Tryptic Soy Broth Research Articles

Beef-Based Medium Influences Biofilm Formation of Escherichia coli O157:H7 Isolated from Beef Processing Plants

Beef-based medium beef extract (BE) and standard medium tryptic soy broth (TSB) are used as minimally processed food models to study the effects on Escherichia coli O157:H7 biofilm formation. The effects of temperatures (4, 10, 25, 37, and 42°C), pH values (4.5, 5.0, 5.5, 6.0, 7.0, and 8.0), strain characteristics, and the expression of functional genes on the biofilm formation ability of the bacteria were determined. The three tested E. coli O157:H7 strains produced biofilm in both media. Biofilm formation was greater in BE than in TSB (P < 0.05). The strongest biofilm formation capacity of E. coli O157:H7 was achieved at 37°C and pH 7.0. Biofilm formation was significantly inhibited for three tested strains incubated at 4°C. Biofilm formation ability was correlated with swarming in TSB. Biofilm formation was significantly and positively correlated with autoaggregation or hydrophobicity in BE (P < 0.05). At the initial stage of biofilm formation, the expressions of luxS, sdiA, csgD, csgA, flhC, adrA, and rpoS were significantly higher in BE than in TSB (P < 0.05). At the maturity stage, the expressions of luxS, sdiA, csgD, csgA, flhC, csrA, adrB, adrA, iraM, and rpoS were significantly higher in TSB than in BE (P < 0.05). Such information could help in the development of effective biofilm removal technologies to deal with risks of E. coli O157:H7 biofilms in the beef industry.

Study of the impact of cultivation conditions and peg surface modification on the in vitro biofilm formation of Staphylococcus aureus and Staphylococcus epidermidis in a system analogous to the Calgary biofilm device.

Introduction. Staphylococcus aureus (SA) and Staphylococcus epidermidis (SE) are the most common pathogens from the genus Staphylococcus causing biofilm-associated infections. Generally, biofilm-associated infections represent a clinical challenge. Bacteria in biofilms are difficult to eradicate due to their resistance and serve as a reservoir for recurring persistent infections.Gap Statement. A variety of protocols for in vitro drug activity testing against staphylococcal biofilms have been introduced. However, there are often fundamental differences. All these differences in methodical approaches can then be reflected in the form of discrepancies between results.Aim. In this study, we aimed to develop optimal conditions for staphylococcal biofilm formation on pegs. The impact of peg surface modification was also studied.Methodology. The impact of tryptic soy broth alone or supplemented with foetal bovine serum (FBS) or human plasma (HP), together with the impact of the inoculum density of bacterial suspensions and the shaking versus the static mode of cultivation, on total biofilm biomass production in SA and SE reference strains was studied. The surface of pegs was modified with FBS, HP, or poly-l-lysine (PLL). The impact on total biofilm biomass was evaluated using the crystal violet staining method and statistical data analysis.Results. Tryptic soy broth supplemented with HP together with the shaking mode led to crucial potentiation of biofilm formation on pegs in SA strains. The SE strain did not produce biofilm biomass under the same conditions on pegs. Preconditioning of peg surfaces with FBS and HP led to a statistically significant increase in biofilm biomass formation in the SE strain.Conclusion. Optimal cultivation conditions for robust staphylococcal biofilm formation in vitro might differ among different bacterial strains and methodical approaches. The shaking mode and supplementation of cultivation medium with HP was beneficial for biofilm formation on pegs for SA (ATCC 29213) and methicillin-resistant SA (ATCC 43300). Peg conditioning with HP and PLL had no impact on biofilm formation in either of these strains. Peg coating with FBS showed an adverse effect on the biofilm formation of these strains. By contrast, there was a statistically significant increase in biofilm biomass production on pegs coated with FBS and HP for SE (ATCC 35983).

