Antimicrobial susceptibility testing of Mycobacterium tuberculosis complex for first and second line drugs by broth dilution in a microtiter plate format.

PubMed

Hall, Leslie; Jude, Kurt P; Clark, Shirley L; Wengenack, Nancy L

2011-06-24

The rapid detection of antimicrobial resistance is important in the effort to control the increase in resistant Mycobacterium tuberculosis (Mtb). Antimicrobial susceptibility testing (AST) of Mtb has traditionally been performed by the agar method of proportion or by macrobroth testing on an instrument such as the BACTEC (Becton Dickinson, Sparks, MD), VersaTREK (TREK Diagnostics, Cleveland, OH) or BacT/ALERT (bioMérieux, Hazelwood, MO). The agar proportion method, while considered the "gold" standard of AST, is labor intensive and requires calculation of resistance by performing colony counts on drug-containing agar as compared to drug-free agar. If there is ≥1% growth on the drug-containing medium as compared to drug-free medium, the organism is considered resistant to that drug. The macrobroth methods require instrumentation and test break point ("critical") drug concentrations for the first line drugs (isoniazid, ethambutol, rifampin, and pyrazinamide). The method described here is commercially available in a 96 well microtiter plate format [MYCOTB (TREK Diagnostics)] and contains increasing concentrations of 12 antimicrobials used for treatment of tuberculosis including both first (isoniazid, rifampin, ethambutol) and second line drugs (amikacin, cycloserine, ethionamide, kanamycin, moxifloxacin, ofloxacin, para-aminosalicylic acid, rifabutin, and streptomycin). Pyrazinamide, a first line drug, is not included in the microtiter plate due to its need for acidic test conditions. Advantages of the microtiter system include both ease of set up and faster turn around time (14 days) compared with traditional agar proportion (21 days). In addition, the plate can be set up from inoculum prepared using either broth or solid medium. Since the microtiter plate format is new and since Mtb presents unique safety challenges in the laboratory, this protocol will describe how to safely setup, incubate and read the microtiter plate.

Evaluation of Cariogenic Bacteria

PubMed Central

Nishikawara, Fusao; Nomura, Yoshiaki; Imai, Susumu; Senda, Akira; Hanada, Nobuhiro

2007-01-01

Objectives The evaluation of Mutans streptococci (MS) is one of the index for caries risk. DentocultTM and CRTTM are commercial kits to detect and evaluate MS, conveniently. However, the evaluation of MS has also been carried out simply using an instruction manual. But the instruction manual is not easy to use for evaluation of MS. The aim of this study was to examine the utility of modified Mitis-Salivalius Bacitracin (MSB) agar medium compared with MSB agar medium and commercial kits, and to establish a convenient kit (mMSB-kit) using modified MSB agar. Methods The MS in stimulated saliva from 27 subjects were detected by MSB, modified MSB agar medium and commercial kits. Laboratory and clinically isolated strains of MS were similarly evaluated. The ratios of MS in detected bacteria were compared by ELISA. Results The scores using an mMSB-kit on the basis of modified MSB agar medium were tabulated. Saliva samples showed different levels of MS between culture methods and the commercial kit. Some samples which were full of MS were not detected by the commercial kit. The detection of MS by modified MSB agar medium and mMSB-kit were significantly higher when compared with MSB agar medium,CRTTM, (P< .01) and Dentocult SMTM (P<.05). Conclusion The sensitivity for detection of MS is higher for modified MSB agar medium when compared with MSB agar medium. The mMSB-kit can be used simply, and can be an important contributor for the evaluation of MS as a caries risk factor. PMID:19212495

Morphological Variations in Conidia of Arthrobotrys oligospora on Different Media.

PubMed

Singh, R K; Kumar, Niranjan; Singh, K P

2005-06-01

Most commonly occurring predacious fungus Arthrobotrys oligospora showed great variation in size and shape of conidia on some media. The formation of larger conidia was recorded on beef extract and nutrient agar media. The length of conidia in Richard's YPSS, Sabouraud's, PDA and corn meal agar media was of medium size while smaller conidia were produced on Czapek's, Jensen's, Martin's medium. Maximum width of conidia was recorded on YPSS medium followed by Sabouraud's medium. The average size of spores on nematode infested corn meal agar medium was slightly increased than those on corn meal agar medium.

[Effect of nutritional stress on autophagy in free-living amoeba].

PubMed

Wang, Nan-Ning; Tan, Yu-Zhen; Wang, Hai-Jie

2010-12-30

To investigate the change of autophagy and morphological characteristics of the autophagic structures in free-living amoeba under nutritional stress. Free-living amoebae were incubated on the agaric solid medium which had been covered with Escherichia cdi in control group. In the experiment group, amoebae incubated on the agaric solid medium with E. coli were collected and moved to another solid medium without E. coli and incubated for 12 h. The morphological changes of free-living amoeba in the medium without E. coli were viewed with scanning electron microscope. The changes of autophagy and the structural features of the autophagosome precursors, autophagosomes and autophagolysosomes in amoeba were examined with transmission electron microscope, and the cross-section areas of the autophagic structures and cytoplasm were measured with an image analyzer. The autophagosomes in the organism were labeled with monodansylcadaverine (MDC) staining and quantitated using laser scanning confocal microscope. In the control group, free-living amoebae were all in the form of trophozoite. In the experiment group, trophozoites were induced to transform to cysts gradually. In control group, amoeba was full of fragment of E. coli. There was merely little autophagy with fewer autophagic structures in amoeba. When compared with the control group, the autophagic abilities of amoeba were enhanced significantly, number of autophagic structures increased in the experiment group. In addition, the ratio of the cross-sectional areas of the autophagic structures to that of the cytoplasm of amoeba was greater (P < 0.05 or 0.01). There was fragment of E. coli that was not digested in some of the amoebae. In the circumstance of nutritional stress, amoebic trophozoites were induced to transform to cysts gradually. The autophagic ability of free-living amoeba significantly enhanced.

Back to the kitchen: food-grade agar is a low-cost alternative to bacteriological agar.

PubMed

Petrovski, Steve; Tillett, Daniel

2012-10-15

Food-grade agar can be used as a low-cost substitute for bacteriological agar in the preparation of solid microbial media. No difference was observed in the colony morphology, growth rate, or viability of bacteria grown on solid media prepared using food-grade agar as compared with using bacteriological-grade agar. This simple tip can reduce the cost of the most common solid media by 80% or more. Copyright © 2012 Elsevier Inc. All rights reserved.

Physical Forces Shape Group Identity of Swimming Pseudomonas putida Cells.

PubMed

Espeso, David R; Martínez-García, Esteban; de Lorenzo, Víctor; Goñi-Moreno, Ángel

2016-01-01

The often striking macroscopic patterns developed by motile bacterial populations on agar plates are a consequence of the environmental conditions where the cells grow and spread. Parameters such as medium stiffness and nutrient concentration have been reported to alter cell swimming behavior, while mutual interactions among populations shape collective patterns. One commonly observed occurrence is the mutual inhibition of clonal bacteria when moving toward each other, which results in a distinct halt at a finite distance on the agar matrix before having direct contact. The dynamics behind this phenomenon (i.e., intolerance to mix in time and space with otherwise identical others) has been traditionally explained in terms of cell-to-cell competition/cooperation regarding nutrient availability. In this work, the same scenario has been revisited from an alternative perspective: the effect of the physical mechanics that frame the process, in particular the consequences of collisions between moving bacteria and the semi-solid matrix of the swimming medium. To this end, we set up a simple experimental system in which the swimming patterns of Pseudomonas putida were tested with different geometries and agar concentrations. A computational analysis framework that highlights cell-to-medium interactions was developed to fit experimental observations. Simulated outputs suggested that the medium is compressed in the direction of the bacterial front motion. This phenomenon generates what was termed a compression wave that goes through the medium preceding the swimming population and that determines the visible high-level pattern. Taken together, the data suggested that the mechanical effects of the bacteria moving through the medium created a factual barrier that impedes to merge with neighboring cells swimming from a different site. The resulting divide between otherwise clonal bacteria is thus brought about by physical forces-not genetic or metabolic programs.

A Novel Double Subculture Method and Its Theory for the Enumeration of Injured Cells in Stressed Microbial Population.

PubMed

Tsuchido, Tetsuaki

2017-01-01

　A novel double subculture method, termed DiVSaL (Differential Viabilities between Solid and Liquid media) method, for the enumeration of injured cell population of a microorganism, which occurs after some sublethal to lethal treatment, was proposed. In this method injured cells were enumerated as the differential value between viabilities determined with two different techniques, the conventional plate counting using a solid agar medium and the growth delay analysis using a liquid medium. In the former technique, the viable cell number is obtained as colony forming unit (CFU) formed on an agar medium where sublethally injured cells are as much rescued as possible. In the latter technique, on the other hand," the integrated viability" defined by Takano and Tsuchido (1982) is introduced and is calculated from the growth delay of a stressed population, referred to unstressed one. For the growth delay analysis, in this paper, not only the original theoretical model, where the specific growth rate (and therefore the defined G 10 value) does not change after the exposure to a stress treatment, but also a novel modified theory, where the parameter changes, is proposed. On the theoretical background, this DiVSaL method as a double subculture method can be used to enumerate the injured cells without selection by addition of some inhibitor or by nutritional shortage.

Degradation of Coal by the Fungi Polyporus versicolor and Poria monticola

PubMed Central

Cohen, Martin S.; Gabriele, Peter D.

1982-01-01

We report that two species of basidiomycete fungi (Polyporus versicolor and Poria monticola) grow in minimal liquid or solid medium when supplemented with crushed lignite coal. The fungi also grow directly on crushed lignite coal. The growth of both fungi was observed qualitatively as the production and extension of hyphae. No fungal growth occurred in minimal agar medium without coal. The fungi degraded solid lignite coal to a black liquid product which never appeared in cultures unless fungi and coal were present together. Apparently, lignite coal can serve as the principal substrate for the growth of the fungi. Infrared analyses of the liquid products of lignite degradation showed both similarities to and differences from the original lignite. Images PMID:16346060

Microbiological characteristics of "androlla", a Spanish traditional pork sausage.

PubMed

García Fontán, María C; Lorenzo, José M; Parada, Ana; Franco, Inmaculada; Carballo, Javier

2007-02-01

Counts of total aerobic mesophilic microflora, lactic acid bacteria, salt-tolerant microflora, Enterobacteriaceae, enterococci, moulds and yeasts, and staphylococci, and some physico-chemical parameters (total solids, NaCl and nitrate contents and pH and aw values) were determined in 20 units of "androlla", a traditional dry-fermented sausage made in the NW of Spain. In general, high counts of all the investigated microbial groups were observed, with average values of 8.99 +/- 0.46 log cfu/g for the total aerobic mesophilic microflora, 9.11 +/- 0.16 log cfu/g for the lactic acid bacteria, 6.87 +/- 0.68 log cfu/g for the salt-tolerant microflora, 2.80+/-1.85 log cfu/g for the Enterobacteriaceae, 3.25 +/- 1.86 log cfu/g for the enterococci, 4.30 +/- 1.73 log cfu/g for the moulds and yeasts, and 3.62 +/- 0.60 log cfu/g for the staphylococci. From MRS agar, SPC agar + 7.5% NaCl, VRBG agar, and KAA agar, 10 colonies were randomly taken from each androlla unit and from each culture medium. A total of 200 strains per culture medium were then identified using the classical methods. Among the isolates from MRS agar, Lactobacillus sakei predominated, followed by Lactobacillus curvatus, Lactobacillus alimentarius and Lactobacillus plantarum. Of the 200 isolates obtained from SPC agar + 7.5% NaCl, only 56 strains belonged to the Staphylococcaceae or Micrococcaceae families. Among the Staphylococcaceae, Staphylococcus xylosus was the main species, followed by Staph. epidermidis; Staph. equorum, Staph. capitis and Staph. saprophyticus were isolated in very low proportions. Among the Micrococcaceae, Micrococcus luteus predominated, followed by Micrococcus lylae, Kocuria varians and Kocuria kristinae. Of the 150 isolates obtained from VRBG agar, Hafnia alvei was the main species, followed by Serratia liquefaciens and Enterobacter amnigenus; six isolates were identified as Salmonella. Among the 190 isolates obtained from KAA agar, 122 were considered enterococci; 20 isolates were identified as Enterococcus faecium, one as Enterococcus faecalis and 101 as Enterococcus inter faecalis-faecium.

Chromogenic agar medium for detection and isolation of Escherichia coli serogroups O26, O45, O103, O111, O121, and O145 from fresh beef and cattle feces.

PubMed

Kalchayanand, Norasak; Arthur, Terrance M; Bosilevac, Joseph M; Wells, James E; Wheeler, Tommy L

2013-02-01

Non-O157 Shiga toxin-producing Escherichia coli (STEC) strains are clinically important foodborne pathogens. Unlike E. coli O157:H7, these foodborne pathogens have no unique biochemical characteristics to readily distinguish them from other E. coli strains growing on plating media. In this study, a chromogenic agar medium was developed in order to differentiate among non-O157 STEC strains of serogroups O26, O45, O103, O111, O121, and O145 on a single agar medium. The ability of this chromogenic agar medium to select and distinguish among these pathogens is based on a combination of utilization of carbohydrates, b -galactosidase activity, and resistance to selective agents. The agar medium in combination with immunomagnetic separation was evaluated and successfully allowed for the detection and isolation of these six serogroups from artificially contaminated fresh beef. The agar medium in combination with immunomagnetic separation also allowed successful detection and isolation of naturally occurring non-O157 STEC strains present in cattle feces. Thirty-five strains of the top six non-O157 STEC serogroups were isolated from 1,897 fecal samples collected from 271 feedlot cattle. This chromogenic agar medium could help significantly in routine screening for the top six non-O157 STEC serogroups from beef cattle and other food.

The Influence of Microgravity on Invasive Growth in Saccharomyces cerevisiae

NASA Astrophysics Data System (ADS)

Van Mulders, Sebastiaan E.; Stassen, Catherine; Daenen, Luk; Devreese, Bart; Siewers, Verena; van Eijsden, Rudy G. E.; Nielsen, Jens; Delvaux, Freddy R.; Willaert, Ronnie

2011-01-01

This study investigates the effects of microgravity on colony growth and the morphological transition from single cells to short invasive filaments in the model eukaryotic organism Saccharomyces cerevisiae. Two-dimensional spreading of the yeast colonies grown on semi-solid agar medium was reduced under microgravity in the Σ1278b laboratory strain but not in the CMBSESA1 industrial strain. This was supported by the Σ1278b proteome map under microgravity conditions, which revealed upregulation of proteins linked to anaerobic conditions. The Σ1278b strain showed a reduced invasive growth in the center of the yeast colony. Bud scar distribution was slightly affected, with a switch toward more random budding. Together, microgravity conditions disturb spatially programmed budding patterns and generate strain-dependent growth differences in yeast colonies on semi-solid medium.

Acceleration of Antimicrobial Susceptibility Testing of Positive Blood Cultures by Inoculation of Vitek 2 Cards with Briefly Incubated Solid Medium Cultures

PubMed Central

Idelevich, Evgeny A.; Schüle, Isabel; Grünastel, Barbara; Wüllenweber, Jörg; Peters, Georg

2014-01-01

Briefly incubated agar cultures from positive blood cultures were used for antimicrobial susceptibility testing (AST) by Vitek 2. The cultivation time until inoculation was 3.8 h for Gram-positive cocci and 2.4 h for Gram-negative rods. The error rates were low, providing early and reliable AST without additional time or cost expenditure. PMID:25165084

Isolation and in vitro culture of trypanosomes from Leptodactylus ocellatus from the Atlantic Forest in a new experimental culture medium.

PubMed

Lemos, M; Souza, C S F; da Costa, S C Gonçalves; Souto-Padrón, T; D'Agosto, M

2013-02-01

The purpose of this study was to verify the in vitro development of Trypanosoma sp. isolated from Leptodactylus ocellatus frogs under a new protocol using a biphasic medium composed of Novy, McNeal, and Nicolle (NNN) blood agar medium as a solid phase and liver infusion, brain heart infusion, and tryptose (LIBHIT) medium as a liquid phase. Blood forms, collected by cardiac puncture or after the maceration of different organs, were inoculated in culture tubes containing the biphasic medium composed by NNN and LIBHIT. Trypanosomes were observed 4 days postinoculation; most bloodstream trypomastigotes had differentiated into epimastigotes and amastigotes by this time. Trypomastigotes were again observed in older cultures (7 days). Parasites were successfully subcultured for 8 mo in this medium and successfully cryopreserved. The present study provides a new protocol medium for the isolation and culture of anuran trypanosomes.

Production of fungal antibiotics using polymeric solid supports in solid-state and liquid fermentation.

PubMed

Bigelis, Ramunas; He, Haiyin; Yang, Hui Y; Chang, Li-Ping; Greenstein, Michael

2006-10-01

The use of inert absorbent polymeric supports for cellular attachment in solid-state fungal fermentation influenced growth, morphology, and production of bioactive secondary metabolites. Two filamentous fungi exemplified the utility of this approach to facilitate the discovery of new antimicrobial compounds. Cylindrocarpon sp. LL-Cyan426 produced pyrrocidines A and B and Acremonium sp. LL-Cyan416 produced acremonidins A-E when grown on agar bearing moist polyester-cellulose paper and generated distinctly different metabolite profiles than the conventional shaken or stationary liquid fermentations. Differences were also apparent when tenfold concentrated methanol extracts from these fermentations were tested against antibiotic-susceptible and antibiotic-resistant Gram-positive bacteria, and zones of inhibition were compared. Shaken broth cultures of Acremonium sp. or Cylindrocarpon sp. showed complex HPLC patterns, lower levels of target compounds, and high levels of unwanted compounds and medium components, while agar/solid support cultures showed significantly increased yields of pyrrocidines A and B and acremonidins A-E, respectively. This method, mixed-phase fermentation (fermentation with an inert solid support bearing liquid medium), exploited the increase in surface area available for fungal growth on the supports and the tendency of some microorganisms to adhere to solid surfaces, possibly mimicking their natural growth habits. The production of dimeric anthraquinones by Penicillium sp. LL-WF159 was investigated in liquid fermentation using various inert polymeric immobilization supports composed of polypropylene, polypropylene cellulose, polyester-cellulose, or polyurethane. This culture produced rugulosin, skyrin, flavomannin, and a new bisanthracene, WF159-A, after fermentation in the presence and absence of polymeric supports for mycelial attachment. The physical nature of the different support systems influenced culture morphology and relative metabolite yields, as determined by HPLC analysis and measurement of antimicrobial activity. The application of such immobilized-cell fermentation methods under solid and liquid conditions facilitated the discovery of new antibiotic compounds, and offers new approaches to fungal fermentation for natural product discovery.

Gentamicin: effect on E. coli in space

NASA Technical Reports Server (NTRS)

Kacena, M. A.; Todd, P.

1999-01-01

Previous investigations have shown that liquid bacterial cultures grown in space flight were not killed as effectively by antibiotic treatments as were cultures grown on Earth. However, the cause for the decreased antibiotic effectiveness remains unknown. Possible explanations include modified cell proliferation and modified antibiotic transport in the culture medium. Escherichia coli cultures were grown in space flight (STS-69 and STS-73), with and without gentamicin, on a solid agar substrate thus eliminating fluid effects and reducing the unknowns associated with space-flight bacterial cultures in suspension. This research showed that E. coli cultures grown in flight on agar for 24 to 27 hours experienced a heightened growth compared to simultaneous controls. However, addition of gentamicin to the agar killed the bacteria such that both flight and ground control E. coli samples had similar final cell concentrations. Therefore, while the reported existence of a decrease in antibiotic effectiveness in liquid cultures remains unexplained, these data suggest that gentamicin in space flight was at least as effective as, if not more effective than, on Earth, when E. coli cells were grown on agar.

Usefulness of Chromogenic CromoCen® AGN agar medium for the identification of the genus Aeromonas: Assessment of faecal samples.

PubMed

Aguilera-Arreola, M G; Portillo-Muñoz, M I; Rodríguez-Martínez, C; Castro-Escarpulli, G

2012-08-01

Selective screening media for the detection and identification of Aeromonas strains are needed to guide primary isolation procedures in the clinical laboratory. This study compared the selective CromoCen® AGN chromogenic agar medium for the detection and identification of Aeromonas strains that were isolated from various samples against the conventional selective agar media that are commonly used for the isolation of this organism in food, environmental and clinical samples. The Miles and Misra and ecometric methods were used to evaluate the microbiological performance of CromoCen® AGN chromogenic agar medium, which was shown to be satisfactory. A total of 14 reference Aeromonas strains, 44 wild strains and 106 clinical stool specimens were examined using both non-chromogenic selective agars that are commonly used for Aeromonas isolation and CromoCen® AGN agar. The latter exhibited 94.73% sensitivity and 100% specificity for the various samples. On CromoCen® AGN agar medium, Aeromonas formed colonies with light green, greenish and salmon pigments with or without a surrounding wide transparent zone (halo) of 2-3mm in diameter around the entire border. This medium is recommended for the isolation and potential identification of the Aeromonas genus. Copyright © 2012 Elsevier B.V. All rights reserved.

Chocolate agar, a differential medium for gram-positive cocci.

PubMed Central

Gunn, B A

1984-01-01

Reactions incurred on chocolate agar by gram-positive cocci were correlated with species identity. Darkening and clearing of the medium was usually associated with the species Staphylococcus epidermidis, Staphylococcus aureus, Staphylococcus simulans, and Streptococcus faecalis. Yellowing of chocolate agar was associated with alpha-hemolytic species of Streptococcus. The study demonstrated that reactions occurring on chocolate agar are useful in identifying gram-positive cocci. PMID:6490866

Induction of a global stress response during the first step of Escherichia coli plate growth.

PubMed

Cuny, Caroline; Lesbats, Maïalène; Dukan, Sam

2007-02-01

We have investigated the first events that occur when exponentially grown cells are transferred from a liquid medium (Luria-Bertani [LB]) to a solid medium (LB agar [LBA]). We observed an initial lag phase of 180 min for the wild type MG1655 without any apparent growth. This lack of growth was independent of the bacterial physiological state (either the stationary or the exponential phase), the solid medium composition, or the number of cells on the plate, but it was dependent on the bacterial genotype. Using lacZ-reporter fusions and two-dimensional electrophoresis analysis, we observed that when cells from exponential-phase cultures were plated on LBA, several global regulons, like heat shock regulons (RpoH, RpoE, CpxAR) and oxidative-stress regulons (SoxRS, OxyR, Fur), were immediately induced. Our results indicate that in order to grow on plates, bacteria must not only adapt to new conditions but also perceive a real stress.

Morphological identification of Candida species on glucose agar, rice extract agar and corn meal agar with and without Tween-80.

PubMed

Joshi, K R; Solanki, A; Prakash, P

1993-01-01

A comparative study for the identification of 32 known strains of Candida species on the basis of morphology on glucose agar, rice extract agar and corn meal agar with and without Tween 80 revealed that when Tween 80 is incorporated in the media identification is possible for 96.8% of the species within 48 hours on rice extract agar and for 96.8% of the species within 48 hours on rice extract agar and for 90.6% of the species on glucose agar. The germ tubes and chlamydospores were also produced more on rice extract agar than on 0.1% glucose agar. Rice extract agar with Tween 80 can be used as single medium for morphologic identification of Candida species. The inoculated medium is first incubated at 37 degrees C for 3 hours and examined for germ tube formation and then incubated at 25 degrees C for 24 to 72 hours and examined for appearance of chlamydospores and mycelial morphology.

Nutrient agar with sodium chloride supplementation for presumptive detection of Moraxella catarrhalis in clinical specimens.

PubMed

Nishiyama, Hiroyuki; Saito, Ryoichi; Chida, Toshio; Sano, Kazumitsu; Tsuchiya, Tatsuyuki; Okamura, Noboru

2012-04-01

We previously reported that Nissui nutrient agar (N medium) promoted the growth of Moraxella catarrhalis but not commensal Neisseria spp. In the present study, we examined which constituent of N medium was responsible for the selective growth of M. catarrhalis using 209 M. catarrhalis and 100 commensal Neisseria spp. clinical strains. We found that peptone, but not meat extract or agar of N medium, had growth-promoting or growth-inhibiting ability with respect to M. catarrhalis and commensal Neisseria spp. Thus, we investigated the amino acid content of N peptone and found it had higher concentrations of amino acids than other commercial peptone products. On varying the sodium chloride concentration of reconstituted N medium, we noted that the concentration was an important factor in bacterial growth differences. Varying the sodium chloride concentration of other commercial nutrient agars achieved similar results to those for N medium. This is, to our knowledge, the first study observing that sodium chloride concentration is responsible for difference in growth between the two organisms. We also successfully isolated colonies of M. catarrhalis from respiratory specimens on N medium, whereas the growth of commensal Neisseria spp. was inhibited, and by adding bovine hematin and β-NAD we were able to isolate Haemophilus influenzae colonies as efficiently as with a chocolate agar. In conclusion, nutrient agar can be used as a medium for the preferential isolation of M. catarrhalis from upper respiratory tract specimens.

Development of a More Sensitive and Specific Chromogenic Agar Medium for the Detection of Vibrio parahaemolyticus and Other Vibrio Species.

PubMed

Yeung, Marie; Thorsen, Trevor

2016-11-08

Foodborne infections in the US caused by Vibrio species have shown an upward trend. In the genus Vibrio, V. parahaemolyticus is responsible for the majority of Vibrio-associated infections. Thus, accurate differentiation among Vibrio spp. and detection of V. parahaemolyticus is critically important to ensure the safety of our food supply. Although molecular techniques are increasingly common, culture-depending methods are still routinely done and they are considered standard methods in certain circumstances. Hence, a novel chromogenic agar medium was tested with the goal of providing a better method for isolation and differentiation of clinically relevant Vibrio spp. The protocol compared the sensitivity, specificity and detection limit for the detection of V. parahaemolyticus between the new chromogenic medium and a conventional medium. Various V. parahaemolyticus strains (n=22) representing diverse serotypes and source of origins were used. They were previously identified by Food and Drug Administration (FDA) and Centers for Disease Control and Prevention (CDC), and further verified in our laboratory by tlh-PCR. In at least four separate trials, these strains were inoculated on the chromogenic agar and thiosulfate-citrate-bile salts-sucrose (TCBS) agar, which is the recommended medium for culturing this species, followed by incubation at 35-37 °C for 24-96 hr. Three V. parahaemolyticus strains (13.6%) did not grow optimally on TCBS, nonetheless exhibited green colonies if there was growth. Two strains (9.1%) did not yield the expected cyan colonies on the chromogenic agar. Non-V. parahaemolyticus strains (n=32) were also tested to determine the specificity of the chromogenic agar. Among these strains, 31 did not grow or exhibited other colony morphologies. The mean recovery of V. parahaemolyticus on the chromogenic agar was ~96.4% relative to tryptic soy agar supplemented with 2% NaCl. In conclusion, the new chromogenic agar is an effective medium to detect V. parahaemolyticus and to differentiate it from other vibrios.

Agar Underlay Method for Recovery of Sublethally Heat-Injured Bacteria

PubMed Central

Kang, D. H.; Siragusa, G. R.

1999-01-01

A method of recovering sublethally heat-injured bacteria was developed. The procedure (termed the agar underlay method) uses a nonselective agar underlaid with a selective medium. In a two-chambered petri dish, the Lutri plate (LP), a nonselective agar is inoculated with a population of sublethally heat-injured bacteria. After a 2-h repair incubation period, selective agar is added to the bottom chamber of the LP and incubated. By diffusing through the nonselective top agar, selective agents from the underlay medium impart selectivity to the system. By the agar underlay method, recovery rates of the heat-injured food-borne pathogens Escherichia coli O157:H7 and Salmonella typhimurium were not different (P > 0.05) from recovery rates determined with nonselective media. Sublethally heat-injured cells (60°C for 1.5 min in buffer or 80°C for 30 s on meat surfaces) grew and produced a typical colony morphology and color reaction when the agar underlay procedure was used with the appropriate respective selective agars. Unlike agar overlay methods for injury repair, the agar underlay procedure allows the typical selective-medium colony morphology to develop and allows colonies to be more easily picked for further characterization. Higher recovery rates of heat-injured fecal enterococci from bovine fecal samples and total coliforms from animal waste lagoons were obtained by the agar underlay method with selective agars than by direct plating on the respective selective media. PMID:10583985

Growth of Desulfovibrio on the surface of agar media.

PubMed

Iverson, W P

1966-07-01

Growth of Desulfovibrio desulfuricans (API strain) was found to take place in an atmosphere of hydrogen on the agar surface of complex media, including yeast extract (Difco), and Trypticase Soy Agar (BBL) without any added reducing agents. For growth on a 2% yeast extract-agar surface in the absence of hydrogen (nitrogen atmosphere), sodium lactate was required in the medium. Growth on the surface of Trypticase Soy Agar (TSA) under nitrogen took place readily in the absence of an added hydrogen donor. A medium (TSA plus salts) is described based upon the addition of sodium lactate (4 ml per liter), magnesium sulfate (2 g per liter), and ferrous ammonium sulfate (0.05%) to TSA, which appears suitable for the isolation and growth of Desulfovibrio on the surface of agar plates in an atmosphere of hydrogen. Sodium lactate does not appear to be essential in this medium for good growth and sulfate reduction in a hydrogen atmosphere, but is essential in a nitrogen atmosphere. Growth of Desulfovibrio (hydrogen atmosphere) on the agar surface of media commonly used for its cultivation as well as on an inorganic medium containing bicarbonate as a source of carbon is poor and erratic unless inoculated (Desulfovibrio) plates of TSA plus salts are incubated in the same container with plates of these media. This stimulatory effect of incubation with inoculated plates of TSA plus salts medium appears to be due to as yet unidentified volatile material produced by D. desulfuricans when growing on this medium. Another volatile material, or possibly the identical material, appears to act similarly to a hydrogen donor.

Rapid Direct Testing of Susceptibility of Mycobacterium tuberculosis to Isoniazid and Rifampin on Nutrient and Blood Agar in Resource-Starved Settings

PubMed Central

Ikram, Aamer; Coban, Ahmet Yilmaz; Martin, Anandi

2012-01-01

In this study, we evaluated the performance of blood agar (by macroscopic growth) and nutrient agar (by a microcolony detection method) for drug susceptibility testing of Mycobacterium tuberculosis against rifampin (RIF) and isoniazid (INH), using 67 smear-positive sputum specimens. The direct proportion method on Lowenstein-Jensen (LJ) medium was used as the “gold standard.” Compared with LJ medium, results for both media were in 100% agreement for RIF, while for INH the agreement levels for blood agar and nutrient agar were 98% and 95%, respectively. Within 2 weeks, 100% of specimens yielded results on blood agar, while 96.8% of specimens yielded results on nutrient agar. Our study showed that blood agar and nutrient agar can be used as alternative media for direct susceptibility testing of RIF and INH, especially in resource-poor settings. PMID:22357498

Rapid direct testing of susceptibility of Mycobacterium tuberculosis to isoniazid and rifampin on nutrient and blood agar in resource-starved settings.

PubMed

Satti, Luqman; Ikram, Aamer; Coban, Ahmet Yilmaz; Martin, Anandi

2012-05-01

In this study, we evaluated the performance of blood agar (by macroscopic growth) and nutrient agar (by a microcolony detection method) for drug susceptibility testing of Mycobacterium tuberculosis against rifampin (RIF) and isoniazid (INH), using 67 smear-positive sputum specimens. The direct proportion method on Lowenstein-Jensen (LJ) medium was used as the "gold standard." Compared with LJ medium, results for both media were in 100% agreement for RIF, while for INH the agreement levels for blood agar and nutrient agar were 98% and 95%, respectively. Within 2 weeks, 100% of specimens yielded results on blood agar, while 96.8% of specimens yielded results on nutrient agar. Our study showed that blood agar and nutrient agar can be used as alternative media for direct susceptibility testing of RIF and INH, especially in resource-poor settings.

A simple and cost effective liquid culture system for the micropropagation of two commercially important apple rootstocks.

PubMed

Mehta, Mohina; Ram, Raja; Bhattacharya, Amita

2014-07-01

The two commercially important apple rootstocks i.e., MM106 and B9 were micropropagated using a liquid culture system. Three different strengths of 0.8% agar solidified PGR free basal MS medium were first tested to optimize the culture media for both the rootstocks. Full strength medium (MS0) supported maximum in vitro growth, multiplication, rooting and survival under field conditions as opposed to quarter and half strength media. When three different volumes of liquid MS0 were tested, highest in vitro growth, multiplication, rooting and also survival under field conditions were achieved in 20 mL liquid MS0. The cost of one litre of liquid medium was also reduced by 8 times to Rs. 6.29 as compared to solid medium. The cost of 20 mL medium was further reduced to Rs. 0.125.

Simplifying Collection of Corneal Specimens in Cases of Suspected Bacterial Keratitis

PubMed Central

Kaye, Stephen B.; Rao, Prasad G.; Smith, Godfrey; Scott, John A.; Hoyles, Sharon; Morton, Clare E.; Willoughby, Colin; Batterbury, Mark; Harvey, Graham

2003-01-01

Identification of the causative organisms in suspected bacterial keratitis traditionally involves collecting multiple corneal scrapes, which are plated directly onto different solid agar culture media. Difficulties have been reported with this practice, so the development of a simpler diagnostic method in suspected bacterial keratitis would be useful. It is unclear whether a single corneal scrape sent to the microbiology laboratory in a liquid transport culture medium (indirect method) is as reliable for the diagnosis of bacterial keratitis as inoculation of multiple scrapes directly onto agar plates (direct method). To investigate this, bacterial recovery was assessed following transfer and transport of different concentrations and types of bacteria from an artificially contaminated surgical blade into brain heart infusion (BHI). Bacterial recovery rates between the proposed (indirect) and standard (direct) method were then compared after the in vitro inoculation of pig corneas and following specimen collection in patients with presumed bacterial ulcerative keratitis. Recovery of bacteria from contaminated surgical blades was found to be the same from both solid and liquid culture media. There was no significant difference in the numbers of positive cultures from solid (direct) and liquid (indirect) culture media, both in the experimental pig cornea inoculation study (P = 0.34) and in experiments with patients with clinical infections (P = 0.4), with an 85.2% agreement between methods (kappa = 0.61, P < 0.0001). In conclusion, therefore, the collection of two corneal scrapes, one used for Gram staining and the other transported in BHI followed by plating and subculturing in an enrichment medium, provides a simple method for the investigation of presumed bacterial keratitis. PMID:12843063

Growth of Desulfovibrio on the Surface of Agar Media

PubMed Central

Iverson, Warren P.

1966-01-01

Growth of Desulfovibrio desulfuricans (API strain) was found to take place in an atmosphere of hydrogen on the agar surface of complex media, including yeast extract (Difco), and Trypticase Soy Agar (BBL) without any added reducing agents. For growth on a 2% yeast extract-agar surface in the absence of hydrogen (nitrogen atmosphere), sodium lactate was required in the medium. Growth on the surface of Trypticase Soy Agar (TSA) under nitrogen took place readily in the absence of an added hydrogen donor. A medium (TSA plus salts) is described based upon the addition of sodium lactate (4 ml per liter), magnesium sulfate (2 g per liter), and ferrous ammonium sulfate (0.05%) to TSA, which appears suitable for the isolation and growth of Desulfovibrio on the surface of agar plates in an atmosphere of hydrogen. Sodium lactate does not appear to be essential in this medium for good growth and sulfate reduction in a hydrogen atmosphere, but is essential in a nitrogen atmosphere. Growth of Desulfovibrio (hydrogen atmosphere) on the agar surface of media commonly used for its cultivation as well as on an inorganic medium containing bicarbonate as a source of carbon is poor and erratic unless inoculated (Desulfovibrio) plates of TSA plus salts are incubated in the same container with plates of these media. This stimulatory effect of incubation with inoculated plates of TSA plus salts medium appears to be due to as yet unidentified volatile material produced by D. desulfuricans when growing on this medium. Another volatile material, or possibly the identical material, appears to act similarly to a hydrogen donor. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 PMID:5955798

Application of agar liquid-gel transition in cultivation and harvesting of microalgae for biodiesel production.

PubMed

Kumar, Vinod; Nanda, Manisha; Verma, Monu

2017-11-01

In order to increase microalgal biomass productivity efficient cultivation and harvesting methods are needed against the available traditional methods. The present study focuses on the same by harvesting microalgae using agar gel. Agar medium containing bold's basal medium (BBM) undergoes a thermoreversible gel transition. As compared to the traditional protocols, this gel is used to cultivate microalgae without even affecting the total productivity. To develop the gel for microalgae cultivation, agar was boiled in BBM. Then the agar was cooled to 35°C and microalgae culture was added to it. After seeding the microalgae the temperature of the agar was further decreased by 10°C to induce gelation. Instead of isolated cells microalgae were grown in clusters within the agar gel. Microalgal clusters gravimetrically settle at the bottom within 2h. In this method agar can be reused. Copyright © 2017 Elsevier Ltd. All rights reserved.

Diagnostic value of morphological, physiological and biochemical tests in distinguishing Trichophyton rubrum from Trichophyton mentagrophytes complex.

PubMed

Ates, Aylin; Ozcan, Kadri; Ilkit, Macit

2008-12-01

The two most frequently encountered dermatophyte etiologic agents of glabrous skin and nail dermatophytoses are Trichophyton rubrum and T. mentagrophytes. This study was aimed to discuss the efficacy of morphological, physiological and biochemical diagnostic tests commonly used in the identification of T. rubrum and members of the T. mentagrophytes complex. In this study, we evaluated; hydrolysis of urea in broth and on urea agar slants and Petri plates incubated at 22 degrees C, 28 degrees C and 37 degrees C, in vitro hair perforation (blond child, sheep and goat hair), pigment production on cornmeal dextrose agar (CMDA) and bromcresol purple-milk solids-glucose agar (BCP-MS-G), Tween opacity, sorbitol assimilation, and salt tolerance. Additionally, the production of micro- and macroconidia was investigated by using brain heart infusion agar (BHIA), Christensen's urea agar in Petri plates (UPA), CMDA, Lowenstein-Jensen agar (LJA), malt extract agar, oatmeal agar, Oxoid chromogenic Candida agar, and potato dextrose agar. All cultures were incubated at 28 degrees C, and conidial production was compared on days 5, 10 and 15. It was found that the urea hydrolysis test yielded more rapid and significant results when urea medium was prepared in Petri plates and incubated at 28 degrees C (P<0.01). LJA supported the highest production of microconidia after 15 days (P<0.001). Additionally, it was found that T. rubrum strains produced red pigment on CMDA (P<0.01) and BCP-MS-G, while strains of the T. mentagrophytes species complex did not. A special algorithm containing the various test procedures employed in these studies is presented which was found to be useful in the differentiation of T. rubrum strains from T. mentagrophytes complex. Our results revealed that UPA, CMDA, BCP-MS-G, LJA, and BHIA may be used as common mycological agars in routine practice.

Rapid identification of microorganisms from positive blood cultures by MALDI-TOF mass spectrometry subsequent to very short-term incubation on solid medium.

PubMed

Idelevich, E A; Schüle, I; Grünastel, B; Wüllenweber, J; Peters, G; Becker, K

2014-10-01

Rapid identification of the causative microorganism is important for appropriate antimicrobial therapy of bloodstream infections. Bacteria from positive blood culture (BC) bottles are not readily available for identification by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Lysis and centrifugation procedures suggested for direct MALDI-TOF MS from positive BCs without previous culture are associated with additional hands-on processing time and costs. Here, we describe an alternative approach applying MALDI-TOF MS from bacterial cultures incubated very briefly on solid medium. After plating of positive BC broth on Columbia blood agar (n = 165), MALDI-TOF MS was performed after 1.5, 2, 3, 4, 5, 6, 7, 8, 12 and (for control) 24 h of incubation until reliable identification to the species level was achieved (score ≥2.0). Mean incubation time needed to achieve species-level identification was 5.9 and 2.0 h for Gram-positive aerobic cocci (GPC, n = 86) and Gram-negative aerobic rods (GNR, n = 42), respectively. Short agar cultures with incubation times ≤2, ≤4, ≤6, ≤8 and ≤12 h yielded species identification in 1.2%, 18.6%, 64.0%, 96.5%, 98.8% of GPC, and in 76.2%, 95.2%, 97.6%, 97.6%, 97.6% of GNR, respectively. Control species identification at 24 h was achieved in 100% of GPC and 97.6% of GNR. Ethanol/formic acid protein extraction performed for an additional 34 GPC isolates cultivated from positive BCs showed further reduction in time to species identification (3.1 h). MALDI-TOF MS using biomass subsequent to very short-term incubation on solid medium allows very early and reliable bacterial identification from positive BCs without additional time and cost expenditure. © 2014 The Authors Clinical Microbiology and Infection © 2014 European Society of Clinical Microbiology and Infectious Diseases.

Broth versus solid agar culture of swab samples of cadaveric allograft musculoskeletal tissue.

PubMed

Varettas, Kerry

2013-12-01

As part of the donor assessment protocol, bioburden assessment must be performed on allograft musculoskeletal tissue samples collected at the time of tissue retrieval. Swab samples of musculoskeletal tissue allografts from cadaveric donors are received at the microbiology department of the South Eastern Area Laboratory Services (Australia) to determine the presence of bacteria and fungi. This study will review the isolation rate of organisms from solid agar and broth culture of swab samples of cadaveric allograft musculoskeletal tissue over a 6-year period, 2006-2011. Swabs were inoculated onto horse blood agar (anaerobic, 35 °C) and chocolate agar (CO2, 35 °C) and then placed into a cooked meat broth (aerobic, 35 °C). A total of 1,912 swabs from 389 donors were received during the study period. 557 (29.1 %) swabs were culture positive with the isolation of 713 organisms, 249 (34.9 %) from solid agar culture and an additional 464 (65.1 %) from broth culture only. This study has shown that the broth culture of cadaveric allograft musculoskeletal swab samples recovered a greater amount of organisms than solid agar culture. Isolates such as Clostridium species and Staphylococcus aureus would not have been isolated from solid agar culture alone. Broth culture is an essential part of the bioburden assessment protocol of swab samples of cadaveric allograft musculoskeletal tissue in this laboratory.

Comparison of four chromogenic media and Hektoen agar for detection and presumptive identification of Salmonella strains in human stools.

PubMed

Perez, J M; Cavalli, P; Roure, C; Renac, R; Gille, Y; Freydiere, A M

2003-03-01

Several chromogenic media have been developed to enhance the specificity of Salmonella detection. We compared the performance of four commercial chromogenic media-namely, ABC medium (Lab M. Ltd., Bury, United Kingdom), COMPASS Salmonella agar (Biokar Diagnostics, Beauvais, France), CHROMagar Salmonella agar (CHROMagar Company, Paris, France), and SM ID agar (bioMerieux, Marcy l'Etoile, France)-with conventional Hektoen medium. Nine hundred sixteen stool samples from inpatients at three hospitals were cultured, in parallel, on the five media, both by direct inoculation and after selective enrichment in selenite broth. Sixty-four Salmonella strains with 12 serotypes were isolated on at least one medium. After 48 h of incubation, sensitivity before and after enrichment was 62.5 and 89.1% with ABC medium, 77.1 and 93.8% with COMPASS agar, 66.7 and 89.1% with CHROMagar, 68.8 and 85.9% with SM ID agar, and 85.4 and 98.4% with Hektoen agar, respectively. Broth enrichment and prolonged incubation (48 versus 24 h) increased the sensitivity of all five media. Only one strain was not isolated on Hektoen agar. The number of false-positive isolates was higher with all five media after enrichment in selenite broth and after incubation for 48 h compared to 24 h. The specificity of the four chromogenic media was better than 91% after incubation for 24 h (77.7% with Hektoen agar) and better than 84% after incubation for 48 h (74.8% with Hektoen agar). This higher specificity reduces the need for confirmatory tests, thereby cutting technical time and reagent requirements. Both COMPASS agar and CHROMagar Salmonella, which after simple additional tests showed close efficiencies (96 and 97%, respectively), can be recommended as single-plate media of choice for the detection and presumptive identification of salmonellae in stools.

Development of novel Alicyclobacillus spp. isolation medium.

PubMed

Chang, S; Kang, D-H

2005-01-01

To develop a new isolation medium with higher recovery rates of Alicyclobacillus spp. SK agar was developed with optimized incubation temperature, pH, acidulant, Tween 80 concentration and divalent cation addition. Results indicate that detection of Alicyclobacillus spp. by SK agar was significantly higher (P > 0.05) than those obtained by K agar, orange serum agar, and potato dextrose agar. Current media used for Alicyclobacillus spp. isolation still resulted in high numbers of false negative products. The sensitivity of SK agar to Alicyclobacillus spp. allows detection of low numbers of Alicyclobacillus spp. and also provides a more higher isolation results compared with currently used media. SK agar will be useful to the fruit juice industry to obtain more accurate numbers of contaminant Alicyclobacillus spp. With this media, false negative samples can be reduced, and the likelihood of exported products being rejected can be greatly reduced.

A Novel Chromogenic Ester Agar Medium for Detection of Salmonellae

PubMed Central

Cooke, Venitia M.; Miles, R. J.; Price, R. G.; Richardson, A. C.

1999-01-01

A novel agar medium, chromogenic Salmonella esterase (CSE) agar, for the differentiation of salmonellae is described. The agar contains peptones and nutrient extracts together with the following (grams per liter unless otherwise specified): 4-[2-(4-octanoyloxy-3,5-dimethoxyphenyl)-vinyl]-quinolinium-1-(propan-3-yl carboxylic acid) bromide (SLPA-octanoate; bromide form), 0.3223; lactose, 14.65; trisodium citrate dihydrate, 0.5; Tween 20, 3.0; ethyl 4-dimethylaminobenzoate, 0.035% (wt/vol), novobiocin, 70 mg liter−1. The key component of the medium is SLPA-octanoate, a newly synthesized ester formed from a C8 fatty acid and a phenolic chromophore. In CSE agar, the ester is hydrolyzed by Salmonella spp. to yield a brightly colored phenol which remains tightly bound within colonies. After 24 h of incubation at 37 or 42°C, colonies of typical Salmonella spp. were burgundy colored on a transparent yellow background, whereas non-Salmonella spp. were white, cream, yellow or transparent. CSE agar was evaluated by using a panel of strains including a high proportion of Salmonella and non-Salmonella strains giving atypical reactions on other differential agars. The sensitivity (93.1%) of CSE agar for non-typhi salmonellae compared favorably with those of Rambach (82.8%), xylose-lysine-deoxycholate (XLD; 91.4%), Hektoen-enteric (89.7%), and SM ID (91.4%) agars. The specificity (93.9%) was also comparable to those of other Salmonella media (SM ID agar, 95.9%; Rambach agar, 91.8%; XLD agar, 91.8%; Hektoen-enteric agar, 87.8%). Strains of Citrobacter freundii and Proteus spp. giving false-positive reactions with other media gave a negative color reaction on CSE agar. CSE agar enabled the detection of >30 Salmonella serotypes, including agona, anatum, enteritidis, hadar, heidelberg, infantis, montevideo, thompson, typhimurium, and virchow, which accounted for 91.8% of the salmonella isolates recorded by the Public Health Laboratory Service (Colindale, London, England) for 1997. PMID:9925620

Prospective Comparison of a New Chromogenic Medium, MRSASelect, to CHROMagar MRSA and Mannitol-Salt Medium Supplemented with Oxacillin or Cefoxitin for Detection of Methicillin-Resistant Staphylococcus aureus

PubMed Central

Stoakes, Luba; Reyes, Romina; Daniel, Janis; Lennox, Gwen; John, Michael A.; Lannigan, Robert; Hussain, Zafar

2006-01-01

MRSASelect agar was compared to CHROMagar, mannitol-salt agar with oxacillin, and mannitol-salt agar with cefoxitin (MSA-CFOX) for the isolation of methicillin-resistant Staphylococcus aureus (MRSA). The sensitivities and specificities were 97.3% and 99.8%, 82.9% and 99.1%, 80.2% and 79%, and 99.1% and 84.8%, respectively. MSA-CFOX and MRSASelect had a high sensitivity. MRSASelect, however, was more specific and proved to be a more reliable and rapid medium for the detection of MRSA. PMID:16455933

What is the most reliable solid culture medium for tuberculosis treatment trials?

PubMed

Joloba, Moses L; Johnson, John L; Feng, Pei-Jean I; Bozeman, Lorna; Goldberg, Stefan V; Morgan, Karen; Gitta, Phineas; Boom, Henry W; Heilig, Charles M; Mayanja-Kizza, Harriet; Eisenach, Kathleen D

2014-05-01

We conducted a prospective study to determine which solid medium is the most reliable overall and after two months of therapy to detect Mycobacterium tuberculosis complex (MTB). MTB isolation and contamination rates on LJ and Middlebrook 7H10 and 7H11 agar with and without selective antibiotics were examined in a single laboratory and compared against a constructed reference standard and MGIT 960 results. Of 50 smear positive adults with pulmonary TB enrolled, 45 successfully completed standard treatment. Two spot sputum specimens were collected before treatment and at week 8 and one spot specimen each at weeks 2, 4, 6, and 12. The MTB recovery rate among all solid media for pre-treatment specimens was similar. After 8 weeks, selective (S) 7H11 had the highest positivity rate. Latent class analysis was used to construct the primary reference standard. The 98.7% sensitivity of 7H11S (95% Wilson confidence interval 96.4%-99.6%) was highest among the 5 solid media (P = 0.003 by bootstrap); the 82.6% specificity of 7H10S (95% CI 75.7%-87.8%) was highest (P = 0.098). Our results support 7H11S as the medium of choice. Further studies in different areas where recovery and contamination are likely to vary, are recommended. Copyright © 2014 Elsevier Ltd. All rights reserved.

Improving agar electrospinnability with choline-based deep eutectic solvents.

PubMed

Sousa, Ana M M; Souza, Hiléia K S; Uknalis, Joseph; Liu, Shih-Chuan; Gonçalves, Maria P; Liu, LinShu

2015-09-01

Very recently our group has produced novel agar-based fibers by an electrospinning technique using water as solvent and polyvinyl alcohol (PVA) as co-blending polymer. Here, we tested the deep eutectic solvent (DES), (2-hydroxyethyl)trimethylammonium chloride/urea prepared at 1:2 molar ratio, as an alternative solvent medium for agar electrospinning. The electrospun materials were collected with an ethanol bath adapted to a previous electrospinning set-up. One weight percent agar-in-DES showed improved viscoelasticity and hence, spinnability, when compared to 1 wt% agar-in-water and pure agar nanofibers were successfully electrospun if working above the temperature of sol-gel transition (∼80 °C). By changing the solvent medium we decreased the PVA concentration (5 wt% starting solution) and successfully produced composite fibers with high agar contents (50/50 agar/PVA). Best composite fibers were formed with the 50/50 and 30/70 agar/PVA solutions. These fibers were mechanically resistant, showed tailorable surface roughness and diverse size distributions, with most of the diameters falling in the sub-micron range. Both nano and micro forms of agar fibers (used separately or combined) may have potential for the design of new and highly functional agar-based materials. Published by Elsevier B.V.

Phylogenetic diversity of Flavobacteria isolated from the North Sea on solid media.

PubMed

Hahnke, Richard L; Harder, Jens

2013-10-01

Flavobacteria are abundant in the North Sea, an epeiric sea on the continental shelf of Europe. However, this abundance has so far not been reflected by the number of strains in culture collections. In this study, Flavobacteria were isolated from pelagic and benthic samples, such as seawater, phytoplankton, sediment and its porewater, and from surfaces of animals and seaweeds on agar plates with a variety of carbon sources. Dilution cultivation with a new medium, incubation at low temperatures and with long incubation times, and colony screening by a Flavobacteria-Cytophagia-specific PCR detecting 16S rRNA gene sequences led to a collection of phylogenetically diverse strains. Two strains affiliated with Flammeovirgaceae and seven strains affiliated with Cyclobacteriaceae, whereas within the Flavobacteriaceae 20 isolated strains presumably represented seven novel candidate genera and 355 strains affiliated with 26 of 80 validly described marine Flavobacteriaceae genera, based on a genus boundary of 95.0% 16S rRNA gene sequence identity. The majority of strains (276) affiliated with 37 known species in 16 genera (based on a boundary of 98.7% 16S rRNA gene sequence identity), whereas 79 strains likely represented 42 novel species in 22 established Flavobacteriaceae genera. Pigmentation, iridescence, gliding motility, agar lysis, and flexirubin as a chemical marker supported the taxonomy at the species level. This study demonstrated the culturability on solid medium of phylogenetically diverse Flavobacteria originating from the North Sea. Copyright © 2013 Elsevier GmbH. All rights reserved.

Three-dimensional characterization of bacterial microcolonies on solid agar-based culture media.

PubMed

Drazek, Laurent; Tournoud, Maud; Derepas, Frédéric; Guicherd, Maryse; Mahé, Pierre; Pinston, Frédéric; Veyrieras, Jean-Baptiste; Chatellier, Sonia

2015-02-01

For the last century, in vitro diagnostic process in microbiology has mainly relied on the growth of bacteria on the surface of a solid agar medium. Nevertheless, few studies focused in the past on the dynamics of microcolonies growth on agar surface before 8 to 10h of incubation. In this article, chromatic confocal microscopy has been applied to characterize the early development of a bacterial colony. This technology relies on a differential focusing depth of the white light. It allows one to fully measure the tridimensional shape of microcolonies more quickly than classical confocal microscopy but with the same spatial resolution. Placing the device in an incubator, the method was able to individually track colonies growing on an agar plate, and to follow the evolution of their surface or volume. Using an appropriate statistical modeling framework, for a given microorganism, the doubling time has been estimated for each individual colony, as well as its variability between colonies, both within and between agar plates. A proof of concept led on four bacterial strains of four distinct species demonstrated the feasibility and the interest of the approach. It showed in particular that doubling times derived from early tri-dimensional measurements on microcolonies differed from classical measurements in micro-dilutions based on optical diffusion. Such a precise characterization of the tri-dimensional shape of microcolonies in their late-lag to early-exponential phase could be beneficial in terms of in vitro diagnostics. Indeed, real-time monitoring of the biomass available in a colony could allow to run well established microbial identification workflows like, for instance, MALDI-TOF mass-spectrometry, as soon as a sufficient quantity of material is available, thereby reducing the time needed to provide a diagnostic. Moreover, as done for pre-identification of macro-colonies, morphological indicators such as three-dimensional growth profiles derived from microcolonies could be used to perform a first pre-identification step, but in a shorten time. Copyright © 2014 Elsevier B.V. All rights reserved.

Differential roles of auxin efflux carrier PIN proteins in hypocotyl phototropism of etiolated Arabidopsis seedlings depend on the direction of light stimulus.

PubMed

Haga, Ken; Sakai, Tatsuya

2013-01-01

In a recent study, we demonstrated that although the auxin efflux carrier PIN-FORMED (PIN) proteins, such as PIN3 and PIN7, are required for the pulse-induced first positive phototropism in etiolated Arabidopsis hypocotyls, they are not necessary for the continuous-light-induced second positive phototropism when the seedlings are grown on the surface of agar medium, which causes the hypocotyls to separate from the agar surface. Previous reports have shown that hypocotyl phototropism is slightly impaired in pin3 single mutants when they are grown along the surface of agar medium, where the hypocotyls always contact the agar, producing some friction. To clarify the possible involvement of PIN3 and PIN7 in continuous-light-induced phototropism, we investigated hypocotyl phototropism in the pin3 pin7 double mutant grown along the surface of agar medium. Intriguingly, the phototropic curvature was slightly impaired in the double mutant when the phototropic stimulus was presented on the adaxial side of the hook, but was not impaired when the phototropic stimulus was presented on the abaxial side of the hook. These results indicate that PIN proteins are required for continuous-light-induced second positive phototropism, depending on the direction of the light stimulus, when the seedlings are in contact with agar medium.

Differential roles of auxin efflux carrier PIN proteins in hypocotyl phototropism of etiolated Arabidopsis seedlings depend on the direction of light stimulus

PubMed Central

Haga, Ken; Sakai, Tatsuya

2013-01-01

In a recent study, we demonstrated that although the auxin efflux carrier PIN-FORMED (PIN) proteins, such as PIN3 and PIN7, are required for the pulse-induced first positive phototropism in etiolated Arabidopsis hypocotyls, they are not necessary for the continuous-light-induced second positive phototropism when the seedlings are grown on the surface of agar medium, which causes the hypocotyls to separate from the agar surface. Previous reports have shown that hypocotyl phototropism is slightly impaired in pin3 single mutants when they are grown along the surface of agar medium, where the hypocotyls always contact the agar, producing some friction. To clarify the possible involvement of PIN3 and PIN7 in continuous-light-induced phototropism, we investigated hypocotyl phototropism in the pin3 pin7 double mutant grown along the surface of agar medium. Intriguingly, the phototropic curvature was slightly impaired in the double mutant when the phototropic stimulus was presented on the adaxial side of the hook, but was not impaired when the phototropic stimulus was presented on the abaxial side of the hook. These results indicate that PIN proteins are required for continuous-light-induced second positive phototropism, depending on the direction of the light stimulus, when the seedlings are in contact with agar medium. PMID:23104115

Measurement of Shear Elastic Moduli in Quasi-Incompressible Soft Solids

NASA Astrophysics Data System (ADS)

Rénier, Mathieu; Gennisson, Jean-Luc; Barrière, Christophe; Catheline, Stefan; Tanter, Mickaël; Royer, Daniel; Fink, Mathias

2008-06-01

Recently a nonlinear equation describing the plane shear wave propagation in isotropic quasi-incompressible media has been developed using a new expression of the strain energy density, as a function of the second, third and fourth order shear elastic constants (respectively μ, A, D) [1]. In such a case, the shear nonlinearity parameter βs depends only from these last coefficients. To date, no measurement of the parameter D have been carried out in soft solids. Using a set of two experiments, acoustoelasticity and finite amplitude shear waves, the shear elastic moduli up to the fourth order of soft solids are measured. Firstly, this theoretical background is applied to the acoustoelasticity theory, giving the variations of the shear wave speed as a function of the stress applied to the medium. From such variations, both linear (μ) and third order shear modulus (A) are deduced in agar-gelatin phantoms. Experimentally the radiation force induced by a focused ultrasound beam is used to generate quasi-plane linear shear waves within the medium. Then the shear wave propagation is imaged with an ultrafast ultrasound scanner. Secondly, in order to give rise to finite amplitude plane shear waves, the radiation force generation technique is replaced by a vibrating plate applied at the surface of the phantoms. The propagation is also imaged using the same ultrafast scanner. From the assessment of the third harmonic amplitude, the nonlinearity parameter βS is deduced. Finally, combining these results with the acoustoelasticity experiment, the fourth order modulus (D) is deduced. This set of experiments provides the characterization, up to the fourth order, of the nonlinear shear elastic moduli in quasi-incompressible soft media. Measurements of the A moduli reveal that while the behaviors of both soft solids are close from a linear point of view, the corresponding nonlinear moduli A are quite different. In a 5% agar-gelatin phantom, the fourth order elastic constant D is found to be 30±10 kPa.

Role of fimV in type II secretion system-dependent protein secretion of Pseudomonas aeruginosa on solid medium.

PubMed

Michel, Gérard P F; Aguzzi, Anthony; Ball, Geneviève; Soscia, Chantal; Bleves, Sophie; Voulhoux, Romé

2011-07-01

Although classical type II secretion systems (T2SSs) are widely present in Gram-negative bacteria, atypical T2SSs can be found in some species. In Pseudomonas aeruginosa, in addition to the classical T2SS Xcp, it was reported that two genes, xphA and xqhA, located outside the xcp locus were organized in an operon (PaQa) which encodes the orphan PaQa subunit. This subunit is able to associate with other components of the classical Xcp machinery to form a functional hybrid T2SS. In the present study, using a transcriptional lacZ fusion, we found that the PaQa operon was more efficiently expressed (i) on solid LB agar than in liquid LB medium, (ii) at 25 °C than at 37 °C and (iii) at an early stage of growth. These results suggested an adaptation of the hybrid system to particular environmental conditions. Transposon mutagenesis led to the finding that vfr and fimV genes are required for optimal expression of the orphan PaQa operon in the defined growth conditions used. Using an original culturing device designed to monitor secretion on solid medium, the ring-plate system, we found that T2SS-dependent secretion of exoproteins, namely the elastase LasB, was affected in a fimV deletion mutant. Our findings led to the discovery of an interplay between FimV and the global regulator Vfr triggering the modulation of the level of Vfr and consequently the modulation of T2SS-dependent secretion on solid medium.

Malt-yeast extract-sucrose agar, a suitable medium for enumeration and isolation of fungi from silage.

PubMed Central

Skaar, I; Stenwig, H

1996-01-01

A general medium named malt-yeast extract-sucrose agar (MYSA) containing oxgall was designed. The medium was intended for the enumeration and isolation of molds and yeasts in routine examinations of animal feed stuffs. In this study MYSA was tested as a general medium for mycological examination of silage. The medium was compared with dichloran-rose bengal medium (DRBC) in an examination of more than 500 specimens of big bale grass silage. Selected characteristics of known fungal species commonly isolated from feeds were examined after growth on MYSA and DRBC and on malt extract agar, used as a noninhibitory control medium. MYSA suppressed bacterial growth, without affecting the growth of fungi common in feeds. The fungi growing on MYSA were easily recognized, and the medium seemed to slow radial growth of fungal colonies, which permitted, easy counting. The number of species found was higher on MYSA than on DRBC. When we compared MYSA with DRBC for mycological examination of grass silage samples, MYSA was found to be the medium of choice. PMID:8837416

Culture Phenotypes of Genomically and Geographically Diverse Mycobacterium avium subsp. paratuberculosis Isolates from Different Hosts▿

PubMed Central

Whittington, Richard J.; Marsh, Ian B.; Saunders, Vanessa; Grant, Irene R.; Juste, Ramon; Sevilla, Iker A.; Manning, Elizabeth J. B.; Whitlock, Robert H.

