A novel biosensing strategy for identification of three important bacteria causing meningitis.

Bacterial meningitis is an acute infection which requires rapid diagnosis and treatment due to the high mortality and serious consequences of the disease. The purpose of this study was to design a homemade multiplex PCR and a novel fluorescence biosensor on chip (FBC) to detect three important agents of meningitis including Streptococcus pneumoniae (S. pneumoniae), Neisseria meningitidis (N. meningitidis), and Haemophilus influenzae (H. influenzae). The homemade multiplex PCR can diagnose three bacterial species simultaneously. Fabrication of FBC was carried out based on the deposition of lead nanoparticles on a quartz slide using the thermal evaporation method. Then, the SH-Cap Probe/Target ssDNA /FAM-Rep probe was loaded on lead film. The evaluation of the fluorescence reaction when the probes bind to the target ssDNA was assessed by a Cytation 5 Cell Imaging Multimode Reader Bio-Tek. The limit of detections (LOD) in homemade PCR and FBC to identify S. pneumoniae were 119 × 102 CFU/mL (0.27 ng/µL) and 380 CFU/mL (9 pg/µL), respectively. The LODs of homemade PCR and FBC for detection of N. meningitidis were 4.49 CFU/mL (1.1 pg/µL) and 13 × 103 CFU/mL (30 pg/µL), respectively. Our results confirmed the LODs of homemade PCR and FBC in detection of H. influenzae were 15.1 CFU/mL (30 fg/µL) and 41 × 102 CFU/mL (90 pg/ µL), respectively. Both techniques had appropriate sensitivity and specificity in detection of S. pneumoniae, N. meningitidis and H. influenzae.

Reduction of UreB and CagA expression level by siRNA construct in Helicobacter pylori strain SS1.

BACKGROUND: Two important virulence factors, urease and cagA, play an important role in Helicobacter pylori (H. pylori) gastric cancer. Aim of this study was to investigate the expression level and function of ureB and cagA using small interfering RNAs (siRNA). METHODS: SS1 strain of H. pylori was considered as host for natural transformation. siRNA designed for ureB and cagA genes were inserted in pGPU6/GFP/Neo siRNA plasmid vector to evaluate using phenotypic and genotypic approaches. Then, qPCR was performed for determining inhibition rate of ureB and cagA gene expression. RESULTS: The expression levels of siRNA-ureB and siRNA-cagA in the recombinant strain SS1 were reduced by about 5000 and 1000 fold, respectively, compared to the native H. pylori strain SS1. Also, preliminary evaluation of siRNA-ureB in vitro showed inhibition of urea enzyme activity. These data suggest that siRNA may be a powerful new tool for gene silencing in vitro, and for the development of RNAi-based anti-H. pylori therapies. CONCLUSION: Our results show that targeting ureB and cagA genes with siRNA seems to be a new strategy to inhibit urease enzyme activity, reduce inflammation and colonization rate.

Evaluation of combined carbon dots and ciprofloxacin on the expression level of pslA, pelA, and ppyR genes and biofilm production in ciprofloxacin-resistant Pseudomonas aeruginosa isolates from burn wound infection in Iran.

OBJECTIVES: Antimicrobial resistance and biofilm formation are increasingly significant public health concerns. This study aimed to examine the antibacterial and antibiofilm properties of carbon dots (C-dots) alone and in combination with antibiotics against biofilm-forming isolates of Pseudomonas aeruginosa. METHODS: The antibacterial property of C-dots was investigated by broth microdilution method against ATCC PAO1 and P. aeruginosa clinical isolates. The antibacterial effect of the C-dots and ciprofloxacin combination was investigated using the checkerboard method. The antibiofilm effect of the C-dots alone and its combination with ciprofloxacin was evaluated using the microtiter plate method. Subsequently, the toxicity of each agent was tested on L929 fibroblast cells. In the end, the effects of C-dots on the expression levels of pslA, pelA, and ppyR genes were determined using real-time quantitative PCR. RESULTS: The combination of C-dots and ciprofloxacin exhibited a synergistic effect. Additionally, this compound substantially decreased bacterial growth (P < 0.0001) and inhibited biofilm formation at MIC (96 µg/mL) and sub-MIC (48 µg/mL) concentrations (P < 0.0053, P < 0.01). After being exposed to C-dots at a concentration of 1mg/mL for 24 hours, the survival rate of L929 cells was 87.3%. The expression of genes pslA, pelA, and ppyR, associated with biofilm formation in P. aeruginosa, was significantly reduced upon exposure to C-dots (P < 0.0023). CONCLUSIONS: The findings demonstrate a promising new treatment method for infections. Furthermore, reducing the dosage of antibiotics can lead to an improvement in the toxic effects caused by dose-dependent antibiotics and antimicrobial activity.

