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Abstract BackgroundAs the most malignant type of cancer in the female reproductive system, ovarian cancer (OC) has become the second leading cause of death among Chinese women. Chemotherapy is the main treatment for patients with OC, and its numerous adverse effects can easily lead to malnutrition. It is difficult to centrally manage patients with OC in the intervals between chemotherapy. The use of WeChat, an effective mobile tool, in chronic disease management has been highlighted. ObjectiveThis study aimed to implement a continuous follow-up strategy and health monitoring based on the WeChat platform for patients with OC undergoing chemotherapy to ensure that each phase of chemotherapy was delivered on schedule and to improve the survival rate of patients with OC. MethodsParticipants were recruited and randomly assigned to either the WeChat-based nutrition intervention group or the usual care group. A self-administered general information questionnaire was used at enrollment to obtain basic information about the patients. The Patient-Generated Subjective Global Assessment (PG-SGA) Scale was used to investigate the nutritional status of the patients at 3 time points (T0=before the first admission to the hospital for chemotherapy, T1=2 weeks after the first chemotherapy, and T6=2 weeks after the sixth chemotherapy). The blood indices of patients were investigated through the inhospital health care system at 3 times（T0=before the first admission to the hospital for chemotherapy, T1=2 weeks after the first chemotherapy, and T6=2 weeks after the sixth chemotherapy). Patients in the intervention group were introduced to the nutrition applet, invited to join the nutrition management group chat, and allowed to consult on nutritional issues in private chats with nutrition management team members. Linear mixed models were used to analyze changes in each nutritional indicator in the 2 groups, with their baseline measurements as covariates; with group, time, and group-time interactions considered as fixed effects; and with patients considered as random effects. ResultsA total of 96 patients with OC undergoing chemotherapy were recruited into the study. Distribution was based on a 1:1 ratio, with 48 patients each in the nutrition intervention group and the usual care group. The attrition rate after the first chemotherapy session was 18.75%. The mixed linear model revealed that the group-based effect and the group-time interaction effect on PG-SGA scores were significant (F38,38PF37,37PF38,38PF38,38PF38,38PF37,37PF37,38PF38,38PF37,37P ConclusionsThis study provided evidence that a WeChat-based, multidisciplinary, full-course nutritional management program can significantly improve the nutritional status of patients with OC during chemotherapy.

Abstract Vaccination is still the most promising strategy for combating influenza virus pandemics. However, the highly variable characteristics of influenza virus make it difficult to develop antibody-based universal vaccines, until now. Lung tissue-resident memory T cells (TRM), which actively survey tissues for signs of infection and react rapidly to eliminate infected cells without the need for a systemic immune reaction, have recently drawn increasing attention towards the development of a universal influenza vaccine. We previously designed a sequential immunization strategy based on orally administered Salmonella vectored vaccine candidates. To further improve our vaccine design, in this study, we used two different dendritic cell (DC)-targeting strategies, including a single chain variable fragment (scFv) targeting the surface marker DC-CD11c and DC targeting peptide 3 (DCpep3). Oral immunization with Salmonella harboring plasmid pYL230 (S230), which displayed scFv-CD11c on the bacterial surface, induced dramatic production of spleen effector memory T cells (TEM). On the other hand, intranasal boost immunization using purified DCpep3-decorated 3M2e-ferritin nanoparticles in mice orally immunized twice with S230 (S230inDC) significantly stimulated the differentiation of lung CD11b+ DCs, increased intracellular IL-17 production in lung CD4+ T cells and elevated chemokine production in lung sections, such as CXCL13 and CXCL15, as determined by RNAseq and qRT‒PCR assays, resulting in significantly increased percentages of lung TRMs, which could provide efficient protection against influenza virus challenge. The dual DC targeting strategy, together with the sequential immunization approach described in this study, provides us with a novel “prime and pull” strategy for addressing the production of protective TRM cells in vaccine design.

