Lantibiotic Nisin and Its Detection Methods

The type A lantibiotic nisin produced by several Lactococcus lactis strains, and one Streptococcus uberis strainis a small antimicrobial peptide that inhibits the growth of a wide range of gram-positive bacteria, such as Bacillus, Clostridium, Listeria and Staphylococcus species. It is nontoxic to humans and used as a food preservative (E234) in more than 50 countries including the EU, the USA, and China. National legislations concerning maximum addition levels of nisin in different foods vary greatly. Therefore, there is a demand for non-laborious and sensitive methods to identify and quantify nisin reliably from different food matrices. The horizontal inhibition assay, based on the inhibitory effect of nisin to Micrococcus luteus is the base for most quantification methods developed so far. However, the sensitivity and accuracy of the agar diffusion method is affected by several parameters. Immunological tests have also been described. Taken into account the sensitivity of immunological methods to interfering substances within sample matrices, and possible cross-reactivities with lantibiotics structurally close to nisin, their usefulness for nisin detection from food samples remains limited. The proteins responsible for nisin biosynthesis, and producer self-immunity are encoded by genes arranged into two inducible operons, nisA/Z/QBTCIPRK and nisFEG, which also contain internal, constitutive promoters PnisI and PnisR. The transmembrane histidine kinase NisK and the response regulator NisR form a two-component signal transduction system, in which NisK autophosphorylates after exposure to extra cellular nisin, and subsequently transfers the phosphate to NisR. The phosphorylated NisR then relays the signal downstream by binding to two regulated promoters in the nisin gene cluster, i.e the nisA/Z/Qand the nisF promoters, thus activating transcription of the structural gene nisA/Z/Q and the downstream genes nisBTCIPRK from the nisA/Z/Q promoter, and the genes nisFEG from the nisF promoter. In this work two novel and highly sensitive nisin bioassays were developed. Both of these quantification methods were based on NisRK mediated, nisin induced Green Fluorescent Protein (GFP) fluorescence. The suitabilities of these assays for quantifica¬tion of nisin from food samples were evaluated in several food matrices. These bioassays had nisin sensitivities in the nanogram or picogram levels. In addition, shelf life of nisin in cooked sausages and retainment of the induction activity of nisin in intestinal chyme (intestinal content) was assessed.Nisiini on eräiden Lactococcus lactis -maitohappobakteerikantojen tuottama antimikrobinen peptidi. Nisiini inhiboi useiden gram-positiivisten elintarvikepatogeenien, kuten Bacillus, Clostridium, Listeria ja Staphylococcus -sukujen kasvua. Nisiini on ihmisille vaaraton, ja siksi sitä käytetään elintarvikelisäaineena (E234) yli 50 maassa, mukaan lukien EU, USA ja Kiina. Nisiinin elintarvikekäyttöä koskevat lait ja säädökset vaihtelevat huomattavasti eri maissa. Tästä syystä on tarpeellista kehittää nisiinimittausmenetelmiä, jotka ovat sekä luotettavia, herkkiä, yksinkertaisia suorittaa, että nisiinispesifisiä. Yhä edelleen eniten käytetty nisiinin mittausmenetelmä, agardiffuusiomenetelmä, on epätarkka, antaa vääriä positiivisia tuloksia ja on työläs suorittaa. Nisiinin synteesiin ja nisiinituottajan immuniteetin kehittämiseen osallistuvat geenit ovat järjestäytyneet kahdeksi nisiini-indusoituvaksi operoniksi, nisA/Z/QBTCIPRK ja nisFEG. Tämän työn kannalta keskeiset nisiinigeenit olivat nisRK: proteiinit NisRK muodostavat kaksikomponenttisen signaalivälitysjärjestelmän, jossa solukalvoassosioitunut histidiinikinaasi NisK fosforyloi NisR-proteiinin, kun solujen kasvuympäristössä on nisiiniä. Fosforyloitunut NisR puolestaan toimii transkriptioaktivaattorina, aktivoiden nisiinioperonien sisältämien nisiini-indusoituvien nisA/Z/Q ja nisFEG promoottorien säätelemien geenien ekspressiota. Tässä työssä kehitettiin kaksi uutta ja erityisen herkkää nisiinin mittausmenetelmää. Molemmissa menetelmissä hyödynnetään NisRK signaalivälitysjärjestelmää siten, että solun ulkopuolinen nisiini aktivoi vihreän fluoresoivan proteiinin tuottoa. Menetelmien soveltuvuus nisiinin mittaamiseen elintarvikenäytteistä verifioitiin useissa elintarvikematriiseissa. Lisäksi kehitettyjen menetelmien avulla määritettiin nisiinin puoliintumisaika sekä keitetyissä makkaroissa että suolisto-oloissa