Designing a novel multi-epitopes pan-vaccine against SARS-CoV-2 and seasonal influenza: in silico and immunoinformatics approach.

The merger of COVID-19 and seasonal influenza infections is considered a potentially serious threat to public health. These two viral agents can cause extensive and severe lower and upper respiratory tract infections with lung damage with host factors. Today, the development of vaccination has been shown to reduce the risk of hospitalization and mortality from the COVID-19 virus and influenza epidemics. Therefore, this study contributes to an immunoinformatics approach to producing a vaccine that can elicit strong and specific immune responses against COVID-19 and influenza A and B viruses. The NCBI, GISAID, and Uniprot databases were used to retrieve sequences. Linear B cell, Cytotoxic T lymphocyte, and Helper T lymphocyte epitopes were predicted using the online servers. Population coverage of MHC I epitopes worldwide for SARS-CoV-2, Influenza virus H3N2, H3N2, and Yamagata/Victoria were 99.93%, 68.67%, 68.38%, and 85.45%, respectively. Candidate epitopes were linked by GGGGS, GPGPG, and KK linkers. Different epitopes were permutated several times to form different peptides and then screened for antigenicity, allergenicity, and toxicity. The vaccine construct was analyzed for physicochemical properties, conformational B-cell epitopes, interaction with Toll-like receptors, and IFN-gamma-induced. Immune stimulation response of final construct was evaluated using C-IMMSIM. Eventually, the final construct sequence was codon-optimized for Escherichia coli K12 and Homo sapiens to design a multi-epitope vaccine and mRNA vaccine. In conclusion, due to the variable nature of SARS-CoV-2 and influenza proteins, the design of a multi-epitope vaccine can protect against all their standard variants, but laboratory validation is required.Communicated by Ramaswamy H. Sarma.

Potentially Infectious Helicobacter pylori in Tap Water in Kermanshah, Western Iran.

Background: Although the pathogenesis of Helicobacter pylori is well-defined, the origin and transmission of the bacterium have remained largely unknown. The water transmission hypothesis suggested that water acts as a carrier in oral-fecal transmission, especially in high-prevalence areas. We aimed to evaluate the possible contamination of tap water with infective H. pylori in Kermanshah, Iran from Sep-Oct 2020. Methods: Tap water samples were collected from varieties of probable high-alert regions and the viability of H. pylori were achieved using culture and real-time PCR techniques (ureA gene expression). Results: Out of 50 tap water samples, 3 were positive for H. pylori before enrichment and 6 were positive after enrichment by RT qPCR, while H. pylori colonies of two samples were observed on brucella agar plates. Conclusion: The results of positive samples demonstrated the probable presence of viable H. pylori in tap water samples, showing that tap water distribution systems could be a potential route for H. pylori transmission.

High burden of MDR, XDR, PDR, and MBL producing Gram negative bacteria causing infections in Kermanshah health centers during 2019-2020.

