Safety evaluation of mutagenicity, genotoxicity, and cytotoxicity of Lactobacillus spp. isolates as probiotic candidates.

BACKGROUND: Probiotics promote a healthy balance of gut bacteria and have many beneficial effects on human digestive physiology. Although, few side effects of probiotics have been reported. This study aimed to assess the safety of five probiotic candidate Lactobacillus strains isolated from healthy individuals by examining mutagenicity, genotoxicity, and oral toxic effects. METHODS: Five selected candidate probiotic (SCPs) strains were evaluated for genotoxicity (Ames test with Salmonella typhimurium), in vitro mammalian chromosome aberration test and an in vivo mouse micronucleus assay on peripheral blood of mice. To evaluate the oral dose toxicity, BALB/c mice models were treated repeatedly (2000, 1000, and 500 mg/kg body weight /day) for 28-days. RESULTS: The Ames test performed for two S. typhimurium strains TA 98 and TA100 (both in the absence and in the presence of S-9 metabolic activation system) did not show an increase in reverse mutation because of exposure to the SCPs in any of the doses (5.0, 2.5, 1.25, 0.625, and 0.3125 mg/plate). There was no genotoxicity in the SCPs treatment in the vitro chromosome aberration assay with Chinese hamster ovary cells (CHO-K1). In addition, none of the tested strains increased the frequency of micronucleated reticulocytes in reticulocytes, the SCPs with the studied doses caused no substantial variation in the experimental groups compared to the negative control group (p > 0.05). SCPs were not acutely toxic when administered to male and female BALB/c mice by single gavage at (2000, 1000, and 500 mg/kg b.w/day) with no mortality or clinical signs, change in body weight or macroscopic abnormalities were observed in this dose range. CONCLUSION: As a result, SCPs did not induce mutagenic potential in vitro with bacterial reverse mutation, clastogenicity, and in vivo tests in the ranges of concentrations evaluated in our study.

Bacteriocins: Properties and potential use as antimicrobials.

A variety of bacteriocins originate from lactic acid bacteria, which have recently been modified by scientists. Many strains of lactic acid bacteria related to food groups could produce bacteriocins or antibacterial proteins highly effective against foodborne pathogens such as Staphylococcus aureus, Pseudomonas fluorescens, P. aeruginosa, Salmonella typhi, Shigella flexneri, Listeria monocytogenes, Escherichia coli O157:H7, and Clostridium botulinum. A wide range of bacteria belonging primarily to the genera Bifidobacterium and Lactobacillus have been characterized with different health-promoting attributes. Extensive studies and in-depth understanding of these antimicrobials mechanisms of action could enable scientists to determine their production in specific probiotic lactic acid bacteria, as they are potentially crucial for the final preservation of functional foods or for medicinal applications. In this review study, the structure, classification, mode of operation, safety, and antibacterial properties of bacteriocins as well as their effect on foodborne pathogens and antibiotic-resistant bacteria were extensively studied.

Interesting probiotic traits of mother's milk Lactobacillus isolates; from bacteriocin to inflammatory bowel disease improvement.

