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An aerobic, mildly acidophilic actinobacterium was isolated from the Ochre Beds bog in Kootenay National Park, Canada. Cells of isolate OB1T were Gram-stain-positive, non-motile, pink- to purple-pigmented filaments. The pH range for growth was pH 3.5–6.5 (optimum pH 5.5), and the temperature range was 13–30°C. The major cellular fatty acids were i-C16 : 0 (28.5 %), i-C15 : 0 (14.6 %) and ai-C15 : 0 (14.3 %), and the major polar lipid was phosphohexose. The major quinone was menaquinone-11 (MK-11), and the peptidoglycan type was A1γ. The DNA G+C content was 70.2 %. Along with growth on complex media including yeast extract, proteose peptone, casamino acids and tryptic soy broth, growth occured on mono- and disaccharides (glucose, sucrose, galactose and xylose) and polysaccharides (starch, gellan, pectin, xylan and alginate). Anaerobic growth was not observed. The cells did not fix atmospheric nitrogen. On the basis of comparative 16S rRNA gene sequence analysis, this isolate belonged to the family Actinospicaceae , in the suborder Catenulisporineae of the order Actinomycetales . The most closely related species was Actinospica robiniae . However, the 16S rRNA gene sequence identity to this bacterium was low (92.8 %) and there were several chemotaxonomic differences from this species. We therefore propose a novel genus and species, Actinocrinis puniceicyclus gen. nov., sp. nov., with strain OB1T (=DSM 45618T=ATCC BAA-2771T) as the type strain.

Vacuum freeze-drying is an important food-processing technology for valid retention of nutrients and bioactive compounds. Cronobacter sakazakii has been reported to be associated with severe infections in neonates through consumption of contaminated powdered infant formula. In this study, effects of vacuum freeze-drying treatment for 12, 24, and 36 h on inactivation of C. sakazakii with different initial inoculum levels in sterile water, tryptic soy broth (TSB), skim milk, and whole milk were determined. Results indicated that the lethality rate of C. sakazakii in each sample increased with the extension of vacuum freeze-drying time. With initial inoculum levels of 102 and 103 cfu/mL, the survival of C. sakazakii in different liquid media was significantly affected by vacuum freeze-drying for 12, 24, and 36 h. In addition, the lethality rates of C. sakazakii in whole milk, skim milk, and TSB was significantly reduced compared with those in sterile water. Furthermore, whole milk showed the strongest protective role for C. sakazakii cells, followed by skim milk and TSB medium. Using the scanning electron microscope, the intracellular damage and obvious distortion of C. sakazakii cells were observed after vacuum freeze-drying for 24 and 36 h compared with the untreated sample, and the injured cells increased with the extension of vacuum-drying time. We concluded that inactivation of vacuum freeze-drying on C. sakazakii cells is related to the food matrix, and a combination with other methods for inactivating C. sakazakii is required for ensuring microbial safety of powdered infant formula.

The effects of a commercial lactate salt formulation—containing potassium lactate (KL) and potassium acetate (KA)—and liquid smoke (LS) on the growth of selected lactic acid bacteria (LAB; Carnobacterium maltaromaticum, Carnobacterium inhibens, Lactococcus lactis, Lactobacillus curvatus, and Enterococcus faecalis), fish spoilage bacteria (Photobacterium phosphoreum, Pseudomonas putida, and Vibrio vulnificus), and Listeria innocua, were examined in a tryptic soy broth model system at 20oC based on BioScreen data. The most pronounced inhibition effect on growth of bacteria was seen in the presence of 6% KL + 0.4% KA, used either combined with LS or alone. Only a minor inhibition effect on growth was found in the presence of LS alone. The only exception was Lactobacillus curvatus, which grew quite well in the presence of LS compared to control medium. The growth of Vibrio vulnificus was prevented in 6% KL + 0.4% KA, and significantly inhibited in the presence of 3% KL + 0.2% KA. When V. vulnificus wa...

The objective of this study was to determine the possible source of predominant Bacillus licheniformis contamination in a whey protein concentrate (WPC) 80 manufacturing plant. Traditionally, microbial contaminants of WPC were believed to grow on the membrane surfaces of the ultrafiltration plant as this represents the largest surface area in the plant. Changes from hot to cold ultrafiltration have reduced the growth potential for bacteria on the membrane surfaces. Our recent studies of WPCs have shown the predominant microflora B. licheniformis would not grow in the membrane plant because of the low temperature (10 °C) and must be growing elsewhere. Contamination of dairy products is mostly due to bacteria being released from biofilm in the processing plant rather from the farm itself. Three different reconstituted WPC media at 1%, 5%, and 20% were used for biofilm growth and our results showed that B. licheniformis formed the best biofilm at 1% (low solids). Further investigations were done using 3 different media; tryptic soy broth, 1% reconstituted WPC80, and 1% reconstituted WPC80 enriched with lactose and minerals to examine biofilm growth of B. licheniformis on stainless steel. Thirty-three B. licheniformis isolates varied in their ability to form biofilm on stainless steel with stronger biofilm in the presence of minerals. The source of biofilms of thermo-resistant bacteria such as B. licheniformis is believed to be before the ultrafiltration zone represented by the 1% WPC with lactose and minerals where the whey protein concentration is about 0.6%.

Aim This study was conducted to evaluate the presence of Aeromonas spp. in raw and ready-to-eat (RTE) fish commonly consumed in Assiut city, Egypt, and to determine virulence factors due to they play a key role in their pathogenicity. Materials and methods A total of 125 samples of raw and RTE fish samples were taken from different fish markets and fish restaurants in Assiut Governorate and screened for the presence of Aeromonas spp. by enrichment on tryptic soy broth then incubated at 30°C for 24 h. Plating unto the sterile Petri dishes containing Aeromonas agar base to which Aeromonas selective supplement was added. The plates were incubated at 37°C for 24 h. Presumptive Aeromonas colonies were biochemically confirmed and analyzed for pathogenicity by hemolysin production, protease, and lipase detection. Results The results indicated that raw fish were contaminated with Aeromonas spp. (40% in wild and 36% in cultured Nile tilapia). Regarding RTE, Aeromonas spp. could be isolated with the percentage of 16%, 28% and 20% in fried Bolti, grilled Bolti and fried Bayad, respectively. Out of 35 isolates obtained, 22 were categorized as Aeromonas hydrophila, 12 were classified as Aeromonas sobria and Aeromonas caviae were found in only one isolate. The virulence factors of Aeromonas spp. were detected and the results showed that all isolates produced of hemolysin (91.4%), protease (77.1%), and lipase enzyme (17.1%). Conclusion This study indicates that the presence of A. hydrophila with virulence potential in fresh and RTE fish may be a major threat to public health.

Minimum Inhibitory Concentration (MIC) of allicin were determined for 11 bacterial strains using 6 culture media including Todd Hewitt Broth (THB), Brain Heart Infusion (BHI) Broth, Tryptic Soy Broth (TSB), Mueller Hinton Broth (MHB), Luria Bertani (LB) Broth, and Brucella Broth (BB) at 37 °C. The chemical stability of allicin as a function of media was measured by HPLC. The MICs for most individual isolates grown at the same temperature varied with the media used. Higher MICs were obtained in BB, THB, BHI, TSB/MHB and LB with geometric mean values of 272.7, 145.2, 90.5, 77.3/72.6 and 46.7 μg ml−1 respectively. In media with higher MICs a big percent of allicin disappeared within the first 4 h of incubation and apparently allicin had shorter half-life compared to those with lower MIC values. The results showed that temperature directly influences the stability of allicin and shorter half-life was found at 37 °C. We conclude that the type of culture media, markedly influences the chemical stability of allicin and accordingly its MIC in a given bacterial species.

Bacteria growing as biofilms are distinct from the same bacteria growing as planktonic cells. Biofilms cells show increased resistance to antimicrobial, immunological, predatory, and chemical attack than planktonic cells. Most studies on bacterial diseases use planktonic bacteria. The objective of this study was to identify expressed proteins that are unique to Staphylococcus aureus biofilm mode of growth. S. aureus was grown in tryptic soy broth and Dulbecco's modified eagle medium at biofilm and planktonic growth conditions. Protein samples were cleaned up and separated according to their electrophoretic mobility using 7 cm IPG strips (pH 3–10 and pH 4–7) on 2D gel electrophoresis. Expressed proteins of both growth conditions were compared. Data analysis revealed that the expression of S. aureus proteins from planktonic and biofilm was higher in TSB media than DMEM media. Biofilm growth condition showed higher intensity of expressed proteins and new expressed proteins were observed. One protein was found to be upregulated in planktonic growth condition. Additionally, the majority of the proteins were clustered in the area of acidic region (pH 4–7). 2D-gel electrophoresis is a powerful and widely used method for the proteomic analysis. Biofilms represent a realistic representation of bacterial behavior and organisms are capable of altering their physiology in the surrounded environments. The results could help to illustrate the differences in pathogenesis between biofilm and planktonic cells in any model of disease. This will identify biological markers to improve the diagnostics, treatment, and prevention of S. aureus biofilms.