African swine fever (ASF) is an acute infectious disease with a high mortality rate in both domestic and wild boars. Commercial vaccines or antiviral drugs for ASF were not available due to the complex diversity of the structure and genome of its pathogen African swine fever virus (ASFV). In recent years, there have been many reports on candidate strains of attenuated vaccines for ASFV. In this study, we obtained a recombinant virus named SY18ΔL60LΔCD2v by simultaneously deleting the L60L gene and CD2v gene from highly virulent strain SY18. In vitro, SY18ΔL60LΔCD2v displayed a decreased growth kinetic compared to that of parental SY18. In vivo, high doses (105 TCID50) of SY18ΔL60LΔCD2v can protect pigs (5/5) from attacks by the parental SY18 strain (102 TCID50). Low doses (102 TCID50) of SY18ΔL60LΔCD2v only protected 20% of pigs (1/5) from attacks by the parental SY18 strain (102 TCID50). The results indicated that the absence of these two genes in SY18 could induce protection against the homologous parental strain, and there were no obvious clinical symptoms or viremia. These results indicate that the SY18ΔL60LΔCD2v strain can serve as a new live attenuated vaccine candidate for the prevention and control of ASFV infection.

IntroductionAfrican swine fever (ASF) is an acute and highly contagious disease and its pathogen, the African swine fever virus (ASFV), threatens the global pig industry. At present, management of ASF epidemic mainly relies on biological prevention and control methods. Moreover, due to the large genome of ASFV, only half of its genes have been characterized in terms of function.MethodsHere, we evaluated a previously uncharacterized viral gene, L60L. To assess the function of this gene, we constructed a deletion strain (SY18ΔL60L) by knocking out the L60L gene of the SY18 strain. To evaluate the growth characteristics and safety of the SY18ΔL60L, experiments were conducted on primary macrophages and pigs, respectively.ResultsThe results revealed that the growth trend of the recombinant strain was slower than that of the parent strain in vitro. Additionally, 3/5 (60%) pigs intramuscularly immunized with a 105 50% tissue culture infectious dose (TCID50) of SY18ΔL60L survived the 21-day observation period. The surviving pigs were able to protect against the homologous lethal strain SY18 and survive. Importantly, there were no obvious clinical symptoms or viremia.DiscussionThese results suggest that L60L could serve as a virulence- and replication-related gene. Moreover, the SY18ΔL60L strain represents a new recombinant live-attenuated ASFV that can be employed in the development of additional candidate vaccine strains and in the elucidation of the mechanisms associated with ASF infection.

ABSTRACT The influenza virus continues to pose a great threat to public health due to the frequent variations in RNA viruses. Vaccines targeting conserved epitopes, such as the extracellular domain of the transmembrane protein M2 (M2e), a nucleoprotein, and the stem region of hemagglutinin proteins, have been developed, but more efficient strategies, such as nanoparticle-based vaccines, are still urgently needed. However, the labor-intensive in vitro purification of nanoparticles is still necessary, which could hinder the application of nanoparticles in the veterinary field in the future. To overcome this limitation, we used regulated lysis Salmonella as an oral vector with which to deliver three copies of M2e (3M2e-H1N1)-ferritin nanoparticles in situ and evaluated the immune response. Then, sequential immunization using Salmonella-delivered nanoparticles followed by an intranasal boost with purified nanoparticles was performed to further improve the efficiency. Compared with 3M2e monomer administration, Salmonella-delivered in situ nanoparticles significantly increased the cellular immune response. Additionally, the results of sequential immunization showed that the intranasal boost with purified nanoparticles dramatically stimulated the activation of lung CD11b dendritic cells (DCs) and elevated the levels of effector memory T (TEM) cells in both spleen and lung tissues as well as those of CD4 and CD8 tissue-resident memory T (TRM) cells in the lungs. The increased production of mucosal IgG and IgA antibody titers was also observed, resulting in further improvements to protection against a virus challenge, compared with the pure oral immunization group. Salmonella-delivered in situ nanoparticles efficiently increased the cellular immune response, compared with the monomer, and sequential immunization further improved the systemic immune response, as shown by the activation of DCs, the production of TEM cells and TRM cells, and the mucosal immune response, thereby providing us with a novel strategy by which to apply nanoparticle-based vaccines in the future. IMPORTANCE Salmonella-delivered in situ nanoparticle platforms may provide novel nanoparticle vaccines for oral administration, which would be beneficial for veterinary applications. The combination of administering Salmonella-vectored, self-assembled nanoparticles and an intranasal boost with purified nanoparticles significantly increased the production of effector memory T cells and lung resident memory T cells, thereby providing partial protection against an influenza virus challenge. This novel strategy could open a novel avenue for the application of nanoparticle vaccines for veterinary purposes.