AIMS AND BACKGROUND: Lactobacillus spp. are an important element in breast milk. This component has a beneficial effect on the composition of the intestinal microflora and the intestinal immune system. The aim of this study was to isolate and identify Lactobacillus strains in breast milk and evaluate some of their probiotic properties, such as presence of bacteriocin genes, adhesion to HT-29 cell line, competition with enteropathogens in cell culture, and effect on serum level of lipids and digestive enzymes, and mice model of inflammatory bowel disease (IBD). MATERIALS AND METHODS: A total of 323 lactic acid bacteria (LAB) were isolated from breast milk samples of healthy mothers with the age ranges from 21 to 45 years old. These isolates were subjected to phenotypic and molecular experiments. The frequency of bacteriocin genes was determined by polymerase chain reaction (PCR). Adhesion of Lactobacillus isolates to HT-29 cells was measured based on the number of attached bacterial cells in 20 fields of the light microscopy. Competition test was done by colony count and real-time PCR procedures. Five strongly adhesive Lactobacillus strains were selected and administered orally to the treatment groups. After 8 days, the serum level of digestive enzymes and improvement in induced IBD, and after 14 days, the serum level of lipids (triglycerides, total cholesterol, HDL, and LDL) in treated mice were surveyed compared to the control groups. RESULTS: Based on the phenotypic and molecular experiments, L. casei, L. plantarum, L. rhamnosus, and L. acidophilus strains were isolated and identified in the breast milk samples. The highest frequency of bacteriocin genes belonged to Plantaricin B (100%), followed by Plantaricin D (84.7%), Plantaricin G (84.7%), and Plantaricin EF (54.3%). Also, 71.8% of the isolates were strongly adhesive, 21.8% were non-adhesive, and 6.4% were adhesive. Lactobacillus strains had a significant effect on the displacement of enteropathogens. The in vitro cholesterol-removing ability of L. casei (L1), L. casei (L2), L. casei (L3), L. plantarum (L4), and L. rhamnosus (L5) was 3.5, 31.5, 21.3, 18.7, and 27.3%, respectively. The serum level of total cholesterol in the L. plantarum (L4) group as well as LDL in the L. casei (L3) (p = .0108) and L. rhamnosus (L5) (p = .0206) groups decreased significantly compared to the control group. The serum level of lipase increased in all the treatment groups compared to the control group, which was significant in the L. plantarum (L4) group (p = .0390). Disease activity index (DAI) scores were improved significantly in L. casei (L3) group compared to the IBD control group (p < .0001). CONCLUSION: It could be concluded that lactobacilli strains isolated from the breast milk samples had good probiotic properties, such as presence of bacteriocin genes, attaching to enterocyte-like HT-29 cells, competing with intestinal pathogens, lowering cholesterol, and improving IBD. Thus, after further studies, they could be considered as probiotic strains.

Analysis of virulence genes and molecular typing of Listeria monocytogenes isolates from human, food, and livestock from 2008 to 2016 in Iran.

The frequency of Listeria monocytogenes isolates collected from a total of 1150 samples including food (n = 300), livestock (n = 50), and human clinical (n = 800) was evaluated during 2008-2016. Antimicrobial resistance patterns, virulence factors, and molecular characteristics of these isolates were analyzed using disk diffusion method, sequencing, serotyping, and pulsed-field gel electrophoresis (PFGE). The analysis of 44 L. monocytogenes isolates showed that 72.7% (32 of 44) of all the isolates belonged to Serotype 1/2c, and 15.9% (7 of 44) belonged to Serotype 3c. All 44 isolates were resistant to one or more antimicrobial agents with the most frequent resistance to penicillin (75%) and tetracycline (47.7%). Of the 44 L. monocytogenes strains, 100, 69.2, and 62.5% of livestock, human, and food strains were resistant to penicillin, respectively. Using pulsed-field gel electrophoresis (PFGE) technique, the isolates' genetic diversity was determined, and 28 PFGE patterns with 8 common (CT) and 20 single types (ST) were identified. This study highlights the high prevalence of Serotype 1/2c in clinical and livestock samples, while different serotypes were observed in food samples. The presence of rare serotypes such as 4c, belonging to the Lineage III, as well as 4e and 1/2c which are infrequent in Iran indicates that paying attention to uncommon serotypes, especially 1/2c, during the listeriosis outbreaks is necessary.

The effect of improved formulation of chitosan-alginate microcapsules of Bifidobacteria on serum lipid profiles in mice.

