A review of methods to determine viability, vitality and metabolic rates in microbiology

Viability and metabolic assays are commonly used as proxies to assess the overall metabolism of microorganisms. The variety of these assays combined with little information provided by some assay kits or online protocols often leads to mistakes or poor interpretation of the results. In addition, the use of some of these assays is restricted to simple systems (mostly pure cultures), and care must be taken in their application to environmental samples. In this review, the necessary data are compiled to understand the reactions or measurements performed in many of the assays commonly used in various aspects of microbiology. Also, their relationships to each other, as metabolism links many of these assays, resulting in correlations between measured values and parameters, are discussed. Finally, the limitations of these assays are discussed

Determinants of stimulated salivary flow among haematopoietic stem cell transplantation recipients

The aetiology of hyposalivation in haematopoietic stem cell transplantation (HSCT) recipients is not fully understood. This study examined the effects of treatment-related aetiological factors, particularly medications, on stimulated salivary flow in HSCT recipients. Adult HSCT recipients (N = 118, 66 males, 27 autologous and 91 allogeneic transplants) were examined. Stimulated whole salivary flow rates (SWSFR) were measured before HSCT and at 6 and 12 months post-HSCT. Linear regression models were used to analyse the associations of medications and transplant-related factors with salivary flow rates, which were compared to salivary flow rates of generally healthy controls (N = 247). The SWSFR of recipients were lower pre-HSCT (mean +/- standard deviation, 0.88 +/- 0.56 ml/min; P <0.001), 6 months post-HSCT (0.84 +/- 0.61; P <0.001) and 12 months post-HSCT (1.08 +/- 0.67; P = 0.005) than the SWSFR of controls (1.31 +/- 0.65). In addition, hyposalivation (<0.7 ml/min) was more frequent among HSCT recipients pre-HSCT (P <0.001), 6 months post-HSCT (P <0.001) and 12 months post-HSCT (P = 0.01) than among controls. The SWSFR was observed to improve over time being significantly higher 12 months post-HSCT compared to pre-HSCT (P <0.001). The observed decrease of salivary flow could not be explained by the examined transplant-related factors and medications. Decreased stimulated salivary flow rates could not be explained by the examined factors alone; these findings indicate that hyposalivation in HSCT recipients exhibits a multifactorial aetiology. All HSCT recipients should be considered to be at high risk of hyposalivation and consequent oral diseases, and they should be treated accordingly.Peer reviewe

Dissociations of oral foci of infections with infectious complications and survival after haematopoietic stem cell transplantation

Introduction Haematopoietic stem cell transplantation (HSCT) recipients are at increased risk for severe infections. This study examined the associations of common oral infections with survival and infectious complications in HSCT recipients. Materials and methods All autologous and allogeneic HSCT recipients transplanted in the University Hospital of Basel, Switzerland, between 2008 and 2016 and referred to oral infection control pre-HSCT were included in this retrospective case-control study. All patients had a clinical and a panoramic radiological dental examination taken immediately prior to HSCT. Presence of acute or chronic oral foci of infections, decayed, missing or filled tooth index (DMFT) and radiological attachment loss (RAL) were examined. Survival and infections of the subjects were followed up for 6 months post-HSCT. Results Altogether 341 allogeneic and 125 autologous HSCT recipients were included in the study. Within 6 months post-HSCT, 47 (14%) of the allogeneic and 4 (3%) of the autologous recipients died. Oral foci of infections (acute or chronic), DMFT or periodontitis pre-HSCT were not associated with survival 6 months post-HSCT. Oral foci of infections were also not associated with hospital treated infectious diseases or blood culture positive bacteremia during the 6 month follow-up period. Untreated oral foci of infections were not associated with survival or severe infectious complications within 6 months post-HSCT. Conclusion The results of this study suggest that radical dental interventions to chronic oral infections could be postponed until post-HSCT.Peer reviewe