2011-01-01

Mycobacterium avium subsp. paratuberculosis causes paratuberculosis (Johne's disease) in ruminants in most countries. Historical data suggest substantial differences in culturability of M. avium subsp. paratuberculosis isolates from small ruminants and cattle; however, a systematic comparison of culture media and isolates from different countries and hosts has not been undertaken. Here, 35 field isolates from the United States, Spain, Northern Ireland, and Australia were propagated in Bactec 12B medium and Middlebrook 7H10 agar, genomically characterized, and subcultured to Lowenstein-Jensen (LJ), Herrold's egg yolk (HEY), modified Middlebrook 7H10, Middlebrook 7H11, and Watson-Reid (WR) agars, all with and without mycobactin J and some with sodium pyruvate. Fourteen genotypes of M. avium subsp. paratuberculosis were represented as determined by BstEII IS900 and IS1311 restriction fragment length polymorphism analysis. There was no correlation between genotype and overall culturability, although most S strains tended to grow poorly on HEY agar. Pyruvate was inhibitory to some isolates. All strains grew on modified Middlebrook 7H10 agar but more slowly and less prolifically on LJ agar. Mycobactin J was required for growth on all media except 7H11 agar, but growth was improved by the addition of mycobactin J to 7H11 agar. WR agar supported the growth of few isolates. The differences in growth of M. avium subsp. paratuberculosis that have historically been reported in diverse settings have been strongly influenced by the type of culture medium used. When an optimal culture medium, such as modified Middlebrook 7H10 agar, is used, very little difference between the growth phenotypes of diverse strains of M. avium subsp. paratuberculosis was observed. This optimal medium is recommended to remove bias in the isolation and cultivation of M. avium subsp. paratuberculosis. PMID:21430104

Vaginal lactobacilli inhibiting growth of Gardnerella vaginalis, Mobiluncus and other bacterial species cultured from vaginal content of women with bacterial vaginosis.

PubMed

Skarin, A; Sylwan, J

1986-12-01

On a solid agar medium the growth-inhibitory effect of 9 Lactobacillus strains cultured from vaginal content was tested on bacteria cultured from vaginal content of women with bacterial vaginosis: Mobiluncus, Gardnerella vaginalis, Bacteroides and anaerobic cocci. Inhibition zones were observed in the growth of all of the strains isolated from women with bacterial vaginosis around all lactobacilli. The inhibitory effect of the lactobacilli was further tested on various anaerobic and facultatively anaerobic species, both type strains and fresh extragenitally cultured strains. Four Bacteroides fragilis strains as well as 2 out of 4 Staphylococcus aureus strains were clearly inhibited by the lactobacilli. The inhibition zones were generally wider at pH 5.5 than at 6.0. For all inhibited strains, (the S. aureus excepted) a low pH on the agar around the lactobacilli correlated to wider growth-inhibition zones.

Media for the aerobic growth of campylobacter

USDA-ARS?s Scientific Manuscript database

The effect of agar and sodium bicarbonate (NaHCO3) concentration on aerobic growth of Campylobacter in a fumarate-pyruvate medium was examined. The broth medium was supplemented with 0.0 to 0.2% agar and inoculated with 106 CFU/ml of Campylobacter coli 33559, Campylobacter fetus 27349, Campylobacter...

The upper surface of an Escherichia coli swarm is stationary.

PubMed

Zhang, Rongjing; Turner, Linda; Berg, Howard C

2010-01-05

When grown in a rich medium on agar, many bacteria elongate, produce more flagella, and swim in a thin film of fluid over the agar surface in swirling packs. Cells that spread in this way are said to swarm. The agar is a solid gel, with pores smaller than the bacteria, so the swarm/agar interface is fixed. Here we show, in experiments with Escherichia coli, that the swarm/air interface also is fixed. We deposited MgO smoke particles on the top surface of an E. coli swarm near its advancing edge, where cells move in a single layer, and then followed the motion of the particles by dark-field microscopy and the motion of the underlying cells by phase-contrast microscopy. Remarkably, the smoke particles remained fixed (diffusing only a few micrometers) while the swarming cells streamed past underneath. The diffusion coefficients of the smoke particles were smaller over the virgin agar ahead of the swarm than over the swarm itself. Changes between these two modes of behavior were evident within 10-20 microm of the swarm edge, indicating an increase in depth of the fluid in advance of the swarm. The only plausible way that the swarm/air interface can be fixed is that it is covered by a surfactant monolayer pinned at its edges. When a swarm is exposed to air, such a monolayer can markedly reduce water loss. When cells invade tissue, the ability to move rapidly between closely opposed fixed surfaces is a useful trait.

New medium for rapid screening and enumeration of Clostridium perfringens in foods.

PubMed Central

Erickson, J E; Deibel, R H

1978-01-01

A rapid and sensitive procedure for estimating low numbers of Clostridium perfringens has been investigated and compared to methods used currently in the food industry. The new liquid medium, RPM (rapid perfringens medium), was compared with sulfite-polymyxin-sulfadiazine agar and tryptose-sulfite-cycloserine agar in recovery studies with naturally contaminated and with inoculated foods. The medium consists of a mixture of litmus milk and fluid thioglycolate medium fortified with glucose, peptone, gelatin, yeast extract, sodium chloride, and ferrous sulfate. Selectivity is based on an antibiotic system (polymyxin B sulfate and neomycin sulfate) incorporated into the medium, coupled with an incubation temprature of 46 to 48 degrees C for 24 h. Tubes were scored as positive if a stormy fermentation was observed. All tubes demonstrating stormy fermentation were confirmed as containing C. perfringens. Of a total of 774 naturally contaminated food samples, 546 samples (71%) were found to contain C. perfringens with RPM, whereas only 168 (22%) of the samples were positive using sulfite-polymyxin-sulfadiazine agar. C. perfringens was isolated from 71% of 85 other samples using RPM as compared to 14% with tryptose-sulfite-cycloserine agar. Enumeration studies on 14 individual samples using the most probable number technique also demonstrated greater sensitivity with RPM. PMID:213019

Study of the lactic acid bacteria throughout the manufacture of dry-cured lacón (a Spanish traditional meat product). Effect of some additives.

PubMed

Lorenzo, José M; García Fontán, María C; Cachaldora, Aida; Franco, Inmaculada; Carballo, Javier

2010-04-01

Total aerobic mesophilic microflora (on SPC agar), lactic acid bacteria (on MRS agar) and lactobacilli (on Rogosa agar) were enumerated in samples from the surface and the interior of the pieces throughout the manufacture of six batches of lacón. Three of the batches were made without additives and three with additives (glucose (2 g/kg), sodium nitrite (E(250)) (125 mg/kg), sodium nitrate (E(251)) (175 mg/kg), sodium ascorbate (E(301)) (500 mg/kg), and sodium citrate (E(331)) (100 mg/kg)). The counts decreased throughout the manufacturing process, particularly after the salting stage. The use of additives did not affect the counts or the evolution of the microbial groups, except for the lactobacilli, which were present in higher numbers in the batches with additives. In four batches (two without and two with additives), from MRS agar and from Rogosa agar plates, 10 colonies were randomly taken from each sampling point of each batch (five from the surface sample and five from the interior sample) and from each culture medium; a total of 224 strains from MRS agar, and 176 strains from Rogosa agar that were identified by classical methods. The MRS agar displayed moderate selectivity for the isolation of lactic acid bacteria, and only 59% of the isolated strains belonged to this microbial group. Homofermentative and facultative heterofermentative lactobacilli (particularly Lactobacillus curvatus and Lactobacillus sakei) were the most abundant species isolated on this medium. The selectivity of the Rogosa agar for lactobacilli was extremely high. The species of lactobacilli isolated on this medium at different stages of manufacture of the four batches of lacón were consistent with those isolated from MRS agar. The use of additives in the lacón did not appreciably affect the kinds and proportions of species isolated on either MRS agar or Rogosa agar.

Highly selective medium for isolation of Listeria monocytogenes from food.

PubMed Central

al-Zoreky, N; Sandine, W E

1990-01-01

A new selective medium (Al-Zoreky-Sandine listeria medium [ASLM]) was formulated to recover Listeria monocytogenes from food specimens; the medium completely inhibited common food microflora. Recognition of Listeria colonies is evident by black discoloration of the medium due to esculin hydrolysis without need for special illuminating equipment. The medium contains acriflavin, ceftazidime, and moxalactam as selective agents. Compared with Listeria Selective Agar, ASLM was equally effective in recovering L. monocytogenes. However, ASLM inhibited micrococci, enterococci, and gram-negative bacteria, especially a strain that mimicked L. monocytogenes on Listeria Selective Agar. The new medium was able to recover heat injured cells with only 15% less count than the nonselective medium. Images PMID:2126701

Evaluation of the oxolinic acid--esculin--azide medium for the isolation and enumeration of faecal streptococci in a routine monitoring programme for bathing waters.

PubMed

Figueras, M J; Inza, I; Polo, F; Guarro, J

1998-10-01

m-Enterococcus agar (m-Ent) has been generally considered the reference medium for faecal streptococci in bathing waters. However, it shows several shortcomings, and therefore it is important to test newly developed media that can guarantee more precise results. In this sense, the recently described oxolinic acid--esculin--azide agar medium (OAA) and m-enterococcus agar (m-Ent) were comparatively evaluated for the detection of faecal streptococci from seawater and fresh water. The OAA medium showed a significantly higher relative recovery percentage and specificity for both types of water than m-Ent. A similar spectrum of species was recorded from both media, Enterococcus faecium being predominant in fresh water and Enterococcus faecalis, in seawater. The superior performance of the OAA medium in both types of bathing waters, added to the fact that it does not require the use of complementary confirmative tests, makes this medium an excellent candidate to be employed for monitoring programmes.

Isolation of Strain MLTeJB From Mono Lake, California, a Dissimilatory Tellurite Respiring Prokaryote.

NASA Astrophysics Data System (ADS)

Baesman, S. M.; Oremland, R. S.

2007-12-01

Previous investigations on the dissimilatory reduction of Te-oxyanions have been constrained by the inhibtory effects of circa 1.0 mM concentrations of either Te(IV) or Te(VI) upon growth of established cultures. Therefore we initiated new enrichments using anoxic Mono Lake mud supplemented with 10 mM Te(IV) as the electron acceptor and lactate as the electron donor. Sediments turned black with time owing to the formation of Te(0), microscopic examination of which confirmed the presence of both shards, rosettes, and nanospheres of Te(0). The enrichment was subcultured several times in liquid medium and then streaked onto solid medium and incubated in an anaerobic chamber. Isolated black colonies were re-streaked several times, and thence inoculated into liquid medium. However, growth in liquid medium required the presence of a small amount of solid phase, which included a plug of either agar, phytagel, or glass beads. Growth resulted in oxidation of lactate to acetate, formate and CO2 with the reduction of Te(IV) to Te(0). The isolate, strain MLTeJB was a non-motile rod that stained Gram positive, and formed copious exogenous deposits of Te(0) nano-shards and rosettes. Further details on the physiology of this organism will be presented.

Use of bile-esculin agar for rapid differentiation of Enterobacteriaceae.

PubMed Central

Lindell, S S; Quinn, P

1975-01-01

Bile-esculin agar has been used for several years for the presumptive identification of group D streptococci. All members of the Enterobacteriaceae family will also grow on this medium, but only certain ones can hydrolyze esculin to 6,7-dihydroxycoumarin, which reacts with iron to produce a characteristic blackening of the medium. One thousand and six cultures from clinical specimens representing 20 genera were isolated and identified. Heavy inocula from fresh pure culture isolates on heart infusion agar were placed on bile-esculin agar slants and incubated at 35 C. The slants were examined at 4 h and again at 18 h for esculin hydrolysis. Shigella, Salmonella, Arizona, Proteus mirabilis, Proteus morganii, Providencia alcalifaciens, and Providencia stuartii all produced negative results. Klebsiella pneumoniae, Enterobacter aerogenes, Serratia marcescens, and Serratia rubidaea produced a positive reaction in 4 h. The other remaining eight genera exhibited varying results. The use of this medium in conjunction with triple sugar iron-lysine iron agar has been of great value in differentiating the Klebsiella-Enterobacter-Serratia group from other Enterobacteriaceae. PMID:1176613

Phosphate-Catalyzed Hydrogen Peroxide Formation from Agar, Gellan, and κ-Carrageenan and Recovery of Microbial Cultivability via Catalase and Pyruvate.

PubMed

Kawasaki, Kosei; Kamagata, Yoichi

2017-11-01

Previously, we reported that when agar is autoclaved with phosphate buffer, hydrogen peroxide (H 2 O 2 ) is formed in the resulting medium (PT medium), and the colony count on the medium inoculated with environmental samples becomes much lower than that on a medium in which agar and phosphate are autoclaved separately (PS medium) (T. Tanaka et al., Appl Environ Microbiol 80:7659-7666, 2014, https://doi.org/10.1128/AEM.02741-14). However, the physicochemical mechanisms underlying this observation remain largely unknown. Here, we determined the factors affecting H 2 O 2 formation in agar. The H 2 O 2 formation was pH dependent: H 2 O 2 was formed at high concentrations in an alkaline or neutral phosphate buffer but not in an acidic buffer. Ammonium ions enhanced H 2 O 2 formation, implying the involvement of the Maillard reaction catalyzed by phosphate. We found that other gelling agents (e.g., gellan and κ-carrageenan) also produced H 2 O 2 after being autoclaved with phosphate. We then examined the cultivability of microorganisms from a fresh-water sample to test whether catalase and pyruvate, known as H 2 O 2 scavengers, are effective in yielding high colony counts. The colony count on PT medium was only 5.7% of that on PS medium. Catalase treatment effectively restored the colony count of PT medium (to 106% of that on PS medium). In contrast, pyruvate was not as effective as catalase: the colony count on sodium pyruvate-supplemented PT medium was 58% of that on PS medium. Given that both catalase and pyruvate can remove H 2 O 2 from PT medium, these observations indicate that although H 2 O 2 is the main cause of reduced colony count on PT medium, other unknown growth-inhibiting substances that cannot be removed by pyruvate (but can be by catalase) may also be involved. IMPORTANCE The majority of bacteria in natural environments are recalcitrant to laboratory culture techniques. Previously, we demonstrated that one reason for this is the formation of high H 2 O 2 levels in media prepared by autoclaving agar and phosphate buffer together (PT medium). In this study, we investigated the factors affecting H 2 O 2 formation from agar. H 2 O 2 formation is pH dependent, and ammonium ions promote this phosphate-catalyzed H 2 O 2 formation. Amendment of catalase or pyruvate, a well-known H 2 O 2 -scavenging agent, effectively eliminated H 2 O 2 Yet results suggest that growth-inhibiting factor(s) that cannot be eliminated by pyruvate (but can be by catalase) are present in PT medium. Copyright © 2017 American Society for Microbiology.

Phosphate-Catalyzed Hydrogen Peroxide Formation from Agar, Gellan, and κ-Carrageenan and Recovery of Microbial Cultivability via Catalase and Pyruvate

PubMed Central

Kamagata, Yoichi

2017-01-01

ABSTRACT Previously, we reported that when agar is autoclaved with phosphate buffer, hydrogen peroxide (H2O2) is formed in the resulting medium (PT medium), and the colony count on the medium inoculated with environmental samples becomes much lower than that on a medium in which agar and phosphate are autoclaved separately (PS medium) (T. Tanaka et al., Appl Environ Microbiol 80:7659–7666, 2014, https://doi.org/10.1128/AEM.02741-14). However, the physicochemical mechanisms underlying this observation remain largely unknown. Here, we determined the factors affecting H2O2 formation in agar. The H2O2 formation was pH dependent: H2O2 was formed at high concentrations in an alkaline or neutral phosphate buffer but not in an acidic buffer. Ammonium ions enhanced H2O2 formation, implying the involvement of the Maillard reaction catalyzed by phosphate. We found that other gelling agents (e.g., gellan and κ-carrageenan) also produced H2O2 after being autoclaved with phosphate. We then examined the cultivability of microorganisms from a fresh-water sample to test whether catalase and pyruvate, known as H2O2 scavengers, are effective in yielding high colony counts. The colony count on PT medium was only 5.7% of that on PS medium. Catalase treatment effectively restored the colony count of PT medium (to 106% of that on PS medium). In contrast, pyruvate was not as effective as catalase: the colony count on sodium pyruvate-supplemented PT medium was 58% of that on PS medium. Given that both catalase and pyruvate can remove H2O2 from PT medium, these observations indicate that although H2O2 is the main cause of reduced colony count on PT medium, other unknown growth-inhibiting substances that cannot be removed by pyruvate (but can be by catalase) may also be involved. IMPORTANCE The majority of bacteria in natural environments are recalcitrant to laboratory culture techniques. Previously, we demonstrated that one reason for this is the formation of high H2O2 levels in media prepared by autoclaving agar and phosphate buffer together (PT medium). In this study, we investigated the factors affecting H2O2 formation from agar. H2O2 formation is pH dependent, and ammonium ions promote this phosphate-catalyzed H2O2 formation. Amendment of catalase or pyruvate, a well-known H2O2-scavenging agent, effectively eliminated H2O2. Yet results suggest that growth-inhibiting factor(s) that cannot be eliminated by pyruvate (but can be by catalase) are present in PT medium. PMID:28821549

Application of solid-phase extraction to agar-supported fermentation.

PubMed

Le Goff, Géraldine; Adelin, Emilie; Cortial, Sylvie; Servy, Claudine; Ouazzani, Jamal

2013-09-01

Agar-supported fermentation (Ag-SF), a variant of solid-state fermentation, has recently been improved by the development of a dedicated 2 m(2) scale pilot facility, Platotex. We investigated the application of solid-phase extraction (SPE) to Ag-SF in order to increase yields and minimize the contamination of the extracts with agar constituents. The selection of the appropriate resin was conducted on liquid-state fermentation and Diaion HP-20 exhibited the highest recovery yield and selectivity for the metabolites of the model fungal strains Phomopsis sp. and Fusarium sp. SPE applied to Ag-SF resulted in a particular compartmentalization of the culture. The mycelium that requires oxygen to grow migrates to the top layer and formed a thick biofilm. The resin beads intercalate between the agar surface and the mycelium layer, and trap directly the compounds secreted by the mycelium through a "solid-solid extraction" (SSE) process. The resin/mycelium layer is easily recovered by scraping the surface and the target metabolites extracted by methanol. Ag-SF associated to SSE represents an ideal compromise for the production of bioactive secondary metabolites with limited economic and environmental impact.

Comparison of chromogenic Biolog Rainbow agar Shigella/Aeromonas with xylose lysine desoxycholate agar for isolation and detection of Shigella spp. from foods.

PubMed

Zhang, Guodong; Lampel, Keith A

2010-08-01

Shigella outbreaks are widely reported throughout the world. However, it remains a challenge to isolate Shigella spp. from foods by using conventional microbiological media. The main objective of this study was to determine the effectiveness of a novel chromogenic medium, Rainbow agar Shigella/Aeromonas (Rainbow agar), for the isolation and detection of Shigella spp. in foods. All four Shigella species, S. sonnei, S. flexneri, S. dysenteriae, and S. boydii, were studied. Rainbow agar was compared with tryptic soy agar, xylose lysine desoxycholate agar (XLD), and Salmonella Shigella agar (SSA) for enumeration of Shigella spp. in pure culture. This chromogenic agar and XLD were also used to isolate Shigella spp. in artificially contaminated foods (4.8 log CFU/g of food), including lettuce, parsley, cilantro, spinach, potato salad, and shrimp. The inhibitory effect on Shigella growth by Rainbow agar was between that of XLD and SSA. All vegetables studied showed a moderately high background microflora on XLD and Rainbow agar. With artificially inoculated produce, Rainbow agar recovered about 1 to 2 log CFU more S. sonnei, S. dysenteriae, and S. boydii per g of food than did XLD. For potato salad and shrimp, which had low background microflora on Rainbow agar, Rainbow agar was slightly better in recovering Shigella spp. than XLD was in most cases. However, we found that the addition of streptomycin (6.25 mg/liter) to Rainbow agar could facilitate the isolation of Shigella in vegetables tested. In conclusion, Rainbow agar was a much more effective medium than was XLD for the isolation of Shigella spp. from foods.

Radiation Resistance of Asporogenous Bacteria in Frozen Beef

DTIC Science & Technology

1976-03-01

Salmonella enteritidis , and Escherichia coli were used. Cultures were grown to the maximum stationary phase for use as an inoculum. Ground beef containing...eosin methylene blue agar, Shigella- Salmonella agar, and growth on plate count agar with 2.5% and 6.5% NaCl was observed. Penicillin susceptibility was...selective media as follows: Staphylococcus Medium No. 110 for S. aureus; Violet Red Bile Agar for E. coli; and Bismuth Sulfite Agar for S. enteritidis

Comparison of culture based methods for the isolation of Clostridium difficile from stool samples in a research setting.

PubMed

Lister, Michelle; Stevenson, Emma; Heeg, Daniela; Minton, Nigel P; Kuehne, Sarah A

2014-08-01

Effective isolation of Clostridium difficile from stool samples is important in the research setting, especially where low numbers of spores/vegetative cells may be present within a sample. In this study, three protocols for stool culture were investigated to find a sensitive, cost effective and timely method of C. difficile isolation. For the initial enrichment step, the effectiveness of two different rich media, cycloserine-cefoxitin fructose broth (CCFB) and cycloserine-cefoxitin mannitol broth with taurocholate and lysozyme (CCMB-TAL) were compared. For the comparison of four different, selective solid media; Cycloserine-cefoxitin fructose agar (CCFA), Cycloserine-cefoxitin egg yolk agar (CCEY), ChromID C. difficile and tryptone soy agar (TSA) with 5% sheep's blood with and without preceding broth enrichment were used. As a means to enable differentiation between C. difficile and other fecal flora, the effectiveness of the inclusion of a pH indictor (1% Neutral Red), was also evaluated. The data derived indicated that CCFB is more sensitive than CCMB-TAL, however, the latter had an improved recovery rate. A broth enrichment step had a reduced sensitivity over direct plating. ChromID C. difficile showed the best recovery rate whereas CCEY egg yolk agar was the most sensitive of the four. The addition of 1% Neutral Red did not show sufficient colour change when added to CCEY egg yolk agar to be used as a differential medium. For a low cost, timely and sensitive method of isolating C. difficile from stool samples we recommend direct plating onto CCEY egg yolk agar after heat shock. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

INTERLABORATORY EVALUATION OF MI AGAR AND THE US ENVIRONMENTAL PROTECTION AGENCY-APPROVED MEMBRANE FILTER METHOD FOR THE RECOVERY OF TOTAL COLIFORMS AND ESCHERICHIA COLI FROM DRINKING WATER

EPA Science Inventory

A new membrane filter (MF) medium, MI agar, recently validated for use in recovering chlorine-damaged total coloiforms (TC) and Escherichia coli from drinking water, was compared to the US Environmental Protection Agency (EPA)-approved MF method(mEndo agar and nutrient agar suppl...

A previously uncharacterized gene stm0551 plays a repressive role in the regulation of type 1 fimbriae in Salmonella enterica serotype Typhimurium

PubMed Central

2012-01-01

Background Salmonella enterica serotype Typhimurium produces surface-associated fimbriae that facilitate adherence of the bacteria to a variety of cells and tissues. Type 1 fimbriae with binding specificity to mannose residues are the most commonly found fimbrial type. In vitro, static-broth culture favors the growth of S. Typhimurium with type 1 fimbriae, whereas non-type 1 fimbriate bacteria are obtained by culture on solid-agar media. Previous studies demonstrated that the phenotypic expression of type 1 fimbriae is the result of the interaction and cooperation of the regulatory genes fimZ, fimY, fimW, and fimU within the fim gene cluster. Genome sequencing revealed a novel gene, stm0551, located between fimY and fimW that encodes an 11.4-kDa putative phosphodiesterase specific for the bacterial second messenger cyclic-diguanylate monophosphate (c-di-GMP). The role of stm0551 in the regulation of type 1 fimbriae in S. Typhimurium remains unclear. Results A stm0551-deleted stain constructed by allelic exchange constitutively produced type 1 fimbriae in both static-broth and solid-agar medium conditions. Quantative RT-PCR revealed that expression of the fimbrial major subunit gene, fimA, and one of the regulatory genes, fimZ, were comparably increased in the stm0551-deleted strain compared with those of the parental strain when grown on the solid-agar medium, a condition that normally inhibits expression of type 1 fimbriae. Following transformation with a plasmid possessing the coding sequence of stm0551, expression of fimA and fimZ decreased in the stm0551 mutant strain in both culture conditions, whereas transformation with the control vector pACYC184 relieved this repression. A purified STM0551 protein exhibited a phosphodiesterase activity in vitro while a point mutation in the putative EAL domain, substituting glutamic acid (E) with alanine (A), of STM0551 or a FimY protein abolished this activity. Conclusions The finding that the stm0551 gene plays a negative regulatory role in the regulation of type 1 fimbriae in S. Typhimurium has not been reported previously. The possibility that degradation of c-di-GMP is a key step in the regulation of type 1 fimbriae warrants further investigation. PMID:22716649

[Investigation on antibacterial activity of Forsythia suspense Vahl in vitro with Mueller-Hinton agar].

PubMed

Li, Z X; Wang, X H; Zhao, J H; Yang, J F; Wang, X

2000-12-01

To evaluate the antibacterial activity of Forsythia suspensa in vitro with different media. MIC determination of Forsythia suspensa against Staphylococci was performed by the agar dilution method. MIC90 of decoction of Forsythia suspensa against Staphylococcus epidermidis in M-H agar was 1:640, but in nutrient agar 1:40, the antibacterial activity with M-H agar being 16 fold higher than nutrient agar. The M-H agar should be recommended to replace nutrient agar as medium in the antibacterial experiment of Traditional Chinese medicine, and it is better to use multipoint inoculating device in the sensitivity test.

The isolation of salmonellas from British pork sausages and sausage meat.

PubMed Central

Roberts, D.; Boag, K.; Hall, M. L.; Shipp, C. R.

1975-01-01

Between 1969 and 1974, 1467 packets (3309 samples) of pork sausages and sausage meat produced by two large and two medium sized manufacturers and several local butchers were examined for the presence of salmonellas. Of these, 435 packets (786 samples) were found to contain salmonellas, but there was a wide variation in the isolation rates according to the producer. The salmonella incidence in samples from several small and two medium sized producers was low (0-11%) while the results from the two large producers investigated showed a striking difference, the rate of salmonella contamination in the product of one was low (about 2%) and in that of the other consistently high (40-60%). A comparison of liquid enrichment media, incubation temperatures and selective agar media was also carried out to determine the most efficient combination for the isolation of salmonellas from minced meat products. The results showed that (a) incubation of enrichment cultures at 43 degrees C. yielded a consistently greater number of salmonella isolations that at 37 degrees C., regardless of plating medium, (b) tetrathionate broth A (Rolfe) was superior to selenite broth as en enrichment medium at both 37 and 43 degrees C. and (c) brilliant green agar gave better results than deoxycholate citrate sucrose agar and bismuth sulphite agar as a selective medium. PMID:1100710

The isolation of salmonellas from British pork sausages and sausage meat.

PubMed

Roberts, D; Boag, K; Hall, M L; Shipp, C R

1975-10-01

Between 1969 and 1974, 1467 packets (3309 samples) of pork sausages and sausage meat produced by two large and two medium sized manufacturers and several local butchers were examined for the presence of salmonellas. Of these, 435 packets (786 samples) were found to contain salmonellas, but there was a wide variation in the isolation rates according to the producer. The salmonella incidence in samples from several small and two medium sized producers was low (0-11%) while the results from the two large producers investigated showed a striking difference, the rate of salmonella contamination in the product of one was low (about 2%) and in that of the other consistently high (40-60%). A comparison of liquid enrichment media, incubation temperatures and selective agar media was also carried out to determine the most efficient combination for the isolation of salmonellas from minced meat products. The results showed that (a) incubation of enrichment cultures at 43 degrees C. yielded a consistently greater number of salmonella isolations that at 37 degrees C., regardless of plating medium, (b) tetrathionate broth A (Rolfe) was superior to selenite broth as en enrichment medium at both 37 and 43 degrees C. and (c) brilliant green agar gave better results than deoxycholate citrate sucrose agar and bismuth sulphite agar as a selective medium.

A novel chromogenic medium for isolation of Pseudomonas aeruginosa from the sputa of cystic fibrosis patients.

PubMed

Laine, Larissa; Perry, John D; Lee, Jenner; Oliver, Michelle; James, Arthur L; De La Foata, Corinne; Halimi, Diane; Orenga, Sylvain; Galloway, Angela; Gould, F Kate

2009-03-01

A novel chromogenic medium for isolation and identification of Pseudomonas aeruginosa from sputa of cystic fibrosis (CF) patients was evaluated and compared with standard laboratory methods. One hundred sputum samples from distinct CF patients were cultured onto blood agar (BA), Pseudomonas CN selective agar (CN) and a Pseudomonas chromogenic medium (PS-ID). All Gram-negative morphological variants from each medium were subjected to antimicrobial susceptibility testing, and identification using a combination of biochemical and molecular methods. P. aeruginosa was isolated from 62 samples after 72 h incubation. Blood agar recovered P. aeruginosa from 56 samples (90.3%) compared with 59 samples (95.2%) using either CN or PS-ID. The positive predictive value of PS-ID (98.3%) was significantly higher than growth on CN (88.5%) for identification of P. aeruginosa (P<0.05). PS-ID is a promising medium allowing for the isolation and simultaneous identification of P. aeruginosa from sputa of CF patients.

Fish meal extract bile esculin agar (FMBE) a selective medium for Bacteroides fragilis group.

PubMed

Beena, V K; Rao, S; Kotian, M; Shivananda, P G

1997-07-01

Fish meal extract bile esculin agar (FMBE) is prepared using Fish meal extract concentrate as the basal substance, for the selective isolation and presumptive identification of B.fragilis group. The efficiency of the medium was evaluated by growing stock cultures of B.fragilis groups as well as inoculating clinical specimens and comparing the results with Bacteroides bile esculin agar (BBE). All the 87 stock cultures of B.fragilis grew on FMBE and BBE. No other anaerobes tested grew on the medium. However 7 out of 65 neomycin resistant aerobes grew on the FMBE. From the 100 clinical samples, 62 strains of B. Fragilis group were recovered on FMBE and BBE, and 53 strains on supplemented BHIBA. The cost effectiveness, selectivity and the ability to detect esculin hydrolysis will enable FMBE as a suitable medium as comparable to that of BBE, if not superior.

Improved Medium for Selecting Nitrate-Nonutilizing (nit) Mutants of Verticillium dahliae.

PubMed

Korolev, N; Katan, T

1997-10-01

ABSTRACT Nitrate-nonutilizing (nit) mutants are commonly used to determine vegetative compatibility between isolates of Verticillium dahliae by complementation (heterokaryon) testing. These mutants emerge spontaneously as chlorate-resistant sectors growing out of partially restricted, wild-type colonies on chlorate-amended media. The commonly used chlorate media are based on minimal medium (MMC) or cornmeal agar (CMC), amended with potassium chlorate. nit mutants recovered on these media constituted 10 to 36%(on MMC) and 25 to 45%(on CMC) of the apparently resistant sectors. An improved water agar chlorate medium (WAC) is described that is more effective for selecting chlorate-resistant nit mutants. WAC medium consists of agar (2%), glucose (0.02%), and potassium chlorate (2 to 5%). On WAC, growth of most V. dahliae isolates was strongly inhibited, and 66 to 100%(average >80%) of the chlorate-resistant sectors formed were nit mutants. Most mutants were characterized as nit1, and about 6% as NitM.

NEW MEDIUM FOR THE SIMULTANEOUS DETECTION OF TOTAL COLIFORMS AND ESCHERICHIA COLI IN WATER (PUBLISHED ERRATUM APPEARS IN APP ENVIRON MICROBIOL 1993 DEC;59(12):4378)

EPA Science Inventory

A new membrane filter agar medium (MI agar) containing a chromogen, indoxyl-beta-D-glucuronide, and a fluorogen, 4-methylumbelliferyl-beta-D-galactopyranoside, was developed to simultaneously detect and enumerate Escherichia coli and total coliforms (TC) in water samples on the b...

Comparison of CHROMagar Salmonella Medium and Xylose-Lysine-Desoxycholate and Salmonella-Shigella Agars for Isolation of Salmonella Strains from Stool Samples

PubMed Central

Maddocks, Susan; Olma, Tom; Chen, Sharon

2002-01-01

The growth and appearance of 115 stock Salmonella isolates on a new formulation of CHROMagar Salmonella (CAS) medium were compared to those on xylose-lysine-desoxycholate agar (XLD), Salmonella-Shigella agar (SS), and Hektoen enteric agar (HEA) media. CAS medium was then compared prospectively to XLD and SS for the detection and presumptive identification of Salmonella strains in 500 consecutive clinical stool samples. All stock Salmonella isolates produced typical mauve colonies on CAS medium. Nine Salmonella strains were isolated from clinical specimens. The sensitivities for the detection of salmonellae after primary plating on CAS medium and the combination of XLD and SS after enrichment were 100%. The specificity for the detection of salmonellae after primary plating on CAS medium (83%) was significantly (P < 0.0001) higher than that after primary plating on the combination of SS and XLD media (55%) (a 28% difference in rates; 95% confidence interval, 23.0 to 34%). Twenty-nine non-Salmonella organisms produced mauve colonies on CAS medium, including 17 Candida spp. (59%) and 8 Pseudomonas spp. (28%). These were easily excluded as salmonellae by colony morphology, microscopic examination of a wet preparation, or oxidase testing. One biochemically inert Escherichia coli isolate required further identification to differentiate it from Salmonella spp. The use of plating on CAS medium demonstrated high levels of sensitivity and specificity and reduced the time to final identification of Salmonella spp., resulting in substantial cost savings. It can be recommended for use for the primary isolation of Salmonella spp. from stool specimens. Other media (e.g., XLD) are required to detect Shigella spp. concurrently. PMID:12149365

Enhanced bioactivity of ZnO nanoparticles—an antimicrobial study

PubMed Central

Padmavathy, Nagarajan; Vijayaraghavan, Rajagopalan

2008-01-01

In this study, we investigate the antibacterial activity of ZnO nanoparticles with various particle sizes. ZnO was prepared by the base hydrolysis of zinc acetate in a 2-propanol medium and also by a precipitation method using Zn(NO3)2 and NaOH. The products were characterized by x-ray diffraction (XRD) analysis, transmission electron microscopy (TEM) and photoluminescence (PL) spectroscopy. Bacteriological tests such as minimum inhibitory concentration (MIC) and disk diffusion were performed in Luria-Bertani and nutrient agar media on solid agar plates and in liquid broth systems using different concentrations of ZnO by a standard microbial method for the first time. Our bacteriological study showed the enhanced biocidal activity of ZnO nanoparticles compared with bulk ZnO in repeated experiments. This demonstrated that the bactericidal efficacy of ZnO nanoparticles increases with decreasing particle size. It is proposed that both the abrasiveness and the surface oxygen species of ZnO nanoparticles promote the biocidal properties of ZnO nanoparticles. PMID:27878001

Enhanced bioactivity of ZnO nanoparticles—an antimicrobial study

NASA Astrophysics Data System (ADS)

Padmavathy, Nagarajan; Vijayaraghavan, Rajagopalan

2008-07-01

In this study, we investigate the antibacterial activity of ZnO nanoparticles with various particle sizes. ZnO was prepared by the base hydrolysis of zinc acetate in a 2-propanol medium and also by a precipitation method using Zn(NO3)2 and NaOH. The products were characterized by x-ray diffraction (XRD) analysis, transmission electron microscopy (TEM) and photoluminescence (PL) spectroscopy. Bacteriological tests such as minimum inhibitory concentration (MIC) and disk diffusion were performed in Luria-Bertani and nutrient agar media on solid agar plates and in liquid broth systems using different concentrations of ZnO by a standard microbial method for the first time. Our bacteriological study showed the enhanced biocidal activity of ZnO nanoparticles compared with bulk ZnO in repeated experiments. This demonstrated that the bactericidal efficacy of ZnO nanoparticles increases with decreasing particle size. It is proposed that both the abrasiveness and the surface oxygen species of ZnO nanoparticles promote the biocidal properties of ZnO nanoparticles.

Effect of Soybean Casein Digest Agar Lot on Number of Bacillus stearothermophilus Spores Recovered †

PubMed Central

Pflug, I. J.; Smith, Geraldine M.; Christensen, Ronald

1981-01-01

In recent years it has become increasingly apparent that Bacillus stearothermophilus spores are affected by various environmental factors that influence the performance of the spores as biological indicators. One environmental factor is the recovery medium. The effect of different lots of commercial soybean casein digest agar on the number of colony-forming units per plate was examined in two series of experiments: (i) several lots of medium from two manufacturers were compared in single experiments, and (ii) paired media experiments with four lots of medium were carried out and yielded three-point survivor curves. The results demonstrate that commercial soybean casein digest agar is variable on a lot-to-lot basis. The variation was lowest when recovering unheated or minimally heated spores and increased greatly with the severity of heating. PMID:16345822

Selective vs. nonselective media and direct plating vs. enrichment technique in isolation of Vibrio cholerae: recommendations for clinical laboratories.

PubMed

Rennels, M B; Levine, M M; Daya, V; Angle, P; Young, C

1980-09-01

The occurrence of human cholera along the Gulf of Mexico and the isolation of Vibrio cholerae O1 from the Gulf and Chesapeake Bay make it imperative that microbiology laboratories along estuaries develop the capabilities to culture for these pathogens. In attempts to devise a simplified but efficient culture procedure, a selective medium, thiosulfate-citrate-bile salts-sucrose (TCBS) agar, was compared with a nonselective medium, gelatin agar (GA), and the utility of enrichment was examined. TCBS agar detected 99% of the stools found to be positive by all techniques combined, whereas GA identified only 80%. Of acute diarrheal stools, 96% were positive on direct plating, whereas only 66% of formed stools containing V. cholerae were detected by direct plating. Stools from patients with acute diarrhea can be plated directly into TCBS agar alone; stools from persons shedding low numbers of organisms (such as contacts, carriers, or patients receiving antibiotics) should be incubated first in an enrichment broth and then on TCBS agar.

Comparison of media for detection of fungi on spacecraft

NASA Technical Reports Server (NTRS)

Herring, C. M.; Brandsberg, J. W.; Oxborrow, G. S.; Puleo, J. R.

1974-01-01

Five media, including Trypticase soy agar (TSA; BBL) pour plates, spread plates of TSA, Mycophil agar with chloromycetin, Mycophil agar with chloromycetin and Actidione, and cornmeal agar with chloromycetin were quantitatively and qualitatively compared for the detection of fungi on spacecraft. Cornmeal agar with chloromycetin yielded the highest number of fungal colonies, although not always significantly higher than Mycophil agar with chloromycetin or TSA spread plates. Cornmeal agar with chloromycetin also gave the best qualitative representation of fungi on the spacecraft, recovering 68% of the genera found from all media. This medium yielded 10 times the number of fungal colonies and 3 times the number of genera found on TSA pour plates as currently used for spacecraft assay.

Comparison of media for detection of fungi on spacecraft.

PubMed

Herring, C M; Brandsberg, J W; Oxborrow, G S; Puleo, J R

1974-03-01

Five media, including Trypticase soy agar (TSA; BBL) pour plates, spread plates of TSA, Mycophil agar with chloromycetin, Mycophil agar with chloromycetin and Actidione, and cornmeal agar with chloromycetin were quantitatively and qualitatively compared for the detection of fungi on spacecraft. Cornmeal agar with chloromycetin yielded the highest number of fungal colonies, although not always significantly higher than Mycophil agar with chloromycetin or TSA spread plates. Cornmeal agar with chloromycetin also gave the best qualitative representation of fungi on the spacecraft, recovering 68% of the genera found from all media. This medium yielded 10 times the number of fungal colonies and 3 times the number of genera found on TSA pour plates as currently used for spacecraft assay.

An improved agar medium for growth of Geobacillus thermoglucosidarius strains.

PubMed

Javed, M; Baghaei-Yazdi, N; Qin, W; Amartey, S

2017-01-01

Geobacillus species have potential applications in many biotechnological processes. They are fastidious in their vitamin and amino acid requirements. A new semi-defined agar medium (SDM) was developed which gave consistently high viable cell counts of various G. thermoglucosidasius strains (5×10 8 -6×10 8 cfu/ml) under aerobic conditions at 70°C. Copyright © 2016 Elsevier B.V. All rights reserved.

Effects of physical factors on the swarming motility of text itPseudomonas aeruginosa

NASA Astrophysics Data System (ADS)

Si, Tieyan; Ma, Zidong; Tang, Wai Shing; Yang, Alexander; Tang, Jay

Many species of bacteria can spread over a semi-solid surface via a particular form of collective motion known as surface swarming. Using Pseudomonas aeruginosa as a model organism, we investigate physical factors that either facilitate or restrict the swarming motility. The semi-solid surface is typically formed by 0.5-1% agar containing essential nutrients for the bacterial growth and proliferation. Most bacterial species, including P. aeruginosa, synthesize bio-surfactants to aid in swarming. We found addition of exogenous surfactants such as triton into the agar matrix enhances the swarming. In contrast, increasing agar percentage, infusing osmolites, and adding viscous agents all decrease swarming. We propose that the swarming speed is restricted by the rate of water supply from within the agar gel and by the line tension at the swarm front involving three materials in contact: the air, the bacteria propelled liquid film, and the agar substrate.

How we determined the most reliable solid medium for studying treatment of tuberculosis

PubMed Central

Heilig, Charles M.; Feng, Pei-Jean I.; Joloba, Moses L.; Johnson, John L.; Morgan, Karen; Gitta, Phineas; Boom, W. Henry; Mayanja-Kizza, Harriet; Eisenach, Kathleen D.; Bozeman, Lorna; Goldberg, Stefan V.

2014-01-01

Phase 2 clinical trials for tuberculosis (TB) treatment require reliable culture methods to determine presence or absence of Mycobacterium tuberculosis (Mtb) over the course of therapy, as these trials are based primarily on bacteriological endpoints. We evaluate which of 5 solid media is most reliable: Lowenstein-Jensen (LJ) egg-based medium and 4 Middlebrook agar media (nonselective 7H10 and 7H11 and selective 7H10 and 7H11). We analyze 393 specimens from 50 HIV-negative Ugandan adults with newly-diagnosed, pulmonary TB and high acid-fast bacillus smear grade. Specimens were collected every 2–4 weeks during the first 12 weeks of therapy. We compare the results for each culture to 2 composite reference standards—one that was deemed positive if any solid culture was positive for Mtb and another based on latent class analysis. Both reference standards established that the 2 selective Middlebrook media most reliably determine the presence or absence of Mtb (P<0.003), largely because of their lower contamination rates. We also showed that results on Middlebrook media were similar to each other, while LJ was most frequently discordant. Contaminated results appeared more likely to be truly negative than to harbor undetected Mtb. PMID:24661816

How we determined the most reliable solid medium for studying treatment of tuberculosis.

PubMed

Heilig, Charles M; Feng, Pei-Jean I; Joloba, Moses L; Johnson, John L; Morgan, Karen; Gitta, Phineas; Boom, W Henry; Mayanja-Kizza, Harriet; Eisenach, Kathleen D; Bozeman, Lorna; Goldberg, Stefan V

2014-05-01

Phase 2 clinical trials for tuberculosis (TB) treatment require reliable culture methods to determine presence or absence of Mycobacterium tuberculosis (Mtb) over the course of therapy, as these trials are based primarily on bacteriological endpoints. We evaluate which of 5 solid media is most reliable: Lowenstein-Jensen (LJ) egg-base medium and 4 Middlebrook agar media (nonselective 7H10 and 7H11 and selective 7H10 and 7H11). We analyze 393 specimens from 50 HIV-negative Ugandan adults with newly-diagnosed, pulmonary TB and high acid-fast bacillus smear grade. Specimens were collected every 2-4 weeks during the first 12 weeks of therapy. We compare the results for each culture to 2 composite reference standards--one that was deemed positive if any solid culture was positive for Mtb and another based on latent-class analysis. Both reference standards established that the 2 selective Middlebrook media most reliably determine the presence or absence of Mtb (P < 0.003), largely because of their lower contamination rates. We also showed that results on Middlebrook media were similar to each other, while LJ was most frequently discordant. Contaminated results appeared more likely to be truly negative than to harbor undetected Mtb. Published by Elsevier Ltd.

Equivalency testing of TTC Tergitol 7 agar (ISO 9308-1:2000) with five culture media for the detection of E. coli in water samples in Greece.

PubMed

Mavridou, A; Smeti, E; Mandilara, G; Mandilara, G; Boufa, P; Vagiona-Arvanitidou, M; Vantarakis, A; Vassilandonopoulou, G; Pappa, O; Roussia, V; Tzouanopoulos, A; Livadara, M; Aisopou, I; Maraka, V; Nikolaou, E; Mandilara, G

2010-01-01

In this study ten laboratories in Greece compared the performance of reference method TTC Tergitol 7 Agar (with the additional test of beta-glucuronidase production) with five alternative methods, to detect E. coli in water, in line with European Water Directive recommendations. The samples were prepared by spiking drinking water with sewage effluent following a standard protocol. Chlorinated and non-chlorinated samples were used. The statistical analysis was based on the mean relative difference of confirmed counts and was performed in line with ISO 17994. The results showed that in total, three of the alternative methods (Chromocult Coliform agar, Membrane Lauryl Sulfate agar and Trypton Bilex-glucuronidase medium) were not different from TTC Tergitol 7 agar (TTC Tergitol 7 agar vs Chromocult Coliform agar, 294 samples, mean RD% 5.55; vs MLSA, 302 samples, mean RD% 1; vs TBX, 297 samples, mean RD% -2.78). The other two alternative methods (Membrane Faecal coliform medium and Colilert 18/ Quantitray) gave significantly higher counts than TTC Tergitol 7 agar (TTC Tergitol 7 agar vs MFc, 303 samples, mean RD% 8.81; vs Colilert-18/Quantitray, 76 samples, mean RD% 18.91). In other words, the alternative methods generated performance that was as reliable as, or even better than, the reference method. This study will help laboratories in Greece overcome culture and counting problems deriving from the EU reference method for E. coli counts in water samples.

Different culture media containing methyldopa for melanin production by Cryptococcus species.

PubMed

Menezes, Ralciane de Paula; Penatti, Mário Paulo Amante; Pedroso, Reginaldo dos Santos

2011-10-01

Melanin production by species of Cryptococcus is widely used to characterize C. neoformans complex in mycology laboratories. This study aims to test the efficacy of methyldopa from pharmaceutical tablet as a substrate for melanin production, to compare the production of melanin using different agar base added with methyldopa, and to compare the melanin produced in those media with that produced in Niger seed agar and sunflower seed agar by C. neoformans, C. laurentii, and C. albidus. Two isolates of each species, C. neoformans, C. laurentii, and C. albidus, and one of Candida albicans were used to experimentally detect conditions for melanin production. The following media were tested: Mueller-Hinton agar (MHA), brain and heart infusion agar (BHIA), blood agar base (BAB), and minimal medium agar (MMA), all added with methyldopa, and the media Niger seed agar (NSA) and sunflower seed agar (SSA). All isolates grew in most of the culture media after 24h. Strains planted on media BAB and BHIA showed growth only after 48h. All isolates produced melanin in MMA, MHA, SSA, and NSA media. Methyldopa in the form pharmaceutical tablet can be used as a substrate for melanin production by Cryptococcus species; minimal medium plus methyldopa was more efficient than the BAB, MHA, and BHIA in the melanin production; and NSA and SSA, followed by MMA added with methyldopa, were more efficient than other media studied for melanin production by all strains studied.

New Chromogenic Agar Medium for the Identification of Candida spp.

PubMed Central

Cooke, Venitia M.; Miles, R. J.; Price, R. G.; Midgley, G.; Khamri, W.; Richardson, A. C.

2002-01-01

A new chromogenic agar medium (Candida diagnostic agar [CDA]) for differentiation of Candida spp. is described. This medium is based on Sabouraud dextrose agar (Oxoid CM41) and contains (per liter) 40.0 g of glucose, 10.0 g of mycological peptone, and 15.0 g of agar along with a novel chromogenic glucosaminidase substrate, ammonium 4-{2-[4-(2-acetamido-2-deoxy-β-d-glucopyranosyloxy)-3-methoxyphenyl]-vinyl}-1-(propan-3-yl-oate)-quinolium bromide (0.32 g liter−1). The glucosaminidase substrate in CDA was hydrolyzed by Candida albicans and Candida dubliniensis, yielding white colonies with deep-red spots on a yellow transparent background after 24 to 48 h of incubation at 37°C. Colonies of Candida tropicalis and Candida kefyr were uniformly pink, and colonies of other Candida spp., including Candida glabrata and Candida parapsilosis, were white. CDA was evaluated by using 115 test strains of Candida spp. and other clinically important yeasts and was compared with two commercially available chromogenic agars (Candida ID agar [bioMerieux] and CHROMagar Candida [CHROMagar Company Ltd.]). On all three agars, colonies of C. albicans were not distinguished from colonies of C. dubliniensis. However, for the group containing C. albicans plus C. dubliniensis, both the sensitivity and the specificity of detection when CDA was used were 100%, compared with values of 97.6 and 100%, respectively, with CHROMagar Candida and 100 and 96.8%, respectively, with Candida ID agar. In addition, for the group containing C. tropicalis plus C. kefyr, the sensitivity and specificity of detection when CDA was used were also 100%, compared with 72.7 and 98.1%, respectively, with CHROMagar Candida. Candida ID agar did not differentiate C. tropicalis and C. kefyr strains but did differentiate members of a broader group (C. tropicalis, C. kefyr, Candida lusitaniae plus Candida guilliermondii); the sensitivity and specificity of detection for members of this group were 94.7 and 93.8%, respectively. In addition to the increased sensitivity and/or specificity of Candida detection when CDA was used, differentiation of colony types on CDA (red spotted, pink, or no color) was unambiguous and did not require precise assessment of colony color. PMID:12089051

Fermentation of Acid-pretreated Corn Stover to Ethanol Without Detoxification Using Pichia stipitis

NASA Astrophysics Data System (ADS)

Agbogbo, Frank K.; Haagensen, Frank D.; Milam, David; Wenger, Kevin S.

In this work, the effect of adaptation on P. stipitis fermentation using acidpretreated corn stover hydrolyzates without detoxification was examined. Two different types of adaptation were employed, liquid hydrolyzate and solid state agar adaptation. Fermentation of 12.5% total solids undetoxified acid-pretreated corn stover was performed in shake flasks at different rotation speeds. At low rotation speed (100 rpm), both liquid hydrolyzate and solid agar adaptation highly improved the sugar consumption rate as well as ethanol production rate compared to the wild-type strains. The fermentation rate was higher for solid agar-adapted strains compared to liquid hydrolyzate-adapted strains. At a higher rotation speed (150 rpm), there was a faster sugar consumption and ethanol production for both the liquid-adapted and the wild-type strains. However, improvements in the fermentation rate between the liquid-adapted and wild strains were less pronounced at the high rotation speed.

Evaluation of standard and modified M-FC, MacConkey, and Teepol media for membrane filtration counting of fecal coliforms in water.

PubMed

Grabow, W O; Hilner, C A; Coubrough, P

1981-08-01

MacConkey agar, standard M-FC agar, M-FC agar without rosolic acid, M-FC agar with a resuscitation top layer, Teepol agar, and pads saturated with Teepol broth, were evaluated as growth media for membrane filtration counting of fecal coliform bacteria in water. In comparative tests on 312 samples of water from a wide variety of sources, including chlorinated effluents, M-FC agar without rosolic acid proved the medium of choice because it generally yielded the highest counts, was readily obtainable, easy to prepare and handle, and yielded clearly recognizable fecal coliform colonies. Identification of 1,139 fecal coliform isolates showed that fecal coliform tests cannot be used to enumerate Escherichia coli because the incidence of E. coli among fecal coliforms varied from an average of 51% for river water to 93% for an activated sludge effluent after chlorination. The incidence of Klebsiella pneumoniae among fecal coliforms varied from an average of 4% for the activated sludge effluent after chlorination to 32% for the river water. The advantages of a standard membrane filtration procedure for routine counting of fecal coliforms in water using M-FC agar without rosolic acid as growth medium, in the absence of preincubation or resuscitation steps, are outlined.

Actinomadura Species: Laboratory Maintenance and Ribosome Engineering.

PubMed

Dhakal, Dipesh; Chung, Nguyen Thanh; Rayamajhi, Vijay; Sohng, Jae Kyung

2017-02-06

Actinomadura spp. are aerobic, Gram-positive, catalase-positive, non-acid fast, non-motile actinomycetes. Some species of Actinomadura are associated with opportunistic infections in humans. However, many bioactive compounds with pharmaceutical applications can be isolated from various Actinomadura spp. This unit includes general protocols for the laboratory maintenance of Actinomadura spp., including growth in liquid medium, growth on solid agar, long-term storage, and generation of a higher producing strain by ribosome engineering. Actinomadura hibisca P157-2 is used as a prototype for explaining the considerations for efficient laboratory maintenance of Actinomadura spp. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley & Sons, Inc.

Factors affecting isolation and identification of Mycobacterium avium subsp. paratuberculosis from fecal and tissue samples in a liquid culture system.

PubMed

Whittington, Richard J

2009-03-01

Culture of Mycobacterium avium subsp. paratuberculosis is the definitive diagnostic test for Johne's disease, a chronic granulomatous enteropathy of animals. Compared to solid media, the identification of all strains of the organism in liquid media can be more difficult because the appearance of colonies and mycobactin dependence are not observable, and the growth of other organisms needs to be distinguished, commonly by PCR. Factors affecting the isolation rate of S strains and the contamination rate in modified Middlebrook 7H9 broth (Bactec 12B) and 7H10 agar were studied using 11,598 fecal samples and 2,577 tissue samples from sheep from 1,421 farms over 10 years. Minimization of contamination in Bactec cultures required the avoidance of the carryover of fecal particles from the first sedimentation step in the double-incubation centrifugation method, and contamination was reduced significantly by incubating the sample in a solution containing vancomycin, amphotericin B, and nalidixic acid for 3 days compared to 2 days. The growth of irrelevant microorganisms confounded the identification of M. avium subsp. paratuberculosis in liquid culture by inhibiting IS900 PCR and in solid medium culture by inhibiting the growth of M. avium subsp. paratuberculosis or obscuring colonies. The contamination of samples was clustered in certain laboratory submissions and was reduced by including ampicillin in Bactec medium without affecting the odds of isolation of M. avium subsp. paratuberculosis. The long-term contamination rate for fecal cultures was about 7%, and that for tissue cultures was <0.2%. Liquid medium was more sensitive than solid medium culture for M. avium subsp. paratuberculosis. The applicability of these findings for C strains is discussed.

Comparison of Media for Detection of Fungi on Spacecraft

PubMed Central

Herring, C. M.; Brandsberg, J. W.; Oxborrow, G. S.; Puleo, J. R.

1974-01-01

Five media, including Trypticase soy agar (TSA; BBL) pour plates, spread plates of TSA, Mycophil agar with chloromycetin, Mycophil agar with chloromycetin and Actidione, and cornmeal agar with chloromycetin were quantitatively and qualitatively compared for the detection of fungi on spacecraft. Cornmeal agar with chloromycetin yielded the highest number of fungal colonies, although not always significantly higher than Mycophil agar with chloromycetin or TSA spread plates. Cornmeal agar with chloromycetin also gave the best qualitative representation of fungi on the spacecraft, recovering 68% of the genera found from all media. This medium yielded 10 times the number of fungal colonies and 3 times the number of genera found on TSA pour plates as currently used for spacecraft assay. PMID:4151044

Study of methods for the improvement of bacterial transport media

NASA Technical Reports Server (NTRS)

Gardner, R. L.; Beakley, J. W.

1973-01-01

A series of 500 transport media recipes was tested for ability to hold pure cultures of Streptococcus equisimilus, Corynebacterium equi, Neisseria perflava, and Haemophilus parainfluenzae for 21 days. Stuart Medium Base with 0.4% agar was used as the control medium for this and the other experiments in the investigation. At the end of the holding period inoculated transport media were quantitatively assayed, and the control media were assayed immediately after inoculation. Three vials of each medium were inoculated with an organism, and each vial's medium was diluted and spread on duplicate plates. Assay media for this experiment included Brain Heart Infusion,(BHIA) Tryptic Soy Agar, and BHIA with 1% Isovitalex enrichment.

Strategy to obtain axenic cultures from field-collected samples of the cyanobacterium Phormidium animalis.

PubMed

Vázquez-Martínez, Guadalupe; Rodriguez, Mario H; Hernández-Hernández, Fidel; Ibarra, Jorge E

2004-04-01

An efficient strategy, based on a combination of procedures, was developed to obtain axenic cultures from field-collected samples of the cyanobacterium Phormidium animalis. Samples were initially cultured in solid ASN-10 medium, and a crude separation of major contaminants from P. animalis filaments was achieved by washing in a series of centrifugations and resuspensions in liquid medium. Then, manageable filament fragments were obtained by probe sonication. Fragmentation was followed by forceful washing, using vacuum-driven filtration through an 8-microm pore size membrane and an excess of water. Washed fragments were cultured and treated with a sequential exposure to four different antibiotics. Finally, axenic cultures were obtained from serial dilutions of treated fragments. Monitoring under microscope examination and by inoculation in Luria-Bertani (LB) agar plates indicated either axenicity or the degree of contamination throughout the strategy.

Differential plating medium for quantitative detection of histamine-producing bacteria.

PubMed Central

Niven, C F; Jeffrey, M B; Corlett, D A

1981-01-01

A histidine-containing agar medium has been devised for quantitative detection of histamine-producing bacteria that are alleged to be associated with scombroid fish poisoning outbreaks. The responsible bacteria produce a marked pH change in the agar, with attendant color change of pH indicator adjacent to the colonies, thus facilitating their recognition. Proteus morganii and Klebsiella pneumoniae were the two most common histidine-decarboxylating species isolated from scombroid fish and mahi mahi. PMID:7013698

Isolation of nontuberculous mycobacteria from soil using Middlebrook 7H10 agar with increased malachite green concentration.

PubMed

Hu, Yuli; Yu, Xinglong; Zhao, Dun; Li, Runcheng; Liu, Yang; Ge, Meng; Hu, Huican

2017-12-01

Environmental exposure is considered to be responsible for nontuberculous mycobacterial infections in humans. To facilitate the isolation of mycobacteria from soil, Middlebrook 7H10 agar was optimized as an enhanced selective medium by increasing the concentration of malachite green. A series of modified Middlebrook 7H10 agar media with malachite green concentrations ranging from 2.5 to 2500 mg/L was evaluated using 20 soil samples decontaminated with 3% sodium dodecyl sulfate plus 2% NaOH for 30 min. Among these modified Middlebrook 7H10 media, the medium with malachite green at a concentration of 250 mg/L, i.e., at the same concentration as in Löwenstein-Jensen medium, was the most effective in terms of the number of plates with mycobacterial growth. This medium was further evaluated with 116 soil samples. The results showed that 87.1% (101/116) of the samples produced mycobacterial growth, and 15 samples (12.9%) produced no mycobacterial growth. Of the plates inoculated with the soil samples, each in duplicate, 5.2% (12/232) showed late contamination. In total, 19 mycobacterial species were isolated, including seven (36.8%) rapidly growing mycobacteria and 12 (63.2%) slowly growing mycobacteria. Our results demonstrate that the modified Middlebrook 7H10 agar with 250 mg/L malachite green is useful for the primary isolation of nontuberculous mycobacteria from soil.

Combination of nutrients in a mammalian cell culture medium kills cryptococci.

PubMed

Granger, Donald L; Call, Donna M

2018-06-06

We found that a large inoculum of Cryptococcus gattii cells, when plated on Dulbecco's modified eagle's medium (DMEM) incorporated into agar, died within a few hours provided that DMEM agar plates had been stored in darkness for approximately 3 days after preparation. Standard conditions were developed for quantification of killing. The medium lost its fungicidal activity when exposed to visible light of wave length ∼400 nm. The amount of energy required was estimated at 5.8 × 104 joules @ 550 nm. Liquid DMEM conditioned by incubation over DMEM agar plates stored in darkness was fungicidal. We found that fungicidal activity was heat-stable (100°C). Dialysis tubing with MWC0 < 100 Daltons retained fungicidal activity. Neutral pH was required. Strains of Cryptococcus were uniformly sensitive, but some Candida species were resistant. Components of DMEM required for killing were pyridoxal and cystine. Micromolar amounts of iron shortened the time required for DMEM agar plates to become fungicidal when stored in the dark. Organic and inorganic compounds bearing reduced sulfur atoms at millimolar concentrations inhibited fungicidal activity. Our results point to a light-sensitive antifungal compound formed by reaction of pyridoxal with cystine possibly by Schiff base formation.

Comparison of culture media for the recovery of airborne yeast in wineries.

PubMed

Ocón, E; Garijo, P; Santamaría, P; López, R; Olarte, C; Gutiérrez, A R; Sanz, S

2013-09-01

The direct air sampling impaction method on agar was evaluated using aerobiocollectors for the recovery of yeasts present in the winery air. Three culture media with different composition and specificity were studied. In addition, a resuscitation phase was included before the culture in the specificity medium [in the case of the Dekkera-Brettanomyces Differential Medium (DBDM) medium]. Sampling was conducted at different times of the year and in different parts of the wineries, which were different in age and design. Both the Chloramphenicol Glucose Agar (CGA) and Agar Lysine AL media recovered yeasts from the air without any prior resuscitation phase. CGA was able to recover a higher number of colony-forming units of yeasts than the other media. Consequently, to estimate the number of yeasts present in winery air, the best choice of medium would be CGA. The AL medium permitted the growth of the greatest range of genera and species. If the aim is to study the diversity of yeasts present in the air, the most suitable medium is AL. Neither CGA nor AL proved suitable for recovering yeasts of the Brettanomyces genus. The DBDM medium was the only one which provided sufficient specificity for their recovery and identification from the air, although their special characteristics made a prior protocol of resuscitation necessary. © 2013 The Society for Applied Microbiology.

Comparison of media for enumeration of coliform bacteria and Escherichia coli in non-disinfected water.

PubMed

Pitkänen, Tarja; Paakkari, Piia; Miettinen, Ilkka T; Heinonen-Tanski, Helvi; Paulin, Lars; Hänninen, Marja-Liisa

2007-03-01

In this work alternative media for detection and enumeration of E. coli and coliform bacteria were compared to the reference method ISO 9308-1 (LTTC) using non-disinfected water samples with background flora. The alternative media included LES Endo agar medium (LES Endo), Colilert-18 with 51-well Quanti-tray (Colilert), Chromocult Coliform agar (CC), Harlequin E. coli/Coliform medium (HECM) and Chromogenic Escherichia coli/Coliform medium (CECM). A total of 110 samples of groundwater, bathing water and spiked water was used. Our results revealed that confirmation of coliform bacteria counts is necessary, not only on lactose-based LTTC and LES Endo media, but also on the chromogenic agar media tested, due to the growth of oxidase positive colonies. LTTC and CC media also allowed the growth of some morphologically typical coliform colonies containing gram-positive bacteria. The recovery of coliform bacteria was lower on LES Endo than on LTTC. In most cases Colilert, CC, HECM and CECM gave higher coliform counts than LTTC. The use of the LTTC medium led to higher E. coli counts than obtained with any of the alternative mediums. There are three explanations for this: (1) high sensitivity of LTTC, (2) false positives on LTTC or (3) false negatives especially with Colilert, but also with chromogenic agar media. Although LTTC was found to be a very sensitive medium, the high degree of background growth of non-disinfected waters disturbed substantially the use of it. In conclusion, our results suggest that Colilert, CC and CECM are potential alternative media for detection of coliform bacteria and E. coli from non-disinfected water.