Protease Production from Bacillus sp. Isolated from Gastrointestinal Tract of Catfish (Clarias sp.) with Different Medium

The need for protease in the industrial field has been increasing. Candidates for producing these enzymes can be isolated from the digestive tract of catfish (Clarias sp.). The purpose of this study was to obtain bacterial isolates that produce proteolytic from the gastrointestinal tract of catfish and determine the effect of different production media on the activity of proteolytic. The first step of this study was isolation, screening, and identification of bacteria. The second step was to test the effect of the media Luria Bertani, trypticase soy broth, and skim milk broth on proteolytic activity. Nineteen isolates were obtained from the screening process of proteolytic bacteria. Isolate no 1, was known as the best isolate in producing enzymes and was known as Bacillus sp. Tests with different growth media gave results that semi-quantitative, nutrient growth media produced the highest activity with a proteolytic index value of 2.09 ± 0.41. In addition, based on quantitative tests, the media Luria Bertani Broth produced the highest specific activity with a value of 36.479 U/mg. The conclusion of this study, Bacillus sp. from the gastrointestinal tract of catfish that cultured on the Luria Bertani Broth medium produced the best activity.

Histological and microbiological evaluation of surgical wound closure in mouse skin with cyanoacrylate (Histoacryl®) in comparison to poliglecaprone (Monocryl®) traditional suture

Incisional wound closure is a key surgical step to facilitate tissue healing, reduce the risk of infection and obtain esthetic and functional recovery. Cyanoacrylates such as Histoacryl® have become a popular choice in surgical veterinary practice. However, how Histoacryl® is affecting tissue regeneration and bacterial load in the wound in comparison to poliglecaprone (Monocryl®) traditional suture methods remains to be determined. This work aimed to evaluate how wounded tissue responds to traditional suture with Monocryl® (poliglecaprone 25/4-0) and Histoacryl®, as well as provide evidence of their effects on wound healing in mice. Fortyeight hours after the incisional procedure, wound tissue biopsies were prepared for histological and microbiological analysis. Biopsies were fixed and colored with Mallory's trichrome and hematoxylin-eosin stains. For microbiological assays, biopsies were suspended in tryptic soy broth (TSB) and 1/10 diluted to evaluate the number of CFU in nutrient agar plates. Our results show no differences between Histoacryl® and Monocryl® traditional suture suggesting that both methods could be used to treat wounds in small animals such as rodents.

Zinc Oxide and Silver Nanoparticle Effects on Intestinal Bacteria.

The application of nanoparticles (NPs) for food safety is increasingly being explored. Zinc oxide (ZnO) and silver (Ag) NPs are inorganic chemicals with antimicrobial and bioactive characteristics and have been widely used in the food industry. However, not much is known about the behavior of these NPs upon ingestion and whether they inhibit natural gut microflora. The objective of this study was to investigate the effects of ZnO and Ag NPs on the intestinal bacteria, namely Escherichia coli, Lactobacillus acidophilus, and Bifidobacterium animalis. Cells were inoculated into tryptic soy broth or Lactobacilli MRS broth containing 1% of NP-free solution, 0, 12, 16, 20 mM of ZnO NPs or 0, 1.8, 2.7, 4.6 mM Ag NPs, and incubated at 37 °C for 24 h. The presence and characterization of the NPs on bacterial cells were investigated by scanning electron microscopy (SEM), transmission electron microscopy (TEM), and energy-dispersive X-ray spectroscopy (EDS). Membrane leakage and cell viability were assessed using a UV-visible spectrophotometer and confocal electron microscope, respectively. Numbers of treated cells were within 1 log CFU/mL less than those of the controls for up to 12 h of incubation. Cellular morphological changes were observed, but many cells remained in normal shapes. Only a small amount of internal cellular contents was leaked due to the NP treatments, and more live than dead cells were observed after exposure to the NPs. Based on these results, we conclude that ZnO and Ag NPs have mild inhibitory effects on intestinal bacteria.