Various Listeria monocytogenes strains may contaminate a single food product, potentially resulting in simultaneous exposure of consumers to multiple strains. However, due to bias in strain recovery, L. monocytogenes strains isolated from foods by selective enrichment (SE) might not always represent those that can better survive the immune system of a patient. We investigated the effect of cocultivation in tryptic soy broth with 0.6% yeast extract (TSB-Y) at 10°C for 8 days on (i) the detection of L. monocytogenes strains during SE with the ISO 11290-1:1996/Amd 1:2004 protocol and (ii) the in vitro virulence of strains toward the Caco-2 human colon epithelial cancer cell line following exposure to simulated gastric fluid (SGF; pH 2.0)-HCl (37°C). We determined whether the strains which were favored by SE would be effective competitors under the conditions of challenges related to gastrointestinal passage of the pathogen. Interstrain competition of L. monocytogenes in TSB-Y determined the relative population of each strain at the beginning of SE. This in turn impacted the outcome of SE (i.e., favoring survival of competitors with better fitness) and the levels exposed subsequently to SGF. However, strong growth competitors could be outcompeted after SGF exposure and infection of Caco-2 cells by strains outgrown in TSB-Y and underdetected (or even missed) during enrichment. Our data demonstrate a preferential selection of certain L. monocytogenes strains during enrichments, often not reflecting a selective advantage of strains during infection. These findings highlight a noteworthy scenario associated with the difficulty of matching the source of infection (food) with the L. monocytogenes isolate appearing to be the causative agent during listeriosis outbreak investigations. IMPORTANCE This report is relevant to understanding the processes involved in selection and prevalence of certain L. monocytogenes strains in different environments (i.e., foods or sites of humans exposed to the pathogen). It highlights the occurrence of multiple strains in the same food as an important aspect contributing to mismatches between clinical isolates and infection sources during listeriosis outbreak investigations.

This study investigated the transfer frequency of the extended-spectrum β-lactamase-encoding gene ( bla SHV18 ) among Klebsiella pneumoniae in tryptic soy broth (TSB), pasteurized milk, unpasteurized milk, alfalfa sprouts and chopped lettuce at defined temperatures. All transconjugants were characterized phenotypically and genotypically. KP04 ΔKM and KP08 ΔKM isolated from seed sprouts and KP342 were used as recipients in mating experiments with K. pneumoniae ATCC 700603 serving as the donor. In mating experiments, no transconjugants were detected at 4 °C in liquid media or chopped lettuce, but detected in all media tested at 15 °C, 24 °C, and 37 °C. At 24 °C, the transfer of bla SHV18 gene occurred more frequently in alfalfa sprouts (5.15E-04 transconjugants per recipient) and chopped lettuce (3.85E-05) than liquid media (1.08E-05). On chopped lettuce, transconjugants were not detected at day 1 post-mating at 15 °C, but observed on day 2 (1.43E-05). Transconjugants carried the bla SHV18 gene transferred from the donor and the virulence gene harbored by recipient. More importantly, a class 1 integrase gene and resistance to tetracycline, trimethoprim/sulfamethoxazole were co-transferred during mating. These quantitative results suggest that fresh produce exposed to temperature abuse may serve as a competent vehicle for the spread of gene encoding for antibiotic resistance, having a potential negative impact on human health.

This purpose of this study was to determine the best enrichment medium for rejuvenating and recovering Salmonella placed in cold temperature prior to the employment of the gold biosensor combined with a light microscopic imaging system. A mixture of nalidixic-resistant Salmonella Typhimurium and Enteritidis were inoculated onto chicken (1,000 CFU/chicken). After cold injury at 4℃ for 24 hr, Salmonella on chicken was enriched for 6 hr with six non-selective media including buffered peptone water broth, lactose broth, brain heart infusion broth (BHI), universal pre-enrichment broth, nutrient broth, and tryptic soy broth, and five selective media including brilliant green broth (BG), rappaport-vassiliadis R10 broth, selenite cystine broth, selenite broth, and tetrathionate brilliant green broth (TBG) for the comparison of Salmonella growth. Various concentrations of Salmonella (10, 50, 100, 500, and 1,000 CFU/chicken) were then enriched for 6 hr in both BHI and BG media to select the best media. BHI was selected as the most effective non-selective enrichment medium, while BG was selected as the most effective selective enrichment medium. Finally, BHI medium was selected as the most efficient enrichment medium for Salmonella growth injured from cold temperature during processing or storage.

Genomic antimicrobial resistance (AMR) prediction tools have the potential to support foodborne illness outbreak investigations through their application in the analysis of bacterial genomes from causative strains. The AMR marker profile of a strain of interest, initially identified in outbreak-associated clinical samples, may serve as the basis for customization of selective enrichment media, facilitating its recovery from samples in a food safety investigation. Different possibilities for AMR analyses include the use of comprehensive AMR gene databases such as the Comprehensive Antibiotic Resistance Database, which can be mined with in-house bioinformatics alignment tools (e.g., Antimicrobial Resistance Marker Identifier), or publicly available tools based on clinically relevant acquired AMR gene databases (e.g., ResFinder). In combination with a previously reported pipeline (SigSeekr) designed to identify specific DNA sequences associated with a particular strain for its rapid identification by PCR, it should be possible to deploy custom recovery and identification tools for the efficient detection of priority pathogens such as Shiga toxigenic Escherichia coli (STEC) outbreak strains within the time frame of an active investigation. Using a laboratory STEC strain as a model, trimethoprim resistance identified by both Antimicrobial Resistance Marker Identifier and ResFinder was used as the basis for its selective recovery against a background of commensal E. coli bacteria in ground beef samples. Enrichment in modified tryptic soy broth containing trimethoprim greatly enhanced the recovery of low numbers of model strain cells inoculated in ground beef samples, as verified by the enumeration of colonies on plating media using a strain-specific PCR method to determine the recovery efficiency for the target strain. We discuss the relative merits of different AMR marker prediction tools for this purpose and describe how such tools can be utilized to good effect in a typical outbreak investigation scenario.

A new method to deliver fluoride using biodegradable poly(lactic-co-glycolic acid) microcapsules to suppress cariogenic bacterial growth during orthodontic treatment was investigated. A coaxial ultrasonic atomizer was used to encapsulate KF solution into microcapsules. The orthodontic adhesive resin disk containing fluoride loaded microcapsules (DFLM) was prepared by LED light curing. The microstructure of microcapsules, successful loading of KF, fracture strength, and shear bonding strength were assessed by FE-SEM, confocal laser scanning microscope, and general purpose testing machine, respectively. Fluoride release from the DFLM in phosphate buffered saline and pH changes were measured after different periods of soaking time. Antibacterial activity of the DFLM was assessed in tryptic soy broth containing mutant streptococci. The starting inoculum and the orthodontic resin disk containing microcapsules not loaded with KF were used as negative and positive controls, respectively. As results, the cumulative amount of KF after 49 days was about 85% of the initial amount of fluoride contained in the microcapsules. The fracture and shear bonding strengths of the orthodontic resin disks with and without the microcapsules were similar to each other. The DFLM showed lower bacterial growth than the control groups, whereas no statistically significant differences were found between the negative and positive controls. It can be concluded that the microcapsules loaded with fluoride prepared by a coaxial ultrasonic atomizer have good potential for application as an antibacterial agent due to their excellent cariogenic antibacterial activity when incorporated into orthodontic adhesive resin. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 31-39, 2018.

Biofertilizers are increasingly available in the market as one of the alternatives to chemical fertilizers. The supply of a high number of viable microorganisms is important for farmers. Lysobacter capsici YS1215 producing chitinases and gelatinases, isolated from soil in Korea, was evaluated for the establishment of an optimal medium condition of its shelf life under an in vitro condition. In this study, the population density of a biofertilizer (L. capsici YS1215) in media containing crab shell and gelatin powder (M1, M2, M3 and M4) was observed to be higher than that of populations grown in TSB (Tryptic soy broth) media (M5, M6 and M7) during experimental period. In addition, the population density at 11 months was over 10 6 CFU mL -1 in M1, M3 and M4, but under 10 6 CFU mL -1 in M2, M5, M6 and M7. The best optimal medium for the shelf life was M1 (2.6 × 10 6 CFU mL -1 ) containing both chitinous and gelatinous materials at 11 months. Therefore, this study provided results of the appropriate medium composition for the enhancement of the shelf life of L. capsici YS1215.

Pedobacter cryoconitis BG5 are psychrophiles isolated from the cold environment and capable of proliferating and growing well at low temperature regime. Their cellular products have found a broad spectrum of applications, including in food, medicine, and bioremediation. Therefore, it is imperative to develop a high-cell density cultivation strategy coupled with optimized growth medium for P. cryoconitis BG5. To date, there has been no published report on the design and optimization of growth medium for P. cryoconitis, hence the objective of this research project. A preliminary screening of four commercially available media, namely tryptic soy broth, R2A, Luria Bertani broth, and nutrient broth, was conducted to formulate the basal medium. Based on the preliminary screening, tryptone, glucose, NaCl, and K2HPO4 along with three additional nutrients (yeast extract, MgSO4, and NH4Cl) were identified to form the basal medium which was further analyzed by Plackett-Burman experimental design. Central composite experimental design using response surface methodology was adopted to optimize tryptone, yeast extract, and NH4Cl concentrations in the formulated growth medium. Statistical data analysis showed a high regression factor of 0.84 with a predicted optimum optical (600 nm) cell density of 7.5 using 23.7 g/L of tryptone, 8.8 g/L of yeast extract, and 0.7 g/L of NH4Cl. The optimized medium for P. cryoconitis BG5 was tested, and the observed optical density was 7.8. The cost-effectiveness of the optimized medium was determined as 6.25 unit prices per gram of cell produced in a 250-ml Erlenmeyer flask.

