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Article ; Online: Immunoinformatics, molecular docking and dynamics simulation approaches unveil a multi epitope-based potent peptide vaccine candidate against avian leukosis virus.

Elshafei, Siham O / Mahmoud, Nuha A / Almofti, Yassir A

2024 Volume 14, Issue 1, Page(s) 2870

Abstract: Lymphoid leukosis is a poultry neoplastic disease caused by avian leukosis virus (ALV) and is characterized by high morbidity and variable mortality rates in chicks. Currently, no effective treatment and vaccination is the only means to control it. This ... ...

Abstract	Lymphoid leukosis is a poultry neoplastic disease caused by avian leukosis virus (ALV) and is characterized by high morbidity and variable mortality rates in chicks. Currently, no effective treatment and vaccination is the only means to control it. This study exploited the immunoinformatics approaches to construct multi-epitope vaccine against ALV. ABCpred and IEDB servers were used to predict B and T lymphocytes epitopes from the viral proteins, respectively. Antigenicity, allergenicity and toxicity of the epitopes were assessed and used to construct the vaccine with suitable adjuvant and linkers. Secondary and tertiary structures of the vaccine were predicted, refined and validated. Structural errors, solubility, stability, immune simulation, dynamic simulation, docking and in silico cloning were also evaluated.The constructed vaccine was hydrophilic, antigenic and non-allergenic. Ramchandran plot showed most of the residues in the favored and additional allowed regions. ProsA server showed no errors in the vaccine structure. Immune simulation showed significant immunoglobulins and cytokines levels. Stability was enhanced by disulfide engineering and molecular dynamic simulation. Docking of the vaccine with chicken's TLR7 revealed competent binding energies.The vaccine was cloned in pET-30a(+) vector and efficiently expressed in Escherichia coli. This study provided a potent peptide vaccine that could assist in tailoring a rapid and cost-effective vaccine that helps to combat ALV. However, experimental validation is required to assess the vaccine efficiency.
MeSH term(s)	Animals ; Molecular Docking Simulation ; Avian Leukosis Virus ; Protein Subunit Vaccines ; Immunoinformatics ; Chickens ; Epitopes, T-Lymphocyte ; Molecular Dynamics Simulation ; Epitopes, B-Lymphocyte ; Vaccines, Subunit ; Computational Biology
Chemical Substances	Protein Subunit Vaccines ; Epitopes, T-Lymphocyte ; Epitopes, B-Lymphocyte ; Vaccines, Subunit
Language	English
Publishing date	2024-02-04
Publishing country	England
Document type	Journal Article
ZDB-ID	2615211-3
ISSN	2045-2322 ; 2045-2322
ISSN (online)	2045-2322
ISSN	2045-2322
DOI	10.1038/s41598-024-53048-6
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Article ; Online: A novel strategy for developing vaccine candidate against Jaagsiekte sheep retrovirus from the envelope and gag proteins: an in-silico approach.

Mahmoud, Nuha Amin / Elshafei, Abdelmajeed M / Almofti, Yassir A

BMC veterinary research

2022 Volume 18, Issue 1, Page(s) 343

Abstract: Background: Sheep pulmonary adenocarcinoma (OPA) is a contagious lung cancer of sheep caused by the Jaagsiekte retrovirus (JSRV). OPA typically has a serious economic impact worldwide. A vaccine has yet to be developed, even though the disease has been ... ...