BACKGROUND: Probiotics have been associated with many beneficial effects in human digestive physiology. The aim of this study was to evaluate the effect of improved formulation of chitosan-alginate microcapsules of Bifidobacterium strains on serum triglycerides, cholesterol, HDL, and LDL in mice. METHODS: Five approved probiotic strains of Bifidobacterium were tested for anti-proliferative effect and interleukin-8 induction on HT-29 cell lines. Bifidobacterium strains plus five approved Lactobacillus were encapsulated in chitosan-alginate microcapsules and tested for its survival in simulated gastrointestinal conditions. These microcapsules were administered to 4 groups of mice (including 1. Bif (Bifidobacterium strains), 2. Lac (Lactobacillus strains), 3. Bif-Lac (Bifidobacterium plus Lactobacillus strains) and 4. Control) for 8 days. At eighth day, the blood of mice were taken and serum levels of triglycerides, cholesterol, HDL, and LDL of them were determined. RESULTS: All of the Bifidobacterium strains significantly (P < 0.001) reduced secretion of IL-8 in HT-29 cells as well as maximum antiproliferative effects (P < 0.001). In addition, all microcapsules showed impressive survival rate in bile (>%94.1) and gastrointestinal (>%78.28) conditions (P < 0.05). Only Bif-Lac group displayed significantly lower serum cholesterol and LDL levels than control group (P < 0.05). Besides, all groups indicate statistically significant weight loss of mice during the 8 days in comparison with the control group (P < 0.05). CONCLUSION: The results of this study showed that the microencapsulated probiotics with alginate and chitosan had an effective mean of delivery of viable bacterial cells and non-pharmacological interventions use to reduce serum cholesterol and LDL levels in in-vivo condition.

Inhibitory effects of Lactobacilli cocktail on HT-29 colon carcinoma cells growth and modulation of the Notch and Wnt/ß-catenin signaling pathways.

Probiotics could be considered as attractive candidates for preventing tumor growth through maintaining homeostasis. The aim of this study was to evaluate the inhibitory effect of a cocktail of five Lactobacillus species on human colorectal carcinoma cell line HT-29. The anti-proliferative and apoptotic effects of Lactobacilli cocktail were evaluated using MTT and flow cytometry tests, respectively. Quantitative real-time polymerase chain reaction (qPCR) was used to analyze the expression of several genes in the Notch (notch, hes1, msi1, and numb) and Wnt/ß-catenin (CTNNB1 and CCND1) pathways, following the treatment of HT-29 cells with Lactobacilli cocktail. The treatment by Lactobacilli cocktail induced a significant anti-proliferative effect and late stage apoptosis among the cancer cells (p < 0.05). Compared to the untreated cells, Lactobacilli cocktail induced the down-regulation of notch, hes1, and msi1 genes and up-regulation of numb gene in the Notch pathway as well as the down-regulation of CTNNB1 and CCND1 genes in the Wnt/ß-catenin pathway in a time-dependent manner (p < 0.05). CONCLUSION: Lactobacilli cocktail was shown to have beneficial anti-tumor effects on HT-29 cells by modulating the Notch and Wnt/ß-catenin pathways; therefore, the use of Lactobacilli probiotics as nutritional supplements may prevent the incidence of colon cancer.

Lactobacillus species inhibitory effect on colorectal cancer progression through modulating the Wnt/ß-catenin signaling pathway.

Probiotic bacteria are known to exert a wide range of anticancer activities on their animal hosts. In the present study, the anticancer effect of a cocktail of several potential probiotic Lactobacillus species (potential probiotic L.C) was investigated in vitro and in vivo. MTT and Flow cytometry tests results showed that administration of live potential probiotic L.C significantly decreased the HT-29 and CT-26 cells proliferation and induced late apoptotis in a time-dependent manner. In addition, quantitative real-time polymerase chain reaction (qPCR) results showed that exposure of potential probiotic L.C to both HT-29 and CT-26 cells during the incubation times resulted in the upregulation (apc and CSNK1ε for HT-29, CSNK1ε and gsk3ß for CT-26) and downregulation (CTNNB1, CCND1, pygo2, axin2 and id2) of the Wnt/ß- catenin pathway-related genes in a time-dependent manner. The significance of in vitro anticancer effect of potential probiotic L.C was further confirmed in an experimental tumor model. Data from the murine model of colorectal cancer (CRC) induced by Azoxymethane (AOM) and Dextran Sulfate Sodium (DSS) showed significantly alleviated inflammation and tumor development in AOM/DSS/L.C-injected mice compared to the AOM/DSS-injected mice. Tumor growth inhibition was accompanied by potential probiotic L.C-driven upregulation and downregulation of the Wnt/ß-catenin pathway-related genes, similar to the in vitro results. These results showed that potential probiotic L.C inhibited the tumor growth, and that its anticancer activity was at least partially mediated through suppressing the Wnt/ß-catenin pathway. Overall, the present study suggested that this probiotic could be used clinically as a supplement for CRC prevention and treatment.