No differences in infections between patient-specific implants and conventional mini-plates in mandibular bilateral sagittal split osteotomy - Up to 3-year follow-up

The use of individually designed osteotomies, combined with individually manufactured osteosynthesis material, is rapidly becoming a standard for more challenging maxillofacial surgery. The benefits of patient-specific implants (PSI) in orthognathic surgery are clear in complex cases. PSIs can enhance precision and ease up the surgical protocol. We previously reported on the benefits of PSIs as reposition and fixation systems during Le Fort I osteotomy. The aim of this study was to evaluate a cohort of 28 patients, treated with bilateral sagittal split osteotomy (BSSO) and PSIs for fixation, with regard to healing for up to 3 years. A retrospective cohort of 48 patients with conventional mini-plate repositioned mandibles was also collected for statistical analysis. No statistically significant differences were found with regard to infection, soft tissue problems, or reoperations between these two groups. (C) 2018 European Association for Cranio-Maxillo-Facial Surgery. (C) Published by Elsevier Ltd. All rights reserved.Peer reviewe

Microbiota and Extracellular Vesicles in Anti-PD-1/PD-L1 Therapy

Cancer is a deadly disease worldwide. In light of the requisite of convincing therapeutic methods for cancer, immune checkpoint inhibition methods such as anti-PD-1/PD-L1 therapy appear promising. Human microbiota have been exhibited to regulate susceptibility to cancer as well as the response to anti-PD-1/PD-L1 therapy. However, the probable contribution of bacterial extracellular vesicles (bEVs) in cancer pathophysiology and treatment has not been investigated much. bEVs illustrate the ability to cross physiological barriers, assemble around the tumor cells, and likely modify the tumor microenvironment (EVs). This systematic review emphasizes the correlation between cancer-associated extracellular vesicles, particularly bEVs and the efficacy of anti-PD-1/PD-L1 therapy. The clinical and pharmacological prospective of bEVs in revamping the contemporary treatments for cancer has been further discussed

Surface Roughness and Streptococcus mutans Adhesion on Metallic and Ceramic Fixed Prosthodontic Materials after Scaling

The aim of this study was to evaluate the surface roughness of fixed prosthodontic materials after polishing or roughening with a stainless steel curette or ultrasonic scaler and to examine the effect of these on Streptococcus mutans adhesion and biofilm accumulation. Thirty specimens (10 x 10 x 3 mm(3)) of zirconia (Zr), pressed lithium disilicate (LDS-Press), milled lithium disilicate glazed (LDS-Glaze), titanium grade V (Ti) and cobalt-chromium (CoCr) were divided into three groups (n = 10) according to surface treatment: polished (C), roughened with stainless steel curette (SC), roughened with ultrasonic scaler (US). Surface roughness values (Sa, Sq) were measured with a spinning disc confocal microscope, and contact angles and surface free energy (SFE) were measured with a contact angle meter. The specimens were covered with sterilized human saliva and immersed into Streptococcus mutans suspensions for bacterial adhesion. The biofilm was allowed to form for 24 h. Sa values were in the range of 0.008-0.139 mu m depending on the material and surface treatment. Curette and ultrasonic scaling increased the surface roughness in LDS-Glaze (p <0.05), Ti (p <0.01) and CoCr (p <0.001), however, surface roughness did not affect bacterial adhesion. Zr C and US had a higher bacterial adhesion percentage compared to LDS-Glaze C and US (p = 0.03). There were no differences between study materials in terms of biofilm accumulation.Peer reviewe

Akkermansia muciniphila Adheres to Enterocytes and Strengthens the Integrity of the Epithelial Cell Layer