Growth of Streptococcus mutans on various selective media.

PubMed

Emilson, C G; Bratthall, D

1976-07-01

The ability of Streptococcus mutans to grow on mitis-salivarius (MS) agar, MC agar, mitis-sucrose-bacitracin (MSB), BCY agar, and MM10 sucrose agar was studied. Batch cultures of S. mutans serotype a demonstrated no growth on MSB agar. Certain serotype d and g strains did not grow on MC agar. The yield for most strains of other serotypes on these selective media was lower compared with that on MS agar. The number of total colony-forming units on BCY and MM10 sucrose agar was similar to the blood agar results. Similar data were obtained when fermenter-grown strains, harvested in the middle or the end of the logarithmic growth phase, were used for inoculation of the various media. Enumeration of S. mutans from plaque samples plated on MC and MSB agar yielded about 75% of the counts obtained on MS or the nonselective medium. When the proportions of S. mutans were expressed as a percentage of the total cultivable flora, the selective media (MC and MSB agar) showed approximately 10% lower values than the MS, BCY, and MM10 sucrose agar.

Tobacco Agar, a New Medium for Differentiating Candida dubliniensis from Candida albicans

PubMed Central

Khan, Zia U.; Ahmad, Suhail; Mokaddas, Eiman; Chandy, Rachel

2004-01-01

Isolates of Candida dubliniensis may be misidentified as Candida albicans in microbiological laboratories if only the germ tube and/or the chlamydospore test is used for identification to the species level. In this study, we have evaluated the efficacy of tobacco agar for the differentiation of C. dubliniensis from C. albicans. On this medium at 28°C, all 30 C. dubliniensis isolates produced yellowish-brown colonies with hyphal fringes and abundant chlamydospores, whereas 54 C. albicans isolates formed smooth, white-to-cream-colored colonies with no chlamydospore production. This medium provides a simple tool for presumptive differentiation of C. dubliniensis from C. albicans. PMID:15472343

Improved method of screening for aflatoxin with a coconut agar medium.

PubMed Central

Davis, N D; Iyer, S K; Diener, U L

1987-01-01

Nine isolates of Aspergillus flavus and Aspergillus parasiticus were screened for aflatoxin production on a coconut extract agar medium. Aflatoxin-producing colonies were detected under long-wave UV light (365 nm) by blue fluorescence on the reverse side after 2 to 5 days of growth. Aflatoxin production was verified by chemical analysis. Several types of shredded coconut available in the United States were tested and found to be satisfactory. No additives were required. Various parameters affecting the test were investigated. PMID:3116928

Identification of non-streptococcal organisms from human dental plaque grown on the Streptococcus-selective medium mitis-salivarius agar.

PubMed

Kim, Yeon-Hee; Lee, Si Young

2015-02-01

Mitis-salivarius (MS) agar has been used widely in microbial epidemiological studies because oral viridans streptococci can be selectively grown on this medium. Even though the previous findings reported the limited selecting power of MS agar for streptococcus strains, the identities of non-streptococcal strains from human oral samples which can grow on this medium are not clear yet. In this study, we identified non-streptococcal organisms grown on MS agar plates by polymerase chain reaction (PCR) amplification and sequencing of the 16S ribosomal RNA (rRNA) gene. Eighty bacterial colonies on MS plates were isolated from plaque samples, and bacterial identification was achieved with the rapid ID 32 Strep system and mini API reader. The bacterial colonies identified as non-streptococci by the API system were selected for further identification. The 16S rRNA gene was amplified by PCR and verified using DNA sequencing analysis for identification. Sequences were compared with those of reference organisms in the genome database of the National Center for Biotechnology Information using the Basic Local Alignment Search Tool (BLAST). Among the 11 isolated non-streptococcal strains on MS plates, 3 strains were identified as Actinomyces naeslundii, 7 strains were identified as Actinomyces oris and 1 strain were identified as Actinomyces sp. using Blastn. In this study, we showed that some oral Actinomyces species can grow on Streptococcus-selective MS agar plates. Copyright © 2014 Elsevier Ltd. All rights reserved.

Gelling agents and culture vessels affect in vitro multiplication of banana plantlets.

PubMed

Kaçar, Y A; Biçen, B; Varol, I; Mendi, Y Y; Serçe, S; Cetiner, S

2010-03-09

Agar is the most commonly used gelling agent in media for plant tissue culture. Because of the high price of tissue-culture-grade agar, attempts have been made to identify suitable alternatives. The type of culture vessel and lid also affects the gaseous composition inside the vessel as well as light penetration. In turn, the vessel affects growth parameters, such as shoot elongation, proliferation and fresh weight, as well as hyperhydric degradation processes. We examined the effects of different culture vessels, including commercial glass jars, magenta boxes, and disposable containers, as well as different gelling agents (agar-agar, Agargel, Phytagel, and plant agar) on the micropropagation of Dwarf Cavendish bananas in an effort to find a combination that yields large numbers of high-quality seedlings. The different culture vessels did not significantly affect seedling culture success. The medium significantly affected shoot weight. Phytagel resulted in the highest shoot weight (overall mean = 2.4 g), while agar, Agargel and plant agar resulted in 1.7, 2.2 and 2.2 g, respectively. Disposable container/Phytagel and Magenta/Agargel combinations yielded the highest shoot weights (2.9 and 3.0 g, respectively). Mean shoot length increased progressively with subculture (four subcultures were made). The highest mean shoot length was obtained with Phytagel and Agargel media (6.4 and 6.3 cm, respectively). Shoot number was significantly affected by medium only at subculture 4. Overall, the highest mean shoot length was obtained with the Magenta/Agargel combination (8.5 cm). Phytagel and plant agar gave higher mean shoot number than agar and Agargel (2.1, 2.1 and 1.7 and 1.9, respectively). The costs of the media and of the culture vessels need to be taken into account for final choice of the banana shoot culture system.

Development of a selective myclobutanil agar (MBA) medium for the isolation of Fusarium species from asparagus fields.

PubMed

Vujanovic, Vladimir; Hamel, Chantal; Jabaji-Hare, Suha; St-Arnaud, Marc

2002-09-01

A new selective myclobutanil agar medium for the detection of Fusarium, species is proposed. Ten media formulations based on various selective agents (pentachloronitrobenzene (PCNB), Rose Bengal, malachite green, sodium hypochlorite, captan, benomyl, chlorotalonil, myclobutanil, thiram, and cupric sulfate) were compared. First, mycelium growth and colony appearance of Alternaria alternata, Aspergillus flavus, Cladosporium cladosporioides, Epicoccum nigrum, Fusarium sp., Fuisarium solani, Fusarium moniliforme, Fusarium oxysporum f.sp. dianthi, Penicillium sp., and Trichoderma viride isolates were compared. Second, the ability of the different media to isolate and enumerate fusaria from asparagus fields was evaluated. The myclobutanil-based medium showed the highest selectivity to Fusarium spp. growth but required a slightly longer incubation time (>5 d) than peptone-pentachloronitrobenzene-based agar (PPA) (< 5 d). PPA allowed a faster fusaria growth but also permited the growth of other moulds. The other media were less selective and did not allow to isolate fusaria or to differenciate them from other growing fungi.

Anticlostridial agent 8-hydroxyquinoline improves the isolation of faecal bifidobacteria on modified Wilkins-Chalgren agar with mupirocin.

PubMed

Novakova, J; Vlkova, E; Salmonova, H; Pechar, R; Rada, V; Kokoska, L

2016-04-01

The need for suitable selective cultivation media for the isolation of Bifidobacterium spp. continues to be a real concern in the field of intestinal microbiology. Isolation of bifidobacteria from human and animal faecal samples using selective agar plating may be problematic especially in samples with increased clostridial counts than bifidobacterial counts. Due to the absence of anticlostridial agents in existing selective media, clostridia can displace bifidobacteria resulting in incorrect estimation of their counts. Therefore, we supplemented the existing selective medium 'modified Wilkins Chalgren agar with mupirocin' (MWM) with 90 mg l(-1) of 8-hydroxyquinoline (8HQ), which was recently proved to act selectively against clostridia. The newly composed 'modified Wilkins-Chalgren agar with 8HQ' (MWMQ) was tested on pure bifidobacterial and clostridial strains, their mixtures, and using faecal samples of mammalian origin; its selectivity was evaluated by genus-specific identification of isolates. The results demonstrated that the presence of 8HQ in this agar eliminated the growth of nonbifidobacterial strains on MWMQ compared to that on MWM, whereas the recovery of bifidobacterial counts was at satisfactory levels. In conclusion, MWMQ could be recommended for bifidobacterial isolation from human and animal faeces especially when bifidobacteria are not numerically dominant and there are chances of clostridial contamination. Routine isolation of bifidobacteria from mammalian faeces does not use a reliable selective agar with an anticlostridial agent. Overgrowth of clostridia may result in incorrect estimation of bifidobacterial counts. Thus, in order to improve the selectivity of existing media for bifidobacterial isolation, we chose the modified Wilkins-Chalgren agar with mupirocin and supplemented it with 8-hydroxyquinoline (8HQ), a molecule that shows anticlostridial activity without affecting the growth of bifidobacteria. This newly composed medium showed enhanced selectivity and specificity compared to the original medium and therefore, can be recommended for the isolation of bifidobacteria from mammal faeces. © 2016 The Society for Applied Microbiology.

Recovery of Sublethally Injured Bacteria Using Selective Agar Overlays.

ERIC Educational Resources Information Center

McKillip, John L.

2001-01-01

This experiment subjects bacteria in a food sample and an environmental sample to conditions of sublethal stress in order to assess the effectiveness of the agar overlay method to recover sublethally injured cells compared to direct plating onto the appropriate selective medium. (SAH)

[Viability of nematophagous fungi Arthrobotrys robusta, Duddingtonia flagrans and Monacrosporium thaumasium after sporulation in different culture media].

PubMed

Maciel, Alessandro S; de Araujo, Jackson V; Campos, Artur K

2006-01-01

Due to the shortage of studies that indicate the culture mediums that optimize the sporulation of namatophagous fungi for use in researche, the sporulation of the fungal isolates A. robusta (I31), D. flagrans (CG768) and M. thaumasium (NF34A) was evaluated in laboratorial conditions for 10 days in the means water-agar 2% (WA 2%), potato-dextrose-agar 2% (PDA 2%), corn-meal-agar 2% (CMA 2%) and yeast-phosphate-sulphate-sucrose-agar (YPSSA). The largest conidia production (P < 0.05) for the isolate CG768 happened in BDA 2% while in the isolates I31 and NF34A produced larger conidia number in YPSSA (P < 0.05). The viability of the conidia to prey infective Ancylostoma spp. larvae did not lose its effectiveness (P < 0.05) independent of the culture medium. The middle of culture did not influence in the viability of the conidia (P > 0.05).

[Presumptive identification of Candida spp. and other clinically important yeasts: usefulness of Brilliance Candida Agar].

PubMed

Alfonso, Claudia; López, Mónica; Arechavala, Alicia; Perrone, María Del Carmen; Guelfand, Liliana; Bianchi, Mario

2010-06-30

Fungal infections caused by yeasts have increased during the last decades and invasive forms represent a serious problem for human health. Candida albicans is the species most frequently isolated from clinical samples. However, other emerging yeast pathogens are increasingly responsible for mycotic infections, and some of them are resistant to some antifungal drugs. Consequently, it is necessary to have methods that can provide a rapid presumptive identification at species level. Numerous chromogenic agar media have been shown to be of value as diagnostic tools. We have compared a chromogenic medium, Brilliance Candida Agar, with CHROMagar Candida, the chromogenic medium most used in our country. A multicentre study was conducted in 16 Hospitals belonging to the Mycology Net of Buenos Aires City Government. A total of 240 yeast isolates were included in this research. The new chromogenic agar showed results very similar to those obtained with CHROMagar Candida. Copyright 2009 Revista Iberoamericana de Micología. Published by Elsevier Espana. All rights reserved.

Studies in the differentiation between Microsporum ferrugineum Ota and Trichophyton soudanense Joyeux.

PubMed

Weitzman, I; Rosenthal, S

1984-02-15

A study, conducted with 20 isolates of Microsporum ferrugineum and 12 isolates of Trichophyton soudanense, revealed that some of the discrepancies in the literature regarding their characteristics and differentiation were due to methodology, strain variation and the use of an insufficient number of isolates. We found all isolates of T. soudanense to be urease negative and gelatinase positive (usually by the first week); to produce brown to black colonies on Lowenstein-Jensen medium; to rapidly decompose casein and more slowly tyrosine; to grow well or better at 37 degrees C as compared to room temperature; to produce reflexive branching on cornmeal Tween agar and abundant microconidia on casero medium and to exhibit no sexual reaction with either mating type of arthroderma simii. All but one isolate demonstrated restricted growth on lactose agar and only three isolates perforated hair. In contrast, we found 18 of 20 isolates of M. ferrugineum to be urease positive in urea broth (most isolates were negative on urea agar); all produced light-colored colonies on Lowenstein-Jensen medium; spreading colonies on lactose agar and failed to perforate hair in vitro or to produce reflexive branching. Most isolates manifested poorer to no growth at 37 degrees C compared to room temperature and all but one failed to decompose casein and tyrosine. A few strains produced macroconidia and/or microconidia on casero medium and some reacted sexually with A. simii (a) (-) mating type. Gelatin hydrolysis was variable.(ABSTRACT TRUNCATED AT 250 WORDS)

New milk medium for germ tube and chlamydoconidia production by Candida albicans.

PubMed

Jitsurong, S; Kiamsiri, S; Pattararangrong, N

1993-08-01

A new medium consisting of UHT milk, tween 80 and agar is described for the development of both germ tube and chlamydoconidia by Candida albicans. In total 172 isolates from clinical specimens, including C. albicans (112), C. guilliermondii (4), C. krusei (3), C. parasilopsis (16). C. tropicalis (28), Torulopsis glabrata (6) and Trichosporon beigellii (3), were examined in this medium by using the standard method. A higher percentage (98.2%) of germ tube production by C. albicans was found in this medium than in undiluted serum (90.2%). In addition, only C. albicans was found to be able to produce a high percentage of chlamydoconidia (95.5%) after 48 hours' incubation. In comparison with the conventional medium, corn meal tween 80 agar (21.4%), this new medium gives a significantly higher percentage and abundance of chlamydoconidia production. Being simple, cheap and easy to prepare, the new milk medium is proposed as very practical in the clinical mycology laboratory.

Selective differential human blood bilayer media for isolation of Gardnerella (Haemophilus) vaginalis.

PubMed Central

Totten, P A; Amsel, R; Hale, J; Piot, P; Holmes, K K

1982-01-01

New selective and differential human blood bilayer agar media with Tween 80 (HBT medium) or without Tween 80 (HB medium), developed for the isolation of Gardnerella (Haemophilus) vaginalis, permitted significantly higher G. vaginalis isolation rates than have been obtained for other media used for this purpose. HB medium consists of a basal layer of Columbia agar base containing colistin and naladixic acid with added amphotericin B and an overlayer of the same composition plus 5% human blood. HBT agar also contains Proteose Peptone No. 3 (Difco Laboratories) and Tween 80 in the basal layer and the overlayer. Both Tween 80 and the bilayer composition enhanced G. vaginalis production of human blood hemolysis, permitting detection of this organism even in the presence of heavy growth of other vaginal flora. The use of HB or HBT medium thus permitted the demonstration that G. vaginalis was present in vaginal fluid from a large percentage (up to 68%) of normal women. However, the concentration of G. vaginalis was found by semiquantitative analysis to be significantly higher in vaginal fluid from women with nonspecific vaginitis than in fluid from normal women. Images PMID:6764766

Media preparation and bacteriological tools.

PubMed

Elbing, Karen; Brent, Roger

2002-08-01

Recipes are provided in this unit for minimal liquid media, rich liquid media, solid media, top agar, and stab agar. Also included are descriptions and useful information about tools used with growth media such as inoculating loops, sterile toothpicks and spreaders.

Borelli's lactritmel agar induces conidiation in rare-macroconidia producing dermatophytic fungi.

PubMed

Ilkit, Macit; Gümral, Ramazan; Döğen, Aylin

2012-10-01

Macroconidia are among the most important indicators used to identify dermatophytic fungi, but several do not usually sporulate and/or produce macroconidia on Sabouraud glucose agar. Specifically, Microsporum audouinii, M. ferrugineum, Trichophyton concentricum, T. schoenleinii, T. verrucosum, and T. violaceum (including T. soudanense and T. yaoundei) rarely form macroconidia and, therefore, cannot be easily identified. In this study, we investigated the production of macroconidia on nine common laboratory media, including Borelli's lactritmel agar (BLA), modified Borelli's lactritmel agar (MBLA), brain heart infusion agar (BHIA), Christensen's urease agar in Petri dishes (UPA), cornmeal dextrose agar (CMDA), Lowenstein-Jensen agar (LJA), malt extract agar (MEA), oatmeal agar (OA), and potato dextrose agar (PDA). The performance of these media was evaluated using 18 rare-macroconidia producing isolates, including representative of the six species mentioned above. All cultures in this study were incubated at 26°C on the bench, and conidia formation on each was investigated at 5, 10, 15, 20, 25, and 30 days of incubation. BLA apparently improved macroconidia production after 15 days and was the most useful nutrient agar medium to induce these phenotypic characters in daily practice, closely followed by OA, PDA, and MBLA.

Comparison of Guizotia abyssinica seed extract (birdseed) agar with conventional media for selective identification of Cryptococcus neoformans in patients with acquired immunodeficiency syndrome.

PubMed Central

Denning, D W; Stevens, D A; Hamilton, J R

1990-01-01

Growth of Cryptococcus neoformans from the sputum of patients with acquired immunodeficiency syndrome may be obscured by oral contamination with Candida albicans on conventional media. We prospectively compared direct plating of sputum and urine onto birdseed agar and compared birdseed agar plating with plating onto Mycosel and Sabouraud dextrose agar cultures. Thirty-two sputum and three urine specimens were compared. C. neoformans was isolated from five specimens. In two specimens, one of sputum and one of urine, C. neoformans was detected only on the birdseed agar plate because of overgrowth on the conventional media by C. albicans. C. neoformans produced dark colonies on birdseed agar, unlike C. albicans, which produces white colonies. The use of birdseed agar as the primary culture medium for sputum and urine specimens from patients with acquired immunodeficiency syndrome increases sensitivity for C. neoformans. Images PMID:2254431

Antibiotic content of selective culture media for isolation of Capnocytophaga species from oral polymicrobial samples.

PubMed

Ehrmann, E; Jolivet-Gougeon, A; Bonnaure-Mallet, M; Fosse, T

2013-10-01

In oral microbiome, because of the abundance of commensal competitive flora, selective media with antibiotics are necessary for the recovery of fastidious Capnocytophaga species. The performances of six culture media (blood agar, chocolate blood agar, VCAT medium, CAPE medium, bacitracin chocolate blood agar and VK medium) were compared with literature data concerning five other media (FAA, LB, TSBV, CapR and TBBP media). To understand variable growth on selective media, the MICs of each antimicrobial agent contained in this different media (colistin, kanamycin, trimethoprim, trimethoprim-sulfamethoxazole, vancomycin, aztreonam and bacitracin) were determined for all Capnocytophaga species. Overall, VCAT medium (Columbia, 10% cooked horse blood, polyvitaminic supplement, 3·75 mg l(-1) of colistin, 1·5 mg l(-1) of trimethoprim, 1 mg l(-1) of vancomycin and 0·5 mg l(-1) of amphotericin B, Oxoid, France) was the more efficient selective medium, with regard to the detection of Capnocytophaga species from oral samples (P < 0·001) and the elimination of commensal clinical species (P < 0·001). The demonstrated superiority of VCAT medium, related to its antibiotic content, made its use indispensable for the optimal isolation of Capnocytophaga species from polymicrobial samples. Isolation of Capnocytophaga species is important for the proper diagnosis and treatment of the systemic infections they cause and for epidemiological studies of periodontal flora. We showed that in pure culture, a simple blood agar allowed the growth of all Capnocytophaga species. Nonetheless, in oral samples, because of the abundance of commensal competitive flora, selective media with antibiotics are necessary for the recovery of Capnocytophaga species. The demonstrated superiority of VCAT medium made its use essential for the optimal detection of this bacterial genus. This work showed that extreme caution should be exercised when reporting the isolation of Capnocytophaga species from oral polymicrobial samples, because the culture medium is a determining factor. © 2013 The Society for Applied Microbiology.

Enhancing Aerobic Growth of Campylobacter in Media Supplemented with Organic Acids

USDA-ARS?s Scientific Manuscript database

The effect of agar and sodium bicarbonate (NaHCO3) concentration on aerobic growth of Campylobacter in was determined. A fumarate-pyruvate medium was supplemented with 0.0 to 0.2% agar and inoculated with Campylobacter coli, Campylobacter fetus, or Campylobacter jejuni. Portions of the inoculated me...

Spoilage potential of Vagococcus salmoninarum in preservative-free, MAP-stored brown shrimp and differentiation from Brochothrix thermosphacta on streptomycin thallous acetate actidione agar.

PubMed

Calliauw, F; Horemans, B; Broekaert, K; Michiels, C; Heyndrickx, M

2016-05-01

During a previous study concerning brown shrimp (Crangon crangon), selective streptomycin thallous acetate actidione (STAA) agar was used to determine the growth of Brochothrix thermosphacta. However, the growth of Vagococcus salmoninarum on this medium was also noticed. This study explores the spoilage potential of this organism when inoculated on sterile shrimp. Isolates growing on STAA were identified using (GTG)5 clustering followed by partial 16S rRNA gene sequence analysis. Their biochemical spoilage potential was analysed for H2 S production and enzymatic activities were tested using an APIZYM test. Headspace solid phase micro-extraction (SPME) and gas chromatography-mass spectrometry (GC-MS) were used to analyse the volatile organic compounds (VOCs) produced during storage of inoculated shrimp. Fifty-five per cent of isolates taken from STAA could be identified as V. salmoninarum, while no apparent morphological difference with B. thermosphacta isolates was identified upon the prescribed incubation conditions. For isolates identified as V. salmoninarum, production of 2-heptanone, 2-nonanone, 2-undecanone was found, as was the possibility to form H2 S. When using the STAA medium for detecting B. thermosphacta, one should consider the possible abundant presence of V. salmoninarum as well. Based on this study, V. salmoninarum does not exhibit great spoilage potential, although it can produce H2 S and formed VOCs which are also found in other spoiled seafood products. © 2016 The Society for Applied Microbiology.

Optimisation of a direct plating method for the detection and enumeration of Alicyclobacillus acidoterrestris spores.

PubMed

Henczka, Marek; Djas, Małgorzata; Filipek, Katarzyna

2013-01-01

A direct plating method for the detection and enumeration of Alicyclobacillus acidoterrestris spores has been optimised. The results of the application of four types of growth media (BAT agar, YSG agar, K agar and SK agar) regarding the recovery and enumeration of A. acidoterrestris spores were compared. The influence of the type of applied growth medium, heat shock conditions, incubation temperature, incubation time, plating technique and the presence of apple juice in the sample on the accuracy of the detection and enumeration of A. acidoterrestris spores was investigated. Among the investigated media, YSG agar was the most sensitive medium, and its application resulted in the highest recovery of A. acidoterrestris spores, while K agar and BAT agar were the least suitable media. The effect of the heat shock time on the recovery of spores was negligible. When there was a low concentration of spores in a sample, the membrane filtration method was superior to the spread plating method. The obtained results show that heat shock carried out at 80°C for 10 min and plating samples in combination with membrane filtration on YSG agar, followed by incubation at 46°C for 3 days provided the optimal conditions for the detection and enumeration of A. acidoterrestris spores. Application of the presented method allows highly efficient, fast and sensitive identification and enumeration of A. acidoterrestris spores in food products. This methodology will be useful for the fruit juice industry for identifying products contaminated with A. acidoterrestris spores, and its practical application may prevent economic losses for manufacturers. Copyright © 2012 Elsevier B.V. All rights reserved.

Evaluation of culture media for counts of Bifidobacterium animalis subsp. lactis Bb 12in yoghurt after refrigerated storage.

PubMed

Fachin, Luciano; Moryia, Juliana; Neves Gândara, Ana Lourdes; Viotto, Walkiria Hanada

2008-04-01

The agar RCPB pH5 has been considered a good alternative for counts of Bifidobacterium in yoghurt. However, during the refrigerated storage of yoghurt it is extremely difficult to count this microorganism due to the size of the colonies, which are so small they require the aid of a stereoscope to count them. Another agar, MRS-LP, has been also recommended for counts of Bifidobacterium in the presence of yoghurt bacteria. This study evaluated the supplementation of RCPB pH5 agar with dehydrated liver extract and the salts KH2PO4, K2HPO4, FeSO47H2O, MnSO4H2O and MgSO47H2O, aiming at improving the differentiation of Bifidobacterium in yoghurt after refrigerated storage, and also evaluated the selective count of Bifidobacterium in yoghurt using the agar MRS-LP. The agar MRS-LP presented the same cell recovery as non-fortified RCPB pH5 agar, used as a standard medium, thus being considered a good option for counts of Bifidobacterium in yoghurt. The fortified RCPB pH5 also presented the same recovery as the standard RCPB pH5 medium, however, the addition of dehydrated liver extract to the RCPB pH5 agar considerably increased the size of the Bifidobacterium colonies after refrigerated storage, making differentiation of the colonies much easier and reliable when compared to the standard non-fortified RPCP pH5. The addition of the salts (KH2PO4, K2HPO4, FeSO47H2O, MnSO4H2O and MgSO47H2O) had no influence on the performance of the RCPB pH5 agar.

Control of the pattern of perithecium development in Sordaria fimicola on agar medium.

PubMed

Pollock, R T

1975-06-01

In a Sordaria fimicola (Rob.) Ces. and de Not. colony grown on agar medium in a petri plate, perithecia developed in a narrow band around the plate edge after the colony margin reached the edge. Physical wounding of the colony carried out shortly before or during the time perithecia were developing around the plate edge stimulated perithecium development in the wound area. Diffusion barriers were created by cutting small trenches in the agar parallel to the plate edge. The trenches were made at several different positions between the plate center and edge using cultures of several different ages, and the resultant distribution of perithecia along the trench edges suggested that the colony center and periphery produce diffusible inhibitors of perithecium development. These inhibitors may be responsible, in part, for the observed pattern of perithecium development in the colony.

Simple low cost differentiation of Candida auris from Candida haemulonii complex using CHROMagar Candida medium supplemented with Pal's medium.

PubMed

Kumar, Anil; Sachu, Arun; Mohan, Karthika; Vinod, Vivek; Dinesh, Kavitha; Karim, Shamsul

Candida auris is unique due to its multidrug resistance and misidentification as Candida haemulonii by commercial systems. Its correct identification is important to avoid inappropriate treatments. To develop a cheap method for differentiating C. auris from isolates identified as C. haemulonii by VITEK2. Fifteen C. auris isolates, six isolates each of C. haemulonii and Candida duobushaemulonii, and one isolate of Candida haemulonii var. vulnera were tested using CHROMagar Candida medium supplemented with Pal's agar for better differentiation. On CHROMagar Candida medium supplemented with Pal's agar all C. auris strains showed confluent growth of white to cream colored smooth colonies at 37°C and 42°C after 24 and 48h incubation and did not produce pseudohyphae. The isolates of the C. haemulonii complex, on the contrary, showed poor growth of smooth, light-pink colonies at 24h while at 48h the growth was semiconfluent with the production of pseudohyphae. C. haemulonii complex failed to grow at 42°C. We report a rapid and cheap method using CHROMagar Candida medium supplemented with Pal's agar for differentiating C. auris from isolates identified as C. haemulonii by VITEK2. Copyright © 2017 Asociación Española de Micología. Publicado por Elsevier España, S.L.U. All rights reserved.

Powdered Chitin Agar as a Selective Medium for Enumeration of Actinomycetes in Water and Soil1

PubMed Central

Hsu, S. C.; Lockwood, J. L.

1975-01-01

Agar media made with 0.4% colloidal chitin plus mineral salts and adjusted to pH 8.0 was superior to four other commonly used media for the isolation and enumeration of actinomycetes from water samples. More actinomycetes developed on chitin agar, and the development of bacteria and fungi was suppressed. Frozen and vacuum-dried chitin from aqueous colloidal suspensions was finely divided and gave results comparable to those obtained with media prepared from colloidal suspensions. Images PMID:234719

Establishing axenic cultures from mature pecan embryo explants on media with low water availability.

PubMed

Obeidy, A A; Smith, M A

1990-12-01

Endophytic fungi associated with mature pecan (Carya illinoensis (Wangenh.) C. Koch) nuts prevented successful, contaminant-free in vitro culture of embryo expiants, even after rigorous surface disinfestation of the nuts and careful aseptic shelling. Disinfestation with sodium hypochlorite after shell removal was also unsuccessful, because even dilute concentrations which were ineffective against the fungal contaminants prevented subsequent growth from the embryo. Explanting media with low water availability which would not sustain growth of fungal contaminants, but supported growth from mature pecan embryos, were developed as an alternative disinfestation method. The explanting media were supplemented with 0.9-1.5% agar, and other media components were selectively omitted to test their influence on water availability and fungal growth. Disinfestation of up to 65% of the cultures was accomplished, depending on the medium formulation, compared to 100% loss to contamination on control medium (0.5% agar). A complete medium (containing sucrose, salts, vitamins, 18 μM BAP, and 5 μM IBA) with 1.5% agar provided control of contamination, and encouraged subsequent regeneration from the embryo expiants, which remained free of contaminant growth through subsequent subcultures.

New culture medium for the presumptive identificaion of Candida albicans and Cryptococcus neoformans.

PubMed Central

Fleming, W H; Hopkins, J M; Land, G A

1977-01-01

A new medium composed of Tween 80, oxgall, caffeic acid, and Davis agar (TOC) that provides for the rapid presumptive identification of Candida albicans and Cryptococcus neoformans is described herein. C. albicans is differentiated from other yeasts by the sequential production of germ tubes and chlamydospores. In a comparison with cormeal agar control plates, there was an increase of chlamydospore-forming strains of C. albicans (97.1% versus 87.2%) and a decrease in the time required for chlamydospore formation (24 h versus 48 h). C. neoformans produced a brown pigment of TOC, which is specific for its identification, thus differentiating it from the other yeasts. A comparison of 24-h pigment production by C. neoformans on TOC with that of birdseed agar showed a dark, coffee brown color in the former cultures and a light brown color in the latter. The change in pigmentation of C. neoformans, as well as morphological changes in C. albicans, can be induced within 3 to 12 h and in not more than 24 h on the TOC medium. Images PMID:321472

Individual based simulations of bacterial growth on agar plates

NASA Astrophysics Data System (ADS)

Ginovart, M.; López, D.; Valls, J.; Silbert, M.

2002-03-01

The individual based simulator, INDividual DIScrete SIMulations (INDISIM) has been used to study the behaviour of the growth of bacterial colonies on a finite dish. The simulations reproduce the qualitative trends of pattern formation that appear during the growth of Bacillus subtilis on an agar plate under different initial conditions of nutrient peptone concentration, the amount of agar on the plate, and the temperature. The simulations are carried out by imposing closed boundary conditions on a square lattice divided into square spatial cells. The simulator studies the temporal evolution of the bacterial population possible by setting rules of behaviour for each bacterium, such as its uptake, metabolism and reproduction, as well as rules for the medium in which the bacterial cells grow, such as concentration of nutrient particles and their diffusion. The determining factors that characterize the structure of the bacterial colony patterns in the presents simulations, are the initial concentrations of nutrient particles, that mimic the amount of peptone in the experiments, and the set of values for the microscopic diffusion parameter related, in the experiments, to the amount of the agar medium.

Characterization of the species Malassezia pachydermatis and re-evaluation of its lipid dependence using a synthetic agar medium

PubMed Central

Puig, Laura; Castellá, Gemma

2017-01-01

The genus Malassezia includes lipophilic yeasts, which are part of the skin microbiota of various mammals and birds. Unlike the rest of Malassezia species, M. pachydermatis is described as non-lipid-dependent, as it is able to grow on Sabouraud glucose agar (SGA) without lipid supplementation. In this study we have examined the phenotypic variability within M. pachydermatis and confirmed its lipid-dependent nature using a synthetic agar medium. We used a selection of representative non-lipid-dependent strains from different animal species and three atypical lipid-dependent strains of this species, which were not able to grow after multiple passages on SGA. More than 400 lipid-dependent Malassezia isolates from animals were studied in order to detect the three lipid-dependent strains of M. pachydermatis. The identity of the atypical strains was confirmed by DNA sequencing. On the other hand, we have modified the Tween diffusion test, which is widely used in the characterization of these yeasts, by using a synthetic agar-based medium instead of SGA. This modification has proved to be useful for differentiation of M. pachydermatis strains, providing reproducible results and a straightforward interpretation. The finding of these peculiar lipid-dependent strains exemplifies the large variability within the species M. pachydermatis, which involves rare atypical strains with particular growth requirements. PMID:28586389

Characterization of the species Malassezia pachydermatis and re-evaluation of its lipid dependence using a synthetic agar medium.

PubMed

Puig, Laura; Bragulat, M Rosa; Castellá, Gemma; Cabañes, F Javier

2017-01-01

The genus Malassezia includes lipophilic yeasts, which are part of the skin microbiota of various mammals and birds. Unlike the rest of Malassezia species, M. pachydermatis is described as non-lipid-dependent, as it is able to grow on Sabouraud glucose agar (SGA) without lipid supplementation. In this study we have examined the phenotypic variability within M. pachydermatis and confirmed its lipid-dependent nature using a synthetic agar medium. We used a selection of representative non-lipid-dependent strains from different animal species and three atypical lipid-dependent strains of this species, which were not able to grow after multiple passages on SGA. More than 400 lipid-dependent Malassezia isolates from animals were studied in order to detect the three lipid-dependent strains of M. pachydermatis. The identity of the atypical strains was confirmed by DNA sequencing. On the other hand, we have modified the Tween diffusion test, which is widely used in the characterization of these yeasts, by using a synthetic agar-based medium instead of SGA. This modification has proved to be useful for differentiation of M. pachydermatis strains, providing reproducible results and a straightforward interpretation. The finding of these peculiar lipid-dependent strains exemplifies the large variability within the species M. pachydermatis, which involves rare atypical strains with particular growth requirements.

Identification and discrimination of Pseudomonas aeruginosa bacteria grown in blood and bile by laser-induced breakdown spectroscopy

NASA Astrophysics Data System (ADS)

Rehse, Steven J.; Diedrich, Jonathan; Palchaudhuri, Sunil

2007-10-01

Pseudomonas aeruginosa bacteria colonies have been analyzed by laser-induced breakdown spectroscopy using nanosecond laser pulses. LIBS spectra were obtained after transferring the bacteria from a nutrient-rich culture medium to a nutrient-free agar plate for laser ablation. To study the dependence of the LIBS spectrum on growth and environmental conditions, colonies were cultured on three different nutrient media: a trypticase soy agar (TSA) plate, a blood agar plate, and a medium chosen deliberately to induce bacteria membrane changes, a MacConkey agar plate containing bile salts. Nineteen atomic and ionic emission lines in the LIBS spectrum, which was dominated by inorganic elements such as calcium, magnesium and sodium, were used to identify and classify the bacteria. A discriminant function analysis was used to discriminate between the P. aeruginosa bacteria and two strains of E. coli: a non-pathogenic environmental strain and the pathogenic strain enterohemorrhagic E. coli 0157:H7 (EHEC). Nearly identical spectra were obtained from P. aeruginosa grown on the TSA plate and the blood agar plate, while the bacteria grown on the MacConkey plate exhibited easily distinguishable differences from the other two. All P. aeruginosa samples, independent of initial growth conditions, were readily discriminated from the two E. coli strains.

Comparative analysis of cryopreservation methods in Chlamydomonas reinhardtii.

PubMed

Scarbrough, Chasity; Wirschell, Maureen

2016-10-01

Chlamydomonas is a model organism used for studies of many important biological processes. Traditionally, strains have been propagated on solid agar, which requires routine passaging for long-term maintenance. Cryopreservation of Chlamydomonas is possible, yet long-term viability is highly variable. Thus, improved cryopreservation methods for Chlamydomonas are an important requirement for sustained study of genetically defined strains. Here, we tested a commercial cryopreservation kit and directly compared it's effectiveness to a methanol-based method. We also tested thaw-back procedures comparing the growth of cells in liquid culture or on solid agar media. We demonstrated that methanol was the superior cryopreservation method for Chlamydomonas compared to the commercial kit and that post-thaw culture conditions dramatically affect viability. We also demonstrated that cryopreserved cells could be successfully thawed and plated directly onto solid agar plates. Our findings have important implications for the long-term storage of Chlamydomonas that can likely be extended to other algal species. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Saccharopolyspora Species: Laboratory Maintenance and Enhanced Production of Secondary Metabolites.

PubMed

Dhakal, Dipesh; Pokhrel, Anaya Raj; Jha, Amit Kumar; Thuan, Nguyen Huy; Sohng, Jae Kyung

2017-02-06

Saccharopolyspora spp. are aerobic, Gram-positive, non-acid-fast, and non-motile actinomycetes. Various species of the genus Saccharopolyspora have been reported with an ability to produce various bioactive compounds for pharmaceutical and agricultural uses. This unit includes general protocols for the laboratory maintenance of Saccharopolyspora species, including growth in liquid medium, growth on solid agar, long-term storage, and generation of a higher producer strain by mutagenesis. Saccharopolyspora spinosa ATCC 49460 is used as a prototype for explaining the considerations for efficient laboratory maintenance of Saccharopolyspora spp. Saccharopolyspora spinosa is a producer of spinosad, a prominent insecticide with selective activity against various insects. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley & Sons, Inc.

[Correlations between the characters of the mycelium vegetative growth and the formation of the fruiting body of Ganoderma luciderm].

PubMed

Wang, Shu-fang; He, Xiu-feng; He, Hui-xia; Zhu, Ping

2006-01-01

To select a proper Ganoderma luciderm strain for the fruiting body production. The strains were cultivated on the agar media and in the liquid media, respectively. Then the strains were inoculated onto the solid medium made from agricultural products (such as wheat bran, corn powder, wood meal, etc.) and cultured for a certain period. Strains, which were easier to produce polyporic tissues at the vegetative growth stage, would be more quickly to form fruiting body with high quality and yield of the spores. Appearance of the polyporic tissues at the mycelium vegetative growth stage could be used as a marker for the strain selection for the G. luciderm substituted cultivation.

Asymptomatic bacteriuria, bacteremia, and other infections due to NSU corynebacteria.

PubMed

Furness, G; Kaminski, Z

1975-11-01

By means of the new medium, nonspecific urethritis (NSU) chocolate agar, NSU corymebacteria were isolated from patients with asymptomatic bacteriuria, bacteremia, cervicitis, conjuctivitis, and pericarditis, and also with bone marrow, wound, and cul-de-sac infections. The NSU corynebacteria were considered the etiologic agents. On the basis of biochemical reactions, antibiotic sensitivity, and complement fixation tests some isolates were the same microorganisms. Both patients with conjunctivitis were infected with the same NSU corynebacteria. A second isolate was cultured from patients with osteomyelitis and cervicitis, while a third was recovered from an infected leg wound and from a patient with pericarditis. Seven of the isolates, when injected into rabbits hypersensitive to four NSU corynebacteria isolated from the inflamed epididymis of patients with epididymitis, elicited delayed hypersensitivity reactions, which indicated that they also were related antigenically. It is suggested that nonspecific urethritis and eididymitis may represent an infection with NSU corynebacteria, or may be an extension of bacteriuria due to these microorganisms, with a delayed hypersensitivity reaction as a possible additional complication. Colony counts on NSU chocolate agar of the bacteria in urines from male and female patients were higher than those obtained on conventional agar media. NSU chocolate agar is superior to other agar media for the isolation of pathogenic and saprophytic bacteria not only from the urogenital tract but also from other foci of infection. It is easily prepared from commercial blood agar plates and its use should be considered when a selective medium is not required.

[FEATURES OF MASS-SPECTROMETRIC PROTEIN PROFILES OF STRAINS OF BRUCELLOSIS CAUSATIVE AGENT DURING PREPARATION OF CULTURE ON VARIOUS NUTRIENT MEDIA].

PubMed

Ulshina, D V; Kovalev, D A; Zhirov, A M; Zharinova, N V; Khudoleev, A A; Kogotkova, O I; Efremenko, V I; Evchenko, N I; Kulichenko, A N

2016-01-01

Carry out comparative analysis using time-of-flight mass-spectrometry with matrix laser desorption/ionization (MALDI-TOF MS) of protein profiles of brucellosis causative agents (Brucella melitensis Rev-1 and Brucella abortus 19BA), cultivated in various nutrient media: Albimi agar, brucellagar and erythrit-agar. Vaccine,strains: Brucella melitensis Rev-1 and Brucella abortus 19BA. Protein profiling in linear mode on Microflex "Bruker Daltonics" MALDI-TOF mass-spectrometer. A number of characteristic features of brucella mass-spectra was detected: in particular, preservation of the total qualitative composition of protein profiles of cultures and significant differences in the intensity of separate peaks depending on the nutrient medium used. Based on the analysis of the data obtained, use of Albimi agar as the nutrient medium for preparation of brucella culture samples for mass-spectrometric analysis was shown to be optimal.

An evaluation of strontium chloride, Rappaport and strontium selenite enrichment for the isolation of salmonellas from man, animals, meat products and abbattoir effluents

PubMed Central

Iveson, J. B.; Mackay-Scollay, E. M.

1972-01-01

Strontium chloride enrichment broth was found to be comparable to Rappaport broth for the recovery of a wide range of Salmonella serotypes from man, animals, meat products and effluents. With the exception of cloacal samples from reptiles, both procedures were superior to selenite F. The performance of strontium chloride Mand selenite F enrichment was improved when effluent samples were incubated at 43° C. Strontium chloride M and Rappaport enrichment were superior to selenite F for the isolation of Arizona species from reptiles. Strontium chloride B, strontium selenite and Rappaport broths were found suitable for the isolation of multiple Salmonella serotypes from sea water contaminated with abattoir effluents. The strontium chloride B and strontium selenite enrichment media were superior to Rappaport broth when samples were incubated at 43° C. Modified bismuth sulphite agar was found superior to Salmonella—Shigella agar as a solid subculture medium. The investigation of a food poisoning outbreak due to Salmonella typhimurium phage type 21 is reported. The significance of the choice of sampling and isolation techniques in salmonellosis in man and animals is discussed. PMID:4503874

A serum-free medium for colony growth and hyaluronic acid production by Streptococcus zooepidemicus NJUST01.

PubMed

Zhang, Jianfa; Ding, Xia; Yang, Liuyan; Kong, Zhiming

2006-08-01

A hyaluronic acid (HA)-producing strain, Streptococcus zooepidemicus NJUST01, can grow in a serum-free agar medium, with starch as exclusive carbon source, but not glucose, sucrose, dextrine, xylose, or lactose. In this starch medium, the strain NJUST01 reproduced successively at 37 degrees C for 60 generations, with no obvious variation on morphology and physiology, but colonies of the strain after 60th generation could not produce a clear hemolytic zone on sheep blood agar plates. Hyaluronic acid production by the strain NJUST01 was analyzed relative to the starch medium. Employing a multifactor cross experiment, an optimum medium revealed for hyaluronic acid production was composed of 5% starch, 0.3% glucose, 0.5% peptone, 0.15% MgSO4, and 2.0% K2HPO4. The amount of HA 6.7 g/l was obtained in batch fermentation on a rotary shaker at 37 degrees C, 220 rpm for 36 h.

Evaluation of a chromogenic culture medium for isolation of Clostridium difficile within 24 hours.

PubMed

Perry, John D; Asir, Kerry; Halimi, Diane; Orenga, Sylvain; Dale, Joanne; Payne, Michelle; Carlton, Ruth; Evans, Jim; Gould, F Kate

2010-11-01

Rapid and effective methods for the isolation of Clostridium difficile from stool samples are desirable to obtain isolates for typing or to facilitate accurate diagnosis of C. difficile-associated diarrhea. We report on the evaluation of a prototype chromogenic medium (ID C. difficile prototype [IDCd]) for isolation of C. difficile. The chromogenic medium was compared using (i) 368 untreated stool samples that were also inoculated onto CLO medium, (ii) 339 stool samples that were subjected to alcohol shock and also inoculated onto five distinct selective agars, and (iii) standardized suspensions of 10 C. difficile ribotypes (untreated and alcohol treated) that were also inoculated onto five distinct selective agars. Two hundred thirty-six isolates of C. difficile were recovered from 368 untreated stool samples, and all but 1 of these strains (99.6%) were recovered on IDCd within 24 h, whereas 74.6% of isolates were recovered on CLO medium after 48 h. Of 339 alcohol-treated stool samples cultured onto IDCd and five other selective agars, C. difficile was recovered from 218 samples using a combination of all media. The use of IDCd allowed recovery of 96.3% of isolates within 24 h, whereas 51 to 83% of isolates were recovered within 24 h using the five other media. Finally, when they were challenged with pure cultures, all 10 ribotypes of C. difficile generated higher colony counts on IDCd irrespective of alcohol pretreatment or duration of incubation. We conclude that IDCd is an effective medium for isolation of C. difficile from stool samples within 24 h.

Evaluation of a Chromogenic Culture Medium for Isolation of Clostridium difficile within 24 Hours ▿

PubMed Central

Perry, John D.; Asir, Kerry; Halimi, Diane; Orenga, Sylvain; Dale, Joanne; Payne, Michelle; Carlton, Ruth; Evans, Jim; Gould, F. Kate

2010-01-01

Rapid and effective methods for the isolation of Clostridium difficile from stool samples are desirable to obtain isolates for typing or to facilitate accurate diagnosis of C. difficile-associated diarrhea. We report on the evaluation of a prototype chromogenic medium (ID C. difficile prototype [IDCd]) for isolation of C. difficile. The chromogenic medium was compared using (i) 368 untreated stool samples that were also inoculated onto CLO medium, (ii) 339 stool samples that were subjected to alcohol shock and also inoculated onto five distinct selective agars, and (iii) standardized suspensions of 10 C. difficile ribotypes (untreated and alcohol treated) that were also inoculated onto five distinct selective agars. Two hundred thirty-six isolates of C. difficile were recovered from 368 untreated stool samples, and all but 1 of these strains (99.6%) were recovered on IDCd within 24 h, whereas 74.6% of isolates were recovered on CLO medium after 48 h. Of 339 alcohol-treated stool samples cultured onto IDCd and five other selective agars, C. difficile was recovered from 218 samples using a combination of all media. The use of IDCd allowed recovery of 96.3% of isolates within 24 h, whereas 51 to 83% of isolates were recovered within 24 h using the five other media. Finally, when they were challenged with pure cultures, all 10 ribotypes of C. difficile generated higher colony counts on IDCd irrespective of alcohol pretreatment or duration of incubation. We conclude that IDCd is an effective medium for isolation of C. difficile from stool samples within 24 h. PMID:20739493

Plant-based culture media: Efficiently support culturing rhizobacteria and correctly mirror their in-situ diversity.

PubMed

Youssef, Hanan H; Hamza, Mervat A; Fayez, Mohamed; Mourad, Elhussein F; Saleh, Mohamed Y; Sarhan, Mohamed S; Suker, Ragab M; Eltahlawy, Asmaa A; Nemr, Rahma A; El-Tahan, Mahmod; Ruppel, Silke; Hegazi, Nabil A

2016-03-01

Our previous publications and the data presented here provide evidences on the ability of plant-based culture media to optimize the cultivability of rhizobacteria and to support their recovery from plant-soil environments. Compared to the tested chemically-synthetic culture media (e.g. nutrient agar and N-deficient combined-carbon sources media), slurry homogenates, crude saps, juices and powders of cactus (Opuntia ficus-indica) and succulent plants (Aloe vera and Aloe arborescens) were rich enough to support growth of rhizobacteria. Representative isolates of Enterobacter spp., Klebsiella spp., Bacillus spp. and Azospirillum spp. exhibited good growth on agar plates of such plant-based culture media. Cell growth and biomass production in liquid batch cultures were comparable to those reported with the synthetic culture media. In addition, the tested plant-based culture media efficiently recovered populations of rhizobacteria associated to plant roots. Culturable populations of >10(6)-10(8) cfu g(-1) were recovered from the ecto- and endo-rhizospheres of tested host plants. More than 100 endophytic culture-dependent isolates were secured and subjected to morphophysiological identification. Factor and cluster analyses indicated the unique community structure, on species, genera, class and phyla levels, of the culturable population recovered with plant-based culture media, being distinct from that obtained with the chemically-synthetic culture media. Proteobacteria were the dominant (78.8%) on plant-based agar culture medium compared to only 31% on nutrient agar, while Firmicutes prevailed on nutrient agar (69%) compared to the plant-based agar culture media (18.2%). Bacteroidetes, represented by Chryseobacterium indologenes, was only reported (3%) among the culturable rhizobacteria community of the plant-based agar culture medium.

Plant-based culture media: Efficiently support culturing rhizobacteria and correctly mirror their in-situ diversity

PubMed Central

Youssef, Hanan H.; Hamza, Mervat A.; Fayez, Mohamed; Mourad, Elhussein F.; Saleh, Mohamed Y.; Sarhan, Mohamed S.; Suker, Ragab M.; Eltahlawy, Asmaa A.; Nemr, Rahma A.; El-Tahan, Mahmod; Ruppel, Silke; Hegazi, Nabil A.

2015-01-01

Our previous publications and the data presented here provide evidences on the ability of plant-based culture media to optimize the cultivability of rhizobacteria and to support their recovery from plant-soil environments. Compared to the tested chemically-synthetic culture media (e.g. nutrient agar and N-deficient combined-carbon sources media), slurry homogenates, crude saps, juices and powders of cactus (Opuntia ficus-indica) and succulent plants (Aloe vera and Aloe arborescens) were rich enough to support growth of rhizobacteria. Representative isolates of Enterobacter spp., Klebsiella spp., Bacillus spp. and Azospirillum spp. exhibited good growth on agar plates of such plant-based culture media. Cell growth and biomass production in liquid batch cultures were comparable to those reported with the synthetic culture media. In addition, the tested plant-based culture media efficiently recovered populations of rhizobacteria associated to plant roots. Culturable populations of >106–108 cfu g−1 were recovered from the ecto- and endo-rhizospheres of tested host plants. More than 100 endophytic culture-dependent isolates were secured and subjected to morphophysiological identification. Factor and cluster analyses indicated the unique community structure, on species, genera, class and phyla levels, of the culturable population recovered with plant-based culture media, being distinct from that obtained with the chemically-synthetic culture media. Proteobacteria were the dominant (78.8%) on plant-based agar culture medium compared to only 31% on nutrient agar, while Firmicutes prevailed on nutrient agar (69%) compared to the plant-based agar culture media (18.2%). Bacteroidetes, represented by Chryseobacterium indologenes, was only reported (3%) among the culturable rhizobacteria community of the plant-based agar culture medium. PMID:26966571

Performance of the EUCAST Disk Diffusion Method, the CLSI Agar Screen Method, and the Vitek 2 Automated Antimicrobial Susceptibility Testing System for Detection of Clinical Isolates of Enterococci with Low- and Medium-Level VanB-Type Vancomycin Resistance: a Multicenter Study

PubMed Central

Giske, Christian G.; Haldorsen, Bjørg; Matuschek, Erika; Schønning, Kristian; Leegaard, Truls M.; Kahlmeter, Gunnar

2014-01-01

Different antimicrobial susceptibility testing methods to detect low-level vancomycin resistance in enterococci were evaluated in a Scandinavian multicenter study (n = 28). A phenotypically and genotypically well-characterized diverse collection of Enterococcus faecalis (n = 12) and Enterococcus faecium (n = 18) strains with and without nonsusceptibility to vancomycin was examined blindly in Danish (n = 5), Norwegian (n = 13), and Swedish (n = 10) laboratories using the EUCAST disk diffusion method (n = 28) and the CLSI agar screen (n = 18) or the Vitek 2 system (bioMérieux) (n = 5). The EUCAST disk diffusion method (very major error [VME] rate, 7.0%; sensitivity, 0.93; major error [ME] rate, 2.4%; specificity, 0.98) and CLSI agar screen (VME rate, 6.6%; sensitivity, 0.93; ME rate, 5.6%; specificity, 0.94) performed significantly better (P = 0.02) than the Vitek 2 system (VME rate, 13%; sensitivity, 0.87; ME rate, 0%; specificity, 1). The performance of the EUCAST disk diffusion method was challenged by differences in vancomycin inhibition zone sizes as well as the experience of the personnel in interpreting fuzzy zone edges as an indication of vancomycin resistance. Laboratories using Oxoid agar (P < 0.0001) or Merck Mueller-Hinton (MH) agar (P = 0.027) for the disk diffusion assay performed significantly better than did laboratories using BBL MH II medium. Laboratories using Difco brain heart infusion (BHI) agar for the CLSI agar screen performed significantly better (P = 0.017) than did those using Oxoid BHI agar. In conclusion, both the EUCAST disk diffusion and CLSI agar screening methods performed acceptably (sensitivity, 0.93; specificity, 0.94 to 0.98) in the detection of VanB-type vancomycin-resistant enterococci with low-level resistance. Importantly, use of the CLSI agar screen requires careful monitoring of the vancomycin concentration in the plates. Moreover, disk diffusion methodology requires that personnel be trained in interpreting zone edges. PMID:24599985

Evaluation of antifungal activity of free fatty acids methyl esters fraction isolated from Algerian Linum usitatissimum L. seeds against toxigenic Aspergillus

PubMed Central

Abdelillah, Amrouche; Houcine, Benmehdi; Halima, Dalile; Meriem, Chabane sari; Imane, Zaaboub; Eddine, Smahi Djamal; Abdallah, Moussaoui; Daoudi, Chabane sari

2013-01-01

Objective The aim of this study was to evaluate the antifungal activity of the major fraction of fatty acids methyl esters (FAMEs) isolated from Linum usitatissimum L. seeds oil collected from Bechar department (Algeria). Methods The assessment of antifungal activity was carried out in terms of percentage of radial growth on solid medium (potatoes dextrose agar PDA) and biomass growth inhibition on liquid medium (potatoes dextrose broth PDB) against two fungi. Results The FAMEs was found to be effective in inhibiting the radial mycelial growth of Aspergillus flavus more than Aspergillus ochraceus on all tested concentrations. The highest antifungal index was found to be (54.19%) compared to Aspergillus ochraceus (40.48%). The results of the antifungal activity of the FAMEs inhibition of biomass on liquid medium gave no discounted results, but this does not exclude the antifungal activity. Conclusions We can assume that the observed antifungal potency may be due to the abundance of linoleic and α-linolenic acids in linseed oil which appears to be promising to treat fungal infections, storage fungi and food spoilage in food industry field. PMID:23730556

[Investigation of some virulence factors in Trichosporon spp. strains].

PubMed

Demir, Feyza; Kuştimur, Semra

2014-10-01

The frequency of fungal infections have increased recently in parallel to prolonged survival of patients with chronical infections, common use of the broad-spectrum antibiotics and cytotoxic drugs and surgical interventions. Fungi such as Trichosporon, Fusarium and Geotrichum that were previously evaluated as contaminant/colonization, become important causes of morbidity and mortality especially in neutropenic patients. The aim of this study was to investigate the presence of virulence factors such as acid proteinase, phospholipase, esterase, coagulase and hemolytic activity among Trichosporon species. A total of 40 Trichosporon strains, of them 24 (60%) were T.asahii, 6 (15%) were T.inkin and 10 (25%) were the other species (one of each of T.aquatile, T.asteroides, T.coremiiforme, T.cutaneum, T.dermatis, T.faecale, T.japonicum, T.montevideense, T.mucoides, T.ovoides) were included in the study. Identification of the isolates was performed according to microscopic morphology (blastospores, arthrospores, pseudohyphae and true hyphae) on corn meal agar media, and carbohydrate assimilation patterns (API ID32C; bioMérieux, France). Secretory acid proteinase, phospholipase and esterase activities of the strains were evaluated by 1% bovine serum albumin containing agar, by egg yolk containing solid medium, and by Tween 80 containing solid medium, respectively. Hemolytic activity of the isolates were evaluated by 5-10% sheep blood Sabouraud dextrose agar. Coagulase enzyme activity was determined by using human and rabbit plasma. In our study, all of the 40 Trichosporon spp. strains were found negative in terms of acid proteinase and phospholipase enzyme activity, however all were positive for esterase enzyme activity. Hemolytic enzyme activity were identified in a total of 15 (37.5%) strains, being "+++" in three strains (2 T.asahii, 1 T.japonicum), and "++" in 12 isolates (9 T.asahii, 1 T.inkin, 1 T.asteroides, 1 T.mentevideense). Although 11 of those 15 positive strains were T.asahii, there was no statistical difference between the species in terms of hemolytic enzyme activity (p> 0.05). Coagulase enzyme activity was detected in 5% (2/40; 1 T.asahii, 1 T.inkin) of the strains with human plasma and in 27.5% (11/40; 9 T.asahii, 1 T.inkin, 1 T.montevideense) with rabbit plasma. In conclusion, our data indicated that esterase, coagulase and hemolytic activities detected in Trichosporon spp. might play role in the pathogenesis of Trichosporon infections, however, further large-scaled clinical and mycological studies are needed to prove this relation.

Comparison of m-Endo LES, MacConkey, and Teepol media for membrane filtration counting of total coliform bacteria in water.

PubMed Central

Grabow, W O; du Preez, M

1979-01-01

Total coliform counts obtained by means of standard membrane filtration techniques, using MacConkey agar, m-Endo LES agar, Teepol agar, and pads saturated with Teepol broth as growth media, were compared. Various combinations of these media were used in tests on 490 samples of river water and city wastewater after different stages of conventional purification and reclamation processes including lime treatment, and filtration, active carbon treatment, ozonation, and chlorination. Endo agar yielded the highest average counts for all these samples. Teepol agar generally had higher counts then Teepol broth, whereas MacConkey agar had the lowest average counts. Identification of 871 positive isolates showed that Aeromonas hydrophila was the species most commonly detected. Species of Escherichia, Citrobacter, Klebsiella, and Enterobacter represented 55% of isolates which conformed to the definition of total coliforms on Endo agar, 54% on Teepol agar, and 45% on MacConkey agar. Selection for species on the media differed considerably. Evaluation of these data and literature on alternative tests, including most probable number methods, indicated that the technique of choice for routine analysis of total coliform bacteria in drinking water is membrane filtration using m-Endo LES agar as growth medium without enrichment procedures or a cytochrome oxidase restriction. PMID:394678

Relative diffusion of paramagnetic metal complexes of MRI contrast agents in an isotropic hydrogel medium.

PubMed

Weerakoon, Bimali Sanjeevani; Osuga, Toshiaki

2017-03-01

The observation of molecular diffusion by means of magnetic resonance imaging (MRI) is significant in the evaluation of the metabolic activity of living tissues. Series of MRI examinations were conducted on a diffusion model to study the behaviour of the diffusion process of different-molecular-weight (MW) paramagnetic MRI contrast agents in an isotropic agar hydrogel medium. The model consisted of a solidified 1 % agar gel with an initial concentration of 0.5 mmol/L contrast solution layered on top of the gel. The diffusion process was monitored at pre-determined time intervals of immediately, 1, 6, 9, 23, and 48 h after introduction of the contrast agents onto the agar gel with a T1-weighted spin-echo (SE) pulse sequence. Three types of paramagnetic contrast agents, Gd-DTPA with a MW of 547.57 g/mol, Prohance with a MW of 558.69 g/mol and MnCl 2 with a MW of 125.84 g/mol, resulted in an approximate average diffusional displacement ratio of 1:1:2 per hour, respectively, within 48 h of the experiment. Therefore, the results of this study supported the hypothesis that the rate of the diffusion process of MRI contrast agents in the agar hydrogel medium is inversely related to their MWs. However, more repetitions are necessary under various types of experimental conditions and also with various types of contrast media of different MWs for further confirmation and validation of these results.

Direct impression on agar surface as a diagnostic sampling procedure for candida balanitis.

PubMed

Lisboa, Carmen; Santos, António; Azevedo, Filomena; Pina-Vaz, Cidália; Rodrigues, Acácio Gonçalves

2010-02-01

The diagnosis of candida balanitis should be based upon both clinical and mycological data. The procedure of material collection is a critical issue to confirm or rule out the clinical diagnosis of candida balanitis. To compare direct impression of the glans on the agar surface of solid culture media with the collection of genital exudates with cotton swab for the diagnosis of candida balanitis. A prospective cross-sectional study was carried out during a 36-month period. Sexually transmitted disease clinic attendees with balanitis and asymptomatic men were included. Specimens for yeast culture were collected from the glans penis and inner preputial layer using the direct impression on CHROMagar candida medium and by swabbing with a sterile cotton swab. Among 478 men enrolled, 189 had balanitis. The prevalence of candida balanitis was 17.8% (85/478) confirmed after culture by direct impression; the swab method detected only 54/85 (63.5%) of these men. Of the 289 asymptomatic men, 36 (12.5%) yielded Candida spp; the swab method detected only 38.9% of these men. The risk of having candida balanitis is 8.9 (IC 95% 2.48 to 32.04) whenever the number of candida colonies recovered by direct impression was greater than 10. Direct impression on CHROMagar candida medium resulted in the highest Candida spp recovery rate. More than 10 colonies yielded by impression culture were statistically associated with candida balanitis. This method shows the ideal profile for sampling the male genital area for yeasts and should be included in the management of balanitis.

Evaluation of Caenorhabditis elegans as an acute lethality and a neurotoxicity screening model

DOE Office of Scientific and Technical Information (OSTI.GOV)

Williams, P.L.