The effect of selected Lactobacillus strains on dextran sulfate sodium-induced mouse colitis model.

Inflammatory bowel disease (IBD) comprises two major illnesses: Crohn's disease (CD) and ulcerative colitis (UC). Dextran sulfate sodium (DSS) mouse colitis model has been used in understanding the mechanism of IBD. This study was conducted to examine selected Lactobacillus spp. as potential IBD treatment in the DSS-induced animal model. Balb/c mice were used and colitis was induced by adding 5% dextran sodium sulfate into the drinking water for 8 days. Colon length, disease activity index (DAI) and histological analysis were measured as markers of inflammation in DSS colitis mice. The majority of the Lactobacillus species significantly prevented the shortening of the colon length compared with the DSS group. The DAI scores of mice were significantly reduced following usage of four Lactobacillus strains included: Lactobacillus plantarum 03 and 06, Lactobacillus brevis 02 and Lactobacillus rhamnosus 01. The histological analysis exhibited that oral administration of Lactobacillus strains had therapeutic effects on mice colitis. L. plantarum and L. brevis showed better therapeutic effect against DSS-induced acute colitis mice. The probiotic activities of these three isolates indicated that the probiotic effects were strain specific and none of these useful bacteria could exhibit all of the valued probiotic properties simultaneously.

A review on anti-adhesion therapies of bacterial diseases.

BACKGROUND: Infections caused by bacteria are a foremost cause of morbidity and mortality in the world. The common strategy of treating bacterial infections is by local or systemic administration of antimicrobial agents. Currently, the increasing antibiotic resistance is a serious and global problem. Since the most important agent for infection is bacteria attaching to host cells, hence, new techniques and attractive approaches that interfere with the ability of the bacteria to adhere to tissues of the host or detach them from the tissues at the early stages of infection are good therapeutic strategies. METHODS: All available national and international databanks were searched using the search keywords. Here, we review various approaches to anti-adhesion therapy, including use of receptor and adhesion analogs, dietary constituents, sublethal concentrations of antibiotics, and adhesion-based vaccines. RESULTS: Altogether, the findings suggest that interference with bacterial adhesion serves as a new means to fight infectious diseases. CONCLUSION: Anti-adhesion-based therapies can be effective in prevention and treatment of bacterial infections, but further work is needed to elucidate underlying mechanisms.

Correction to: A review on anti-adhesion therapies of bacterial diseases.

The presentation of Table  was incorrect. The corrected table is given below.

Rapid Simultaneous Molecular Stool-Based Detection of Toxigenic Clostridioides difficile by Quantitative TaqMan Real-Time PCR Assay.