Exopolysaccharides Regulate Calcium Flow in Cariogenic Biofilms

Caries-associated biofilms induce loss of calcium from tooth surfaces in the presence of dietary carbohydrates. Exopolysaccharides (EPS) provide a matrix scaffold and an abundance of primary binding sites within biofilms. The role of EPS in binding calcium in cariogenic biofilms is only partially understood. Thus, the aim of the present study is to investigate the relationship between the calcium dissolution rates and calcium tolerance of caries-associated bacteria and yeast as well as to examine the properties of EPS to quantify its binding affinity for dissolved calcium. Calcium dissolution was measured by dissolution zones on Pikovskaya’s agar. Calcium tolerance was assessed by isothermal microcalorimetry (IMC) by adding CaCl2 to the bacterial cultures. Acid-base titration and Fourier transform infrared (FTIR) spectroscopy were used to identify possible functional groups responsible for calcium binding, which was assessed by isothermal titration calorimetry (ITC). Lactobacillus spp. and mutans streptococci demonstrated calcium dissolution in the presence of different carbohydrates. All strains that demonstrated high dissolution rates also revealed higher rates of calcium tolerance by IMC. In addition, acidic functional groups were predominantly identified as possible binding sites for calcium ions by acid-base titration and FTIR. Finally, ITC revealed EPS to have a higher binding affinity for calcium compared, for example, to lactic acid. In conclusion, this study illustrates the role of EPS in terms of the calcium tolerance of cariogenic microbiota by determining the ability of EPS to control free calcium concentrations within the biofilms as a self-regulating mode of action in the pathogenesis of dental caries

Syöpäpotilaan suun ja hampaiston hoito

Vertaisarvioitu.Syöpähoitoa edeltävän suun tarkastuksen tavoite on ehkäistä infektiokomplikaatioita sekä parantaa suun terveydentilaa hoidon aikana ja sen jälkeen. Näyttö suun kroonisten infektiopesäkkeiden vaikutuksesta syöpähoidon ennusteeseen on vähäistä ja akuutit hammasperäiset infektiot hoidon aikana harvinaisia. Silti suun tarkastuksen sisällyttäminen hoitokokonaisuuteen heti diagnoosin jälkeen on suositeltavaa sairauden tai hoitojen takia suurentuneen infektioriskin vuoksi. Syöpähoitoja edeltävän hammashoidon tulee painottua sairautta ehkäiseviin toimenpiteisiin. Kroonisten infektiopesäkkeiden radikaaliin saneeraukseen on yleensä syytä suhtautua pidättyvästi. Suuhygienian omaehtoinen ja ammattimainen ylläpito hoidon aikana ja jälkeen on tärkeää, sillä kuiva suu, limakalvovauriot, karies sekä luustolääkkeiden ja sädehoidon aiheuttama kudosten heikentynyt paranemiskyky altistavat uusille suun sairauksille. Lisäksi ne vaikuttavat hoidosta toipumiseen ja elämänlaatuun.Peer reviewe

Human leukocyte antigens are associated with salivary level of active MMP-8

Objectives: This case control study examined the associations of HLA antigens and periodontitis with the salivary level of active MMP-8 (aMMP-8). Materials and Methods: A total of 202 subjects, registered as Swiss bone marrow donors, participated in the study. HLA-A, -B, and -C types were determined by serology or PCR. Saliva samples were collected from subjects, followed by a periodontal examination. The salivary level of aMMP-8 was determined with immunofluorometric assay. Results: The mean salivary level of aMMP-8 was directly comparable to the grade of periodontitis and increased from healthy to mild/moderate to severe (125.0 +/- 132.1, 200.6 +/- 170.2, 290.1 +/- 202.3 ng/ml; p < 0.001 between each group, respectively). The only association between the HLA types and the salivary level of aMMP-8 was observed in subjects with HLA-A11. Subjects with healthy periodontium and HLA-A11 had a lower level of aMMP-8 (49.2 +/- 32.5 ng/ml) compared with subjects without HLA-A11 (123.6 +/- 119.2; p = 0.048). Among subjects with periodontitis, a higher level of aMMP-8 (394.2 +/- 255.6 ng/ml) was observed in subjects with HLA-A11 compared with subjects without HLA-A11 (201.1 +/- 146.1 ng/ml; p < 0.002). This finding was statistically significant also after adjusting for sex, age, smoking, tooth brushing and the number of medications (p < 0.05). Conclusions: HLA-A11 is associated with the salivary level of aMMP-8 which contributes to the subject's immune and inflammatory response in periodontium.Peer reviewe

Isothermal microcalorimetry provides new insights into biofilm variability and dynamics