Application of growth rate from kinetic model to calculate stochastic growth of a bacteria population at low contamination level

Traditional predictive microbiology is not suited for cell growth predictions for low-level contamination, where individual cell heterogeneity becomes apparent. Accordingly, we simulated a stochastic birth process of bacteria population using kinetic parameters. We predicted the variation in behavior of Salmonella enterica serovar Typhimurium cells at low inoculum density. The modeled cells were grown in tryptic soy broth at 25 °C. Kinetic growth parameters were first determined empirically for an initial cell number of 104 cells. Monte Carlo simulation based on the growth kinetics and Poisson distribution for different initial cell numbers predicted the results of 50 replicate growth experiments with the initial cell number of 1, 10, and 64 cells. Indeed, measured behavior of 85% cells fell within the 95% prediction area of the simulation. The calculations link the kinetic and stochastic birth process with Poisson distribution. The developed model can be used to calculate the probability distribution of population size for exposure assessment and for the evaluation of a probability that a pathogen would exceed critical contamination level during food storage.

Radiation sensitivity and inactivation of antibiotic-resistant Salmonella spp. in fresh chicken legs

In the Philippines, a total of 21,000 typhoid cases with 40 deaths caused by Salmonella typhi were reported in 2016. Previous studies have shown that multi-drug resistant Salmonella (MDRS) exists in fresh chicken meat purchased from retail stalls and supermarkets. MDRS has become a global health concern because of its resistance to treatment with the usual battery of antibiotics. Some studies show that gamma irradiation can eliminate antibiotic-resistant bacteria in food. This study was aimed at determining the radiation sensitivity and inactivation of antibiotic-resistant Salmonella spp. in fresh chicken legs. In phase one, four strains of freshly prepared Salmonella spp. were identified for resistance/susceptibility using disk diffusion method and minimum inhibitory concentration (MIC) with a different spectrum of Ampicillin (AMP), Chloramphenicol (CHL), Gentamicin (GN), Tetracycline (TET) Sulfamethoxazole/Trimethoprim (STX), based on the breakpoints established by CLSI (Clinical and Laboratory Standards Institute). Results showed that the S. enterica ser. Enteritidis 1963, S. enterica ser. Typhi 1768 and S. enterica ser. Typhimurium 1741were resistant to AMP and GN, while S. enterica ser. Typhimurium ATCC 14028 was resistant to GN. In phase two, the freshly prepared pure strains of antibiotic-resistant Salmonella spp. were grown in tryptic soy broth (TSB) for 24 h, centrifuged, and cell pellets were re-suspended in phosphate buffer saline (PBS) and adjusted at 0.5 McFarland (1.0 × 108 CFU per mL). One ml of Salmonella spp. was inoculated into fresh chicken meat then irradiated at different doses of control (0, 0.2, 0.4, 0.6, 0.8 1.0, and 2.0 kGy) at the Gamma Irradiation Facility of the Philippine Nuclear Research Institute. The D10-values, for the four strains of Salmonella, ranged from 0.14 to 0.19 kGy. This indicates that low dose radiation can induce a 1-log reduction of Salmonella spp. in fresh chicken legs and at 2 kGy doses, effectively eliminate the Salmonella spp. by 6 logs cycle. The four strains of Salmonella were mostly resistant to AMP and GN. Therefore, S. enterica serovar Enteritidis 1963, S. enterica serovar Typhi 1768, and S. enterica serovar Typhimurium 1741 exhibited multi-drug resistance. Gamma radiation can be used as a feasible technique to inactivate the presence of Salmonella spp. including the antibiotic-resistant Salmonella in fresh chicken meat.