Abstract	Background: Sheep pulmonary adenocarcinoma (OPA) is a contagious lung cancer of sheep caused by the Jaagsiekte retrovirus (JSRV). OPA typically has a serious economic impact worldwide. A vaccine has yet to be developed, even though the disease has been globally spread, along with its complications. This study aimed to construct an effective multi-epitopes vaccine against JSRV eliciting B and T lymphocytes using immunoinformatics tools. Results: The designed vaccine was composed of 499 amino acids. Before the vaccine was computationally validated, all critical parameters were taken into consideration; including antigenicity, allergenicity, toxicity, and stability. The physiochemical properties of the vaccine displayed an isoelectric point of 9.88. According to the Instability Index (II), the vaccine was stable at 28.28. The vaccine scored 56.51 on the aliphatic index and -0.731 on the GRAVY, indicating that the vaccine was hydrophilic. The RaptorX server was used to predict the vaccine's tertiary structure, the GalaxyWEB server refined the structure, and the Ramachandran plot and the ProSA-web server validated the vaccine's tertiary structure. Protein-sol and the SOLPro servers showed the solubility of the vaccine. Moreover, the high mobile regions in the vaccine's structure were reduced and the vaccine's stability was improved by disulfide engineering. Also, the vaccine construct was docked with an ovine MHC-1 allele and showed efficient binding energy. Immune simulation remarkably showed high levels of immunoglobulins, T lymphocytes, and INF-γ secretions. The molecular dynamic simulation provided the stability of the constructed vaccine. Finally, the vaccine was back-transcribed into a DNA sequence and cloned into a pET-30a ( +) vector to affirm the potency of translation and microbial expression. Conclusion: A novel multi-epitopes vaccine construct against JSRV, was formed from B and T lymphocytes epitopes, and was produced with potential protection. This study might help in controlling and eradicating OPA.
MeSH term(s)	Adenocarcinoma of Lung/veterinary ; Animals ; Epitopes ; Jaagsiekte sheep retrovirus ; Lung Neoplasms/veterinary ; Sheep ; Sheep Diseases/prevention & control ; Vaccines
Chemical Substances	Epitopes ; Vaccines
Language	English
Publishing date	2022-09-10
Publishing country	England
Document type	Journal Article
ZDB-ID	2191675-5
ISSN	1746-6148 ; 1746-6148
ISSN (online)	1746-6148
ISSN	1746-6148
DOI	10.1186/s12917-022-03431-0
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Article ; Online: Vaccinomic approach for novel multi epitopes vaccine against severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2).

Almofti, Yassir A / Abd-Elrahman, Khoubieb Ali / Eltilib, Elsideeq E M

BMC immunology

2021 Volume 22, Issue 1, Page(s) 22

Abstract: Background: The spread of a novel coronavirus termed severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in China and other countries is of great concern worldwide with no effective vaccine. This study aimed to design a novel vaccine construct ... ...

Abstract	Background: The spread of a novel coronavirus termed severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in China and other countries is of great concern worldwide with no effective vaccine. This study aimed to design a novel vaccine construct against SARS-CoV-2 from the spike S protein and orf1ab polyprotein using immunoinformatics tools. The vaccine was designed from conserved epitopes interacted against B and T lymphocytes by the combination of highly immunogenic epitopes with suitable adjuvant and linkers. Results: The proposed vaccine composed of 526 amino acids and was shown to be antigenic in Vaxigen server (0.6194) and nonallergenic in Allertop server. The physiochemical properties of the vaccine showed isoelectric point of 10.19. The instability index (II) was 31.25 classifying the vaccine as stable. Aliphatic index was 84.39 and the grand average of hydropathicity (GRAVY) was - 0.049 classifying the vaccine as hydrophilic. Vaccine tertiary structure was predicted, refined and validated to assess the stability of the vaccine via Ramachandran plot and ProSA-web servers. Moreover, solubility of the vaccine construct was greater than the average solubility provided by protein sol and SOLpro servers indicating the solubility of the vaccine construct. Disulfide engineering was performed to reduce the high mobile regions in the vaccine to enhance stability. Docking of the vaccine construct with TLR4 demonstrated efficient binding energy with attractive binding energy of - 338.68 kcal/mol and - 346.89 kcal/mol for TLR4 chain A and chain B respectively. Immune simulation significantly provided high levels of immunoglobulins, T-helper cells, T-cytotoxic cells and INF-γ. Upon cloning, the vaccine protein was reverse transcribed into DNA sequence and cloned into pET28a(+) vector to ensure translational potency and microbial expression. Conclusion: A unique vaccine construct from spike S protein and orf1ab polyprotein against B and T lymphocytes was generated with potential protection against the pandemic. The present study might assist in developing a suitable therapeutics protocol to combat SARSCoV-2 infection.
MeSH term(s)	B-Lymphocytes/immunology ; COVID-19/immunology ; COVID-19/prevention & control ; COVID-19 Vaccines/chemistry ; COVID-19 Vaccines/genetics ; COVID-19 Vaccines/immunology ; Epitopes, B-Lymphocyte/chemistry ; Epitopes, B-Lymphocyte/genetics ; Epitopes, B-Lymphocyte/immunology ; Epitopes, T-Lymphocyte/chemistry ; Epitopes, T-Lymphocyte/genetics ; Epitopes, T-Lymphocyte/immunology ; Humans ; Polyproteins/chemistry ; Polyproteins/genetics ; Polyproteins/immunology ; SARS-CoV-2/chemistry ; SARS-CoV-2/genetics ; SARS-CoV-2/immunology ; Spike Glycoprotein, Coronavirus/chemistry ; Spike Glycoprotein, Coronavirus/genetics ; Spike Glycoprotein, Coronavirus/immunology ; T-Lymphocytes/immunology ; Viral Proteins/chemistry ; Viral Proteins/genetics ; Viral Proteins/immunology
Chemical Substances	COVID-19 Vaccines ; Epitopes, B-Lymphocyte ; Epitopes, T-Lymphocyte ; ORF1ab polyprotein, SARS-CoV-2 ; Polyproteins ; Spike Glycoprotein, Coronavirus ; Viral Proteins ; spike protein, SARS-CoV-2
Language	English
Publishing date	2021-03-25
Publishing country	England
Document type	Journal Article
ISSN	1471-2172
ISSN (online)	1471-2172
DOI	10.1186/s12865-021-00412-0
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Article: Immunoinformatics Approach for Multiepitopes Vaccine Prediction against Glycoprotein B of Avian Infectious Laryngotracheitis Virus.

