella stability, virulence, and oxidative stress tolerance and plays critical roles in the pathogenesis of A. veronii infections.

Lp-pPG-611.1-LPS as an immune enhancer provides effective protection against Aeromonas veronii infection in Carassius auratus.

Aeromonas veronii (A. veronii) is an important zoonotic pathogen that causes substantial economic losses in aquaculture. In this study, we aimed to develop a safe and effective immune enhancer to protect Carassius auratus (C. auratus) from A. veronii infections. With recognized safety, lactic acid bacteria are used as antigen delivery vehicles to present antigens. Lipopolysaccharide (LPS), a protective antigen, induces immune responses in animals. Therefore, we created recombinant Lactobacillus plantarum (L. plantarum) with surface-displayed LPS of A. veronii TH0426 and tested its effects on immune responses in C. auratus. The results showed that recombinant L. plantarum Lp-pPG-611.1-LPS, as an immune enhancer, could improve the innate and adaptive immune responses of C. auratus when it was added to the diet of C. auratus. The challenge test showed that the survival rate of C. auratus fed with L. plantarum Lp-pPG-611.1-LPS was higher than that of the control groups, indicating that the recombinant L. plantarum Lp-pPG-611.1-LPS increased the resistance of C. auratus to A. veronii infection. The present results provide a theoretical basis for the development of recombinant L. plantarum Lp-pPG-611.1-LPS as an immune enhancer in aquaculture.

Functional analysis of ascP in Aeromonas veronii TH0426 reveals a key role in the regulation of virulence.

Aeromonas veronii is a pathogen which can induce diseases in humans, animals and aquatic organisms, but its pathogenic mechanism and virulence factors are still elusive. In this study, we successfully constructed a mutant strain (ΔascP) by homologous recombination. The results showed that the deletion of the ascP gene significantly down-regulated the expression of associated effector proteins in A. veronii compared to its wild type. The adhesive and invasive abilities of ΔascP to EPC cells were 0.82-fold lower in contrast to the wild strain. The toxicity of ΔascP to cells was decreased by about 2.91-fold (1 h) and 1.74-fold (2 h). Furthermore, the LD50 of the mutant strain of crucian carp was reduced by 19.94-fold, and the virulence was considerably attenuated. In contrast to the wild strain, the ΔascP content in the liver and spleen was considerably lower. The titers of serum cytokines (IL-8, TNF-α, and IL-1ß) in crucian carp after the infection of the ΔascP strain were considerably lower in contrast to the wild strain. Hence, the ascP gene is essential for the etiopathogenesis of A. veronii TH0426.

Terrestrial runoff influences the transport and contamination levels of Toxoplasma gondii in marine organisms.

There is a growing concern regarding the potential adverse impact of Toxoplasma gondii contamination of the marine environment on marine wildlife and public health. Terrestrial runoff is a significant route for dissemination of T. gondii oocysts from land to sea. Yet, the influence of terrestrial runoff on T. gondii prevalence in marine animals in China is largely unknown. To address this concern, we examined the presence of T. gondii in marine oysters Crassostrea spp., rockfish Sebastes schlegelii (S. schlegelii), fat greenling fish Hexagrammos otakii (H. otakii), and Asian paddle crab Charybdis japonica (C. japonica) using a PCR assay targeting T. gondii B1 gene. A total of 1920 samples were randomly collected, in Jan-Dec 2020, from terrestrial runoff areas (TRA, TRB, and TRC) and non-terrestrial runoff area (Grape bay) in Weihai, China. T. gondii prevalence in TRB and TRC was 6.04 % and 5.83 %, respectively, which was higher than 2.29 % detected in the non-terrestrial runoff area. The highest prevalence was detected in Crassostrea spp., and a correlation was observed between T. gondii prevalence and weight of Crassostrea spp. The temperature, but not precipitation, significantly correlated with T. gondii prevalence. Understanding the fate of T. gondii delivered to oceans by terrestrial runoff is critical for predicting future disease risks for marine wildlife and humans.

Essential role of ascO for virulence of Aeromonas veronii and inducing apoptosis.