Antimicrobial Properties of Thyme Essential Oil on Salmonella Typhimurium

Antimicrobial properties of thyme essential oil solutions (EO) oil have been identified against foodborne bacteria, but the effectiveness has not yet been fully determined. The objective of this study is to analyze the effect of thyme essential oil under standardized conditions on Salmonella Typhimurium. The Salmonella Typhimurium (ATCC 53648™) inoculum was prepared by growing the bacteria in tryptic soy broth for 24 hours at 37°C, yielding 6.9 x10 CFU/ml. One hundred microliters of the starting bacterial culture were spread onto Mueller Hinton agar plates and left to dry for 15 minutes. Treatments were formed by creating Thyme essential oil solutions (EO) of 0, 2.5, 10, 25% using 70% ethyl alcohol as the diluent. Each EO was vortexed and filtered using a 0.2 microliter sterile syringe filter. Twenty microliters of each EO were pipetted into a sterile paper disc (weight ranged of 0.0086-0.009 g). Three paper discs of each treatment containing the EO were placed on standardized locations on each inoculated plate. Each treatment consisted of 9 inoculated plates. The plates were then incubated at 37C for 24 hours. Additionally, three manufactured standard antibiotic paper discs of 5 mcg Ciprofloxacin, 10 mcg Ampicillin, and 10 mcg Streptomycin were placed on separate plates as controls. The zone of inhibition created was measured using an image processing program, Image J. All data were analyzed using the general linear model procedure for analysis of variance (ANOVA). The antimicrobial property of the EO observed as the zone of inhibition increased as the concentration of the EO increased (R= 0.91, P < 0.0001). EO at 25% showed the greatest antimicrobial effect against Salmonella Typhimurium compared to 10, 2.5, and 0% (21.5, 14.8, 10.5, and 0.0 mm, respectively; P < 0.0001). There was no significant difference (P=0.20) between the zone of inhibition of 25% E0 and ampicillin. All antibiotics had some effect against Salmonella Typhimurium where ciprofloxacin had the greatest effect; its effect was even greater than the 25% EO (29.5 vs. 21.5 mm; P< 0.0001). The 25% and 10% EO was more effective against Salmonella than Streptomycin (21.5, 14.5 vs. 12.7 mm respectively; P< 0.0001). However, the 2.5% EO was not as effective as Streptomycin (10.5 vs. 12.7 mm; P< 0.0001). In conclusion, thyme EO showed increasing antimicrobial properties against Salmonella Typhimurium as the concentration of oil increased. It proved to be more effective than streptomycin antibiotic control at concentration as low as 10% EO. Further evaluation of the effects of thyme EO is needed to determine the extent of its effectiveness against additional commensal and pathogenic organisms.

Potential of Actinomycetes Isolates as Antimicrobials for Salmonella typhi

Actinomycetes are aerobic bacteria and Gram-positive bacilli. Acinomycets have an antimicrobial, antitumor, immunosuppressive, and antiparasitic acivity. This study aims to determine the antimicrobial potency of Actinomycetes against Salmonella typhi. The research design was a true experimental design with Actinomycetes isolates obtained from the Microbiology Laboratory of the Veteran National Development University, Jakarta. Rejuvenation of Actinomycetes isolates used spread plate method on media Starch Casein Agar (SCA) with six serial dilutions of 10-1, 10-2, 10-3, 10-4, 10-5, and 10-6. The antimicrobial test method used the liquid dilution method to determine the value of the minimum inhibitory concentration (MIC) with Trypticase Soy Broth (TSB) media followed by colony calculations on Salmonella Shigella Agar (SSA) media to determine the value of the minimum bactericidal concentration (MBC). The results showed that the antimicrobial activity of Actinomycetes isolates is bacteriostatic with MIC value of 10-2. MBC value cannot be determined. It can be concluded that there is a potential of Actinomycetes isolates as an antimicrobial against Salmonella typhi (p &lt;0.05).

Detection of KPC, NDM and VIM-Producing Organisms Directly from Rectal Swabs by a Multiplex Lateral Flow Immunoassay.