Background and Objectives: Microorganisms producing Metallo-Beta-Lactamase (MBL) are a threat and cause of concern as they have become one of the most feared resistance mechanisms. This study was designed to explore the prevalence of MBL production in clinical isolates of Gram negative bacteria using phenotypic MBL detection. Materials and Methods: A total of 248 isolates were collected from various clinical samples and were evaluated for carbapenem resistance and MBL production. All strains were screened for MBL production using Double Disk Confirmatory Test (DDCT). Results: The results of screening for MBL production using phenotypic disk diffusion method showed that in the 85 isolates were carbapenemase positive; including, 10 (16.1%) Klebsiella pneumoniae, 9 (14.5%) Escherichia coli, 58 (93.6%) Acinetobacter baumannii, and 8 (12.9%) Pseudomonas aeruginosa isolates. Also, 83 (97.6) Carbapenemase-producing isolates were resistant to at least four classes of antimicrobials (MDR). Conclusion: A. baumannii was the most common carbapenem resistant bacterium in medical centers in Kermanshah. Significant multiple drug resistance (MDR) incidence was observed compared to different classes of antibiotics.

In silico designing and immunoinformatics analysis of a novel peptide vaccine against metallo-beta-lactamase (VIM and IMP) variants.

The rapid spread of acquired metallo-beta-lactamases (MBLs) among gram negative pathogens is becoming a global concern. Improper use of broad-spectrum antibiotics can trigger the colonization and spread of resistant strains which lead to increased mortality and significant economic loss. In the present study, diverse immunoinformatic approaches are applied to design a potential epitope-based vaccine against VIM and IMP MBLs. The amino acid sequences of VIM and IMP variants were retrieved from the GenBank database. ABCpred and BCPred online Web servers were used to analyze linear B cell epitopes, while IEDB was used to determine the dominant T cell epitopes. Sequence validation, allergenicity, toxicity and physiochemical analysis were performed using web servers. Seven sequences were identified for linear B cell dominant epitopes and 4 sequences were considered as dominant CD4+ T cell epitopes, and the predicted epitopes were joined by KK and GPGPG linkers. Stabilized multi-epitope protein structure was obtained using molecular dynamics simulation. Molecular docking showed that the designed vaccine exhibited sustainable and strong binding interactions with Toll-like receptor 4 (TLR4). Finally, codon adaptation and in silico cloning studies were performed to design an effective vaccine production strategy. Immune simulation significantly provided high levels of immunoglobulins, T helper cells, T-cytotoxic cells and INF-γ. Even though the introduced vaccine candidate demonstrates a very potent immunogenic potential, but wet-lab validation is required to further assessment of the effectiveness of this proposed vaccine candidate.

Prevalence of Mycobacterium tuberculosis mutations associated with isoniazid and rifampicin resistance: A systematic review and meta-analysis.

Tuberculosis (TB) is still one of the leading causes of worldwide death, especially following the emergence of strains resistant to isoniazid (INH) and rifampicin (RIF). This study aimed to systematically review published articles focusing on the prevalence of INH and/or RIF resistance-associated mutations of Mycobacterium tuberculosis isolates in recent years. Literature databases were searched using appropriate keywords. The data of the included studies were extracted and used for a random-effects model meta-analysis. Of the initial 1442 studies, 29 were finally eligible to be included in the review. The overall resistance to INH and RIF was about 17.2% and 7.3%, respectively. There was no difference between the frequency of INH and RIF resistance using different phenotypic or genotypic methods. The INH and/or RIF resistance was higher in Asia. The S315T mutation in KatG (23.7 %), C-15 T in InhA (10.7 %), and S531L in RpoB (13.5 %) were the most prevalent mutations. Altogether, the results showed that due to S531L in RpoB, S315T in KatG, and C-15 T in InhA mutations INH- and RIF-resistant M. tuberculosis isolates were widely distributed. Thus, it would be diagnostically and epidemiologically beneficial to track these gene mutations among resistant isolates.

Global prevalence of Clostridioides difficile in 17,148 food samples from 2009 to 2019: a systematic review and meta-analysis.

