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1.
Arch Virol ; 164(11): 2819-2822, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31482204

RESUMO

A recent study by Ghosh et al. compared the gut microbiomes of 20 preschool children from India and found an association between the gut microbiome and the nutritional status of the child. Here, we explored these metagenomes for the presence of genomic signatures of prokaryotic and eukaryotic viruses. Several of the viral signatures found in all 20 metagenomes belonged to giant viruses (GVs). In addition, we found hits for bacteriophages to several major human pathogens, including Shigella, Salmonella, Escherichia, and Enterobacter. Concurrently, we also detected several antibiotic resistance genes (ARGs) in the metagenomes. All of the ARGs detected in this study (beta-lactam, macrolide, metronidazole, and tetracycline) are associated with mobile genetic elements (MGEs) and have been reported to cause high levels of resistance to their respective antibiotics. Despite recent reports of giant viruses and their genomic signatures in gut microbiota, their role in human physiology remains poorly understood. The effect of cooccurrence of ARGs and GVs in the gut needs further investigation.


Assuntos
Bacteriófagos/genética , Microbioma Gastrointestinal/genética , Genoma Viral/genética , Vírus Gigantes/genética , Metagenoma/genética , Pré-Escolar , Resistência Microbiana a Medicamentos/genética , Enterobacter/genética , Escherichia/virologia , Vírus Gigantes/isolamento & purificação , Humanos , Índia , Sequências Repetitivas Dispersas/genética , Salmonella/virologia , Shigella/virologia
2.
Arch Virol ; 164(12): 3115-3119, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31535209

RESUMO

A virulent phage, named ST20, infecting Escherichia coli O165:H8 was isolated from wastewater and subjected to genomic sequencing using the Illumina HiSeq system. Genomic analysis revealed that it contains double-stranded DNA, and its complete genome consists of 44,517 nucleotides with an average GC content of 50.81%. Morphological observations showed that phage ST20 belongs to the order Caudovirales and the family Siphoviridae due to its characteristic icosahedral capsid and a long noncontractile tail. This phage was further characterized by one-step growth curve analysis and measurement of its stability at 4 °C. The study has implications for the development of potential biocontrol agents.


Assuntos
Escherichia coli Shiga Toxigênica/virologia , Siphoviridae/classificação , Sequenciamento Completo do Genoma/métodos , Bacteriófagos/classificação , Bacteriófagos/genética , Bacteriófagos/isolamento & purificação , Bacteriófagos/patogenicidade , Composição de Bases , Tamanho do Genoma , Siphoviridae/genética , Siphoviridae/isolamento & purificação , Siphoviridae/patogenicidade , Virulência , Águas Residuárias/microbiologia
3.
Arch Virol ; 164(10): 2627-2630, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31363923

RESUMO

A lytic bacteriophage, designated Vibrio phage vB_VpP_BA6, was isolated from sewage collected in Guangzhou, China. The double-stranded DNA genome of phage BA6 is composed of 50,520 bp with a G+C content of 41.77%. It possesses 64 open reading frames relating to phage structure, packaging, host lysis, DNA metabolism, and additional functions. Three tRNAs genes (encoding Pro, Ile and Trp) were detected. Comparison of its genomic features and phylogenetic analysis revealed that phage BA6 is a novel member of the family Podoviridae. This phage may represent a potential therapeutic agent against multidrug-resistant Vibrio parahaemolyticus.


Assuntos
Bacteriófagos/genética , Bacteriófagos/isolamento & purificação , Genoma Viral , Podoviridae/genética , Podoviridae/isolamento & purificação , Vibrio parahaemolyticus/virologia , Bacteriólise , Bacteriófagos/classificação , Bacteriófagos/crescimento & desenvolvimento , Composição de Bases , China , DNA/química , DNA/genética , Fases de Leitura Aberta , Filogenia , Podoviridae/classificação , Podoviridae/crescimento & desenvolvimento , RNA de Transferência/genética , Esgotos/virologia
4.
Arch Virol ; 164(10): 2637-2640, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31372754

RESUMO

A novel lytic Raoultella phage, RP180, was isolated and characterized. The RP180 genome has 44,851 base pairs and contains 65 putative genes, 35 of them encoding proteins whose functions were predicted based on sequence similarity to known proteins. The RP180 genome possesses a gene synteny typical of members of the subfamily Guernseyvirinae. Phylogenetic analysis of the RP180 genome and similar phage genomes revealed that phage RP180 is the first member of the genus Kagunavirus, subfamily Guernseyvirinae, that is specific for Raoultella sp. The genome of RP180 encodes a putative protein with similarity to CRISPR-like Cas4 nucleases, which belong to the pfam12705/PDDEXK_1 family. Cas4-like proteins of this family have been shown to interfere with the bacterial host type II-C CRISPR-Cas system.