We report a preliminary evaluation of the NG-Test CARBA 5 immunochromatographic assay for detecting carbapenemases directly from rectal swabs on the same day of sampling. Thirty fecal swabs were examined for carbapenemase-producing organisms (CPOs) by conventional culture, PCR, and NG-Test CARBA 5. Each sample was tested by the immunochromatographic assay five times, including direct testing and incubation in trypticase soy broth for 1, 2, 3, and 4 h. Twenty patients yielded CPOs by culture. Immunochromatographic and PCR results were concordant and detected the same 25 carbapenemases (11 KPC, 8 VIM, and 6 NDM). In five cases, we detected co-carriage of KPC and VIM. Compared with PCR, the sensitivity of NG-Test CARBA 5 for the detection of KPC, VIM, and NDM was 80% without incubation, 88% with one hour, 92% with two, and 100% with three hours incubation, while specificity was 100% for all time points. All samples containing adequate fecal content were detected by NG-Test CARBA 5 concordantly with PCR, without incubation. NG-Test CARBA 5 is a reliable test that rapidly detects the presence of carbapenemases at the same day of sampling, directly from rectal swabs. It thus provides early information to guide antimicrobial treatment and infection control interventions.

227 Effect of triamcinolone & manuka honey on Staphylococcus aureus growth & hemolytic activity

Atopic dermatitis (AD) is an inflammatory skin disease commonly treated with topical corticosteroids (TCS). Manuka honey (MH) is an alternative therapy made from nectar of the Leptospermum scoparium, native to New Zealand. The high abundance of Staphyloccocus aureus (S. aureus) on AD skin enhances disease severity, in part due to the effects of hemolysins. To address the effect TCS & MH have on S. aureus, we quantified growth & hemolytic activity of USA300 (MRSA strain) with & without exposure to triamcinolone acetonide (TAC) or MH. USA300 was cultured in Tryptic soy broth (TSB) containing 0.1% w/v TAC or MH (ManukoraTM; 1.25%, 2.5%, 5% & 10% w/v). Growth after MH exposure was evaluated by measuring optical density at 600nm (OD600). Due to TAC’s low solubility which interfered with OD readings, we evaluated growth by colony forming units (CFUs) after 5 hrs. After 24 hrs, supernatants were filtered (0.22-μm) and spotted onto rabbit blood agar (RBA) to quantify hemolytic activity (diameter of zones of clearance). TAC exposure resulted in a slightly lower CFUs (avg 0.82 x 109) than TSB alone (avg 1.26 x 109 CFU; n=2/condition), suggesting that TAC did not significantly inhibit USA300 growth. In contrast, MH markedly reduced OD600 values by 5, 23, 82 & 92% in 1.25, 2.5, 5, & 10% MH, respectively, compared to TSB alone (mean; n=3, p<0.03) after 5 hrs. This inhibitory effect of MH on USA300 growth was notable with the most significant reduction (mean 35%; n=3, p=0.001) observed at the highest concentration (10%) that was still observed at 24 hrs. Supernatants from TAC-incubated USA300 reduced clearance zones on RBA (avg 1.8±0.4 mm) compared to TSB alone (avg 5.8±0.4 mm; n=2/condition). Whereas, MH, inhibited all hemolytic activity, even at the lowest concentration 1.25% (n=3). Collectively, these findings demonstrate that at even at low MH concentrations, commonly found in cosmetic products, there is a greater effect on S. aureus growth as well as nearly complete inhibition of hemolytic activity compared to TAC, the most commonly used topical TCS.

Biofilm Formation by Multidrug-Resistant Serotypes of Salmonella Isolated from Fresh Products: Effects of Nutritional and Environmental Conditions