BACKGROUND: Clostridioides (Clostridium) difficile is an important infectious pathogen, which causes mild-to-severe gastrointestinal infections by creating resistant spores and producing toxins. Spores contaminated foods might be one of the most significant transmission ways of C. difficile-associated infections. This systematic review and meta-analysis study were conducted to investigate the prevalence of C. difficile in food. METHODS: Articles that published the prevalence of C. difficile in food in PubMed, Web of Science, and Scopus databases were retrieved using selected keywords between January 2009 and December 2019. Finally, 17,148 food samples from 60 studies from 20 countries were evaluated. RESULTS: The overall prevalence of C. difficile in various foods was 6.3%. The highest and lowest levels of C. difficile contamination were detected to seafood (10.3%) and side dishes (0.8%), respectively. The prevalence of C. difficile was 4% in cooked food, 6.2% in cooked chicken and 10% in cooked seafood. CONCLUSIONS: There is still little known concerning the food-borne impact of C. difficile, but the reported contamination might pose a public health risk. Therefore, to improve the food safety and prevent contamination with C. difficile spores, it is necessary to observe hygienic issues during foods preparation, cooking and transfer.

Modified gold nanoparticle colorimetric probe-based biosensor for direct and rapid detection of Mycobacterium tuberculosis in sputum specimens.

The incidence of Mycobacterium tuberculosis (MTB) is increasing due to lack of appropriate diagnostic and therapeutic methods. Therefore, early and accurate detection of this bacteria plays a significant role in controlling tuberculosis. This study aimed to design, develop, and implement a direct and rapid detection method of MTB using modified gold nanoparticle (AuNP) colorimetric probe-based biosensor in sputum specimens. Spherical AuNPs were synthesized by the citrate reduction method and were functionalized using thiol-modified oligonucleotides (AuNP-biosensor). AuNP-biosensor and IS6110 PCR were compared to the gold standard in terms of analytical and clinical sensitivity and specificity, positive predictive value (PPV), negative predictive value (NPV), diagnostic odds ratio (DOR), and accuracy in 52 clinical specimens. Gold standard was defined as a positive result in concentrated sputum smear microscopy (SSM), culture, or Xpert MTB/RIF.The AuNP-biosensor had 100% sensitivity and specificity for detection of total sputum DNA in less than 15 min with ready-to-use AuNP-biosensor. PPV, NPV, DOR and accuracy of this method were 100%, 100%, 2325 and 100%, respectively. Considering the promising results of the diagnostic value indices of the AuNP-biosensor, the designed method is an affordable, rapid, reliable, and cost-beneficial way for direct detection of MTB in sputum specimens.

Designing multi-epitope vaccine against important colorectal cancer (CRC) associated pathogens based on immunoinformatics approach.

BACKGROUND: It seems that several members of intestinal gut microbiota like Streptococcus bovis, Bacteroides fragilis, Helicobacter pylori, Fusobacterium nucleatum, Enterococcus faecalis, Escherichia coli, Peptostreptococcus anaerobius may be considered as the causative agents of Colorectal Cancer (CRC). The present study used bioinformatics and immunoinformatics approaches to design a potential epitope-based multi-epitope vaccine to prevent CRC with optimal population coverage. METHODS: In this study, ten amino acid sequences of CRC-related pathogens were retrieved from the NCBI database. Three ABCpred, BCPREDS and LBtope online servers were considered for B cells prediction and the IEDB server for T cells (CD4+ and CD8+) prediction. Then, validation, allergenicity, toxicity and physicochemical analysis of all sequences were performed using web servers. A total of three linkers, AAY, GPGPG, and KK were used to bind CTL, HTL and BCL epitopes, respectively. In addition, the final construct was subjected to disulfide engineering, molecular docking, immune simulation and codon adaptation to design an effective vaccine production strategy. RESULTS: A total of 19 sequences of different lengths for linear B-cell epitopes, 19 and 18 sequences were considered as epitopes of CD4+ T and CD8+ cells, respectively. The predicted epitopes were joined by appropriate linkers because they play an important role in producing an extended conformation and protein folding. The final multi-epitope construct and Toll-like receptor 4 (TLR4) were evaluated by molecular docking, which revealed stable and strong binding interactions. Immunity simulation of the vaccine showed significantly high levels of immunoglobulins, helper T cells, cytotoxic T cells and INF-γ. CONCLUSION: Finally, the results showed that the designed multi-epitope vaccine could serve as an excellent prophylactic candidate against CRC-associated pathogens, but in vitro and animal studies are needed to justify our findings for its use as a possible preventive measure.

Variation analysis of SARS-CoV-2 complete sequences from Iran.

