Developing a Live Probiotic Vaccine Based on the Enterococcus faecium L3 Strain Expressing Influenza Neuraminidase

Probiotic microorganisms are currently considered as a promising platform for the development of recombinant vaccines expressing foreign antigens. In this study, we generated and evaluated the live mucosal recombinant vaccine by integrating genes encoding influenza virus neuraminidase (NA) of the N2 subtype into the DNA of the probiotic strain Enterococcus faecium L3 (L3). We confirmed NA expression in the pili of L3 using immune electron microscopy. Mice were fed with a probiotic vaccine containing the NA gene (L3-NA) or pure L3. Oral administration of L3-NA caused detectable increase in virus-specific serum IgG and local IgA after the third feeding. Immunization with L3-NA increased the survival rate by 34% when the mice were infected using A(H1N1)pdm09 influenza virus after the third feeding. After S. pneumoniae post-influenza infection, the L3-NA-immunized mice were 50% more protected from lethality in comparison with L3-fed mice. Thus, a live probiotic vaccine candidate based on L3 induced the formation of systemic and local immunity and provide partial protection against complicated influenza.

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Development of live vaccine based on the Enterococcus faecium L3 vector for prevention influenza infections

10.21203/rs.3.rs-749575/v1 ◽

2021 ◽

Author(s):

Yulia Desheva ◽

Galina Leontieva ◽

Tatiana Kramskaya ◽

Tatiana Gupalova ◽

Olga Kopteva ◽

...

Keyword(s):

Influenza Virus ◽

Enterococcus Faecium ◽

Influenza Infection ◽

Probiotic Strain ◽

Recombinant Vaccines ◽

Local Immunity ◽

Epidemic Season ◽

The Third ◽

Genes Encoding ◽

Influenza Virus Neuraminidase

Abstract Probiotic microorganisms are currently considered as a promising platform for the development of recombinant vaccines expressing foreign antigens. In this study we generated and evaluated the live mucosal recombinant vaccine by integrating genes encoding influenza virus neuraminidase (NA) of N2 subtype into the DNA of the probiotic strain Enterococcus faecium L3 (L3). We confirmed NA expression in the pili of L3 using immune electron microscopy. Mice were fed with a probiotic vaccine containing the NA gene (L3-NA) or pure L3. Oral administration of L3-NA caused detectable increase of virus-specific serum IgG and local IgA after the third feeding. A the same time, single spleen cell suspensions were stimulated with whole A(H1N1)pdm09 virus followed by flow cytometry. In mice received L3-NA, the content of cytotoxic T-lymphocytes was more pronounced compared to mice receiving pure L3. Immunization with L3-NA increased the survival rate by 34% when the mice were infected using A(H1N1)pdm09 influenza virus after the third feeding. After S. pneumonia post-influenza infection, the L3-NA immunized mice were 50% more protected from lethality in comparison with L3 fed mice. Thus, a live probiotic vaccine candidate based on L3 induced the formation of systemic and local immunity and provide protection against complicated influenza. The approach based on a probiotic vaccine expressing viral epitopes can allow repeated immunization during epidemic season.

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Infection of influenza virus neuraminidase-vaccinated mice with homologous influenza virus leads to strong protection against heterologous influenza viruses

Journal of General Virology ◽

10.1099/vir.0.067736-0 ◽

2014 ◽

Vol 95 (12) ◽

pp. 2627-2637 ◽

Cited By ~ 4

Author(s):

Biao He ◽

Haiyan Chang ◽

Zhihua Liu ◽

Chaoyang Huang ◽

Xueying Liu ◽

...

Keyword(s):

Influenza Virus ◽

Influenza Infection ◽

Lethal Dose ◽

Influenza Viruses ◽

Virus Challenge ◽

Influenza Pandemics ◽

Homologous Virus ◽

Major Antigen ◽

Influenza Virus Neuraminidase ◽

Antibody Levels

Vaccination is the best measure to prevent influenza pandemics. Here, we studied the protective effect against heterologous influenza viruses, including A/reassortant/NYMC X-179A (pH1N1), A/Chicken/Henan/12/2004 (H5N1), A/Chicken/Jiangsu/7/2002 (H9N2) and A/Guizhou/54/89×A/PR/8/34 (A/Guizhou-X) (H3N2), in mice first vaccinated with a DNA vaccine of haemagglutinin (HA) or neuraminidase (NA) of A/PR/8/34 (PR8) and then infected with the homologous virus. We showed that PR8 HA or NA vaccination both protected mice against a lethal dose of the homologous virus; PR8 HA or NA DNA vaccination and then PR8 infection in mice offered poor or excellent protection, respectively, against a second, heterologous influenza virus challenge. In addition, before the second heterologous influenza infection, the highest antibody level against nucleoprotein (NP) and matrix (M1 and M2) proteins was found in the PR8 NA-vaccinated and PR8-infected group. The level of induced cellular immunity against NP and M1 showed a trend consistent with that seen in antibody levels. However, PR8 HA+NA vaccination and then PR8 infection resulted in limited protection against heterologous influenza virus challenge. Results of the present study demonstrated that infection of the homologous influenza virus in mice already immunized with a NA vaccine could provide excellent protection against subsequent infection of a heterologous influenza virus. These findings suggested that NA, a major antigen of influenza virus, could be an important candidate antigen for universal influenza vaccines.