In this study, we examined the antimicrobial effects of citrus extract (Citrox(®)) and chitosan on lactic acid bacteria (LAB) and the pathogens Escherichia coli O157:H7 and Salmonella enterica on turkey meat during storage under vacuum packaging (VP) at 4 and 10 °C. We also examined the effects of Citrox and chitosan on pathogen contamination in tryptic soy broth (TSB). Chitosan alone or in combination with Citrox inhibited the growth of endogenous LAB in turkey meat, whereas citrus extract did not cause a major reduction in bacterial density. Citrus extract combined with chitosan yielded the lowest mesophilic total viable counts (TVCs), irrespective of temperature, showing major declines in all treated turkey samples from days 0-21 of storage. The shelf-lives of untreated, Citrox-treated, and chitosan and Citrox/chitosan-treated samples (as determined by TVC and sensory data) were 13, 17, and >21 days, respectively, at 4 °C for VP turkey. The addition of Citrox was more effective against S. enterica than E. coli in turkey, causing reductions of >0.5 and 2 log cfu/g at 4 and 10 °C, respectively, after 21 days of storage. Interestingly, the addition of chitosan had a significant inhibitory effect on E. coli at 4 °C and S. enterica at 10 °C as compared with the control (inoculated samples) resulting in dramatic reductions in E. coli (2 log) and S. enterica (5 log) cell counts on day 21. Of all the treatments examined, citrus extract in combination with chitosan showed an additive inhibitory effect against both pathogens, reducing E. coli and S. enterica populations, by approximately 2.7 or 4.5 and 2.2 or 5.6 log cfu/g, respectively, at 4 and 10 °C on day 21 of storage.

Lauric arginate (LAE) is a water-soluble cationic surfactant which has antimicrobial activity against a broad spectrum of foodborne pathogens. Some spice essential oils are effective lipophilic antimicrobials. Combining both antimicrobials may reduce their usage levels and possible negative sensory impacts when applied in complex food matrices. The objective of this study was to combine a nonionic surfactant (Tween 80) with LAE to form stable nanoemulsions with cinnamon bark essential oil (CBO) and to characterize the antimicrobial activity of these nanoemulsions. CBO was homogenized at 1% w/w in the aqueous phase with 3% w/w Tween 80 and 0.05-0.375% w/w LAE, followed by heating at 90°C for 30min to obtain final emulsions. With 0.125% and higher LAE, transparent emulsions with ~100nm in hydrodynamic diameter were observed to be stable during 30-day storage at 21°C. Antimicrobial activities of the nanoemulsion prepared with Tween 80 and 0.375% w/w LAE were studied. The respective minimum inhibitory concentrations (MICs) of the nanoemulsion in tryptic soy broth (TSB) were 12, 7, and 8ppm LAE for Salmonella enteritidis, Escherichia coli O157:H7, and Listeria monocytogenes, while those of free LAE were 11, 6, and 6ppm, respectively. MICs of CBO were 400ppm for the tested bacteria and Tween 80 at 6% w/w did not show inhibitory effect. Growth kinetics of the bacteria in TSB treated with the nanoemulsion or individual components at concentrations corresponding to the MICs of free LAE showed that binding among the LAE and Tween 80 and CBO components resulted in the antibacterial activity of nanoemulsion being lower than same concentrations of free LAE and CBO. Conversely, little difference was observed for the individual antimicrobials and the nanoemulsion in 2% reduced fat milk, and 750ppm LAE and 2000ppm CBO were observed to be the dominant antimicrobial against Gram-positive and Gram-negative bacteria, respectively. The growth of L. monocytogenes in 2% reduced fat milk at 4°C was not observed when treated by the nanoemulsion corresponding to 187.5ppm LAE and 500ppm CBO. Therefore, stable and transparent nanoemulsions of EOs can be prepared with the combination of LAE and Tween 80 without compromising antimicrobial activities.

Meat products are clearly associated with foodborne pathogens including, antibiotic-resistant strains. Population growth and growing consumer demand facilitate the transmission of foodborne pathogens, particularly in developing countries. To determine the prevalence of antibiotic-resistant Salmonella in goat meat, a study was done in Tanzania (June to July, 2015). Overall 120 goat meat samples were collected from five large and five small slaughter facilities (n = 60, respectively). Pre-enrichment for Salmonella isolation was done in Tryptic Soy Broth followed by selective enrichment in Modified Semisolid Rappaport-Vassiliadis agar. Isolation of Salmonella was done in xylose-Lysine-Deoxycholate agar followed by biochemical confirmation in triple sugar iron agar. The average prevalence of Salmonella was 60 and 63% in large and small facilities, respectively. Breakpoint assays indicated an overall low prevalence of resistance (2 to 4%; n = 219 isolates) to ampicillin, amoxicillin, streptomycin, sulphamethoxazole and trimethoprim with complete susceptibility to ciprofloxacin, ceftazidime and cefotaxime. No significant difference (p > 0.05) in the prevalence of resistance between large and small facilities was observed. High probability of Salmonella contamination of goat meat from Arusha area of Tanzania can pose risks to consumers. Antibiotic resistance appears minimal in this population. Improved hygienic slaughter and meat-handling practices are encouraged to reduce the burden of Salmonella-positive meat products. Key words: Antibiotic resistance, goat meat, Salmonella, Arusha, Tanzania.

Several enzymes are involved in the process of converting milk to lactic acid and coagulated milk to curd and, therefore, are important in dairy fermented products. Amylase, proteinase, and lipase are enzymes that play an important role in degrading milk into monomeric molecules such as oligosaccharides, amino acids, and fatty acids, which are the main molecules responsible for flavors in cheese. In the current study, we determined the amylase, proteinase, and lipase activities of Lactococcus chungangensis CAU 28(T), a bacterial strain of nondairy origin, and compared them with those of the reference strain, Lactococcus lactis ssp. lactis KCTC 3769(T), which is commonly used in the dairy industry. Lactococcus chungangensis CAU 28(T) and L. lactis ssp. lactis KCTC 3769(T) were both found to have amylase, proteinase, and lipase activities in broth culture, cream cheese, and yogurt. Notably, the proteinase and lipase activities of L. chungangensis CAU 28(T) were higher than those of L. lactis ssp. lactis KCTC 3769(T), with proteinase activity of 10.50 U/mL in tryptic soy broth and 8.64 U/mL in cream cheese, and lipase activity of 100 U/mL of tryptic soy broth, and 100 U/mL of cream cheese. In contrast, the amylase activity was low, with 5.28 U/mL in tryptic soy broth and 8.86 U/mL in cream cheese. These enzyme activities in L. chungangensis CAU 28(T) suggest that this strain has potential to be used for manufacturing dairy fermented products, even though the strain is of nondairy origin.

Among various preventive approaches, non-invasive phototherapy/photodynamic therapy is one of the methods used to control oral biofilm. Studies indicate that light at specific wavelengths has a potent antibacterial effect. The objective of this study was to determine the effectiveness of violet-blue light at 380-440 nm to inhibit biofilm formation of Streptococcus mutans or kill S. mutans. S. mutans UA159 biofilm cells were grown for 12-16 h in 96-well flat-bottom microtiter plates using tryptic soy broth (TSB) or TSB with 1 % sucrose (TSBS). Biofilm was irradiated with violet-blue light for 5 min. After exposure, plates were re-incubated at 37 °C for either 2 or 6 h to allow the bacteria to recover. A crystal violet biofilm assay was used to determine relative densities of the biofilm cells grown in TSB, but not in TSBS, exposed to violet-blue light. The results indicated a statistically significant (P 0.05) decrease compared to the non-treated groups after the 2 or 6 h recovery period. Growth rates of planktonic and biofilm cells indicated a significant reduction in the growth rate of the violet-blue light-treated groups grown in TSB and TSBS. Biofilm viability assays confirmed a statistically significant difference between violet-blue light-treated and non-treated groups in TSB and TSBS. Visible violet-blue light of the electromagnetic spectrum has the ability to inhibit S. mutans growth and reduce the formation of S. mutans biofilm. This in vitro study demonstrated that violet-blue light has the capacity to inhibit S. mutans biofilm formation. Potential clinical applications of light therapy in the future remain bright in preventing the development and progression of dental caries.

Optical density (OD) measurement is applied universally to estimate cell numbers of microorganisms growing in liquid cultures. It is a fast and reliable method but is based on the assumption that the bacteria grow as single cells of equal size and that the cells are dispersed evenly in the liquid culture. When grown in such liquid cultures, the human pathogen Staphylococcus aureus is characterized by its aggregation of single cells into clusters of variable size. Here, we show that aggregation during growth in the laboratory standard medium tryptic soy broth (TSB) is common among clinical and laboratory S. aureus isolates and that aggregation may introduce significant bias when applying standard enumeration methods on S. aureus growing in laboratory batch cultures. We provide a simple and efficient sonication procedure, which can be applied prior to optical density measurements to give an accurate estimate of cellular numbers in liquid cultures of S. aureus regardless of the aggregation level of the given strain. We further show that the sonication procedure is applicable for accurate determination of cell numbers using agar plate counting of aggregating strains.