Salmonella serotypes can develop biofilms in fresh food products. This study focused on determining the antimicrobial resistance profile and the effects of different growth media and environmental conditions on biofilm formation by multidrug-resistant serotypes of Salmonella. All 49.4% of the Salmonella strains (five serotypes) were multidrug resistant. Assessment of the ability to form biofilms using the crystal violet staining method revealed that 95.6% of the strains of Salmonella were strong biofilm producers in 96-well polystyrene microtiter plates. Overall, 59.3% of the Salmonella strains showed the rdar (red dry and rough colony) morphotype, 2.1% pdar (pink dry and rough colony), 27.4% bdar (brown dry and rough colony) and 10.9% saw (smooth and white colony), at two temperatures (22 and 35 °C). Mono-species biofilms of Salmonella serotypes showed a mean cell density of 8.78 log10 CFU/cm2 ± 0.053 in TSBS (1/20 diluted TSB (tryptic soy broth) + 1% strawberry residues) and 8.43 log10 CFU/cm2 ± 0.050 in TSBA (1/20 diluted TSB + 1% avocado residues) on polypropylene type B (PP) (p &lt; 0.05). In addition, epifluorescence microscopy and scanning electron microscopy (SEM) enabled visualizing the bacteria and extracellular polymeric substances of biofilms on PP. Salmonella form biofilms depending on the serotype of the strains and the environmental conditions. Mono-species biofilms formed by Salmonella serotypes respond to nutrient limitation with the use of simplified culture media such as TSBA and TSBS.

Neonatal sepsis due to non-albicans Candida species and their susceptibility to antifungal agents: first report from Bangladesh

Background and objectives: Frequency of neonatal sepsis in Neonatal Intensive Care Units (NICU) has been increasing worldwide over the last decades. The emergence of non-albicans Candida (NAC) species and their resistance to common antifungal agents become an important preventive and therapeutic issue. The present study was undertaken to find out the role of NAC species in neonatal sepsis/candidemia in the NICUs of hospitals of Dhaka city. The susceptibility pattern of NAC species to antifungal agents was also determined.&#x0D; Materials and methods: Suspected cases of neonatal sepsis admitted in NICU of four tertiary care hospitals of Dhaka city, from March to December 2018 were enrolled. In this cross sectional study, blood samples were collected from neonates with suspected sepsis for culture. Identification of Candida species was done by carbohydrate (CHO) assimilation tests using swab auxanographic technique, CHO impregnated yeast nitrogen base plate method (YNB), microtiter plate based miniaturized method and by HiCromeTM Candida Differential Media. Susceptibility of the isolated Candida species to antifungal agents was determined by disk diffusion (DD) and by minimum inhibitory concentration (MIC) methods. MIC was determined by broth microdilution method using RPMI 1640 and trypticase soy broth (TSB).&#x0D; Results: In the present study, NAC species were isolated from 39.7% neonates. C. tropicalis was the predominant species (81.0%) followed by C. parapsilosis (12.1%), C. auris (5.2%) and C. dubliniensis (1.7%). Isolated NAC species were 98.3% sensitive to voriconazole. Sensitivity to fluconazole, ketoconazole, itraconazole, and clotrimazole was 3.5%, 15.5%, 86.2% and 56.9% respectively by DD method. All the isolates (100%) were sensitive to miconazole and nystatin. All the C. tropicalis, C. auris and C. dubliniensis were sensitive to amphotericin B and anidulafungin. One and four C. parapsilosis were found resistant to amphotericin B and anidulafungin respectively. The MIC results obtained by using RPMI 1640 and TSB as growth medium were concordant suggesting that TSB media was a good alternative to expensive RPMI 1640.&#x0D; Conclusion: The advent of NAC species merits attention as they are highly resistant to most of the azoles. Therefore, speciation of Candida in neonatal candidemia is essential to institute appropriate antifungal therapy.&#x0D; Ibrahim Med. Coll. J. 2020; 14(2): 19-26

Fate of Salmonella and Enterococcus faecium during Pilot-Scale Spray Drying of Soy Protein Isolate