Aim: SARS-CoV-2 is an emerging coronavirus that was discovered in China and rapidly spread throughout the world. The authors looked at nucleotide and amino acid variations in SARS-CoV-2 genomes, as well as phylogenetic and evolutionary events in viral genomes, in Iran. Materials & methods: All SARS-CoV-2 sequences that were publicly released between the start of the pandemic and 15 October 2021 were included. Results: The majority of mutations were found in vaccine target proteins, Spike and Nucleocapsid proteins, and nonstructural proteins. The majority of the viruses that circulated in the early stages of the pandemic belonged to the B.4 lineage. Conclusion: We discovered the prevalence of viral populations in Iran. As a result, tracking the virus's variation in Iran and comparing it with a variety of nearby neighborhoods may reveal a pattern for future variant introductions.

Rapid Detection of vanA Resistance Gene from E. faecalis Clinical Isolates Using Duplex Loop-Mediated Isothermal Amplification and Triplex PCR Assay.

Today, the spread of vancomycin-resistant strains isolated from Enterococcus faecalis (E. faecalis) has become a major health concern worldwide. Therefore, it is essential to provide a rapid and sensitive assay for identifying vanA gene for timely and appropriate antimicrobial control of resistant enterococcal infections. For this purpose, a cross-sectional study was performed on different clinical specimens of enterococci from Imam Reza hospital, Kermanshah, Iran. The antimicrobial susceptibility testing was determined by disk diffusion and MIC methods. Triplex-PCR and duplex-LAMP assays were also used to identify vanA E. faecalis resistance gene isolates. The results of this study shown that out of 108 Enterococcus isolates, 86, 18, 2, 1, and one isolates of E. faecalis, E. faecium, E. avium, E. psudoavium, and E. raffinosus were identified, respectively. On the other hand, E. faecalis was confirmed in 87 and 88 isolates using duplex-LAMP and triplex PCR, respectively. The LAMP primer set designed in this study can reliably identify seven distinct regions of the vanA gene, and finally the sensitivity, specificity, and the positive and negative predictive values of LAMP assay were shown to be 94.19%, 72.73%, 76.19%, and 93.10%, respectively. In general, sample processing, isothermal reaction and result reporting were completed using the LAMP assay in 75 minutes. Our findings suggest that LAMP assay has been approved as an alternative to the vancomycin resistance Enterococcus genotype (vanA and vanB) compared to other methods and has the advantage of being rapid, time-consuming, and easy for diagnosis.

Characterization of the resistome in Lactobacillus genomic sequences from the human gut.

OBJECTIVES: The gut is a complex environment inhabited by a wide range of bacterial species. Lactobacillus species constitute a significant proportion of this environment and, due to their mobile genetic elements such as plasmids and transposons, are more likely to acquire and transfer antibiotic resistance genes through horizontal gene transfer (HGT). METHODS: The current study obtained and analysed 321 genome assemblies to determine the prevalence of intrinsic and acquired antibiotic resistance genes (ARGs) among Lactobacillus species colonizing the human gastrointestinal tract. RESULTS: A total of four high-frequency resistance genes were identified, including dfra42 (42%), poxtA (17.4%), lmrB (12%), and BJP-1 (7.7%); aside from dfra42, which is an intrinsic resistance gene, the other genes are acquired resistance genes. PoxtA was found in several different species, mainly in L. paracasei, whereas BJP-1 and lmrB were found in only one species, L. rhamnosus. IS5-like elements family transposase flanked 11% and 8% of detected lmrB and BJP-1, respectively, while a variety of insertion sequences surrounded 22% of identified poxtA. Furthermore, to the best of our knowledge, this is the first report of BJP-1 in lactobacilli that would suggest it has transferred from soil microbiota to humans. CONCLUSION: According to the 'One Health' perspective, early detection of a new reservoir would control the global spread of the antibiotic-resistant bacterial species among the three environments, which include humans, the environment, and animals. Finally, the study's findings may then highlight the possibility of lactobacilli acquiring or transmitting resistance to other species within or outside the human intestine.