Salmonella serovars, one of the leading contributors to foodborne illness and are especially problematic for foods that are not cooked before consumption, such as fresh produce. The shipping containers that are used to transport and store fresh produce may play a role in cross contamination and subsequent illnesses. However, methods for quantitatively attached cells are somewhat variable. The overall goal of this study was to compare conventional plating with molecular methods for quantitating attached representative strains for Salmonella Typhimurium and Heidelberg on reusable plastic containers (RPC) coupons, respectively. We attached Salmonella enterica serovar Typhimurium ATCC 14028 and serovar Heidelberg SL486 (parent and an antibiotic resistant marker strain) to plastic coupons (2.54 cm(2)) derived from previously used shipping containers by growing for 72 h in tryptic soy broth. The impact of the concentration of sanitizer on log reductions between unsanitized and sanitized coupons was evaluated by exposing attached S. Typhimurium cells to 200 ppm and 200,000 ppm sodium hypochlorite (NaClO). Differences in sanitizer effectiveness between serovars were also evaluated with attached S. Typhimurium compared to attached S. Heidelberg populations after being exposed to 200 ppm peracetic acid (PAA). Treatment with NaClO caused an average of 2.73 ± 0.23 log CFU of S. Typhimurium per coupon removed with treatment at 200 ppm while 3.36 ± 0.54 log CFU were removed at 200,000 ppm. Treatment with PAA caused an average of 2.62 ± 0.15 log CFU removed for S. Typhimurium and 1.41 ± 0.17 log CFU for S. Heidelberg (parent) and 1.61 ± 0.08 log CFU (marker). Lastly, scanning electron microscopy (SEM) was used to visualize cell attachment and coupon surface topography. SEM images showed that remaining attached cell populations were visible even after sanitizer application. Conventional plating and qPCR yielded similar levels of enumerated bacterial populations indicating a high concordance between the two methods. Therefore, qPCR could be used for the rapid quantification of Salmonella attached on RPC.

A new approach to microbiological analysis of several pharmaceuticals, in particular, those that can be used in manufacturing sterile medical products, is presented. The proposed approach was verified using a series of 23 samples of various products, three of which were obtained by biotechnological methods. The scheme includes inoculation of samples with test strains followed by isolation and analysis of the contaminants. Validation results showed that the amount of products for microbiological analysis could be reduced to 3 g, which was sufficient for obtaining reliable results. Tryptic soy broth was used as the solvent (diluent) and enrichment broth for microbe growth.

Objective Streptococcus mutans is an etiological agent of human dental caries and produces glucosyltransferase (GTF), encoded by genes gtfB and gtfC, an enzyme that synthesizes insoluble glucan from sucrose to form sticky biofilms. However, GTF functions are limited by pH. To observe the effects of low pH in primary culture conditions, we performed biofilm formation assays with S. mutans and compared the results obtained at pH 6.0 and pH 7.0. Methods Biofilm formation of S. mutans MT8148 was observed after 16h in 96-well microtiter plates containing Tryptic soy broth (TSB) without dextrose and with 0.25% sucrose. Roles of extracellular DNA (eDNA) in biofilm formation, with and without insoluble glucans, were also examined by addition of DNase and colony stimulating peptide (CSP) to the biofilm and gtfB mutant. Results S. mutans formed significant biofilms in the presence of eDNA after 16h culture at pH 6.0, which showed lower production of insoluble glucans than culture at pH 7.0. Moreover, the S. mutans MT8148 gtfB mutant produced significant biofilms in pH 6.0 condition, and biofilm formation was inhibited by DNase and stimulated by addition of CSP. Insoluble glucan-independent and eDNA-dependent biofilm formation was observed in the stress condition of low pH. Conclusions Insoluble glucan-independent biofilm formation by S. mutans was significantly influenced by increasing eDNA in primary low pH culture conditions.

Listeria monocytogenes cells may induce alkali stress adaptation when exposed to sublethal concentrations of alkaline cleaners and sanitizers that may be frequently used in the food processing environment. In the present study, the effect of temperature on the induction and the stability of such alkali stress adaptation in L. monocytogenes were investigated. Our results demonstrate that temperature plays a critical role in the induction of alkali stress adaptation in L. monocytogenes under sublethal alkaline conditions. Pre-exposure to pH 9.0 tryptic soy broth supplemented with 0.6% yeast extract (TSB-YE) at 37 °C induced pronounced alkali stress adaptation whereas sublethal alkaline pre-exposure at 4 °C failed to induce any alkali stress adaptation. Furthermore, this pattern of alkali stress adaptation in L. monocytogenes was not dependent on the length of pre-exposure time, the concentration of sublethal alkali, the types of alkaline agents and the growth phases of cells. In addition, alkali stress adaptation induced at 37 °C was completely reversed in pH 7.2 TSB-YE within 2 h at 37 °C or within 4 h at 22 °C. However, once it was induced at a higher temperature, alkali stress adaptation in L. monocytogenes remained stable at 4 °C for at least 4 h. Our findings suggest that even though cold temperatures do not induce alkali stress adapted phenotypes, but it can maintain the previously acquired alkali stress adaptation much longer in L. monocytogenes.

This study investigated biofilm formation, cell surface hydrophobicity, colony spreading, and slime production for 112 Staphylococcus aureus strains isolated from various sources (leaf vegetables, pea leaf, perilla leaf, Kim-bab, person, and animal). When biofilm formation was classified by origin, S. aureus isolated from animal origin showed a significantly higher level of biofilm formation than others (p≤0.05). When S. aureus groups with different levels of biofilm formation (very strong, strong, moderate, and weak) were evaluated for the correlation with cell surface properties, there was a positive correlation between biofilm formation and hydrophobicity (r=0.926). Biofilm formation and colony spreading on tryptic soy broth (without dextrose) also showed positive correlation (r=0.863). In contrast, biofilm formation and slime production were negatively correlated (r=−0.973). Based on these results, the biofilm forming ability of S. aureus differs depending on their origin and might be affected by cell surface properties such as cell surface hydrophobicity.

Tofu liquid waste contains high levels of organic matter, especially protein and amino acids. These organic compounds can be proteins, carbohydrates and fats. Most of the bacteria are able to use oil or fat as a source of carbon and energy, bacteria that have this ability are often known as lipolytic bacteria. This study aims to obtain biosurfactant producing bacteria from tofu wastewater. This research was conducted from July - September 2020. The method used in this research is survey method and emulsification method. The media used for bacterial isolation were Tryptic Soy Broth (TSB) and Tryptic Soy Agar (TSA). 6 of the isolates are able to produce biosurfactants. Morphological and biochemical characteristics indicate six bacterial genera, namely Genus Agrobacterium, Proteus, Proteus, Citrobacter, Enterobacter, and Serratia. The bacteria obtained are bacteria that have the potential to degrade oil in a polluted environment.

The slaughterhouse wastewater is rich of oil and fat and it may inhabit by biosurfactant producing bacteria. Isolation of the bacteria is needed in order to find out a biosurfactant producing bacteria that can be used to maintain the oil and fat polluted area through bioremediation process. To isolate and identify the biosurfactant-producing bacteria, a study has been conducted on July to September 2020. The samples were obtained from slaughterhouse wastewater at Cipta Karya street Pekanbaru, Riau Province and samplings were conducted three times. The isolation was conducted by TSB (Tryptic Soy Broth) and incubated for 24 hours. Then the bacterial samples were planted by TSA (Tryptic Soy Agar) and isolated for 24 hours. Then, it was identified using biochemical and morphological tests were carried out to find out the types of bacteria. The Emulsification Index was then calculated by using TSB (Tryptic Soy Broth) and kerosene. Results shown that the emulsification index were Flavobacterium 53%, Agrobacterium 53%, Serratia 60%, Salmonella 52. 5%, Salmonella 52.4%, Proteus 65.7%, Clostridium 62% and Aeromonas 59.5%. As Proteus is shown the highest Emulsification Index, it was check using a PCR ( Polymerase Chain Reaction) and it is proved that the species is Proteus vulgaris

Bio-indicators are nowadays being increasingly used by the food and drug industry to detect the changes in environmental conditions. In this study, a time-temperature indicator (TTI) is fabricated by using an office thermal inkjet printer to print a new custom-made bio-ink for detecting changes in color that occur with time and temperature variation. In lieu of the regular inks, the TTI uses solutions containing Geobacillus stearothermophilus (G. stearothermophilus) and Bacillus atrophaeus (B. atrophaeus) spores, tryptic soy broth, pH indicators, and a combination of polyvinyl alcohol and borax as the gelling agent, to be dispensed from the printer cartridge. Following the printing process, the samples thus obtained are coated with a protective film to avoid moisture loss. The effects of bio-ink gelatinization, pH, color intensity, and print resolution on bacterial printing are determined by varying the level of each parameter. The presence of the protective film on printed bacteria is found not only to lead to the growth and proliferation of bacteria but also to reveal color with sharper contrast. The indicator’s initial and final colors are modified by varying the pH levels of the solutions. Increasing color intensity (20%, 40%, 60%, 80%, and 100%) in the software settings is used to induce significant enhancements in the printed bacterial populations, thereby controlling the rate of color change. It is, moreover, observed that changes in print resolution do not have any significant effects on the populations of printed bacterial spores. In general, the TTI’s endpoint may be modified so as to suit various pharmaceutical and food applications.