Outbreaks and recalls associated with microbial contamination of powdered foods have raised concern for the safety of the spray-drying process and its products. However, little research on the fate of bacteria during the spray-drying process has been done, leaving much unknown about the risks of contamination in spray dryers. Therefore, quantifying the contamination levels of Salmonella and Enterococcus faecium (as a surrogate) in various locations within a pilot-scale spray dryer can help illustrate the distribution of bacterial contamination, including in the final product. A 10% (w/w) dispersion of water and soy protein isolate was mixed with tryptic soy broth containing yeast extract inoculated with Salmonella enterica serovar Enteritidis phage type 30 (PT30) or E. faecium strain NRRL B-2354. This dispersion was spray dried using a pilot-scale tall-form cocurrent spray dryer at an inlet air temperature of 180, 200, or 220°C. After drying, samples of powder from eight locations within the system were collected or surface swabbed, plated, and enumerated. Spray drying achieved 2.40 to 4.15 and 2.33 to 2.83 log reductions in the concentrations of Salmonella and E. faecium, respectively, in the final powder product accumulated in the dryer's collectors. Salmonella and E. faecium were found in various concentrations in all locations within the spray dryer after a complete drying cycle. Differences in inlet air temperature between 180 and 220°C had no significant effect on the inactivation levels. As a surrogate, E. faecium was more resistant to spray drying than Salmonella. Overall, spray drying is capable of significant bacterial reduction in the final powder product, which can be combined with other hurdle technologies. However, adequate cleaning and sanitization procedures must be taken into consideration to prevent cross-contamination.

Identifikasi Bakteri pada Ikan Pindang Tongkol (Euthynnus affinis) di Pasar Tradisional Semarapura, Klungkung, Bali

Mackerel tuna (Euthynnus affinis) brine salting is a famous seafood product in Indonesia due to its good nutrition content. Bacterial contamination on this product may increase risk of foodborne diseases to happen, in addition to a decrease in quality and nutritionous value of the product. The main objective of this research was to identify bacteria contaminating the brine salting Mackerel tuna (Euthynnus affinis) sold in Tradisional Markets in Klungkung, Bali, Indonesia. Polymerase Chain Reaction (PCR) was apllied in the process of identification of bacterial contaminants. Samples were collected from 5 fish kiosks at the Tradisional market in Klungkung, Bali Indonesia. Each sample was cultured in Blood Agar or Nutrient Agar. Colonies with different morfology were selected, enriched in Tryptic Soy Broth (TSB), and their DNA was extracted before being amplified with 16S rRNA primers. This PCR products were then sequenced, and the results were compared with those available in the geneBank. Basic Local Alignment Search Tool was conducted with help of MEGA software version 6. Seven bacterial species were identified, and these included Serratia nematodiphila, Bacillus cereus, Shewanella seohaensis, Vibrio alginolyticus, Kurthia gibsonii, Enterobacter cloacae, and Staphylococcus sciuri. All but one Kurthia gibsonii are potensial pathogens in human. Staphylococcus sciuri is not commonly found in seafood. The present of such species in the mackerel tuna brine salting product was probably due to poor handling during the process and distribution.

Isolation and Identification of Biosurfactant Producing Bacteria from Slaughterhouse Wastewater

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

Isolation and Identification of Biosurfactant Producing Bacteria from Tofu Liquid Waste UD. Dika Putra, Riau Province

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.

PhageDx™ E. coli O157:H7 Assay in Ground Beef Level 3 Modification to Add 375 g Beef Trim and New Method Procedure for 25 g Ground Beef: AOAC Performance Tested MethodSM 081601.

BackgroundThePhageDx™ E. coli O157:H7 Assay originally required a 5 h enrichment period and a 10 mL test sample for 25 g ground beef. The proposed method modifications seek to include beef trim (375 g) matrix and a new method procedure for 25 g ground beef comprised of 6–7 h enrichment and 1 mL test sample.ObjectiveTo validate the method modifications for PhageDx E. coli O157:H7 Assay to include beef trim (375 g) matrix and modify enrichment time and test sample size for 25 g ground beef.MethodFor both matrixes, pre-warmed tryptic soya broth (TSB; 42 ± 1°C) was added in a 3:1 (media:sample size) ratio, blended, and enriched for 9–10 h (beef trim, 375 g) or 6 h (ground beef, 25 g) at 42 ± 1°C. One milliliter samples were transferred to a microfuge tube, centrifuged, and the supernatant removed. The pellet was resuspended in 0.2 mL of TSB and phage reagent was added. Samples were incubated for 2 h at 37 ± 1°C. After infection, the sample was centrifuged, and 150 µl of the supernatant was transferred to a 96-well plate. Then, lysis buffer and luciferase substrate were added and the sample read on a luminometer to determine the presence or absence of E. coli O157:H7.ResultsNo significant differences were found between the PhageDx E. coli O157:H7 Assay method modifications and the U.S. Department of Agriculture Food Safety and Inspection Service (FSIS) Microbiology Laboratory Guidebook (MLG) reference method.ConclusionsThe independent study demonstrated that the PhageDx E. coli O157:H7 Assay method modifications meet the qualifications for Performance Tested Method SM status.HighlightsThe PhageDx E. coli O157:H7 Assay is a rapid detection method capable of detecting E. coli O157:H7 in 25 g ground beef and 375 g beef trim samples in as little as 8.5 h and 11.5 h total turn around time, respectively.