Aeromonas veronii is a significant pathogen that is capable of infecting humans, animals, and aquatic animals. The type III secretion system (T3SS) is intimately associated with bacterial pathogenicity. The ascO gene is an important core component of T3SS in A. veronii, but its function is still unclear. The ascO gene of A. veronii TH0426 was deleted by using the pRE112 suicide plasmid to study its function. The study results showed that the ability of ∆ascO to adhere and invade EPC cells was significantly reduced by 1.28 times. The toxicity of the mutant strain ΔascO to EPC cells was consistently significantly lower than wild-type strain TH0426 at 1, 2, and 4 h. The LD50 values of ∆ascO against zebrafish and Carassius auratus (C. auratus) were 53 and 15 times that of the wild-type strain. In addition, the bacterial load of the mutant strain ΔascO in blood, heart, liver, and spleen was lower than wild-type strain TH0426. The Hoechst staining showed that the apoptotic degree of EPC cells induced by the mutant strain ΔascO was lower than that of the wild-type strain TH0426. Furthermore, real-time quantitative PCR (RT-qPCR) analysis revealed lower expression levels of pro-apoptotic genes (including cytC, cas3, cas9, TNF-α, and IL-1ß) in C. auratus tissues infected with the mutant strain ΔascO compared to the wild-type strain TH0426. The results of in vivo and in vitro experiments have shown that ascO gene mutation can reduce the adhesion and toxicity of A. veronii to EPC and reduce the level of apoptosis induced by A. veronii. As a result, these insights will help further elucidate the function of the ascO gene and thus contribute to understanding the pathogenesis of A. veronii.

Dietary supplementation with Weissella cibaria C-10 and Bacillus amyloliquefaciens T-5 enhance immunity against Aeromonas veronii infection in crucian carp (Carassiu auratus).

With the aim to discover novel lactic acid bacteria and Bacillus strains from fish as potential probiotics to replace antibiotics in aquaculture, the present study was conducted to isolate lactic acid bacteria and Bacillus from intestinal tract of healthy crucian carp (Carassiu auratus) and largemouth bass (Micropterus salmoides) and evaluate their resistance against Aeromonas veronii. Based on the evaluation of antibacterial activity and tolerance test, one strain of lactic acid bacteria (Weissella cibaria C-10) and one strain of Bacillus (Bacillus amyloliquefaciens T-5) with strong environmental stability were screened out. The safety evaluation showed that these two strains were non-toxic to crucian carp and were sensitive to most antibiotics. In vivo study, the crucian carps were fed a basal diet supplemented with W. cibaria C-10 (C-10), B. amyloliquefaciens T-5 (T-5) and W. cibaria C-10 + B. amyloliquefaciens T-5 (C-10+T-5), respectively, for 5 weeks. Then, various immune parameters were measured at 35 days of post-feeding. Results showed both probiotics could improve the activities of related immune enzymes, immune factors and non-specific immune antibodies in blood and organs (gill, gut, kidney, liver, and spleen) of crucian carp in varying degrees. Moreover, after 7 days of challenge experiment, the survival rates after challenged with A. veronii of W. cibaria C-10 (C-10), B. amyloliquefaciens T-5 (T-5) and W. cibaria C-10 + B. amyloliquefaciens T-5 (C-10+T-5) supplemented groups to the crucian carps were 20%, 33% and 22%, respectively. Overall, W. cibaria C-10 and B. amyloliquefaciens T-5 could be considered to be developed into microecological preparations for the alternatives of antibiotics in aquaculture.

Focus on the Mechanisms and Functions of Pyroptosis, Inflammasomes, and Inflammatory Caspases in Infectious Diseases.

Eukaryotic cells can initiate several distinct self-destruction mechanisms to display essential roles for the homeostasis maintenance, development, and survival of an organism. Pyroptosis, a key response mode in innate immunity, also referred to as caspase-1-dependent proinflammatory programmed necrotic cell death activated by human caspase-1/4/5, or mouse caspase-1/11, plays indispensable roles in response to cytoplasmic insults and immune defense against infectious diseases. These inflammatory caspases are employed by the host to eliminate pathogen infections such as bacteria, viruses, protozoans, and fungi. Gasdermin D requires to be cleaved and activated by these inflammatory caspases to trigger the pyroptosis process. Physiological rupture of cells results in the release of proinflammatory cytokines, the alarmins IL-1ß and IL-18, symbolizing the inflammatory potential of pyroptosis. Moreover, long noncoding RNAs play direct or indirect roles in the upstream of the pyroptosis trigger pathway. Here, we review in detail recently acquired insights into the central roles of inflammatory caspases, inflammasomes, and pyroptosis, as well as the crosstalk between pyroptosis and long noncoding RNAs in mediating infection immunity and pathogen clearance.

Effects of four different adjuvants separately combined with Aeromonas veronii inactivated vaccine on haematoimmunological state, enzymatic activity, inflammatory response and disease resistance in crucian carp.