BACKGROUND: Clostridioides difficile is a major cause of nosocomial infectious diarrhea in hospitalized patients throughout the world. METHODS: A multiplex real-time PCR assay was developed and evaluated in comparison with toxigenic culture (TC) (as gold standard method) for direct detection of toxigenic C. difficile in fecal specimens. The multiplex real-time PCR assay simultaneously detected glutamate dehydrogenase (gluD), toxin A (tcdA), toxin B (tcdB), and binary toxin (cdtB) genes in stool samples. RESULTS: The results of multiplex real-time PCR were compared to those of the TC method in 250 patients suspected of C. difficile infection. The prevalence of positive TC was 13.6%. Forty-two stool samples (16.8%) were determined to be gluD+ using multiplex real-time PCR. These included 35 (83.3%) toxigenic (32 tcdA+, tcdB+ and three tcdB+) and 7 (20.0%) were cdtB+. The multiplex real-time PCR assay had a sensitivity of 91.45%, specificity of 99.54%, and positive and negative predictive values of 97% and 98.6%, respectively, compared to the TC method for diagnosis of C. difficile. The analytical sensitivity of the multiplex real-time PCR assay was estimated to be 102 CFU/g of stools and 0.0200 pg of genomic DNA from culture. The analytical specificity was determined to be 100% by using enteric and non-C. difficile standard bacterial strains. CONCLUSIONS: The molecular method developed in the study was rapid, sensitive, and specific for detection of toxigenic C. difficile. It is applicable to be performed in clinical laboratories and correlated well with the results obtained by TC.

Characterization of Clostridioides difficile isolates recovered from hospitalized patients and the hospitals environment and air: A multicenter study.

In healthcare settings, contamination of environment with toxigenic and hypervirulent Clostridioides difficile strains is a serious concern. Here, we assessed whether patients with C. difficile have a role to play in the dissemination of C. difficile in our settings or other sources are implicated in its circulation. A total of 700 fecal specimens and 1435 environmental samples from surfaces, equipment and air of rooms occupied by patients suspected of C. difficile infection were taken from 4 tertiary hospitals in Tehran, Iran between April 2016 and August 2017. Antibiotic susceptibility testing and detection of resistance genes were performed for the environmental isolates. The clinical and environmental isolates of C. difficile were subjected to Pulsed Field Gel Electrophoresis (PFGE) analysis. Forty three (6.14%) and 2 (0.13%) isolates of C. difficile were recovered from the clinical and environmental samples, respectively. In the clinical settings, 2 patients were suspected of recurrent C. difficile infection. Thirty distinct pulsotypes were found among the C. difficile isolates including 28 singletons and 2 common types. One of the two environmental isolates was isolated from floor in the Medical ward, of pulsotype/ribotype/toxinotype PT10/New ribotype/toxinotype V, harbored cdtA/B and tcdC-A, and resistant to ciprofloxacin. The other one was isolated from air of a room in ICU, assigned to PT11/RT001/toxinotype 0, belonged to tcdC-sc3 genotypes and resistant to metronidazole. The environmental isolates did not generate amplicons in PCR assays targeting vanA and nim genes. This study provided evidence for dissemination of genetically diverse strains of C. difficile in hospitalized patients, presence of C. difficile in hospital air, existence of binary toxin positive/antibiotic-resistant isolate on the floor and intra-hospital dissemination of this pathogen.

Characterization of bacteriocin production in Lactobacillus spp. isolated from mother's milk.

The purpose of the present study was to isolate Lactobacillus bacteria from mother's milk and to assess their probiotic potential. Sixty breast milk samples were collected from the volunteered mothers aged from 19 to 35 and from rural areas of Lorestan and Markazi Provinces, Iran. At first, 970 bacill-shaped bacterial colonies were isolated from these samples and stored in proper condition. Two hundred isolates were randomly selected and investigated for their ability to tolerate acidic condition and to tolerate bile salt as well. Only 33 isolates could withstand the exposure to low pH and bile salt. The isolates were identified using PCR primer specific to Lactobacillus and it was demonstrated that eighteen of thirty-three isolates were belonged to the Lactobacillus. Among the isolates, 16 and 2 of them were Lactobacillus reuteri and L. gasseri, respectively. In addition, the antibiotic resistance of the isolates was determined using disc diffusion method and all of the isolates were shown to be sensitive to eight out of the twelve investigated antibiotics. Moreover, the antagonistic effect of the isolates was inspected on ten indicator pathogens. Interestingly, all of the pathogenic bacteria were inhibited by Lactobacillus isolates. In addition, to partially understand the nature of inhibition mechanism, well diffusion deployed for two randomly-selected indicator bacteria and the resulting halos of three isolates were statistically significant compared to other lactobacillus (p < 0.05). Subsequently, bacteriocin genes (plnS, Laf, gasA) were identified by PCR among the isolates. The results showed that only 2 isolates possessed the gasA gene which were in accordance with well diffusion test. Consequently, eighteen Lactobacillus isolated from breast milk samples which all of them were able to tolerate low pH and bile salt. Similarly, all of the Lactobacillus isolates were proved to inhibit the growth of pathogen strains and two of them possess a bacteriocin-related gene.