Assuntos
Bacteriófagos/classificação , Bacteriófagos/isolamento & purificação , Enterobacteriaceae/virologia , Filogenia , Siphoviridae/classificação , Siphoviridae/isolamento & purificação , Bacteriólise , Bacteriófagos/genética , Genoma Viral , Microscopia Eletrônica de Transmissão , Análise de Sequência de DNA , Siphoviridae/genética , Sintenia , Proteínas Virais/genética , Vírion/ultraestrutura
5.
J Agric Food Chem ; 67(40): 11219-11229, 2019 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-31408330

RESUMO

Peanut allergy is a major health problem worldwide. Detection of food allergens is a critical aspect of food safety. The VHH domain of single chain antibody from camelids, also known as nanobody (Nb), showed its advantages in the development of biosensors because of its high stability, small molecular size, and ease of production. However, no nanobody specific to peanut allergens has been developed. In this study, we constructed a library with random triplets (NNK) in its CDR regions of a camel nanobody backbone. We screened the library with peanut allergy Ara h 3 and obtained several candidate nanobodies. One of the promising nanobodies, Nb16 was further biochemical characterization by gel filtration, isothermal titration calorimetry (ITC), cocrystallization, and Western blot in terms of its interaction with Ara h 3. Nb16 specifically binds to peanut major allergen Ara h 3 with a dissociation constant of 400 nM. Furthermore, we obtained the Ara h 3-Nb16 complex crystals. Structure analysis shows the packing mode is completely different between the Ara h 3-Nb16 complex crystal and the native Ara h 3 crystal. Structural determination of Ara h 3-Nb16 will provide the necessary information to understand the allergenicity of this important peanut allergen. The nanobody Nb16 may have application in the development of biosensors for peanut allergen detection.


Assuntos
Antígenos de Plantas/imunologia , Arachis/imunologia , Proteínas de Armazenamento de Sementes/imunologia , Anticorpos de Domínio Único/genética , Anticorpos de Domínio Único/imunologia , Sequência de Aminoácidos , Antígenos de Plantas/química , Antígenos de Plantas/genética , Arachis/química , Arachis/genética , Bacteriófagos/genética , Bacteriófagos/metabolismo , Western Blotting , Técnicas de Visualização da Superfície Celular , Proteínas de Armazenamento de Sementes/química , Proteínas de Armazenamento de Sementes/genética , Anticorpos de Domínio Único/análise
6.
Nat Commun ; 10(1): 2925, 2019 07 02.
Artigo em Inglês | MEDLINE | ID: mdl-31266960

RESUMO

Bacteriophage Q protein engages σ-dependent paused RNA polymerase (RNAP) by binding to a DNA site embedded in late gene promoter and renders RNAP resistant to termination signals. Here, we report a single-particle cryo-electron microscopy (cryo-EM) structure of an intact Q-engaged arrested complex. The structure reveals key interactions responsible for σ-dependent pause, Q engagement, and Q-mediated transcription antitermination. The structure shows that two Q protomers (QI and QII) bind to a direct-repeat DNA site and contact distinct elements of the RNA exit channel. Notably, QI forms a narrow ring inside the RNA exit channel and renders RNAP resistant to termination signals by prohibiting RNA hairpin formation in the RNA exit channel. Because the RNA exit channel is conserved among all multisubunit RNAPs, it is likely to serve as an important contact site for regulators that modify the elongation properties of RNAP in other organisms, as well.


Assuntos
Bacteriófagos/enzimologia , Códon de Terminação/genética , RNA Polimerases Dirigidas por DNA/química , RNA Polimerases Dirigidas por DNA/metabolismo , Transcrição Genética , Proteínas Virais/química , Proteínas Virais/metabolismo , Bacteriófagos/química , Bacteriófagos/genética , Códon de Terminação/metabolismo , Microscopia Crioeletrônica , RNA Polimerases Dirigidas por DNA/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Escherichia coli/virologia , Regiões Promotoras Genéticas , Proteínas Virais/genética
8.
Nat Commun ; 10(1): 3048, 2019 07 11.
Artigo em Inglês | MEDLINE | ID: mdl-31296855