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A Cluster of Cuticle Protein Genes of Drosophila melanogaster at 65A: Sequence, Structure and Evolution

Genetics ◽

10.1093/genetics/147.3.1213 ◽

1997 ◽

Vol 147 (3) ◽

pp. 1213-1224

Author(s):

Jean-Philippe Charles ◽

Carol Chihara ◽

Shamim Nejad ◽

Lynn M Riddiford

Keyword(s):

Drosophila Melanogaster ◽

Genomic Dna ◽

Multiple Copy ◽

Sequence Structure ◽

Coding Regions ◽

The Third ◽

Genetic Complexity ◽

Genes Encoding ◽

Cuticle Protein ◽

Cuticle Proteins

A 36-kb genomic DNA segment of the Drosophila melanogaster genome containing 12 clustered cuticle genes has been mapped and partially sequenced. The cluster maps at 65A 5-6 on the left arm of the third chromosome, in agreement with the previously determined location of a putative cluster encompassing the genes for the third instar larval cuticle proteins LCP5, LCP6 and LCP8. This cluster is the largest cuticle gene cluster discovered to date and shows a number of surprising features that explain in part the genetic complexity of the LCP5, LCP6 and LCP8 loci. The genes encoding LCP5 and LCP8 are multiple copy genes and the presence of extensive similarity in their coding regions gives the first evidence for gene conversion in cuticle genes. In addition, five genes in the cluster are intronless. Four of these five have arisen by retroposition. The other genes in the cluster have a single intron located at an unusual location for insect cuticle genes.

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Performance of case definitions and clinical predictors for influenza surveillance among patients followed in a rural cohort in Senegal

BMC Infectious Diseases ◽

10.1186/s12879-020-05724-x ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Mamadou Aliou Barry ◽

Florent Arinal ◽

Cheikh Talla ◽

Boris Gildas Hedible ◽

Fatoumata Diene Sarr ◽

...

Keyword(s):

Influenza Virus ◽

Virus Infection ◽

Sore Throat ◽

Predictive Value ◽

Influenza Infection ◽

Clinical Signs ◽

Influenza Virus Infection ◽

Nasal Discharge ◽

Age Group ◽

Case Definitions

Abstract Background Influenza is a major cause of morbidity and mortality in Africa. However, a lack of epidemiological data remains for this pathology, and the performances of the influenza-like illness (ILI) case definitions used for sentinel surveillance have never been evaluated in Senegal. This study aimed to i) assess the performance of three different ILI case definitions, adopted by the WHO, USA-CDC (CDC) and European-CDC (ECDC) and ii) identify clinical factors associated with a positive diagnosis for Influenza in order to develop an algorithm fitted for the Senegalese context. Methods All 657 patients with a febrile pathological episode (FPE) between January 2013 and December 2016 were followed in a cohort study in two rural villages in Senegal, accounting for 1653 FPE observations with nasopharyngeal sampling and influenza virus screening by rRT-PCR. For each FPE, general characteristics and clinical signs presented by patients were collected. Sensitivity, Specificity, Positive Predictive Value (PPV) and Negative Predictive Value (NPV) for the three ILI case definitions were assessed using PCR result as the reference test. Associations between clinical signs and influenza infection were analyzed using logistic regression with generalized estimating equations. Sore throat, arthralgia or myalgia were missing for children under 5 years. Results WHO, CDC and ECDC case definitions had similar sensitivity (81.0%; 95%CI: 77.0–85.0) and NPV (91.0%; 95%CI: 89.0–93.1) while the WHO and CDC ILI case definitions had the highest specificity (52.0%; 95%CI: 49.1–54.5) and PPV (32.0%; 95%CI: 30.0–35.0). These performances varied by age groups. In children < 5 years, the significant predictors of influenza virus infection were cough and nasal discharge. In patients from 5 years, cough, nasal discharge, sore throat and asthenia grade 3 best predicted influenza infection. The addition of “nasal discharge” as a symptom to the WHO case definition decreased sensitivity but increased specificity, particularly in the pediatric population. Conclusion In summary, all three definitions studies (WHO, ECDC & CDC) have similar performance, even by age group. The revised WHO ILI definition could be chosen for surveillance purposes for its simplicity. Symptomatic predictors of influenza virus infection vary according the age group.