Improvement of semen parameters after antibiotic therapy in asymptomatic infertile men infected with Mycoplasma genitalium.

OBJECTIVES: To elucidate the association between asymptomatic infections caused by Mycoplasma genitalium and male infertility, and evaluate the role of antibiotic therapy in treatment of this failure. METHODS: A total of 165 infertile males having abnormal semen parameters (study group) and 165 healthy fertile men (control group) were included. Semen samples were taken from all participants and after analyzing for semen parameters, undergone real-time PCR, microbial culture, and reactive oxygen species (ROS), as well as total antioxidant capacity (TAC) assays. Infected individuals of study group were treated with antibiotic. One month after the treatment completion, second semen samples were taken and subjected to all the tests mentioned. The data were analyzed using SPSS statistical software, version 22.0. RESULTS: The frequency of M. genitalium was significantly higher in the infertile men compared with the fertile ones (9.7% vs. 1.2%; p = 0.001). Mean cycle threshold (C t) value was lower in infected infertile than infected fertile men (p < 0.001). All semen parameters, except volume, pH, and viscosity, were improved (p < 0.05), most of which reached their normal range; leukocytes in seminal fluid decreased (p = 0.02), the level of TAC was elevated (p = 0.002), and ROS level as well as ROS/TAC ratio reduced after antibiotic treatment (p = 0.03). Wives of seven infected infertile men (43.8%) became pregnant 4 months after the treatment completion. CONCLUSIONS: Asymptomatic infection caused by M. genitalium is correlated with male infertility and antibiotic therapy can improve the semen quality and be used to treat male infertility.

Bifidobacterium obtained from mother's milk and their infant stool; A comparative genotyping and antibacterial analysis.

Antibacterial activity of Bifidobacterium species has been considered as an important probiotic property for development of human gut immunity. This study was conducted to assess the genotypes and antibacterial activities of the native Bifidobacterium isolates obtained from the human's breast milk and the feces of their paired infants. Fifty-six samples from twenty-eight mothers' milk and paired infants feces were collected and cultured. Suspicious colonies were picked up and confirmed by phenotypic and molecular identifications. Randomly amplified polymorphic DNA (RAPD-PCR) and antibacterial activity were carried out. Amongst 56 samples, 41 different Bifidobacterium species including 12 B. breve, 14 B. longum, and 15 B. bifidum were isolated. Out of which, 12 isolates including B. longum (6), B. breve (4) and B. bifidum (2) were shared between six mother-infant pairs. Only three strains of B. longum showed 100% similarity in their RAPD-PCR. No significant difference was observed in the antibacterial activity of the Bifidobacterium isolates, with the same or different RAPD-PCR profile, against the enteric bacteria. Overall, 29% of the Bifidobacteria species isolated from the mothers milk and their paired infants feces were shared. All species of Bifidobacteria showed the universal role of antipathogens activities irrespective of the host and the isolation site.

The association of type II diabetes with gut microbiota composition.