Abstract Aeromonas veronii (A. veronii) is a pathogenic that can infect human, animal and aquatic organisms, in which poses a huge threat to the health of many aquatic organisms such as Cyprinus carpio. In this study, Lactobacillus casei (L. casei) strain CC16 was used as antigen deliver carrier and fused with cholera toxin B subunit (CTB) as an adjuvant to construct the recombinant L. casei pPG-Aha1/Lc CC16(surface-displayed) and pPG-Aha1-CTB/Lc CC16(surface-displayed) expressing Aha1 protein of A. veronii, respectively. And the immune responses in Cyprinus carpio by oral route was explored. Our results demonstrated that the recombinant strains could stimulate high serum specific antibody immunoglobulin M (IgM) and induce a stronger acid phosphatase (ACP), alkaline phosphatase (AKP), C3, C4, lysozyme (LZM), Lectin and superoxide dismutase (SOD) activity in Cyprinus carpio compared with control groups. Meanwhile, the expression of Interleukin-10 (IL-10), Interleukin-1β (IL-1β), Tumor Necrosis Factor-α (TNF-α), immunoglobulin Z1 (IgZ1) and immunoglobulin Z2 (IgZ2) in the tissues were significantly upregulated compared with Lc-pPG or PBS groups, indicating that humoral and cell immune response were triggered. Additionally, recombinant L. casei could survive and colonize in fish intestine. Significantly, recombinant L. casei provides immune protection against A. veronii infection, which Cyprinus carpio received pPG-Aha1-CTB/Lc CC16 (64.29%) and pPG-Aha1/Lc CC16 (53.57%) had higher survival rates compared with the controls. Thus, we demonstrated that recombinant pPG-Aha1/Lc CC16 and pPG-Aha1-CTB/Lc CC16 may be the promising strategy for the development of an oral vaccine against A. veronii.

(1) Background: African swine fever (ASF) is a highly contagious disease that causes high pig mortality. Due to the absence of vaccines, prevention and control are relatively challenging. The pathogenic African swine fever virus (ASFV) has a complex structure and encodes over 160 proteins, many of which still need to be studied and verified for their functions. In this study, we identified one of the unknown functional genes, C84L. (2) Methods: A gene deficient strain was obtained through homologous recombination and several rounds of purification, and its replication characteristics and virulence were studied through in vitro and in vivo experiments, respectively. (3) Results: Deleting this gene from the wild-type virulent strain SY18 did not affect its replication in porcine primary macrophages but reduced its virulence in pigs. In animal experiments, we injected pigs with a 102 TCID50, 105 TCID50 deletion virus, and a 102 TCID50 wild-type strain SY18 intramuscularly. The control group pigs reached the humane endpoint on the ninth day (0/5) and were euthanized. Two pigs in the 102 TCID50(2/5) deletion virus group survived on the twenty-first day, and one in the 105 TCID50(1/5) deletion virus group survived. On the twenty-first day, the surviving pigs were euthanized, which was the end of the experiment. The necropsies of the survival group and control groups’ necropsies showed that the surviving pigs’ liver, spleen, lungs, kidneys, and submaxillary lymph nodes did not show significant lesions associated with the ASFV. ASFV-specific antibodies were first detected on the seventh day after immunization; (4) Conclusions: This is the first study to complete the replication and virulence functional exploration of the C84L gene of SY18. In this study, C84L gene was preliminarily found not a necessary gene for replication, gene deletion strain SY18ΔC84L has similar growth characteristics to SY18 in porcine primary alveolar macrophages. The C84L gene affects the virulence of the SY18 strain.

Phage therapy is an alternative approach to overcome the problem of multidrug resistance in bacteria. In this study, a bacteriophage named PZL-Ah152, which infects Aeromonas hydrophila, was isolated from sewage, and its biological characteristics and genome were studied. The genome contained 54 putative coding sequences and lacked known putative virulence factors, so it could be applied to phage therapy. Therefore, we performed a study to (i) investigate the efficacy of PZL-Ah152 in reducing the abundance of pathogenic A. hydrophila strain 152 in experimentally infected crucian carps, (ii) evaluate the safety of 12 consecutive days of intraperitoneal phage injection in crucian carps, and (iii) determine how bacteriophages impact the normal gut microbiota. The in vivo and in vitro results indicated that the phage could effectively eliminate A. hydrophila. Administering PZL-Ah152 (2 × 109 PFU) could effectively protect the fish (2 × 108 CFU/carp). Furthermore, a 12-day consecutive injection of PZL-Ah152 did not cause significant adverse effects in the main organs of the treated animals. We also found that members of the genus Aeromonas could enter and colonize the gut. The phage PZL-Ah152 reduced the number of colonies of the genus Aeromonas. However, no significant changes were observed in α-diversity and β-diversity parameters, which suggested that the consumed phage had little effect on the gut microbiota. All the results illustrated that PZL-Ah152 could be a new therapeutic method for infections caused by A. hydrophila.