Ali, Sumaia A / Almofti, Yassir A / Abd-Elrahman, Khoubieb A

Advances in bioinformatics

2019 Volume 2019, Page(s) 1270485

Abstract: Infectious laryngotracheitis virus (ILTV) is ... ...

Abstract	Infectious laryngotracheitis virus (ILTV) is a
Language	English
Publishing date	2019-03-18
Publishing country	Egypt
Document type	Journal Article
ZDB-ID	2448875-6
ISSN	1687-8035 ; 1687-8027
ISSN (online)	1687-8035
ISSN	1687-8027
DOI	10.1155/2019/1270485
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Article ; Online: A novel strategy for developing vaccine candidate against Jaagsiekte sheep retrovirus from the envelope and gag proteins: an in-silico approach

Mahmoud, Nuha Amin / Elshafei, Abdelmajeed M. / Almofti, Yassir A.

BMC Vet Res. 2022 Dec., v. 18, no. 1 p.343-343

2022

Abstract: BACKGROUND: Sheep pulmonary adenocarcinoma (OPA) is a contagious lung cancer of sheep caused by the Jaagsiekte retrovirus (JSRV). OPA typically has a serious economic impact worldwide. A vaccine has yet to be developed, even though the disease has been ... ...

Abstract	BACKGROUND: Sheep pulmonary adenocarcinoma (OPA) is a contagious lung cancer of sheep caused by the Jaagsiekte retrovirus (JSRV). OPA typically has a serious economic impact worldwide. A vaccine has yet to be developed, even though the disease has been globally spread, along with its complications. This study aimed to construct an effective multi-epitopes vaccine against JSRV eliciting B and T lymphocytes using immunoinformatics tools. RESULTS: The designed vaccine was composed of 499 amino acids. Before the vaccine was computationally validated, all critical parameters were taken into consideration; including antigenicity, allergenicity, toxicity, and stability. The physiochemical properties of the vaccine displayed an isoelectric point of 9.88. According to the Instability Index (II), the vaccine was stable at 28.28. The vaccine scored 56.51 on the aliphatic index and -0.731 on the GRAVY, indicating that the vaccine was hydrophilic. The RaptorX server was used to predict the vaccine's tertiary structure, the GalaxyWEB server refined the structure, and the Ramachandran plot and the ProSA-web server validated the vaccine's tertiary structure. Protein-sol and the SOLPro servers showed the solubility of the vaccine. Moreover, the high mobile regions in the vaccine’s structure were reduced and the vaccine’s stability was improved by disulfide engineering. Also, the vaccine construct was docked with an ovine MHC-1 allele and showed efficient binding energy. Immune simulation remarkably showed high levels of immunoglobulins, T lymphocytes, and INF-γ secretions. The molecular dynamic simulation provided the stability of the constructed vaccine. Finally, the vaccine was back-transcribed into a DNA sequence and cloned into a pET-30a (+) vector to affirm the potency of translation and microbial expression. CONCLUSION: A novel multi-epitopes vaccine construct against JSRV, was formed from B and T lymphocytes epitopes, and was produced with potential protection. This study might help in controlling and eradicating OPA.
Keywords	Jaagsiekte sheep retrovirus ; adenocarcinoma ; alleles ; allergenicity ; computer simulation ; disulfides ; economic impact ; energy ; epitopes ; hydrophilicity ; immunoglobulins ; immunoinformatics ; isoelectric point ; lung neoplasms ; nucleotide sequences ; ovine pulmonary adenomatosis ; sheep ; solubility ; toxicity ; vaccines
Language	English
Dates of publication	2022-12
Size	p. 343.
Publishing place	BioMed Central
Document type	Article ; Online
ZDB-ID	2191675-5
ISSN	1746-6148
ISSN	1746-6148
DOI	10.1186/s12917-022-03431-0
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Article ; Online: Immunoinformatics Approach for Multiepitope Vaccine Prediction from H, M, F, and N Proteins of Peste des Petits Ruminants Virus.