1988-01-01

This investigation evaluated C. elegans as a lethality and neurotoxicity screening model. The lethality experiments were performed in both agar and an aquatic medium. The salts of 8 metals (Hg, Be, Al, Cu, Zn, Pb, Cd, and Sr) were used in the agar studies and the salts of 14 metals (Ag, Hg, Cu, Be, Al, Pb, Cr, As, Tl, Zn, Cd, Ni, Sr, and Sb) were used in the aquatic tests. In each of these tests an LC50 value was determined. The data from the agar plates were compared to the published mammalian oral LD50 values for salts of themore » same metals. Within this set of chemicals C. elegans was found to be a predictor of mammalian acute lethality, generating LC50 values parallel to the rat and mouse LD50 values. The aquatic data were compared to data from EPA Ambient Water Quality Criteria documents. C. elegans was found to be less sensitive than Daphnia but generally more sensitive than the other invertebrate organisms that are presently used. The neurotoxicity testing also was performed in both agar and an aquatic media. The testing in agar was conducted with the salts of 4 metals (Cu, Be, Pb, and Hg) and 2 organophosphate pesticides (malathion and vapona). The studies in an aquatic medium tested the salts of 4 metals (Cu, Be, Pb, and Hg).« less

Production of microbial medium from defatted brebra (Milletia ferruginea) seed flour to substitute commercial peptone agar

PubMed Central

Andualem, Berhanu; Gessesse, Amare

2013-01-01

Objective To investigate and optimize microbial media that substitute peptone agar using brebra seed defatted flour. Methods 'Defatted process, inoculums preparation, evaluation of bacterial growth, preparation of cooked and hydrolyzed media and growth turbidity of tested bacteria were determined. Results Two percent defatted flour was found to be suitable concentration for the growth of pathogenic bacteria: Escherichia coli (ATCC 25922) (E. coli), Pseudomonas aeruginosa (ATCC 27853), Salmonella (NCTC 8385) and Shigella flexneri (ATCC 12022) (S. flexneri), while 3% defatted flour was suitable for Staphylococcus aureus (ATCC 25923) (S. aureus). E. coli (93±1) and S. flexneri (524±1) colony count were significantly (P≤0.05) greater in defatted flour without supplement than in supplemented medium. E. coli [(3.72×109±2) CFU/mL], S. aureus [(7.4×109±2) CFU/mL], S. flexneri [(4.03×109±2) CFU/mL] and Salmonella [(2.37×109±1) CFU/mL] in non-hydrolyzed sample were statistically (P≤0.05) greater than hydrolyzed one and commercial peptone agar. Colony count of Salmonella [(4.55×109±3) CFU/mL], S. flexneri [(5.40×109±3) CFU/mL] and Lyesria moncytogenes (ATCC 19116) [(5.4×109±3) CFU/mL] on raw defatted flour agar was significantly (P≤0.05) greater than cooked defatted flour and commercial peptone agar. Biomass of E. coli, S. aureus, Salmonella and Enterococcus faecalis in non-hydrolyzed defatted flour is highly increased over hydrolyzed defatted flour and commercial peptone broth. Conclusions The defatted flour agar was found to be better microbial media or comparable with peptone agar. The substances in it can serve as sources of carbon, nitrogen, vitamins and minerals that are essential to support the growth of microorganisms without any supplements. Currently, all supplements of peptone agar are very expensive in the market. PMID:24075344

Evaluation of CHROMagar KPC for Rapid Detection of Carbapenem-Resistant Enterobacteriaceae▿

PubMed Central

Samra, Zmira; Bahar, Judi; Madar-Shapiro, Liora; Aziz, Nazi; Israel, Sara; Bishara, Jihad

2008-01-01

A new CHROMagar KPC medium was compared to MacConkey agar with carbapenem discs and PCR for the blaKPC gene for rapid detection of carbapenem-resistant Klebsiella pneumoniae. The sensitivity and specificity relative to PCR were 100% and 98.4%, respectively, for CHROMagar KPC and 92.7% and 95.9%, respectively, for MacConkey agar. PMID:18632915

Identification of phanerosporic acid in birch degraded by Phanerochaete chrysosporium

Treesearch

Michael D. Mozuch; Philip J. Kersten

2003-01-01

Extracts of Phanerochaete chrysosporium cultures grown on birch or on a malt extract-peptone-glucose agar medium were analysed by HPLC. A major component from the two sources appears to be identical by HPLC and UV- visible spectrometry. The product isolated from agar-grown cultures was purified to apparent homogeneity and structure analysis by NMR indicates that the...

Growth inhibition of heat-injured Enterococcus faecium by oligophosphates in a cured meat model.

PubMed

Houben, J H; Tjeerdsma-van Bokhoven, J L M

2004-12-01

Cells of two heat-resistant strains of Enterococcus faecium were heated and incubated in meat suspensions containing curing ingredients. The concentrations of the curing ingredients were those frequently used for pasteurized ham-type products, except that the concentrations of the oligophosphates (triphosphate and diphosphate) varied. Heating tests at 69 degrees C were performed with inoculated meat suspensions in heat-sealed plastic pouches. Numbers of bacteria were counted immediately after heating and in parallel series of heated pouches incubated at 37 degrees C. Plating was performed in Tryptone Dextrose Yeast Meat Peptonised Milk Agar (TDYMP); in TDYMP Agar to which the curing ingredients were added; and in TDYMP Agar to which the curing ingredients except oligophosphates were added. The inclusion of oligophosphates in the heating medium increased the heat-injury sustained by the E. faecium cells, and in combination with rather severe heat treatment even completely blocked the growth of surviving organisms in the meat suspension incubated at 37 degrees C. The presence of oligophosphates in the culture medium TDYMP Agar severely reduced the counts of freshly heated cells; however, this effect disappeared after repair and growth of the surviving organisms in the meat suspension.

Arabidopsis alcohol dehydrogenase expression in both shoots and roots is conditioned by root growth environment

NASA Technical Reports Server (NTRS)

Chung, H. J.; Ferl, R. J.

1999-01-01

It is widely accepted that the Arabidopsis Adh (alcohol dehydrogenase) gene is constitutively expressed at low levels in the roots of young plants grown on agar media, and that the expression level is greatly induced by anoxic or hypoxic stresses. We questioned whether the agar medium itself created an anaerobic environment for the roots upon their growing into the gel. beta-Glucuronidase (GUS) expression driven by the Adh promoter was examined by growing transgenic Arabidopsis plants in different growing systems. Whereas roots grown on horizontal-positioned plates showed high Adh/GUS expression levels, roots from vertical-positioned plates had no Adh/GUS expression. Additional results indicate that growth on vertical plates closely mimics the Adh/GUS expression observed for soil-grown seedlings, and that growth on horizontal plates results in induction of high Adh/GUS expression that is consistent with hypoxic or anoxic conditions within the agar of the root zone. Adh/GUS expression in the shoot apex is also highly induced by root penetration of the agar medium. This induction of Adh/GUS in shoot apex and roots is due, at least in part, to mechanisms involving Ca2+ signal transduction.

Effect of impact stress on microbial recovery on an agar surface.

PubMed Central

Stewart, S L; Grinshpun, S A; Willeke, K; Terzieva, S; Ulevicius, V; Donnelly, J

1995-01-01

Microbial stress due to the impaction of microorganisms onto an agar collection surface was studied experimentally. The relative recovery rates of aerosolized Pseudomonas fluorescens and Micrococcus luteus were determined as a function of the impaction velocity by using a moving agar slide impactor operating over a flow rate range from 3.8 to 40 liters/min yielding impaction velocities from 24 to 250 m/s. As a reference, the sixth stage of the Andersen Six-Stage Viable Particle Sizing Sampler was used at its operating flow rate of 28.3 liters/min (24 m/s). At a collection efficiency of close to 100% for the agar slide impactor, an increase in sampling flow rate and, therefore, in impaction velocity produced a significant decline in the percentage of microorganisms recovered. Conversely, when the collection efficiency was less than 100%, greater recovery and lower injury rates occurred. The highest relative rate of recovery (approximately 51% for P. fluorescens and approximately 62% for M. luteus) was obtained on the complete (Trypticase soy agar) medium at 40 and 24 m/s (6.4 and 3.8 liters/min), respectively. M. luteus demonstrated less damage than P. fluorescens, suggesting the hardy nature of the gram-positive strain versus that of the gram-negative microorganism. Comparison of results from the agar slide and Andersen impactors at the same sampling velocity showed that recovery and injury due to collection depends not only on the magnitude of the impaction velocity but also on the degree to which the microorganisms may be embedded in the collection medium.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7747946

Random transposon mutagenesis of the Saccharopolyspora erythraea genome reveals additional genes influencing erythromycin biosynthesis

PubMed Central

Fedashchin, Andrij; Cernota, William H.; Gonzalez, Melissa C.; Leach, Benjamin I.; Kwan, Noelle; Wesley, Roy K.; Weber, J. Mark

2015-01-01

A single cycle of strain improvement was performed in Saccharopolyspora erythraea mutB and 15 genotypes influencing erythromycin production were found. Genotypes generated by transposon mutagenesis appeared in the screen at a frequency of ∼3%. Mutations affecting central metabolism and regulatory genes were found, as well as hydrolases, peptidases, glycosyl transferases and unknown genes. Only one mutant retained high erythromycin production when scaled-up from micro-agar plug fermentations to shake flasks. This mutant had a knockout of the cwh1 gene (SACE_1598), encoding a cell-wall-associated hydrolase. The cwh1 knockout produced visible growth and morphological defects on solid medium. This study demonstrated that random transposon mutagenesis uncovers strain improvement-related genes potentially useful for strain engineering. PMID:26468041

Development of a selective agar plate for the detection of Campylobacter spp. in fresh produce.

PubMed

Yoo, Jin-Hee; Choi, Na-Young; Bae, Young-Min; Lee, Jung-Su; Lee, Sun-Young

2014-10-17

This study was conducted to develop a selective medium for the detection of Campylobacter spp. in fresh produce. Campylobacter spp. (n=4), non-Campylobacter (showing positive results on Campylobacter selective agar) strains (n=49) isolated from fresh produce, indicator bacteria (n=13), and spoilage bacteria isolated from fresh produce (n=15) were plated on four Campylobacter selective media. Bolton agar and modified charcoal cefoperazone deoxycholate agar (mCCDA) exhibited higher sensitivity for Campylobacter spp. than did Preston agar and Hunt agar, although certain non-Campylobacter strains isolated from fresh produce by using a selective agar isolation method, were still able to grow on Bolton agar and mCCDA. To inhibit the growth of non-Campylobacter strains, Bolton agar and mCCDA were supplemented with 5 antibiotics (rifampicin, polymyxin B, sodium metabisulfite, sodium pyruvate, ferrous sulfate) and the growth of Campylobacter spp. (n=7) and non-Campylobacter strains (n=44) was evaluated. Although Bolton agar supplemented with rifampicin (BR agar) exhibited a higher selectivity for Campylobacter spp. than did mCCDA supplemented with antibiotics, certain non-Campylobacter strains were still able to grow on BR agar (18.8%). When BR agar with various concentrations of sulfamethoxazole-trimethoprim were tested with Campylobacter spp. (n=8) and non-Campylobacter (n=7), sulfamethoxazole-trimethoprim was inhibitory against 3 of 7 non-Campylobacter strains. Finally, we validated the use of BR agar containing 50mg/L sulfamethoxazole (BRS agar) or 0.5mg/L ciprofloxacin (BRCS agar) and other selective agars for the detection of Campylobacter spp. in chicken and fresh produce. All chicken samples were positive for Campylobacter spp. when tested on mCCDA, BR agar, and BRS agar. In fresh produce samples, BRS agar exhibited the highest selectivity for Campylobacter spp., demonstrating its suitability for the detection of Campylobacter spp. in fresh produce. Copyright © 2014 Elsevier B.V. All rights reserved.

Growth and differentiation of a murine interleukin-3-producing myelomonocytic leukemia cell line in a protein-free chemically defined medium.

PubMed

Kajigaya, Y; Ikuta, K; Sasaki, H; Matsuyama, S

1990-10-01

We established the continuous growth of WEHI-3B D+ cells in protein-free chemically defined F-12 medium by stepwise decreases in the concentration of fetal calf serum. This cell line, designated as WEHI-3B-Y1, has now been propagated in protein-free F-12 medium for 3 years. The population-doubling time of the cells in culture is about 24 hr. WEHI-3B-Y1 cells are immature undifferentiated cells which show positive staining for naphthol ASD chloroacetate esterase and alpha-naphthyl butyrate esterase and spontaneously exhibit a low level of differentiation to mature granulocytes and macrophages. Medium conditioned by WEHI-3B-Y1 cells stimulated the proliferation of an interleukin-3 (IL-3)-dependent FDCP-2 cell line. This conditioned medium was shown to have erythroid burst-promoting activity when assayed using normal murine bone marrow. The colony formation of WEHI-3B-Y1 cells in semi-solid agar culture was not stimulated by purified recombinant human granulocyte colony-stimulating factor (rhG-CSF). However, in the presence of human transferrin, rhG-CSF enhanced the number of colonies of WEHI-3B-Y1 cells but did not induce their differentiation. These results suggest that WEHI-3B-Y1 cells cultured in protein-free medium produced murine IL-3. In addition, human G-CSF enhanced the clonal growth but did not induce the differentiation of WEHI-3B-Y1 cells cultured in serum-free medium.

Effect of the fungus Pochonia chlamydosporia on Echinostoma paraensei (Trematoda: Echinostomatidae).

PubMed

Lelis, Rosane Teixeira; Braga, Fabio Ribeiro; de Carvalho, Lorendane Millena; de Paula, Alessandra Teixeira; Araujo, Juliana Milani; Fausto, Mariana Costa; Junior, Arnaldo Maldonado; Rodrigues, João Victor Facchini; de Freitas Soares, Filippe Elias; Garcia, Juberlan Silva; de Araújo, Jackson Victor

2014-11-01

Echinostoma paraensei is a trematode of the genus Echinostoma that causes echinostomiasis in humans. The objectives of this study were to: evaluate the ovicidal activity of the nematophagous fungus Pochonia chlamydosporia (VC1 and VC4) on a solid medium 2% water-agar (2% WA) against E. paraensei eggs (assay A); evaluate ovicidal effect (destruction of eggs) of the isolate VC4 in supplemented culture media (assay B); and evaluate the ovicidal ability of the crude extract (VC4) on E. paraensei eggs (assay C). Eggs of E. paraensei (assay A) were placed in Petri dishes containing 2% WA with an isolate of the fungus P. chlamydosporia (VC1 and VC4) grown for 10 days, and without fungus as a control and evaluated regarding their destruction. In assay B, eggs of E. paraensei were placed in Petri dishes with different supplemented culture media and with VC4 isolate and the destruction of eggs was examined at the end of 25 days of interaction. In assay C, effects of the crude extract of P. chlamydosporia (VC4) on eggs were evaluated at the end of 7 days. In assay A, there was no difference (p>0.05) in ovicidal activity among the tested isolates (VC1 and VC4); however, the highest percentage for ovicidal activity (type 3 effect) was demonstrated by the isolate VC4. In assay B, the culture medium starch-agar showed the best results for the destruction of the eggs, with a percentage of 46.6% at the end of the assay. In assay C, the crude extract of VC4 was effective in the destruction of E. paraensei eggs, with a percentage reduction of 53%. The results of this study demonstrate that a rich culture medium with a greater availability of carbon and nitrogen may interfere directly in the predatory characteristics of ovicidal fungi. Copyright © 2014 Elsevier B.V. All rights reserved.

New method for the isolation of Streptococcus mutans and its differentiation from other oral streptococci.

PubMed

Linke, H A

1977-06-01

A new, improved agar medium for the isolation of Streptococcus mutans, the etiological agent of dental caries, was developed. In contrast to mitis-salivarius agar, this medium not only recovers a greater number of S. mutans strains from most oral specimens but, because of its mannitol and sorbitol content, it also facilitates the differentiation of S. mutans from other oral streptococci, e.g., S. salivarius, S. mitis, and S. sanguis, which do not grow or produce scanty growth only after 10 days of incubation. The medium is easy to prepare because of its simple and unique composition, is characterized by the presence of an acid indicator, and can be utilized under aerobic and anaerobic conditions as well. The medium cannot be used to distinguish among the eight serotypes, a to g and SL-1, of S. mutans. Mannitol-utilizing bacteria such as streptococci (e.g., S. faecalis) and other microorganisms (e.g., Staphylococcus aureus) are able to grow on this medium and can be distinguished from S. mutans by their unique colony morphology.

Analysis of soybean tissue culture protein dynamics using difference gel electrophoresis

USDA-ARS?s Scientific Manuscript database

Excised hypocotyls from developing soybean (Glycine max (L.) merr. cv. Jack) were cultivated on agar-solidified medium until callus formed. The calli were then propagated in liquid medium until stable, relatively uniform, finely-divided suspension cultures were obtained. Cells were typically transfe...

Standard Nutrient Agar 1 as a substitute for blood-supplemented Müller-Hinton agar for antibiograms in developing countries.

PubMed

Niederstebruch, N; Sixt, D

2013-02-01

In the industrial world, the agar diffusion test is a standard procedure for the susceptibility testing of bacteria isolates. Beta-hemolytic Streptococcus spp. are tested with Müller-Hinton agar supplemented with 5 % blood, a so-called blood agar. The results are interpreted using standardized tables, which only exist for this type of nutrient matrix. Because of a number difficulties, both with respect to technical issues and to manual skills, blood agar is not a feasible option in many developing countries. Beta-hemolytic Streptococcus spp. also grow on Standard Nutrient Agar 1 (StNA1). This suggests using that type of nutrient medium for running agar diffusion tests. However, there are no standardized tables that can be used for interpreting the diameters of the zones of inhibition on StNA1 1. Using the existing standardized tables for blood agar to interpret cultures on StNA1 1 would be of great benefit under such circumstances where blood agar is not available. With this in mind, we conducted comparative tests to evaluate the growth characteristics of beta-hemolytic Streptococcus spp. on StNA1 1 compared to Müller-Hinton agar supplemented with 5 % sheep blood. In this study, we were able to show that beta-hemolytic Streptococcus spp. develop similar zones of inhibition on blood agar and on StNA1 1. Therefore, it is suggested that, for the interpretation of antibiograms of beta-hemolytic Streptococcus spp. performed on StNA1 1, the standard tables for blood agar can be used.

The routine use of modified Borelli's lactritmel agar (MBLA).

PubMed

Kaminski, G W

1985-07-01

The original formula of Borelli's lactritmel agar (BLA)(3) which contains wheat flour, milk and honey, has been modified by replacing the wheat flour with dehydrated Bacto Corn Meal Agar (Difco) and by slightly altering the concentrations of the milk and honey. The modified medium (MBLA) is less turbid, less particulate, and easier to prepare than BLA. Although Trichophyton rubrum usually produces a wine-red pigment with BLA, most strains initially produce a yellow pigment, with the red pigment developing later. The corn meal in MBLA reduces this tendency and stimulates the early formation of deep wine red pigment, MBLA enhances sporulation of dermatophytes and various fungi which fail to sporulate on other media, and maintains characteristic growth without developing pleomorphic degeneration. It has been used routinely since 1972 as a reliable aid to the differentiation of T. rubrum and T. mentagrophytes. Since 1975 selective MBLA has been used as a routine primary isolation medium for dermatophytes, and has proved to be most useful.

Comparison of three different C18 HPLC columns with different particle sizes for the optimization of aflatoxins analysis.

PubMed

Medina, A; Magan, N

2012-03-15

In this work we compared the performance of chromatography columns with particles of 5 and 3 μm with the new 2.7 μm solid core particles for the analysis of aflatoxins B1, G1, B2, and G2 using trifluoroacetic acid pre-column derivatization. Three different columns have been used and chromatographic parameters as retention time, resolution, limit of detection (LOD), limit of quantification (LOQ) were obtained from all of them and compared. The results show that comparing with the traditional columns, shorter columns (100 mm × 4.6 mm) with the new solid core particles are suitable for the analysis of these mycotoxins and allowed the reduction of the analysis time by 45.5% and 33.3% with respect to columns with particle size 5 μm (150 mm × 4.6 mm) and 3 μm (150 mm × 4.6 mm) respectively, without any detrimental effect on performance. This leads to the reduction of the analysis costs by saving on organic solvents and increasing the total number of analyses per day. The capability of these columns for analyzing samples, in different culture media, was assessed by analyzing different samples from: yeasts extract sucrose medium, corn meal agar medium and fresh hazelnut media. Copyright © 2012 Elsevier B.V. All rights reserved.

Supplementation of CHROMagar Candida Medium with Pal's Medium for Rapid Identification of Candida dubliniensis

PubMed Central

Sahand, Ismail H.; Moragues, María D.; Eraso, Elena; Villar-Vidal, María; Quindós, Guillermo; Pontón, José

2005-01-01

CHROMagar Candida medium is used for the isolation and identification of Candida species, but it does not differentiate Candida albicans from Candida dubliniensis. This differentiation can be achieved by using Pal's agar, which cannot be used in primary isolation. We have combined both media to obtain a new medium that can be used for the isolation and identification of C. dubliniensis in primary cultures. PMID:16272515

Direct identification and recognition of yeast species from clinical material by using albicans ID and CHROMagar Candida plates.

PubMed

Baumgartner, C; Freydiere, A M; Gille, Y

1996-02-01

Two chromogenic media, Albicans ID and CHROMagar Candida agar plates, were compared with a reference medium, Sabouraud-chloramphenicol agar, and standard methods for the identification of yeast species. This study involved 951 clinical specimens. The detection rates for the two chromogenic media for polymicrobial specimens were 20% higher than that for the Sabouraud-chloramphenicol agar plates. The rates of identification of Candida albicans for Albicans ID and CHROMagar Candida agar plates were, respectively, 37.0 and 6.0% after 24 h of incubation and 93.6 and 92.2% after 72 h of incubation, with specificities of 99.8 and 100%. Furthermore, CHROMagar Candida plates identified 13 of 14 Candida tropicalis and 9 of 12 Candida krusei strains after 48 h of incubation.

Tissue culture of three species of Laurencia complex

NASA Astrophysics Data System (ADS)

Shen, Songdong; Wu, Xunjian; Yan, Binlun; He, Lihong

2010-05-01

To establish a micropropagation system of three Laurencia complex species ( Laurencia okamurai, Laurencia tristicha, and Chondrophycus undulatus) by tissue culture techniques, we studied the regeneration characteristics and optimal culture conditions of axenic algal fragments cultured on solid medium and in liquid medium. Regeneration structures were observed and counted regularly under a reverse microscope to investigate the regeneration process, polarity and optimal illumination, and temperature and salinity levels. The results show that in most cultures of the three species, we obtained bud regeneration on solidified medium with 0.5% agar and in liquid medium. Rhizoid-like regeneration was filamentous and developed from the lower cut surface of fragments in L. okamurai, but was discoid and developed from the apical back side of bud regeneration in L. tristicha and C. undulatus. Regeneration polarity was localized to the apical part of algal fronds in all three species, and on fragments cut from the basal part of algae buds could develop from both the upper and the lower cut surfaces. Buds could develop from both the medullary and the cortical portions in L. okamurai and C. undulatus, while in L. tristicha, buds only emerged from the cortex. The optimal culture conditions for L. okamurai were 4 500 lx, 20°C and 35 (salinity); for C. undulatus, 4 500 lx, 20°C and 30; and for L. tristicha, 4 500 lx, 25°C and 30.

Dynamic speckle study of microbial growth

NASA Astrophysics Data System (ADS)

Vincitorio, F. M.; Mulone, C.; Marcuzzi, P. A.; Budini, N.; Freyre, C.; Lopez, A. J.; Ramil, A.

2015-08-01

In this work we present a characterization of yeast dynamic speckle activity during growth in an isolated agar culture medium. We found that it is possible to detect the growth of the microorganisms even before they turn out to be visible. By observing the time evolution of the speckle activity at different regions of the culture medium we could extract a map of the growth process, which served to analyze how the yeast develops and spreads over the agar's medium. An interesting point of this study concerns with the influence of the laser light on the yeast growth rate. We have found that yeast finds hard to develop at regions with higher laser light illumination, although we used a synchronous system to capture the speckle pattern. The results obtained in this work would serve us as a starting point to fabricate a detector of growing microorganism colonies, with obvious interesting applications in diverse areas.

Effect of medium composition and light on root and rhinacanthin formation in Rhinacanthus nasutus cultures.

PubMed

Panichayupakaranant, P; Meerungrueang, W

2010-11-01

Rhinacanthus nasutus (L.) Kurz (Acanthaceae) has long been used in Thai traditional medicine for treatment of tinea versicolor, ringworm, pruritic rash, and abscess. The active constituents are known as a group of naphthoquinone esters, rhinacanthins. This work focused on establishment of R. nasutus root cultures and determination of rhinacanthin production. Induction of R. nasutus root formation was accomplished on solid Gamborg's B5 (B5) medium, supplied with 0.1 mg/L indole-3-butyric acid (IBA) and 20 g/L sucrose. The effects of explants (whole leaf explants and four-side excised leaf explants), light and medium composition on root and rhinacanthin formation were investigated. The root formation from the whole leaf explants was 10 times higher than that from the four-side excised leaf explants. In addition, light possessed an inhibitory effect on the root and rhinacanthin formation of R. nasutus. Medium manipulation found that Murashige and Skoog (MS) medium supplied with 3 mg/L IBA and 30 g/L sucrose was the most suitable for induction of the root formation. Unfortunately, the obtained root cultures produced only rhinacanthin-C in very low amount, 0.026 mg/g dry weight (DW), when they were transferred into the same MS liquid medium. With semisolid medium (4 g/L agar) of the same MS composition, however, the root cultures appeared to produce higher content of rhinacanthin-C, -D and -N (3.45, 0.07 and 0.07 mg/g DW, respectively). Our finding suggests that culturing in semisolid medium is capable of improving of rhinacanthin production in R. nasutus root cultures.

Novel Single-Tube Agar-Based Test System for Motility Enhancement and Immunocapture of Escherichia coli O157:H7 by H7 Flagellar Antigen-Specific Antibodies

PubMed Central

Murinda, Shelton E.; Nguyen, Lien T.; Ivey, Susan J.; Almeida, Raul A.; Oliver, Stephen P.

2002-01-01

This paper describes a novel single-tube agar-based technique for motility enhancement and immunoimmobilization of Escherichia coli O157:H7. Motility indole ornithine medium and agar (0.4%, wt/vol) media containing either nutrient broth, tryptone broth, or tryptic soy broth (TSBA) were evaluated for their abilities to enhance bacterial motility. Twenty-six E. coli strains, including 19 O157:H7 strains, 1 O157:H− strain, and 6 generic E. coli strains, were evaluated. Test bacteria were stab inoculated in the center of the agar column, and tubes were incubated at 37°C for 18 to 96 h. Nineteen to 24 of the 26 test strains (73.1 to 92.3%) were motile in the different media. TSBA medium performed best and was employed in subsequent studies of motility enhancement and H7 flagellar immunocapture. H7 flagellar antiserum (30 and 60 μl) mixed with TSBA was placed as a band (1 ml) in the middle of an agar column separating the top (3-ml) and bottom (3-ml) agar layers. The top agar layer was inoculated with the test bacterial strains. The tubes were incubated at 37°C for 12 to 18 h and for 18 to 96 h. The specificity and sensitivity of the H7 flagellar immunocapture tests were 75 and 100%, respectively. The procedure described is simple and sensitive and could be adapted easily for routine use in laboratories that do not have sophisticated equipment and resources for confirming the presence of H7 flagellar antigens. Accurate and rapid identification of H7 flagellar antigen is critical for the complete characterization of E. coli O157:H7, owing to the immense clinical, public health, and economic significance of this food-borne pathogen. PMID:12454173

Media for the isolation and enumeration of bifidobacteria in dairy products.

PubMed

Roy, D

2001-09-28

Bifidobacteria are commonly used for the production of fermented milks, alone or in combination with other lactic acid bacteria. Bifidobacteria populations in fermented milks should be over 10(6) bifidobacteria/g at the time of consumption of strain added to the product. Hence, rapid and reliable methods are needed to routinely determine the initial inoculum and to estimate the storage time period bifidobacteria remain viable. Plate count methods are still preferable for quality control measurements in dairy products. It is, therefore, necessary to have a medium that selectively promotes the growth of bifidobacteria, whereas other bacteria are suppressed. The present paper is an overview of media and methods including summaries of published comparisons between different selective media. Culture media for bifidobacteria may be divided into basal, elective, differential and selective culture medium. Non-selective media are useful for routine enumeration of bifidobacteria when present in non-fermented milks. Reinforced Clostridial Agar and De Man Rogosa Sharpe (MRS) supplemented with cysteine and agar available commercially are the media of choice for industrial quality control laboratories. Several media for selective or differential isolation have been described for enumeration of bifidobacteria from other lactic acid bacteria. From the large number of selective media available, it can be concluded that there is no standard medium for the detection of bifidobacteria. However, Columbia agar base media supplemented with lithium chloride and sodium propionate and MRS medium supplemented with neomycin, paromomycin, nalidixic acid and lithium chloride can be recommended for selective enumeration of bifidobacteria in dairy products.

Induction of anchorage-independent growth in primary human cells exposed to protons or HZE ions separately or in dual exposures.

PubMed

Sutherland, B M; Cuomo, N C; Bennett, P V

2005-10-01

Travelers on space missions will be exposed to a complex radiation environment that includes protons and heavy charged particles. Since protons are present at much higher levels than are heavy ions, the most likely scenario for cellular radiation exposure will be proton exposure followed by a hit by a heavy ion. Although the effects of individual ion species on human cells are being investigated extensively, little is known about the effects of exposure to both radiation types. One useful measure of mammalian cell damage is induction of the ability to grow in a semi-solid agar medium highly inhibitory to the growth of normal human cells, termed neoplastic transformation. Using primary human cells, we evaluated induction of soft-agar growth and survival of cells exposed to protons only or to heavy charged particles (600 MeV/nucleon silicon) only as well as of cells exposed to protons followed after a 4-day interval by silicon ions. Both ions alone efficiently transformed the human cells to anchorage-independent growth. Initial experiments indicate that the dose responses for neoplastic transformation of cells exposed to protons and then after 4 days to silicon ions appear similar to that of cells exposed to silicon ions alone.

Rhizobacterial characterization for quality control of eucalyptus biogrowth promoter products.

PubMed

Zarpelon, Talyta Galafassi; Guimarães, Lúcio Mauro da Silva; Alfenas-Zerbini, Poliane; Lopes, Eli Sidney; Mafia, Reginaldo Gonçalves; Alfenas, Acelino Couto

Plant growth-promoting rhizobacteria strains from special formulations have been used to optimize eucalyptus cutting production. To undertake quality control for the formulated products, the rhizobacterial strains should be characterized to assess their purity and authentication. In the present study, we characterized nine strains of rhizobacteria, including three Bacillus subtilis (S1, S2 and 3918), two Pseudomonas sp. (MF4 and FL2), P. putida (MF2), P. fulva (Ca), Frateuria aurantia (R1), and Stenotrophomonas maltophilia (CIIb). The strains were differentiated by colony morphology after 24h of incubation in three different solid state culture media (glucose-nutritive agar, 523 medium and yeast extract-mannitol agar), sensitivity to a panel of 28 antibiotics (expressed according to the formation of inhibition halos of bacterial growth in the presence of antibiotics), and PCR-RFLP profiles of the 16S rDNA gene produced using nine restriction enzymes. It was possible to differentiate all nine strains of rhizobacteria using their morphological characteristics and sensitivity to antibiotics. The molecular analysis allowed us to separate the strains CIIb, FL2 and R1 from the strains belonging to the genera Bacillus and Pseudomonas. By using these three methods concomitantly, we were able to determine strain purity and perform the authentication. Copyright © 2016. Published by Elsevier Editora Ltda.

Rheological and structural characterization of agar/whey proteins insoluble complexes.

PubMed

Rocha, Cristina M R; Souza, Hiléia K S; Magalhães, Natália F; Andrade, Cristina T; Gonçalves, Maria Pilar

2014-09-22

Complex coacervation between whey proteins and carboxylated or highly sulphated polysaccharides has been widely studied. The aim of this work was to characterise a slightly sulphated polysaccharide (agar) and whey protein insoluble complexes in terms of yield, composition and physicochemical properties as well as to study their rheological behaviour for better understanding their structure. Unlike other sulphated polysaccharides, complexation of agar and whey protein at pH 3 in the absence of a buffering agent resulted in a coacervate that was a gel at 20°C with rheological properties and structure similar to those of simple agar gels, reinforced by proteins electrostatically aggregated to the agar network. The behaviour towards heat treatment was similar to that of agar alone, with a high thermal hysteresis and almost full reversibility. In the presence of citrate buffer, the result was a "flocculated solid", with low water content (75-81%), whose properties were governed by protein behaviour. Copyright © 2014 Elsevier Ltd. All rights reserved.

Agar Sediment Test for Assessing the Suitability of Organic Waste Streams for Recovering Nutrients by the Aquatic Worm Lumbriculus variegatus.

PubMed

Laarhoven, Bob; Elissen, H J H; Temmink, H; Buisman, C J N

2016-01-01

An agar sediment test was developed to evaluate the suitability of organic waste streams from the food industry for recovering nutrients by the aquatic worm Lumbriculus variegatus (Lv). The effects of agar gel, sand, and food quantities in the sediment test on worm growth, reproduction, and water quality were studied. Agar gel addition ameliorated growth conditions by reducing food hydrolysis and altering sediment structure. Best results for combined reproduction and growth were obtained with 0.6% agar-gel (20 ml), 10 g. fine sand, 40 g. coarse sand, and 105 mg fish food (Tetramin). With agar gel, ingestion and growth is more the result of addition of food in its original quality. Final tests with secondary potato starch sludge and wheat bran demonstrated that this test is appropriate for the comparison of solid feedstuffs and suspended organic waste streams. This test method is expected to be suitable for organic waste studies using other sediment dwelling invertebrates.

Agar Sediment Test for Assessing the Suitability of Organic Waste Streams for Recovering Nutrients by the Aquatic Worm Lumbriculus variegatus

PubMed Central

Laarhoven, Bob; Elissen, H. J. H.; Temmink, H.; Buisman, C. J. N.

2016-01-01

An agar sediment test was developed to evaluate the suitability of organic waste streams from the food industry for recovering nutrients by the aquatic worm Lumbriculus variegatus (Lv). The effects of agar gel, sand, and food quantities in the sediment test on worm growth, reproduction, and water quality were studied. Agar gel addition ameliorated growth conditions by reducing food hydrolysis and altering sediment structure. Best results for combined reproduction and growth were obtained with 0.6% agar-gel (20 ml), 10 g. fine sand, 40 g. coarse sand, and 105 mg fish food (Tetramin). With agar gel, ingestion and growth is more the result of addition of food in its original quality. Final tests with secondary potato starch sludge and wheat bran demonstrated that this test is appropriate for the comparison of solid feedstuffs and suspended organic waste streams. This test method is expected to be suitable for organic waste studies using other sediment dwelling invertebrates. PMID:26937632

[The use of polymer gel dosimetry to measure dose distribution around metallic implants].

PubMed

Nagahata, Tomomasa; Yamaguchi, Hajime; Monzen, Hajime; Nishimura, Yasumasa

2014-10-01

A semi-solid polymer dosimetry system using agar was developed to measure the dose distribution close to metallic implants. Dosimetry of heterogeneous fields where electron density markedly varies is often problematic. This prompted us to develop a polymer gel dosimetry technique using agar to measure the dose distribution near substance boundaries. Varying the concentration of an oxygen scavenger (tetra-hydroxymethyl phosphonium chloride) showed the absorbed dose and transverse relaxation rate of the magnetic resonance signal to be linear between 3 and 12 Gy. Although a change in the dosimeter due to oxidization was observed in room air after 24 hours, no such effects were observed in the first 4 hours. The dose distribution around the metal implants was measured using agar dosimetry. The metals tested were a lead rod, a titanium hip joint, and a metallic stent. A maximum 30% dose increase was observed near the lead rod, but only a 3% increase in the absorbed dose was noted near the surface of the titanium hip joint and metallic stent. Semi-solid polymer dosimetry using agar thus appears to be a useful method for dosimetry around metallic substances.

Influence of Sodium Chloride on Growth of Neisseria meningitidis

PubMed Central

Mitzel, John R.; Hunter, Jack A.; Beam, Walter E.

1972-01-01

Nasopharyngeal isolates of Neisseria meningitidis were tested for growth on nutrient agar with and without the addition of 0.8% sodium chloride. Of the 822 strains tested, 1.3% grew on the salt-free medium, and 74.1% grew on the medium supplemented with sodium chloride. PMID:4626905

Echinocandin Susceptibility Testing of Candida Species: Comparison of EUCAST EDef 7.1, CLSI M27-A3, Etest, Disk Diffusion, and Agar Dilution Methods with RPMI and IsoSensitest Media▿

PubMed Central

Arendrup, Maiken Cavling; Garcia-Effron, Guillermo; Lass-Flörl, Cornelia; Lopez, Alicia Gomez; Rodriguez-Tudela, Juan-Luis; Cuenca-Estrella, Manuel; Perlin, David S.

2010-01-01

This study compared nine susceptibility testing methods and 12 endpoints for anidulafungin, caspofungin, and micafungin with the same collection of blinded FKS hot spot mutant (n = 29) and wild-type isolates (n = 94). The susceptibility tests included EUCAST Edef 7.1, agar dilution, Etest, and disk diffusion with RPMI-1640 plus 2% glucose (2G) and IsoSensitest-2G media and CLSI M27A-3. Microdilution plates were read after 24 and 48 h. The following test parameters were evaluated: fks hot spot mutants overlapping the wild-type distribution, distance between the two populations, number of very major errors (VMEs; fks mutants misclassified as susceptible), and major errors (MEs; wild-type isolates classified as resistant) using a wild-type-upper-limit value (WT-UL) (two twofold-dilutions higher than the MIC50) as the susceptibility breakpoint. The methods with the lowest number of errors (given as VMEs/MEs) across the three echinocandins were CLSI (12%/1%), agar dilution with RPMI-2G medium (14%/0%), and Etest with RPMI-2G medium (8%/3%). The fewest errors overall were observed for anidulafungin (4%/1% for EUCAST, 4%/3% for CLSI, and 3%/9% for Etest with RPMI-2G). For micafungin, VME rates of 10 to 71% were observed. For caspofungin, agar dilution with either medium was superior (VMEs/MEs of 0%/1%), while CLSI, EUCAST with IsoSensitest-2G medium, and Etest were less optimal (VMEs of 7%, 10%, and 10%, respectively). Applying the CLSI breakpoint (S ≤ 2 μg/ml) for CLSI results, 89.2% fks hot spot mutants were classified as anidulafungin susceptible, 60.7% as caspofungin susceptible, and 92.9% as micafungin susceptible. In conclusion, no test was perfect, but anidulafungin susceptibility testing using the WT-UL to define susceptibility reliably identified fks hot spot mutants. PMID:19884370

Development of novel agar media for isolating guaiacol producing Alicyclobacillus spp.

PubMed

Chang, S S; Park, S H; Kang, D H

2013-06-03

The purpose of this study is to develop a selective and differential medium (SK2 agar) for isolating guaiacol producing Alicyclobacillus. Forty-one selected dyes and vanillic acid were incorporated in SK agar for screening selective and differential agents. Two guaiacol producing (1016, 1101) and two non-guaiacol producing (19220, C-GD 1-1) Alicyclobacillus isolates were streaked onto media and color differentiation of the isolates was assessed. Among 41 tested dyes, Chrome Azurol S (CAS) allowed color differentiation of the two types of Alicyclobacillus. Colonies of guaiacol producing Alicyclobacillus isolates appeared as dark purple to royal blue color with yellow background, whereas non-guaiacol producing Alicyclobacillus isolates produced cream colored colonies with yellow background. Vanillic acid not only served as a precursor for guaiacol formation but also inhibited non-guaiacol producing Alicyclobacillus. Non-guaiacol producing isolates did not grow on SK agar containing more than 70 ppm vanillic acid, whereas the recovery of guaiacol producing isolates was unaffected. When compared with other Alicyclobacillus isolation media, not only was SK2 agar capable of selectively recovering guaiacol-producing Alicyclobacillus, the degree of growth was also approximately equal if not better than orange serum agar, potato dextrose agar, and K agar. The development of SK2 agar provides the fruit juice industry with an inexpensive, simple to use alternative for the detection of guaiacol producing Alicyclobacillus. Copyright © 2013 Elsevier B.V. All rights reserved.

Rapid multiplication of Dalbergia sissoo Roxb.: a timber yielding tree legume through axillary shoot proliferation and ex vitro rooting.

PubMed

Vibha, J B; Shekhawat, N S; Mehandru, Pooja; Dinesh, Rachana

2014-01-01

An efficient and improved method for in vitro propagation of mature tree of Dalbergia sissoo, an ecologically and commercially important timber yielding species, has been developed through axillary shoot proliferation. Bud breaking occurred from nodal shoot segments derived from rejuvenated shoots produced during early spring from a 20-25-year-old lopped tree, on MS medium containing 8.88 μM benzylaminopurine (BAP). Multiple shoots differentiated (20-21shoots/node) on re-culture of explants on half-strength agar gelled amended MS medium with a combination of 2.22 μM of BAP and 0.002 μM of thidiazuron (TDZ) with 1.0 mM each of Ca(NO3)2, K2SO4, KCl, and NH4(SO4)2. The maximum shoot multiplication (29-30 shoots/node) was achieved on subculturing in the above mentioned but liquid medium. Furthermore, the problem of shoot tip necrosis and defoliation observed on solid medium were overcome by the use of liquid medium. Ex vitro rooting was achieved on soilrite after basal treatment of microshoots with 984 μM of indole-3-butyric acid (IBA) for 2 min. About 90 % microshoots were rooted on soilrite within 2-3 weeks under the greenhouse conditions. From 20 nodal shoot segments, about 435 hardened plants were acclimatized and transplanted. This is the first report for rapid in vitro propagation of mature trees of D. sissoo on liquid medium followed by ex vitro rooting.

Production of microbial medium from defatted brebra (Milletia ferruginea) seed flour to substitute commercial peptone agar.

PubMed

Andualem, Berhanu; Gessesse, Amare

2013-10-01

To investigate and optimize microbial media that substitute peptone agar using brebra seed defatted flour. Defatted process, inoculums preparation, evaluation of bacterial growth, preparation of cooked and hydrolyzed media and growth turbidity of tested bacteria were determined. Two percent defatted flour was found to be suitable concentration for the growth of pathogenic bacteria: Escherichia coli (ATCC 25922) (E. coli), Pseudomonas aeruginosa (ATCC 27853), Salmonella (NCTC 8385) and Shigella flexneri (ATCC 12022) (S. flexneri), while 3% defatted flour was suitable for Staphylococcus aureus (ATCC 25923) (S. aureus). E. coli (93±1) and S. flexneri (524±1) colony count were significantly (P≤0.05) greater in defatted flour without supplement than in supplemented medium. E. coli [(3.72×10(9)±2) CFU/mL], S. aureus [(7.4×10(9)±2) CFU/mL], S. flexneri [(4.03×10(9)±2) CFU/mL] and Salmonella [(2.37×10(9)±1) CFU/mL] in non-hydrolyzed sample were statistically (P≤0.05) greater than hydrolyzed one and commercial peptone agar. Colony count of Salmonella [(4.55×10(9)±3) CFU/mL], S. flexneri [(5.40×10(9)±3) CFU/mL] and Lyesria moncytogenes (ATCC 19116) [(5.4×10(9)±3) CFU/mL] on raw defatted flour agar was significantly (P≤0.05) greater than cooked defatted flour and commercial peptone agar. Biomass of E. coli, S. aureus, Salmonella and Enterococcus faecalis in non-hydrolyzed defatted flour is highly increased over hydrolyzed defatted flour and commercial peptone broth. The defatted flour agar was found to be better microbial media or comparable with peptone agar. The substances in it can serve as sources of carbon, nitrogen, vitamins and minerals that are essential to support the growth of microorganisms without any supplements. Currently, all supplements of peptone agar are very expensive in the market. Copyright © 2013 Asian Pacific Tropical Biomedical Magazine. Published by Elsevier B.V. All rights reserved.

Real-time optical monitoring of microbial growth using optimal combination of light-emitting diodes

NASA Astrophysics Data System (ADS)

Kobayashi, Ken-ichi; Yamada, Takeshi; Hiraishi, Akira; Nakauchi, Shigeki

2012-12-01

We developed a real-time optical monitoring system consisting of a monochrome complementary metal-oxide semiconductor (CMOS) camera and two light-emitting diodes (LEDs) with a constant temperature incubator for the rapid detection of microbial growth on solid media. As a target organism, we used Alicyclobacillus acidocaldarius, which is an acidophilic thermophilic endospore-forming bacterium able to survive in pasteurization processes and grow in acidic drink products such as apple juice. This bacterium was cultured on agar medium with a redox dye applied to improve detection sensitivity. On the basis of spectroscopic properties of the colony, medium, and LEDs, an optimal combination of two LED illuminations was selected to maximize the contrast between the colony and medium areas. We measured A. acidocaldarius and Escherichia coli at two different dilution levels using these two LEDs. From the results of time-course changes in the number of detected pixels in the detection images, a similar growth rate was estimated amongst the same species of microbes, regardless of the dilution level. This system has the ability to detect a colony of approximately 26 μm in diameter in a detection image, and it can be interpreted that the size corresponds to less than 20 μm diameter in visual inspection.

A novel agar formulation for isolation and direct enumeration of Vibrio vulnificus from oyster tissue.

PubMed

Griffitt, Kimberly J; Grimes, D Jay

2013-08-01

A new selective and differential medium, Vibrio vulnificus X-Gal (VVX), was developed for direct enumeration of V. vulnificus (Vv) from oyster samples. This agar utilizes cellobiose and lactose as carbon sources, and the antibiotics colistin and polymyxin B as selective agents. Hydrolysis of 5-bromo-4-chloro-3-indolyl- beta-d-galactopyranoside (x-gal), used in the agar as a lactose analog, produces an insoluble blue dye that makes lactose positive colonies easily distinguishable from any non-lactose fermenting bacteria. Various bacterial species were spot plated onto thiosulfate-citrate-bile salts-sucrose agar (TCBS), and CHROMagar Vibrio, two vibrio-specific selective agars, non-selective agar, and VVX to compare selectivity of VVX to other widely used media. A V. vulnificus pure culture was serially diluted on VVX and non-selective agar to determine the VVX percent recovery. Water and oyster samples were spread plated on VVX agar and allowed to incubate for 16-18 h at 33 °C. Blue and white colonies from VVX agar were picked and screened by end point PCR for the Vv hemolysin vvhA. VVX agar showed a significant improvement over TCBS and CHROMagar at preventing non-target growth. There was an 87.5% recovery compared to non-selective plating and a 98% positivity rate of blue colonies picked from oyster tissue plating. The findings suggest that this new agar is a fast, distinctive, and accurate method for enumeration of V. vulnificus from the environment. Copyright © 2013 Elsevier B.V. All rights reserved.

Identification of the Staphylococcus aureus vfrAB operon, a novel virulence factor regulatory locus.

PubMed

Bose, Jeffrey L; Daly, Seth M; Hall, Pamela R; Bayles, Kenneth W

2014-05-01

During a screen of the Nebraska Transposon Mutant Library, we identified 71 mutations in the Staphylococcus aureus genome that altered hemolysis on blood agar medium. Although many of these mutations disrupted genes known to affect the production of alpha-hemolysin, two of them were associated with an apparent operon, designated vfrAB, that had not been characterized previously. Interestingly, a ΔvfrB mutant exhibited only minor effects on the transcription of the hla gene, encoding alpha-hemolysin, when grown in broth, as well as on RNAIII, a posttranscriptional regulatory RNA important for alpha-hemolysin translation, suggesting that VfrB may function at the posttranscriptional level. Indeed, a ΔvfrB mutant had increased aur and sspAB protease expression under these conditions. However, disruption of the known secreted proteases in the ΔvfrB mutant did not restore hemolytic activity in the ΔvfrB mutant on blood agar. Further analysis revealed that, in contrast to the minor effects of VfrB on hla transcription when strains were cultured in liquid media, the level of hla transcription was decreased 50-fold in the absence of VfrB on solid media. These results demonstrate that while VfrB represses protease expression when strains are grown in broth, hla regulation is highly responsive to factors associated with growth on solid media. Intriguingly, the ΔvfrB mutant displayed increased pathogenesis in a model of S. aureus dermonecrosis, further highlighting the complexity of VfrB-dependent virulence regulation. The results of this study describe a phenotype associated with a class of highly conserved yet uncharacterized proteins found in Gram-positive bacteria, and they shed new light on the regulation of virulence factors necessary for S. aureus pathogenesis.

Evaluation of a New Selective Medium, BD BBL CHROMagar MRSA II, for Detection of Methicillin-Resistant Staphylococcus aureus in Stool Specimens ▿

PubMed Central

Havill, Nancy L.; Boyce, John M.

2010-01-01

We compared the recovery of methicillin-resistant Staphylococcus aureus (MRSA) on a new selective chromogenic agar, BD BBL CHROMagar MRSA II (CMRSAII), to that on traditional culture media with 293 stool specimens. The recovery of MRSA was greater on the CMRSAII agar. Screening of stool samples can identify patients who were previously unknown carriers of MRSA. PMID:20392908

Performance of the chromID Salmonella Elite chromogenic agar in comparison with CHROMagar™ Salmonella, Oxoid™ Brilliance™ Salmonella and Hektoen agars for the isolation of Salmonella from stool specimens.

PubMed

Martiny, Delphine; Dediste, Anne; Anglade, Claire; Vlaes, Linda; Moens, Catherine; Mohamed, Souad; Vandenberg, Olivier

2016-10-01

chromID™ Salmonella Elite is compared with 3 culture media commonly used for Salmonella isolation from stool specimens. As results were equivalent to other chromogenic media (100% sensitivity, 98% specificity), only financial arguments should guide the choice for a medium with respect to another. Copyright © 2016 Elsevier Inc. All rights reserved.

A one-step matrix application method for MALDI mass spectrometry imaging of bacterial colony biofilms.

PubMed

Li, Bin; Comi, Troy J; Si, Tong; Dunham, Sage J B; Sweedler, Jonathan V

2016-11-01

Matrix-assisted laser desorption/ionization imaging of biofilms cultured on agar plates is challenging because of problems related to matrix deposition onto agar. We describe a one-step, spray-based application of a 2,5-dihydroxybenzoic acid solution for direct matrix-assisted laser desorption/ionization imaging of hydrated Bacillus subtilis biofilms on agar. Using both an optimized airbrush and a home-built automatic sprayer, region-specific distributions of signaling metabolites and cannibalistic factors were visualized from B. subtilis cells cultivated on biofilm-promoting medium. The approach provides a homogeneous, relatively dry coating on hydrated samples, improving spot to spot signal repeatability compared with sieved matrix application, and is easily adapted for imaging a range of agar-based biofilms. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Hichrom candida agar for identification of Candida species.

PubMed

Baradkar, V P; Mathur, M; Kumar, S

2010-01-01

Chromogenic media are frequently used in direct and rapid identification of yeasts because different Candida species produce unique colors on these media. We used 60 isolates of Candida species including 30 C. albicans, 10 C. parapsilosis, 11 C. glabrata, five C. tropicalis, and four C. dubliniensis, isolated from various clinical specimens, to evaluate the performance of HiChrome Candida agar. These strains had been identified by germ tube test, morphology on cornmeal agar, chlamydospore formation on tobacco agar and sugar assimilation tests. The sensitivity and specificity results were: C. albicans (96.55 and 96.42%); C. parapsilosis (80 and 98.03%), C. glabrata (90.90 and 88.23%), C. tropicalis (100 and 100%) and C. dubliniensis (60 and 96.55%) respectively. HiChrom Candida agaris medium has been useful and capable of presumptive, rapid identification of Candida species within 48 hours.

Colony formation by sublethally heat-injured Zygosaccharomyces rouxii as affected by solutes in the recovery medium and procedure for sterilizing medium.

PubMed Central

Golden, D A; Beuchat, L R

1990-01-01

Recovery and colony formation by healthy and sublethally heat-injured cells of Zygosaccharomyces rouxii as influenced by the procedure for sterilizing recovery media (YM agar [YMA], wort agar, cornmeal agar, and oatmeal agar) were investigated. Media were supplemented with various concentrations of glucose, sucrose, glycerol, or sorbitol and sterilized by autoclaving (110 degrees C, 15 min) and by repeated treatment with steam (100 degrees C). An increase in sensitivity was observed when heat-injured cells were plated on glucose-supplemented YMA at an aw of 0.880 compared with aws of 0.933 and 0.998. Colonies which developed from unheated and heated cells on YMA at aws of 0.998 and 0.933 generally exceeded 0.5 mm in diameter within 3.5 to 4 days of incubation at 25 degrees C, whereas colonies formed on YMA at an aw of 0.880 typically did not exceed 0.5 mm in diameter until after 5.5 to 6.5 days of incubation. The number of colonies exceeding 0.5 mm in diameter which were formed by heat-injured cells on YMA at an aw of 0.880 was 2 to 3 logs less than the total number of colonies detected, i.e., on YMA at an aw of 0.933 and using no limits of exclusion based on colony diameter. A substantial portion of cells which survived heat treatment were sublethally injured as evidenced by increased sensitivity to a suboptimum aw (0.880). In no instance was recovery of Z. rouxii significantly affected by medium sterilization procedure when glucose or sorbitol was used as the aw-suppressing solute.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:2403251

Colony formation by sublethally heat-injured Zygosaccharomyces rouxii as affected by solutes in the recovery medium and procedure for sterilizing medium.

PubMed

Golden, D A; Beuchat, L R

1990-08-01

Recovery and colony formation by healthy and sublethally heat-injured cells of Zygosaccharomyces rouxii as influenced by the procedure for sterilizing recovery media (YM agar [YMA], wort agar, cornmeal agar, and oatmeal agar) were investigated. Media were supplemented with various concentrations of glucose, sucrose, glycerol, or sorbitol and sterilized by autoclaving (110 degrees C, 15 min) and by repeated treatment with steam (100 degrees C). An increase in sensitivity was observed when heat-injured cells were plated on glucose-supplemented YMA at an aw of 0.880 compared with aws of 0.933 and 0.998. Colonies which developed from unheated and heated cells on YMA at aws of 0.998 and 0.933 generally exceeded 0.5 mm in diameter within 3.5 to 4 days of incubation at 25 degrees C, whereas colonies formed on YMA at an aw of 0.880 typically did not exceed 0.5 mm in diameter until after 5.5 to 6.5 days of incubation. The number of colonies exceeding 0.5 mm in diameter which were formed by heat-injured cells on YMA at an aw of 0.880 was 2 to 3 logs less than the total number of colonies detected, i.e., on YMA at an aw of 0.933 and using no limits of exclusion based on colony diameter. A substantial portion of cells which survived heat treatment were sublethally injured as evidenced by increased sensitivity to a suboptimum aw (0.880). In no instance was recovery of Z. rouxii significantly affected by medium sterilization procedure when glucose or sorbitol was used as the aw-suppressing solute.(ABSTRACT TRUNCATED AT 250 WORDS)

Interaction of the nematophagous fungus Pochonia chlamydosporia and Parascaris equorum eggs in different culture media.

PubMed

de Carvalho, Lorendane Millena; Braga, Fabio Ribeiro; Domingues, Rafael Reis; Araujo, Juliana Milani; Lelis, Rosane Teixeira; de Paula, Alessandra Teixeira; da Silveira, Wendeo Ferreira; de Araújo, Jackson Victor

2014-07-01

Research involving the use of nematophagous fungi in the biological control of parasites of interest to veterinarians has occurred over recent years, with promising results. This article reports the infection of Parascaris equorum eggs by the fungus Pochonia chlamydosporia (isolates VC1 and VC4). Six groups were formed for each isolate, with six different culture media: 2% water-agar (2% WA); agar-chitin (AC); YPSSA (yeast extract, K2HPO4, MgSO4 ·7H2O, soluble starch); AELA extract (starch + water + agar); 2% corn-meal-agar (2% CMA); and 2% potato dextrose-agar (2% PDA). A total of 1000 eggs of P. equorum were transferred to each plate containing isolates grown for a period of 7 days (treatment group). Also, 1000 eggs were added to each plate without fungus (controlgroup). The plates were kept in an environmental chamber at 25 °C in the dark for 21 days. After, we analyzed the effects on ovicidal activity: effect 1 (accession shell); effect 2 (penetration hyphae); and effect 3 (destruction of the eggs). No differences were observed in the destruction of eggs between the two isolates. The decreasing effectiveness of the different culture media was: PDA (38.9%); CMA (38.3%); WA (36.7%); YPSSA (36.45%); and AC (32.5%). The highest percentage egg destruction was observed when the strains were grown in culture medium AELA (44.9%); this was the best medium. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Influence of culture media and environmental factors on mycelial growth and pycnidial production of Sphaeropsis pyriputrescens.

PubMed

Kim, Y K; Xiao, C L; Rogers, J D

2005-01-01

Sphaeropsis pyriputrescens, the causal agent of Sphaeropsis rot of pears and apples, is a recently described species. In this study the effects of culture media, temperature, water potential, pH and light on mycelial growth and pycnidial production of S. pyriputrescens were evaluated. Apple juice agar and pear juice agar were most suitable for mycelial growth of all six isolates tested. Cornmeal agar was not suitable for either mycelial growth or pycnidial production. The fungus grew from -3 to 25 C, with optimum growth at 20 C and no growth at 30 C. The fungus grew at water potential as low as -5.6 MPa on potassium chloride-amended potato-dextrose agar (PDA). Hyphal extension was not observed at -7.3 MPa after 10 d incubation, but growth resumed when the inoculum plugs were placed on PDA. The fungus grew at pH 3.3-6.3 and optimum growth was at pH 3.3-4.2. No mycelial growth was observed at pH above 7.2 after 10 d incubation, but growth resumed when the inoculum plugs were transferred onto PDA. Regardless of medium tested, few pycnidia formed at 20 C in the dark. Pycnidial production was enhanced significantly by fluorescent light, but continuous light appeared to reduce pycnidial production, depending on the medium. Oatmeal agar (OMA) was most suitable for production of pycnidia and conidia. Pycnidia that formed on 3 wk old OMA cultures at 20 C under 12 h light/12 h dark produced abundant conidia, and the technique is recommended for inoculum production.

[Selective-differential nutrient medium "Shewanella IRHLS agar" for isolation of Shewanella genus bacteria].

PubMed

Sivolodsky, E P

2015-01-01

Development of a selective-differential nutrient medium for isolation of Shewanella genus bacteria. 73 strains of Shewanella bacteria (S. algae--3, S. baltica--26, S. putrefaciens--44) and 80 strains of 22 other bacteria genera were used. Shewanella species were identified by methods and criteria proposed by Nozue H. et al., 1992; Khashe S. et al., 1998. Nutrient media "Shewanella IRHLS Agar" for shewanella isolation was developed. Medium selective factors: irgazan DP-300 (I). 0.14-0.2 g/l and rifampicin (R) 0.0005-0.001 g/l. Shevanella colonies were detected by the production of hydrogen sulfide (H), lipase presence (L), lack of sorbitol fermentation (S). The medium suppressed the growth of hydrogen sulfide producers (Salmonella, Proteus) and blocked hydrogen sulfide production by Citrobacter. Growth of Escherichia, Enterobacter, Klebsiella, Shigella, Staphylococcus, Bacillus was also suppressed, Analytical sensitivity of the medium was 1-2 CFU/ml for Shewanella and Stenotrophomonas, Aerombnas, Serratia genera bacteria. 72 strains of Shewanella were isolated from water of Neva river in this medium, 91.7 ± 3.2% of those produced H2S. 1 strain of S. algae was isolated from clinical material. The developed media allows to use it in a complex for Stenotrophomo- nas sp., Aeromonas sp., Serratia sp., Citrobactersp. and Shewanella bacteria isolation.

Use of headspace SPME-GC-MS for the analysis of the volatiles produced by indoor molds grown on different substrates.

PubMed

Van Lancker, Fien; Adams, An; Delmulle, Barbara; De Saeger, Sarah; Moretti, Antonio; Van Peteghem, Carlos; De Kimpe, Norbert

2008-10-01

An automated headspace solid phase microextraction method followed by GC-MS analysis was used to evaluate and compare the in vitro production of microbial volatile organic compounds (MVOCs) on malt extract agar, plasterboard and wallpaper. Five fungal strains were isolated from the walls of water-damaged houses and identified. In addition, four other common molds were studied. In general, MVOC production was the highest on malt extract agar. On this synthetic medium, molds typically produced 2-methylpropanol, 2-methylbutanol and 3-methylbutanol. On wallpaper, mainly 2-ethylhexanol, methyl 2-ethylhexanoate and compounds of the C8-complex such as 1-octene-3-ol, 3-octanone, 3-octanol and 1,3-octadiene were detected. The detection of 2-ethylhexanol and methyl 2-ethylhexanoate indicates an enhanced degradation of the substrate by most fungi. For growth on plasterboard, no typical metabolites were detected. Despite these metabolite differences on malt extract agar, wallpaper and plasterboard, some molds also produced specific compounds independently of the used substrate, such as trichodiene from Fusarium sporotrichioides and aristolochene from Penicillium roqueforti. Therefore, these metabolites can be used as markers for the identification and maybe also mycotoxin production of these molds. All five investigated Penicillium spp. in this study were able to produce two specific diterpenes, which were not produced by the other species studied. These two compounds, which remain unidentified until now, therefore seem specific for Penicillium spp. and are potentially interesting for the monitoring of this fungal genus. Further experiments will be performed with other Penicillium spp. to study the possibility that these two compounds are specific for this group of molds.

[TMOSKOVHE COMPARATIVE CHARACTERISTIC OF GROWTH MEDIUMS FOR SEPARATION OF CORYNEBACTERIA].

PubMed

Shepelin, A P; Polosenko, O V; Borisova, O Yu; Pimenova, A S; Gadua, N T

2016-01-01

The comparative tests of growth mediums for isolation and accumulation of diphtheria bacteria were implemented. The testing consisted of six series of growth medium "Corynebacagar" produced by the state research center of applied microbiology and biotechnology and three series of blood tellurite agar. The concluding results of identification of biological indicators of all series of growth nutrient mediums are presented The "Corynebacagar" is recommended for application in health care practice for primary inoculation of pathological material during implementation of cultural analysis on diphtheria.

[Production of infectious units of Isaria fumosorosea (Hypocreales: Cordycipitaceae) from different indigenous isolates of northeastern Mexico using 3 propagation strategies].

PubMed

Gandarilla-Pacheco, Fatima L; Morales-Ramos, Lilia H; Pereyra-Alférez, Benito; Elías-Santos, Myriam; Quintero-Zapata, Isela

The aim of this study was to evaluate the production of blastospores and conidia of different native isolates and a strain of Isaria fumosorosea using different propagation techniques. Two liquid culture media of casamino acids and peptone as nitrogen sources and glucose as carbon source for both media cultures were respectively used in the production of blastospores, while for the production of conidia, the fungi were grown in potato dextrose agar; from these cultures, solutions of conidia to a concentration of 1×10 6 per milliliter were prepared to inoculate flasks with Sabouraud dextrose broth for the liquid phase of the biphasic culture, also known as preculture. Subsequently, rice grain bags were inoculated with the preculture and the conidia solutions, which were incubated for 14 days for solid fermentation and biphasic culture, respectively. The HIB-23 isolate recorded a concentration of 4.90×10 8 blastospores/ml in the casamino acid medium, while a concentration of 2.15×10 8 blastospores/ml was obtained in the peptone collagen medium. For the Pfr-612 strain, the conidia production in solid-state fermentation was 1.58×10 9 conidia/g, and for HIB-30 in the biphasic culture of 9.00×10 6 conidia/g. Solid-state fermentation proved to be the most effective method with an average of 1.09×10 9 conidia/g, whereas the biphasic culture was the least effective method with 2.76×10 6 conidia/g; no significant difference was reported for the submerged production media. Copyright © 2017 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.