The global emergence of carbapenem-resistant Enterobacteriaceae (CRE) is a major concern for public health. The Centers for Disease Control and Prevention (CDC)-recommended procedure for surveillance is overnight incubation of a rectal/perianal swab in 5 ml of tryptic soy broth (TSB) supplemented

Background: Dental caries is one of the most common infectious diseases and often occurs in the community caused by bacteria. Attached bacteria in the tooth surface for a long time will form a biofilm and will lead to demineralization characterized by damage in the structure of the tooth enamel. The bacteria that cause dental caries and can form biofilms is Streptococcus mutans. The bacteria inside biofilms are more resistant to antibacterial agents. Flavonoids in mangosteen pericarp extract can be a cleaner alternative for the anti-biofilm cavity that has properties against Streptococcus mutans. Purpose: To determine the activity of flavonoids in mangosteen pericarp extract at a certain concentration against Streptococcus mutans bacteria. Methods: This study was a laboratory experimental study with a post-test only control group design. Streptococcus mutans were diluted according to the Mc Farland dilution standard 106 in Tryptic Soy Broth (TSB) medium and put in a flexible U-bottom microtiter plate. Then it was incubated for 5x24 hours and checked using crystal violet simple staining to see the formation of biofilms. Flavonoid extract of mangosteen pericarp performed serial dilution in a concentration of 100%, 50%, 25%, 12.5%, 6.25%, 3.125%, 1.56%, and 0.78% was added, and the incubation process were conducted for 1x24 hours. OD (Optical Density) readings were done with a wavelength of 595 nm. Results: There was a significant difference between the test groups and the positive control group. The concentration of 100% had the anti-biofilm activity and showed the value of the highest percentage of inhibition, whilst the concentration of 0.78% showed a minimum biofilm inhibition concentration. The results were demonstrated by a statistical analysis test. Conclusion: Flavonoid extract of mangosteen pericarp at a certain concentration has anti-biofilm activity against Streptococcus mutans biofilm.

The resistance of lactobacilli to oxidative stress is of great importance for their applicability as probiotics. This study aimed to evaluate the response of Lactobacillus plantarum strain ATCC® 14917™, grown in either de Man, Rogosa, and Sharpe agar (MRS medium) or tryptic soy broth (TSB medium), to 1–2 mM H2O2 after the exposure to different doses of ionising radiation. Two bacterial extracellular enzyme groups, dehydrogenases (DHAs) and quinone reductases (QRs), served as the criteria of viability and antioxidant activity, respectively. The irradiated L. plantarum culture grown in TSB showed increased QR activity at irradiation doses of 2–50 Gy, with the maximum activity at 10 Gy, compared to non-irradiated cells. The QR activity of the MRS-grown culture was increased significantly (p

We delineate the excellent bactericidal efficacy of stable heterojunction nanocomposites composed of single-walled carbon nanotubes (SWCNTs) and copper(II) oxide (CuO) synthesized via facile recrystallization and calcination. The bactericidal effectiveness of the fabricated nanocomposites was examined using the standard broth-dilution method and the growth-inhibition-zone analysis method, in which bacteria cultured in an incubator in tryptic soy broth medium were subjected to the prepared samples. The bactericidal activity of all of the as-synthesized samples is evident in both methods, displaying a substantial decrease in bacterial colonies and resulting in clear inhibition zones, respectively. Among the CuO–SWCNT nanocomposites, the sample subjected to calcination at 500 °C for 5 h was found to exhibit the best performance against Staphylococcus aureus and Escherichia coli, forming inhibition zones 182% and 162% larger than those formed by pure CuO, respectively.

The development of cost-effective bioreactors and scalable methods for the high quantities of bacteriophage production meets the global demand for phage therapy. This study demonstrates the potential application of a novel PES-STiO2 nanocomposite membrane (NCM) to maintain the bacteriophages, increase the adsorption rate, and productivity in the phage enrichment reactor (PER). A constant volume of freshly grown Escherichia coli cells in tryptic soy broth (TSB) medium (first phase) was tainted with Escherichia phage in PER (second phase) in a cyclic process. Subsequently, the bacteria-phage filtrate and phage lysate samples were removed from the PER through the external cross-flow membrane reactor and the outlet of the PER at each cycle, respectively. The PER performed with the membrane had shown higher phage titers and productivity in the phage lysate at each cycle than those achieved from the PER w/o NCM (control). The phage productivity was increased (∼107 to 1013 PFU mL−1 cycle−1) with the increase in cycles and was higher than those from the control, and other reported reactors. As detected with the electron scanning microscope (SEM) and flux test, the NCM operated with bacteria-phage could able to prevent biofouling without their loss. Thus, the PES-STiO2 nanocomposite membrane integrated PER is simple and can be used at an industrial scale to maintain the volumetric phage production and higher phage productivity.

Pseudomonas fluorescens group bacterial strains with high biofilm-forming ability were isolated from cold raw milk, identified based on molecular data, and their motility, biofilm-forming ability and proteolytic activity were characterised. After 48 h incubation at 7 °C, 57 out of 67 P. fluorescens group strains produced biofilms in vitro on polystyrene microplates. Six isolates with high-ability to produce biofilms were identified as Pseudomonas simiae, Pseudomonas veronii, Pseudomonas cedrina and three strains of P. fluorescens. They were able to produce high amounts of biofilm on stainless steel surface in tryptic soy broth and UHT milk. AdnA gene was detected in the six tested strains, but in two strains that did not show any modes of motility until 96 h of incubation at 7 °C, fliC gene was not detected. AprX gene and different levels of total and heat-resistant proteolytic activity were detected in the six tested strains.

Biocide susceptibility testing (BST) of bacteria lacks standardised methods. Based on a recently established broth macrodilution BST method, a broth microdilution method for BST was developed. To establish the respective protocol, four reference strains Staphylococcus aureus ATCC® 6538, Enterococcus hirae ATCC® 10541, Escherichia coli ATCC® 10536 and Pseudomonas aeruginosa ATCC® 15442 were investigated for their minimal inhibitory concentrations (MICs) towards quaternary ammonium compounds (benzalkonium chloride), cationic compounds (chlorhexidine), aldehydes (glutardialdehyde) and alcohols (isopropanol) using tryptic soy broth. All combinations of (i) inoculum preparation according to the German Veterinary Medical Society (DVG) or the Clinical and Laboratory Standards Institute (CLSI) with some modifications, (ii) use of 1st subculture (SC) and 2nd SC, (iii) direct colony suspension (DCS) with/without glass beads, and (iv) incubation at 37 °C for 24 h, 48 h, and 72 h were compared using seven independent replications. Overall, the reproducibility was high for all abovementioned strain/biocide/parameter combinations. In total, 86.9 % - 100 % of the results were within ± one dilution step of the mode value. The proposed method for a standardised BST protocol comprises (i) two different inoculum densities, (ii) the use of a fresh overnight culture (1st SC or 2nd SC), (iii) the preparation of the inoculum suspension by either of the two methods using DCS with or without glass beads, and (iv) the incubation at 37 °C for 24 h. This broth microdilution method will help to harmonize BST of bacterial pathogens in routine diagnostics.

In vitro cultivation of staphylococci is fundamental to both clinical and research microbiology, but few studies, to-date, have investigated how the differences in rich media can influence the volatilome of cultivated bacteria. The objective of this study was to determine the influence of rich media composition on the chemical characteristics of the volatilomes of Staphylococcus aureus and Staphylococcus epidermidis. S. aureus (ATCC 12600) and S. epidermidis (ATCC 12228) were cultured in triplicate in four rich complex media (brain heart infusion (BHI), lysogeny broth (LB), Mueller Hinton broth (MHB), and tryptic soy broth (TSB)), and the volatile metabolites produced by each culture were analyzed using headspace solid-phase microextraction combined with comprehensive two-dimensional gas chromatography&mdash, time-of-flight mass spectrometry (HS-SPME-GC&times, GC-TOFMS). When comparing the chemical compositions of the staph volatilomes by the presence versus absence of volatiles produced in each medium, we observed few differences. However, when the relative abundances of volatiles were included in the analyses, we observed that culturing staph in media containing free glucose (BHI and TSB) resulted in volatilomes dominated by acids and esters (67%). The low-glucose media (LB and MHB) produced ketones in greatest relative abundances, but the volatilome compositions in these two media were highly dissimilar. We conclude that the staphylococcal volatilome is strongly influenced by the nutritional composition of the growth medium, especially the availability of free glucose, which is much more evident when the relative abundances of the volatiles are analyzed, compared to the presence versus absence.