Akkermansia muciniphila is a Gram-negative mucin-degrading bacterium that resides in the gastrointestinal tracts of humans and animals. A. muciniphila has been linked with intestinal health and improved metabolic status in obese and type 2 diabetic subjects. Specifically, A. muciniphila has been shown to reduce high-fat-diet-induced endotoxemia, which develops as a result of an impaired gut barrier. Despite the accumulating evidence of the health-promoting effects of A. muciniphila, the mechanisms of interaction of the bacterium with the host have received little attention. In this study, we used several in vitro models to investigate the adhesion of A. muciniphila to the intestinal epithelium and its interaction with the host mucosa. We found that A. muciniphila adheres strongly to the Caco-2 and HT-29 human colonic cell lines but not to human colonic mucus. In addition, A. muciniphila showed binding to the extracellular matrix protein laminin but not to collagen I or IV, fibronectin, or fetuin. Importantly, A. muciniphila improved enterocyte monolayer integrity, as shown by a significant increase in the transepithelial electrical resistance (TER) of cocultures of Caco-2 cells with the bacterium. Further, A. muciniphila induced interleukin 8 (IL-8) production by enterocytes at cell concentrations 100-fold higher than those for Escherichia coli, suggesting a very low level of proinflammatory activity in the epithelium. In conclusion, our results demonstrate that A. muciniphila adheres to the intestinal epithelium and strengthens enterocyte monolayer integrity in vitro, suggesting an ability to fortify an impaired gut barrier. These results support earlier associative in vivo studies and provide insights into the interaction of A. muciniphila with the host.Peer reviewe

Effect of Surface Tooling Techniques of Medical Titanium Implants on Bacterial Biofilm Formation In Vitro

The aim of this study was to assess the biofilm formation of Streptococcus mutans, Staphylococcus aureus, Enterococcus faecalis, and Escherichia coli on titanium implants with CAD-CAM tooling techniques. Twenty specimens of titanium were studied: Titanium grade 2 tooled with a Planmeca CAD-CAM milling device (TiGrade 2), Ti6Al4V grade 5 as it comes from CAD-DMLS device (computer aided design-direct metal laser sintering device) (TiGrade 5), Ti6Al4V grade 23 as it comes from a CAD-CAM milling device (TiGrade 23), and CAD-DMLS TiGrade 5 polished with an abrasive disc (TiGrade 5 polished). Bacterial adhesion on the implants was completed with and without saliva treatment to mimic both extraoral and intraoral surgical methods of implant placement. Five specimens/implant types were used in the bacterial adhesion experiments. Autoclaved implant specimens were placed in petri plates and immersed in saliva solution for 30 min at room temperature and then washed 3× with 1× PBS. Bacterial suspensions of each strain were made and added to the specimens after saliva treatment. Biofilm was allowed to form for 24 h at 37 °C and the adhered bacteria was calculated. Tooling techniques had an insignificant effect on the bacterial adhesion by all the bacterial strains studied. However, there was a significant difference in biofilm formation between the saliva-treated and non-saliva-treated implants. Saliva contamination enhanced S. mutans, S. aureus, and E. faecalis adhesion in all material types studied. S. aureus was found to be the most adherent strain in the saliva-treated group, whereas E. coli was the most adherent strain in the non-saliva-treated group. In conclusion, CAD-CAM tooling techniques have little effect on bacterial adhesion. Saliva coating enhances the biofilm formation; therefore, saliva contamination of the implant must be minimized during implant placement. Further extensive studies are needed to evaluate the effects of surface treatments of the titanium implant on soft tissue response and to prevent the factors causing implant infection and failure

Effect of Surface Tooling Techniques of Medical Titanium Implants on Bacterial Biofilm Formation In Vitro

The aim of this study was to assess the biofilm formation of Streptococcus mutans, Staphylococcus aureus, Enterococcus faecalis, and Escherichia coli on titanium implants with CAD-CAM tooling techniques. Twenty specimens of titanium were studied: Titanium grade 2 tooled with a Planmeca CAD-CAM milling device (TiGrade 2), Ti6Al4V grade 5 as it comes from CAD-DMLS device (computer aided design-direct metal laser sintering device) (TiGrade 5), Ti6Al4V grade 23 as it comes from a CAD-CAM milling device (TiGrade 23), and CAD-DMLS TiGrade 5 polished with an abrasive disc (TiGrade 5 polished). Bacterial adhesion on the implants was completed with and without saliva treatment to mimic both extraoral and intraoral surgical methods of implant placement. Five specimens/implant types were used in the bacterial adhesion experiments. Autoclaved implant specimens were placed in petri plates and immersed in saliva solution for 30 min at room temperature and then washed 3x with 1x PBS. Bacterial suspensions of each strain were made and added to the specimens after saliva treatment. Biofilm was allowed to form for 24 h at 37 degrees C and the adhered bacteria was calculated. Tooling techniques had an insignificant effect on the bacterial adhesion by all the bacterial strains studied. However, there was a significant difference in biofilm formation between the saliva-treated and non-saliva-treated implants. Saliva contamination enhanced S. mutans, S. aureus, and E. faecalis adhesion in all material types studied. S. aureus was found to be the most adherent strain in the saliva-treated group, whereas E. coli was the most adherent strain in the non-saliva-treated group. In conclusion, CAD-CAM tooling techniques have little effect on bacterial adhesion. Saliva coating enhances the biofilm formation; therefore, saliva contamination of the implant must be minimized during implant placement. Further extensive studies are needed to evaluate the effects of surface treatments of the titanium implant on soft tissue response and to prevent the factors causing implant infection and failure.Peer reviewe