Performance characteristics and estimation of measurement uncertainty of three plating procedures for Campylobacter enumeration in chicken meat.

PubMed

Habib, I; Sampers, I; Uyttendaele, M; Berkvens, D; De Zutter, L

2008-02-01

In this work, we present an intra-laboratory study in order to estimate repeatability (r), reproducibility (R), and measurement uncertainty (U) associated with three media for Campylobacter enumeration, named, modified charcoal cefoperazone deoxycholate agar (mCCDA); Karmali agar; and CampyFood ID agar (CFA) a medium by Biomérieux SA. The study was performed at three levels: (1) pure bacterial cultures, using three Campylobacter strains; (2) artificially contaminated samples from three chicken meat matrixes (total n=30), whereby samples were spiked using two contamination levels; ca. 10(3)cfuCampylobacter/g, and ca. 10(4)cfuCampylobacter/g; and (3) pilot testing in naturally contaminated chicken meat samples (n=20). Results from pure culture experiment revealed that enumeration of Campylobacter colonies on Karmali and CFA media was more convenient in comparison with mCCDA using spread and spiral plating techniques. Based on artificially contaminated samples testing, values of repeatability (r) were comparable between the three media, and estimated as 0.15log(10)cfu/g for mCCDA, 0.14log(10)cfu/g for Karmali, and 0.18log(10)cfu/g for CFA. As well, reproducibility performance of the three plating media was comparable. General R values which can be used when testing chicken meat samples are; 0.28log(10), 0.32log(10), and 0.25log(10) for plating on mCCDA, Karmali agar, and CFA, respectively. Measurement uncertainty associated with mCCDA, Karmali agar, and CFA using spread plating, for combination of all meat matrixes, were +/-0.24log(10)cfu/g, +/-0.28log(10)cfu/g, and +/-0.22log(10)cfu/g, respectively. Higher uncertainty was associated with Karmali agar for Campylobacter enumeration in artificially inoculated minced meat (+/-0.48log(10)cfu/g). The general performance of CFA medium was comparable with mCCDA performance at the level of artificially contaminated samples. However, when tested at naturally contaminated samples, non-Campylobacter colonies gave similar deep red colour as that given by the typical Campylobacter growth on CFA. Such colonies were not easily distinguishable by naked eye. In general, the overall reproducibility, repeatability, and measurement uncertainty estimated by our study indicate that there are no major problems with the precision of the International Organization for Standardization (ISO) 10272-2:2006 protocol for Campylobacter enumeration using mCCDA medium.

Detection of Agar, by Analysis of Sugar Markers, Associated with Bacillus Anthracis Spores, After Culture

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wunschel, David S.; Colburn, Heather A.; Fox, Alvin

2008-08-01

Detection of small quantities of agar associated with spores of Bacillus anthracis could provide key information regarding its source or growth characteristics. Agar, widely used in growth of bacteria on solid surfaces, consists primarily of repeating polysaccharide units of 3,6-anhydro-L-galactose (AGal) and galactose (Gal) with sulfated and O-methylated galactoses present as minor constituents. Two variants of the alditol acetate procedure were evaluated for detection of potential agar markers associated with spores. The first method employed a reductive hydrolysis step, to stabilize labile anhydrogalactose, by converting to anhydrogalactitol. The second eliminated the reductive hydrolysis step simplifying the procedure. Anhydrogalactitol, derived frommore » agar, was detected using both derivatization methods followed by gas chromatography-mass spectrometry (GC-MS) analysis. However, challenges with artefactual background (reductive hydrolysis) or marker destruction (hydrolysis) lead to the search for alternative sugar markers. A minor agar component, 6-O-methyl galactose (6-O-M gal), was readily detected in agar-grown but not broth-grown bacteria. Detection was optimized by the use of gas chromatography-tandem mass spectrometry (GC-MS-MS). With appropriate choice of sugar marker and analytical procedure, detection of sugar markers for agar has considerable potential in microbial forensics.« less

Isolation and characterization of a novel agarase-producing Pseudoalteromonas spp. bacterium from the guts of spiny turban shells.

PubMed

Oh, Young Hoon; Jung, Changkyou; Lee, Jinwon

2011-08-01

An agar-degrading bacterium was isolated from the guts of spiny turban shells. It was identified as a Pseudoalteromonas species and named Pseudoalteromonas sp. JYBCL 1. The viscosity of the inoculated agar medium decreased by more than 60% after 20 h cultivation. The agarase produced by the isolate had optimal activities at 35 degrees C and pH 7. The enzyme had extremely strong resistance to ionic stress compared with other known agarases. Its molecular mass was estimated at about 60 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The agarase could saccharify Gelidium amansii directly, with an efficiency about half that compared with agar saccharification.

Fabrication and characterization of biotissue-mimicking phantoms in the THz frequency range

NASA Astrophysics Data System (ADS)

Liakhov, E.; Smolyanskaya, O.; Popov, A.; Odlyanitskiy, E.; Balbekin, N.; Khodzitsky, M.

2016-08-01

The study revealed the most promising candidates for phantoms mimicking different biological tissues in the terahertz frequency range. Closest to biological tissues in terms of the refractive index appeared to be gelatin-based gels; in terms of the absorption coefficient they were agar-based gels. Gelatin is more stable in time, but requires special storage conditions to limit water evaporation. The dense structure of the agar-based phantom allows its use without mold and risk of damage. However, agar is a nutrient medium for bacteria and its parameters degrade even when the phantom form and water content are retained. Use of liquid suspensions of lecithin and milk powder are found to be extremely limited.

Development and validation of a large, modular test meal with liquid and solid components for assessment of gastric motor and sensory function by non-invasive imaging.

PubMed

Parker, H L; Tucker, E; Hoad, C L; Pal, A; Costigan, C; Hudders, N; Perkins, A; Blackshaw, E; Gowland, P; Marciani, L; Fox, M R

2016-04-01

Current investigations of stomach function are based on small test meals that do not reliably induce symptoms and analysis techniques that rarely detect clinically relevant dysfunction. This study introduces the large 'Nottingham Test Meal' (NTM) for assessment of gastric motor and sensory function by non-invasive imaging. NTM comprises 400 mL liquid nutrient (0.75 kcal/mL) and 12 solid agar-beads (0 kcal) with known breaking strength. Gastric fullness and dyspeptic sensations were documented by 100 mm visual analogue scale (VAS). Gastric emptying (GE) were measured in 24 healthy volunteers (HVs) by gastric scintigraphy (GS) and magnetic resonance imaging (MRI). The contribution of secretion to gastric volume was assessed. Parameters that describe GE were calculated from validated models. Inter-observer agreement and reproducibility were assessed. NTM produced moderate fullness (VAS ≥30) but no more than mild dyspeptic symptoms (VAS <30) in 24 HVs. Stable binding of meal components to labels in gastric conditions was confirmed. Distinct early and late-phase GE were detected by both modalities. Liquid GE half-time was median 49 (95% CI: 36-62) min and 68 (57-71) min for GS and MRI, respectively. Differences between GS and MRI measurements were explained by the contribution of gastric secretion. Breaking strength for agar-beads was 0.8 N/m(2) such that median 25 (8-50) % intact agar-beads and 65 (47-74) % solid material remained at 120 min on MRI and GS, respectively. Good reproducibility for liquid GE parameters was present and GE was not altered by agar-beads. The NTM provided an objective assessment of gastric motor and sensory function. The results were reproducible and liquid emptying was not affected by non-nutrient agar-beads. The method is potentially suitable for clinical practice. © 2016 John Wiley & Sons Ltd.

Spatiotemporal Patterns Produced by Bacteria

NASA Astrophysics Data System (ADS)

Shimada, Yuji; Nakahara, Akio; Matsushita, Mitsugu; Matsuyama, Tohey

1995-06-01

Spatiotemporal patterns formed by a bacterial colony of Proteus mirabilis on an agar plate were observed. About half or one hour after the colony spread over the entire surface of the agar medium in a petridish, various patterns including target and spiral patterns appeared. They are very similar to those seen in other dissipative systems, such as chemical oscillations and electrohydrodynamic convective systems. Microscopic observations revealed that the collective motion of bacterial cells is responsible for the formation of these spatiotemporal patterns.

Broth and agar hop-gradient plates used to evaluate the beer-spoilage potential of Lactobacillus and Pediococcus isolates.

PubMed

Haakensen, M; Schubert, A; Ziola, B

2009-03-15

Identification of the beer-spoilage Lactobacillus and Pediococcus bacteria has largely taken two approaches; identification of spoilage-associated genes or identification of specific species of bacteria regardless of ability to grow in beer. The problem with these two approaches is that they are either overly inclusive (i.e., detect all bacteria of a given species regardless of spoilage potential) or overly selective (i.e., rely upon individual, putative spoilage-associated genes). Our goal was to design a method to assess the ability of Lactobacillus and Pediococcus to spoil beer that is independent of speciation or genetic background. In searching for a method by which to differentiate between beer-spoilage bacteria and bacteria that cannot grow in beer, we explored the ability of lactobacilli and pediococci isolates to grow in the presence of varying concentrations of hop-compounds and ethanol in broth medium versus on agar medium. The best method for differentiating between bacteria that can grow in beer and bacteria that do not pose a threat as beer-spoilage organisms was found to be a hop-gradient agar plate containing ethanol. This hop-gradient agar plate technique provides a rapid and simple solution to the dilemma of assessing the ability of Lactobacillus and Pediococcus isolates to grow in beer, and provides new insights into the different strategies used by these bacteria to survive under the stringent conditions of beer.

Anti-L. donovani activity in macrophage/amastigote model of palmarumycin CP18 and its large scale production.

PubMed

Ortega, Humberto E; Teixeira, Eliane de Morais; Rabello, Ana; Higginbotham, Sarah; Cubilla-Ríos, Luis

2014-01-01

Palmarumycin CP18, isolated from an extract of the fermentation broth and mycelium of the Panamanian endophytic fungus Edenia sp., was previously reported with strong and specific activity against Leishmania donovani. Here we report that when the same strain was cultured on different solid media--Harrold Agar, Leonian Agar, Potato dextrose Agar (PDA), Corn Meal Agar, Honey Peptone Agar, and eight vegetables (V8) Agar--in order to determine the optimal conditions for isolation of palmarumycin CP18, no signal for this compound was observed in any of the 1H NMR spectra of fractions obtained from these extracts. However, one extract, prepared from the fungal culture in PDA contained significant amounts of CJ-12,372, a possible biosynthetic precursor of palmarumycin CP18. Edenia sp. was cultivated on a large scale on PDA and CJ-12,372 was converted to palmarumycin CP18 by oxidation of its p-hydroquinone moiety with DDQ in dioxane. Palmarumycin CP18 showed anti-leishmanial activity against L. donovani in a macrophage/amastigote model, with IC50 values of 23.5 microM.

[Screening and identification of a bacterium capable of converting agar to neoagaro oligosaccharides].

PubMed

Han, Junping; Huang, Yayan; Ye, Jing; Xiao, Meitian

2015-09-04

To screen and identify a bacterium capable of converting agar to neoagaro oligosaccharides. We took samples of porphyra haitanensis and nearby seawater, and then used the medium containing 1 per thousand agar to enrich the target bacteria. The target isolates were obtained by dilution-plate method, of which crude enzymes were further obtained by liquid culture. We adopted DNS method to determine the target bacteria which can convert agar to neoagaro oligosaccharides. The phylogenetics was identified by analyzing 16S rDNA sequence and combining the strain's morphological and bacterial colonial physiological biochemical characteristics. We isolated a gram-negative bacterial strain HJPHYXJ-1 capable of transforming agar to neoagaro oligosaccharides. Basic Local Alignment Search Tool (BLAST) search of HJPHYXJ-1's 16S rDNA sequence on GenBank suggested that the similarity between this strain and Vibrio natriegens reached 99% . In addition, the morphological and physiological biochemical characteristics of HJPHYXJ-1 also showed highly similarity to Vibrio natriegens. So we identified HJPHYXJ-1 as Vibrio natriegens. The results of HPLC suggested that the metabolite of enzymatic degradation was neoagaro oligosaccharides. HJPHYXJ-1 or the new isolate of Vibrio natriegens was capable of converting agar to neoagaro oligosaccharides.

Effect of specimen storage, antibiotics, and feminine hygiene products on the detection of group B Streptococcus by culture and the STREP B OIA test.

PubMed

Ostroff, R M; Steaffens, J W

1995-07-01

Agar culture from vaginal swabs is the routine method for diagnosis of maternal Group B Streptococcus (GBS) colonization. Swab specimens are often transported to a clinical laboratory for processing. In these studies, specimen transport was simulated by inoculating swabs with GBS and storing them at selected temperatures and with or without transport medium. The recovery of viable GBS was assessed by agar culture. GBS antigen was detected immunologically with an Optical ImmunoAssay (OIA) method. Swabs that were stored with transport medium harbored viable but rapidly declining numbers of GBS. In contrast, a strong OIA signal was maintained. Recovery of viable GBS organisms declined more quickly when swabs were stored in the absence of transport medium, whereas detection of GBS antigen remained consistent. Both methods were tested for interference from either antibiotics or feminine hygiene products. These compounds inhibited the detection of GBS by culture but had no detrimental effect on the OIA result.

Characterization of nitrate-reducing and amino acid-using bacteria prominent in nitrotoxin-enriched equine cecal populations

USDA-ARS?s Scientific Manuscript database

In the present study, populations of equine cecal microbes enriched for enhanced rates of 3-nitro-1-propionic acid (NPA) or nitrate metabolism were diluted and cultured for NPA-metabolizing bacteria on a basal enrichment medium (BEM) or tryptose soy agar (TSA) medium supplemented with either 5 mM NP...

Intelligent pH indicator film composed of agar/potato starch and anthocyanin extracts from purple sweet potato.

PubMed

Choi, Inyoung; Lee, Jun Young; Lacroix, Monique; Han, Jaejoon

2017-03-01

A new colorimetric pH indicator film was developed using agar, potato starch, and natural dyes extracted from purple sweet potato, Ipomoea batatas. Both agar and potato starch are solid matrices used to immobilize natural dyes, anthocyanins. The ultraviolet-visible (UV-vis) spectrum of anthocyanin extract solutions and agar/potato starch films with anthocyanins showed color variations to different pH values (pH 2.0-10.0). Fourier transform infrared (FT-IR) and UV-vis region spectra showed compatibility between agar, starch, and anthocyanin extracts. Color variations of pH indicator films were measured by a colorimeter after immersion in different pH buffers. An application test was conducted for potential use as a meat spoilage sensor. The pH indicator films showed pH changes and spoilage point of pork samples, changing from red to green. Therefore, the developed pH indicator films could be used as a diagnostic tool for the detection of food spoilage. Copyright © 2016 Elsevier Ltd. All rights reserved.

Periodic Pattern Formation of Bacterial Colonies

NASA Astrophysics Data System (ADS)

Itoh, Hiroto; Wakita, Jun-ichi; Matsuyama, Tohey; Matsushita, Mitsugu

1999-04-01

We have experimentally investigated pattern formation of colonies ofbacterial species Proteus mirabilis, which is famous forforming concentric-ring-like colonies.The colony grows cyclically with the interface repeating an advance anda stop alternately on a surface of a solid agar medium.We distinguish three phases (initial lag phase, the followingmigration and consolidation phases that appear alternately) for the colony growth.When we cut a colony just behind a migrating front shortly after the migrationstarted, the migration ended earlier and the following consolidationlasted longer.However, the following cycles were not influenced by the cut, i.e., thephases of the migration and consolidation were not affected.Global chemical signals governing the colony formation from thecenter were not found to exist.We also quantitatively checked phase entrainment by letting two coloniescollide with each other and found that it does not take place in macroscopic scales.All these experimental results suggest that the most important factorfor the migration is the cell population density.

Screening for Indian isolates of egg-parasitic fungi for use in biological control of fascioliasis and amphistomiasis in ruminant livestock.

PubMed

De, S; Sanyal, P K; Sarkar, A K; Patel, N K; Pal, S; Mandal, S C

2008-09-01

Wild isolates of the egg-parasitic fungi Paecilomyces lilacinus and Verticillium chlamydosporium, obtained from the organic environment of Durg, Chhattisgarh, India, were subjected to screening for in vitro growth using different media types, range of incubation temperature and pH, and their predatory activity to the eggs of Fasciola gigantica and Gigantocotyle explanatum. Maximum growth of P. lilacinus was obtained in corn-meal agar compared to any other media types. The preferred medium for growth of V. chlamydosporium was corn-meal agar, followed by potato-dextrose agar. After initial growth for 16 h of incubation, no growth was observed in water agar for both the fungi. Six different temperatures--4 degrees C, 10 degrees C, 18 degrees C, 26 degrees C, 34 degrees C and 40 degrees C--were used to observe growth profiles of the fungi in corn-meal agar medium. While no and very little growth of P. lilacinus and V. chlamydosporium was observed at 4 degrees C and 10 degrees C, respectively, growth profiles of both the fungi were optimal at 26-40 degrees C. A range of pH (pH 4-8) supported growth of both P. lilacinus and V. chlamydosporium. Full-grown plates of the fungi baited with viable eggs of F. gigantica and G. explanatum revealed that V. chlamydosporium was more vigorous in its egg-parasitic ability compared to P. lilacinus. Distortion of the eggs started on day 2-3 of egg baiting in culture plates of V. chlamydosporium, with complete distortion by day 7. On the contrary, P. lilacinus exhibited very limited egg-parasitic ability and some of the baited eggs even showed development of miracidia.

Performance of Chromogenic Candida agar and CHROMagar Candida in recovery and presumptive identification of monofungal and polyfungal vaginal isolates.

PubMed

Ozcan, Kadri; Ilkit, Macit; Ates, Aylin; Turac-Bicer, Aygul; Demirhindi, Hakan

2010-02-01

Chromogenic Candida agar (OCCA) is a novel medium facilitating isolation and identification of Candida albicans, C. tropicalis, and C. krusei, as well as indicating polyfungal population in clinical samples. We compare the performance of OCCA, to CHROMagar Candida (CAC) and Sabouraud chloramphenicol agar (SCA). Vaginal swab samples from 392 women were simultaneously inoculated onto three study media. A total of 161 (41.1%) were found to be positive for fungi of which 140 (87%) were monofungal, and 21 (13%) polyfungal. One-hundred and fifty-seven samples (97.5%) were positive on CAC, 156 (96.9%) on OCCA, 148 (91.9%) on SCA and 144 (89.4%) samples were positive on all three media. The yeasts were identified by conventional methods including germ tube test, microscopic morphology on cornmeal-Tween 80 agar, and the commercial API 20C AUX. The 182 isolates were C. albicans (n = 104), C. glabrata (n = 51), C. krusei (n = 7), C. tropicalis (n = 5), C. famata (n = 3), C. kefyr (n = 3), C. zeylanoides (n = 3), C. colliculosa (n = 2), and other species of Candida (n = 4). Among the 21 polyfungal populations, 20 (95.2%) were detected in OCCA, 14 (66.7%) in CAC, and 13 (61.9%) in CAC and OCCA (P <0.05). Most polyfungal populations (47.6%) yielded C. albicans + C. glabrata. The efficiency of both chromogenic media for C. albicans was >or=92.9% at 72 h. OCCA is more efficient and reliable for rapidly identifying C. albicans and polyfungal populations than CAC. However, CAC is more efficient for identifying C. krusei and C. tropicalis. A chromogenic agar with a higher isolation rate of yeasts and better detection of polyfungal populations than SCA, is suggested as a medium of first choice when available.

Incubation at room temperature may be an independent factor that induces chlamydospore production in Candida dubliniensis.

PubMed

Sancak, Banu; Colakoglu, Sule; Acikgoz, Ziya Cibali; Arikan, Sevtap

2005-08-01

Production of chlamydospores is one of the phenotypic features used to differentiate Candida albicans and Candida dubliniensis. C. albicans produces few chlamydospores on only cornmeal/rice-Tween agar at room temperature, whereas C. dubliniensis produces abundant chlamydospores at this temperature both on cornmeal agar and some other commonly used media. We tried to determine whether the room temperature is the main factor that induces chlamydospore production of C. dubliniensis, regardless of the medium used. For this purpose, 100 C. albicans and 24 C. dubliniensis isolates were tested for chlamydospore production at room temperature and at 37 degrees C on some routinely used media, including eosin-methylene blue agar (EMB), nutrient agar (NA), nutrient broth (NB), and also on an investigational medium, phenol red agar (PR). At 37 degrees C, none of the isolates produced chlamydospores on any of the tested media. At 26 degrees C, all C. dubliniensis isolates produced abundant chlamydospores and pseudohyphae after 24-48 h on all tested media. At this incubation temperature, all C. albicans isolates failed to produce chlamydospores and pseudohyphae on EMB, NA, and NB, whereas 2 of the C. albicans isolates produced a few chlamydospores on PR. We also observed that all C. dubliniensis isolates tested on EMB and PR produced rough colonies with a hyphal fringe around the colonies, whereas none of the C. albicans isolates showed this property. In conclusion, incubation at 26 degrees C may play the key role for production of abundant chlamydospores and pseudohyphae by C. dubliniensis. Comprehensive molecular studies are needed to clarify the genetic basis of this observation. Using EMB and PR may be an inexpensive, a time-saving, and a simple way of presumptive identification of C. dubliniensis based on chlamydospore formation and colony morphology.

Enrichment and isolation of Bacillus beveridgei sp. nov., a facultative anaerobic haloalkaliphile from Mono Lake, California, that respires oxyanions of tellurium, selenium, and arsenic

USGS Publications Warehouse

Baesman, S.M.; Stolz, J.F.; Kulp, T.R.; Oremland, R.S.

2009-01-01

Mono Lake sediment slurries incubated with lactate and tellurite [Te(IV)] turned progressively black with time because of the precipitation of elemental tellurium [Te(0)]. An enrichment culture was established from these slurries that demonstrated Te(IV)-dependent growth. The enrichment was purified by picking isolated black colonies from lactate/Te(IV) agar plates, followed by repeated streaking and picking. The isolate, strain MLTeJB, grew in aqueous Te(IV)-medium if provided with a small amount of sterile solid phase material (e.g., agar plug; glass beads). Strain MLTeJB grew at high concentrations of Te(IV) (~8 mM) by oxidizing lactate to acetate plus formate, while reducing Te(IV) to Te(0). Other electron acceptors that were found to sustain growth were tellurate, selenate, selenite, arsenate, nitrate, nitrite, fumarate and oxygen. Notably, growth on arsenate, nitrate, nitrite and fumarate did not result in the accumulation of formate, implying that in these cases lactate was oxidized to acetate plus CO2. Strain MLTeJB is a low G + C Gram positive motile rod with pH, sodium, and temperature growth optima at 8.5-9.0, 0.5-1.5 M, and 40??C, respectively. The epithet Bacillus beveridgei strain MLTeJBT is proposed. ?? 2009 Springer.

Inhibition of Listeria monocytogenes by Food-Borne Yeasts†

PubMed Central

Goerges, Stefanie; Aigner, Ulrike; Silakowski, Barbara; Scherer, Siegfried

2006-01-01

Many bacteria are known to inhibit food pathogens, such as Listeria monocytogenes, by secreting a variety of bactericidal and bacteriostatic substances. In sharp contrast, it is unknown whether yeast has an inhibitory potential for the growth of pathogenic bacteria in food. A total of 404 yeasts were screened for inhibitory activity against five Listeria monocytogenes strains. Three hundred and four of these yeasts were isolated from smear-ripened cheeses. Most of the yeasts were identified by Fourier transform infrared spectroscopy. Using an agar-membrane screening assay, a fraction of approximately 4% of the 304 red smear cheese isolates clearly inhibited growth of L. monocytogenes. Furthermore, 14 out of these 304 cheese yeasts were cocultivated with L. monocytogenes WSLC 1364 on solid medium to test the antilisterial activity of yeast in direct cell contact with Listeria. All yeasts inhibited L. monocytogenes to a low degree, which is most probably due to competition for nutrients. However, one Candida intermedia strain was able to reduce the listerial cell count by 4 log units. Another four yeasts, assigned to C. intermedia (three strains) and Kluyveromyces marxianus (one strain), repressed growth of L. monocytogenes by 3 log units. Inhibition of L. monocytogenes was clearly pronounced in the cocultivation assay, which simulates the conditions and contamination rates present on smear cheese surfaces. We found no evidence that the unknown inhibitory molecule is able to diffuse through soft agar. PMID:16391059

In Vitro Effect of Zingiber officinale Extract on Growth of Streptococcus mutans and Streptococcus sanguinis.

PubMed

Azizi, Arash; Aghayan, Shabnam; Zaker, Saeed; Shakeri, Mahdieh; Entezari, Navid; Lawaf, Shirin

2015-01-01

Background and Objectives. Tooth decay is an infectious disease of microbial origin. Considering the increasing prevalence of antibiotic resistance due to their overuse and also their side effects, medicinal plants are now considered for use against bacterial infections. This study aimed to assess the effects of different concentrations of Zingiber officinale extract on proliferation of Streptococcus mutans and Streptococcus sanguinis in vitro. Materials and Methods. In this experimental study, serial dilutions of the extract were prepared in two sets of 10 test tubes for each bacterium (total of 20). Standard amounts of bacterial suspension were added; 100ƛ of each tube was cultured on prepared solid agar plates and incubated at 37°C for 24 hours. Serial dilutions of the extract were prepared in another 20 tubes and 100ƛ of each tube was added to blood agar culture medium while being prepared. The mixture was transferred to the plates. The bacteria were inoculated on plates and incubated as described. Results. The minimum inhibitory concentration (MIC) was 0.02 mg/mL for S. mutans and 0.3 mg/mL for S. sanguinis. The minimum bactericidal concentration (MBC) was 0.04 mg for S. mutans and 0.6 mg for S. sanguinis. Conclusion. Zingiber officinale extract has significant antibacterial activity against S. mutans and S. sanguinis cariogenic microorganisms.

In Vitro Effect of Zingiber officinale Extract on Growth of Streptococcus mutans and Streptococcus sanguinis

PubMed Central

Azizi, Arash; Aghayan, Shabnam; Zaker, Saeed; Shakeri, Mahdieh; Entezari, Navid; Lawaf, Shirin

2015-01-01

Background and Objectives. Tooth decay is an infectious disease of microbial origin. Considering the increasing prevalence of antibiotic resistance due to their overuse and also their side effects, medicinal plants are now considered for use against bacterial infections. This study aimed to assess the effects of different concentrations of Zingiber officinale extract on proliferation of Streptococcus mutans and Streptococcus sanguinis in vitro. Materials and Methods. In this experimental study, serial dilutions of the extract were prepared in two sets of 10 test tubes for each bacterium (total of 20). Standard amounts of bacterial suspension were added; 100ƛ of each tube was cultured on prepared solid agar plates and incubated at 37°C for 24 hours. Serial dilutions of the extract were prepared in another 20 tubes and 100ƛ of each tube was added to blood agar culture medium while being prepared. The mixture was transferred to the plates. The bacteria were inoculated on plates and incubated as described. Results. The minimum inhibitory concentration (MIC) was 0.02 mg/mL for S. mutans and 0.3 mg/mL for S. sanguinis. The minimum bactericidal concentration (MBC) was 0.04 mg for S. mutans and 0.6 mg for S. sanguinis. Conclusion. Zingiber officinale extract has significant antibacterial activity against S. mutans and S. sanguinis cariogenic microorganisms. PMID:26347778

Evaluation of RGM Medium for Isolation of Nontuberculous Mycobacteria from Respiratory Samples from Patients with Cystic Fibrosis in the United States.

PubMed

Plongla, Rongpong; Preece, Clair L; Perry, John D; Gilligan, Peter H

2017-05-01

A novel selective agar (RGM medium) has been advocated for the isolation of rapidly growing mycobacteria from the sputa of cystic fibrosis (CF) patients. The aim of this study was to compare RGM medium to Burkholderia cepacia selective agar (BCSA) and a standard acid-fast bacillus (AFB) culture method for the isolation of nontuberculous mycobacteria (NTM) from patients with CF. The applicability of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) for the identification of NTM isolated on RGM medium was also assessed. Respiratory samples ( n = 869) were collected from 487 CF patients and inoculated directly onto RGM medium and BCSA. Cultures were incubated at 30°C and examined for up to 28 days. A subset of 212 samples (from 172 patients) was also cultured by using a mycobacterial growth indicator tube (MGIT) and on Lowenstein-Jensen medium following dual decontamination. By using a combination of all methods, 98 mycobacteria were isolated from 869 samples (11.3%). The sensitivity of RGM medium (96.9%) was significantly higher than that of BCSA (35.7%) for the isolation of mycobacteria ( P < 0.0001). The sensitivity of RGM medium was also superior to that of standard AFB culture for the isolation of mycobacteria (92.2% versus 47.1%; P < 0.0001). MALDI-TOF MS was effective for the identification of mycobacteria in RGM medium. RGM medium offers a simple and highly effective tool for the isolation of NTM from patients with CF. Extended incubation of RGM medium for 28 days facilitates the isolation of slow-growing species, including members of the Mycobacterium avium complex (MAVC). Copyright © 2017 American Society for Microbiology.

Research on Candida dubliniensis in a Brazilian yeast collection obtained from cardiac transplant, tuberculosis, and HIV-positive patients, and evaluation of phenotypic tests using agar screening methods.

PubMed

Ribeiro, Patrícia Monteiro; Querido, Silvia Maria Rodrigues; Back-Brito, Graziela Nueremberg; Mota, Adolfo José; Koga-Ito, Cristiane Yumi; Jorge, Antonio Olavo Cardoso

2011-09-01

The aim of this study was to research Candida dubliniensis among isolates present in a Brazilian yeast collection and to evaluate the main phenotypic methods for discrimination between C. albicans and C. dubliniensis from oral cavity. A total of 200 isolates, presumptively identified as C. albicans or C. dubliniensis obtained from heart transplant patients under immunosuppressive therapy, tuberculosis patients under antibiotic therapy, HIV-positive patients under antiretroviral therapy, and healthy subjects, were analyzed using the following phenotypic tests: formation and structural arrangement of chlamydospores on corn meal agar, casein agar, tobacco agar, and sunflower seed agar; growth at 45 °C; and germ tube formation. All strains were analyzed by polymerase chain reaction (PCR). In a preliminary screen for C. dubliniensis, 48 of the 200 isolates on corn meal agar, 30 of the 200 on casein agar, 16 of the 200 on tobacco agar, and 15 of the 200 on sunflower seed agar produced chlamydoconidia; 27 of the 200 isolates showed no or poor growth at 45 °C. All isolates were positive for germ tube formation. These isolates were considered suggestive of C. dubliniensis. All of them were subjected to PCR analysis using C. dubliniensis-specific primers. C. dubliniensis isolates were not found. C. dubliniensis isolates were not recovered in this study done with immunocompromised patients. Sunflower seed agar was the medium with the smallest number of isolates of C. albicans suggestive of C. dubliniensis. None of the phenotypic methods was 100% effective for discrimination between C. albicans and C. dubliniensis. Copyright © 2011 Elsevier Inc. All rights reserved.

Development and use of a selective medium for isolation of Leuconostoc spp. from vegetables and dairy products.

PubMed Central

Benkerroum, N; Misbah, M; Sandine, W E; Elaraki, A T

1993-01-01

A selective medium (LUSM medium) for the isolation of Leuconostoc spp. was developed. This medium contained 1.0% glucose, 1.0% Bacto Peptone (Difco), 0.5% yeast extract (BBL), 0.5% meat extract (Difco), 0.25% gelatin (Difco), 0.5% calcium lactate, 0.05% sorbic acid, 75 ppm of sodium azide (Sigma), 0.25% sodium acetate, 0.1% (vol/vol) Tween 80, 15% tomato juice, 30 micrograms of vancomycin (Sigma) per ml, 0.20 microgram of tetracycline (Serva) per ml, 0.5 mg of cysteine hydrochloride per ml, and 1.5% agar (Difco). LUSM medium was used successfully for isolation and enumeration of Leuconostoc spp. in dairy products and vegetables. Of 116 colony isolates obtained from fresh raw milk, curdled milk, or various vegetables, 115 were identified as members of the genus Leuconostoc. A total of 89 of these isolates were identified to species; 13.5% of the isolates were Leuconostoc cremoris, 7.9% were Leuconostoc mesenteroides subsp. mesenteroides, 11.2% were Leuconostoc mesenteroides subsp. dextranicum, 16.9% were Leuconostoc mesenteroides subsp. paramesenteroides, 10.1% were leuconostoc lactis, and 40.4% were Leuconostoc oenos. When we compared the counts obtained for two Leuconostoc strains, Leuconostoc dextranicum 181 and L. cremoris JLL8, on MRS agar and LUSM medium, we found no significant difference between the values obtained on the two media. PMID:8434926

Bis(2,3-dibromo-4,5-dihydroxybenzyl) Ether, a Marine Algae Derived Bromophenol, Inhibits the Growth of Botrytis cinerea and Interacts with DNA Molecules

PubMed Central

Liu, Ming; Wang, Genzhu; Xiao, Lin; Xu, Xuanli; Liu, Xiaohui; Xu, Pingxiang; Lin, Xiukun

2014-01-01

Bis(2,3-dibromo-4,5-dihydroxybenzyl) ether (BDDE) is a bromophenol isolated from marine algae. Previous reports have shown that BDDE possesses cytotoxic and antibacterial activity. In the present study, we demonstrate that BDDE displays broad-spectrum antifungal activities, especially on Botrytis cinerea. BDDE inhibits the growth of B. cinerea cultured on a solid medium of potato dextrose agar (PDA) as well as on the potato dextrose broth (PDB) medium. Moreover, BDDE decreases the incidence of fruit decay and severity of strawberries infected with B. cinerea. Further studies have revealed that BDDE decreases the germination rate and inhibits the mycelial growth of B. cinerea. The inhibition mechanisms are related to the disruption of the cell membrane integrity in B. cinerea spores and newly formed germ tubes. This study also suggests that BDDE possibly interacts with DNA via intercalation and minor groove binding. The studies provide evidence that BDDE has potential application in the control of gray mold after fruit harvest and the compound could serve as a candidate or lead template for rational drug design and for the development of antifungal agents. PMID:24979270

[Counting colonies of micro-organisms on solid media (author's transl)].

PubMed

Stonebrink, B

1978-11-01

The base of the system consists in a stainless steel cup with a shallow recess into which about 3 ml. of fluid agar medium may be poured, which produces a flat rectangular surface of about 8.65 sq. cm. after solidification. In the majority of cases, inoculation was performed by flooding the medium with a diluted suspension of micro-organisms. The cups were then put upright and carried over into a centrifuge tube which was closed with a rubber stopper. After the required incubation period, the colonies were counted, if necessary, using a specially developed colony counter connected to an electromechanical counting relay. Dilutions were prepared in screw capped bottles of various sizes. Pipettes were used only with connected "Volac" pipette holders. Cleaning and sterilisation could be done in an efficient and reliable way. In addition to the above method, the cups containing media may also be used for: impression counts, examination of fluids by immersion, inoculation by swabs or wire loops, studying air pollution. the system requires a rather large investment, but the daily cost and quantities of material used are small. The underlying study of the literature and the discussion of statistical methods used are available to the interested reader.

Selective effects of two systemic fungicides on soil fungi.

PubMed

Abdel-Fattah, H M; Abdel-Kader, M I; Hamida, S

1982-08-20

BAS 317 00F was not toxic to the total count of fungi after 2 days but was regularly significantly toxic at the three doses after 5, 20 and 40 days and toxic at the low and the high doses after 80 days. In the agar medium, it was toxic to the counts of total fungi. Aspergillus, A. terreus, Rhizopus oryzae and Mucor racemosus at the high dose. Only the mycelial growth of Trichoderma viride which was significantly inhibited by the three doses when this fungicide was added to the liquid medium. Polyram-Combi induced two effects on the total population of soil fungi. One inhibitory and this was demonstrated almost regularly after 2, 10 and 40 days and the other stimulatory after 80 days of treatment with the low and the high doses. In the agar medium, this fungicide was very toxic to total fungi and to almost all fungal genera and species at the three doses. Several fungi could survive the high dose. In liquid medium, the test fungi showed variable degree of sensitivity and the most sensitive was Gliocladium roseum which was completely eradicated by the three doses.

Addition of Carbon to the Culture Medium Improves the Detection Efficiency of Aflatoxin Synthetic Fungi

PubMed Central

Suzuki, Tadahiro; Iwahashi, Yumiko

2016-01-01

Aflatoxin (AF) is a harmful secondary metabolite that is synthesized by the Aspergillus species. Although AF detection techniques have been developed, techniques for detection of AF synthetic fungi are still required. Techniques such as plate culture methods are continually being modified for this purpose. However, plate culture methods require refinement because they suffer from several issues. In this study, activated charcoal powder (carbon) was added to a culture medium containing cyclodextrin (CD) to enhance the contrast of fluorescence and improve the detection efficiency for AF synthetic fungi. Two culture media, potato dextrose agar and yeast extract sucrose agar, were investigated using both plate and liquid cultures. The final concentrations of CD and carbon in the media were 3 mg/mL and 0.3 mg/mL, respectively. Addition of carbon improved the visibility of fluorescence by attenuating approximately 30% of light scattering. Several fungi that could not be detected with only CD in the medium were detected with carbon addition. The carbon also facilitated fungal growth in the potato dextrose liquid medium. The results suggest that addition of carbon to media can enhance the observation of AF-derived fluorescence. PMID:27854283

The role of gravity in the nutrition and formation of Bacillus colonies

NASA Astrophysics Data System (ADS)

Puzyr, A.; Tirranen, L.; Krylova, T.

The soil-like substrate is used to cultivate higher plants in man-made closed ecosystems. It allows increasing the closeness of the systems and decreasing the plant solid residues and human wastes. Unusual funnel-shaped bacterial colonies of Bacillus species have been observed during analysis of microflora of plant nutritional solution. The colonies have the following characteristics: a) the diameter of "funnel socket" (the biomass contacting with nutritional agar) is 10.0-15.0 mm; b) the thickness of "funnel socket" is 0.5-2.5 mm; c) the diameter of the middle part of the "funnel spout" (the biomass contacting with the gas phase) is 1,0-1,5 mm; d) the length of the "funnel spout" is 10.0-15.0 mm. In the socket and the middle part of the "funnel spout" there is a gas cavity which is most probably formed by bacterial gas metabolites. It has been shown that: i) the surface of these funnel-shaped colonies of Bacillus species is hydrophobic, as is the surface of other Bacillus species ( . brevis, B. cellulomonos, B. flavus, B.B formosus, B. subtilis); ii) the forms of colonies can be changed by varying the position of the growing biomass in relation to the gravitation forces. The experiment proved that the form of the "funnel sockets" and the length of the "funnel spouts" of the colonies are determined by hydrophobic air-contacting surface layer, which does not leak and stretches under the weight of accumulated water. A hypothesis has been suggested that the gravity force plays the role of a "pump" supplying and holding water within the colony. Thus, the water that comes under the gravity force contains dissolved nutrients and bacterial cells in the hydrophobic layer. These cells that are situated far away from the nutrient agar have no nutrient deficiency. The water accumulated by the colonies might be free water of agar media or it can be produced by metabolic disruption of medium fat. Hence, when growing a colony in agar media the water-soluble nutrient substances enter the growing colonies not only due to diffusion processes but also with the directional water flow under the gravity force.

Inactivation of Salmonella typhimurium and Escherichia coli O157:H7 in apple juice by a combination of nisin and cinnamon.

PubMed

Yuste, J; Fung, D Y C

2004-02-01

Pasteurized apple juice with nisin (0, 25, 50, 100, and 200 ppm, wt/vol) and cinnamon (0 and 0.3%, wt/vol) was inoculated with Salmonella Typhimurium and Escherichia coli O157:H7 at 10(4) CFU/ml and stored at 5 and 20 degrees C. Counts on tryptic soy agar (TSA), selective medium (xylose Lysine desoxycholate agar for Salmonella Typhimurium, and MacConkey sorbitol agar for E. coli O157:H7), and thin agar layer (TAL) were determined at 1 h and 1, 3, 7, and 14 days. The TAL method (selective medium overlaid with TSA) was used for recovery of sublethally injured cells. The pathogens were gradually inactivated by the acidic pH of apple juice. Nisin and cinnamon greatly contributed to the inactivation. The killing effect was more marked at 20 degrees C, with counts in all treated samples being undetectable by direct plating in 3 days for Salmonella Typhimurium and 7 days for E. coli O157:H7. Thus, several factors influenced the decrease in counts: low pH, addition of nisin and cinnamon, and storage temperature. The TAL method was as effective as TSA in recovering injured cells of the pathogens. The combination of nisin and cinnamon accelerates death of Salmonella Typhimurium and E. coli O157:H7 in apple juice and so enhances the safety of the product.

[Gum-like exudate from Laguncularia racemosa (white mangrove) as culture media for fungi].

PubMed

Mesa, L M; León-Pinto, G

1993-01-01

Morphological studies of eight species of fungus: Aspergillus flavus Microsporum canis, Epidermophyton floccosum, Curvularia lunata, Cladosporium carrionii, Natrassia mangífera (Edo. Scytalidium), Sporotrix schenckii y Rhizophus oligosporus, which belong to families Mucedinaceae, Dematiaceae and Mucoraceae have been carried out in support medium based in gum exudate from Laguncularia racemosa (mangle blanco). This native polimer contains galactose, arabinose, rhamnose, uronic acid and proteins. Nitrogen calcium and magnesium are microconstituents of the gum. An economical substrate which contained gum exudate (4%) and agar (1.5%) was used in these studies. The results obtained showed that gum exudate-agar medium (EGA) permits an adequate identification of the studied species, therefore, it is a possible substitute for Sabouraud. It is important to know that the gum exudate is a natural product, economical and easy to obtain.

In vitro survival of fresh and frozen/thawed bovine demi-embryos.

PubMed

Lucas-Hahn, A; Niemann, H

1991-10-01

Three experiments were conducted to investigate the effects of type of culture medium in freshly bisected bovine embryos and the effects of agar embedding and of 1.2 propanediol (PROH) as the cryoprotectant in frozen/thawed bisected bovine embryos on development in vitro. A total of 265 bovine embryos were used as controls or were microsurgically bisected and were cultured in vitro for 48 hours and development was determined 24 and 48 hours after the onset of culture. Whitten's medium supported more (P<0.05) intact and demi-embryos to grow to expanded blastocysts (92.9 and 73.1%, respectively) compared with Ham's F10 (43.8 and 26.3%, respectively) and PBS (53.8 and 12.5%, respectively). Embedding in agar and culture in Whitten's medium resulted in a higher (P<0.05) percentage of in vitro development of frozen/thawed demi-embryos after 24 hours than the freezing of nonembedded demi-embryos (44.1 versus 19.6%, respectively). This difference disappeared, however, after a 48 hours culture period (17.6 versus 11.8%, respectively). Following freezing in PROH, survival rates of 40 and 28%, respectively after 24 hours of culture were obtained for intact and demi-embryos. The respective percentages after 48 hours were 8.6 and 16%. Since neither embedding in agar nor the use of PROH as the cryoprotectant resulted in high survival rates of frozen/thawed demi-embryos in vitro, new freezing procedures are needed to overcome the sensitivity of demi-embryos to freezing and thawing.

Improved agar diffusion method for detecting residual antimicrobial agents.

PubMed

Tsai, C E; Kondo, F

2001-03-01

The improved agar diffusion method for determination of residual antimicrobial agents was investigated, and the sensitivities of various combinations of test organisms and assay media were determined using 7 organisms, 5 media, and 31 antimicrobial agents. Bacillus stearothermophilus and synthetic assay medium (SAM) showed the greatest sensitivity for screening penicillins (penicillin G and ampicillin). The combination of Bacillus subtilis and minimum medium (MM) was the most sensitive for tetracyclines (oxytetracycline and chlortetracycline), B. stearothermophilus and SAM or Micrococcus luteus and Mueller-Hinton agar (MHA) for detecting tylosin and erythromycin, B. subtilis and MHA for aminoglycosides (streptomycin, kanamycin, gentamicin, and dihydrostreptomycin), B. stearothermophilus and SAM for polyethers (salinomycin and lasalocid), and B. subtilis and MM or Clostridium perfringens and GAM for polypeptides (thiopeptin, enramycin, virginiamycin, and bacitracin). However, gram-negative bacterium Escherichia coli ATCC 27166 and MM were better for screening for colistin and polymixin-B. For detecting the synthetic drugs tested, the best combination was B. subtilis and MM for sulfonamides, E. coli 27166 and MM for quinolones (oxolinic acid and nalidixic acid), B. subtilis and MM for furans (furazolidone), and the bioluminescent bacterium Photobacterium phosphoreum and luminescence assay medium for chloramphenicol and oxolinic acid. The results showed that the use of four assay plates, B. stearothermophilus and SAM, B. subtilis and MM, M. luteus and MHA, and E. coli 27166 and MM, was superior to the currently available techniques for screening for residual antimicrobial agents in edible animal tissues.

Long-term biological hydrogen production by agar immobilized Rhodobacter capsulatus in a sequential batch photobioreactor.

PubMed

Elkahlout, Kamal; Alipour, Siamak; Eroglu, Inci; Gunduz, Ufuk; Yucel, Meral

2017-04-01

In this study, agar immobilization technique was employed for biological hydrogen production using Rhodobacter capsulatus DSM 1710 (wild type) and YO3 (hup-mutant) strains in sequential batch process. Different agar and glutamate concentrations were tested with defined nutrient medium. Agar concentration 4% (w/v) and 4 mM glutamate were selected for bacterial immobilization in terms of rate and longevity of hydrogen production. Acetate concentration was increased from 40 to 60-100 and 60 mM gave best results with both bacterial strains immobilized in 4% (w/v) agar. Cell concentration was increased from 2.5 to 5 mg dcw mL -1 agar and it was found that increasing cell concentration of wild-type strain caused decrease in yield and productivity while these parameters improved by increasing cell concentration of mutant strain. Also, the hydrogen production time has extended from 17 days up to 60 days according to the process conditions and parameters. Hydrogen production by immobilized photosynthetic bacteria is a convenient technology for hydrogen production as it enables to produce hydrogen with high organic acid concentrations comparing to suspended cultures. Besides, immobilization increases the stability of the system and allowed sequential batch operation for long-term application.

Effect of sporulation medium on wet-heat resistance and structure of Alicyclobacillus acidoterrestris DSM 3922-type strain spores and modeling of the inactivation kinetics in apple juice.

PubMed

Molva, Celenk; Baysal, Ayse Handan

2014-10-17

Alicyclobacillus acidoterrestris is a spoilage bacterium in fruit juices leading to high economic losses. The present study evaluated the effect of sporulation medium on the thermal inactivation kinetics of A. acidoterrestris DSM 3922 spores in apple juice (pH3.82±0.01; 11.3±0.1 °Brix). Bacillus acidocaldarius agar (BAA), Bacillus acidoterrestris agar (BATA), malt extract agar (MEA), potato dextrose agar (PDA) and B. acidoterrestris broth (BATB) were used for sporulation. Inactivation kinetic parameters at 85, 87.5 and 90°C were obtained using the log-linear model. The decimal reduction times at 85°C (D85°C) were 41.7, 57.6, 76.8, 76.8 and 67.2min; D87.5°C-values were 22.4, 26.7, 32.9, 31.5, and 32.9min; and D90°C-values were 11.6, 9.9, 14.7, 11.9 and 14.1min for spores produced on PDA, MEA, BATA, BAA and BATB, respectively. The estimated z-values were 9.05, 6.60, 6.96, 6.15, and 7.46, respectively. The present study suggests that the sporulation medium affects the wet-heat resistance of A. acidoterrestris DSM 3922 spores. Also, the dipicolinic acid content (DPA) was found highest in heat resistant spores formed on mineral containing media. After wet-heat treatment, loss of internal volume due to the release of DPA from spore core was observed by scanning electron microscopy. Since, there is no standardized media for the sporulation of A. acidoterrestris, the results obtained from this study might be useful to determine and compare the thermal resistance characteristics of A. acidoterrestris spores in fruit juices. Copyright © 2014 Elsevier B.V. All rights reserved.

[Efficacy of topical 0.3% ciprofloxacin application in reducing the conjunctival biota of patients undergoing cataract extraction].

PubMed

Carron, A; Samudio, M; Laspina, F; Fariña, N; Sanabria, R R; Cibils, D; Ramirez, L; Carron, J; Mino de Kaspar, H

2013-09-01

To determine the efficacy of topical 0.3% ciprofloxacin in reducing conjunctival biota in patients undergoing cataract surgery. Experimental, prospective, randomized, controlled and single-blind study. Forty-six eyes of 46 patients were randomized into 2 groups, the study group (n=23) received topical 0.3% ciprofloxacin one day before surgery for six times, and on the day of the surgery one drop every 15minutes starting one hour before surgery until 3 doses were completed. The control group (n=23) did not receive any antibiotics. For both groups for the surgical field 10% povidone-iodine was applied. Samples from the conjunctiva were taken at four different times and then cultured on solid media (chocolate agar, blood agar) and enrichment broth (thioglycolate). The aqueous humor samples were also cultured in thioglycolate. The presence of bacteria was identified quantitatively and qualitatively, and the frequency of contamination was measured by considering the presence of bacteria in liquid and solid culture media. The number of colony forming units (CFU) was counted in the solid culture medium. Positive cultures were obtained in 82.6% and 78.2% of the patients in the study and control groups, respectively, before the administration of 0.3% ciprofloxacin. The administration of 0.3% ciprofloxacin significantly reduced the CFU compared to the control group (P<.05). Immediately after the use of povidone-iodine, the proportion of patients with a positive culture decreased to 21.7% in the study group, and 8.7% in the control group. At the end of the surgery, this percentage was 26% and 30.4%, respectively. The most common isolated pathogen was negative-coagulase Staphylococcus (66.7%). The administration of 0.3% ciprofloxacin reduces conjunctival bacterial load in the preoperative period. However, it was unable to eradicate the bacteria completely. The administration of povidone-iodine reduced conjunctival biota in 50%-70% of patients undergoing cataract surgery. Copyright © 2012 Sociedad Española de Oftalmología. Published by Elsevier Espana. All rights reserved.

Thermal inactivation and sublethal injury kinetics of Salmonella enterica and Listeria monocytogenes in broth versus agar surface.

PubMed

Wang, Xiang; Devlieghere, Frank; Geeraerd, Annemie; Uyttendaele, Mieke

2017-02-21

The objective of the present study was to compare the thermal inactivation and sublethal injury kinetics of Salmonella enterica and Listeria monocytogenes in broth (suspended cells) and on solid surface (agar-seeded cells). A 3-strain cocktail of S. enterica or L. monocytogenes inoculated in broth or on agar was subjected to heating in a water bath at various set temperatures (55.0, 57.5 and 60.0°C for S. enterica and 60.0, 62.5 and 65°C for L. monocytogenes). The occurrence of sublethally injured cells was determined by comparing enumerations on nonselective (TSAYE) and selective (XLD or ALOA) media. Results showed that the inactivation curves obtained from selective media were log-linear, and significant shoulders (p<0.05) were observed on some of the inactivation curves from TSAYE media. The D-values derived from the total population were higher than those from the uninjured cells. Generally, cells on agar surface exhibited higher heat resistance than those in broth. For S. enterica, cell injury increased with the exposure time, no difference was observed when treated at temperatures from 55.0 to 60.0°C, while for L. monocytogenes, cell injury increased significantly with heating time and treatment temperature (from 60.0 to 65°C). Moreover, the degree of sublethal injury affected by thermal treatment in broth or on agar surface depended upon the target microorganism. Higher proportions of injured S. enterica cells were observed for treatment in broth than on agar surface, while the opposite was found for L. monocytogenes. The provided information may be used to assess the efficacy of thermal treatment processes on surfaces for inactivation of S. enterica and L. monocytogenes, and it provides insight into the sublethally injured survival state of S. enterica and L. monocytogenes treated in liquid or on solid food. Copyright © 2016 Elsevier B.V. All rights reserved.

Evaluation of Bruker Biotyper and Vitek MS for the identification of Candida tropicalis on different solid culture media.

PubMed

Wang, He; Li, Ying; Fan, Xin; Chiueh, Tzong-Shi; Xu, Ying-Chun; Hsueh, Po-Ren

2017-11-11

The aim of this study was to investigate the performance of the Bruker Biotyper matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and Vitek MS systems for identification of genetically-confirmed blood isolates of Candida tropicalis that had been grown on several types of culture media commonly used for primary fungal isolation. Isolates included 105 from the National China Hospital Invasive Fungal Surveillance Net program (CHIF-NET) and 120 from National Taiwan University Hospital (NTUH). Culture media tested for CHIF-NET isolates included trypticase soy agar supplemented with 5% sheep blood (BAP), Sabouraud dextrose agar (SDA-C), CHROMagar, China blue agar (CBA), chocolate agar supplemented with vancomycin (CAP-VA), and MacConkey agar (MAC). Culture media used for NTUH isolates included BAP, SDA, CHROMagar, eosin methylene blue (EMB), inhibitory mold agar (IMA), Mycosel agar, and cornmeal agar (CMA). The Bruker Biotyper correctly identified all CHIF-NET isolates to the species level on all six agar media tested and correctly identified the majority of NTUH isolates with the exception of isolates grown on SDA (85.8%) and CMA (52.5%). The Vitek MS system correctly identified all CHIF-NET isolates to the species level with the exception of isolates grown on CHROMagar (84.8%), and correctly identified the majority of NTUH isolates with the exception of isolates grown on SDA (51.7%), Mycosel agar (57.5%), and CMA (9.2%) for NTUH isolates. Clinical microbiologists should be aware that different culture media can affect the performance of the Bruker Biotyper MALDI-TOF MS and Vitek MS systems in identifying C. tropicalis. Copyright © 2017. Published by Elsevier B.V.

Gravity, chromosomes, and organized development in aseptically cultured plant cells

NASA Technical Reports Server (NTRS)

Krikorian, Abraham D.

1993-01-01

The objectives of the PCR experiment are: to test the hypothesis that microgravity will in fact affect the pattern and developmental progression of embryogenically competent plant cells from one well-defined, critical stage to another; to determine the effects of microgravity in growth and differentiation of embryogenic carrot cells grown in cell culture; to determine whether microgravity or the space environment fosters an instability of the differentiated state; and to determine whether mitosis and chromosome behavior are adversely affected by microgravity. The methods employed will consist of the following: special embryogenically competent carrot cell cultures will be grown in cell culture chambers provided by NASDA; four cell culture chambers will be used to grow cells in liquid medium; two dishes (plant cell culture dishes) will be used to grow cells on a semi-solid agar support; progression to later embryonic stages will be induced in space via crew intervention and by media manipulation in the case of liquid grown cell cultures; progression to later stages in case of semi-solid cultures will not need crew intervention; embryo stages will be fixed at a specific interval (day 6) in flight only in the case of liquid-grown cultures; and some living cells and somatic embryos will be returned for continued post-flight development and 'grown-out.' These will derive from the semi-solid grown cultures.

Trace Amounts of Furan-2-Carboxylic Acids Determine the Quality of Solid Agar Plates for Bacterial Culture

PubMed Central

Hara, Shintaro; Isoda, Reika; Tahvanainen, Teemu; Hashidoko, Yasuyuki

2012-01-01

Background Many investigators have recognised that a significant proportion of environmental bacteria exist in a viable but non-culturable state on agar plates, and some researchers have also noticed that some of such bacteria clearly recover their growth on matrices other than agar. However, the reason why agar is unsuitable for the growth of some bacteria has not been addressed. Methodology/Principal Findings According to the guide of a bioassay for swarming inhibition, we identified 5-hydroxymethylfuran-2-carboxylic acid (5-HMFA) and furan-2-carboxylic acid (FA) as factors that inhibit bacterial swarming and likely inhibit extracellular polysaccharide production on agar. The furan-2-carboxylic acids 5-HMFA and FA effectively inhibited the swarming and swimming of several environmental bacteria at concentrations of 1.8 and 2.3 µg L−1 (13 and 21 nmol L−1), respectively, which are equivalent to the concentrations of these compounds in 0.3% agar. On Luria-Bertani (LB) plates containing 1.0% agar that had been previously washed with MeOH, a mixture of 5-HMFA and FA in amounts equivalent to their original concentrations in the unwashed agar repressed the swarming of Escherichia coli K12 strain W3110, a representative swarming bacterium. Conclusions/Significance Agar that contains trace amounts of 5-HMFA and FA inhibits the proliferation of some slow-growing or difficult-to-culture bacteria on the plates, but it is useful for single colony isolation due to the ease of identification of swarmable bacteria as the non-swarmed colonies. PMID:22848437

Chemical Growth Regulators for Guayule Plants

NASA Technical Reports Server (NTRS)

Dastoor, M. N.; Schubert, W. W.; Petersen, G. R.

1982-01-01

Test Tubes containing Guayule - tissue cultures were used in experiments to test effects of chemical-growth regulators. The shoots grew in response to addition of 2-(3,4-dichlorophenoxy)-triethylamine (triethylamine (TEA) derivative) to agar medium. Preliminary results indicate that a class of compounds that promotes growth in soil may also promote growth in a culture medium. Further experiments are needed to define the effect of the TEA derivative.

Esculin hydrolysis by Vibrio vulnificus.

PubMed

Tison, D L

1986-01-01

A clinical isolate of Vibrio vulnificus was found to hydrolyze esculin when tested on bile-esculin-azide agar during the initial characterization of the strain. Reports in the literature of esculin hydrolysis by V. vulnificus are conflicting. We tested herein 52 strains of V. vulnificus from clinical and environmental sources for the ability to hydrolyze esculin. Seventy-eight percent of the strains hydrolyzed esculin on bile-esculin-azide agar, whereas all strains of V. vulnificus tested were positive for esculin hydrolysis in a noninhibitory medium, whereas some strains failed to hydrolyze esculin on media containing inhibitory compounds.

Practical direct plaque assay for coliphages in 100-ml samples of drinking water.

PubMed Central

Grabow, W O; Coubrough, P

1986-01-01

A practical single-agar-layer plaque assay for the direct detection of coliphages in 100-ml samples of water was designed and evaluated. With this assay a 100-ml sample of water, an agar medium containing divalent cations, and the host Escherichia coli C (ATCC 13706) were mixed in a single container, and the mixture was plated on 10 14-cm-diameter petri dishes. It was more sensitive, reliable, and accurate than various other methods and proved rapid, simple, and economic. PMID:3532952

An improved procedure for shipboard enumeration of faecal indicator bacteria in marine sediments from sewage sludge disposal areas.

PubMed

West, P A

1988-04-01

An improved membrane filtration procedure for use on board ship to enumerate Escherichia coli and Group D faecal streptococci in marine sediments is described. Ultrasonication extraction combined with resuscitation of sublethally-injured cells yielded significantly higher counts of E. coli than sediments shaken by hand. Counts of E. coli were also higher on mFC agar (without rosalic acid) after a period of resuscitation on tryptone-soy agar supplemented with 0.1% yeast extract than on a 4% Teepol-lactose medium. Ultrasonication of sediments made no significant difference to counts of Group D faecal streptococci on KF-streptococcus agar. These improved isolation procedures allowed better discrimination of the area affected by sewage sludge at a disposal site off the northeast coast of England.

Comparison of CHROMagar Salmonella Medium and Hektoen Enteric Agar for Isolation of Salmonellae from Stool Samples

PubMed Central

Gaillot, Olivier; Di Camillo, Patrick; Berche, Patrick; Courcol, René; Savage, Colette

1999-01-01

CHROMagar Salmonella (CAS), a new chromogenic medium, was retrospectively compared to Hektoen enteric agar (HEA) with 501 Salmonella stock isolates and was then prospectively compared to HEA for the detection and presumptive identification of Salmonella spp. with 508 stool samples before and after enrichment. All stock cultures (100%), including cultures of H2S-negative isolates, yielded typical mauve colonies on CAS, while 497 (99%) isolates produced typical lactose-negative, black-centered colonies on HEA. Following overnight incubation at 37°C, a total of 20 Salmonella strains were isolated from the 508 clinical samples. Sensitivities for primary plating and after enrichment were 95% (19 isolates) and 100% (20 isolates), respectively, for CAS and 80% (16 isolates) and 100% (20 isolates), respectively, for HEA. The specificity of CAS (88.9%) was significantly higher than that of HEA (78.5%; P < 0.0001). On the basis of its good sensitivity and specificity, CAS medium can be recommended for use for primary plating when human stool samples are screened for Salmonella spp. PMID:9986847

Ensifer meliloti overexpressing Escherichia coli phytase gene ( appA) improves phosphorus (P) acquisition in maize plants

NASA Astrophysics Data System (ADS)

Sharma, Vikas; Kumar, Ajit; Archana, G.; Kumar, G. Naresh

2016-10-01

The Escherichia coli phytase gene appA encoding enzyme AppA was cloned in a broad host range plasmid pBBR1MCS2 ( lac promoter), termed pVA1, and transformed into the Ensifer meliloti 1020. Transformation of pVA1 in Ensifer meliloti { E. m (pVA1)} increased its phosphatase and phytase activity by ˜9- and ˜50-fold, respectively, compared to the transformants containing empty plasmid as control { E. m (pBBR1MCS2)}. The western blot experiments using rabbit anti-AppA antibody showed that AppA is translocated into the periplasm of the host after its expression. Ensifer meliloti harboring AppA protein { E. m (pVA1)} and { E. m (pBBR1MCS2)} could acidify the unbuffered phytate minimal media (pH 8.0) containing Ca-phytate or Na-phytate as sole organic P (Po) source to below pH 5.0 and released P. However, both { E. m (pVA1)} and { E. m (pBBR1MCS2)} neither dropped pH of the medium nor released P when the medium was buffered at pH 8.0 using Tris-Cl, indicating that acidification of medium was important for the enzymatic hydrolysis of phytate. Further experiments proved that maize plants inoculated with { E. m. (pVA1)} showed increase in growth under sterile semi solid agar (SSA) medium containing Na-phytate as sole P source. The present study could be helpful in generating better transgenic bioinoculants harboring phosphate mineralization properties that ultimately promote plant growth.

Ensifer meliloti overexpressing Escherichia coli phytase gene (appA) improves phosphorus (P) acquisition in maize plants.