Gaafar, Bothina B M / Ali, Sumaia A / Abd-Elrahman, Khoubieb Ali / Almofti, Yassir A

Journal of immunology research

2019 Volume 2019, Page(s) 6124030

Abstract: Background: Small ruminant morbillivirus or peste des petits ruminants virus (PPRV) is an acute and highly contagious viral disease of goats, sheep, and other livestock. This study aimed at predicting an effective multiepitope vaccine against PPRV from ... ...

Abstract	Background: Small ruminant morbillivirus or peste des petits ruminants virus (PPRV) is an acute and highly contagious viral disease of goats, sheep, and other livestock. This study aimed at predicting an effective multiepitope vaccine against PPRV from the immunogenic proteins haemagglutinin (H), matrix (M), fusion (F), and nucleoprotein (N) using immunoinformatics tools. Materials and methods: The sequences of the immunogenic proteins were retrieved from GenBank of the National Center for Biotechnology Information (NCBI). BioEdit software was used to align each protein from the retrieved sequences for conservancy. Immune Epitope Database (IEDB) analysis resources were used to predict B and T cell epitopes. For B cells, the criteria for electing epitopes depend on the epitope linearity, surface accessibility, and antigenicity. Results: Nine epitopes from the H protein, eight epitopes from the M protein, and ten epitopes from each of the F and N proteins were predicted as linear epitopes. The surface accessibility method proposed seven surface epitopes from each of the H and F proteins in addition to six and four epitopes from the M and N proteins, respectively. For antigenicity, only two epitopes Conclusion: Fourteen epitopes were predicted as promising vaccine candidates against PPRV from four immunogenic proteins. These epitopes should be validated experimentally through in vitro and in vivo studies.
MeSH term(s)	Amino Acid Sequence ; Animals ; Antigens, Viral/chemistry ; Antigens, Viral/genetics ; Antigens, Viral/immunology ; Base Sequence ; Computational Biology/methods ; Epitope Mapping ; Epitopes/chemistry ; Epitopes/genetics ; Epitopes/immunology ; Epitopes, T-Lymphocyte/chemistry ; Epitopes, T-Lymphocyte/genetics ; Epitopes, T-Lymphocyte/immunology ; Models, Molecular ; Peste-des-Petits-Ruminants/prevention & control ; Peste-des-petits-ruminants virus/classification ; Peste-des-petits-ruminants virus/genetics ; Peste-des-petits-ruminants virus/immunology ; Phylogeny ; Protein Conformation ; Structure-Activity Relationship ; Viral Proteins/chemistry ; Viral Proteins/genetics ; Viral Proteins/immunology ; Viral Vaccines/administration & dosage ; Viral Vaccines/immunology
Chemical Substances	Antigens, Viral ; Epitopes ; Epitopes, T-Lymphocyte ; Viral Proteins ; Viral Vaccines
Language	English
Publishing date	2019-10-30
Publishing country	Egypt
Document type	Journal Article
ZDB-ID	2817541-4
ISSN	2314-7156 ; 2314-7156
ISSN (online)	2314-7156
ISSN	2314-7156
DOI	10.1155/2019/6124030
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Article ; Online: Impact of erythromycin resistance on the virulence properties and fitness of Campylobacter jejuni.

Almofti, Yassir A / Dai, Menghong / Sun, Yawei / Haihong, Hao / Yuan, Zonghui

Microbial pathogenesis

2011 Volume 50, Issue 6, Page(s) 336–342

Abstract: Epidemiological studies of macrolide resistance in Campylobacter jejuni demonstrated that infections with macrolide-resistant C. jejuni could be associated with an increased risk of adverse events, development of invasive illness or death compared to ... ...