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Influenza vaccine failure in the tropics: a retrospective cohort study of waning effectiveness

Epidemiology and Infection ◽

10.1017/s0950268820002952 ◽

2020 ◽

Vol 148 ◽

Author(s):

B. E. Young ◽

T. M. Mak ◽

L. W. Ang ◽

S. Sadarangani ◽

H. J. Ho ◽

...

Keyword(s):

Influenza Virus ◽

Cohort Study ◽

Influenza Vaccine ◽

Retrospective Cohort Study ◽

Retrospective Cohort ◽

Influenza Infection ◽

Winter Season ◽

Temperate Climate ◽

Vaccine Failure ◽

Virus Activity

Abstract Influenza vaccine effectiveness (VE) wanes over the course of a temperate climate winter season but little data are available from tropical countries with year-round influenza virus activity. In Singapore, a retrospective cohort study of adults vaccinated from 2013 to 2017 was conducted. Influenza vaccine failure was defined as hospital admission with polymerase chain reaction-confirmed influenza infection 2–49 weeks after vaccination. Relative VE was calculated by splitting the follow-up period into 8-week episodes (Lexis expansion) and the odds of influenza infection in the first 8-week period after vaccination (weeks 2–9) compared with subsequent 8-week periods using multivariable logistic regression adjusting for patient factors and influenza virus activity. Records of 19 298 influenza vaccinations were analysed with 617 (3.2%) influenza infections. Relative VE was stable for the first 26 weeks post-vaccination, but then declined for all three influenza types/subtypes to 69% at weeks 42–49 (95% confidence interval (CI) 52–92%, P = 0.011). VE declined fastest in older adults, in individuals with chronic pulmonary disease and in those who had been previously vaccinated within the last 2 years. Vaccine failure was significantly associated with a change in recommended vaccine strains between vaccination and observation period (adjusted odds ratio 1.26, 95% CI 1.06–1.50, P = 0.010).

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Combined administration of bacteriocin- -producing, probiotic strain Enterococcus faecium CCM7420 with Eleutherococcus senticosus and their effect in rabbits

Polish Journal of Veterinary Sciences ◽

10.2478/pjvs-2013-0089 ◽

2013 ◽

Vol 16 (4) ◽

pp. 619-627 ◽

Cited By ~ 9

Author(s):

M. Pogány Simonová ◽

A. Lauková ◽

L. Chrastinová ◽

I. Plachá ◽

V. Strompfová ◽

...

Keyword(s):

Growth Performance ◽

Enterococcus Faecium ◽

Average Daily Gain ◽

Probiotic Strain ◽

Cholesterol Concentration ◽

Control Group ◽

Feed Conversion ◽

Eleutherococcus Senticosus ◽

Commercial Diet ◽

And Control

Abstract The effect of Enterococcus faecium CCM7420 (EF) - enterocin-producing and probiotic strain of rabbit origin, Eleutherococcus senticosus extract (ES) and their combination (ES+EF) was determined on selected bacteria in faeces and caecum content, leukocytes phagocytosis, blood biochemistry and growth performance. Ninety-six weaned rabbits were divided into 3 experimental (ES, EF, ES+EF) and control group (CG). The rabbits in the groups ES and EF+ES were fed commercial diet enriched with E. senticosus extract (30 g/100 kg feed), rabbits in groups EF and CG were fed untreated diet. The rabbits in the EF and ES+EF groups were administered with an overnight culture of E. faecium CCM7420 strain (500 μl/animal/day into water, 109 CFU/ml). The treatment period lasted 21 days. The microbiological examinations in faecal samples confirmed the presence of E. faecium CCM7420 strain. In groups EF and ES+EF, the reduction of faecal coliforms, Pseudomonas-like sp., Clostridium-like sp. and S. aureus was recorded. Leucocyte phagocytosis significantly increased in all experimental groups (P<0.0001) compared to CG. The lowest GPx values were measured in the ES+EF group. Higher total protein, triglycerides and calcium concentrations were detected in experimental groups compared to CG. The cholesterol concentration decreased in the ES group. The highest average daily gain was recorded in EF group; in ES+EF the better feed conversion ratio and no mortality was recorded. These results indicated that the dietary supplementation with the E. faecium CCM7420 and E. senticosus extract stimulate the leukocytes phagocytosis and reduces the potential pathogens in rabbits digestive tract without oxidative stress and improve the growth performance.

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Construction of a weight-based seed sorting system for the third-generation hybrid rice

Rice ◽

10.1186/s12284-021-00510-y ◽

2021 ◽

Vol 14 (1) ◽

Author(s):

Jianxin Wu ◽

Shijun Qiu ◽

Menglong Wang ◽

Chunjue Xu ◽

Xing Wang Deng ◽

...