PubMed

Sharma, Vikas; Kumar, Ajit; Archana, G; Kumar, G Naresh

2016-10-01

The Escherichia coli phytase gene appA encoding enzyme AppA was cloned in a broad host range plasmid pBBR1MCS2 (lac promoter), termed pVA1, and transformed into the Ensifer meliloti 1020. Transformation of pVA1 in Ensifer meliloti {E. m (pVA1)} increased its phosphatase and phytase activity by ∼9- and ∼50-fold, respectively, compared to the transformants containing empty plasmid as control {E. m (pBBR1MCS2)}. The western blot experiments using rabbit anti-AppA antibody showed that AppA is translocated into the periplasm of the host after its expression. Ensifer meliloti harboring AppA protein {E. m (pVA1)} and {E. m (pBBR1MCS2)} could acidify the unbuffered phytate minimal media (pH 8.0) containing Ca-phytate or Na-phytate as sole organic P (Po) source to below pH 5.0 and released P. However, both {E. m (pVA1)} and {E. m (pBBR1MCS2)} neither dropped pH of the medium nor released P when the medium was buffered at pH 8.0 using Tris-Cl, indicating that acidification of medium was important for the enzymatic hydrolysis of phytate. Further experiments proved that maize plants inoculated with {E. m. (pVA1)} showed increase in growth under sterile semi solid agar (SSA) medium containing Na-phytate as sole P source. The present study could be helpful in generating better transgenic bioinoculants harboring phosphate mineralization properties that ultimately promote plant growth.

Isolation and Characterization of Isogenic Pairs of Domed Hemolytic and Flat Nonhemolytic Colony Types of Bordetella pertussis

PubMed Central

Peppler, Mark S.

1982-01-01

Four different serotype strains of Bordetella pertussis, 3779BL2S4, Tohama I, 353/Z, and 2753, were plated on Bordet-Gengou agar, where they grew as domed, hemolytic (D+H+) wild-type colonies. Cloned D+H+ colony types of all four strains were passed onto modified Stainer-Scholte medium solidified with 1% Noble Agar. Colonies were selected from Stainer-Scholte agar, and these subsequently grew as flat, nonhemolytic (D−H−) colonies when transferred back onto Bordet-Gengou agar. The frequency of D−H− organisms within a population of cloned D+H+ was determined to be between 5 × 10−5 and 5 × 10−6. The D−H− colony types maintained their flat, nonhemolytic characteristics for over 80 single-colony passages on Bordet-Gengou agar. The isogenic pairs of D+H+ and D−H− colony types from the four strains were compared for hemagglutination titer, lymphocytosis-promoting activity, adenylate cyclase activity, and presence of agglutinogens by agglutination. In all cases the D−H− colony types showed reduced activities or amounts of antigen compared with their D+H+ parents. Freely diffusible antigens were markedly different between the two phenotypes as noted by double diffusion of antisera added to plates on which colonies of the variants were growing. Antigens solubilized from the two colony types by Triton X-100 were also markedly different as judged by radial immunodiffusion with antifimbrial hemagglutinin, antilymphocytosis-promoting factor, and anti-353/Z adsorbed with autoclaved 353/Z. In addition, autoradiographs of 125I-surface-labeled whole cells separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed unique banding patterns for each colony type. Since all organisms, regardless of colony type, were grown on Bordet-Gengou agar, the differences reported could not be due to medium composition. Differences between phenotypes were also independent of passage number on Bordet-Gengou agar. By analogy to previous studies, the D−H− organisms appear to fulfill the criteria for phase III or phase IV in the system of Leslie and Gardner (P. H. Leslie and A. D. Gardner, J. Hyg. 31:423-434, 1931) or phase III in the system of Kasuga et al. (T. Kasuga, Y. Nakase, K. Ukishima, and K. Takatsu, Kitasato Arch. Exp. Med. 26:121-134, 1954). Images PMID:6279517

Alginate-encapsulation of shoot tips of jojoba [Simmondsia chinensis (Link) Schneider] for germplasm exchange and distribution.

PubMed

Kumar, Sunil; Rai, Manoj K; Singh, Narender; Mangal, Manisha

2010-12-01

Shoot tips excised from in vitro proliferated shoots derived from nodal explants of jojoba [Simmondsia chinensis (Link) Schneider] were encapsulated in calcium alginate beads for germplasm exchange and distribution. A gelling matrix of 3 % sodium alginate and 100 mM calcium chloride was found most suitable for formation of ideal calcium alginate beads. Best response for shoot sprouting from encapsulated shoot tips was recorded on 0.8 % agar-solidified full-strength MS medium. Rooting was induced upon transfer of sprouted shoots to 0.8 % agar-solidified MS medium containing 1 mg l(-1) IBA. About 70 % of encapsulated shoot tips were rooted and converted into plantlets. Plants regenerated from encapsulated shoot tips were acclimatized successfully. The present encapsulation approach could also be applied as an alternative method of propagation of desirable elite genotype of jojoba.

Chlamydospore production and germ-tube formation by auxotrophs of Candida albicans.

PubMed

Balish, E

1973-04-01

A prototrophic strain and 21 auxotrophic strains of Candida albicans were assessed for their capacity to produce chlamydospores and germ tubes. All of the mutants were able to produce germ-tubes in human serum but only two mutants produced them in defined medium with L-alpha-amino-n-butyric acid as the sole source of nitrogen. Most auxotrophs were not able to produce chlamydospores on corn meal agar with 1% Tween 80, but they could be induced to do so if the medium was supplemented with their growth requirement(s). Although L-cysteine was able to support the growth of two methionine mutants, it did not support chlamydospore formation when added to corn meal agar with 1% Tween 80. Mutants of C. albicans that do not form chlamydospores could be incorrectly identified in laboratories that rely on chlamydospore formation for identification.

High efficiency induction of callus and regeneration of sporophytes of Laminaria japonica (Phaeophyta)

NASA Astrophysics Data System (ADS)

Wang, Xi-Hua; Qin, Song; Li, Xin-Ping; Jiang, Peng; Zeng, Cheng-Kui; Qin, Mei

1998-03-01

Four media (PESI solid, MS liquid, MS solid and ASP-C-I solid medium) were used to induce callus from excised tissues of the kelp Laminaria japonica. Only PESI solid medium and MS solid medium produced calli. Modified MS solid medium supplemented with mannitol (3%,W/V), yeast extract (0.1%, W/V), VB2 (0.5 mg/ml), VB12 (0.5 mg/ml), kinetin (0.108 μg/ml) and NAA (1.860μg/ml) showed much better effect on callus induction than non-modified MS solid medium. After 24 days of induction 75.5% of tissues in PESI solid medium showed callus formation. For modified MS solid medium, after three months of induction 67.3% of tissues dedifferentiated into calli. No callus could be found after five months of induction in either MS liquid or ASP-C-I solid medium. When calli were squashed and cultured in N-P enriched autoclaved seawater, MS liquid medium and ASP12-NTA liquid medium (both modified with kelp extract), differentiation of cells and regeneration of sporophytes were only observed in ASP12-NTA medium supplemented with kelp extract. Gametophyte-like filaments formed first, then eggs were released. It was suggested that sporophyte formation could be a process of parthenogenesis. Sterilization techniques in tissue culture of L. japonica were also tested in this study.

Collaborative investigation of broth microdilution and semisolid agar dilution for in vitro susceptibility testing of Candida albicans.

PubMed Central

Shawar, R; Paetznick, V; Witte, Z; Ensign, L G; Anaissie, E; LaRocco, M

1992-01-01

A study was performed in two laboratories to evaluate the effect of growth medium and test methodology on inter- and intralaboratory variations in the MICs of amphotericin B (AMB), flucytosine (5FC), fluconazole (FLU), itraconazole (ITRA), and the triazole Sch 39304 (SCH) against 14 isolates of Candida albicans. Testing was performed by broth microdilution and semisolid agar dilution with the following media, buffered to pH 7.0 with morpholinepropanesulfonic acid (MOPS): buffered yeast nitrogen base (BYNB), Eagle's minimal essential medium (EMEM), RPMI 1640 medium (RPMI), and synthetic amino acid medium for fungi (SAAMF). Inocula were standardized spectrophotometrically, and endpoints were defined by the complete absence of growth for AMB and by no more than 25% of the growth in the drug-free control for all other agents. Comparative analyses of median MICs, as determined by each test method, were made for all drug-medium combinations. Both methods yielded similar (+/- 1 twofold dilution) median MICs for AMB in EMEM and RPMI, 5FC in all media, and FLU in EMEM, RPMI, and SAAMF. In contrast, substantial between-method variations in median MICs were seen for AMB in BYNB and SAAMF, FLU In BYNB, and ITRA and SCH in all media. Interlaboratory concordance of median MICs was good for AMB, 5FC, and FLU but poor for ITRA and SCH in all media. Endpoint determinations were analyzed by use of kappa statistical analyses for evaluating the strength of observer agreement. Moderate to almost perfect interlaboratory agreement occurred with AMB and 5FC in all media and with FLU in EMEM, RPMI, and SAAMF, irrespective of the test method. Slight to almost perfect interlaboratory agreement occurred with ITRA and SCH in EMEM, RPMI, and SAAMF when tested by semisolid agar dilution but not broth microdilution. Kappa values assessing intralaboratory agreement between methods were high for 5FC in all media, for AMB in BYNB, ENEM, and RPMI, and for FLU in EMEM, RPMI, and SAAMF. One laboratory, but not the other, reported substantial to almost perfect agreement between methods for ITRA, and SCH in EMEM, RPMI, and SAAMF. Both laboratories reported poor agreement between methods for the azoles in BYNB. Discrepancies noted in azole-BYNB combinations were largely due to the greater inhibitory effect of these agents in BYNB than in other media. These results indicate that the semisolid agar dilution and broth microdilution methods with EMEM or RPMI yield equivalent and reproducible MICs for AMB, 5FC, and FLU but not ITRA and SCH. PMID:1500502

Collaborative investigation of broth microdilution and semisolid agar dilution for in vitro susceptibility testing of Candida albicans.

PubMed

Shawar, R; Paetznick, V; Witte, Z; Ensign, L G; Anaissie, E; LaRocco, M

1992-08-01

A study was performed in two laboratories to evaluate the effect of growth medium and test methodology on inter- and intralaboratory variations in the MICs of amphotericin B (AMB), flucytosine (5FC), fluconazole (FLU), itraconazole (ITRA), and the triazole Sch 39304 (SCH) against 14 isolates of Candida albicans. Testing was performed by broth microdilution and semisolid agar dilution with the following media, buffered to pH 7.0 with morpholinepropanesulfonic acid (MOPS): buffered yeast nitrogen base (BYNB), Eagle's minimal essential medium (EMEM), RPMI 1640 medium (RPMI), and synthetic amino acid medium for fungi (SAAMF). Inocula were standardized spectrophotometrically, and endpoints were defined by the complete absence of growth for AMB and by no more than 25% of the growth in the drug-free control for all other agents. Comparative analyses of median MICs, as determined by each test method, were made for all drug-medium combinations. Both methods yielded similar (+/- 1 twofold dilution) median MICs for AMB in EMEM and RPMI, 5FC in all media, and FLU in EMEM, RPMI, and SAAMF. In contrast, substantial between-method variations in median MICs were seen for AMB in BYNB and SAAMF, FLU In BYNB, and ITRA and SCH in all media. Interlaboratory concordance of median MICs was good for AMB, 5FC, and FLU but poor for ITRA and SCH in all media. Endpoint determinations were analyzed by use of kappa statistical analyses for evaluating the strength of observer agreement. Moderate to almost perfect interlaboratory agreement occurred with AMB and 5FC in all media and with FLU in EMEM, RPMI, and SAAMF, irrespective of the test method. Slight to almost perfect interlaboratory agreement occurred with ITRA and SCH in EMEM, RPMI, and SAAMF when tested by semisolid agar dilution but not broth microdilution. Kappa values assessing intralaboratory agreement between methods were high for 5FC in all media, for AMB in BYNB, ENEM, and RPMI, and for FLU in EMEM, RPMI, and SAAMF. One laboratory, but not the other, reported substantial to almost perfect agreement between methods for ITRA, and SCH in EMEM, RPMI, and SAAMF. Both laboratories reported poor agreement between methods for the azoles in BYNB. Discrepancies noted in azole-BYNB combinations were largely due to the greater inhibitory effect of these agents in BYNB than in other media. These results indicate that the semisolid agar dilution and broth microdilution methods with EMEM or RPMI yield equivalent and reproducible MICs for AMB, 5FC, and FLU but not ITRA and SCH.

Evaluation of Chromogenic Medium for Selective Isolation of Yersinia.

PubMed

Thuan, Nguyen Khanh; Naher, Kamrun; Kubo, Ryoichi; Taniguchi, Takahide; Hayashidani, Hideki

2016-01-01

Cefsulodin-irgasan-novobiocin agar (CIN) has been used as a selective agar to detect Yersinia in food or human patients; however, its components can inhibit the growth of some strains of Yersinia enterocolitica serovar O3 and Y. pseudotuberculosis. Recently, a new Yersinia selective agar, CHROMagar Yersinia enterocolitica (CAYe), was developed and evaluated as a novel selective agar for pathogenic Y. enterocolitica. In this research, a total of 251Yersinia strains (176 pathogenic Y. enterocolitica, 59 Y. pseudotuberculosis, and 16 non-pathogenic Yersinia) were cultured on both CIN and CAYe for comparison. Except for 10 of 104 pathogenic Y. enterocolitica O3 strains and 59 Y. pseudotuberculosis strains, 198 Yersinia isolates grew on both media after 48 hr of incubation at 32℃. Of the 10 pathogenic Y. enterocolitica O3 which could not grow on CIN or CAYe, 9 strains could not grow on CIN with supplements and 1 strain could not grow CAYe with supplements. Of 9 strains which did not grow on CIN with supplements, 3 strains could not grow on CIN without supplements. However, 1 strain which did not grow on CAYe with supplements could grow on CAYe without supplements. All of the Y. pseudotuberculosis strains could grow on CIN with/without supplements and on CAYe without supplements. The results indicate that the inhibition of the growth of Y. enterocolitica O3 on CIN is related to the components of CIN; however, the inhibition on CAYe appears to be related to the supplements in CAYe. Therefore, CAYe may be a more useful selective medium than CIN for pathogenic Y. enterocolitica .

Germ tube and chlamydospore formation by Candida albicans on a new medium.

PubMed

Beheshti, F; Smith, A G; Krause, G W

1975-10-01

A new medium composed of "cream of rice" infusion, oxgall, Tween 80, and agar is described for the sequential development of germ tubes and chlamydospores by Candida albicans. The procedure used (Dalmau's technique) is an improvement over the fluid substrate procedures previously advocated for germ tube formation. That the same preparation is then used for chlamydospore production is of practical importance for the clinical mycology laboratory.

Performance of Candida ID, a New Chromogenic Medium for Presumptive Identification of Candida Species, in Comparison to CHROMagar Candida

PubMed Central

Willinger, Birgit; Hillowoth, Cornelia; Selitsch, Brigitte; Manafi, Mammad

2001-01-01

Candida ID agar allows identification of Candida albicans and differentiation of other Candida species. In comparison with CHROMagar Candida, we evaluated the performance of this medium directly from 596 clinical specimens. In particular, detection of C. albicans after 24 h of incubation was easier on Candida ID (sensitivity, 96.8%) than on CHROMagar (sensitivity, 49.6%). PMID:11574621

Development of selective media for the isolation and enumeration of Alternaria species from soil and plant debris.

PubMed

Hong, Soon Gyu; Pryor, Barry M

2004-07-01

A new semi-selective medium, acidified weak potato-dextrose agar (AWPDA) with Mertect (active ingredient: thiabendazole), was developed for the isolation and enumeration of Alternaria species from samples of soil and plant debris. The medium was selected based on growth inhibition tests against Alternaria and several other commonly encountered saprobic fungi utilizing three antifungal agents, Botran (active ingredient: dichloran), Bayleton (active ingredient: triadimefon), and Mertect, and two basal media, acidified potato-dextrose agar (APDA) and AWPDA. Botran inhibited growth of Rhizopus stolonifer moderately, but had little effect on Cladosporium cladosporoides, Fusarium oxysporum, Penicillium chrysogenum, or Trichoderma harzianum. Bayleton inhibited growth of R. stolonifer and C. cladosporoides severely, and inhibited growth of F. oxysporum, P. chrysogenum, and T. harzianum moderately. Mertect inhibited growth of C. cladosporoides, F. oxysporum, P. chrysogenum, and T. harzianum completely, but had little or moderate effect on R. stolonifer. All three antifungal agents inhibited growth of Alternaria species slightly or moderately. The combination of Bayleton and Mertect inhibited growth of all fungi severely. A comparison of recovery rates of Alternaria from soil and plant debris samples on AWPDA with Mertect and weak potato-dextrose agar (WPDA) revealed that Alternaria spp. accounted for 63.6%-81.0% of recovered fungal isolates on AWPDA with Mertect as compared to 0.6%-2.7% of recovered isolates on WPDA. The AWPDA medium with Mertect exhibited superior selective growth of Alternaria species from samples of soil and plant debris, and will be useful in studies where the recovery and enumeration of Alternaria species is necessary.

Inactivation of pathogenic bacteria inoculated onto a Bacto™ agar model surface using TiO2-UVC photocatalysis, UVC and chlorine treatments.

PubMed

Yoo, S; Ghafoor, K; Kim, S; Sun, Y W; Kim, J U; Yang, K; Lee, D-U; Shahbaz, H M; Park, J

2015-09-01

The aim of this study was to study inactivation of different pathogenic bacteria on agar model surface using TiO2-UV photocatalysis (TUVP). A unified food surface model was simulated using Bacto(™) agar, a routinely used microbial medium. The foodborne pathogenic bacteria Escherichia coli K12 (as a surrogate for E. coli O157:H7), Salmonella Typhimurium, Staphylococcus aureus and Listeria monocytogenes were inoculated onto the agar surface, followed by investigation of TUVP-assisted inactivation and morphological changes in bacterial cells. The TUVP process showed higher bacterial inactivation, particularly for Gram-negative bacteria, than UVC alone and a control (dark reaction). A TUVP treatment of 17·2 mW cm(-2) (30% lower than the UVC light intensity) reduced the microbial load on the agar surface by 4·5-6·0 log CFU cm(-2). UVC treatment of 23·7 mW cm(-2) caused 3·0-5·3 log CFU cm(-2) reduction. The use of agar model surface is effective for investigation of bacterial disinfection and TUVP is a promising nonthermal technique. The results showing effects of photocatalysis and other treatments for inactivation of bacterial pathogens on model surface can be useful for applying such processes for disinfection of fruit, vegetables and other similar surfaces. © 2015 The Society for Applied Microbiology.

Chemical test for mammalian feces in grain products: collaborative study.

PubMed

Gerber, H R

1989-01-01

A collaborative study was conducted to validate the use of the AOAC alkaline phosphatase method for mammalian feces in corn meal, 44.B01-44.B06, for 7 additional products: brown rice cream, oat bran, grits, semolina, pasta flour, farina, and barley plus (a mixture of barley, oat bran, and brown rice). The proposed method determines the presence of alkaline phosphatase, an enzyme contained in mammalian feces, by using phenolphthalein diphosphate as the enzyme substrate in a test agar medium. Fecal matter is separated from the grain products by specific gravity differences in 1% test agar. As the product is distributed on liquid test agar, fecal fragments float while the grain products sink. The alkaline phosphatase cleaves phosphate radicals from phenolphthalein diphosphate, generating free phenolphthalein, which produces a pink to red-purple color around the fecal particles in the previously colorless medium. Collaborators' recovery averages ranged from 21.7 particles (72.3%) for oat bran to 25.3 particles (84.3%) for semolina at the 30 particle spike level. Overall average background was 0.4 positive reactions per food type. The collaborators reported that the method was quick, simple, and easy to use. The method has been approved interim official first action for all 7 grain products.

Preparation, Imaging, and Quantification of Bacterial Surface Motility Assays

PubMed Central

Morales-Soto, Nydia; Anyan, Morgen E.; Mattingly, Anne E.; Madukoma, Chinedu S.; Harvey, Cameron W.; Alber, Mark; Déziel, Eric; Kearns, Daniel B.; Shrout, Joshua D.

2015-01-01

Bacterial surface motility, such as swarming, is commonly examined in the laboratory using plate assays that necessitate specific concentrations of agar and sometimes inclusion of specific nutrients in the growth medium. The preparation of such explicit media and surface growth conditions serves to provide the favorable conditions that allow not just bacterial growth but coordinated motility of bacteria over these surfaces within thin liquid films. Reproducibility of swarm plate and other surface motility plate assays can be a major challenge. Especially for more “temperate swarmers” that exhibit motility only within agar ranges of 0.4%-0.8% (wt/vol), minor changes in protocol or laboratory environment can greatly influence swarm assay results. “Wettability”, or water content at the liquid-solid-air interface of these plate assays, is often a key variable to be controlled. An additional challenge in assessing swarming is how to quantify observed differences between any two (or more) experiments. Here we detail a versatile two-phase protocol to prepare and image swarm assays. We include guidelines to circumvent the challenges commonly associated with swarm assay media preparation and quantification of data from these assays. We specifically demonstrate our method using bacteria that express fluorescent or bioluminescent genetic reporters like green fluorescent protein (GFP), luciferase (lux operon), or cellular stains to enable time-lapse optical imaging. We further demonstrate the ability of our method to track competing swarming species in the same experiment. PMID:25938934

Comparison of the BBL CHROMagar Staph aureus Agar Medium to Conventional Media for Detection of Staphylococcus aureus in Respiratory Samples

PubMed Central

Flayhart, Diane; Lema, Clara; Borek, Anita; Carroll, Karen C.

2004-01-01

Screening for Staphylococcus aureus has become routine in certain patient populations. This study is the first clinical evaluation of the BBL CHROMagar Staph aureus agar (CSA) medium (BD Diagnostics, Sparks, Md.) for detection of S. aureus in nasal surveillance cultures and in respiratory samples from cystic fibrosis (CF) patients. S. aureus colonies appear mauve on CSA. Other organisms are inhibited or produce a distinctly different colony color. S. aureus was identified from all media by slide coagulase, exogenous DNase, and mannitol fermentation assays. Susceptibility testing was performed using the agar dilution method. A total of 679 samples were evaluated. All samples were inoculated onto CSA. Nasal surveillance cultures were inoculated onto sheep blood agar (SBA) (BD Diagnostics), and samples from CF patients were inoculated onto mannitol salt agar (MSA) (BD Diagnostics). Of the 679 samples cultured, 200 organisms produced a mauve color on CSA (suspicious for S. aureus) and 180 were positive for S. aureus on SBA or MSA. Of 200 CSA-positive samples 191 were identified as S. aureus. Nine mauve colonies were slide coagulase negative and were subsequently identified as Staphylococcus lugdunensis (one), Staphylococcus epidermidis (three), Staphylococcus haemolyticus (one), and Corynebacterium species (four). CSA improved the ability to detect S. aureus by recovering 12 S. aureus isolates missed by conventional media. Of the 192 S. aureus isolates recovered, 122 were methicillin susceptible and 70 were methicillin resistant. Overall, the sensitivity and specificity of CSA in this study were 99.5 and 98%, respectively. There was no difference in the performance of the slide coagulase test or in susceptibility testing performed on S. aureus recovered from CSA compared to SBA or MSA. Our data support the use of CSA in place of standard culture media for detection of S. aureus in heavily contaminated respiratory samples. PMID:15297498

Exophiala pisciphila. A study of its development.

PubMed

Gaskins, J E; Cheung, P J

1986-03-01

Exophiala pisciphila is a dematiaceous fungus that belongs to a group of fungi known as the 'black yeasts'. It was isolated from the skin lesions of a smooth dogfish, Mustelus canis Mitchill, that had been born in the shark exhibit tank of the New York Aquarium. The different stages of development of this fungus were studied by light microscopy and scanning electron microscopy to illustrate the morphology and surface structures of conidia and mycelium. The list of marine and fresh water fish, which have been infected by Exophiala spp. and Exophiala-like fungi has been up-dated. Potato Dextrose Agar and Malt Agar proved to be the best growth media, while Corn Meal Agar proved to be the best medium for studying the morphological features of the conidia and mycelial development of E. pisciphila, which exhibited polymorphic conidiogenesis.

Evaluation of the Granada agar plate for detection of vaginal and rectal group B streptococci in pregnant women.

PubMed

Gil, E G; Rodríguez, M C; Bartolomé, R; Berjano, B; Cabero, L; Andreu, A

1999-08-01

Granada medium was evaluated for the detection of group B streptococci (GBS) in vaginal and rectal swabs compared with selective Columbia blood agar and selective Lim broth. From May 1996 to March 1998, 702 pregnant women (35 to 37 weeks of gestation) participated in this three-phase study; 103 (14.7%) of these women carried GBS. In the first phase of the experiment (n = 273 women), vaginorectal specimens were collected on the same swab; the sensitivities of Granada tube, selective Columbia blood agar, and Lim broth were 31.4, 94.3, and 74.3%, respectively. In the second and third phases (n = 429 women), vaginal and rectal specimens were collected separately; the sensitivities of Granada plate, selective Columbia blood agar, and Lim broth (subcultured at 4 h on selective Columbia agar in the second phase and at 18 to 24 h in Granada plate in the third phase) were 91.1, 83.9, and 75%, respectively, in the second phase and 88.5, 90.4, and 63.5%, respectively, in the third phase. There were no statistically significant differences in GBS recovery between the Granada agar plate and selective Columbia blood agar, but the Granada plate provided a clear advantage; the characteristic red-orange colonies produced overnight by GBS can be identified by the naked eye and is so specific that further identification is unnecessary. The use of the Granada tube and Lim broth did not result in increased isolation of GBS. In conclusion, the Granada agar plate is highly sensitive for detecting GBS in vaginal and rectal swabs from pregnant women and can provide results in 18 to 24 h.

Phacidiopycnis washingtonensis--a new species associated with pome fruits from Washington State.

PubMed

Xiao, C L; Rogers, J D; Kim, Y K; Liu, Q

2005-01-01

A new species of Phacidiopycnis associated with pome fruits is described. The fungus causes fruit rot on apples during storage and is associated with a twig dieback and canker disease of crabapple trees and dead twigs of pear trees. To characterize the biology of the fungus and compare it with Ph. piri, the type species of the genus, effects of nine media and light on mycelial growth and pycnidial production, mycelial growth in response to temperature and mode of conidial germination in response to nutrient were determined. Apple-juice agar, pear-juice agar, prune-juice agar, potato-dextrose agar (PDA) and malt-extract agar, Czapek-Dox agar and oatmeal agar (OMA) favored mycelial growth. Cornmeal agar (CMA) did not favor mycelial growth. Light effect on pycnidial formation was medium dependent. Abundant pycnidia with mature conidia formed in 14 d old PDA and OMA cultures at 20 C, regardless of light, whereas none or very few pycnidia formed on other media in the dark. Fluorescent light stimulated formation of pycnidia except on CMA. The fungus grew at -3-25 C, with optimum growth at 15-20 C. Conidia germinated either by forming germ tubes or less often by budding. Budding of conidia occurred in 1 and 10% pear-juice solutions but not in 100% pear-juice solution. Six isolates of Ph. washingtonensis from different species of pome fruits had identical ITS sequences. The sizes of the ITS region were the same for both Ph. washingtonensis and Ph. piri, and four polymorphic nucleotide sites were found in the ITS region between Ph. washingtonensis and Ph. piri. The similarity in ITS sequences between these two taxa is confirmatory evidence for the erection of the new species of Phacidiopycnis associated with pome fruits we describe here.

Rapid staining and enumeration of small numbers of total bacteria in water by solid-phase laser cytometry

NASA Technical Reports Server (NTRS)

Broadaway, Susan C.; Barton, Stephanie A.; Pyle, Barry H.

2003-01-01

The nucleic acid stain SYBR Green I was evaluated for use with solid-phase laser cytometry to obtain total bacterial cell counts from several water sources with small bacterial numbers. Results were obtained within 30 min and exceeded or equaled counts on R2A agar plates incubated for 14 days at room temperature.

Identification of exopolysaccharides produced by fluorescent pseudomonads associated with commercial mushroom (Agaricus bisporus) production.

PubMed

Fett, W F; Wells, J M; Cescutti, P; Wijey, C

1995-02-01

The acidic exopolysaccharides (EPSs) from 63 strains of mushroom production-associated fluorescent pseudomonads which were mucoid on Pseudomonas agar F medium (PAF) were isolated, partially purified, and characterized. The strains were originally isolated from discolored lesion which developed postharvest on mushroom (Agaricus bisporus) caps or from commercial lots of mushroom casing medium. An acidic galactoglucan, previously named marginalan, was produced by mucoid strains of the saprophyte Pseudomonas putida and the majority of mucoid strains of saprophytic P. fluorescens (biovars III and V) isolated from casing medium. One biovar II strain (J1) of P. fluorescens produced alginate, a copolymer of mannuronic and guluronic acids, and one strain (H13) produced an apparently unique EPS containing neutral and amino sugars. Of 10 strains of the pathogen "P. gingeri," the causal agent of mushroom ginger blotch, 8 gave mucoid growth on PAF. The "P. gingeri" EPS also was unique in containing both neutral sugar and glucuronic acid. Mucoid, weakly virulent strains of "P. reactans" produced either alginate or marginalan. All 10 strains of the pathogen P. tolaasii, the causal agent of brown blotch of mushrooms were nonnmucoid on PAF. Production of EPS by these 10 strains plus the 2 nonmucoid strains of "P. gingeri" also was negative on several additional solid media as well as in two broth media tested. The results support our previous studies indicating that fluorescent pseudomonads are a rich source of novel EPSs.

Identification of exopolysaccharides produced by fluorescent pseudomonads associated with commercial mushroom (Agaricus bisporus) production.

PubMed Central

Fett, W F; Wells, J M; Cescutti, P; Wijey, C

1995-01-01

The acidic exopolysaccharides (EPSs) from 63 strains of mushroom production-associated fluorescent pseudomonads which were mucoid on Pseudomonas agar F medium (PAF) were isolated, partially purified, and characterized. The strains were originally isolated from discolored lesion which developed postharvest on mushroom (Agaricus bisporus) caps or from commercial lots of mushroom casing medium. An acidic galactoglucan, previously named marginalan, was produced by mucoid strains of the saprophyte Pseudomonas putida and the majority of mucoid strains of saprophytic P. fluorescens (biovars III and V) isolated from casing medium. One biovar II strain (J1) of P. fluorescens produced alginate, a copolymer of mannuronic and guluronic acids, and one strain (H13) produced an apparently unique EPS containing neutral and amino sugars. Of 10 strains of the pathogen "P. gingeri," the causal agent of mushroom ginger blotch, 8 gave mucoid growth on PAF. The "P. gingeri" EPS also was unique in containing both neutral sugar and glucuronic acid. Mucoid, weakly virulent strains of "P. reactans" produced either alginate or marginalan. All 10 strains of the pathogen P. tolaasii, the causal agent of brown blotch of mushrooms were nonnmucoid on PAF. Production of EPS by these 10 strains plus the 2 nonmucoid strains of "P. gingeri" also was negative on several additional solid media as well as in two broth media tested. The results support our previous studies indicating that fluorescent pseudomonads are a rich source of novel EPSs. PMID:7574589

Performance of a New Chromogenic Medium, BBL CHROMagar MRSA II (BD), for Detection of Methicillin-Resistant Staphylococcus aureus in Screening Samples ▿

PubMed Central

Van Vaerenbergh, Kristien; Cartuyvels, Reinoud; Coppens, Guy; Frans, Johan; Van den Abeele, Anne-Marie; De Beenhouwer, Hans

2010-01-01

Two chromogenic media for the detection of MRSA were compared: BBL CHROMagar MRSA II (BD) and MRSA ID agar (bioMérieux). Following overnight nonselective enrichment, 1,919 screening samples were inoculated on both chromogenic agars. After 24 h, the sensitivities of both media were high and comparable. Both media showed an important decrease in specificity after 48 h of incubation (decreases of 8% for MRSA II and 10% for MRSA ID), but MRSA II was significantly more specific at both time points. PMID:20181915

Inhibitory effects of terpene alcohols and aldehydes on growth of green alga Chlorella pyrenoidosa

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ikawa, Miyoshi; Mosley, S.P.; Barbero, L.J.

1992-10-01

The growth of the green alga Chlorella pyrenoidosa was inhibited by terpene alcohols and the terpene aldehyde citral. The strongest activity was shown by citral. Nerol, geraniol, and citronellol also showed pronounced activity. Strong inhibition was linked to acyclic terpenes containing a primary alcohol or aldehyde function. Inhibition appeared to be taking place through the vapor phase rather than by diffusion through the agar medium from the terpene-treated paper disks used in the system. Inhibition through agar diffusion was shown by certain aged samples of terpene hydrocarbons but not by recently purchased samples.

Germ tube and chlamydospore formation by Candida albicans on a new medium.

PubMed Central

Beheshti, F; Smith, A G; Krause, G W

1975-01-01

A new medium composed of "cream of rice" infusion, oxgall, Tween 80, and agar is described for the sequential development of germ tubes and chlamydospores by Candida albicans. The procedure used (Dalmau's technique) is an improvement over the fluid substrate procedures previously advocated for germ tube formation. That the same preparation is then used for chlamydospore production is of practical importance for the clinical mycology laboratory. Images PMID:1102561

Use of sucrose-agar globule with root exudates for mass production of vesicular arbuscular mycorrhizal fungi.

PubMed

Selvaraj, Thangaswamy; Kim, Hoon

2004-03-01

A sucrose-agar globule (SAG) was newly introduced to increase production of the vesicular arbuscular mycorrhizal (VAM) fungal spores, Gigaspora gigantea and Glomus fasciculatum. An SAG inoculum and a sucrose-agar globule with root exudates (SAGE) inoculum were prepared, and their spore productions were compared with a soil inoculum. When the SAGE was used as the inoculum on sucrose-agar medium plates the number of spores was increased (35% more than the soil inoculum). After the soil inoculum and SAGE were inoculated on an experimental plant, Zingiber officinale, the percentage root colonization, number of VAM spores, and dry matter content were analyzed. It was observed that the SAGE showed a higher percentage of root colonization (about 10% more), and increases in the number of spores (about 26%) and dry matter (more than 13%) for the two VAM fungal spores than the soil inoculum. The results of this study suggested that the SAGE inoculum may be useful for the mass production of VAM fungi and also for the large scale production of VAM fungal fertilizer.

Development of blood-yolk-polymyxin B-trimethoprim agar for the enumeration of Bacillus cereus in various foods.

PubMed

Kim, Dong-Hyeon; Kim, Hyunsook; Chon, Jung-Whan; Moon, Jin-San; Song, Kwang-Young; Seo, Kun-Ho

2013-07-15

Blood-yolk-polymyxin B-trimethoprim agar (BYPTA) was developed by the addition of egg yolk, laked horse blood, sodium pyruvate, polymyxin B, and trimethoprim, and compared with mannitol-yolk-polymyxin B agar (MYPA) for the isolation and enumeration of Bacillus cereus (B. cereus) in pure culture and various food samples. In pure culture, there was no statistical difference (p>0.05) between the recoverability and sensitivity of MYPA and BYPTA, whereas BYPTA exhibited higher specificity (p<0.05). To evaluate BYPTA agar with food samples, B. cereus was experimentally spiked into six types of foods, triangle kimbab, sandwich, misugaru, Saengsik, red pepper powder, and soybean paste. No statistical difference was observed in recoverability (p>0.05) between MYPA and BYPTA in all tested foods, whereas BYPTA exhibited higher selectivity than MYPA, especially in foods with high background microflora, such as Saengsik, red pepper powder, and soybean paste. The newly developed selective medium BYPTA could be a useful enumeration tool to assess the level of B. cereus in foods, particularly with high background microflora. Copyright © 2013 Elsevier B.V. All rights reserved.

Mupirocin-mucin agar for selective enumeration of Bifidobacterium bifidum.

PubMed

Pechar, Radko; Rada, Vojtech; Parafati, Lucia; Musilova, Sarka; Bunesova, Vera; Vlkova, Eva; Killer, Jiri; Mrazek, Jakub; Kmet, Vladimir; Svejstil, Roman

2014-11-17

Bifidobacterium bifidum is a bacterial species exclusively found in the human intestinal tract. This species is becoming increasingly popular as a probiotic organism added to lyophilized products. In this study, porcine mucin was used as the sole carbon source for the selective enumeration of B. bifidum in probiotic food additives. Thirty-six bifidobacterial strains were cultivated in broth with mucin. Only 13 strains of B. bifidum utilized the mucin to produce acids. B. bifidum was selectively enumerated in eight probiotic food supplements using agar (MM agar) containing mupirocin (100 mg/L) and mucin (20 g/L) as the sole carbon source. MM agar was fully selective if the B. bifidum species was presented together with Bifidobacterium animalis subsp. lactis, Bifidobacterium breve, and Bifidobacterium longum subsp. longum species and with lactic acid bacteria (lactobacilli, streptococci). Isolated strains of B. bifidum were identified using biochemical, PCR, MALDI-TOF procedures and 16S rRNA gene sequencing. The novel selective medium was also suitable for the isolation of B. bifidum strains from human fecal samples. Copyright © 2014 Elsevier B.V. All rights reserved.

Evaluation of the chitin-binding dye Congo red as a selection agent for the isolation, classification, and enumeration of ascomycete yeasts.

PubMed

Linder, Tomas

2018-05-01

Thirty-nine strains of ascomycete yeasts representing 35 species and 33 genera were tested for their ability to grow on solid agar medium containing increasing concentrations of the chitin-binding dye Congo red. Six strains were classified as hypersensitive (weak or no growth at 10 mg/l Congo red), five were moderately sensitive (weak or no growth at 50 mg/l), three were moderately tolerant (weak or no growth at 100 mg/l), while the remaining 25 strains were classified as resistant (robust growth at ≥ 100 mg/l) with 20 of these strains classified as hyper-resistant (robust growth at 200 mg/l). Congo red growth phenotypes were consistent within some families but not others. The frequency of Congo red resistance among ascomycete yeasts was deemed too high for the practical use of Congo red as a selection agent for targeted isolation, but can be useful for identification and enumeration of yeasts.

Natural Competence of Xylella fastidiosa Occurs at a High Frequency Inside Microfluidic Chambers Mimicking the Bacterium's Natural Habitats

PubMed Central

Kandel, Prem P.; Lopez, Samantha M.; Almeida, Rodrigo P. P.

2016-01-01

ABSTRACT Xylella fastidiosa is a xylem-limited bacterium that is the causal agent of emerging diseases in a number of economically important crops. Genetic diversity studies have demonstrated homologous recombination occurring among X. fastidiosa strains, which has been proposed to contribute to host plant shifts. Moreover, experimental evidence confirmed that X. fastidiosa is naturally competent for recombination in vitro. Here, as an approximation of natural habitats (plant xylem vessels and insect mouthparts), recombination was studied in microfluidic chambers (MCs) filled with media amended with grapevine xylem sap. First, different media were screened for recombination in solid agar plates using a pair of X. fastidiosa strains that were previously reported to recombine in coculture. The highest frequency of recombination was obtained with PD3 medium, compared to those with the other two media (X. fastidiosa medium [XFM] and periwinkle wilt [PW] medium) used in previous studies. Dissection of the media components led to the identification of bovine serum albumin as an inhibitor of recombination that was correlated to its previously known effect on inhibition of twitching motility. When recombination was performed in liquid culture, the frequencies were significantly higher under flow conditions (MCs) than under batch conditions (test tubes). The recombination frequencies in MCs and agar plates were not significantly different from each other. Grapevine xylem sap from both susceptible and tolerant varieties allowed high recombination frequency in MCs when mixed with PD3. These results suggest that X. fastidiosa has the ability to be naturally competent in the natural growth environment of liquid flow, and this phenomenon could have implications in X. fastidiosa environmental adaptation. IMPORTANCE Xylella fastidiosa is a plant pathogen that lives inside xylem vessels (where water and nutrients are transported inside the plant) and the mouthparts of insect vectors. This bacterium causes emerging diseases in various crops worldwide, including recent outbreaks in Europe. The mechanisms by which this bacterium adapts to new hosts is not understood, but it was previously shown that it is naturally competent, meaning that it can take up DNA from the environment and incorporate it into its genome (recombination). In this study, we show that the frequency of recombination is highest when the bacterium is grown under flow conditions in microfluidic chambers modeled after its natural habitats, and recombination was still high when the medium was amended with grapevine sap. Our results suggest that this bacterium is able to recombine when growing inside plants or insects, and this can be a mechanism of adaptation of this pathogen that causes incurable diseases. PMID:27316962

Identification of thermophilic bacteria in solid-waste composting.

PubMed Central

Strom, P F

1985-01-01

The thermophilic microbiota of solid-waste composting, with major emphasis on Bacillus spp., was examined with Trypticase soy broth (BBL Microbiology Systems) with 2% agar as the initial plating medium. Five 4.5-liter laboratory units at 49 to 69 degrees C were fed a mixture of dried table scraps and shredded newspaper. The composting plants treating refuse at Altoona, Pa., and refuse-sludge at Leicester, England, were also sampled. Of 652 randomly picked colonies, 87% were identified as Bacillus spp. Other isolates included two genera of unidentified nonsporeforming bacteria (one of gram-negative small rods and the other of gram-variable coccobacilli), the actinomycetes Streptomyces spp. and Thermoactinomyces sp., and the fungus Aspergillus fumigatus. Among the Bacillus isolates, the following, in order of decreasing frequency, were observed: B. circulans complex, B. stearothermophilus, B. coagulans types A and B, B. licheniformis, B. brevis, B. sphaericus, Bacillus spp. types i and ii, and B. subtilis. About 15% of the Bacillus isolates could be assigned to species only by allowing for greater variability in one or more characteristics than has been reported by other authors for their strains. In particular, growth at higher temperatures than previously reported was found for strains of several species. A small number of Bacillus isolates (less than 2%) could not be assigned to any recognized species. PMID:4083886

Role of water-soluble polysaccharides in bacterial cellulose production.

PubMed

Ishida, Takehiko; Mitarai, Makoto; Sugano, Yasushi; Shoda, Makoto

2003-08-20

Acetobacter xylinum BPR2001 produces water-insoluble bacterial cellulose (BC) and a water-soluble polysaccharide called acetan in corn steep liquor-fructose medium. Acetobacter xylinum EP1, which is incapable of acetan production was derived by disrupting the aceA gene of BPR2001. The BC production by EP1 (2.88 g/L) was lower than that by BPR2001 (4.6 g/L) in baffled-flask culture. When purified acetan or agar was added to the medium from the start of cultivation, the BC production by EP1 was enhanced and the final BC yield of EP1 was almost the same as that of BPR2001. A similar improvement of BC production by EP1 by the addition of agar was also confirmed by cultivation in a 50-L airlift reactor. From these results, the role of acetan in BC production is associated with the increase in the viscosity of the culture medium which may hinder coagulation of BC and cells in the culture, thereby accelerating the growth of BPR2001 and BC production by BPR2001. Copyright 2003 Wiley Periodicals, Inc. Biotechnol Bioeng 83: 474-478, 2003.

The molecular bacterial load assay replaces solid culture for measuring early bactericidal response to antituberculosis treatment.

PubMed

Honeyborne, Isobella; Mtafya, Bariki; Phillips, Patrick P J; Hoelscher, Michael; Ntinginya, Elias N; Kohlenberg, Anke; Rachow, Andrea; Rojas-Ponce, Gabriel; McHugh, Timothy D; Heinrich, Norbert

2014-08-01

We evaluated the use of the molecular bacterial load (MBL) assay, for measuring viable Mycobacterium tuberculosis in sputum, in comparison with solid agar and liquid culture. The MBL assay provides early information on the rate of decline in bacterial load and has technical advantages over culture in either form. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Use of Selective Fungal Culture Media Increases Rates of Detection of Fungi in the Respiratory Tract of Cystic Fibrosis Patients.

PubMed

Hong, Gina; Miller, Heather B; Allgood, Sarah; Lee, Richard; Lechtzin, Noah; Zhang, Sean X

2017-04-01

The prevalence of fungi in the respiratory tracts of cystic fibrosis (CF) patients has risen. However, fungal surveillance is not routinely performed in most clinical centers in the United States, which may lead to an underestimation of the true prevalence of the problem. We conducted a prospective study comparing the rates of detection for clinically important fungi (CIF), defined as Aspergillus , Scedosporium , and Trichosporon species and Exophiala dermatitidis , in CF sputa using standard bacterial and selective fungal culture media, including Sabouraud dextrose agar with gentamicin (SDA), inhibitory mold agar (IMA), and brain heart infusion (BHI) agar with chloramphenicol and gentamicin. We described the prevalence of these fungi in an adult CF population. A total of 487 CF respiratory samples were collected from 211 unique participants. CIF were detected in 184 (37.8%) samples. Only 26.1% of CIF-positive samples were detected in bacterial culture medium, whereas greater rates of detection for fungi were found in IMA (65.8%; P < 0.001), in SDA (at 30°C, 64.7%; P = 0.005), and in BHI agar (63.0%; P = 0.001). The prevalences of Aspergillus and Scedosporium species were 40.8% and 5.2%, respectively, which are greater than the nationally reported prevalence numbers of 20.4% and 1.9%. Selective fungal culture media and longer incubation periods yielded higher rates of detection for CIF in CF sputum samples compared with that detected in bacterial culture medium, resulting in an underdetection of fungi by bacterial culture alone. The prevalence of fungi in CF may be better estimated by using selective fungal culture media, and this may translate to important clinical decisions. Copyright © 2017 American Society for Microbiology.

Use of Selective Fungal Culture Media Increases Rates of Detection of Fungi in the Respiratory Tract of Cystic Fibrosis Patients

PubMed Central

Hong, Gina; Miller, Heather B.; Allgood, Sarah; Lee, Richard; Lechtzin, Noah

2017-01-01

ABSTRACT The prevalence of fungi in the respiratory tracts of cystic fibrosis (CF) patients has risen. However, fungal surveillance is not routinely performed in most clinical centers in the United States, which may lead to an underestimation of the true prevalence of the problem. We conducted a prospective study comparing the rates of detection for clinically important fungi (CIF), defined as Aspergillus, Scedosporium, and Trichosporon species and Exophiala dermatitidis, in CF sputa using standard bacterial and selective fungal culture media, including Sabouraud dextrose agar with gentamicin (SDA), inhibitory mold agar (IMA), and brain heart infusion (BHI) agar with chloramphenicol and gentamicin. We described the prevalence of these fungi in an adult CF population. A total of 487 CF respiratory samples were collected from 211 unique participants. CIF were detected in 184 (37.8%) samples. Only 26.1% of CIF-positive samples were detected in bacterial culture medium, whereas greater rates of detection for fungi were found in IMA (65.8%; P < 0.001), in SDA (at 30°C, 64.7%; P = 0.005), and in BHI agar (63.0%; P = 0.001). The prevalences of Aspergillus and Scedosporium species were 40.8% and 5.2%, respectively, which are greater than the nationally reported prevalence numbers of 20.4% and 1.9%. Selective fungal culture media and longer incubation periods yielded higher rates of detection for CIF in CF sputum samples compared with that detected in bacterial culture medium, resulting in an underdetection of fungi by bacterial culture alone. The prevalence of fungi in CF may be better estimated by using selective fungal culture media, and this may translate to important clinical decisions. PMID:28100601

Diagnostic accuracy of a standardized scheme for identification of Streptococcus uberis in quarter milk samples: A comparison between conventional bacteriological examination, modified Rambach agar medium culturing, and 16S rRNA gene sequencing.

PubMed

Wald, Regina; Baumgartner, Martina; Urbantke, Verena; Stessl, Beatrix; Wittek, Thomas

2017-02-01

Bacteriological examination of milk samples is a prerequisite for pathogen-specific therapy and aids in limiting antimicrobial resistance. The aims of this study were to establish a standardized scheme for reliable Streptococcus uberis identification in routine diagnosis and to evaluate the accuracy of conventional tests and growing patterns of Strep. uberis on a selective medium (modified Rambach agar medium, MRAM) using 16S rRNA gene sequencing analysis as a reference method. We obtained isolates of presumptive Strep. uberis (n = 336) from quarter milk samples of dairy cows with intramammary infections and classified the isolates into 2 clusters using biochemical characterization. In cluster 1 (n = 280), cocci grew as non-hemolytic colonies, hydrolyzing esculin, carrying no Lancefield antigen (A/B/C/D/G) or Christie Atkins Munch-Petersen factor, and their growth was inhibited on an Enterococcus agar. Production of β-d-galactosidase on MRAM was shown by 257 of the cluster 1 isolates (91.79%), and 16S rRNA gene sequencing verified 271 (96.79%) of the isolates to be Strep. uberis. In 264 isolates (94.29%), MRAM agreed with the sequencing results. In cluster 2 (n = 56), isolates showed different characteristics: 37 (66.07%) were β-d-galactosidase-positive, and based on 16S sequencing results, 36 (64.29%) were identified correctly as Strep. uberis using biochemical methods. Identification success in this group differed significantly between routine diagnosis and MRAM application: MRAM agreed with sequencing results in 47 isolates (83.93%). To identify Strep. uberis and differentiate it from other lactic acid bacteria in routine diagnosis, we suggest using catalase reaction, hemolysis, esculin hydrolysis, and growth on enterococci agar. Isolates that show a typical biochemical profile can be identified satisfactorily with these tests. For Strep. uberis isolates with divergent patterns, application of MRAM as a follow-up test increased the diagnostic accuracy to 94.64%. Copyright © 2017 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Comparison of Chromogenic Selective Media for the Detection of Cronobacter spp. (Enterobacter sakazakii).

PubMed

Teramura, Hajime; Fukuda, Noriko; Okada, Yumiko; Ogihara, Hirokazu

2018-01-01

　The four types of chromogenic selective media that are commercially available in Japan were compared for establishing a Japanese standard method for detecting Cronobacter spp. based on ISO/TS 22964:2006. When assessed using 9 standard Cronobacter spp. strains and 29 non-Cronobacter strains, Enterobacter sakazakii isolation agar, Chromocult TM Enterobacter sakazakii agar, CHROMagar TM E. sakazakii, and XM-sakazakii agar demonstrated excellent inclusivity and exclusivity. Using the ISO/TS 22964:2006 method, the recovered numbers of 38 Cronobacter spp. strains, including 29 C. sakazakii isolates obtained from each medium, were equivalent, indicating that there was no significant difference (p > 0.05) among the four types of chromogenic selective media. Thus, we demonstrated that these four chromogenic selective media are suitable alternatives when using the standard method for detecting Cronobacter spp. in Japan, based on the ISO/TS 22964:2006.

Study of cellulolytic soil fungi and two nova species and new medium

PubMed Central

Khalid, Mahmood; Yang, Wei-jun; Kishwar, Nazir; Rajput, Zahid Iqbal; Arijo, Abdullah G.

2006-01-01

This study is aimed at identifying and determining the percentage of occurrence frequency of cellulose decomposing soil fungi. The soil samples were inoculated into culture plates prepared in Sabouraud medium under sterilized conditions and incubated at 30 °C for 4 to 7 d. The identified fungal species were incubated in self-designed cellulose medium for testing their cellulolytic ability. Forty-two species, including 2 nova species, representing sixteen genera showed growth and sporulation in the cellulose medium. Most of the isolated species were from genus Aspergillus and Penicillium. Aspergillus niger and Mucor hiemalis showed highest occurrence frequency (45% and 36% respectively), as these species were collected from about 80% of soil samples. Being agar free and cheaper, the new fungal medium designed showed results equivalent to Sabouraud medium. PMID:16691640

Cellulase production by pink pigmented facultative methylotrophic strains (PPFMs).

PubMed

Jayashree, Shanmugam; Lalitha, Rajendran; Vadivukkarasi, Ponnusamy; Kato, Yuko; Seshadri, Sundaram

2011-07-01

Pink pigmented facultative methylotrophs (PPFM) isolated from water samples of Cooum and Adyar rivers in Chennai and soil samples of forests located in various districts of Tamil Nadu, India were screened for cellulase production using carboxymethylcellulose agar (CMC agar) medium. The strains showed wide variations in the production of clearing zones around the colonies on CMC agar medium flooded with Congo red. CMCase and filter paper assays were used to quantitatively measure the cellulase activity of 13 PPFM strains. Among the strains, Methylobacterium gregans, MNW 60, MHW 109, MSF 34, and MSF 40 showed cellulolytic activity ranging from 0.73 to 1.16 U mL(-1) with wide temperature (35-65°C) and pH (5 to 8) tolerance. SDS-PAGE analysis of the crude enzyme of PPFM strain MNW 60 exhibited several protein bands, and zymogram analysis revealed two dimeric cellulase bands with molecular mass of ~92 and 42 kDa. Scanning electron microscopic studies revealed significant morphological differences between the cells grown in normal and CMC amended medium. The strain MNW 60 was identified as Methylobacterium sp. based on biochemical, physiological, and morphological analyses, and the methylotrophic nature was authenticated by the presence of mxaF gene, encoding methanol dehydrogenase as a key indicator enzyme of methylotrophs, with 99% similarity to Methylobacterium lusitanum. With the 16S ribosomal RNA sequence showing 97% similarity to M. lusitanum strain MP2, this can be proposed as a novel taxon of the genus Methylobacterium. The study forms the first detailed report on the extracellular cellulase production by pink pigmented Methylobacterium sp., and it is expected that this might be the basis for further studies on cellulase production by PPFMs to explore the molecular mechanism, strain improvement, and large-scale cellulase production for its application.

Growth on solid media.

PubMed

Elbing, Karen; Brent, Roger

2002-08-01

Detailed protocols are provided for titering and isolating bacterial colonies by serial dilutions, or alternatively by streaking or spreading a plate. Support protocols describe replica plating as well as methods for storing strains as agar stabs or frozen glycerol stocks.

Proteomic signatures differentiating Bacillus anthracis Sterne sporulation on soil relative to laboratory media.

PubMed

Wunschel, D S; Hutchison, J R; Deatherage Kaiser, B L; Merkley, E D; Hess, B M; Lin, A; Warner, M G

2017-12-18

The process of sporulation is vital for the stability and infectious cycle of Bacillus anthracis. The spore is the infectious form of the organism and therefore relevant to biodefense. While the morphological and molecular events occurring during sporulation have been well studied, the influence of growth medium and temperature on the proteins expressed in sporulated cultures is not well understood. Understanding the features of B. anthracis sporulation specific to natural vs. laboratory production will address an important question in microbial forensics. In an effort to bridge this knowledge gap, a system for sporulation on two types of agar-immobilized soils was used for comparison to cultures sporulated on two common types of solid laboratory media, and one liquid sporulation medium. The total number of proteins identified as well as their identity differed between samples generated in each medium and growth temperature, demonstrating that sporulation environment significantly impacts the protein content of the spore. In addition, a subset of proteins common in all of the soil-cultivated samples was distinct from the expression profiles in laboratory medium (and vice versa). These differences included proteins involved in thiamine and phosphate metabolism in the sporulated cultures produced on soils with a notable increase in expression of ATP binding cassette (ABC) transporters annotated to be for phosphate and antimicrobial peptides. A distinct set of ABC transporters for amino acids, sugars and oligopeptides were found in cultures produced on laboratory media as well as increases in carbon and amino acid metabolism-related proteins. These protein expression changes indicate that the sporulation environment impacts the protein profiles in specific ways that are reflected in the metabolic and membrane transporter proteins present in sporulated cultures.

Screening of micro-organisms for decolorization of melanins produced by bluestain fungi.

PubMed

Rättö, M; Chatani, M; Ritschkoff, A C; Viikari, L

2001-03-01

A total of 17 fungi and four bacteria were screened for their ability to decolorize melanin, using isolated extracellular melanin of the bluestain fungus Aureobasidium pullulans as substrate. On agar media, decolorization was observed by four fungal strains: Bjerkandera adusta VTT-D-99746, Galactomyces geotrichum VTT-D-84228, Trametes hirsuta VTT-D-95443 and Trametes versicolor VTT-D-99747. The four fungi were more efficient on nitrogen-limited medium than on complete medium. The melanin-decolorizing activity of G. geotrichum appeared to be located on the mycelium and could be liberated into the medium enzymatically.

Performance of transport and selective media for swine Bordetella bronchiseptica recovery and it comparison to polymerase chain reaction detection

PubMed Central

Coutinho, Tania Alen; Bernardi, Mari Lourdes; de Itapema Cardoso, Marisa Ribeiro; Borowski, Sandra Maria; Moreno, Andrea Micke; de Barcellos, David Emilio Santos Neves

2009-01-01

Three comparative assays were performed seeking to improve the sensitivity of the diagnosis of Bordetella bronchiseptica infection analyzing swine nasal swabs. An initial assay compared the recovery of B. bronchiseptica from swabs simultaneously inoculated with B. bronchiseptica and some interfering bacteria, immersed into three transport formulations (Amies with charcoal, trypticase soy broth and phosphate buffer according to Soerensen supplemented with 5% of bovine fetal serum) and submitted to different temperatures (10°C and 27°C) and periods of incubation (24, 72 and 120 hours). A subsequent assay compared three selective media (MacConkey agar, modified selective medium G20G and a ceftiofur medium) for their recovery capabilities from clinical specimens. One last assay compared the polymerase chain reaction to the three selective media. In the first assay, the recovery of B. bronchiseptica from transport systems was better at 27°C and the three formulations had good performances at this temperature, but the collection of qualitative and quantitative analysis indicated the advantage of Amies medium for nasal swabs transportation. The second assay indicated that MacConkey agar and modified G20G had similar results and were superior to the ceftiofur medium. In the final assay, polymerase chain reaction presented superior capability of B. bronchiseptica detection to culture procedures. PMID:24031390

Grain dust originating from organic and conventional farming as a potential source of biological agents causing respiratory diseases in farmers.

PubMed

Zukiewicz-Sobczak, Wioletta A; Cholewa, Grażyna; Krasowska, Ewelina; Chmielewska-Badora, Jolanta; Zwoliński, Jacek; Sobczak, Paweł

2013-12-01

Agricultural producers are exposed to a number of different health risks associated with their work environment. The objective of the study was to assess the degree of colonization by fungi in terms of quantity and in terms of variety of species the samples taken from the settled dust from combine threshing of rye cultivation from organic and conventional farms in the Province of Lublin. This paper is a preliminary quantitative assessment of the species of fungi colonizing the samples of settled dust collected during combine threshing from organic and conventional farms in the Province of Lublin. One of the stages of the project was the classification of biosafety BSL (biosafety level) of selected isolates and API ZYM tests to evaluate the potential ability of isolates to cause adverse health effects. To determine the concentration and composition of fungi in collected samples plate dilution method was used with two media: Malt Agar and Potato Dextrose Agar. MOST COMMONLY ISOLATED FUNGI IN SETTLED DUST SAMPLES COLLECTED DURING COMBINE THRESHING FROM ORGANIC FARMS, ON PDA MEDIUM WERE: Alternaria alternata and Aureobasidium pullulans. Cultures on MA medium were dominated by Alternaria alternata, Mycelia sterilia and Fusarium poae. In samples of dust from conventional crops, the predominant species was Alternaria alternata on PDA medium and on MA medium. The obtained results show a potential risk of people involved in agricultural work.

STUDIES ON THE USE OF AMPICILLIN-DEXTRIN AGAR ASAS AEROMONAS RECOVERY MEDIUM

EPA Science Inventory

The Contaminant Candidate List (CCL) includes the unregulated chemical and microbiological contaminants the EPA has identified as possibly posing a significant public risk to consumers if present in drinking water (1). There are three bacterial species listed in the CCL (Aeromon...

[Performance evaluation of Rapid™ Yeast Plus (Remel) system from two different culture media].

PubMed

Romeo, Ana M; Snitman, Gabriela V; Marucco, Andrea P; Ponce, Graciela Del V; Cataldi, Silvana P; Guelfand, Liliana I; Arechavala, Alicia

Within the genus Candida, Candida albicans is the most commonly isolated species from clinical samples. Due to the emergence of other species which can show a higher index of antifungal resistance, a fast identification of these species is necessary. The aim of this work was to evaluate the performance of the RapID Yeast Plus system from two different subculture media formulations: Sabouraud dextrose agar adjusted by Emmons (the medium is indicated in the equipment insert) and Sabouraud glucose agar, which is the most frequently used in Buenos Aires City laboratories. One hundred and sixty-six clinical sample strains coming from different hospitals belonging to the Mycology Network of Buenos Aires City were studied. From the obtained results, we conclude that the conditions and culture medium indicated by the manufacturer should be followed. Copyright © 2016 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.

Phenotypic and genotypic detection of Candida albicans and Candida dubliniensis strains isolated from oral mucosa of AIDS pediatric patients

PubMed Central

Livério, Harisson Oliveira; Ruiz, Luciana da Silva; de Freitas, Roseli Santos; Nishikaku, Angela; de Souza, Ana Clara; Paula, Claudete Rodrigues; Domaneschi, Carina

2017-01-01

ABSTRACT The aim of this study was to assess a collection of yeasts to verify the presence of Candida dubliniensis among strains isolated from the oral mucosa of AIDS pediatric patients which were initially characterized as Candida albicans by the traditional phenotypic method, as well as to evaluate the main phenotypic methods used in the discrimination between the two species and confirm the identification through genotypic techniques, i.e., DNA sequencing. Twenty-nine samples of C. albicans isolated from this population and kept in a fungi collection were evaluated and re-characterized. In order to differentiate the two species, phenotypic tests (Thermotolerance tests, Chromogenic medium, Staib agar, Tobacco agar, Hypertonic medium) were performed and genotypic techniques using DNA sequencing were employed for confirmation of isolated species. Susceptibility and specificity were calculated for each test. No phenotypic test alone was sufficient to provide definitive identification of C. dubliniensis or C. albicans, as opposed to results of molecular tests. After amplification and sequencing of specific regions of the 29 studied strains, 93.1% of the isolates were identified as C. albicans and 6.9% as C. dubliniensis. The Staib agar assay showed a higher susceptibility (96.3%) in comparison with other phenotypic techniques. Therefore, genotypic methods are indispensable for the conclusive identification and differentiation between these species. PMID:28423089

Phenotypic and genotypic detection of Candida albicans and Candida dubliniensis strains isolated from oral mucosa of AIDS pediatric patients.

PubMed

Livério, Harisson Oliveira; Ruiz, Luciana da Silva; Freitas, Roseli Santos de; Nishikaku, Angela; Souza, Ana Clara de; Paula, Claudete Rodrigues; Domaneschi, Carina

2017-04-13

The aim of this study was to assess a collection of yeasts to verify the presence of Candida dubliniensis among strains isolated from the oral mucosa of AIDS pediatric patients which were initially characterized as Candida albicans by the traditional phenotypic method, as well as to evaluate the main phenotypic methods used in the discrimination between the two species and confirm the identification through genotypic techniques, i.e., DNA sequencing. Twenty-nine samples of C. albicans isolated from this population and kept in a fungi collection were evaluated and re-characterized. In order to differentiate the two species, phenotypic tests (Thermotolerance tests, Chromogenic medium, Staib agar, Tobacco agar, Hypertonic medium) were performed and genotypic techniques using DNA sequencing were employed for confirmation of isolated species. Susceptibility and specificity were calculated for each test. No phenotypic test alone was sufficient to provide definitive identification of C. dubliniensis or C. albicans, as opposed to results of molecular tests. After amplification and sequencing of specific regions of the 29 studied strains, 93.1% of the isolates were identified as C. albicans and 6.9% as C. dubliniensis. The Staib agar assay showed a higher susceptibility (96.3%) in comparison with other phenotypic techniques. Therefore, genotypic methods are indispensable for the conclusive identification and differentiation between these species.