Abstract	Epidemiological studies of macrolide resistance in Campylobacter jejuni demonstrated that infections with macrolide-resistant C. jejuni could be associated with an increased risk of adverse events, development of invasive illness or death compared to macrolide-susceptible isolates. In this study, an in vitro induction experiment was conducted using susceptible C. jejuni strain and erythromycin as a selecting agent to obtain Ery-resistant mutant with 23S rRNA gene mutation (A2074C). Changes in the virulence characteristics and fitness between the susceptible parent strain and Ery-resistant mutant were examined. Ery-resistant mutant demonstrated slightly more resistance to bile in the bile tolerance assay compared to the susceptible strain but with no statistical significant difference. However Ery-resistant mutant apparently demonstrated reduced adhesion and invasion characteristics to intestinal epithelial cells, murine macrophage and short time intracellular survivability within macrophage compared to the susceptible strain. Co-inoculation of the two strains in the mice resulted in low colonization level of the resistant strain compared to the susceptible strain. Competition experiments resulted in mutant that grew significantly slower than the susceptible parent strain and the mutation imposed a fitness cost in Ery-resistant mutant. Taken together these findings demonstrated the increment of the virulence characteristics of Ery-susceptible strain rather than Ery-resistant strain. The adverse events previously observed in the epidemiological studies for macrolide-resistant strains infection, we suggested this maybe attributed to the resistivity of the resistant strains to the treatment and consequently prolonged the symptoms and compromised the disease in patients.
MeSH term(s)	Animals ; Anti-Bacterial Agents/pharmacology ; Campylobacter jejuni/drug effects ; Campylobacter jejuni/genetics ; Campylobacter jejuni/growth & development ; Campylobacter jejuni/pathogenicity ; Drug Resistance, Bacterial ; Erythromycin/pharmacology ; Gene Deletion ; Genetic Variation ; Humans ; Mice ; Microbial Sensitivity Tests ; Mutation/drug effects ; RNA, Ribosomal/drug effects ; Virulence/drug effects ; Virulence/genetics
Chemical Substances	Anti-Bacterial Agents ; RNA, Ribosomal ; Erythromycin (63937KV33D)
Language	English
Publishing date	2011-06
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	632772-2
ISSN	1096-1208 ; 0882-4010
ISSN (online)	1096-1208
ISSN	0882-4010
DOI	10.1016/j.micpath.2011.02.009
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Article: The physiologic and phenotypic alterations due to macrolide exposure in Campylobacter jejuni

Almofti, Yassir A / Dai, Menghong / Sun, Yawei / Hao, Haihong / Liu, Zhenli / Cheng, Guyue / Yuan, Zonghui

International journal of food microbiology. 2011 Nov. 15, v. 151, no. 1

2011

Abstract: Physiologic and phenotypic alterations in the context of antibiotic resistance have been extensively studied in some bacteria. However there are not enough data addressing these alterations due to macrolide resistance in Campylobacter jejuni. The present ...

Abstract	Physiologic and phenotypic alterations in the context of antibiotic resistance have been extensively studied in some bacteria. However there are not enough data addressing these alterations due to macrolide resistance in Campylobacter jejuni. The present study examined the fitness cost imposed by different macrolide resistance mutations and the phenotypic alterations due to exposure to macrolides in C. jejuni. C. jejuni was induced with different macrolide agents to obtain different macrolide resistance mutations. The results revealed that the mutations significantly imposed defect variations on the doubling time and the relative fitness in the resistant strains when competed against the susceptible strain. Furthermore macrolides through induction or exposure to sub-MIC concentrations impaired the motility of C. jejuni in 0.4% MH agar plates. Electron microscope analysis revealed the absence of flagellar filaments from strains exposed to macrolides. SDS-PAGE demonstrated that macrolides have no effect on protein synthesis and immunoblotting analysis further confirmed that flagellin was fully synthesized within C. jejuni strains exposed to macrolides. Nevertheless C. jejuni strains exposed to macrolides demonstrated defect in their excreted flagellin into the supernatant compared to strains not exposed to macrolides. Accordingly we speculated that macrolides inhibited flagellar filament formation in the strains exposed to macrolides via affecting the secretion of flagellin without affecting the amount synthesized within the bacteria. Taken together these findings demonstrated that different macrolide resistance mutations imposed different fitness costs and exposure to macrolides resulted in phenotypic alterations such as inhibition of flagellar filament formation and loss of motility in C. jejuni.
Keywords	Campylobacter jejuni ; agar ; antibiotic resistance ; bacteria ; electron microscopes ; flagellin ; immunoblotting ; macrolides ; mutation ; polyacrylamide gel electrophoresis ; protein synthesis ; secretion
Language	English
Dates of publication	2011-1115
Size	p. 52-61.
Publishing place	Elsevier B.V.
Document type	Article
ZDB-ID	87122-9
ISSN	1879-3460 ; 0168-1605
ISSN (online)	1879-3460
ISSN	0168-1605
DOI	10.1016/j.ijfoodmicro.2011.08.002
Database	NAL-Catalogue (AGRICOLA)