Polymicrobial biofilms are major complications of various chronic infections. Therefore, in vitro biorelevant polymicrobial biofilm models are essential tools for medical studies. This study presents an in vitro model for dual species biofilm of Pseudomonas aeruginosa and Staphylococcus aureus developed on cell-derived matrices (CDMs), in order to simulate the microenvironment of in vivo biofilms. P. aeruginosa and S. aureus are two of the most frequent pathogens in polymicrobial biofilms of wound infections. Although they are commonly isolated from polymicrobial biofilms, their interaction is antagonistic; and there is severe battle between them for nutrients and space. We introduced a nutritious formulation supporting co-cultures of P. aeruginosa and S. aureus in order to study the interaction of these gram-positive and gram-negative bacterial species. Quantitative analyses demonstrated that the enrichment of tryptic soy broth (TSB) with NaCl and glucose facilitate dual-species biofilm formation of P. aeruginosa and S. aureus when it is mixed with fetal bovine serum (FBS). Furthermore, the dual species biofilm was incubated on CDMs. Characterization of the model by fluorescent and electron microscopy techniques revealed realistic features of chronic multi-species biofilms, including competitive distribution pattern of two bacterial species and small-colony variants (SCVs) morphology of S. aureus.

Bacillus subtilis AB2.0 showed to produce a surface-active compound that was identified by mass spectrometry analysis as a surfactin. Four different isoforms were identified in the protonated form [M+H]+ with m/z of 1030.6350, 1044.6511, 1058.6661 and 1072.6848. They were constituted of fatty acids varying from C13 to C16; however, about 70% of the isoforms showed m/z of 1058.6661 and the fatty acid C15. Based on the AB2.0 genome sequence, only one gene group related to surfactin biosynthesis was found. Strain AB2.0 produced 99.6 mg/l of surfactin in Tryptic Soy Broth after 48 h. The biosurfactant presented an emulsification activity of 45.7% in this growth condition. It considerably decreased the water surface tension from 72.0 mN/m to 24.7 mN/m and the interfacial tension of n-hexadecane from 30.1 mN/m to 7.5 mN/m, as well as of a medium crude oil from 42.3 mN/m to 6.9 mN/m and of a light crude oil from 36.4 to 3.8 mN/m. Calcite flotation test was conducted to access the wettability reversal of the biosurfactant in high salinity condition similar to that observed in an oil production reservoir, and AB2.0 reversed 100% using concentrations as low as 25 ppm of biosurfactant. Oil contaminated sand columns were used to simulate microbial enhanced oil recovery at different temperatures and high salinity. When surfactin from AB2.0 was added to the sand columns, the TPH concentration in liquid phases recovered was enhanced up to 24× indicating that AB2.0 is an excellent candidate for its application in MEOR in subsurface environments.

In this study, we examined the change in resistance of Salmonella enterica serovar Typhimurium and Escherichia coli O157:H7 to 222-nm krypton-chlorine (KrCl) excilamp treatment as influenced by acid adaptation and identified a mechanism of resistance change. In addition, we measured changes in apple juice quality indicators, such as color, total phenols, and 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging activity, during treatment. Non-acid-adapted and acid-adapted pathogens were induced by growing the cells in tryptic soy broth without dextrose (TSB w/o D) at pH 7.3 and in TSB w/o D at pH 5.0 (adjusted with HCl), respectively. For the KrCl excilamp treatment, acid-adapted pathogens exhibited significantly (P 0.05) difference in quality indicators compared to those of untreated controls during treatment at 1,773 mJ/cm(2). IMPORTANCE There is a need for novel, mercury-free UV lamp technology to replace germicidal lamps containing harmful mercury, which are routinely utilized for UV pasteurization of apple juice. In addition, consideration of the changes in response to antimicrobial treatments that may occur when pathogens are adapted to the acid in an apple juice matrix is critical to the practical application of this technology. Based on this, an investigation using 222-nm KrCl excilamp technology, an attractive alternative to mercury lamps, was conducted. Our study demonstrated increased resistance to 222-nm KrCl excilamp treatment as pathogens adapted to acids, and this was due to changes in reactivity to ROS with changes in the fatty acid composition of the cell membrane. Despite increased resistance, the 222-nm KrCl excilamp achieved pathogen reductions of 5 log or more at laboratory scale without affecting apple juice quality. These results provide valuable baseline data for application of 222-nm KrCl excilamps in the apple juice industry.

To evaluate total absorbance, planktonic growth, biofilm formation, viability, metabolic activity, and pH of Streptococcus mutans UA159 cultures when different dilutions of Stevia rebaudiana Bertoni were applied and to determine the minimum inhibitory concentration (MIC) and the minimum biofilm inhibitory concentration (MBIC) of Stevia on S. mutans. The effects of different dilutions of Stevia (0–400 mg/ml) on S. mutans total growth, planktonic growth, biofilm formation, viability, metabolic activity, and pH during a 72-h growth period were evaluated in this in vitro study. A stock solution was prepared by mixing 10 ml of tryptic soy broth (TSB) supplemented with 1% sucrose (TSBS) and 4 g of Stevia. S. mutans total growth and biofilm formation decreased with reduced concentrations of Stevia. Furthermore, the MIC was 25 mg/ml and the MBIC was 6.25 mg/ml. Complete eradication of S. mutans was not observed with any of the Stevia concentrations. Planktonic growth of S. mutans was not repressed by high concentrations of Stevia and most of the Stevia concentrations generated an increased pH. Because Stevia reduces biofilm and acid production, Stevia can be considered a non-cariogenic sweetener. This study confirms the anticariogenic effect of Stevia, like it has been previously reported, but more studies on the most effective concentration are needed, and in the present study, the minimum inhibitory concentration (MIC) and the minimum biofilm inhibitory concentration (MBIC) was determined in the presence of sucrose. Additionally, this is the first study to evaluate the effect of Stevia on S. mutans metabolic activity.

Evidence is presented suggesting that the decreased enumeration of heat-stressed Staphylococcus aureus cells on selective media is the result of accumulation of metabolic H2O2. It accumulates due to the decreased activity of catalase caused by the synergistic effects of heat and NaCl. Heated cells enumerated anaerobically on tryptic soy agar (TSA) containing 6.5% NaCl (TSAS 6.5) exhibited a 200-fold increase compared to cells enumerated aerobically on the same medium. The anaerobic counts on TSAS 6.5 were similar to the aerobic counts on TSA. Increases in both death and injury occurred when S. aureus was propagated in tryptic soy broth (TSB) plus 10% NaCl (TSBS) instead of TSB before thermal injury. Addition of catalase to TSA and TSA containing 7.5% NaCl (TSAS) increased the count to approximately the same levels on TSA and TSAS as that found following thermal injury after propagation in TSB. Catalase activity was 12-fold higher in stationary phase cells propagated in TSB than in TSBS. Indirect evidence indicates that toxic levels of H2O2 accumulated rapidly, causing one to two log decreases in enumeration after 30 to 60 min incubation on TSAS.

Background. N ecrosis teeth need a procedure called the root canal treatment to eliminate microorganisms in root canal , s o the inflammation process did not continue to the periapical.One of the microbes that exist i n thenecrosis teethis P orphyromonas gingivalis , that capable of forming biofilm because it has the ability to communicate with bacteria signaling ( quorum sensing ) . It is necessary to discover another antibiofilm agent as an irrigation material in root canal preparation deriving from nature. Flavonoidmangosteen pericarp extract can disrupt acyl-homoserine lactones ( AHLs ) thus quorum sensing bacteria will be disrupted and causing permanent attachment and the establishment of colonies on the surface of the teeth any barrier . Purpose. The aim of this study found out the antibiofilmactivity of flavonoid mangosteen pericarp extract against Porphyromonas gingivalis bacteria. Method. This study has a laboratory experimental with post-test only group design. Porphyromonas gingivalis ATCC 33277 was diluted according to Mc. Farland standard 10 6 CFU/ml in Tryptic Soy Broth (TSB) medium and inserted into microtitterplate flexible U bottom. Porphyromonas gingivalis was incubated for 6x24 hour and checked with a simple staining to see the formation of biofilm. Flavonoid mangosteen pericarp extract was added in various concentrations and OD (optical density) readings done with a wavelength of 595 nm. Result. Flavonoid mangosteen pericarp extractshowed a decrease of OD value from control in all concentrations. Conclusion. Flavonoid mangosteen pericarp extract has antibiofilm activity against Porphyromonas gingivalis bacteria.

Rapid detection of bacterial pathogens is a critical unmet need for both food and environmental samples such as irrigation water. As a part of the Food safety Modernization Act (FSMA), The Produce Safety rule has established several requirements for testing for the presence of generic Escherichia coli in water, but the current method available for testing (EPA M1603) demands specified multiple colony verification and highly trained personnel to perform these tests. The purpose of the study was to assess a phage induced bacterial lysis using quantitative image analysis to achieve rapid detection of E. coli at low concentrations within 8 hours. This study aimed to develop a simple yet highly sensitive and specific approach to detect target bacteria in complex matrices. In the study, E. coli cells were first enriched in tryptic soy broth (TSB), followed by T7 phage induced lysis, concentration, staining and fluorescent imaging. Image analysis was conducted including image pre-processing, image segmentation and quantitatively analysis of cellular morphological features (area, eccentricity and full width at half maximum). Challenge experiments using realistic matrices, including simulated fresh produce wash water, coconut water and spinach wash water, demonstrated the method can be applied for use in situations that occur in food processing facilities. The results indicated E. coli cells that are lysed by T7 phages demonstrated significantly (P < 0.05) higher extracellular DNA release, altered cellular shape (from rod to circular) and diffused fluorescent signal intensity. Using this biosensing strategy, a sensitivity to detect Escherichia coli at 10 CFU/ml within 8 hours was achieved, both in laboratory medium and in complex matrices. The proposed phage based biosensing strategy enables rapid detection of bacteria and is applicable to analysis of food systems. Furthermore, the steps involved in this assay can be automated to enable detection of target bacteria in food facilities without extensive resources.