It is known that type 2 diabetes (T2D) in humans could be linked to the composition of gut microbiota. The aim of this study was to evaluate three faecal bacterial species, including Bacteroides fragilis, Bifidobacterium longum and Faecalibacterium prausnitzii in patients with T2D. This case control study included 18 patients with T2D and 18 matched persons without diabetes. The concentrations of B. fragilis, B. longum and F. prausnitzii were determined by quantitative Real-Time PCR. Quantitative PCR analysis revealed that the gut bacterial composition in patients with T2D was partially different from that in the healthy individuals. Faecalibacterium prausnitzii was significantly lower in patients with T2D (P-value = 0.038). Bacteroides fragilis was under-represented in the microbiota of the group with diabetes, but its difference between two groups was not significant (P-value = 0.38). No difference was observed for B. longum community between the both groups (P-value = 0.99). Characterization of specific species of intestinal microbiota shows some compositional changes in patients with T2D. The results may be valuable for developing strategies to control type 2 diabetes by modifying the intestinal microbiota. Long-term studies with emphasis on other bacterial groups are suggested to clarify the association of T2D with gut microbiota.

Antibacterial potential and genetic profile of Enterococcus faecium strains isolated from human normal flora.

Enterococci have a widespread attendance in the circumference and belongs to the enteric commensal microbiota. Most of them produce the antimicrobial compounds and have an inhibition effect on pathogenic microorganisms. The objective of this study was to characterize the enterococcal strains isolated from human normal flora and assess their antibacterial activity. Enterococcal isolates were obtained from the feces of eighteen healthy humans. All enterococcal species were identified by biochemical and species-specific polymerase chain reaction (PCR). These isolates were investigated further to examine their ability to inhibit growth of Salmonella typhi, Shigella flexneri and Escherichia coli by well diffusion assay. Furthermore, antibiotic susceptibility test was performed and genetic relatedness of all isolates was evaluated by Pulse Field Gel Electrophoresis (PFGE). In all, 432 isolates were obtained from fecal samples. All of the isolates identified as Enterococcus faecium by biochemical and molecular (PCR) methods. Using repetitive element palindromic (REP)-PCR method 54 patterns have been obtained and were selected for further evaluation. The results indicated that 66%, 38% and 24% of our isolates had antimicrobial effect against S. typhi, S flexneri and enteroaggregative Escherichia coli (EAEC), respectively. On the other hand, there was no significant inhibition effect against enteropathogenic E. coli (EPEC) and enterotoxigenic E. coli (ETEC). All isolates were sensitive to vancomycin, teicoplanin, linezolid, ampicillin, chloramphenicol and gentamicin. On the other hand, the resistance rates for erythromycin, tetracycline and ciprofloxacin were 20%, 22%, and 1.8% respectively. In addition, the analysis of PFGE showed forty patterns with eight (40.7%) common types (CT) and thirty two (59.2%) single types (ST). Among eight common types, only one common type (CT5) had similar antimicrobial effect. These results suggested that enterococcal isolates obtained from human normal flora have potential antibacterial effect against S. typhi, S. flexneri and E. coli.

Molecular characterization of vancomycin-resistant Enterococcus faecium isolated from intensive care units.

To assess the molecular characterization of disseminated vancomycin-resistant enterococci (VRE) in the intensive care units, 546 enterococci isolates were collected from different clinical samples in a prospective observational study. The results showed that a total number of 33 isolates (6 %) were resistant to vancomycin. Most of the VRE isolates 11 (34 %) were isolated from intensive care units (ICUs). 18 (55 %) VRE isolates were obtained from urinary tract infections. The results from pulsed-field gel electrophoresis showed five common types (CT) and 13 single types (ST) among the VRE isolates. The analysis showed two and one major CTs and ST among the ICUs isolates, respectively. Tn1546 transposon was analyzed using ClaI-digested long PCR (L-PCR) RFLP followed by sequencing. The results showed the presence of two different lineages of transposon among the two clonal groups. Lineage 1 with the arrangement of Tn1546 prototype in the first clonal group and the second lineage with 13 kb harboring two insertion sequences, IS1216 V and IS1542. DNA hybridization showed that vanA gene in all VRE isolates, with an exception of one isolate, was present in the same location on the genome. Overall, the results suggest that a few VRE clonal types were disseminated in ICUs in hospitals in Iran which were able to transfer their vanA with high conjugation frequency.