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Article ; Online: The physiologic and phenotypic alterations due to macrolide exposure in Campylobacter jejuni.

Almofti, Yassir A / Dai, Menghong / Sun, Yawei / Hao, Haihong / Liu, Zhenli / Cheng, Guyue / Yuan, Zonghui

International journal of food microbiology

2011 Volume 151, Issue 1, Page(s) 52–61

Abstract	Physiologic and phenotypic alterations in the context of antibiotic resistance have been extensively studied in some bacteria. However there are not enough data addressing these alterations due to macrolide resistance in Campylobacter jejuni. The present study examined the fitness cost imposed by different macrolide resistance mutations and the phenotypic alterations due to exposure to macrolides in C. jejuni. C. jejuni was induced with different macrolide agents to obtain different macrolide resistance mutations. The results revealed that the mutations significantly imposed defect variations on the doubling time and the relative fitness in the resistant strains when competed against the susceptible strain. Furthermore macrolides through induction or exposure to sub-MIC concentrations impaired the motility of C. jejuni in 0.4% MH agar plates. Electron microscope analysis revealed the absence of flagellar filaments from strains exposed to macrolides. SDS-PAGE demonstrated that macrolides have no effect on protein synthesis and immunoblotting analysis further confirmed that flagellin was fully synthesized within C. jejuni strains exposed to macrolides. Nevertheless C. jejuni strains exposed to macrolides demonstrated defect in their excreted flagellin into the supernatant compared to strains not exposed to macrolides. Accordingly we speculated that macrolides inhibited flagellar filament formation in the strains exposed to macrolides via affecting the secretion of flagellin without affecting the amount synthesized within the bacteria. Taken together these findings demonstrated that different macrolide resistance mutations imposed different fitness costs and exposure to macrolides resulted in phenotypic alterations such as inhibition of flagellar filament formation and loss of motility in C. jejuni.
MeSH term(s)	Anti-Bacterial Agents/pharmacology ; Campylobacter jejuni/drug effects ; Campylobacter jejuni/genetics ; Campylobacter jejuni/physiology ; DNA Mutational Analysis ; DNA, Bacterial/genetics ; Drug Resistance, Bacterial/genetics ; Flagella/physiology ; Flagellin/genetics ; Flagellin/metabolism ; Genetic Fitness ; Macrolides/pharmacology ; Microbial Sensitivity Tests ; Mutation/drug effects ; Phenotype
Chemical Substances	Anti-Bacterial Agents ; DNA, Bacterial ; Macrolides ; Flagellin (12777-81-0)
Language	English
Publishing date	2011-11-15
Publishing country	Netherlands
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	87122-9
ISSN	1879-3460 ; 0168-1605
ISSN (online)	1879-3460
ISSN	0168-1605
DOI	10.1016/j.ijfoodmicro.2011.08.002
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Article ; Online: The role of RamA on the development of ciprofloxacin resistance in Salmonella enterica serovar Typhimurium.

Sun, Yawei / Dai, Menghong / Hao, Haihong / Wang, Yulian / Huang, Lingli / Almofti, Yassir A / Liu, Zhenli / Yuan, Zonghui

PloS one

2011 Volume 6, Issue 8, Page(s) e23471

Abstract: Active efflux pump is a primary fluoroquinolone resistant mechanism of clinical isolates of Salmonella enterica serovar Typhimurium. RamA is an essential element in producing multidrug resistant (MDR) S. enterica serovar Typhimurium. The aim of the ... ...