An estimated 179 million cases of acute gastroenteritis (AGE) occur each year in the United States. Escherichia coli O157:H7 and Listeria monocytogenes, common etiologic agents of AGE, both have low-infectious doses and high environmental resistance. Surfaces contaminated with either pathogen, such as the exterior surface of a food package, could serve as a vehicle for AGE transmission. The aim of this study was to determine the persistence of E. coli O157:H7 and L. monocytogenes on three common food packaging materials–oriented polyethylene terephthalate (OPET), oriented polypropylene (OPP), and nylon-6. Coupons (5 × 5 cm2) from each material were sterilized under ultraviolet light for 5 min, inoculated with ca. 7 log CFU of a three-strain-mixture of green fluorescent protein (GFP)-labeled E. coli O157:H7, then incubated according to the Technical Association of the Pulp and Paper Industry standards. Surviving E. coli O157:H7 cells were recovered in saline at 0, 0.25, 0.5, 1, 2, 3, 5, 7, 14, 15 days then enumerated on tryptic soy broth (TSB) supplemented with ampicillin using the three-tube most probable number (MPN) method. The same procedure using a three-strain mixture of rifampin-resistant (Rif)-L. monocytogenes was followed except TSB was supplemented with rifampin. E. coli O157:H7 and L. monocytogenes both survived on OPET, OPP, and nylon-6 for 15 days. The log reductions for E. coli O157:H7 were 5.7 log MPN, 5.3 log MPN, and 5.1 log MPN across the three materials respectively. The log reduction for L. monocytogenes was 6.5 log MPN, 6.3 log MPN, and 4.8 log across the same three materials. The number of surviving E. coli cells was significantly higher (P > 0.05) than L. monocytogenes at sampling times 1, 2, 3, 5, 7, 14, and 15 days. No significant difference was shown between the two bacteria between 0 and 0.5 days. Survival of both bacteria across all three materials was not significantly different (P > 0.05). These results suggest that a highly contaminated (107 CFU/coupon) exterior surface of a food package could be a potential vehicle for the transmission of bacteria associated with AGE.

Pseudomonas spp. are commonly isolated from dogs with clinical otitis and have been shown to produce biofilm. There is a paucity of studies demonstrating biofilm growth in veterinary medicine.To compare biofilm production of Pseudomonas spp. isolated from dogs with otitis using three different enrichment broths at two different time points. Speciation was performed.One hundred isolates from 98 dogs with clinical otitis were assessed for biofilm production.One hundred isolates were assessed for biofilm production using a microtitre plate assay. Biofilm production in Luria-Bertani Broth (LBB), Mueller-Hinton Broth (MHB) and Tryptic Soy Broth (TSB) were assessed after 18 and 24 h of incubation.At 18 h, biofilm production was demonstrated in 87% of LBB, 91% of TSB and 93% of MHB grown isolates. By 24 h, this was 92% of LBB, 96% of TSB and 99% of MHB isolates. Biofilm production was significantly increased after 24 h incubation compared to 18 h. A significant difference was noted in biofilm production between LBB and MHB (P = 0.0349), but not between LBB and TSB (P = 0.3727) or MHB and TSB (P = 0.3687) at 24 h incubation. Two isolates were speciated as P. fluorescens and 98 as P. aeruginosa.Not all enrichment broths were equivalent to one another and 24 h incubation was superior to 18 h. Biofilm production was high in this population of Pseudomonas spp. isolates.Les Pseudomonas spp. sont fréquemment isolés de chiens atteints d'otite clinique et sont capables de produire du biofilm. Il y a peut d’étude montrant la croissance de biofilm en médecine vétérinaire. HYPOTHÈSES/OBJECTIFS: Comparer la production de biofilm par des Pseudomonas spp. isolés de chiens avec otite à l'aide de trois milieux différemment enrichis à deux moments distincts. Une spéciation a été réalisée.Cent souches de 98 animaux avec otite clinique ont été évaluées pour la production de biofilm. MATÉRIEL ET MÉTHODE: Cent souches ont été testées pour la production de biofilm à l'aide de plaques de microtitration. La production de biofilm a été évaluée après 18 et 24h d'incubation dans LBB (Luria-Bertani Broth), MHB (Mueller-Hinton Broth) et TSB (Tryptic Soy Broth). RÉSULTATS: A 18h, une production de biofilm a été montrée à 87% sur LBB, 91% sur TSB et 93% sur MHB. A 24h, il y avait 92% sur LBB, 96% sur TSB et 99% sur MHB des souches. La production de biofilm a augmenté significativement après une incubation de 24h par rapport à 18h. Une différence significative a été notée dans la production de biofilm entre LBB et MHB (P = 0.0349), mais pas entre LBB et TSB (P = 0.3727) ou MHB et TSB (P = 0.3687) à 24 h d'incubation. Des souches ont été spéciées comme P. fluorescens et 98 comme P. aeruginosa.Tous les milieux n’étaient pas équivalents et une incubation de 24h était supérieure à 18h. La présence d'un biofilm est fréquente dans les otites à Pseudomonas spp.INTRODUCCIÓN: Pseudomonas spp. comúnmente se aíslan de perros con otitis clínica y se ha demostrado que producen biomembranas. Hay una escasez de estudios que demuestren el crecimiento de biomembranas en medicina veterinaria. HIPÓTESIS/OBJETIVOS: comparar la producción de biomembranas de Pseudomonas spp. aisladas de perros con otitis utilizando tres caldos de enriquecimiento diferentes en dos puntos temporales diferentes. Se realizó especiación. ANIMALES: se evaluó la producción de biomembranas de cien aislados obtenidos de 98 animales con otitis clínica. MÉTODOS Y MATERIALES: se evaluaron cien aislamientos para la producción de biomembranas utilizando un ensayo de placa de microtitulación. La producción de biomembranas en caldo Luria-Bertani (LBB), caldo Mueller-Hinton (MHB) y caldo tríptico de soja (TSB) se evaluó después de 18 y 24 h de incubación. RESULTADOS: a las 18 h, se demostró la producción de biomembranas en el 87% de cultivos en LBB, el 91% en TSB y el 93% de los aislados en MHB. A las 24 h, estos resultados fueron 92% en LBB, 96% en TSB y 99% en aislamientos en MHB. La producción de biomembranas aumentó significativamente después de 24 h de incubación en comparación con 18 h. Se observó una diferencia significativa en la producción de biomembranas entre LBB y MHB (P = 0,0349), pero no entre LBB y TSB (P = 0,3727) o MHB y TSB (P = 0,3687) a las 24 h de incubación. Se identificaron dos aislados como P. fluorescens y 98 como P. aeruginosa. CONCLUSIÓN E IMPORTANCIA CLÍNICA: no todos los caldos de enriquecimiento fueron equivalentes entre sí y las 24 h de incubación fueron superiores a las 18 h. La presencia de un biomembranas es probable para Pseudomonas spp. aisladas de casos de otitis.Pseudomonas spp. werden häufig bei Hunden mit einer klinischen Otitis isoliert. Es konnte gezeigt werden, dass sie einen Biofilm produzieren. Es gibt nur wenige Studien, die das Wachstum des Biofilms in der Veterinärmedizin dokumentieren.Ein Vergleich der Biofilmproduktion von Pseudomonas spp., welcher bei Hunden mit einer Otitis isoliert wurde mittels dreier verschiedener Anreicherungsbouillons zu zwei verschiedenen Zeitpunkten. Es wurde eine Speziesbestimmung durchgeführt.Es wurden einhundert Isolate von 98 Tieren mit einer klinischen Otitis auf Biofilmproduktion untersucht.Einhundert Isolate wurde auf Biofilmproduktion mittels Mikrotiterplattenassay untersucht. Es wurde die Biofilmproduktion in Luria-Bertani Bouillon (LBB), in Mueller-Hinton Bouillon (MHB) und Tryptischer Sojabouillon (TSB) nach einer 18- und 24-stündigen Inkubation beurteilt.Nach 18h konnte eine Biofilmproduktion in 87% der in LBB gewachsenen Isolate, in 91% der in TSB gewachsenen Isolate und in 93% der in MHB gewachsenen Isolate gezeigt werden. Nach 24h war dies bei 92% der Isolate in LBB, 96% der Isolate in TSB und 99% der Isolate in MHB der Fall. Die Biofilmproduktion war nach 24h Inkubation im Vergleich zu einer 18-stündigen Inkubation signifikant erhöht. Nach 24-stündiger Inkubation wurde ein signifikanter Unterschied bei der Biofilmproduktion zwischen LBB und MHB (P = 0,0349), aber nicht zwischen LBB und TSB (P = 0,3727) oder MHB und TSB (P = 0,3687) festgestellt. Zwei Isolate wurden als P. fluorescens und 98 als P. aeruginosa identifiziert.Nicht alle Anreicherungsbouillons waren gleichwertig und eine 24-stündige war besser als eine 18-stündige Inkubationsdauer. Das Auftreten von Biofilm ist bei Pseudomonas spp. Isolaten aus dem Ohr sehr wahrscheinlich.Pseudomonas spp são comumente isoladas em cães com otite e têm demonstrado ser produtoras de biofilme. Há uma escassez de estudos demonstrando o crescimento de biofilme em medicina veterinária. HIPÓTESE/OBJETIVOS: Comparar a produção de biofilme de Pseudomonas spp isoladas de cães com otite utilizando três meios de cultura em caldo diferentes em tempos distintos. Realizou-se especiação.Cem isolados de 98 animais com otite foram avaliados para a produção de biofilme. MÉTODOS E MATERIAIS: Os cem isolados foram avaliados para a produção de biofilme utilizando um ensaio em placa de microtitulação. Avaliou-se a produção de biofilme em caldo de Luria-Bertani (LBB), caldo Mueller-Hinton (MHB) e caldo de soja tríptica (TSB) após 18 e 24 horas de incubação.Após 18 horas, a produção de biofilme foi demonstrada em 87% dos isolados em LBB, 91% em TSB e 93% em MHB. A produção de biofilme foi significativamente maior após 24 horas de incubação, comparado a 18 horas. Observou-se uma diferença significativa na produção de biofilme entre LBB e MHB (P = 0.0349), mas não entre LBB e TSB (P = 0.3727) ou MHB e TSB (P = 0.3687) após 24 horas de incubação. Dois isolados foram especiados como P. fluorescens e 98 as P. aeruginosa. CONCLUSÕES E IMPORTÂNCIA CLÍNICA: Nem todos os meios de cultura em caldo foram equivalentes entre si e o tempo de incubação de 24 horas foi superior a 18 horas. É provável a presença de biofilme em isolados auriculares de Pseudomonas spp.背景: シュードモナス属菌は臨床的に中耳炎を有する犬から一般的に分離され、バイオフィルムを形成することが明らかにされている。獣医学においてバイオフィルムの発達を実証する研究は不足している。 仮説/目的: 本研究の目的は、異なる2時点において3つの異なる濃縮ブロスを用いて中耳炎を有する犬から分離したシュードモナス属菌のバイオフィルム生成能を比較することである。さらにスペシエーションを実施した。 被験動物: 臨床的に中耳炎を有する98頭の動物から採取された100分離株に対し、バイオフィルム産生能について評価した。 方法および材料: マイクロタイタープレートアッセイを用いて100株をバイオフィルム産生能について評価した。 Luria-Bertani Broth(LBB)、Mueller-Hinton Broth(MHB)およびTryptic Soy Broth(TSB)においてバイオフィルム産生能を、培養18および24時間後に評価した。 結果: 培養後18時間の時点で、87%のLBB、91%のTSB、および93%のMHB増殖分離株がバイオフィルム産生を示した。 24時間までに、92%のLBB、96%のTSBおよび99%のMHB分離株がバイオフィルムを産生した。バイオフィルム産生は、18時間と比較して24時間培養後に有意に増加した。 24時間培養で、LBBとMHB間にバイオフィルム産生に有意差が見られた(P = 0.0349)が、LBBとTSB間(P = 0.3727)またはMHBとTSB間(P = 0.3687)では見られなかった。分離株のうち2株をP. fluorescens、98株をP. aeruginosaとして分類した。 結論と臨床的重要性: すべての濃縮ブロスが互いに同等ではなく、24時間培養が18時間より優れていた。バイオフィルムは、耳におけるシュードモナス属株に存在しそうである。.背景: 临床中,经常能从耳炎患犬的耳道中分离出有生物膜的假单胞菌。但很少有研究阐明兽医学中生物膜的生长。 假设/目标: 从耳炎患犬分离出来假单胞菌,分别使用三种不同的富集液培养,比较两个不同时间点的生物膜形成,并进行形态分析。 动物: 评估采自98只耳炎动物的一百个菌株形成的生物膜。 方法和材料: 使用微量滴定板测定法评估一百个菌株形成的生物膜。分别在Luria-Bertani肉汤(LBB)、Mueller-Hinton肉汤(MHB)和Tryptic Soy肉汤(TSB)中培养18和24小时,随后对生物膜的形成进行评估。 结果: 培养18小时后,87%的LBB、91%的TSB和93%的MHB培养的菌株形成了生物膜;培养24小时后,92%的LBB、96%的TSB和99%的MHB分离株有生物膜的形成。与18小时相比,培养24小时后的生物膜形成显著增加。LBB与MHB的生物膜形成有显著差异(P = 0.0349),但在培养24小时后,LBB和TSB(P = 0.3727)或MHB和TSB(P = 0.3687)之间无显著差异。两种菌株被鉴定为荧光假单胞菌,而98株为铜绿假单胞菌。 结论和临床价值: 并非所有富集液彼此一致,但培养24小时优于18小时。耳道假单胞菌菌株很可能形成生物膜。.