RESUMO

Bacteriophages typically hijack the host bacterial transcriptional machinery to regulate their own gene expression and that of the host bacteria. The structural basis for bacteriophage protein-mediated transcription regulation-in particular transcription antitermination-is largely unknown. Here we report the 3.4 Å and 4.0 Å cryo-EM structures of two bacterial transcription elongation complexes (P7-NusA-TEC and P7-TEC) comprising the bacteriophage protein P7, a master host-transcription regulator encoded by bacteriophage Xp10 of the rice pathogen Xanthomonas oryzae pv. Oryzae (Xoo) and discuss the mechanisms by which P7 modulates the host bacterial RNAP. The structures together with biochemical evidence demonstrate that P7 prevents transcription termination by plugging up the RNAP RNA-exit channel and impeding RNA-hairpin formation at the intrinsic terminator. Moreover, P7 inhibits transcription initiation by restraining RNAP-clamp motions. Our study reveals the structural basis for transcription antitermination by phage proteins and provides insights into bacterial transcription regulation.


Assuntos
Proteínas de Bactérias/metabolismo , Bacteriófagos/genética , Fatores de Elongação da Transcrição/metabolismo , Proteínas Virais/metabolismo , Xanthomonas/genética , Proteínas de Bactérias/isolamento & purificação , Proteínas de Bactérias/ultraestrutura , Microscopia Crioeletrônica , RNA Polimerases Dirigidas por DNA/isolamento & purificação , RNA Polimerases Dirigidas por DNA/metabolismo , RNA Polimerases Dirigidas por DNA/ultraestrutura , Regulação Bacteriana da Expressão Gênica , Regulação Viral da Expressão Gênica , Interações entre Hospedeiro e Microrganismos/genética , Oryza/microbiologia , Estrutura Secundária de Proteína , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/ultraestrutura , Regiões Terminadoras Genéticas/genética , Transcrição Genética , Fatores de Elongação da Transcrição/isolamento & purificação , Fatores de Elongação da Transcrição/ultraestrutura , Proteínas Virais/isolamento & purificação , Proteínas Virais/ultraestrutura , Xanthomonas/virologia
9.
Arch Virol ; 164(10): 2599-2603, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31278422

RESUMO

This work describes the characterization and genome annotation of a new lytic Enterococcus faecalis siphovirus, vB_EfaS_AL3 (referred to as AL3), isolated from wastewater samples collected in Liaoning Province, China. The genome of phage AL3 is composed of linear double-stranded DNA that is 40,789 bp in length with a G + C content of 34.84% and 61 putative protein-coding genes. Phylogenetic and comparative genomic analyses indicate that phage AL3 should be considered a novel phage.


Assuntos
Bacteriófagos/genética , Enterococcus faecalis/virologia , Genoma Viral , Filogenia , Análise de Sequência de DNA , Águas Residuárias/virologia , Bacteriólise , Composição de Bases , China , DNA/química , DNA/genética , DNA Viral/química , DNA Viral/genética , Microscopia Eletrônica de Transmissão , Anotação de Sequência Molecular , Ensaio de Placa Viral , Vírion/ultraestrutura
11.
Arch Virol ; 164(9): 2265-2275, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31197549

RESUMO

Proteus mirabilis is responsible for a wide range of infections that affect the urinary tract, the respiratory tract, burns, wounds and the feet of individuals with diabetes. They are highly resistant to antimicrobial agents, and new therapeutic options are therefore needed to combat this pathogen. The use of bacteriophages is one option that may be useful in treating multidrug-resistant (MDR) Proteus mirabilis infections, especially biofilm-based infections. The aim of this study was to control biofilms formed by MDR Proteus mirabilis using bacteriophages. Proteus mirabilis isolates were identified based on biochemical tests, and their resistance profiles were determined by the disk diffusion method. The biofilm-forming capacity of the isolates was assessed by the spectrophotometric method. Bacteriophages attacking Proteus mirabilis were isolated from sewage. The effect of phage on biofilm formation was investigated by the viable count method. A high rate of drug resistance was found (87.2%). Strong biofilm formation was observed in 80.5% of isolates, while moderate production was found in 19.5%. Five bacteriophages were isolated from sewage and were tested for their ability to eliminate biofilms. Significant disruption of pre-formed biofilms was observed that reached up to 99.9% decrease in the number of viable cells. The use of bacteriophages is considered a promising strategy against the biofilm infections caused by MDR Proteus mirabilis isolates.