Multilocus Sequence Typing for Molecular Epidemiology of Stenotrophomonas maltophilia Clinical and Environmental Isolates from a Tertiary Hospital in West of Iran

Background: Stenotrophomonas maltophilia is an opportunistic bacterium, contributing to different hospital-acquired infections and can be acquired from different hospital setting sources. Epidemiological study of S. maltophilia in the hospital also demonstrates the intrahospital distribution of certain strains of bacteria in healthcare facilities. The aim of the current study was to identify the molecular epidemiology of S. maltophilia isolates from clinical and environmental sources within a hospital. Methods: A total of 400 samples (clinical and environmental) were collected from the different settings of hospital. Following the standard biochemical testing and 23S rRNA genotyping, the molecular typing of S. maltophilia isolates was determined using the multilocus sequence typing (MLST) technique. Also, the frequencies of zot and entF virulence genes among S. maltophilia isolates were examined by PCR technique. Results: Based on the biochemical testes and PCRs targeting 23S rRNA gene, 22 S. maltophilia isolates were identified. The MLST analysis demonstrated that these isolates were assigned to 14 ST, and 6 out of 14 STs were common among clinical and environmental samples. All 22 isolates were identified in the PubMLST database. The PCR screening demonstrated that none of 22 S. maltophilia isolates had zot virulence gene, while the entF gene with the 59% frequency was observed in 13 out of 22 isolates. Among these 13 isolates, 6 STs were common in clinical and environmental isolates. Conclusion: Our study showed the clonal relatedness between clinical and environmental sources of the S. maltophilia isolates in a hospital. Further studies are required to understand the epidemic situation of this pathogen in the clinic and the environment.

Prevalence of Clostridioides difficile contamination in the healthcare environment and instruments: A systematic review and meta-analysis.

Introduction: Worldwide, Clostridioides difficile infection is becoming one of the most common healthcare-associated infections. Management and control of this infection in healthcare facilities are associated with screening for environmental and instrumental C. difficile contamination. This systematic review and meta-analysis aimed to assess the overall prevalence of C. difficile in hospital settings, medical devices, and instruments. Methods: Four main databases, PubMed, Web of Science, Google Scholar, and Scopus, were searched using the keywords Clostridioides difficile, Clostridium difficile, C. difficile, clostridia, Clostridium spp., hospital environments, antibiotic associate colitis, intensive care unit, and ward in combination as a search strategy. The PRISMA checklist was used for selecting eligible studies. Results: A total of 11 eligible articles published between 2012 and 2021 were included. The overall pooled prevalence of C. difficile in hospital environments was 14.9%. The highest and lowest prevalence were reported for India (51.1%) and the USA (1.6%), respectively. The highest prevalence was reported for beds (46.3%). A significant heterogeneity was seen between C. difficile prevalence in hospital environments in different samples. The highest and lowest prevalence was reported for floor corners (63.2%) and privacy curtains (1.4%), respectively. Conclusions: In conclusion, hospitals' medical devices and environmental surfaces are considered a crucial source of Clostridioides difficile infection. In this regard, we strongly recommend revising and improving the cleaning and disinfection methods in hospitals and quality control of cleaning adequacy.

A worldwide systematic review and meta-analysis of bacteria related to antibiotic-associated diarrhea in hospitalized patients.

INTRODUCTION: Antibiotic-associated diarrhea (AAD) is a major hospital problem and a common adverse effect of antibiotic treatment. The aim of this study was to investigate the prevalence of the most important bacteria that cause AAD in hospitalized patients. MATERIALS AND METHODS: PubMed, Web of Science and Scopus databases were searched using multiple relevant keywords and screening carried out based on inclusion/exclusion criteria from March 2001 to October 2021. The random-effects model was used to conduct the meta-analysis. RESULTS: Of the 7,377 identified articles, 56 met the inclusion criteria. Pooling all studies, the prevalence of Clostridioides (Clostridium) difficile, Clostridium perfringens, Klebsiella oxytoca, and Staphylococcus aureus as AAD-related bacteria among hospitalized patients were 19.6%, 14.9%, 27%, and 5.2%, respectively. The prevalence of all four bacteria was higher in Europe compared to other continents. The highest resistance of C. difficile was estimated to ciprofloxacin and the lowest resistances were reported to chloramphenicol, vancomycin, and metronidazole. There was no or little data on antibiotic resistance of other bacteria. CONCLUSIONS: The results of this study emphasize the need for a surveillance program, as well as timely public and hospital health measures in order to control and treat AAD infections.