Investigating the antimicrobial and anti-inflammatory effects of Lactobacillus and Bifidobacterium spp. on cariogenic and periodontitis pathogens.

Background: The use of probiotics is emerging as an innovative approach to managing oral health issues and mediating the immune system. The current study assessed the in vitro impacts of non-orally isolated probiotics on periodontitis and tooth decay pathogens. Methods: Briefly, the persistence of probiotics in exposure to oral cavity enzymes, hydrogen peroxide, and saliva samples was examined. It was also investigated the biofilm formation and aggregation ability of probiotics, the adherence of probiotics in human gingival fibroblast cell (HGFC) lines and molar teeth samples, and the potential of probiotics to co-aggregate with oral pathogens. Additionally, the current study evaluated the effects of live probiotics on virulence gene expression, biofilm production of main oral pathogens, and changes in inflammation markers. Results: The probiotics remained alive when exposed to enzymes in the oral cavity, hydrogen peroxide, and saliva at baseline, 1, 3, and 5 h after incubation at 37°C (p-value <0.05). Probiotics demonstrated to produce biofilm and aggregation, as well as adherence to HGFCs and maxillary molars (p-value >0.05). They showed significant co-aggregation with oral pathogens, which were recorded as 65.57% for B. bifidum 1001 with S. mutans, 50.06% for B. bifidum 1005 with P. gingivalis, 35.6% for L. plantarum 156 with F. nucleatum, and 18.7% for B. longum 1044 with A. actinomycetemcomitans after 8 h of incubation. A balance between pro-inflammatory and anti-inflammatory cytokines, along with inhibition of biofilm formation and changes in virulence gene transcripts, were observed. However, most of these changes were not statistically significant (p-value >0.05). Conclusion: This study demonstrated the direct link between adhesiveness, aggregation, and biofilm formation with probiotic antibacterial activity. In addition to the careful selection of suitable probiotic strains, the concentration and origin of probiotic isolates should be considered.

Exploring the interplay between Fusobacterium nucleatum with the expression of microRNA, and inflammatory mediators in colorectal cancer.

Background: Fusobacterium nucleatum has been recognized as an important key bacterium in the cause and spread of colorectal carcinogenesis. Nevertheless, the clinical relevance of F. nucleatum in colorectal cancer (CRC) and its effect on immune factors and the tumor microenvironment have not been fully elucidated. Materials and methods: The frequency of F. nucleatum was measured in 100 paired tumor and normal tissue specimens by TaqMan quantification Real-Time Polymerase Chain Reaction (qPCR). The mRNA expression levels of cytokines (IL-6, IL-10, IL-12ß, IL-17, TNF-α, TLR-2, and TLR-4), and miRNAs (miR-21, miR-31) were examined. Eventually, any potential correlations between the molecular and clinicopathological features of the neoplastic samples and the abundance of F. nucleatum were analyzed. Results: The relative frequency of F. nucleatum was significantly increased in cancerous tissue compared to adjacent non-tumor tissues. Furthermore, the high level of F. nucleatum was significantly associated with histological grade III and IV CRC tissues (P = 0.027 and P = 0.022, respectively) and perineural invasion-positive patients (P = 0.037). In addition, the expression levels of IL-6, IL-17, TNF-α,IL-12ß, TLR-2, and TLR-4 as well as miR-21 and miR-31 showed a significant increase in the cancer group. A notable correlation was also observed between the high status of F. nucleatum and the expression of IL-6, TNF-α and miR-21. Conclusion: Our results emphasize the importance of F. nucleatum and changes in the expression of genes involved in CRC. Studying the microbial profile and gene expression changes in CRC patients may be a promising approach to improve screening methods and provide therapeutic strategies.