Assuntos
Bacteriófagos/fisiologia , Biofilmes , Proteus mirabilis/fisiologia , Proteus mirabilis/virologia , Antibacterianos/farmacologia , Bacteriófagos/genética , Bacteriófagos/isolamento & purificação , Farmacorresistência Bacteriana , Proteus mirabilis/efeitos dos fármacos , Proteus mirabilis/genética , Esgotos/microbiologia , Esgotos/virologia
12.
Arch Virol ; 164(9): 2339-2343, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31214785

RESUMO

We isolated a novel lytic phage of Ralstonia solanacearum, GP4. The GP4 phage has a latent period of ~ 2 h at its optimal multiplicity of infection and is stable at temperatures ranging from 40-70 °C. GP4 lysed 16 strains of R. solanacearum belonging to phylotype IV. High-throughput sequencing revealed that GP4 has a linear dsDNA genome that consists of 61,129 bp, contains 83 open reading frames, and encodes a tRNA for cysteine. The GP4 genome has low similarity to other phage sequences in the GenBank database. Phylogenetic analysis indicated that GP4 can be taxonomically classified as a member of the Bcep22-like subfamily of the family Podoviridae.


Assuntos
Bacteriófagos/genética , Bacteriófagos/isolamento & purificação , Genoma Viral , Podoviridae/isolamento & purificação , Ralstonia solanacearum/virologia , Bacteriófagos/classificação , Sequenciamento de Nucleotídeos em Larga Escala , Fases de Leitura Aberta , Filogenia , Podoviridae/genética
13.
Arch Virol ; 164(9): 2389-2393, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31214784

RESUMO

Klebsiella pneumoniae is an important human pathogen that is associated with a wide range of diseases, including pneumonia and septicemia. Because of the threat of drug-resistant K. pneumoniae to humans, especially carbapenem-resistant K. pneumoniae, which is becoming a growing threat to hospitalized patients, the potential use of phage therapy has generated considerable interest. Henu1, isolated from a sewage sample, was identified as a linear double-stranded DNA phage of 40,352 bp with 53.14% G + C content and 143-bp terminal repeats. The Henu1 genome contains 45 open reading frames, and no tRNA genes were found. K. pneumoniae clinical strains with the capsular types K-1, K-2, and K-57 could be infected by Henu1. No human-virulence-related genes or lysogen-formation gene clusters were detected in this phage genome, suggesting that Henu1 is a virulent phage in its bacterial host and is safe for humans.


Assuntos
Bacteriófagos/isolamento & purificação , Genoma Viral , Klebsiella pneumoniae/virologia , Bacteriófagos/classificação , Bacteriófagos/genética , Bacteriófagos/fisiologia , Composição de Bases , Humanos , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/fisiologia , Fases de Leitura Aberta , Filogenia
14.
Arch Virol ; 164(9): 2277-2284, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31222428

RESUMO

To gain insight into the presence and nature of prophages in the black soldier fly (BSF; Hermetia illucens L. [Diptera: Stratiomyidae]) gut, we isolated and characterized a novel, temperate Escherichia bacteriophage designated vB_EcoS_PHB10 (PHB10). Electron microscopy analysis revealed that phage PHB10 has a long, flexible, non-contractile tail and belongs to the family Siphoviridae. The phage was found to be stable over a wide range of temperatures (4-37 °C) and pH values (pH 5-9), and it lysed two out of 13 Escherichia strains tested. The genome of PHB10 contains genes encoding a putative transcriptional regulator and an integrase, and it shows a high degree of similarity to a region of the Enterobacter cloacae MBRL1077 genome. Induction experiments revealed that phage PHB10 could be induced by different gut substrates, suggesting that diet might be a potential regulator of lytic/lysogenic switches in commensal lysogens.


Assuntos
Bacteriófagos/isolamento & purificação , Escherichia/virologia , Intestinos/virologia , Simuliidae/microbiologia , Simuliidae/virologia , Siphoviridae/isolamento & purificação , Animais , Bacteriófagos/classificação , Bacteriófagos/genética , Bacteriófagos/fisiologia , Genoma Viral , Intestinos/microbiologia , Larva/microbiologia , Larva/virologia , Lisogenia , Filogenia , Siphoviridae/classificação , Siphoviridae/genética , Siphoviridae/fisiologia
15.
Environ Int ; 129: 488-496, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31158595