The purpose of the current study was to explore the immunomodulatory effects of different adjuvants combined with inactivated vaccines under Aeromonas veronii TH0426 infection in crucian carp. This study explored the best conditions for A. veronii as an inactivated vaccine, and included an animal safety test. Furthermore, we expressed the flagellin FlaA of the A. veronii TH0426 strain for use as an adjuvant supplemented in the diet. Crucian carp were fed 12 different experimental diets for 35 days, including the administration of 10 different adjuvants and inactivated vaccine combinations (50% aluminum hydroxide gel and inactivated vaccine combination, and inactivated vaccine with 20%, 30%, or 50% glucan, astragalus polysaccharide or flagellin), inactivated vaccine alone, and PBS control without adjuvant and inactivated vaccine. After the 42 day feeding trials, the fish were challenged with A. veronii TH0426, and the survival rate over 14 days was recorded. In addition, flagellin FlaA can be expressed normally in large amounts. All experimental groups produced higher levels of IgM serum titres than the control group in the different feeding cycles. Moreover, the activity of serum ACP, AKP, SOD, and LZM, and the expression of inflammatory factors were significantly increased in the experimental groups compared with the control group. The results of qRT-PCR analysis showed that the transcription levels of the IL-10, IL-1ß, IFN-γ and TNF-α genes in heart, liver, spleen and kidney tissues were significantly enhanced by adjuvant treatment, indicating that the addition of adjuvants can significantly promote the body's inflammatory response. In addition, the phagocytic activity of leukocytes in each adjuvant treated group was significantly enhanced compared to that in the groups without adjuvant. After the A. veronii challenge, the survival rate of all adjuvant-treated groups was significantly higher than that of the control group, and the 50% flagellin adjuvant group had the highest rate of 78.37%. Overall, our findings strongly indicate that adjuvants not only significantly improve the body's immunity, but also exhibit a strong anti-infection ability. Importantly, this work provides a new perspective for the prevention and control of aquaculture diseases.

Functional analysis of fis in Aeromonas veronii TH0426 reveals a key role in the regulation of virulence.

Aeromonas veronii is a comorbid pathogen that can infect humans, and animals including various aquatic organisms. In recent years, an increasing number of cases of A. veronii infection has been reported, indicating serious risks. This bacterium not only threatens public health and safety but also causes considerable economic loss in the aquaculture industry. Currently, some understanding of the pathogenic mechanism of A. veronii has been obtained. In this study, we first constructed the A. veronii TH0426 fis gene deletion strain Δfis and the complementation strain C-fis through homologous recombination technology. The results showed that the adhesion and invasion ability of the Δfis strain towards Epithelioma papulosum cyprini (EPC) cells and the cytotoxicity were 3.8-fold and 1.38-fold lower, respectively, than those of the wild-type strain. In the zebrafish infection model, the lethality of the deleted strain is 3-fold that of the wild strain. In addition, the bacterial load of the deletion strain Δfis in crucian carp was significantly lower than the wild-type strain, and the load decreased with time. In summary, deletion of the fis gene led to a decrease in the virulence of A. veronii. Our research results showed that the deletion of the fis gene significantly reduces the virulence and adhesion ability of A. veronii TH0426. Therefore, the fis gene plays a vital role in the pathogenesis of A. veronii TH0426. This preliminary study of the function of the fis gene in A. veronii will help researchers further understand the pathogenic mechanism of A. veronii.

Comparative genomics analysis of strains from diverse sources reveals the evolutionary relationship of Aeromonas veronii.

Aeromonas veronii (A. veronii, AV) strains are emerging zoonotic and aquatic pathogens, yet we know very little about their genomics. This study aims to utilize comparative genomics to investigate the intraspecific genetic diversity, differences in virulence factors and evolutionary mechanisms of A. veronii strains from diverse sources and to fundamentally demonstrate their pathogenic mechanisms. We conducted comparative genomics analysis of 39 A. veronii strains from different sources and found that 1993 core genes are shared by these strains and that these shared core genes may be necessary to maintain the basic characteristics of A. veronii. Additionally, phylogenetic relationship analysis based on these shared genes revealed that a distant relationship between the AMC34 strain and the other 38 strains but that, the genetic relationship among the 38 strains is relatively close, indicating that AMC34 may not belong to A. veronii. Furthermore, analysis of shared core genes and average nucleotide identity (ANI) values showed no obvious correlation with the location of A. veronii isolation and genetic relationship. Our research indicates the evolutionary mechanism of A. veronii from different sources and provides new insights for a deeper understanding of its pathogenic mechanism.

Bacillus subtilis BSH has a protective effect on Salmonella infection by regulating the intestinal flora structure in chickens.