Heat-stable antifungal factor (HSAF), which was first isolated from Lysobacter enzymogenes, exhibits inhibitory activities against a wide range of pathogens; however, a low level of HSAF was obtained from L. enzymogenes cultured in 0.1 × tryptic soy broth (TSB), an amount that does not satisfy HSAF application in disease control. In this study, the optimization of media components and environmental conditions were examined for improving the production of HSAF from L. enzymogenes OH11. The one factor at a time method was used to screen optimal nitrogen and carbon sources and inorganic salt. Then the orthogonal matrix method was used to determine the optimal concentration of the media components and environmental factors. The results showed that the maximum level of HSAF (23361 mAU·s) was achieved when OH11 cultured in the media of 0.7% (w/v) soybean powder, 0.5% (w/v) glucose and 0.08% CaCl2 at 200 rpm at 30°C for 60 h, which is much higher than that cultured in 0.1 × TSB. This opens up the possibility of HSAF or L. enzymogenes utilization for biological control of plant disease.

Under stressful conditions, Salmonella enterica forms multinucleated elongated filaments. The triggers and outcomes of filamentation are not well characterized. S. enterica serotypes Newport, Javiana, and Typhimurium were evaluated for their ability to form filaments upon exposure to 20 mM pelargonic acid. S. Newport was used as a model to investigate the progression and fate of filamentation via culturable population size, cell length, and viability assays. All serotypes displayed filament formation after 16 h of incubation. Pelargonic acid amendment of tryptic soy broth (TSBpel) produced a 5-log CFU reduction compared to TSB after 24 h (P

This paper presents the effects of the composition of different media (i.e., Tryptic soy broth (TSB), Brain heart infusion (BHI), Listeria enrichment broth (LEB), Fraser broth (FB) and University of Vermont medium (UVM)) on the detection of a short peptide fragment PepD specific to the p60 protein (p60) of L. monocytogenes by a monoclonal antibody (anti-PepD mAb). Expression of the p60 obtained was demonstrated to be proportional to the cellular growth of Listeria monocytogenes regardless of the tested growth medium. However, the early growth of L monocytogenes and the expression of the p60 were negatively affected by the presence of selective agents present in LEB, FB and UVM. Among those three selective enrichment media commonly used for L. monocytogenes, LEB allowed a better expression of L. monocytogenes p60 after an incubation period of 18 h. Optimization of the LEB revealed that the dextrose concentration was the critical factor for improving the expression of p60 and promotes the early expression of p60. Moreover, an optimal dextrose concentration of 0.5% (w/v) in LEB, coupled with anti-PepD mAb immobilized to solid support, reduced the detection of p60 from 18 h to 9 h for an initial concentration of L. monocytogenes of 10(8) CFU/ml. (C) 2019 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

Aims. Microbial colonization of silicone voice prostheses by bacteria andCandidaspecies limits the device lifetime of modern voice prostheses in laryngectomized patients. Thus, research focuses on biofilm inhibitive properties of novel materials, coatings, and surface enhancements. Goal of thisin vitrostudy was the evaluation of seven commonly used growth media to simulate growth of mixed oropharyngeal species as mesoscale biofilms on prosthetic silicone for future research purposes.Methods and Results. Yeast Peptone Dextrose medium (YPD), Yeast Nitrogen Base medium (YNB), M199 medium, Spider medium, RPMI 1640 medium, Tryptic Soy Broth (TSB), and Fetal Bovine Serum (FBS) were used to culture combined mixedCandidastrains and mixed bacterial-fungal compositions on silicone over the period of 22 days. The biofilm surface spread and the microscopic growth showed variations fromin vivobiofilms depending on the microbial composition and growth medium.Conclusion. YPD and FBS prove to support continuousin vitrogrowth of mixed bacterial-fungal oropharyngeal biofilms deposits over weeks as needed for longtermin vitrotesting with oropharyngeal biofilm compositions.Significance and Impact of Study. The study provides data on culture conditions for mixed multispecies biofilm compositions that can be used for future prosthesis designs.

Bacteriophages specifically targeting different strains of bacteria can be isolated from urban sewage using properly modified enrichment techniques. This chapter provides a detailed protocol for isolation of Proteus mirabilis-specific bacteriophages. Briefly, prefiltered sewage is mixed with double-concentrated tryptic soy broth containing the target strain and incubated. Subsequently, the suspension is spread on phage nutrient agar, and if needed, supplemented with swarming motility inhibitor, for the induction of bacterial growth and phage multiplication. Phages infecting bacteria are identified by plaques (patches of dead bacteria) in the confluent bacterial lawn. A pure phage preparation is obtained by cutting out a single plaque from a double-layer agar plate and subsequent virus propagation five times on a given P. mirabilis strain.