RESUMO

The emerging contamination of pathogenic bacteria in the soil has caused a serious threat to public health and environmental security. Therefore, effective methods to inactivate pathogenic bacteria and decrease the environmental risks are urgently required. As a century-old technique, bacteriophage (phage) therapy has a high efficiency in targeting and inactivating pathogenic bacteria in different environmental systems. This review provides an update on the status of bacteriophage therapy for the inactivation of pathogenic bacteria in the soil environment. Specifically, the applications of phage therapy in soil-plant and soil-groundwater systems are summarized. In addition, the impact of phage therapy on soil functioning is described, including soil function gene transmission, soil microbial community stability, and soil nutrient cycling. Soil factors, such as soil temperature, pH, clay mineral, water content, and nutrient components, influence the survival and activity of phages in the soil. Finally, the future research prospects of phage therapy in soil environments are described.


Assuntos
Bactérias/virologia , Bacteriófagos/fisiologia , Doenças das Plantas/prevenção & controle , Microbiologia do Solo , Bactérias/genética , Bacteriófagos/genética , Doenças das Plantas/microbiologia , Temperatura Ambiente
17.
Ann Agric Environ Med ; 26(2): 203-209, 2019 Jun 17.
Artigo em Inglês | MEDLINE | ID: mdl-31232046

RESUMO

The antibiotic resistance in many pathogenic bacteria has become a major clinical problem, therefore, the necessity arises to search for new therapeutic strategies. The most promising solution lies in bacteriophages, phage endolysins and antimicrobial peptides. The aim of this study is to review the possibilities for the common use of bacteriophages, phage endolysins and antimicrobial peptides, both in the form of combined therapies and new strategies for the production of peptide drugs. Bacteriophages are viruses that specifically infect and destroy pathogenic bacteria by penetration into bacterial cells, causing metabolism disorders and, consequently, cell lysis. Phage-encoded endolysins are bacteriolytic proteins produced at the end of the phage lytic cycle that destroy elements of bacterial cell wall and enable the release of phage progeny from host cells. Antimicrobial peptides (AMPs) constitute an element of the innate immunity of living organisms and are characterized by the activity against a broad spectrum of bacteria. In the literature, there are only a few reports on the direct interaction of bacteriophages, phage endolysins and antimicrobial peptides against pathogenic bacteria. In each of them, a synergistic effect was observed, and Phage-encoded antimicrobial peptides as a specific group of AMPs have were also discussed. Phage-display technique was also reviewed in terms of its applications to produce and deliver biologically active peptides. The literature data also suggest that bacteriophages, phage endolysins and antimicrobial peptides can be used in combined therapy, thus negating many of the limitations resulting from their specificity as a single antimicrobial agent.


Assuntos
Antibacterianos/farmacologia , Peptídeos Catiônicos Antimicrobianos/farmacologia , Bactérias/efeitos dos fármacos , Infecções Bacterianas/tratamento farmacológico , Bacteriófagos/química , Endopeptidases/farmacologia , Animais , Antibacterianos/química , Antibacterianos/metabolismo , Peptídeos Catiônicos Antimicrobianos/química , Peptídeos Catiônicos Antimicrobianos/metabolismo , Bactérias/virologia , Infecções Bacterianas/microbiologia , Bacteriófagos/enzimologia , Bacteriófagos/genética , Bacteriófagos/fisiologia , Desenho de Drogas , Endopeptidases/química , Endopeptidases/metabolismo , Humanos
18.
BMC Evol Biol ; 19(1): 124, 2019 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-31215393

RESUMO

BACKGROUND: Mycobacteria occupy various ecological niches and can be isolated from soil, tap water and ground water. Several cause diseases in humans and animals. To get deeper insight into our understanding of mycobacterial evolution focusing on tRNA and non-coding (nc)RNA, we conducted a comparative genome analysis of Mycobacterium mucogenicum (Mmuc) and Mycobacterium neoaurum (Mneo) clade members. RESULTS: Genome sizes for Mmuc- and Mneo-clade members vary between 5.4 and 6.5 Mbps with the complete MmucT (type strain) genome encompassing 6.1 Mbp. The number of tRNA genes range between 46 and 79 (including one pseudo tRNA gene) with 39 tRNA genes common among the members of these clades, while additional tRNA genes were probably acquired through horizontal gene transfer. Selected tRNAs and ncRNAs (RNase P RNA, tmRNA, 4.5S RNA, Ms1 RNA and 6C RNA) are expressed, and the levels for several of these are higher in stationary phase compared to exponentially growing cells. The rare tRNAIleTAT isoacceptor and two for mycobacteria novel ncRNAs: the Lactobacillales-derived GOLLD RNA and a homolog to the antisense Salmonella typhimurium phage Sar RNA, were shown to be present and expressed in certain Mmuc-clade members. CONCLUSIONS: Phages, IS elements, horizontally transferred tRNA gene clusters, and phage-derived ncRNAs appears to have influenced the evolution of the Mmuc- and Mneo-clades. While the number of predicted coding sequences correlates with genome size, the number of tRNA coding genes does not. The majority of the tRNA genes in mycobacteria are transcribed mainly from single genes and the levels of certain ncRNAs, including RNase P RNA (essential for the processing of tRNAs), are higher at stationary phase compared to exponentially growing cells. We provide supporting evidence that Ms1 RNA represents a mycobacterial 6S RNA variant. The evolutionary routes for the ncRNAs RNase P RNA, tmRNA and Ms1 RNA are different from that of the core genes.