Salmonellosis is a worldwide zoonotic disease that poses a serious threat to the reproduction of livestock and poultry and the health of young animals. Probiotics including Bacillus species, have received increasing attention as a substitute for antibiotics. In this study, chicks infected with Salmonella were fed feed supplemented with the BSH to observe the pathological changes in the liver, detect the number of viable bacteria in the liver and spleen, and record the death of the chicks. The results showed that BSH could reduce the pathological changes in the liver and the invasion of Salmonella into the liver and spleen of chicks. In addition, the survival rate of chicks in the BSH experimental group was 60%, while that in the infected control group was 26%, indicating that BSH had a protective effect on chicks infected with Salmonella. Finally, the fecal microflora of 9-day-old chicks was analyzed by 16S rRNA high-throughput sequencing. The results showed that Salmonella infection could cause intestinal flora changes, while BSH could alleviate this change. In addition, BSH also promoted the proliferation of Lactobacillus salivarius in the cecum of chick. This study emphasized that BSH has anti- Salmonella infection effects in chickens and can be used as a candidate microecological preparation strain.

Antitumour metastasis and the antiangiogenic and antitumour effects of a Eimeria stiedae soluble protein.

Some protozoa (Plasmodium falciparum, Toxoplasma gondii, etc) are used to treat cancer because they can improve tumour-induced immunosuppression. This study aims to evaluate the antitumour effect of Eimeria stiedae oocyst soluble protein (ESSP). ESSP was extracted, and mice were injected with 5 × 105 CT26 cells in the right axilla, and then, 50 µg of ESSP was intraperitoneally injected for 5 continuous days. The effect of ESSP on tumour immunity was detected by flow cytometry 25 days after the CT26 inoculation. The results showed that ESSP can inhibit the growth of CT26 subcutaneous tumours; significantly increase the expression of MHC I, MHC II, CD80 and CD86 on the surface of splenic dendritic cells; and enhance the level of IL-12 secretion. ESSP induced an increase in the number of NK cells in the mouse spleen, and the levels of IFN-γ and CD107 were upregulated in the NK cells and CD8+ T cells. The number of metastatic nodules in the lung tumours in the mice was significantly reduced, and the number of tubes, area of the loops and total length of the tubes were significantly reduced. ESSP enhances the antitumour immune response and inhibits tumour growth, metastasis and angiogenesis.

Comparative proteomic analysis reveals novel potential virulence factors of Aeromonas veronii.

Aeromonas veronii is an important zoonotic and aquatic pathogen. An increasing number of reports indicate that it has caused substantial economic losses in the aquaculture industry, in addition to threatening human health. However, little is known about its pathogenesis. Exploration of new virulence factors of A. veronii would be helpful for further understanding its pathogenesis. Hence, we comparatively analyzed the proteomes of virulent, attenuated, and avirulent strains of A. veronii using tandem mass tag (TMT) protein labeling and found numerous proteins either up- or downregulated in the virulent strain. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses showed that these differentially expressed proteins (DEPs) were involved mainly in pathways associated with bacterial chemotaxis and microbial metabolism in diverse environments. Furthermore, the expression levels of lysine decarboxylase, endoribonuclease, maltoporin, pullulanase, and aerolysin were positively correlated with the virulence of the strains, suggesting that their function may be closely related to the virulence of A. veronii. The results of qRT-PCR and multiple reaction monitoring for some DEPs were consistent with the results of TMT protein labeling. These results suggest that these DEPs may be novel potential virulence factors and will help to further understand the pathogenesis of A. veronii.

Comparative genomic analysis of different virulence strains reveals reasons for the increased virulence of Aeromonas veronii.

Aeromonas veronii is an important zoonotic and aquatic agent. More and more cases have shown that it has caused huge economic losses in the aquaculture industry in addition to threatening human health. But the reasons for the increasing virulence of A. veronii are still unclear. In order to further understand the reasons for the increased virulence of A. veronii, we conducted a comparative analysis of the genomes of A. veronii with different virulence. The analysis revealed that there are multiple virulence factors, such as those related to fimbriae, flagella, toxins, iron ion uptake systems and type II, type III and type VI secretion systems in the virulent strain TH0426 genome. And comparative analysis showed that there were two complete type III secretion systems (API1 and API2), of which the API2 and iron ion transport system were unique to the TH0426 strain. In addition, TH0426 strain also has unique functional gene clusters, which may play important roles in terms of resisting infection, adapting to different environments and genetic evolution. These particular virulence factors and gene clusters may be the important reasons for the increased virulence. These insights will provide a reference for the study of the pathogenesis of A. veronii.