Abstract Background The inner membrane protein DotU of Aeromonas veronii is an important component of the minimal core conserved membrane proteome required for the formation of an envelope-transmembrane complex. This protein functions in a type VI secretion system (T6SS), and the role of this T6SS during the pathogenic process has not been clearly described. Results A recombinant A. veronii with a partial disruption of the dotU gene (720 bp of the in-frame sequence) (defined as ∆dotU) was constructed by two conjugate exchanges. We found that the mutant ∆dotU allele can be stably inherited for more than 50 generations. Inactivation of the A. veronii dotU gene resulted in no significant changes in growth or resistance to various environmental changes. However, compared with the wild-type strain colony, the mutant ∆dotU colony had a rough surface morphology. In addition, the biofilm formation ability of the mutant ∆dotU was significantly enhanced by 2.1-fold. Conversely, the deletion of the dotU gene resulted in a significant decrease in pathogenicity and infectivity compared to those of the A. veronii wild-type strain. Conclusions Our findings indicated that the dotU gene was an essential participant in the pathogenicity and invasiveness of A. veronii TH0426, which provides a novel perspective on the pathogenesis of TH0426 and lays the foundation for discovering potential T6SS effectors.

Citrobacter freundii refers to a fish pathogen extensively reported to be able to cause injury and high mortality. Phage therapy is considered a process to alternatively control bacterial infections and contaminations. In the present study, the isolation of a virulent bacteriophage IME-JL8 isolated from sewage was presented, and such bacteriophage was characterized to be able to infect Citrobacter freundii specifically. Phage IME-JL8 has been classified as the member of the Siphoviridae family, which exhibits the latent period of 30–40 min. The pH and thermal stability of phage IME-JL8 demonstrated that this bacteriophage achieved a pH range of 4–10 as well as a temperature range of 4, 25, and 37°C. As revealed from the results of whole genomic sequence analysis, IME-JL8 covers a double-stranded genome of 49,838 bp (exhibiting 47.96% G+C content), with 80 putative coding sequences contained. No bacterial virulence- or lysogenesis-related ORF was identified in the IME-JL8 genome, so it could be applicable to phage therapy. As indicated by the in vitro experiments, phage IME-JL8 is capable of effectively removing bacteria (the colony count decreased by 6.8 log units at 20 min), and biofilm can be formed in 24 h. According to the in vivo experiments, administrating IME-JL8 (1 × 107 PFU) was demonstrated to effectively protect the fish exhibiting a double median lethal dose (2 × 109 CFU/carp). Moreover, the phage treatment led to the decline of pro-inflammatory cytokines in carp with lethal infections. IME-JL8 was reported to induce efficient lysis of Citrobacter freundii both in vitro and in vivo, thereby demonstrating its potential as an alternative treatment strategy for infections attributed to Citrobacter freundii.

Abstract Background Porcine circovirus type 1 (PCV1) was discovered in 1974 as a contaminant of a porcine kidney (PK-15) cell line and was generally accepted to be nonpathogenic. But recently it was shown to cause lesions in experimentally infected pig fetuses. Serological evidence and genetic studies suggested that PCV1 was widespread in domestic pigs. Thus, the molecular epidemiology and genetic variation of PCV1 are still necessary to understand. Results Here 247 tissue samples were collected from piglets in Guangxi Province, China and performed whole-genome sequencing of the PCV1 genome. Thirteen PCV1 strains were sequenced from the samples. Similarity analysis showed that there were 97.8% to 99.6% nucleotide similarity to each other and 97.1% to 99.8% nucleotide similarity to the 40 reference strains. Besides, based on sequence analysis, we found one putative recombinant virus named GXdx84 strain contained the open-reading frame 1 (ORF1) of PCV1 and the ORF2 of PCV2d-2, which was consistent with the results of phylogenetic analysis that compared PCV1 and PCV2 strains. Variation analysis of the amino acids of the capsid protein revealed that the GXyl224 strain, which encoded 235 amino acids, had two amino acids more than other strains. This is the first study to report that a cap gene mutation resulted in lengthening of in the gene sequence. Conclusions These data contribute to the understanding of PCV1 evolution and molecular epidemiology that will facilitate programs for its control and prevention.