Production of Membrane Vesicles in Listeria monocytogenes Cultured with or without Sub-Inhibitory Concentrations of Antibiotics and Their Innate Immune Responses In Vitro.

Listeriosis is a food-borne illness caused by Listeria monocytogenes. Ampicillin (AMP) alone or in combination with gentamicin (GEN) is the first-line treatment option. Membrane vesicle (MV) production in L. monocytogenes under antibiotic stress conditions and pathologic roles of these MVs in hosts have not been reported yet. Thus, the aim of this study was to investigate the production of MVs in L. monocytogenes cultured with sub-minimum inhibitory concentrations (MICs) of AMP, GEN, or trimethoprim/sulfamethoxazole (SXT) and determine pathologic effects of these MVs in colon epithelial Caco-2 cells. L. monocytogenes cultured in tryptic soy broth with 1/2 MIC of AMP, GEN, or SXT produced 6.0, 2.9, or 1.5 times more MV particles, respectively, than bacteria cultured without antibiotics. MVs from L. monocytogenes cultured with AMP (MVAMP), GEN (MVGEN), or SXT (MVSXT) were more cytotoxic to Caco-2 cell than MVs obtained from cultivation without antibiotics (MVTSB). MVAMP induced more expression of tumor necrosis factor (TNF)-α gene than MVTSB, MVGEN and MVSXT, whereas MVTSB induced more expression of interleukin (IL)-1β and IL-8 genes than other MVs. Expression of pro-inflammatory cytokine genes by L. monocytogenes MVs was significantly inhibited by proteinase K treatment of MVs. In conclusion, antibiotic stress can trigger the biogenesis of MVs in L. monocytogenes and MVs produced by L. monocytogenes exposed to sub-MIC of AMP can induce strong pro-inflammatory responses by expressing TNF-α gene in host cells, which may contribute to the pathology of listeriosis.

High concentration of sodium chloride could induce the viable and culturable states of Escherichia coli O157:H7 and Salmonella enterica serovar Enteritidis.

In the present study, Escherichia coli O157:H7 and Salmonella enterica serovar Enteritidis were transferred into Luria-Bertani medium without NaCl (LBWS) and adjusted to various pHs (4, 5, 6 and 7) with lactic acid containing 0·75, 5, 10 and 30% NaCl, and stored at 25°C until the bacterial populations reached below detectable levels on tryptic soy agar (TSA). Although E.coli O157:H7 and S.Enteritidis did not grow on TSA when incubated in LBWS with 30% NaCl for 35 and 7days, more than 60 and 70% of the bacterial cells were shown to be viable via fluorescent staining with SYTO9 and propidium iodide (PI), respectively, suggesting that a number of cells could be induced into the viable but nonculturable (VBNC) state. These bacteria that were induced into a VBNC state were transferred to a newly prepared tryptic soy broth (TSB) and then incubated at 37°C for several days. After more than 7days, E.coli O157:H7 and S.Enteritidis regained their culturability. We, therefore, suggest that E.coli O157:H7 and S.Enteritidis entered the VBNC state under the adverse condition of higher salt concentrations and were revived when these conditions were reversed.