Wood and humus decay strategies by white-rot basidiomycetes correlate with two different dye decolorization and enzyme secretion patterns on agar plates.

PubMed

Barrasa, José M; Blanco, María N; Esteve-Raventós, Fernando; Altés, Alberto; Checa, Julia; Martínez, Angel T; Ruiz-Dueñas, Francisco J

2014-11-01

During several forays for ligninolytic fungi in different Spanish native forests, 35 white-rot basidiomycetes growing on dead wood (16 species from 12 genera) and leaf litter (19 species from 10 genera) were selected for their ability to decolorize two recalcitrant aromatic dyes (Reactive Blue 38 and Reactive Black 5) added to malt extract agar medium. In this study, two dye decolorization patterns were observed and correlated with two ecophysiological groups (wood and humus white-rot basidiomycetes) and three taxonomical groups (orders Polyporales, Hymenochaetales and Agaricales). Depending on the above groups, different decolorization zones were observed on the dye-containing plates, being restricted to the colony area or extending to the surrounding medium, which suggested two different decay strategies. These two strategies were related to the ability to secrete peroxidases and laccases inside (white-rot wood Polyporales, Hymenochaetales and Agaricales) and outside (white-rot humus Agaricales) of the fungal colony, as revealed by enzymatic tests performed directly on the agar plates. Similar oxidoreductases production patterns were observed when fungi were grown in the absence of dyes, although the set of enzyme released was different. All these results suggest that the decolorization patterns observed could be related with the existence of two decay strategies developed by white-rot basidiomycetes adapted to wood and leaf litter decay in the field. Published by Elsevier Inc.

INFLUENCE OF IODOFORM ON ANTIMICROBIAL POTENTIAL OF CALCIUM HYDROXIDE

PubMed Central

Estrela, Carlos; Estrela, Cyntia Rodrigues de Araújo; Hollanda, Augusto César Braz; Decurcio, Daniel de Almeida; Pécora, Jesus Djalma

2006-01-01

The purpose of this research was to verify the influence of Iodoform on antimicrobial potential of calcium hydroxide. S. aureus, E. faecalis, P. aeruginosa, B. subtilis, C. albicans were the biological indicators. The substances tested were: calcium hydroxide + saline; calcium hydroxide + Iodoform + saline; Iodoform + saline. For the agar diffusion test, 18 Petri plates with 20 ml of BHI agar were inoculated with the microbial suspensions. Fifty-four cavities were made and filled with the substances tested. The diameters of microbial inhibition were then measured. In direct exposure test, 162 #50 sterile absorbent paper points were immersed in the experimental suspensions for 5 min, and covered with the pastes. At intervals of 24, 48 and 72 hours, the paper points were immersed in 10 ml of Letheen Broth, followed by incubation at 37°°C for 48h. Microbial growth was evaluated by turbidity of the culture medium. A 0.1 ml inoculum obtained from the Letheen Broth was transferred to 7 ml of BHI, and incubated at 37°°C for 48h. Bacterial growth was again evaluated by turbidity of the culture medium. The calcium hydroxide associated with the saline or the iodoform plus saline showed antimicrobial effectiveness in both experimental methods. The iodoform paste presented antimicrobial ineffectiveness for the agar diffusion test on all biological microorganisms and for the direct exposure test on B. subtilis and on the mixture. PMID:19089027

Visualization of the Charcoal Agar Resazurin Assay for Semi-quantitative, Medium-throughput Enumeration of Mycobacteria.

PubMed

Gold, Ben; Roberts, Julia; Ling, Yan; Lopez Quezada, Landys; Glasheen, Jou; Ballinger, Elaine; Somersan-Karakaya, Selin; Warrier, Thulasi; Nathan, Carl

2016-12-14

There is an urgent need to discover and progress anti-infectives that shorten the duration of tuberculosis (TB) treatment. Mycobacterium tuberculosis, the etiological agent of TB, is refractory to rapid and lasting chemotherapy due to the presence of bacilli exhibiting phenotypic drug resistance. The charcoal agar resazurin assay (CARA) was developed as a tool to characterize active molecules discovered by high-throughput screening campaigns against replicating and non-replicating M. tuberculosis. Inclusion of activated charcoal in bacteriologic agar medium helps mitigate the impact of compound carry-over, and eliminates the requirement to pre-dilute cells prior to spotting on CARA microplates. After a 7-10 day incubation period at 37 °C, the reduction of resazurin by mycobacterial microcolonies growing on the surface of CARA microplate wells permits semi-quantitative assessment of bacterial numbers via fluorometry. The CARA detects approximately a 2-3 log10 difference in bacterial numbers and predicts a minimal bactericidal concentration leading to ≥99% bacterial kill (MBC≥99). The CARA helps determine whether a molecule is active on bacilli that are replicating, non-replicating, or both. Pilot experiments using the CARA facilitate the identification of which concentration of test agent and time of compound exposure require further evaluation by colony forming unit (CFU) assays. In addition, the CARA can predict if replicating actives are bactericidal or bacteriostatic.

Comparison of different sampling techniques and of different culture methods for detection of group B streptococcus carriage in pregnant women.

PubMed

El Aila, Nabil A; Tency, Inge; Claeys, Geert; Saerens, Bart; Cools, Piet; Verstraelen, Hans; Temmerman, Marleen; Verhelst, Rita; Vaneechoutte, Mario

2010-09-29

Streptococcus agalactiae (group B streptococcus; GBS) is a significant cause of perinatal and neonatal infections worldwide. To detect GBS colonization in pregnant women, the CDC recommends isolation of the bacterium from vaginal and anorectal swab samples by growth in a selective enrichment medium, such as Lim broth (Todd-Hewitt broth supplemented with selective antibiotics), followed by subculture on sheep blood agar. However, this procedure may require 48 h to complete. We compared different sampling and culture techniques for the detection of GBS. A total of 300 swabs was taken from 100 pregnant women at 35-37 weeks of gestation. For each subject, one rectovaginal, one vaginal and one rectal ESwab were collected. Plating onto Columbia CNA agar (CNA), group B streptococcus differential agar (GBSDA) (Granada Medium) and chromID Strepto B agar (CA), with and without Lim broth enrichment, were compared. The isolates were confirmed as S. agalactiae using the CAMP test on blood agar and by molecular identification with tDNA-PCR or by 16S rRNA gene sequence determination. The overall GBS colonization rate was 22%. GBS positivity for rectovaginal sampling (100%) was significantly higher than detection on the basis of vaginal sampling (50%), but not significantly higher than for rectal sampling (82%). Direct plating of the rectovaginal swab on CNA, GBSDA and CA resulted in detection of 59, 91 and 95% of the carriers, respectively, whereas subculturing of Lim broth yielded 77, 95 and 100% positivity, respectively. Lim broth enrichment enabled the detection of only one additional GBS positive subject. There was no significant difference between GBSDA and CA, whereas both were more sensitive than CNA. Direct culture onto GBSDA or CA (91 and 95%) detected more carriers than Lim broth enrichment and subculture onto CNA (77%). One false negative isolate was observed on GBSDA, and three false positives on CA. In conclusion, rectovaginal sampling increased the number GBS positive women detected, compared to vaginal and/or rectal sampling. Direct plating on CA and/or GBSDA provided rapid detection of GBS that was at least as sensitive and specific as the CDC recommended method of Lim broth subcultured onto non chromogenic agar.

Detection of Salmonella by indicator agar media and PCR as affected by alfalfa seed homogenates and native bacteria.

PubMed

Liao, C-H; Shollenberger, L M

2003-01-01

To investigate and prevent the undesirable effect of native bacteria and alfalfa seed homogenates on detection of Salmonella in alfalfa seeds by indicator agar media and polymerase chain reaction (PCR). The relative sensitivity of five indicator agar media, including modified semisolid RV (MSRV), xylose-lysine-Tergitol 4 (XLT4), Hektoen enteric agar (HEA), brilliant green agar (BGA) and bismuth sulphite agar (BSA), for detection of Salmonella in the presence of a large number of native bacteria from alfalfa seeds was examined. The detection limit as measured by the ratio between the numbers of native bacteria and Salmonella was estimated to be 10(6) to 1 for MSRV and 10(3) to 1 for XLT4, HEA, BGA or BSA. Presence of alfalfa seed homogenates markedly reduced the sensitivity of Salmonella detection by PCR. The minimal number of Salmonella detectable by PCR was determined to be 1-10 and 100-1000 CFU in the absence and presence of seed homogenate, respectively. Application of anti-Salmonella immunomagnetic beads permitted detection of 2-5 CFU of heat-injured cells in 25 g of seeds within 24 h by PCR. The MSRV medium is more sensitive than other indicator agars for detecting a small number of motile Salmonella in samples containing a large number of native bacteria. Application of immunomagnetic beads eliminates the PCR-inhibitory activity of seed homogenates and improves the detection of Salmonella in inoculated seeds. The results generated from this study will aid the seed distributors, sprout growers and public health officials to identify and recall the Salmonella-contaminated seed lots to be used for sprout production.

In Vitro Activities of Gemifloxacin versus Five Quinolones and Two Macrolides against 271 Spanish Isolates of Legionella pneumophila: Influence of Charcoal on Susceptibility Test Results

PubMed Central

García, M. T.; Pelaz, C.; Giménez, M. J.; Aguilar, L.

2000-01-01

The MICs at which 90% of isolates are inhibited for gemifloxacin, trovafloxacin, and grepafloxacin were low (≤0.01 μg/ml) for 271 Legionella isolates when they were determined by the broth microdilution method but increased (≥6 dilutions) when they were determined by the agar dilution method. This was due to the charcoal in the agar dilution medium, as shown by the progressive decrease in the MICs when the charcoal concentrations decreased. As free drug is the active fraction, charcoal binding should be considered. PMID:10898695

Evaluation of methods for the microbiological control of natural corks for sparkling wine bottles.

PubMed

Centeno, S; Calvo, M A

2000-01-01

The various parameters proposed in Norm 0.20/95 of Catalunya (Spain) for the microbiological analysis of natural corks for sparkling wines were evaluated. The best results were obtained through the use of 1/4 Ringer's solution or saline for rinsing with an agitation time of 30 min, and an agitation speed of 150-200 rpm. Tryptone soya agar (TSA) and Sabouraud dextrose agar (SDA) were used as a culture medium for the bacteria and fungi, respectively, and a cultivation time of 48 h and incubation temperatures of 37 +/- 2 degrees C for bacteria and 28 degrees C for yeast and filamentous fungi.

Effect of Sodium Fluorescein and Plating Medium on Recovery of Irradiated Escherichia coli and Serratia marcescens from Aerosols

PubMed Central

Dorsey, Emerson L.; Berendt, Richard F.; Neff, Everett L.

1970-01-01

Irradiation of aerosols of either Escherichia coli or Serratia marcescens with simulated solar (xenon) radiation caused a significant decrease in viability. When sodium fluorescein was employed to determine the physical loss of organisms from the aerosol, an additional adverse effect upon survival was noted. The decay curves indicated that at least two mechanisms of inactivation were operative, one due to aerosolization, the other to irradiation. After collection from aerosols, both species of microorganisms grew better on blood agar base than on Casitone agar, but this finding did not appear to be related to the effect of irradiation. PMID:4922085

Tinea cruris: diagnostic confusion due to isolation of Candida albicans alone.

PubMed Central

Kane, J.; Blakeman, J. M.; Fischer, J. B.

1976-01-01

The diagnostic importance of the isolation of Candida albicans from a skin lesion is often uncertain. In a 68-year-old man from whose lesions only C. albicans was originally isolated Trichophyton rubrum and Epidermophyton floccosum were also isolated when the growth of the yeast was inhibited in a selective medium. The use of this selective medium, casamino acids erythritol albumin agar, ensures the proper interpretation of the significance of the presence of C. albicans in skin lesions. PMID:773524

Axenic cultivation of a pathogenic Phytomonas species isolated from tomato fruit, and from its phytophagic insect vector, Phthia picta (Hemiptera: Coreidae).

PubMed

Fiorini, J E; de Faria e Silva, P M; Brazil, R P; Attias, M; Esteves, M J; Angluster, J

1993-01-01

Axenic cultures of Phytomonas sp. were obtained from naturally infected tomatoes and from Phthia picta, a predator of tomato plants, by using a biphasic medium with Roitman's complex medium overlaying rabbit blood-agar slants. Light and electron microscopy of both isolates showed a similarity of morphological characteristics among the flagellates in fresh material or after cultivation. Other properties, including their agglutinability with the haemolymph of Phthia picta, suggest that these isolates are virtually identical.

[Swarming phenomenon of an aeromonas spec (author's transl)].

PubMed

Müller, H E; Lenz, W

1975-05-01

A genuine swarming phenomenon, such as has previously been known to occur in Proteus, Bacillus and Clostridium species only, was observed in an Aeromonas species. Fig. 1 shows the terraced swarming zones of the Aeromonas species on nutrient agar. The swarming rate, expressed as the growth of the swarming zone per time unit, was measured to be 70-120 mum/min on blood agar at 30 degrees C. The swarming could be inhibited by incubation at 37 degrees C (Table 2), by low saline concentrations (Table 3) as well as by addition of 4-nitro-phenylglycerol to the medium (Table 4). A DIENES-phenomenon between the swarming zones of Proteus strains and that of the Aeromonas species could not be observed (Fig. 2). The manner of swarming as seen in phase contrast microscopy was the same kind as that of Proteus. Furthermore, it could be shown by means of light- and electronmicroscopical investigations that the swarming phenomenon is connected with changes in the cell morphology and the form of flagellation (Figs. 4 and 5). Whereas in broth cultures (Fig. 3) as well as in the centre of colonies on solid media (Fig. 5a) the cells appeared as cocoid rods with polar flagellation, they developed elongated forms at the edge of the swarming zone, which - either in addition to or devoid of the polar flagella - were peritrichously populated with thin, flagella-like filaments (Figs. tb, 6, 7 and 8). The discussion deals with the various forms of bacterial surface translocation and investigates into the role of peritrichous flagella or fimbriae in the swarming phenomenon.

Evaluation of CP Chromo Select Agar for the enumeration of Clostridium perfringens from water.

PubMed

Manafi, Mammad; Waldherr, Kerstin; Kundi, Michael

2013-10-01

The European Directive on drinking water quality has included mCP agar as the reference method for recovering Clostridium perfringens from drinking waters. In the present study, three media (mCP, TSCF and CP Chromo Select Agar) were evaluated for recovery of C. perfringens in different surface water samples. Out of 139 water samples, using a membrane filtration technique, 131 samples (94.2%) were found to be presumptively positive for C. perfringens in at least one of the culture media. Green colored colonies on CP Chromo Select Agar (CCP agar) were counted as presumptive C. perfringens isolates. Out of 483 green colonies on CCP agar, 96.3% (465 strains, indole negative) were identified as C. perfringens, and 15 strains (3.1%) were indole positive and were identified as Clostridium sordellii, Clostridium bifermentans or Clostridium tetani. Only 3 strains (0.6%) gave false positive results and were identified as Clostridium fallax, Clostridium botulinum, and Clostridium tertium. Variance analysis of the data obtained shows statistically no significant differences in the counts obtained between media employed in this work. The mCP method is very onerous for routine screening and bacterial colonies could not be used for further biochemical testing. The colonies on CCP and TSCF were easy to count and subculture for confirmation tests. TSCF detects sulfite-reducing clostridia, including species other than C. perfringens, and in some cases excessive blackening of the agar frustrated counting of the colonies. If the contamination was too high, TSCF did not consistently produce black colonies and as a consequence, the colonies were white and gave false negative results. On the other hand, the identification of typical and atypical colonies isolated from all media demonstrated that CCP agar was the most useful medium for C. perfringens recovery in water samples. Copyright © 2013 Elsevier B.V. All rights reserved.

Evaluation of five selective media for the detection of Pseudomonas aeruginosa using a strain panel from clinical, environmental and industrial sources.

PubMed

Weiser, Rebecca; Donoghue, Denise; Weightman, Andrew; Mahenthiralingam, Eshwar

2014-04-01

Isolation and correct identification of the opportunistic pathogen and industrial contaminant Pseudomonas aeruginosa are very important and numerous selective media are available for this purpose. A novel comparison of five selective media having positive (acetamide-based agars), negative (Pseudomonas CN selective agar [Oxoid Ltd.] and Pseudomonas Isolation agar [Sigma-Aldrich Company Ltd.]) and chromogenic (chromID® P. aeruginosa [bioMérieux]) selection strategies was performed using a systematically designed bacterial test panel (58 P. aeruginosa and 90 non-P. aeruginosa strains including those commonly misidentified as P. aeruginosa by culture-dependent techniques). Standardised inocula were added to the selective media and the results were recorded after 24 and 72h. After 72h of incubation at 37°C chromID® P. aeruginosa displayed the highest specificity (70%) and had good sensitivity (95%), although the sensitivity was negatively impacted by the large variation in colour of P. aeruginosa colonies, which hampered interpretation. Both media containing inhibitory selective agents performed very similarly, both having 100% sensitivity and a specificity of approximately 30%. Raising the incubation temperature to 42°C increased the specificity of Pseudomonas CN selective agar and Pseudomonas isolation agar (61% and 47% respectively after 72h), but increased the number of false positives encountered with the chromogenic medium, decreasing its specificity to 68% after 72h. Growth on the acetamide agars was weak for all strains and it was often difficult to determine whether true growth had occurred. This, compounded by the low specificity of the acetamide agars (<26%), suggested they were less suitable for application to clinical or industrial settings without further modification. Overall, the chromogenic agar was the most selective but further consideration is required to optimise interpretation of results. Copyright © 2014 Elsevier B.V. All rights reserved.

Proliferation and glucosinolates accumulation of broccoli adventitious roots in liquid medium

NASA Astrophysics Data System (ADS)

Nhut, Nguyen Minh; Tien, Le Thi Thuy

2017-09-01

Cotyledons from 7-day-old in vitro broccoli seedling were used as explant source in adventitious root induction on MS medium supplemented with 30 g/l sucrose, 1.6 mg/l IBA and 7 g/l agar. Adventitious roots from cotyledons were transferred to liquid medium containing the same components as rooting medium for two weeks, then subcultured to MS medium with diferent sugar, macrominerals and casein hydrolysate concentrations. The best adventitious root growth was observed in half-strength MS medium supplemented with 40 g/l sucrose, 600 mg/l casein hydrolysate and 1.6 mg/l IBA (growth index of 4.00 in about 14 culture days with inoculum density of 1.0 g fresh weight / 30 ml of culture medium). The culturing process can be stopped on the 28th day for root biomass and on the 35th day for glucosinolates.

Procedures involving lipid media for detection of bacterial contamination in breweries.

PubMed

Van Vuuren, H J; Louw, H A; Loos, M A; Meisel, R

1977-02-01

The liquid equivalent of universal beer agar, designated universal beer liquid medium, and its beer-free equivalent, universal liquid medium (UL), were equally effective in demonstrating bacterial contamination in 120 of 200 samples from different stages of commercial brewing process. Growth of the contaminants after 3 days was consistently more luxuriant in the UL medium. A yeast-water substrate medium failed to reveal many contaminants detected with UL in 392 samples from three breweries and revealed only a few not detected with UL. The use of UL and a lactose-peptone medium, with microscope examination of the media for bacterial growth, permitted detection of 93% of the known contaminants compared to 87%, detected with UL alone; this combination or universal beer liquid medium plus lactose-peptone medium can therefore be recommended for the detection of bacterial contaminants in brewery samples. Bacterial contamination of pitching yeasts appeared to be a particular problem in the breweries investigated.

Effect of genotype, gelling agent, and auxin on the induction of somatic embryogenesis in sweet potato (Ipomoea batatas Lam.).

PubMed

El Abidine Triqui, Zine; Guédira, Abdelkarim; Chlyah, Averil; Chlyah, Hassane; Souvannavong, Vongthip; Haïcour, Robert; Sihachakr, Darasinh

2008-03-01

Lateral buds of six cultivars of sweet potato were induced to form embryogenic callus in a culture medium solidified with two types of gelling agents, Agar or Gelrite, and supplemented with various concentrations of auxins, 2,4-D, 2,4,5-T and Picloram. Of the six cultivars screened, only three gave an embryogenic response. Best results with an average of 3.53% embryogenic response were obtained with the medium solidified with Agar, while in Gelrite only 0.45% of lateral buds gave rise to embryogenic callus. The interaction between the genotype and auxins was highly significant; particularly the optimal response was obtained with cv. Zho and 865 yielding 10.7 and 14.7% somatic embryogenesis, respectively, in the medium containing 2,4,5-T or Picloram. The plant conversion was dramatically improved by subculture of the embryogenic callus on the medium with the combination of 1 microM 2,4-D and 1 microM Kinetin or 5 microM ABA alone before transfer of mature embryos onto hormone-free medium. The embryogenic callus of sweet potato and its sustained ability to further regenerate plants have regularly been maintained for several years by frequent subculture in 5 microM 2,4,5-T or the combination of 10 microM 2,4-D and 1 microM BAP or kinetin. The embryo-derived plants seemed apparently genetically stable and similar to the hexaploid parental plants, based on morphological analysis and their ploidy level determined by using flow cytometry.

In-vitro and in-vivo anti-Trichophyton activity of essential oils by vapour contact.

PubMed

Inouye, S; Uchida, K; Yamaguchi, H

2001-05-01

The minimum inhibitory doses (MIDs) of essential oils by vapour contact to inhibit the growth of Trichophyton mentagrophytes and Trichophyton rubrum on agar medium were determined using airtight boxes. Among seven essential oils examined, cinnamon bark oil showed the least MID, followed by lemongrass, thyme and perilla oils. Lavender and tea tree oils showed moderate MID, and citron oil showed the highest MID, being 320 times higher than that of cinnamon bark oil. The MID values were less than the minimum inhibitory concentration (MIC) values determined by agar dilution assay. Furthermore, the minimum agar concentration (MAC) of essential oils absorbed from vapour was determined at the time of MID determination as the second antifungal measure. The MAC value by vapour contact was 1.4 to 4.7 times less than the MAC remaining in the agar at the time of MIC determination by agar dilution assay. Using selected essential oils, the anti-Trichophyton activity by vapour contact was examined in more detail. Lemongrass, thyme and perilla oils killed the conidia, inhibited germination and hyphal elongation at 1-4 micrograms ml-1 air, whereas lavender oil was effective at 40-160 micrograms ml-1 air. The in-vivo efficacy of thyme and perilla oils by vapour contact was shown against an experimental tinea pedis in guinea pigs infected with T. mentagrophytes. These results indicated potent anti-Trichophyton action of essential oils by vapour contact.

Surprisal analysis of genome-wide transcript profiling identifies differentially expressed genes and pathways associated with four growth conditions in the microalga Chlamydomonas.

PubMed

Bogaert, Kenny A; Manoharan-Basil, Sheeba S; Perez, Emilie; Levine, Raphael D; Remacle, Francoise; Remacle, Claire

2018-01-01

The usual cultivation mode of the green microalga Chlamydomonas is liquid medium and light. However, the microalga can also be grown on agar plates and in darkness. Our aim is to analyze and compare gene expression of cells cultivated in these different conditions. For that purpose, RNA-seq data are obtained from Chlamydomonas samples of two different labs grown in four environmental conditions (agar@light, agar@dark, liquid@light, liquid@dark). The RNA seq data are analyzed by surprisal analysis, which allows the simultaneous meta-analysis of all the samples. First we identify a balance state, which defines a state where the expression levels are similar in all the samples irrespectively of their growth conditions, or lab origin. In addition our analysis identifies additional constraints needed to quantify the deviation with respect to the balance state. The first constraint differentiates the agar samples versus the liquid ones; the second constraint the dark samples versus the light ones. The two constraints are almost of equal importance. Pathways involved in stress responses are found in the agar phenotype while the liquid phenotype comprises ATP and NADH production pathways. Remodeling of membrane is suggested in the dark phenotype while photosynthetic pathways characterize the light phenotype. The same trends are also present when performing purely statistical analysis such as K-means clustering and differentially expressed genes.

Oxytetracycline-Resistant Coliforms in Commercial Poultry Products

PubMed Central

Corey, R. Reece; Byrnes, Joseph M.

1963-01-01

The presence of oxytetracycline-resistant bacteria was investigated with commercially frozen chicken thighs and drumsticks. Bacterial flora were surveyed by means of total and coliform counts with Tryptone Glucose Extract Agar and Desoxycholate Agar, respectively. After counting, the Desoxycholate Agar plates were replicated on the same medium containing 25, 50, 75, and 100 ppm of oxytetracycline. Resistant colonies were found on all samples that were replicated. Of 2613 colonies isolated on Desoxycholate Agar, 47.8% grew in the presence of 25 ppm of oxytetracycline. From 50 to 100 ppm, the number of resistant isolates remained essentially the same, near 34%. Of 812 colonies of antibiotic-resistant bacteria identified with dulcitol-lactose-iron-agar, 82.5% were paracolons, 13.7% were pseudomonads, and 3.8% were Escherichia or Aerobacter. Bacteria resistant to oxytetracycline were shown to be present on commercially processed chicken. The origin of the resistance to oxytetracycline was not established; however, since the antibiotic was not used during processing, it appeared that these antibiotic-resistant bacteria arose in the intestines of the chickens as a result of feed which contained antibiotic. This is supported by a comparison with the antibiotic resistance of coliforms from chickens raised on feed both with and without oxytetracycline, for the percentages of resistant colonies are similar in both commercial chicken and chicken raised on feed containing the antibiotic. PMID:14075046

Use of agar agar stabilized milled zero-valent iron particles for in situ groundwater remediation

NASA Astrophysics Data System (ADS)

Schmid, Doris; Velimirović, Milica; Wagner, Stephan; Micić Batka, Vesna; von der Kammer, Frank; Hofmann, Thilo

2015-04-01

A major obstacle for use of nanoscale zero-valent iron (nZVI) particles as a nontoxic material for effective in situ degradation of chlorinated aliphatic hydrocarbons (CAHs) is the high production cost. For that reason, submicro-scale milled zero-valent iron particles were recently developed (milled ZVI, UVR-FIA, Germany) by grinding macroscopic raw materials of elementary iron as a cheaper alternative to products produced by solid-state reduction. However, milled ZVI particles tend to aggregate and due to the rather large particle size (d50= 11.9 µm) also rapidly sediment. To prevent aggregation and consequently sedimentation of milled ZVI particles and therefore improve the mobility after in situ application, the use of a stabilizer is considered in literature as a most promising option. In this study, milled ZVI particles (1 g L-1 of particle concentration) were stabilized by environmentally friendly polymer agar agar (>0.5 g L-1), which had a positive impact on the milled ZVI stability. Sedimentation rate was significantly decreased by increasing the suspension viscosity. Column transport experiments were performed for bare and agar agar stabilized milled ZVI particles in commercially available fine grained quartz sand (DORSILIT® Nr.8, Gebrüder Dorfner GmbH Co, Germany) and different porous media collected from brownfields. The experiments were carried out under field relevant injection conditions of 100 m d-1. The maximal travel distance (LT) of less than 10 cm was determined for non-stabilized suspension in fine grained quartz sand, while agar agar (1 g L-1) stabilized milled ZVI suspension revealed LT of 12 m. Similar results were observed for porous media from brownfields showing that mobility of agar agar stabilized particle suspensions was significantly improved compared to bare particles. Based on the mobility data, agar agar stabilized milled zero-valent iron particles could be used for in situ application. Finally, lab-scale batch degradation experiments were performed to determine the impact of agar agar on the reactivity of milled ZVI and investigate the apparent corrosion rate of particles by quantifying the hydrogen gas generated by anaerobic corrosion of milled ZVI. The results indicate that agar agar had a positive impact on the milled ZVI stability and mobility, however adverse impact on the reactivity towards trichloroethene (TCE) was observed compared to the non-stabilized material. On the other hand, this study shows that the apparent corrosion rate of non-stabilized and agar agar stabilized milled ZVI particles is in the same order of magnitude. These data indicate that the dechlorination pathway of TCE by agar agar stabilized milled ZVI particles is possibly impacted by blocking of the reactive sites and not hydrogen revealed during particles corrosion. Finally, calculated longevity of the particles based on the apparent corrosion rate is significantly prolonged compared to the longevity of the nZVI particles reported in previous studies. This research receives funding from the European Union's Seventh Framework Programme FP7/2007-2013 under grant agreement n°309517.

Color features as an approach for the automated screening of Salmonella strain

NASA Astrophysics Data System (ADS)

Trujillo, Alejandra Serrano; González, Viridiana Contreras; Andrade Rincón, Saulo E.; Palafox, Luis E.

2016-11-01

We present the implementation of a feature extraction approach for the automated screening of Salmonella sp., a task visually carried out by a microbiologist, where the resulting color characteristics of the culture media plate indicate the presence of this strain. The screening of Salmonella sp. is based on the inoculation and incubation of a sample on an agar plate, allowing the isolation of this strain, if present. This process uses three media: Xylose lysine deoxycholate, Salmonella Shigella, and Brilliant Green agar plates, which exhibit specific color characteristics over the colonies and over the surrounding medium for a presumed positive interpretation. Under a controlled illumination environment, images of plates are captured and the characteristics found over each agar are processed separately. Each agar is analyzed using statistical descriptors for texture, to determine the presence of colonies, followed by the extraction of color features. A comparison among the color features seen over the three media, according to the FDA Bacteriological Analytical Manual, determines the presence of Salmonella sp. on a given sample. The implemented process proves that the task addressed can be accomplished under an image processing approach, leading to the future validation and automation of additional screening processes.

Isolation of amylolytic, xylanolytic, and cellulolytic microorganisms extracted from the gut of the termite Reticulitermes santonensis by means of a micro-aerobic atmosphere.

PubMed

Tarayre, Cédric; Brognaux, Alison; Bauwens, Julien; Brasseur, Catherine; Mattéotti, Christel; Millet, Catherine; Destain, Jacqueline; Vandenbol, Micheline; Portetelle, Daniel; De Pauw, Edwin; Eric, Haubruge; Francis, Frédéric; Thonart, Philippe

2014-05-01

The aim of this work was to isolate enzyme-producing microorganisms from the tract of the termite Reticulitermes santonensis. The microorganisms were extracted from the guts and anaerobic (CO₂ or CO₂/H₂) and micro-aerobic atmospheres were used to stimulate growth. Three different strategies were tried out. First, the sample was spread on Petri dishes containing solid media with carboxymethylcellulose, microcrystalline cellulose or cellobiose. This technique allowed us to isolate two bacteria: Streptomyces sp. strain ABGxAviA1 and Pseudomonas sp. strain ABGxCellA. The second strategy consisted in inoculating a specific liquid medium containing carboxymethylcellulose, microcrystalline cellulose, or cellobiose. The samples were then spread on Petri dishes with the same specific medium containing carboxymethylcellulose, microcrystalline cellulose, or cellobiose. This led to the isolation of the mold Aspergillus sp. strain ABGxAviA2. Finally, the third strategy consisted in heating the first culture and spreading samples on agar plates containing rich medium. This led to the isolation of the bacterium Bacillus subtilis strain ABGx. All those steps were achieved in controlled atmospheres. The four enzyme-producing strains which were isolated were obtained by using a micro-aerobic atmosphere. Later, enzymatic assays were performed on the four strains. Streptomyces sp. strain ABGxAviA1 was found to produce only amylase, while Pseudomonas sp. strain ABGxCellA was found to produce β-glucosidase as well. Aspergillus sp. strain ABGxAviA2 showed β-glucosidase, amylase, cellulase, and xylanase activities. Finally, B. subtilis strain ABGx produced xylanase and amylase.

An improved embryo-rescue protocol for hybrid progeny from seedless Vitis vinifera grapes × wild Chinese Vitis species.

PubMed

Li, Gui Rong; Ji, Wei; Wang, Gang; Zhang, Jian Xia; Wang, Yue Jin

A highly efficient technique of embryo rescue is critical when using stenospermocarpic Vitis vinifera cultivars (female parents) to breed novel, disease-resistant, seedless grape cultivars by hybridizing with wild Chinese Vitis species (male parents) having many disease-resistance alleles. The effects of various factors on the improvement of embryo formation, germination, and plantlet development for seven hybrid combinations were studied. The results indicated that Beichun and Shuangyou were the best male parents. The best sampling time for ovule inoculation differed among the female parents. When hybrid ovules were cultured on a double-phase medium with five different solid medium types, percent embryo formation was highest (11.3-28.3%) on a modified MM3 medium. Percentages of embryo germination (15.4-55.4%) and plantlet development (11.15-44.6%) were all highest when embryos were cultured on Woody Plant Medium + 5.7 μM indole-3-acetic acid + 4.4 μM 6-benzylaminopurine + 1.4 μM gibberellic acid + 2% sucrose + 0.05% casein hydrolysate + 0.3% activated charcoal + 0.7% agar. In the absence of other amino acids, the addition of proline significantly increased embryo formation (36.1%), embryo germination (64.6%), and plantlet development (90.5%). A highly efficient protocol has been developed for hybrid embryo rescue from seedless V. vinifera grapes × wild Chinese Vitis species that results in a significant improvement in breeding efficiency for new disease-resistant seedless grapes.

Efficacy of the thin agar layer method for the recovery of stressed Cronobacter spp. (Enterobacter sakazakii).

PubMed

Osaili, Tareq M; Al-Nabulsi, Anas A; Shaker, Reyad R; Al-Holy, Murad M; Al-Haddaq, Mohammed S; Olaimat, Amin N; Ayyash, Mutamed M; Al Ta'ani, Mahmoud K; Forsythe, Stephen J

2010-10-01

Cronobacter spp. (Enterobacter sakazakii) are emerging opportunistic pathogens for all age groups, and are of particular concern when it comes to infants. Prior to contaminating food, the organism may be exposed to a variety of stresses, leading to a generation of sublethally injured cells that may not be detected by selective media unless a protracted recovery period is included in the isolation procedure. This study evaluated the efficacy of the thin agar layer (TAL) method for the recovery of Cronobacter cells that had been exposed to various stress conditions. Five strains of C. sakazakii and C. muytjensii were exposed to starvation, heat, cold, acid, alkaline, chlorine, or ethanol, with or without further exposure to desiccation stress. The recovery of the stressed cells was determined on tryptone soy agar (TSA; nonselective control medium), violet red bile glucose agar (VRBGA; selective agar), Druggan-Forsythe-Iversen (DFI; selective agar), and TAL media (viz., VRBGA overlaid with TSA, and DFI overlaid with TSA). Regardless of stress type, there were no significant differences among the recoveries of stressed desiccated Cronobacter spp. cultures on TSA, DFI+TSA, and VRBGA+TSA, but there was significantly less recovery on VRBGA. The recovery of prestressed desiccated Cronobacter spp. on DFI+TSA was similar to that on TSA, whereas the recovery on VRBGA+TSA was lower. DFI+TSA performed better than VRBGA+TSA did in differentiating Cronobacter spp. within mixed bacterial cultures. The results of this study suggest the use of the TAL method DFI+TSA as an improved method for the direct recovery of stressed Cronobacter spp.

Use of Dehydrated Agar to Estimate Microbial Water Quality for Horticulture Irrigation.

PubMed

Meador, Dustin P; Fisher, Paul R; Guy, Charles L; Harmon, Philip F; Peres, Natalia A; Teplitski, Max

2016-07-01

Petrifilms are dehydrated agar culture plates that have been used to quantify colony forming units (CFU) mL of either aerobic bacteria (Petrifilm-AC) or fungus (Petrifilm-YM), depending on substrate composition. Microbes in irrigation systems can indicate biofilm risk and potential clogging of irrigation emitters. The research objective was to compare counts on Petrifilms versus traditional, hydrated-agar plates using samples collected from recirculated irrigation waters and cultures of isolated known species. The estimated count (in CFU mL) from a recirculated irrigation sample after 7 d of incubation on Petrifilm-YM was only 5.5% of the count quantified using sabouraud dextrose agar (SDA) with chloramphenicol after 14 d. In a separate experiment with a known species, Petrifilm-YM did not successfully culture zoospores of . Isolates of viable zoospores were cultured successfully on potato-dextrose agar (PDA), with comparable counts with a vegetable juice medium supplemented with the antibiotics pimaricin, ampicillin, rifamycin, pentochloronitrobenzene and hymexazol (PARP-H). The quantification of pv. Begoniaceae on Petrifilm-AC was not significantly different ( < 0.05) than on PDA, but was lower than on Reasoner and Goldrich agar (R2A) or with a hemocytometer. The current formulation of Petrifilm-YM is unlikely to be a useful monitoring method for plant pathogens in irrigation water because of the inability to successfully culture oomycetes. However, Petrifilm-AC was an effective method to quantify bacteria and can provide an easy-to-use on-farm tool to monitor biofilm risk and microbial density. Copyright © by the American Society of Agronomy, Crop Science Society of America, and Soil Science Society of America, Inc.

[Dynamics of genome changes in Rauwolfia serpentina callus tissue upon the switch to conditions of submerged cultivation].

PubMed

Spiridonova, E V; Adnof, D M; Andreev, I O; Kunakh, V A

2008-01-01

Genome of Rauwolfia serpentina callus cells was found to fail undergo the noticeable changes for several early passages upon the switch from surface to submerged cultivation in the liquid medium of special composition. After subsequent 4-6 passages in submerged culture RAPD spectra polymorphism was revealed which may reflect the changes in DNA sequence as well as in the structure of cell population that forms the strain. Introduction of the intermediary passage on the agar-solidified medium of more simple composition prior to transfer into liquid medium appeared not to affect essentially the level and the pattern of genome changes.

Biochemical Characterization of Extracellular Cellulase from Tuber maculatum Mycelium Produced Under Submerged Fermentation.

PubMed

Bedade, Dattatray K; Singhal, Rekha S; Turunen, Ossi; Deska, Jan; Shamekh, Salem

2017-02-01

Interaction of truffle mycelium with the host plant involves the excretion of extracellular enzymes. The ability of Tuber maculatum mycelium to produce an extracellular cellulase during submerged fermentation was demonstrated for the first time. T. maculatum mycelia were isolated and tested for extracellular cellulase production at variable pH on solid agar medium, and the highest activity was observed at pH 7.0. Furthermore, T. maculatum was subjected to submerged fermentation in basal salt medium for cellulase production. Under optimized conditions using sodium carboxymethyl cellulose (0.5 % w/v) as carbon source and an initial pH of 7.0, the enzyme production yielded 1.70 U/mL of cellulase in the cell-free supernatant after 7 days of incubation time. The optimum of the obtained cellulase's activity was at pH 5.0 and a temperature of 50 °C. The enzyme showed good thermostability at 50 °C by retaining 99 % of its maximal activity over an incubation time of 100 min. The cellulase activity was inhibited by Fe 2+ and slightly activated by Mn 2+ and Cu 2+ at 1 mM concentration. The results indicated that truffle mycelium is utilizing cellulosic energy source in the root system, and the optimal conditions are those existing in the acidic Finnish soil.

Relationship between pH and Medium Dissolved Solids in Terms of Growth and Metabolism of Lactobacilli and Saccharomyces cerevisiae during Ethanol Production

PubMed Central

Narendranath, Neelakantam V.; Power, Ronan

2005-01-01

The specific growth rates of four species of lactobacilli decreased linearly with increases in the concentration of dissolved solids (sugars) in liquid growth medium. This was most likely due to the osmotic stress exerted by the sugars on the bacteria. The reduction in growth rates corresponded to decreased lactic acid production. Medium pH was another factor studied. As the medium pH decreased from 5.5 to 4.0, there was a reduction in the specific growth rate of lactobacilli and a corresponding decrease in the lactic acid produced. In contrast, medium pH did not have any significant effect on the specific growth rate of yeast at any particular concentration of dissolved solids in the medium. However, medium pH had a significant (P < 0.001) effect on ethanol production. A medium pH of 5.5 resulted in maximal ethanol production in all media with different concentrations of dissolved solids. When the data were analyzed as a 4 (pH levels) by 4 (concentrations of dissolved solids) factorial experiment, there was no synergistic effect (P > 0.2923) observed between pH of the medium and concentration of dissolved solids of the medium in reducing bacterial growth and metabolism. The data suggest that reduction of initial medium pH to 4.0 for the control of lactobacilli during ethanol production is not a good practice as there is a reduction (P < 0.001) in the ethanol produced by the yeast at pH 4.0. Setting the mash (medium) with ≥30% (wt/vol) dissolved solids at a pH of 5.0 to 5.5 will minimize the effects of bacterial contamination and maximize ethanol production by yeast. PMID:15870306

Evaluation of different conditions and culture media for the recovery of Aeromonas spp. from water and shellfish samples.

PubMed

Latif-Eugenín, F; Beaz-Hidalgo, R; Figueras, M J

2016-09-01

To perform a comparative study for determining the optimum culture method (direct plating or enrichment) and medium (ampicillin dextrin agar (ADA), starch ampicillin agar (SAA), bile salts irgasan brilliant green modified (BIBG-m)) for recovering Aeromonas species from water and shellfish samples. By direct culture, Aeromonas was detected in 65% (13/20) of the water samples and in 54·5% (6/11) of the shellfish samples. However, when a pre-enrichment step was included, the number of positive water samples increased to 75% (15/20) and the ones of shellfish to 90·1% (10/11). The enriched culture significantly favoured (P < 0·05) the isolation of Aeromonas allosaccharophila from water, Aeromonas salmonicida from shellfish and Aeromonas caviae from both types of samples. The most specific (P < 0·05) culture medium for detecting Aeromonas from water was ADA. However, no differences were observed in the case of shellfish samples (P > 0·05). Isolation of Aeromonas media from water was favoured (P < 0·05) in the ADA medium, while SAA enhanced (P < 0·05) the isolation of Aer. salmonicida from shellfish. The culture method and medium used influenced the recovery of some Aeromonas species from water and shellfish samples. This fact should be considered in future prevalence studies to avoid overestimating the above mentioned Aeromonas species. © 2016 The Society for Applied Microbiology.

Rapid biochemical screening for Salmonella, Shigella, Yersinia, and Aeromonas isolates from stool specimens.

PubMed Central

De Ryck, R; Struelens, M J; Serruys, E

1994-01-01

Four screens for the rapid (4 to 6 h) biochemical detection of pathogens from enteric isolation media are described. The Salmonella screen consisted of Kligler iron agar (KIA), motility-indole-urea-tryptophan-deamination semisolid medium (MIU-TDA), and the o-nitrophenyl-beta-D-galactopyranoside (ONPG) test; the Shigella screen consisted of KIA, MIU-TDA, the ONPG test, and the lysine decarboxylation-indole test; the Yersinia screen consisted of a rhamnose broth; the Aeromonas screen consisted of a xylose agar plate. When tested on 2,102 fresh isolates and 71 stock strains, the screens correctly detected 212 enteric pathogens (sensitivity, 100%), with a specificity of 98.1%. PMID:8077408

Evaluation of BBL CHROMagar VanRE for detection of vancomycin-resistant Enterococci in rectal swab specimens.

PubMed

Stamper, Paul D; Shulder, Stephanie; Bekalo, Pearl; Manandhar, Deepika; Ross, Tracy L; Speser, Sharon; Kingery, Julie; Carroll, Karen C

2010-11-01

A study was performed on 517 surveillance rectal swabs to evaluate a selective and differential chromogenic medium, the BBL CHROMagar VanRE (CVRE), which enables recovery and identification of VanA- and VanB-containing Enterococcus faecium (ENFM) and Enterococcus faecalis (ENFS) isolates. Compared to BBL Enterococcosel agar, a bile-esculin-azide-vancomycin (BEAV) agar, the initial overall sensitivity, specificity, and positive and negative predictive values of CVRE for the detection of vancomycin-resistant ENFM and ENFS were 99.1% and 94.8% and 84.2% and 99.7%, respectively. Among our patient population, more vancomycin-resistant enterococci (VRE) were recovered with CVRE than BEAV.

Evaluation of BBL CHROMagar VanRE for Detection of Vancomycin-Resistant Enterococci in Rectal Swab Specimens▿

PubMed Central

Stamper, Paul D.; Shulder, Stephanie; Bekalo, Pearl; Manandhar, Deepika; Ross, Tracy L.; Speser, Sharon; Kingery, Julie; Carroll, Karen C.

2010-01-01

A study was performed on 517 surveillance rectal swabs to evaluate a selective and differential chromogenic medium, the BBL CHROMagar VanRE (CVRE), which enables recovery and identification of VanA- and VanB-containing Enterococcus faecium (ENFM) and Enterococcus faecalis (ENFS) isolates. Compared to BBL Enterococcosel agar, a bile-esculin-azide-vancomycin (BEAV) agar, the initial overall sensitivity, specificity, and positive and negative predictive values of CVRE for the detection of vancomycin-resistant ENFM and ENFS were 99.1% and 94.8% and 84.2% and 99.7%, respectively. Among our patient population, more vancomycin-resistant enterococci (VRE) were recovered with CVRE than BEAV. PMID:20739492

Optimal conditions of mycelia growth of Laetiporus sulphureus sensu lato

PubMed Central

Luangharn, Thatsanee; Karunarathna, Samantha C.; Hyde, Kevin D.; Chukeatirote, Ekachai

2014-01-01

Laetiporus sulphureus is an edible wood-rotting basidiomycete, growing on decaying logs, stumps, and trunks of many deciduous and coniferous tree species. This fungus produces relatively large striking yellowish or orange-coloured bracket-like fruitbodies. L. sulphureus is widely consumed as a nutritional food because of its fragrance and texture. In this study, two L. sulphureus strains, MFLUCC 12-0546 and MFLUCC 12-0547, isolated from Chiang Rai, Thailand, were investigated for optimal conditions of mycelia growth. Potato dextrose agar and malt extract agar were observed as the favourable medium for mycelia growth. The optimum pH and temperature for the mushroom mycelia were 6–8 and 25–30°C, respectively. PMID:25544934

Probiotic culture survival and implications in fermented frozen yogurt characteristics.

PubMed

Davidson, R H; Duncan, S E; Hackney, C R; Eigel, W N; Boling, J W

2000-04-01

Low-fat ice cream mix was fermented with probiotic-supplemented and traditional starter culture systems and evaluated for culture survival, composition, and sensory characteristics of frozen product. Fermentations were stopped when the titratable acidity reached 0.15% greater than the initial titratable acidity (end point 1) or when the pH reached 5.6 (end point 2). Mix was frozen and stored for 11 wk at -20 degrees C. The traditional yogurt culture system contained the strains Streptococcus salivarius ssp. thermophilus and Lactobacillus delbrueckii ssp. bulgaricus. The probiotic-supplemented system contained the traditional cultures as well as Bifidobacterium longum and Lactobacillus acidophilus. We compared recovery of Bifodobacterium by three methods, a repair-detection system with roll-tubes and plates on modified bifid glucose medium and plates with maltose + galactose reinforced clostridial medium. Culture bacteria in both systems did not decrease in the yogurt during frozen storage. The roll-tube method with modified bifid glucose agar and repair detection system provided at least one-half log10 cfu/ml higher recovery of B. longum compared with recoveries using modified bifid glucose agar or maltose + galactose reinforced clostridial agar on petri plates. No change in concentrations of lactose or protein for products fermented with either culture system occurred during storage. Acid flavor was more intense when product was fermented to pH 5.6, but yogurt flavor was not intensified. The presence of probiotic bacteria in the supplemented system seemed to cause no differences in protein and lactose concentration and sensory characteristics.

Microbial contamination of soft contact lenses & accessories in asymptomatic contact lens users.

PubMed

Thakur, Deeksha V; Gaikwad, Ujjwala N

2014-08-01

With increasing use of soft contact lenses the incidence of contact lens induced infections is also increasing. This study was aimed to assess the knowledge of new and existing contact lens users about the risk of microbial contamination associated with improper use and maintenance of contact lenses, type of microbial flora involved and their potential to cause ophthalmic infections. Four samples each from 50 participants (n=200) were collected from the lenses, lens care solutions, lens care solution bottles and lens cases along with a questionnaire regarding their lens use. The samples were inoculated onto sheep blood agar, Mac Conkey's agar and Sabouraud's dextrose agar. Organisms were identified using standard laboratory protocols. Overall rate of microbial contamination among the total samples was 52 per cent. The most and the least contaminated samples were found to be lens cases (62%) and lens care solution (42%), respectively. The most frequently isolated contaminant was Staphylococcus aureus (21%) followed by Pseudomonas species (19.5%). Majority (64%) of the participants showed medium grade of compliance to lens cleaning practices. Rate of contamination was 100 and 93.75 per cent respectively in those participants who showed low and medium compliance to lens care practices as compared to those who had high level of compliance (43.75%) (P<0.05). Lens care practices amongst the participants were not optimum which resulted into high level contamination. Hence, creating awareness among the users about the lens care practices and regular cleaning and replacements of lens cases are required.

Evaluation of Five Chromogenic Agar Media and the Rosco Rapid Carb Screen Kit for Detection and Confirmation of Carbapenemase Production in Gram-Negative Bacilli

PubMed Central

Gilmour, Matthew W.; DeGagne, Pat; Nichol, Kim; Karlowsky, James A.

2014-01-01

An efficient workflow to screen for and confirm the presence of carbapenemase-producing Gram-negative bacilli was developed by evaluating five chromogenic screening agar media and two confirmatory assays, the Rapid Carb screen test (Rosco Diagnostica A/S, Taastrup, Denmark) and the modified Hodge test. A panel of 150 isolates was used, including 49 carbapenemase-producing isolates representing a variety of β-lactamase enzyme classes. An evaluation of analytical performance, assay cost, and turnaround time indicated that the preferred workflow (screening test followed by confirmatory testing) was the chromID Carba agar medium (bioMérieux, Marcy l'Étoile, France), followed by the Rapid Carb screen test, yielding a combined sensitivity of 89.8% and a specificity of 100%. As an optional component of the workflow, a determination of carbapenemase gene class via molecular means could be performed subsequent to confirmatory testing. PMID:25355764

A new selective enrichment procedure for isolating Pasteurella multocida from avian and environmental samples

USGS Publications Warehouse

Moore, M.K.; Cicnjak-Chubbs, L.; Gates, R.J.

1994-01-01

A selective enrichment procedure, using two new selective media, was developed to isolate Pasteurella multocida from wild birds and environmental samples. These media were developed by testing 15 selective agents with six isolates of P. multocida from wild avian origin and seven other bacteria representing genera frequently found in environmental and avian samples. The resulting media—Pasteurella multocida selective enrichment broth and Pasteurella multocida selective agar—consisted of a blood agar medium at pH 10 containing gentamicin, potassium tellurite, and amphotericin B. Media were tested to determine: 1) selectivity when attempting isolation from pond water and avian carcasses, 2) sensitivity for detection of low numbers of P. multocida from pure and mixed cultures, 3) host range specificity of the media, and 4) performance compared with standard blood agar. With the new selective enrichment procedure, P. multocida was isolated from inoculated (60 organisms/ml) pond water 84% of the time, whereas when standard blood agar was used, the recovery rate was 0%.

Chemically defined medium for cultivation of several epiphytic and phytopathogenic spiroplasmas.

PubMed

Lee, I M; Davis, R E

1983-12-01

A chemically defined medium, LD82, was formulated for in vitro cultivation of spiroplasmas. Medium LD82 supported good growth for four epiphytic and insect-pathogenic spiroplasmas, Spiroplasma floricola 23-6, Spiroplasma sp. strain SR3, Spiroplasma sp. strain brevi, and Spiroplasma sp. strain AS576, and of the phytopathogenic spiroplasmas Spiroplasma citri Maroc R8A2 and PC1. Titers of all six strains grown in defined medium LD82 reached 2.0 x 10 to 6.0 x 10 CFU/ml of culture. All spiroplasma strains tested formed colonies readily on agar medium LD82. None of the spiroplasmas formed typical fried-egg colonies. All formed diffuse colonies, but the forms of colonies differed somewhat among the spiroplasma strains. In preliminary studies of nutritional requirements, phospholipids slightly enhanced the growth of the epiphytic and insect-pathogenic strains in medium LD82 and were found essential for good growth of S. citri.

The visual assessment of broth cultures for tissue bank samples.

PubMed

Varettas, Kerry

2017-09-01

The bioburden screening process of allograft musculoskeletal tissue samples received at the South Eastern Area Laboratory Services includes the routine use of solid agar and cooked meat (CM) broth media. CM has been routinely sub-cultured onto solid agar plates after aerobic incubation at 35 °C. This study will evaluate whether a visual assessment of CM can replace sub-culture by an in vitro inoculation and a prospective study. Eight challenge organisms were serially diluted and inoculated into CM. The average inoculum of 0.5-5.5 CFU produced visible turbidity of CM after 24-h incubation for 7 of the challenge organisms with one organism producing turbidity after 48-h incubation. The prospective study evaluated 222 CM of which 213 were visually clear and no-growth on sub-culture and 9 turbid CM which were culture positive. Broth cultures are an integral part of the bioburden screening process of allograft musculoskeletal tissue and swab samples and visual assessment of CM can replace sub-culture.

Conventional and microwave pyrolysis of a macroalgae waste from the Agar-Agar industry. Prospects for bio-fuel production.

PubMed

Ferrera-Lorenzo, N; Fuente, E; Bermúdez, J M; Suárez-Ruiz, I; Ruiz, B

2014-01-01

A comparative study of the pyrolysis of a macroalgae industrial solid waste (algae meal) in an electrical conventional furnace and in a microwave furnace has been carried out. It was found that the chars obtained from both pyrolyses are similar and show good properties for performing as a solid bio-fuel and as a precursor of activated carbon. Bio-oils from conventional pyrolysis have a greater number of phenolic, pyrrole and alkane compounds whereas benzene and pyridine compounds are more predominant in microwave pyrolysis with a major presence of light compounds. The bio-gas fraction from microwave pyrolysis presents a much higher syngas content (H2+CO), and a lower CO2 and CH4 proportion than that obtained by conventional pyrolysis. Yields are similar for both treatments with a slightly higher gas yield in the case of microwave pyrolysis due to the fact that microwave heating favors heterogeneous reactions between the gases and the char. Copyright © 2013 Elsevier Ltd. All rights reserved.

Natural Competence of Xylella fastidiosa Occurs at a High Frequency Inside Microfluidic Chambers Mimicking the Bacterium's Natural Habitats.

PubMed

Kandel, Prem P; Lopez, Samantha M; Almeida, Rodrigo P P; De La Fuente, Leonardo

2016-09-01

Xylella fastidiosa is a xylem-limited bacterium that is the causal agent of emerging diseases in a number of economically important crops. Genetic diversity studies have demonstrated homologous recombination occurring among X. fastidiosa strains, which has been proposed to contribute to host plant shifts. Moreover, experimental evidence confirmed that X. fastidiosa is naturally competent for recombination in vitro Here, as an approximation of natural habitats (plant xylem vessels and insect mouthparts), recombination was studied in microfluidic chambers (MCs) filled with media amended with grapevine xylem sap. First, different media were screened for recombination in solid agar plates using a pair of X. fastidiosa strains that were previously reported to recombine in coculture. The highest frequency of recombination was obtained with PD3 medium, compared to those with the other two media (X. fastidiosa medium [XFM] and periwinkle wilt [PW] medium) used in previous studies. Dissection of the media components led to the identification of bovine serum albumin as an inhibitor of recombination that was correlated to its previously known effect on inhibition of twitching motility. When recombination was performed in liquid culture, the frequencies were significantly higher under flow conditions (MCs) than under batch conditions (test tubes). The recombination frequencies in MCs and agar plates were not significantly different from each other. Grapevine xylem sap from both susceptible and tolerant varieties allowed high recombination frequency in MCs when mixed with PD3. These results suggest that X. fastidiosa has the ability to be naturally competent in the natural growth environment of liquid flow, and this phenomenon could have implications in X. fastidiosa environmental adaptation. Xylella fastidiosa is a plant pathogen that lives inside xylem vessels (where water and nutrients are transported inside the plant) and the mouthparts of insect vectors. This bacterium causes emerging diseases in various crops worldwide, including recent outbreaks in Europe. The mechanisms by which this bacterium adapts to new hosts is not understood, but it was previously shown that it is naturally competent, meaning that it can take up DNA from the environment and incorporate it into its genome (recombination). In this study, we show that the frequency of recombination is highest when the bacterium is grown under flow conditions in microfluidic chambers modeled after its natural habitats, and recombination was still high when the medium was amended with grapevine sap. Our results suggest that this bacterium is able to recombine when growing inside plants or insects, and this can be a mechanism of adaptation of this pathogen that causes incurable diseases. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Distribution and survival of Pseudomonas sp. on Italian ryegrass and Curly dock in Georgia

USDA-ARS?s Scientific Manuscript database

Yellow bud, caused by Pseudomonas sp. is an emerging bacterial disease of onion. Polymerase chain reaction (PCR) assay based on the coronafacate ligase (cfl) and HrpZ genes were used to detect initial suspected bacteria on weeds. Growth on an agar medium, ability to cause a hypersensitive response i...

Effect of Carbon Dioxide on Testing of Susceptibilities of Respiratory Tract Pathogens to Macrolide and Azalide Antimicrobial Agents

PubMed Central

Johnson, M. M.; Hill, S. L.; Piddock, Laura J. V.

1999-01-01

The in vitro activities of erythromycin, azithromycin, and clarithromycin against 178 clinical isolates from the lower respiratory tract of patients with chronic obstructive pulmonary disease were determined by an agar dilution method. The plates were incubated in air alone or in 5% carbon dioxide. The MICs measured in air alone were lower for most isolates than those measured in 5% carbon dioxide, illustrating the “pH effect” of incubation in carbon dioxide. Testing of isolates in 5% carbon dioxide on pH-adjusted medium (pH 8.4) resulted in MICs of one or two doubling dilutions lower than those obtained on agar with a neutral pH. A bioassay of the three agents incubated in air and in 5% carbon dioxide resulted in a significant loss of activity of all three agents in the carbon dioxide-enriched atmosphere. However, this loss-of-activity effect was significantly reduced when the bioassay medium was adjusted to pH 8.4 prior to incubation in 5% carbon dioxide. PMID:10428903

Quantitative assessment of mycological air pollution in selected rooms of residential and dormitory housing facilities.

PubMed

Lonc, Elzbieta; Plewa, Kinga; Kiewra, Dorota; Szczepańska, Anna; Firling, Conrad E

2013-01-01

The qualitative and quantitative mycological composition of indoor areas of three private residencies and an academic dormitory in Wroclaw, Poland was investigated. Seasonal fungal samples were obtained using a MAS-100 air sampler. The samples were cultured on three different media: Sabouraud Agar (SAB), Dichloran Glycerol Selective Medium (DG18) and Malt Extract Agar (MEA). The number of colony forming unit (CFU) values ranged from 10 CFU/m3 to 490 CFU/m3 depending on the culture medium, season, and sampling site. The identification of the cultured fungi was performed using macro- and microscopic observations and diagnostic keys. Eleven fungal genera were identified. The most common fungi were members of genera Cladosporium, Penicillium, Aspergillus, Alternaria, and Fusarium; the least common fungi were members of genera Geotrichum and Paecilomyces. Seasonal variations in the concentration of fungi were observed with the highest concentration of fungi in the spring and the lowest concentration of fungi in the winter. There were no statistically significant correlations between fungal concentrations and the temperature or the relative humidity of the sample sites.

Comparison of different agar diffusion methods for the detection of residues in the kidneys of pigs treated with antimicrobial drugs.

PubMed

Korkeala, H; Sorvettula, O; Mäki-Petäys, O; Hirn, J

1983-01-01

Residue analyses of the kidneys of twenty-six pigs treated with various antimicrobial drugs 20 h before slaughter and of eleven untreated pigs were performed. The effects of storage temperature of the kidneys, and of sampling location, on the residue analysis were also studied. No method alone was sufficient for the detection of residues. Oxytetracycline residues could be detected at pH 6, dihydrostreptomycin residues at pH 8, and sulphonamide residues if trimethoprim was present in the medium. Chloramphenicol, penicillin G procaine, tylosin and lincomycin residues were not detectable with the methods used. The concentration of ampicillin decreased during the storage of samples at +4°C. Most methods also yielded zones of inhibition for the frozen kidneys from untreated pigs. It seems necessary to use agar media of two different pH values: the addition of trimethoprim to the medium is also needed. The use of fresh pig kidneys, and samples containing both kidney medulla and kidney cortex, is recommended in residue analysis. Copyright © 1983. Published by Elsevier Ltd.

Assessment of bioburden encapsulated in bulk materials

NASA Astrophysics Data System (ADS)

Schubert, Wayne W.; Newlin, Laura; Chung, Shirley Y.; Ellyin, Raymond

2016-05-01

The National Aeronautics and Space Administration (NASA) imposes bioburden limitations on all spacecraft destined for solar system bodies that might harbor evidence of extant or extinct life. The subset of microorganisms trapped within solid materials during manufacture and assembly is referred to as encapsulated bioburden. In the absence of spacecraft-specific data, NASA relies on specification values to estimate total spacecraft encapsulated bioburden, typically 30 endospores/cm3 or 300 viable cells/cm3 in non-electronic materials. Specification values for endospores have been established conservatively, and represent no less than an order of magnitude greater abundance than that derived from empirical assessments of actual spacecraft materials. The goal of this study was to generate data germane to determining whether revised bulk encapsulated material values (lower than those estimated by historical specifications) tailored specifically to the materials designated in modern-day spacecraft design could be used, on a case-by-case basis, to comply with planetary protection requirements. Organic materials having distinctly different chemical properties and configurations were selected. This required more than one experimental and analytical approach. Filtration was employed for liquid electrolytes, lubricants were suspended in an aqueous solution and solids (wire and epoxy sealant) were cryogenically milled. The final data characteristic for all bioburden estimates was microbial colony formation in rich agar growth medium. To assess survival potential, three non-spore-forming bacterial cell lines were systematically encapsulated in an epoxy matrix, liberated via cryogenic grinding, and cultured. Results suggest that bulk solid materials harbor significantly fewer encapsulated microorganisms than are estimated by specification values. Lithium-ion battery electrolyte reagents housed fewer than 1 CFU/cm3. Results also demonstrated that non-spore-forming microorganisms are capable of surviving encapsulation within, and liberation from, epoxy solids. It must be noted, however, that all purposely spiked experimental solids, resulted in very low recovery (1 × 10-3-1 × 10-5 CFU/cm3) of viable organisms.