The Cre-recombinase mediated in vivo minicircle DNA vaccine platform (CRIM) provided a novel option to replace a traditional DNA vaccine. To further improve the immune response of our CRIM vaccine, we designed a dual promoter expression plasmid named pYL87 which could synthesize short HN protein under a prokaryotic in vivo promoter PpagC and full length HN protein of genotype VII Newcastle disease virus (NDV) under the previous eukaryotic CMV promoter at the same time. Making use of the self-lysed Salmonella strain as a delivery vesicle, chickens immunized with the pYL87 construction showed an increased serum haemagglutination inhibition antibody response, as well as an increased cell proliferation level and cellular IL-4 and IL-18 cytokines, compared with the previous CRIM vector pYL47. After the virus challenge, the pYL87 vector could provide 80% protection compared to 50% protection against genotype VII NDV in pYL47 immunized chickens, indicating a promising dual promoter strategy used in vaccine design.

Aeromonas veronii is an important pathogen causing freshwater fish sepsis and ulcer syndrome. An increasing number of cases have demonstrated its significance as an aquatic zoonotic agent. The purpose of this study was to ensure the safety of freshwater products by evaluating the infection status of edible freshwater fish. In this experiment, we isolated A. veronii from several species of apparently healthy freshwater fish, including Carassius auratus, Cyprinus carpio, Ctenopharyngodon idella, and Silurus asotus. A. veronii was identified through bacterial staining, culture characteristics, and 16S rDNA gene sequence. In addition, polymerase chain reaction (PCR) was used to investigate the distribution of seven major virulence genes, including aerolysin (aer: 88.51%), cytotoxic enterotoxin (act: 71.26%), serine proteinase (ser: 54.02%), adhesin (Aha: 40.23%), phospholipase (lip: 45.98%), nuclease (exu: 51.72%), and quorum sensing-controlled virulence factor (LuxS: 59.77%). In total, 496 strains of Aeromonas were isolated, including 87 strains of A. veronii. The isolates of A. veronii were Gram-negative, rod-shaped bacteria, and the colonies are yellow on Rimler-Shotts (RS) medium and showed greater than 99% homology with A. veronii ATCC35624 according to analyses of the 16S rDNA sequence. Nearly 50% of the A. veronii isolates carried at least four or more virulence genes, 25% of the isolates carried at least five types of virulence genes, and 59.77% isolates carried the LuxS gene, and the isolates carrying more virulence genes were found to be more virulent. These results are of great significance for further improving the food safety assessment of freshwater aquatic products.

With some advances in modern medicine (such as cancer chemotherapy, broad exposure to antibiotics, and immunosuppression), the incidence of opportunistic fungal pathogens such as Candida albicans has increased. Cases of drug resistance among these pathogens have become more frequent, requiring the development of new drugs and a better understanding of the targeted enzymes. Sterol 14α-demethylase (CYP51) is a cytochrome P450 enzyme required for biosynthesis of sterols in eukaryotic cells and is the major target of clinical drugs for managing fungal pathogens, but some of the CYP51 key features important for rational drug design have remained obscure. We report the catalytic properties, ligand-binding profiles, and inhibition of enzymatic activity of C. albicans CYP51 by clinical antifungal drugs that are used systemically (fluconazole, voriconazole, ketoconazole, itraconazole, and posaconazole) and topically (miconazole and clotrimazole) and by a tetrazolebased drug candidate, VT-1161 (oteseconazole: (R)-2-(2,4-difluorophenyl)-1,1-difluoro-3-(1H-tetrazol-1-yl)-1-(5-(4-(2,2,2-trifluoroethoxy)phenyl)pyridin-2-yl)propan-2-ol). Among the compounds tested, the first-line drug fluconazole was the weakest inhibitor, whereas posaconazole and VT-1161 were the strongest CYP51 inhibitors. We determined the X-ray structures of C. albicans CYP51 complexes with posaconazole and VT-1161, providing a molecular mechanism for the potencies of these drugs, including the activity of VT-1161 against Candida krusei and Candida glabrata, pathogens that are intrinsically resistant to fluconazole. Our comparative structural analysis outlines phylum-specific CYP51 features that could direct future rational development of more efficient broadspectrum antifungals. [ABSTRACT FROM AUTHOR]