Assuntos
Genoma Bacteriano , Mycobacterium/crescimento & desenvolvimento , Mycobacterium/genética , RNA Bacteriano/genética , RNA de Transferência/genética , RNA não Traduzido/genética , Aminoacil-tRNA Sintetases/genética , Bacteriófagos/genética , Tamanho do Genoma , Genômica , Anotação de Sequência Molecular , Mycobacterium/classificação , Filogenia , Plasmídeos/genética , RNA não Traduzido/química , Ribonuclease P/genética , Inversão de Sequência
19.
PLoS Genet ; 15(6): e1008221, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31242186

RESUMO

Wolbachia are maternally inherited bacteria that infect arthropod species worldwide and are deployed in vector control to curb arboviral spread using cytoplasmic incompatibility (CI). CI kills embryos when an infected male mates with an uninfected female, but the lethality is rescued if the female and her embryos are likewise infected. Two phage WO genes, cifAwMel and cifBwMel from the wMel Wolbachia deployed in vector control, transgenically recapitulate variably penetrant CI, and one of the same genes, cifAwMel, rescues wild type CI. The proposed Two-by-One genetic model predicts that CI and rescue can be recapitulated by transgenic expression alone and that dual cifAwMel and cifBwMel expression can recapitulate strong CI. Here, we use hatch rate and gene expression analyses in transgenic Drosophila melanogaster to demonstrate that CI and rescue can be synthetically recapitulated in full, and strong, transgenic CI comparable to wild type CI is achievable. These data explicitly validate the Two-by-One model in wMel-infected D. melanogaster, establish a robust system for transgenic studies of CI in a model system, and represent the first case of completely engineering male and female animal reproduction to depend upon bacteriophage gene products.


Assuntos
Bacteriófagos/genética , Drosophila melanogaster/genética , Proteínas Virais/genética , Wolbachia/genética , Animais , Animais Geneticamente Modificados/genética , Animais Geneticamente Modificados/crescimento & desenvolvimento , Citoplasma/genética , Citoplasma/metabolismo , Citoplasma/microbiologia , Vetores de Doenças , Drosophila melanogaster/crescimento & desenvolvimento , Drosophila melanogaster/microbiologia , Feminino , Regulação da Expressão Gênica/genética , Masculino , Herança Materna/genética , Reprodução/genética , Wolbachia/patogenicidade , Wolbachia/virologia
20.
Biosci Biotechnol Biochem ; 83(9): 1683-1696, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31094670

RESUMO

The fully synthetic humanized phage antibody library has the advantages including the minimized immunogenicity, which frequently happened in hybridoma cell-based antibody production. In this paper, using the constructed diverse complementarity determining region gene library and the germline gene as the backbone, we constructed eight single-chain antibody libraries and a combinatorial antibody library with a big capacity of 1.41 × 1010. M13EEA helper phage that was engineered from M13KO7 was applied to prepare phage antibody library. The eukaryotic expression of T-cell immune receptor with Ig and ITIM domain (TIGIT) antigen was used as a target antigen for screening. The screening of antigen-specific single-chain Fc-fused protein was performed through evaluation of binding affinity based on ELISA analysis. The IgG antibody was prepared with the screened single-chain protein. Finally, the CB3 antibody was screened out which exhibits the highest binding affinity with TIGIT with the Kd value of 8.155 × 10-10 M.


Assuntos
Bacteriófagos/genética , Imunoterapia/métodos , Neoplasias/terapia , Ensaio de Imunoadsorção Enzimática , Humanos , Neoplasias/imunologia , Ressonância de Plasmônio de Superfície
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