Abstract	Active efflux pump is a primary fluoroquinolone resistant mechanism of clinical isolates of Salmonella enterica serovar Typhimurium. RamA is an essential element in producing multidrug resistant (MDR) S. enterica serovar Typhimurium. The aim of the present study was to elucidate the roles of RamA on the development of ciprofloxacin, the first choice for the treatment of salmonellosis, resistance in S. enterica serovar Typhimurium. Spontaneous mutants were selected via several passages of S. enterica serovar Typhimurium CVCC541 susceptible strain (ST) on M-H agar with increasing concentrations of ciprofloxacin (CIP). Accumulation of ciprofloxacin was tested by the modified fluorometric method. The expression levels of MDR efflux pumps were determined by real time RT-PCR. In ST and its spontaneous mutants, the ramA gene was inactivated by insertion of the kan gene and compensated on a recombinant plasmid pGEXΦ(gst-ramA). The mutant prevention concentration (MPC) and mutant frequencies of ciprofloxacin against ST and a spontaneous mutant in the presence, absence and overexpression of RamA were tested. Four spontaneous mutants (SI1-SI4) were obtained. The SI1 (CIP MICs, 0.1 mg/L) without any target site mutation in its quinolone resistant determining regions (QRDRs) and SI3 (CIP MICs, 16 mg/L) harboring the Ser83→Phe mutation in its QRDR of GyrA strains exhibited reduced susceptibility and resistance to multidrugs, respectively. In SI1, RamA was the main factor that controlled the susceptibility to ciprofloxacin by activating MdtK as well as increasing the expression level of acrAB. In SI3, RamA played predominant role in ciprofloxacin resistance via increasing the expression level of acrAB. Likewise, the deficiency of RamA decreased the MPCs and mutant frequencies of ST and SI2 to ciprofloxacin. In conclusion, the expression of RamA promoted the development of ciprofloxacin resistant mutants of S. enterica serovar Typhimurium. The inhibition of RamA could decrease the appearance of the ciprofloxacin resistant mutants.
MeSH term(s)	Anti-Infective Agents/metabolism ; Anti-Infective Agents/pharmacology ; Bacterial Proteins/genetics ; Bacterial Proteins/physiology ; Biological Transport/drug effects ; Carbonyl Cyanide m-Chlorophenyl Hydrazone/pharmacology ; Ciprofloxacin/metabolism ; Ciprofloxacin/pharmacology ; Dipeptides/pharmacology ; Dose-Response Relationship, Drug ; Drug Resistance, Bacterial/genetics ; Drug Resistance, Bacterial/physiology ; Drug Resistance, Multiple, Bacterial/genetics ; Drug Resistance, Multiple, Bacterial/physiology ; Escherichia coli/drug effects ; Gene Expression Regulation, Bacterial ; Membrane Transport Proteins/genetics ; Membrane Transport Proteins/metabolism ; Microbial Sensitivity Tests ; Molecular Sequence Data ; Multidrug Resistance-Associated Proteins/genetics ; Multidrug Resistance-Associated Proteins/physiology ; Mutation ; Nalidixic Acid/pharmacology ; Promoter Regions, Genetic/genetics ; Proton Ionophores/pharmacology ; Reverse Transcriptase Polymerase Chain Reaction ; Salmonella typhimurium/drug effects ; Salmonella typhimurium/genetics ; Salmonella typhimurium/metabolism ; Trans-Activators/genetics ; Trans-Activators/physiology
Chemical Substances	Anti-Infective Agents ; Bacterial Proteins ; Dipeptides ; Membrane Transport Proteins ; Multidrug Resistance-Associated Proteins ; Proton Ionophores ; RamA protein, Salmonella typhimurium ; Trans-Activators ; phenylalanine arginine beta-naphthylamide ; Nalidixic Acid (3B91HWA56M) ; Carbonyl Cyanide m-Chlorophenyl Hydrazone (555-60-2) ; Ciprofloxacin (5E8K9I0O4U)
Language	English
Publishing date	2011-08-12
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ISSN	1932-6203
ISSN (online)	1932-6203
DOI	10.1371/journal.pone.0023471
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In stock of ZB MED Cologne/Königswinter

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In stock of ZB MED Bonn / Germany

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In stock of ZB MED Cologne/Königswinter

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Order via subito

Inter-library loan at ZB MED