Keyword(s):

Hybrid Rice ◽

Pollen Grains ◽

Third Generation ◽

Male Sterile ◽

Specific Expression ◽

The Third ◽

Genes Encoding ◽

Wild Type Male ◽

Sorting System ◽

Generation Hybrid

Abstract Background The third-generation hybrid rice technology can be constructed by transforming a recessive nuclear male sterile (NMS) mutant with a transgenic cassette containing three functional modules: the wild type male fertility gene to restore the fertility of the mutant, the pollen killer gene that specifically kills the pollen grains carrying the transgene, and the red fluorescence protein (RFP) gene to mark the transgenic seed (maintainer). The transgenic plant produces 1:1 NMS seeds and maintainer seeds that can be distinguished by the RFP signal. However, the RFP signals in the partially filled or pathogen-infected maintainer seeds are often too weak to be detected by RFP-based seed sorting machine, resulting in intermingling of the maintainer seeds with NMS seeds. Results Here we constructed a weight-based seed sorting system for the third-generation hybrid rice technology by silencing the genes encoding ADP-glucose pyrophosphorylase (AGP) essential for endosperm starch biosynthesis via endosperm-specific expression of artificial microRNAs (amiRNAs). In this system, the NMS seeds have normal endosperm and are heavy, but the maintainer seeds have shrunken endosperms and are light-weighted. The maintainer seeds can be easily and accurately sorted out from the NMS seeds by weight-sorting machines, so pure and fully filled NMS seeds are available. Conclusions The weight-based seed sorting system shows obvious advantages over the RFP-based seed sorting system in accuracy, efficiency, and cost for propagation of pure male sterile seeds. These characteristics will significantly increase the value and transgenic safety of the third-generation hybrid rice technology.

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Docking and 3D QSAR study of thiourea analogs as potent inhibitors of influenza virus neuraminidase

Journal of Molecular Modeling ◽

10.1007/s00894-010-0685-9 ◽

2010 ◽

Vol 16 (12) ◽

pp. 1809-1818 ◽

Cited By ~ 12

Author(s):

Jiaying Sun ◽

Shaoxi Cai ◽

Hu Mei ◽

Jian Li ◽

Ning Yan ◽

...

Keyword(s):

Influenza Virus ◽

3D Qsar ◽

Qsar Study ◽

Influenza Virus Neuraminidase

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Anti-adhesion of probiotic Enterococcus faecium WEFA23 against five pathogens and the beneficial effect of its S-layer proteins against Listeria monocytogenes

Canadian Journal of Microbiology ◽

10.1139/cjm-2018-0031 ◽

2019 ◽

Vol 65 (3) ◽

pp. 175-184

Author(s):

Yao He ◽

Xiongpeng Xu ◽

Fen Zhang ◽

Di Xu ◽

Zhengqi Liu ◽

...

Keyword(s):

Listeria Monocytogenes ◽

Enterococcus Faecium ◽

Probiotic Strain ◽

Antagonistic Activity ◽

Bacterial Surface ◽

Colony Forming Units ◽

Infant Feces ◽

Adhesion Activity ◽

Displacement Assays ◽

Different Levels

Enterococcus faecium WEFA23 is a potential probiotic strain isolated from Chinese infant feces. In this study, the antagonistic activity of E. faecium WEFA23 on adhesion to pathogens was investigated. Enterococcus faecium WEFA23 was able to compete, exclude, and displace the adhesion of Escherichia coli O157:H7, Salmonella Typhimurium ATCC 13311, Listeria monocytogenes CMCC54007, Staphylococcus aureus CMCC26003, and Shigella sonnei ATCC 25931 to Caco-2 cells. Among them, L. monocytogenes achieved the strongest inhibition rate in both competition and displacement assays. Those anti-adhesion capacities were related to the bacterial physicochemical properties (hydrophobicity, auto-aggregation, and co-aggregation) of the bacterial surface. For L. monocytogenes, the anti-adhesion capacity was affected by the heat treatment, cell density, and growth phase of E. faecium WEFA23; 108 colony-forming units of viable cells per millilitre at the stationary phase exhibited the strongest anti-adhesion activity. In addition, removal of S-layer proteins of E. faecium WEFA23 by treatment with 5 mol/L LiCl significantly decreased its adhesion capacity, and those S-layer proteins were able to compete, displace, and exclude L. monocytogenes at different levels. Both cells and S-layer proteins of E. faecium WEFA23 significantly reduced the apoptosis of Caco-2 cells induced by L. monocytogenes, which was mediated by caspase-3 activation. This study might be helpful in understanding the anti-adhesion mechanism of probiotics against pathogens.

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