The purpose of this study was to investigate a three-species in vitro biofilm with peri-implantitis-related bacteria for its variability and metabolic activity. Streptococcus sanguinis, Fusobacterium nucleatum, and Porphyromonas gingivalis were suspended in simulated body fluid containing 0.2% glucose to form biofilms on polished, protein-coated implant-grade titanium disks over 72 h using a flow chamber system. Thereafter, biofilm-coated disks were characterized by scanning electron microscopy and fluorescence in situ hybridization/confocal laser scanning microscopy. To assess metabolic activity within the biofilms, their heat flow was recorded for 480 h at 37 °C by IMC. The microscopic methods revealed that the total number of bacteria in the biofilms varied slightly among specimens (2.59 × 104 ± 0.67 × 104 cells mm−2), whereas all three species were found constantly with unchanged proportions (S. sanguinis 41.3 ± 4.8%, F. nucleatum 17.7 ± 2.1%, and P. gingivalis 41.0 ± 4.9%). IMC revealed minor differences in time-to-peak heat flow (20.6 ± 4.5 h), a trend consistent with the small variation in bacterial species proportions as shown by microscopy. Peak heat flow (35.8 ± 42.6 µW), mean heat flow (13.1 ± 22.0 µW), and total heat over 480 h (23.5 ± 37.2 J) showed very high variation. These IMC results may be attributed to differences in the initial cell counts and relative proportions of the three species, their distribution and embedment in exopolysaccharide matrix on the test specimens. The present results provide new insights into variability and dynamics of biofilms on titanium disks, aspects that should be explored in future studies of dental surface

Quantification of vital adherent Streptococcus sanguinis cells on protein-coated titanium after disinfectant treatment

The quantification of vital adherent bacteria is challenging, especially when efficacy of antimicrobial agents is to be evaluated. In this study three different methods were compared in order to quantify vital adherent Streptococcus sanguinis cells after exposure to disinfectants. An anaerobic flow chamber model accomplished initial adhesion of S. sanguinis on protein-coated titanium. Effects of chlorhexidine, Betadine®, Octenidol®, and ProntOral® were assessed by quantifying vital cells using Live/Dead BacLight™, conventional culturing and isothermal microcalorimetry (IMC). Results were analysed by Kruskal-Wallis one-way analysis of variance. Live/dead staining revealed highest vital cell counts (P0.05), indicating equivalent numbers of bacteria were created and disinfectants delayed growth but did not eliminate it. In conclusion, contrary to culturing, live/dead staining enables detection of cells that may be viable but non-cultivable. Microcalorimetry allows unique evaluation of relative disinfectant effects by quantifying differences in time delay of regrowth of remaining vital cell

Suun mukosiitti

English summaryPeer reviewe

Influence of time, toothpaste and saliva in the retention of Streptococcus mutans and Streptococcus sanguinis on different toothbrushes

Objectives: The intraoral transmission of cariogenic and periodontopathogenic species seems to be facilitated by contaminated toothbrushes and other oral hygiene devices. The aim of this investigation was to analyze the in vitro retention and survival rate of Streptococcus mutans and Streptococcus sanguinis on different toothbrushes. The impacts of human saliva and antimicrobial toothpaste on these parameters were further evaluated. Material and Methods: Part I: Four toothbrushes (Colgate 360°, Curaprox CS5460 ultra soft, elmex InterX, Trisa Flexible Head3) were contaminated by S. mutans DSM 20523 or S. sanguinis DSM 20068 suspensions for three minutes. Bacteria were removed from the toothbrushes after either three minutes (T0) or 24 hours (T24) of dry storage and grown on Columbia blood agar plates for the quantification of colony-forming units (CFUs). Part II: The effects of saliva from a caries-active or a caries-inactive person and of toothpaste containing 0.12% chlorhexidine digluconate were also tested. Results: Part I: After three minutes of dry storage, approximately one percent of the bacteria were still detectable on the toothbrushes. After 24 hours, S. sanguinis exhibited a more pronounced decrease in viable cell numbers compared with S. mutans but the differences were not significant (Kruskal-Wallis test, p>;0.05). Part II: The addition of human saliva from a caries-active or caries-inactive person slightly increased the retention of both streptococcal species at T0. The use of toothpaste had no influence on the amount of viable streptococci at T0, but it reduced the microbial load after 24 hours of storage. There were only slight nonsignificant differences (p>;0.05) between the four toothbrushes. Conclusions: In vitro bacterial retention and survival of S. sanguinis and S. mutans on different toothbrushes occurred. Within the limitations of this study, the use of human saliva or an antimicrobial toothpaste did not lead to significant differences in the microbial load on toothbrushes