Bacterial variation in the oral microbiota in multiple sclerosis patients.

BACKGROUND: Microorganisms in oral cavity are called oral microbiota, while microbiome consists of total genome content of microorganisms in a host. Interaction between host and microorganisms is important in nervous system development and nervous diseases such as Autism, Alzheimer, Parkinson and Multiple Sclerosis (MS). Bacterial infections, as an environmental factor in MS pathogenesis play role in T helper 17(Th17) increase and it enhancing the production of pro-inflammatory cytokines such as Interlukin-21(IL-21), IL-17 and IL -22. Oral microbiota consists diverse populations of cultivable and uncultivable bacterial species. Denaturing gradient gel electrophoresis (DGGE) is an acceptable method for identification of uncultivable bacteria. In this study, we compared the bacterial population diversity in the oral cavity between MS and healthy people. METHODS: From October to March 2019, samples were taken at Kermanshah University of Medical Sciences' MS patients center. A total of 30 samples were taken from MS patients and another 30 samples were taken from healthy people. Phenotypic tests were used to identify bacteria after pure cultures were obtained. DNA was extracted from 1 mL of saliva, and PCR products produced with primers were electrophoresed on polyacrylamide gels. RESULTS: The genera Staphylococcus, Actinomyces, Fusobacterium, Bacteroides, Porphyromonas, Prevotella, Veillonella, Propionibacterium and uncultivable bacteria with accession number MW880919-25, JQ477416.1, KF074888.1 and several other un-culturable strains were significantly more abundant in the MS group while Lactobacillus and Peptostreptococcus were more prevalent in the normal healthy group according to logistic regression method. CONCLUSION: Oral micro-organisms may alleviate or exacerbate inflammatory condition which impact MS disease pathogenesis. It may be assumed that controlling oral infections may result in reduction of MS disease progression.

In silico vaccine design and epitope mapping of New Delhi metallo-beta-lactamase (NDM): an immunoinformatics approach.

BACKGROUND: Antibiotic resistance is a global health crisis. The adage that "prevention is better than cure" is especially true regarding antibiotic resistance because the resistance appears and spreads much faster than the production of new antibiotics. Vaccination is an important strategy to fight infectious agents; however, this strategy has not attracted sufficient attention in antibiotic resistance prevention. New Delhi metallo-beta-lactamase (NDM) confers resistance to many beta-lactamases, including important carbapenems like imipenem. Our goal in this study is to use an immunoinformatics approach to develop a vaccine that can elicit strong and specific immune responses against NDMs that prevent the development of antibiotic-resistant bacteria. RESULTS: In this study, 2194 NDM sequences were aligned to obtain a conserved sequence. One continuous B cell epitope and three T cell CD4+ epitopes were selected from NDMs conserved sequence. Epitope conservancy for B cell and HLA-DR, HLA-DQ, and HLA-DP epitopes was 100.00%, 99.82%, 99.41%, and 99.86%, respectively, and population coverage of MHC II epitopes for the world was 99.91%. Permutation of the four epitope fragments resulted in 24 different peptides, of which 6 peptides were selected after toxicity, allergenicity, and antigenicity assessment. After primary vaccine design, only one vaccine sequence with the highest similarity with discontinuous B cell epitope in NDMs was selected. The final vaccine can bind to various Toll-like receptors (TLRs). The prediction implied that the vaccine would be stable with a good half-life. An immune simulation performed by the C-IMMSIM server predicted that two doses of vaccine injection can induce a strong immune response to NDMs. Finally, the GC-Content of the vaccine was designed very similar to E. coli K12. CONCLUSIONS: In this study, immunoinformatics strategies were used to design a vaccine against different NDM variants that could produce an effective immune response against this antibiotic-resistant factor.