Effect of Surface Tooling Techniques of Medical Titanium Implants on Bacterial Biofilm Formation In Vitro

The aim of this study was to assess the biofilm formation of Streptococcus mutans, Staphylococcus aureus, Enterococcus faecalis, and Escherichia coli on titanium implants with CAD-CAM tooling techniques. Twenty specimens of titanium were studied: Titanium grade 2 tooled with a Planmeca CAD-CAM milling device (TiGrade 2), Ti6Al4V grade 5 as it comes from CAD-DMLS device (computer aided design-direct metal laser sintering device) (TiGrade 5), Ti6Al4V grade 23 as it comes from a CAD-CAM milling device (TiGrade 23), and CAD-DMLS TiGrade 5 polished with an abrasive disc (TiGrade 5 polished). Bacterial adhesion on the implants was completed with and without saliva treatment to mimic both extraoral and intraoral surgical methods of implant placement. Five specimens/implant types were used in the bacterial adhesion experiments. Autoclaved implant specimens were placed in petri plates and immersed in saliva solution for 30 min at room temperature and then washed 3× with 1× PBS. Bacterial suspensions of each strain were made and added to the specimens after saliva treatment. Biofilm was allowed to form for 24 h at 37 °C and the adhered bacteria was calculated. Tooling techniques had an insignificant effect on the bacterial adhesion by all the bacterial strains studied. However, there was a significant difference in biofilm formation between the saliva-treated and non-saliva-treated implants. Saliva contamination enhanced S. mutans, S. aureus, and E. faecalis adhesion in all material types studied. S. aureus was found to be the most adherent strain in the saliva-treated group, whereas E. coli was the most adherent strain in the non-saliva-treated group. In conclusion, CAD-CAM tooling techniques have little effect on bacterial adhesion. Saliva coating enhances the biofilm formation; therefore, saliva contamination of the implant must be minimized during implant placement. Further extensive studies are needed to evaluate the effects of surface treatments of the titanium implant on soft tissue response and to prevent the factors causing implant infection and failure

Characterization of Highly Mucus-Adherent Non-GMO Derivatives of Lacticaseibacillus rhamnosus GG

Lacticaseibacillus rhamnosusGG is one of the best studied lactic acid bacteria in the context of probiotic effects.L. rhamnosusGG has been shown to prevent diarrhea in children and adults and has been implicated to have mitigating or preventive effects in several disorders connected to microbiota dysbiosis. The probiotic effects are largely attributed to its adhesive heterotrimeric sortase-dependent pili, encoded by thespaCBA-srtC1gene cluster. Indeed, the strain-specific SpaCBA pili have been shown to contribute to adherence, biofilm formation and host signaling. In this work we set out to generate non-GMO derivatives ofL. rhamnosusGG that adhere stronger to mucus compared to the wild-type strain using chemical mutagenesis. We selected 13 derivatives that showed an increased mucus-adherent phenotype. Deep shotgun resequencing of the strains enabled division of the strains into three classes, two of which revealed SNPs (single nucleotide polymorphisms) in thespaAandspaCgenes encoding the shaft and tip adhesive pilins, respectively. Strikingly, the other class derivatives demonstrated less clear genotype - phenotype relationships, illustrating that pili biogenesis and structure is also affected by other processes. Further characterization of the different classes of derivatives was performed by PacBio SMRT sequencing and RNAseq analysis, which resulted in the identification of molecular candidates driving pilin biosynthesis and functionality. In conclusion, we report on the generation and characterization of three classes of strongly adherentL. rhamnosusGG derivatives that show an increase in adhesion to mucus. These are of special interest as they provide a window on processes and genes driving piliation and its control inL. rhamnosusGG and offer a variety of non-GMO derivatives of this key probiotic strain that are applicable in food products.Peer reviewe

The canine isolate Lactobacillus acidophilus LAB20 adheres to intestinal epithelium and attenuates LPS-induced IL-8 secretion of enterocytes in vitro

Peer reviewe