CHROMagar Candida as the Sole Primary Medium for Isolation of Yeasts and as a Source Medium for the Rapid-Assimilation-of-Trehalose Test

PubMed Central

Murray, Melissa P.; Zinchuk, Riva; Larone, Davise H.

2005-01-01

The chromogenic medium BBL CHROMagar Candida (CAC) was evaluated as a sole primary medium for the isolation of yeasts from clinical specimens in which yeasts are the primary concern. Additionally, the reliability of the rapid-assimilation-of-trehalose (RAT) test in yielding correct results with isolates taken from CAC was assessed. A total of 270 throat, urine, and genital (TUG) specimens were streaked onto CAC, Sabouraud dextrose agar (SDA), inhibitory mold agar (IMA), and Mycosel (MYC). A total of 69 blood culture broths that were smear positive for yeast were streaked onto CAC and SDA. A 1-h RAT test (NCCLS M35-A) was performed simultaneously on isolates from CAC and SDA. A total of 112 TUG specimens yielded yeast colonies (CAC, 111 colonies; IMA, 105; SDA, 103; MYC, 91). The 69 blood culture yeasts grew on both CAC and SDA. Mixed cultures of yeasts were detected on 11 CAC plates but were unrecognized on other media. Colonies suspected of being C. glabrata on 32 CAC plates were all RAT test positive and confirmed to be C. glabrata; of 59 colonies with various characteristics of color and morphology on CAC, none were RAT positive, and all were conventionally identified as yeasts other than C. glabrata (sensitivity and specificity, 100%). The same isolates from SDA tested for RAT produced six false negatives and no false positives (sensitivity, 81%; specificity, 100%). The results show that CAC can be used as the sole primary medium for recovery of yeasts from clinical specimens. Additionally, isolates grown on CAC yield excellent results with the RAT test utilized in this study. PMID:15750085

CHROMagar Candida as the sole primary medium for isolation of yeasts and as a source medium for the rapid-assimilation-of-trehalose test.

PubMed

Murray, Melissa P; Zinchuk, Riva; Larone, Davise H

2005-03-01

The chromogenic medium BBL CHROMagar Candida (CAC) was evaluated as a sole primary medium for the isolation of yeasts from clinical specimens in which yeasts are the primary concern. Additionally, the reliability of the rapid-assimilation-of-trehalose (RAT) test in yielding correct results with isolates taken from CAC was assessed. A total of 270 throat, urine, and genital (TUG) specimens were streaked onto CAC, Sabouraud dextrose agar (SDA), inhibitory mold agar (IMA), and Mycosel (MYC). A total of 69 blood culture broths that were smear positive for yeast were streaked onto CAC and SDA. A 1-h RAT test (NCCLS M35-A) was performed simultaneously on isolates from CAC and SDA. A total of 112 TUG specimens yielded yeast colonies (CAC, 111 colonies; IMA, 105; SDA, 103; MYC, 91). The 69 blood culture yeasts grew on both CAC and SDA. Mixed cultures of yeasts were detected on 11 CAC plates but were unrecognized on other media. Colonies suspected of being C. glabrata on 32 CAC plates were all RAT test positive and confirmed to be C. glabrata; of 59 colonies with various characteristics of color and morphology on CAC, none were RAT positive, and all were conventionally identified as yeasts other than C. glabrata (sensitivity and specificity, 100%). The same isolates from SDA tested for RAT produced six false negatives and no false positives (sensitivity, 81%; specificity, 100%). The results show that CAC can be used as the sole primary medium for recovery of yeasts from clinical specimens. Additionally, isolates grown on CAC yield excellent results with the RAT test utilized in this study.

Development of selective and differential medium for Shigella sonnei using three carbohydrates (lactose, sorbitol, and xylose) and X-Gal.

PubMed

Na, G N; Kim, S A; Kwon, O C; Rhee, M S

2015-08-01

The aim of this study was to develop a new selective and differential medium for isolating Shigella sonnei (designated 3SD medium). The new medium was based on three carbohydrates (lactose, sorbitol, and xylose) and a chromogenic substrate (5-bromo-4-chloro-3-indolyl-β-D-galactopyranoside, X-Gal). S. sonnei cannot ferment lactose, sorbitol, or xylose, but can ferment X-Gal, which generates turquoise-blue colonies with rough edges. Other bacteria (54 strains of foodborne pathogens and spoilage bacteria) produced visually distinct colonies on 3SD medium (colorless or pink-violet colonies), or their growth was inhibited on 3SD medium. The optimum concentration of 50 mg/L X-Gal was selected because it yielded the highest level of morphological discrimination between S. sonnei and other bacteria, and this concentration was cost-effective. Bile salt concentration optimization was performed using healthy, heat-injured, and acid-injured S. sonnei. The recovery rate differed significantly depending on the bile salt concentration; media containing >1.0 g/L bile salt showed significantly lower recovery of stress-injured cells than medium containing 0.5 g/L bile salt (P<0.05). Growth of all Gram-positive bacteria was inhibited on medium containing 0.5 g/L bile salt; therefore, this concentration was used as the optimal concentration. Previous media used to isolate Shigella spp. (MacConkey, xylose lysine desoxycholate, and Salmonella-Shigella agar) showed poor performance when used to support the growth of injured S. sonnei cells, whereas 3SD medium supported a high growth rate of injured and healthy cells (equivalent to that obtained with nutrient-rich tryptic soy agar). To validate the performance of 3SD medium with real specimens, S. sonnei and other bacteria were spiked into samples such as untreated water, carrot, salad, and oyster. 3SD medium showed superior specificity (100%) and sensitivity (100%) for S. sonnei, and yielded no false-positive or false-negative results. Thus, the novel 3SD medium described herein is a powerful tool for the rapid and efficient selective isolation of S. sonnei in research and clinical laboratories, and the food industry. Copyright © 2015 Elsevier B.V. All rights reserved.

Misinterpretation of Gram Stain from the Stationary Growth Phase of Positive Blood Cultures for Brucella and Acinetobacter Species.

PubMed

Bazzi, Ali M; Al-Tawfiq, Jaffar A; Rabaan, Ali A

2017-01-01

Acinetobacter baumannii and Brucella species are Gram-negative organisms that are vulnerable to misinterpretation as Gram-positive or Gram-variable in blood cultures. We assess the random errors in gram stain interpretation to reduce the likelihood of such errors and therefore patient harm. Aerobic and anaerobic blood cultures from two patients in an acute care facility in Saudi Arabia were subjected to preliminary Gram-staining. In case 1, VITEK-2 Anaerobe Identification, repeat Gram staining from a blood agar plate, Remel BactiDrop™ Oxidase test, Urea Agar urease test and real-time PCR were used to confirm presence of Brucella and absence of Coryneform species. In case 2, repeat Gram- staining from the plate and the vials, VITEK-2 Gram-Negative Identification, real-time PCR and subculture on to Columbia agar, blood agar, and MacConkey agar were carried out to identify A. baumannii . In case 1, initially pleomorphic Gram-positive bacteria were identified. Coryneform species were suspected. Tiny growth was observed after 24 h on blood agar plates, and good growth by 48 h. Presence of Brucella species was ultimately confirmed. In case 2, preliminary Gram-stain results suggested giant Gram-positive oval cocci. Further testing over 18-24 h identified A. baumannii . Oxidase test from the plate and urease test from the culture vial is recommended after apparent identification of pleomorphic Gram-positive bacilli from blood culture, once tiny growth is observed, to distinguish Brucella from Corynebacterium species. If giant Gram-positive oval cocci are indicated by preliminary Gram-staining, it is recommended that the Gram stain be repeated from the plate after 4-6 h, or culture should be tested in Triple Sugar Iron (TSI) medium and the Gram stain repeated after 2-4 h incubation.

Commercial Lysogeny Broth culture media and oxidative stress: a cautious tale.

PubMed

Ezraty, Benjamin; Henry, Camille; Hérisse, Marion; Denamur, Erick; Barras, Frédéric

2014-09-01

Lysogeny Broth (LB), most often misnamed Luria-Bertani medium, ranks among the most commonly used growth media in microbiology. Surprisingly, we observed that oxidative levels vary with the commercial origin of the LB ready to use powder. Indeed, growth on solid media of Escherichia coli and Salmonella derivatives lacking antioxidative stress defenses, such as oxyR mutant devoid of the H2O2-sensing transcriptional activator or Hpx(-) strains lacking catalases and peroxidases, exhibit different phenotypes on LB-Sigma or LB-Difco. Using gene fusion and exogenously added catalase, we found that LB-Sigma contains higher levels of H2O2 than LB-Difco. Also we observed differences in population counts of 82 clinical and environmental isolates of E. coli, depending on the LB used. Further investigations revealed a significant influence of the commercial origin of agar as well. Besides being a warning to the wide population of LB users, our observations provide researchers in the oxidative stress field with a tool to appreciate the severity of mutations in antioxidative stress defenses. Copyright © 2014 Elsevier Inc. All rights reserved.

Direct Biotransformation of Dioscin into Diosgenin in Rhizome of Dioscorea zingiberensis by Penicillium dioscin.

PubMed

Dong, Jingzhou; Lei, Can; Lu, Dayan; Wang, Ying

2015-06-01

Diosgenin is an important precursor for synthesis of more than 200 steroidal hormone medicines. Rhizome of Dioscorea zingiberensis C. H. Wright (RDZ) contained the highest content of diosgenin in Dioscorea plant species. Diosgenin is traditionally extracted by acid hydrolysis from RDZ. However, the acid hydrolysis process produces massive wastewater which caused serious environment pollution. In this study, diosgenin extraction by direct biotransformation with Penicillium dioscin was investigated. The spawn cultivation conditions were optimized as: Czapeks liquid culture medium without sugar and agar (1,000 ml) + 6.0 g dioscin/6.0 g DL, 30 °C, 36 h; solid fermentation of RDZ: mycelia/RDZ of 0.05 g/kg, 30 °C, 50 h; the yield of diosgenin was over 90 %. Spawn cultivation was crucial for the direct biotransformation. In the spawn cultivation, amount and ratio of dioscin/DL were the key factors to promote biotransformation activity of P. dioscin. This biotransformation method was environment-friendly, simple and energy saving, and might be a potential substitute for acid hydrolysis in diosgenin extraction industry.

Selection and identification of fungi isolated from sugarcane bagasse and their application for phenanthrene removal from soil.

PubMed

Cortés-Espinosa, D V; Fernández-Perrino, F J; Arana-Cuenca, A; Esparza-García, F; Loera, O; Rodríguez-Vázquez, R

2006-01-01

This work investigated the identification and selection of fungi isolated from sugarcane bagasse and their application for phenanthrene (Phe) removal from soil. Fungi were identified by PCR amplification of ITS regions as Aspergillus terrus, Aspergillus fumigatus and Aspergillus niger, Penicillium glabrum and Cladosporium cladosporioides. A primary selection of fungi was accomplished in plate, considering Phe tolerance of every strain in two different media: potato dextrose agar (PDA) and mineral medium (MM). The radial extension rate (r(r)) in PDA exhibited significant differences (p<0.05) at 200 and 400 ppm of Phe. A secondary selection of A. niger, C. cladosporoides, and P. glabrum sp. was achieved based on their tolerance to 200, 400, 600 and 800 ppm of Phe, in solid culture at a sugarcane bagasse/contaminated soil ratio of 95:5, in Toyamas, Czapeck and Wunder media. Under these conditions, a maximum (70%) Phe removal by A. niger was obtained. In addition C. cladosporioides and A. niger were able to remove high (800 ppm) Phe concentrations.

Synthesis of Silver Nanoparticles from Microbial Source-A Green Synthesis Approach, and Evaluation of its Antimicrobial Activity against Escherichia coli

NASA Astrophysics Data System (ADS)

Behera, S. S.; Jha, S.; Arakha, M.; Panigrahi, T. K.

2012-03-01

TRACT Nanoparticles synthesis by biological methods using various microorganisms, plants, and plant extracts and enzymes have attracted a great attention as these are cost effective, nontoxic, eco-friendly and an alternative to physical and chemical methods. In this research, Silver nanoparticles (Ag-NPs) were synthesized from AgNO3 solution by green synthesis process with the assistance of microbial source only. The detailed characterization of the Ag NPs were carried out using UV-visible spectroscopy, Scanning electron microscopy (SEM), Energy dispersive X-ray Spectroscopy (EDS), Dynamic light scattering (DLS) analysis, and their antimicrobial evaluation was done against Escherichia coli. The UV-visible spectroscopy analysis showed the surface plasmon resonance property of nanoparticles. The DLS analysis showed the particle distribution of synthesized silver nanoparticles in solution, and SEM analysis showed the morphology of nanoparticles. The elemental composition of synthesized sample was confirmed by EDS analysis. Antibacterial assay of synthesized Ag NP was carried out in solid (Nutrient Agar) growth medium against E.coli. The presence of zone of inhibition clearly indicated the antibacterial activity of silver nanoparticles.

Utilization of Bacillus sp. strain TAT105 as a biological additive to reduce ammonia emissions during composting of swine feces.

PubMed

Kuroda, Kazutaka; Waki, Miyoko; Yasuda, Tomoko; Fukumoto, Yasuyuki; Tanaka, Akihiro; Nakasaki, Kiyohiko

2015-01-01

Bacillus sp. strain TAT105 is a thermophilic, ammonium-tolerant bacterium that grows assimilating ammonium nitrogen and reduces ammonia emission during composting of swine feces. To develop a practical use of TAT105, a dried solid culture of TAT105 (5.3 × 10(9) CFU/g of dry matter) was prepared as an additive. It could be stored for one year without significant reduction of TAT105. Laboratory-scale composting of swine feces was conducted by mixing the additive. When the additive, mixed with an equal weight of water one day before use, was added to obtain a TAT105 concentration of above 10(7) CFU/g of dry matter in the initial material, the ammonia concentration emitted was lower and nitrogen loss was approximately 22% lower in the treatment with the additive than in the control treatment without the additive. The colony formation on an agar medium containing high ammonium could be used for enumeration of TAT105 in the composted materials.

HS-SPME-GC-MS analysis of body odor to test the efficacy of foot deodorant formulations.

PubMed

Caroprese, Alessandro; Gabbanini, Simone; Beltramini, Claudia; Lucchi, Elena; Valgimigli, Luca

2009-11-01

Foot malodor is mostly due to short-chain fatty acids produced by bacterial metabolism of eccrine sweating. We aimed to develop a protocol for an objective (instrumental) efficacy evaluation of foot deodorant formulations. Head-space solid-phase microextractions of target fatty acids from the feet of six healthy volunteers were analyzed by GC-MS. A comparative analysis of the treated vs. the untreated foot was performed in each subject after washing the feet with a physiologic solution and incubating at 36 degrees C for 24-72 h in tryptic soy agar growth medium. Acetic, butyric, isobutyric and isovaleric acids were identified as the main contributors to foot malodor in the majority of volunteers. Propionic, valeric and isocaproic acids were also detected in some subjects. Comparative analysis according to the protocol developed showed a statistically significant (P<0.01) reduction of target fatty acids ranging from -26.6% to -77.0%. The protocol developed is a convenient, sensitive and non-invasive method to test the efficacy of foot deodorant formulations in human volunteers.

Temporary Immersion System for Date Palm Micropropagation.

PubMed

Othmani, Ahmed; Bayoudh, Chokri; Sellemi, Amel; Drira, Noureddine

2017-01-01

The temporary immersion system (TIS) is being used with tremendous success for automation of micropropagation of many plant species. TIS usually consists of a culture vessel comprising two compartments, an upper one with the plant material and a lower one with the liquid culture medium and an automated air pump. The latter enables contact between all parts of the explants and the liquid medium by setting overpressure to the lower part of the container. These systems are providing the most satisfactory conditions for date palm regeneration via shoot organogenesis and allow a significant increase of multiplication rate (5.5-fold in comparison with that regenerated on agar-solidified medium) and plant material quality, thereby reducing production cost.

Selective medium for the isolation of Bacteroides gingivalis.

PubMed

Hunt, D E; Jones, J V; Dowell, V R

1986-03-01

Bacteroides gingivalis has been implicated in various forms of periodontal disease and may be responsible for other diseases in humans. The role of B. gingivalis in disease has been difficult to assess, because it is inhibited by most selective media commonly used by clinical laboratories to aid in isolating gram-negative, nonsporeforming anaerobes. We have developed a new medium, Bacteroides gingivalis agar, which contains bacitracin, colistin, and nalidixic acid as selective agents. This medium allowed B. gingivalis to be isolated from oral specimens with little difficulty and also allowed B. gingivalis to be isolated from phenotypically similar Bacteroides species, such as B. asaccharolyticus and B. endodontalis, with which it can easily be confused.

Bacillus licheniformis BT5.9 Isolated from Changar Hot Spring, Malang, Indonesia, as a Potential Producer of Thermostable α-amylase

PubMed Central

Ibrahim, Darah; Zhu, Han Li; Yusof, Nuraqilah; Isnaeni; Hong, Lim Sheh

2013-01-01

A total of 34 bacterial isolates were obtained from soil samples collected from Changar Hot Spring, Malang, Indonesia. Of these, 13 isolates produced a zone of hydrolysis in starch-nutrient agar medium and generated various amylases in liquid medium. One isolate was selected as the best amylase producer and was identified as Bacillus licheniformis BT5.9. The improvement of culture conditions (initial medium pH of 5.0, cultivation temperature of 50°C, agitation speed of 100 rpm and inoculum size of 1.7 × 109 cells/ml) provided the highest amylase production (0.327 U/ml). PMID:24575243

Surgical site infections due to rapidly growing mycobacteria in puducherry, India.

PubMed

Kannaiyan, Kavitha; Ragunathan, Latha; Sakthivel, Sulochana; Sasidar, A R; Muralidaran; Venkatachalam, G K

2015-03-01

Rapidly growing Mycobacteria are increasingly recognized, nowadays as an important pathogen that can cause wide range of clinical syndromes in humans. We herein describe unrelated cases of surgical site infection caused by Rapidly growing Mycobacteria (RGM), seen during a period of 12 months. Nineteen patients underwent operations by different surgical teams located in diverse sections of Tamil Nadu, Pondicherry, Karnataka, India. All patients presented with painful, draining subcutaneous nodules at the infection sites. Purulent material specimens were sent to the microbiology laboratory. Gram stain and Ziehl-Neelsen staining methods were used for direct examination. Culture media included blood agar, chocolate agar, MacConkey agar, Sabourauds agar and Lowenstein-Jensen medium for Mycobacteria. Isolated microorganisms were identified and further tested for antimicrobial susceptibility by standard microbiologic procedures. Mycobacterium fortuitum and M.chelonae were isolated from the purulent drainage obtained from wounds by routine microbiological techniques from all the specimens. All isolates analyzed for antimicrobial susceptibility pattern were sensitive to clarithromycin, linezolid and amikacin but were variable to ciprofloxacin, rifampicin and tobramycin. Our case series highlights that a high level of clinical suspicion should be maintained for patients presenting with protracted soft tissue lesions with a history of trauma or surgery as these infections not only cause physical but also emotional distress that affects both the patients and the surgeon.

Spaceflight exposure effects on transcription, activity, and localization of alcohol dehydrogenase in the roots of Arabidopsis thaliana.

PubMed Central

Porterfield, D M; Matthews, S W; Daugherty, C J; Musgrave, M E

1997-01-01

Although considerable research and speculation have been directed toward understanding a plant's perception of gravity and the resulting gravitropic responses, little is known about the role of gravity-dependent physical processes in normal physiological function. These studies were conducted to determine whether the roots of plants exposed to spaceflight conditions may be experiencing hypoxia. Arabidopsis thaliana (L.) Heynh. plants were grown in agar medium during 6 or 11 d of spaceflight exposure on shuttle missions STS-54 (CHROMEX-03) and STS-68 (CHROMEX-05), respectively. The analysis included measurement of agar redox potential and root alcohol dehydrogenase (ADH) activity, localization, and expression. ADH activity increased by 89% as a result of spaceflight exposure for both CHROMEX-03 and -05 experiments, and ADH RNase protection assays revealed a 136% increase in ADH mRNA. The increase in ADH activity associated with the spaceflight roots was realized by a 28% decrease in oxygen availability in a ground-based study; however, no reduction in redox potential was observed in measurements of the spaceflight bulk agar. Spaceflight exposure appears to effect a hypoxic response in the roots of agar-grown plants that may be caused by changes in gravity-mediated fluid and/or gas behavior. PMID:9085569

Spaceflight exposure effects on transcription, activity, and localization of alcohol dehydrogenase in the roots of Arabidopsis thaliana

NASA Technical Reports Server (NTRS)

Porterfield, D. M.; Matthews, S. W.; Daugherty, C. J.; Musgrave, M. E.

1997-01-01

Although considerable research and speculation have been directed toward understanding a plant's perception of gravity and the resulting gravitropic responses, little is known about the role of gravity-dependent physical processes in normal physiological function. These studies were conducted to determine whether the roots of plants exposed to spaceflight conditions may be experiencing hypoxia. Arabidopsis thaliana (L.) Heynh. plants were grown in agar medium during 6 or 11 d of spaceflight exposure on shuttle missions STS-54 (CHROMEX-03) and STS-68 (CHROMEX-05), respectively. The analysis included measurement of agar redox potential and root alcohol dehydrogenase (ADH) activity, localization, and expression. ADH activity increased by 89% as a result of spaceflight exposure for both CHROMEX-03 and -05 experiments, and ADH RNase protection assays revealed a 136% increase in ADH mRNA. The increase in ADH activity associated with the spaceflight roots was realized by a 28% decrease in oxygen availability in a ground-based study; however, no reduction in redox potential was observed in measurements of the spaceflight bulk agar. Spaceflight exposure appears to effect a hypoxic response in the roots of agar-grown plants that may be caused by changes in gravity-mediated fluid and/or gas behavior.

Improving culture media for the isolation of Clostridium difficile from compost.

PubMed

Dharmasena, Muthu; Jiang, Xiuping

2018-06-01

This study was to optimize the detection methods for Clostridium difficile from the animal manure-based composts. Both autoclaved and unautoclaved dairy composts were inoculated with a 12-h old suspension of a non-toxigenic C. difficile strain (ATCC 43593) and then plated on selected agar for vegetative cells and endospores. Six types of enrichment broths supplemented with taurocholate and l-cysteine were assessed for detecting a low level of artificially inoculated C. difficile (ca. 5 spores/g) from dairy composts. The efficacy of selected enrichment broths was further evaluated by isolating C. difficile from 29 commercial compost samples. Our results revealed that using heat-shock was more effective than using ethanol-shock for inducing endospore germination, and the highest endospore count (p < 0.05) was yielded at 60 °C for 25 min. C. difficile agar base, supplemented with 0.1% l-cysteine, 7% defibrinated horse blood, and cycloserine-cefoxitin (CDA-CYS-H-CC agar) was the best medium (p < 0.05) for recovering vegetative cells from compost. C. difficile endospore populations from both types of composts enumerated on both CDA-CYS-H-CC agar supplemented with 0.1% sodium taurocholate (CDA-CYS-H-CC-T agar) and brain heart infusion agar supplemented with 0.5% yeast extract, 0.1% l-cysteine, cycloserine-cefoxitin, and 0.1% sodium taurocholate (BHIA-YE-CYS-CC-T agar) media were not significantly different from each other (p > 0.05). Overall, enrichment of inoculated compost samples in broths containing moxalactam-norfloxacin (MN) produced significantly higher (p < 0.05) spore counts than in non-selective broths or broths supplemented with CC. Enrichment in BHIB-YE-CYS-MN-T broth followed by culturing on an agar containing 7% horse blood and 0.1% taurocholate provided a more sensitive and selective combination of media for detecting a low population of C. difficile from environmental samples with high background microflora. Copyright © 2018 Elsevier Ltd. All rights reserved.

Comparative analyses of viable bacterial counts in foods and seawater under microplate based liquid- and conventional agar plate cultivation: increased culturability of marine bacteria under liquid cultivation.

PubMed

Shigematsu, Toru; Ueno, Shigeaki; Tsuchida, Yasuharu; Hayashi, Mayumi; Okonogi, Hiroko; Masaki, Haruhiko; Fujii, Tomoyuki

2007-12-01

Bacterial counts under liquid cultivation using 96-well microplates were performed. The counts under liquid and under solid cultivation were equivalent in foods, although the counts under liquid cultivation exceeded those under solid cultivation in seawater, suggesting that some bacteria in seawater were viable but did not form detectable colonies. Phylogenetic analysis of bacteria obtained under liquid cultivation was also performed.

Regulation of coal polymer degradation by fungi. Eighth quarterly report, [January--March 1996

DOE Office of Scientific and Technical Information (OSTI.GOV)

Irvine, R.L.; Bumpus, J.A.

1996-07-28

Progress is reported on solubilization of low-rank coal by enzyme activity derived from Trametes versicolor or P. chrysosporium. Specifically during the reporting period efforts were directed towards the determining the effect of pH on solubilization of leonardite, the role of laccase in low coal solubilization and metabolism, the decolorization of soluble coal macromolecule by P. chrysosprium and T. versicolor in solid agar gel, and the solubilization of low rank coal in slurry cultures and solid phase reactors.

Antibacterial activity and effects of Colla corii asini on Salmonella typhimurium invasion in vitro and in vivo.

PubMed

Park, Kwang-Il; Lee, Mi-Ra; Oh, Tae-Woo; Kim, Kwang-Youn; Ma, Jin-Yeul

2017-12-04

Salmonella enterica serovar Typhimurium is a foodborne pathogen that triggers inflammatory responses in the intestines of humans and livestock. Colla corii asini is a traditional medicine used to treat gynecologic and chronic diseases in Korea and China. However, the antibacterial activity of Colla corii asini has been unknown. In this study, we investigated the antibacterial activity and effects of Colla corii asini extract on Salmonella typhimurium invasion. To tested for antibacterial effects of Colla corii asini extracts, we confirmed the agar diffusion using Luria solid broth medium. Also, we determined the MIC (minimum inhibitory concentration) and the MBC (minimum bactericidal concentration) value of the Colla corii asini ethanol extract (CEE) by using two-fold serial dilution methods. We evaluated the expression of salmonella invasion proteins including SipA, SipB and SipC by using Western blot and qPCR at the concentration of CEE without inhibition of bacterial growth. In vitro and vivo, we determined the inhibitory effect of invasion of S. typhimurium on CEE by using gentamicin assay and S. typhimurium-infected mice. CEE significantly inhibited the growth of Salmonella typhimurium in an agar diffuse assay and had an MIC of 0.78 mg/ml and an MBC of 1.56 mg/ml. Additionally, CEE reduced Salmonella typhimurium cell invasion via the inhibition of Salmonella typhimurium invasion proteins, such as SipA, SipB and SipC. Furthermore, CEE significantly suppressed invasion in the small intestines (ilea) of mice injected with Salmonella typhimurium. These findings show that Colla corii asini exerts antibacterial activity and suppresses Salmonella typhimurium invasion in vitro and in vivo. Together, these findings demonstrate that Colla corii asini is a potentially useful therapeutic herbal medicine for treating salmonella-mediated diseases.

Isolation and Purification of Antibiotic Material from Physarum gyrosum

PubMed Central

Schroeder, H. R.; Mallette, M. F.

1973-01-01

The myxomycete Physarum gyrosum was cultured in its plasmodial stage on agar plates containing 0.025 M phosphate buffer at pH 6.5, 2% bakers' yeast, and 0.2% glucose and was supplemented with live Escherichia coli. Extracts of these plasmodia contained several antibiotic substances. Antibiotic materials were partially purified by dialysis of the agar medium-mold mixture, evaporation of the dialyzate, and butanol extraction of the residue. Further purification in two paper and two thin-layer chromatographic systems gave one product which was pure in six thin-layer chromatographic systems. Antibiotic activity against some gram-positive and gram-negative bacteria and yeasts was demonstrated with partially purified extracts and a paper-chromatographically separated fraction. One pure antibiotic was effective against Bacillus cereus. PMID:4799591

Plating Bacteriophage M13.

PubMed

Green, Michael R; Sambrook, Joseph

2017-10-03

A plaque of bacteriophage M13 derives from infection of a single bacterium by a single virus particle. The progeny particles infect neighboring bacteria, which, in turn, release another generation of daughter virus particles. If the bacteria are growing in semisolid medium (e.g., containing agar or agarose), then the diffusion of the progeny particles is limited. Cells infected with bacteriophage M13 are not killed, but have a longer generation time than uninfected Escherichia coli In consequence, plaques appear as areas of slower-growing cells on a faster-growing lawn of bacterial cells. This protocol describes plating of bacteriophage M13 stocks. Plaques are readily detectable on top agar after 4-8 h of incubation at 37°C. © 2017 Cold Spring Harbor Laboratory Press.

A new chromogenic medium for isolation of Bacteroides fragilis suitable for screening for strains with antimicrobial resistance.

PubMed

Tierney, Daniel; Copsey, Sarah D; Morris, Trefor; Perry, John D

2016-06-01

There have been an increasing number of reports describing the acquisition of antimicrobial resistance by Bacteroides fragilis including the occurrence of strains with resistance to multiple antimicrobials that are relied upon for treatment of infections. The aim of this study was to design a chromogenic selective medium for isolation of B. fragilis that could be adapted for specific isolation of antimicrobial-resistant strains. Bacteroides chromogenic agar (BCA) was the result of this endeavour and allowed growth of Bacteroides spp. as black colonies and the efficient inhibition of almost all other genera tested. The medium also allowed some differentiation of B. fragilis from other members of the B. fragilis group. When compared with an adaptation of Bacteroides bile-esculin agar (BBE) for the isolation of B. fragilis from 100 stool samples, 30 isolates of B. fragilis were recovered on BCA compared with 19 isolates recovered on BBE (P = 0.022). When supplemented with meropenem (4 μg/ml) or metronidazole (2 μg/ml), BCA could be used to select for the growth of B. fragilis isolates with resistance to these agents. We conclude that BCA is a useful research tool for surveillance studies to assess the prevalence of B. fragilis and, in particular, the occurrence of antimicrobial-resistant strains. Copyright © 2016 Elsevier Ltd. All rights reserved.

Isolation and characterization of rhizosphere bacteria for the biocontrol of the damping-off disease of tomatoes in Tunisia.

PubMed

Hammami, Inés; Ben Hsouna, Anis; Hamdi, Naceur; Gdoura, Radhouane; Triki, Mohamed Ali

2013-01-01

Fluorescent Pseudomonas spp., isolated from tomato and pepper plants rhizosphere soil, was evaluated in vitro as a potential antagonist of fungal pathogens. Pseudomonas strains were tested against the causal agents of tomatoes damping-off (Sclerotinia sclerotiorum), root rot (Fusarium solani), and causal agents of stem canker and leaf blight (Alternaria alternata). For this purpose, dual culture antagonism assays were carried out on 25% tryptic soy agar, King B medium and potato dextrose agar to determine the effect of the strains on mycelial growth of the pathogens. In addition, strains were screened for their ability to produce exoenzymes and siderophores. All the strains significantly inhibited Alternaria alternata, particularly in 25% TSA medium. Antagonistic effect on Sclerotinia sclerotiorum and Fusarium solani was greater on King B medium. Protease was produced by 30% of the strains, but no strain produced cellulase or chitinase. Finally, the selected Pseudomonas strain, Psf5, was evaluated on tomato seedling development and as a potential candidate for controlling tomato damping-off caused by Sclerotinia sclerotiorum, under growth chamber conditions. In vivo studies resulted in significant increases in plant stand as well as in root dry weight. Psf5 was able to establish and survive in tomato plants rhizosphere after 40days following the planting of bacterized seeds. © 2013 Académie des sciences. Published by Elsevier SAS. All rights reserved.

Changes in soluble sugar, starch, and alcohol dehydrogenase in Arabidopsis thaliana exposed to N2 diluted atmospheres

NASA Technical Reports Server (NTRS)

Porterfield, D. M.; Crispi, M. L.; Musgrave, M. E.

1997-01-01

Proper exchange of atmospheric gases is important for normal root and shoot metabolism in plants. This study was conducted to determine how restricted air supply affects foliar carbohydrates, while using the marker enzyme alcohol dehydrogenase (ADH) to report on the oxygenation status of the rootzone. Fourteen-day-old Arabidopsis thaliana (L.) Heynh. plants grown singly in 7-ml tubes containing agarified nutrient medium were placed in coupled Magenta vessels and exposed for six days to either ambient air or one of six different air/nitrogen dilutions. Redox potential of the agar medium was measured immediately after harvesting and freezing leaf tissue, and then root systems were quickly extracted from the agar and frozen for subsequent analyses. Redox potential measurements indicated that this series of gas mixtures produced a transition from hypoxia to anoxia in the root zones. Root ADH activity increased at higher rates as the redox potential neared anoxic levels. In contrast, ADH mRNA expression quickly neared its maximum as the medium became hypoxic and showed little further increase as it became anoxic. Foliar carbohydrate levels increased 1.5- to 2-fold with decreased availability of metabolic gases, with starch increasing at higher concentrations of air than soluble carbohydrate. The results serve as a model for plant performance under microgravity conditions, where absence of convective air movement prevents replenishment of metabolic gases.

An alternative method for the analysis of melanin production in Cryptococcus neoformans sensu lato and Cryptococcus gattii sensu lato.

PubMed

Brilhante, Raimunda S N; España, Jaime D A; de Alencar, Lucas P; Pereira, Vandbergue S; Castelo-Branco, Débora de S C M; Pereira-Neto, Waldemiro de A; Cordeiro, Rossana de A; Sidrim, José J C; Rocha, Marcos F G

2017-10-01

Melanin is an important virulence factor for several microorganisms, including Cryptococcus neoformans sensu lato and Cryptococcus gattii sensu lato, thus, the assessment of melanin production and its quantification may contribute to the understanding of microbial pathogenesis. The objective of this study was to standardise an alternative method for the production and indirect quantification of melanin in C. neoformans sensu lato and C. gattii sensu lato. Eight C. neoformans sensu lato and three C. gattii sensu lato, identified through URA5 methodology, Candida parapsilosis ATCC 22019 (negative control) and one Hortaea werneckii (positive control) were inoculated on minimal medium agar with or without L-DOPA, in duplicate, and incubated at 35°C, for 7 days. Pictures were taken from the third to the seventh day, under standardised conditions in a photographic chamber. Then, photographs were analysed using grayscale images. All Cryptococcus spp. strains produced melanin after growth on minimal medium agar containing L-DOPA. C. parapsilosis ATCC 22019 did not produce melanin on medium containing L-DOPA, while H. werneckii presented the strongest pigmentation. This new method allows the indirect analysis of melanin production through pixel quantification in grayscale images, enabling the study of substances that can modulate melanin production. © 2017 Blackwell Verlag GmbH.

Diverse bacteria isolated from microtherm oil-production water.

PubMed

Sun, Ji-Quan; Xu, Lian; Zhang, Zhao; Li, Yan; Tang, Yue-Qin; Wu, Xiao-Lei

2014-02-01

In total, 435 pure bacterial strains were isolated from microtherm oil-production water from the Karamay Oilfield, Xinjiang, China, by using four media: oil-production water medium (Cai medium), oil-production water supplemented with mineral salt medium (CW medium), oil-production water supplemented with yeast extract medium (CY medium), and blood agar medium (X medium). The bacterial isolates were affiliated with 61 phylogenetic groups that belong to 32 genera in the phyla Actinobacteria, Firmicutes, and Proteobacteria. Except for the Rhizobium, Dietzia, and Pseudomonas strains that were isolated using all the four media, using different media led to the isolation of bacteria with different functions. Similarly, nonheme diiron alkane monooxygenase genes (alkB/alkM) also clustered according to the isolation medium. Among the bacterial strains, more than 24 % of the isolates could use n-hexadecane as the sole carbon source for growth. For the first time, the alkane-degrading ability and alkB/alkM were detected in Rhizobium, Rhodobacter, Trichococcus, Micrococcus, Enterococcus, and Bavariicoccus strains, and the alkM gene was detected in Firmicutes strains.

Assay for adhesion and agar invasion in S. cerevisiae.

PubMed

Guldal, Cemile G; Broach, James

2006-11-08

Yeasts are found in natural biofilms, where many microorganisms colonize surfaces. In artificial environments, such as surfaces of man-made objects, biofilms can reduce industrial productivity, destroy structures, and threaten human life. 1-3 On the other hand, harnessing the power of biofilms can help clean the environment and generate sustainable energy. 4-8 The ability of S. cerevisiae to colonize surfaces and participate in complex biofilms was mostly ignored until the rediscovery of the differentiation programs triggered by various signaling pathways and environmental cues in this organism. 9, 10 The continuing interest in using S. cerevisiae as a model organism to understand the interaction and convergence of signaling pathways, such as the Ras-PKA, Kss1 MAPK, and Hog1 osmolarity pathways, quickly placed S. cerevisiae in the junction of biofilm biology and signal transduction research. 11-20 To this end, differentiation of yeast cells into long, adhesive, pseudohyphal filaments became a convenient readout for the activation of signal transduction pathways upon various environmental changes. However, filamentation is a complex collection of phenotypes, which makes assaying for it as if it were a simple phenotype misleading. In the past decade, several assays were successfully adopted from bacterial biofilm studies to yeast research, such as MAT formation assays to measure colony spread on soft agar and crystal violet staining to quantitatively measure cell-surface adherence. 12, 21 However, there has been some confusion in assays developed to qualitatively assess the adhesive and invasive phenotypes of yeast in agar. Here, we present a simple and reliable method for assessing the adhesive and invasive quality of yeast strains with easy-to-understand steps to isolate the adhesion assessment from invasion assessment. Our method, adopted from previous studies, 10, 16 involves growing cells in liquid media and plating on differential nutrient conditions for growth of large spots, which we then wash with water to assess adhesion and rub cells completely off the agar surface to assess invasion into the agar. We eliminate the need for streaking cells onto agar, which affects the invasion of cells into the agar. In general, we observed that haploid strains that invade agar are always adhesive, yet not all adhesive strains can invade agar medium. Our approach can be used in conjunction with other assays to carefully dissect the differentiation steps and requirements of yeast signal transduction, differentiation, quorum sensing, and biofilm formation.

Assay for Adhesion and Agar Invasion in S. cerevisiae

PubMed Central

Guldal, Cemile G; Broach, James

2006-01-01

Yeasts are found in natural biofilms, where many microorganisms colonize surfaces. In artificial environments, such as surfaces of man-made objects, biofilms can reduce industrial productivity, destroy structures, and threaten human life. 1-3 On the other hand, harnessing the power of biofilms can help clean the environment and generate sustainable energy. 4-8 The ability of S. cerevisiae to colonize surfaces and participate in complex biofilms was mostly ignored until the rediscovery of the differentiation programs triggered by various signaling pathways and environmental cues in this organism. 9, 10 The continuing interest in using S. cerevisiae as a model organism to understand the interaction and convergence of signaling pathways, such as the Ras-PKA, Kss1 MAPK, and Hog1 osmolarity pathways, quickly placed S. cerevisiae in the junction of biofilm biology and signal transduction research. 11-20 To this end, differentiation of yeast cells into long, adhesive, pseudohyphal filaments became a convenient readout for the activation of signal transduction pathways upon various environmental changes. However, filamentation is a complex collection of phenotypes, which makes assaying for it as if it were a simple phenotype misleading. In the past decade, several assays were successfully adopted from bacterial biofilm studies to yeast research, such as MAT formation assays to measure colony spread on soft agar and crystal violet staining to quantitatively measure cell-surface adherence. 12, 21 However, there has been some confusion in assays developed to qualitatively assess the adhesive and invasive phenotypes of yeast in agar. Here, we present a simple and reliable method for assessing the adhesive and invasive quality of yeast strains with easy-to-understand steps to isolate the adhesion assessment from invasion assessment. Our method, adopted from previous studies, 10, 16 involves growing cells in liquid media and plating on differential nutrient conditions for growth of large spots, which we then wash with water to assess adhesion and rub cells completely off the agar surface to assess invasion into the agar. We eliminate the need for streaking cells onto agar, which affects the invasion of cells into the agar. In general, we observed that haploid strains that invade agar are always adhesive, yet not all adhesive strains can invade agar medium. Our approach can be used in conjunction with other assays to carefully dissect the differentiation steps and requirements of yeast signal transduction, differentiation, quorum sensing, and biofilm formation. PMID:18704175

Comparison of plate counts, Petrifilm, dipslides, and adenosine triphosphate bioluminescence for monitoring bacteria in cooling-tower waters.

PubMed

Mueller, Sherry A; Anderson, James E; Kim, Byung R; Ball, James C

2009-04-01

Effective bacterial control in cooling-tower systems requires accurate and timely methods to count bacteria. Plate-count methods are difficult to implement on-site, because they are time- and labor-intensive and require sterile techniques. Several field-applicable methods (dipslides, Petrifilm, and adenosine triphosphate [ATP] bioluminescence) were compared with the plate count for two sample matrices--phosphate-buffered saline solution containing a pure culture of Pseudomonas fluorescens and cooling-tower water containing an undefined mixed bacterial culture. For the pure culture, (1) counts determined on nutrient agar and plate-count agar (PCA) media and expressed as colony-forming units (CFU) per milliliter were equivalent to those on R2A medium (p = 1.0 and p = 1.0, respectively); (2) Petrifilm counts were not significantly different from R2A plate counts (p = 0.99); (3) the dipslide counts were up to 2 log units higher than R2A plate counts, but this discrepancy was not statistically significant (p = 0.06); and (4) a discernable correlation (r2 = 0.67) existed between ATP readings and plate counts. For cooling-tower water samples (n = 62), (1) bacterial counts using R2A medium were higher (but not significant; p = 0.63) than nutrient agar and significantly higher than tryptone-glucose yeast extract (TGE; p = 0.03) and PCA (p < 0.001); (2) Petrifilm counts were significantly lower than nutrient agar or R2A (p = 0.02 and p < 0.001, respectively), but not statistically different from TGE, PCA, and dipslides (p = 0.55, p = 0.69, and p = 0.91, respectively); (3) the dipslide method yielded bacteria counts 1 to 3 log units lower than nutrient agar and R2A (p < 0.001), but was not significantly different from Petrifilm (p = 0.91), PCA (p = 1.00) or TGE (p = 0.07); (4) the differences between dipslides and the other methods became greater with a 6-day incubation time; and (5) the correlation between ATP readings and plate counts varied from system to system, was poor (r2 values ranged from < 0.01 to 0.47), and the ATP method was not sufficiently sensitive to measure counts below approximately 10(4) CFU/mL.

Microbial contamination of soft contact lenses & accessories in asymptomatic contact lens users

PubMed Central

Thakur, Deeksha V.; Gaikwad, Ujjwala N.

2014-01-01

Background & objectives: With increasing use of soft contact lenses the incidence of contact lens induced infections is also increasing. This study was aimed to assess the knowledge of new and existing contact lens users about the risk of microbial contamination associated with improper use and maintenance of contact lenses, type of microbial flora involved and their potential to cause ophthalmic infections. Methods: Four samples each from 50 participants (n=200) were collected from the lenses, lens care solutions, lens care solution bottles and lens cases along with a questionnaire regarding their lens use. The samples were inoculated onto sheep blood agar, Mac Conkey's agar and Sabouraud's dextrose agar. Organisms were identified using standard laboratory protocols. Results: Overall rate of microbial contamination among the total samples was 52 per cent. The most and the least contaminated samples were found to be lens cases (62%) and lens care solution (42%), respectively. The most frequently isolated contaminant was Staphylococcus aureus (21%) followed by Pseudomonas species (19.5%). Majority (64%) of the participants showed medium grade of compliance to lens cleaning practices. Rate of contamination was 100 and 93.75 per cent respectively in those participants who showed low and medium compliance to lens care practices as compared to those who had high level of compliance (43.75%) (P<0.05). Interpretation & conclusions: Lens care practices amongst the participants were not optimum which resulted into high level contamination. Hence, creating awareness among the users about the lens care practices and regular cleaning and replacements of lens cases are required. PMID:25297366

CD3-T cell receptor modulation is selectively induced in CD8 but not CD4 lymphocytes cultured in agar.

PubMed Central

Oudrhiri, N; Farcet, J P; Gourdin, M F; M'Bemba, E; Gaulard, P; Katz, A; Divine, M; Galazka, A; Reyes, F

1990-01-01

The CD3-T cell receptor (TcR) complex is central to the immune response. Upon binding by specific ligands, internalized CD3-TcR molecules increase, and either T cell response or unresponsiveness may ensue depending on the triggering conditions. Using semi-solid agar culture, we have shown previously that quiescent CD4 but not CD8 lymphocytes generate clonal colonies under phytohaemagglutinin stimulation. Here we have demonstrated that the agar induces selective CD3-TcR modulation in the CD8 and not in the CD4 subset. CD8 lymphocytes preactivated in liquid culture and recultured in agar with exogenous recombinant interleukin-2 generate colonies with a modulated CD3-TcR surface expression. The peptides composing the CD3-TcR complex are synthesized in CD8 colonies as well as in CD4; however, the CD3 gamma chain is phosphorylated at a higher level in CD8 colonies. A component of the agar polymer, absent in agarose, appears to be the ligand that induces differential CD3-TcR modulation in the CD8 subset. In contrast to agar culture, CD8 colonies can be derived from quiescent CD8 lymphocytes in agarose. These CD8 colonies express unmodulated CD-TcR. CD3-TcR modulation with anti-CD3 monoclonal antibody prior to culturing in agarose inhibits the colony formation. We conclude that given triggering conditions can result in both CD3-TcR modulation and inhibition of the proliferative response selectively in the CD8 lymphocyte subset and not in the CD4. Images Fig. 3 Fig. 4 Fig. 5 PMID:2146997

Evaluation of the Bruker Biotyper Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry System for Identification of Aspergillus Species Directly from Growth on Solid Agar Media

PubMed Central

Li, Ying; Wang, He; Zhao, Yu-Pei; Xu, Ying-Chun; Hsueh, Po-Ren

2017-01-01

We evaluated the accuracy of the Bruker Biotyper matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) system at identifying clinical isolates of Aspergillus species that were grown on agar media. A total of 381 non-duplicate Aspergillus isolates representing 21 different Aspergillus species identified by molecular analysis were included in this study. The Bruker Biotyper MALDI-TOF MS system was able to identify 30.2% (115/381) of the isolates to the species level (score values of ≥2.000) and 49.3% to the genus level (score values of 1.700–1.999). When the identification cutoff value was lowered from ≥2.000 to ≥1.700, the species-level identification rate increased to 79.5% with a slight rise of false identification from 2.6 to 5.0%. From another aspect, a correct species-level identification rate of 89% could be reached by the Bruker Biotyper MALDI-TOF MS system regardless of the score values obtained. The Bruker Biotyper MALDI-TOF MS system had a moderate performance in identification of Aspergillus directly inoculated on solid agar media. Continued expansion of the Bruker Biotyper MALDI-TOF MS database and adoption of alternative cutoff values for interpretation are required to improve the performance of the system for identifying highly diverse species of clinically encountered Aspergillus isolates. PMID:28706514

Evaluation of the Bruker Biotyper Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry System for Identification of Aspergillus Species Directly from Growth on Solid Agar Media.

PubMed

Li, Ying; Wang, He; Zhao, Yu-Pei; Xu, Ying-Chun; Hsueh, Po-Ren

2017-01-01

We evaluated the accuracy of the Bruker Biotyper matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) system at identifying clinical isolates of Aspergillus species that were grown on agar media. A total of 381 non-duplicate Aspergillus isolates representing 21 different Aspergillus species identified by molecular analysis were included in this study. The Bruker Biotyper MALDI-TOF MS system was able to identify 30.2% (115/381) of the isolates to the species level (score values of ≥2.000) and 49.3% to the genus level (score values of 1.700-1.999). When the identification cutoff value was lowered from ≥2.000 to ≥1.700, the species-level identification rate increased to 79.5% with a slight rise of false identification from 2.6 to 5.0%. From another aspect, a correct species-level identification rate of 89% could be reached by the Bruker Biotyper MALDI-TOF MS system regardless of the score values obtained. The Bruker Biotyper MALDI-TOF MS system had a moderate performance in identification of Aspergillus directly inoculated on solid agar media. Continued expansion of the Bruker Biotyper MALDI-TOF MS database and adoption of alternative cutoff values for interpretation are required to improve the performance of the system for identifying highly diverse species of clinically encountered Aspergillus isolates.

Genetic Analysis of Collective Motility of Paenibacillus sp. NAIST15-1

PubMed Central

Kobayashi, Kazuo; Kanesaki, Yu

2016-01-01

Bacteria have developed various motility mechanisms to adapt to a variety of solid surfaces. A rhizosphere isolate, Paenibacillus sp. NAIST15-1, exhibited unusual motility behavior. When spotted onto 1.5% agar media, Paenibacillus sp. formed many colonies, each of which moved around actively at a speed of 3.6 μm/sec. As their density increased, each moving colony began to spiral, finally forming a static round colony. Despite its unusual motility behavior, draft genome sequencing revealed that both the composition and organization of flagellar genes in Paenibacillus sp. were very similar to those in Bacillus subtilis. Disruption of flagellar genes and flagellar stator operons resulted in loss of motility. Paenibacillus sp. showed increased transcription of flagellar genes and hyperflagellation on hard agar media. Thus, increased flagella and their rotation drive Paenibacillus sp. motility. We also identified a large extracellular protein, CmoA, which is conserved only in several Paenibacillus and related species. A cmoA mutant could neither form moving colonies nor move on hard agar media; however, motility was restored by exogenous CmoA. CmoA was located around cells and enveloped cell clusters. Comparison of cellular behavior between the wild type and cmoA mutant indicated that extracellular CmoA is involved in drawing water out of agar media and/or smoothing the cell surface interface. This function of CmoA probably enables Paenibacillus sp. to move on hard agar media. PMID:27764113

Solid colloidal optical wavelength filter

DOEpatents

Alvarez, Joseph L.

1992-01-01

A solid colloidal optical wavelength filter includes a suspension of spheal particles dispersed in a coagulable medium such as a setting plastic. The filter is formed by suspending spherical particles in a coagulable medium; agitating the particles and coagulable medium to produce an emulsion of particles suspended in the coagulable medium; and allowing the coagulable medium and suspended emulsion of particles to cool.

Reflected scatterometry for noninvasive interrogation of bacterial colonies

NASA Astrophysics Data System (ADS)

Kim, Huisung; Doh, Iyll-Joon; Sturgis, Jennifer; Bhunia, Arun K.; Robinson, J. Paul; Bae, Euiwon

2016-10-01

A phenotyping of bacterial colonies on agar plates using forward-scattering diffraction-pattern analysis provided promising classification of several different bacteria such as Salmonella, Vibrio, Listeria, and E. coli. Since the technique is based on forward-scattering phenomena, light transmittance of both the colony and the medium is critical to ensure quality data. However, numerous microorganisms and their growth media allow only limited light penetration and render the forward-scattering measurement a challenging task. For example, yeast, Lactobacillus, mold, and several soil bacteria form colorful and dense colonies that obstruct most of the incoming light passing through them. Moreover, blood agar, which is widely utilized in the clinical field, completely blocks the incident coherent light source used in forward scatterometry. We present a newly designed reflection scatterometer and validation of the resolving power of the instrument. The reflectance-type instrument can acquire backward elastic scatter patterns for both highly opaque media and colonies and has been tested with three different bacterial genera grown on blood agar plates. Cross-validation results show a classification rate above 90% for four genera.

Integration of Stable Isotope and other Mass Spectral Data for Microbial Forensics

NASA Astrophysics Data System (ADS)

Kreuzer-Martin, H. W.; Jarman, K. H.

2008-12-01

The nascent field of microbial forensics requires the development of diverse signatures as indicators of various aspects of the production environment of microorganisms. We have characterized isotopic relationships between Bacillus subtilis ATCC 6051 spores and their growth environment, using as a database the carbon, nitrogen, oxygen and hydrogen stable isotope ratios of a total of 247 separate cultures of spores produced on a total of 32 different culture media. We have analyzed variation within individual samples, between cultures produced in tandem, and between cultures produced in the same medium but at different times in the context of using stable isotope ratios as a signature for sample matching. We have correlated the stable isotope ratios of carbon, nitrogen, oxygen, and hydrogen of growth medium nutrients or water and spores and show examples of how these relationships can be used to exclude nutrient or water samples as possible growth substrates for specific cultures. The power of stable isotope ratio data can be greatly enhanced by combining it with orthogonal datasets that speak to different aspects of an organism's production environment. We developed a Bayesian network that follows the causal relationship from culture medium recipe to spore elemental content as measured by secondary ion mass spectrometry (SIMS), carbon and nitrogen stable isotope ratios, and to the presence of residual agar by electrospray ionization MS (ESI-MS). The network was developed and tested on data from three replicate cultures of B. subtilis ATCC 49760 in broth and agar-containing versions of four different nutrient media. To test the network, data from SIMS analyses of B. subtilis 49760 produced in a different medium, from approximately 200 ESI MS analyses of B. thuringensis ATCC 58890 and B. anthracis Sterne grown in five additional media, and the stable isotope data from the 247 cultures of B. subtilis 6051 spores were used. This network was able to characterize Bacillus spores grown under multiple culture conditions with an error rate of less than 0.07 in characterizing carbon and nitrogen source, addition of metals, and presence of agar, and an error rate of 0.19 in characterizing the culture medium recipe. The integration of multiple analytical techniques allowed us to maximize the amount of information obtained from unknown source microorganisms. The Bayesian network approach allowed us to combine scientific understanding with well established statistical methodologies to characterize a microbe's growth environment without the need for reference signatures. Similar approaches could be applied to data from other scientific disciplines, as well as to other problems of attribution.

Physical structure changes of solid medium by steam explosion sterilization.

PubMed

Zhao, Zhi-Min; Wang, Lan; Chen, Hong-Zhang

2016-03-01

Physical structure changes of solid medium were investigated to reveal effects of steam explosion sterilization on solid-state fermentation (SSF). Results indicated that steam explosion changed the structure of solid medium at both molecular and three-dimensional structural levels, which exposed hydrophilic groups and enlarged pores and cavities. It was interesting to find that pores where capillary water located were the active sites for SSF, due to the close relationship among capillary water relaxation time, specific surface area and fermentation performance. Therefore, steam explosion sterilization increased the effective contact area for microbial cells on solid medium, which contributed to improving SSF performance. Combined with the previous research, mechanisms of SSF improvement by steam explosion sterilization contained both chemical and physical effects. Copyright © 2015 Elsevier Ltd. All rights reserved.

Clonal propagation of Stevia rebaudiana Bertoni by stem-tip culture.

PubMed

Tamura, Y; Nakamura, S; Fukui, H; Tabata, M

1984-10-01

Clonal propagation of Stevia rebaudiana has been established by culturing stem-tips with a few leaf primordia on an agar medium supplemented with a high concentration (10 mg/l) of kinetin. Anatomical examination has suggested that these multiple shoots originate from a number of adventitious buds formed on the margin of the leaf. Innumerable shoots can be obtained by repeating the cycle of multiple-shoot formation from a single stem-tip of Stevia. These shoots produce roots when transferred to a medium containing NAA (0.1 mg/l) without kinetin. The regenerated plantlets can be transplanted to soil.

Somatic embryogenesis and plant regeneration in tissue cultures of sweet potato (Ipomea batatas Poir.).

PubMed

Liu, J R; Cantliffe, D J

1984-06-01

Leaf, shoot-tip, stem, and root explants of sweet potato (Ipomea batatas Poir.) gave rise to two kinds of callus on nutrient agar medium containing 0.5 to 2.0 mg/l 2,4-D. One callus, bright- to pale-yellow, was compact and organized, while the other was dull-yellow and friable. The former callus gave rise to numerous globular and heart-shaped embryoids. When transferred onto hormone-free medium, the embryoids readily developed into a torpedo-shape before germination. The plantlets were transplanted to soil where they flowered and formed storage roots at maturity.

Impact of Salt and Nutrient Content on Biofilm Formation by Vibrio fischeri.

PubMed

Marsden, Anne E; Grudzinski, Kevin; Ondrey, Jakob M; DeLoney-Marino, Cindy R; Visick, Karen L

2017-01-01

Vibrio fischeri, a marine bacterium and symbiont of the Hawaiian bobtail squid Euprymna scolopes, depends on biofilm formation for successful colonization of the squid's symbiotic light organ. Here, we investigated if culture conditions, such as nutrient and salt availability, affect biofilm formation by V. fischeri by testing the formation of wrinkled colonies on solid media. We found that V. fischeri forms colonies with more substantial wrinkling when grown on the nutrient-dense LBS medium containing NaCl relative to those formed on the more nutrient-poor, seawater-salt containing SWT medium. The presence of both tryptone and yeast extract was necessary for the production of "normal" wrinkled colonies; when grown on tryptone alone, the colonies displayed a divoting phenotype and were attached to the agar surface. We also found that the type and concentration of specific seawater salts influenced the timing of biofilm formation. Of the conditions assayed, wrinkled colony formation occurred earliest in LBS(-Tris) media containing 425 mM NaCl, 35 mM MgSO4, and 5 mM CaCl2. Pellicle formation, another measure of biofilm development, was also enhanced in these growth conditions. Therefore, both nutrient and salt availability contribute to V. fischeri biofilm formation. While growth was unaffected, these optimized conditions resulted in increased syp locus expression as measured by a PsypA-lacZ transcriptional reporter. We anticipate these studies will help us understand how the natural environment of V. fischeri affects its ability to form biofilms and, ultimately, colonize E. scolopes.

[Factors inducing transition from growth to dormancy in rhizobacteria Azospirillum brasilense].

PubMed

Kushneruk, M A; Tugarova, A V; Il'chukova, A V; Slavkina, E A; Starichkova, N I; Bogatyrev, V A; Antoniuk, L P

2013-01-01

The factors suppressing division of the cells of the rhizobacterium Azospirillum brasilense and inducing their transition to a dormant state were analyzed. These included the presence of hexylresorcinol or heavy metals (Cu and Co) in the medium, oxygen stress, and transfer of the cells into the physiological saline or phosphate buffer solution. The results were used to develop a protocol for obtaining of uncultured cells of A. brasilense Sp245, a natural symbiont of wheat. The cells lost their ability to grow on synthetic agar medium, but could revert to growth when incubated in freshly prepared liquid medium. Needle-shaped crystals differing from struvite, which has been previously reported for this strain, were found in the dormant culture of A. brasilense Sp245.

Media for identification of Gibberella zeae and production of F-2-(Zearalenone).

PubMed

Bacon, C W; Robbins, J D; Porter, J K

1977-02-01

Media are described for the isolaton of Fusarium graminearum in the perithecial state, Gibberella zeae, and for the production of F-2 (zearalenone) by Fusarium species. On soil extract-corn meal agar isolated medium, G. Zeae produced perithecia in 9 to 14 days under a 12-h photoperiod. Species of Fusarium were screened for F-2 production on a liquid medium. From strains that produced F-2, the yields, from stationary cultures of G. zeae and F. culmorum after 12 days of incubation, ranged from 22 to 86 mg/liter. Three strains produced no F-2. Glumatic acid, starch, yeast extract,and the proper ratio of medium volume-to-flask volume were necessary for F-2 synthesis.

Two rapid pigmentation tests for identification of Cryptococcus neoformans.

PubMed Central

Kaufmann, C S; Merz, W G

1982-01-01

Two tests were developed for the rapid identification of Cryptococcus neoformans based on pigment produced by the organism's phenoloxidase activity. Caffeic acid was incorporated into cornmeal agar, a medium used routinely for yeast identification. When tested on this medium, only C. neoformans isolates produced brown pigment. All other yeasts maintained their normal morphology and did not produce the reaction product. A non-medium-based test was developed for same-day identification of C. neoformans isolates. Paper strips saturated with a buffered L-beta-3,4-dihydroxyphenylalanine-ferric citrate solution were inoculated with isolates and incubated at 37 degrees C. Pigment production occurred only with C. neoformans isolates, many within 60 to 90 min. All other yeasts remained negative. PMID:7040452

Volatile profiles of toxigenic and non-toxigenic Aspergillus flavus using SPME for solid phase extraction

USDA-ARS?s Scientific Manuscript database

Toxigenic and atoxigenic strains of Aspergillus flavus were grown on potato dextrose agar (PDA) and wetted sterile, cracked corn for 21 and 14 days, respectively. Volatile compounds produced by A. flavus, as well as those present in the PDA controls and sterile cracked corn, were collected using sol...

Development of a Novel Screening Method for the Isolation of “Cronobacter” spp. (Enterobacter sakazakii)▿

PubMed Central

Iversen, Carol; Druggan, Patrick; Schumacher, Sandra; Lehner, Angelika; Feer, Claudia; Gschwend, Karl; Joosten, Han; Stephan, Roger

2008-01-01

A differential medium, “Cronobacter” screening broth, has been designed to complement agars based on hydrolysis of chromogenic α-glucopyranoside substrates. The broth was evaluated using 329 Enterobacteriaceae strains (229 target isolates), spiked/naturally contaminated samples, and a parallel comparison with current methods for raw materials, line/end products, and factory environment samples. PMID:18310415

An ability of endophytic bacteria from nutgrass (cyperus rotundus) from lafau beach of north nias in producing indole acetic acid and in solubilizing phosphate

NASA Astrophysics Data System (ADS)

Zega, Atriani; Suryanto, Dwi; Yurnaliza

2018-03-01

Endophytic bacteria have taken much attention for their potency to promote plant growth. This study was aimed to isolate endophytic bacteria from nutgrass (Cyperus rotundus) and to examine their potency in producing indole acetic acid (IAA) and in solubilizing phosphate. Isolation of endophytic bacteria was done by slicing and sterilizing root, stem, and leaf sample surface with alcohol 70% and sodium hypochlorite 2%, followed by incubation of the sliced samples in nutrient agar medium. Morphological characterization and simple biochemical tests were performed on bacterial isolates. All bacterial isolates were examined for their ability to produce indole acetic acid and to solubilize phosphate. Three isolates (AZ5, AZ12 and AZ6) out of fifteen indicated the ability to produce indole acetic acid and to solubilize phosphate. IAA producing test using spectrophotometry method showed that AZ5, AZ12,and AZ6 produce more IAA with concentration of 49,91, 48,18, and 44,45 ppm, respectively. Phosphate solubilizing test using Pikovskaya agar medium showed that the three isolates were able to solubilize phosphate with index of 6.27, 3,31, and 3.41 respectively.

Waving and skewing: how gravity and the surface of growth media affect root development in Arabidopsis.

PubMed

Oliva, Michele; Dunand, Christophe

2007-01-01

Arabidopsis seedlings growing on inclined agar surfaces exhibit characteristic root behaviours called 'waving' and 'skewing': the former consists of a series of undulations, whereas the latter is a deviation from the direction of gravity. Even though the precise basis of these growth patterns is not well understood, both gravity and the contact between the medium and the root are considered to be the major players that result in these processes. The influence of these forces on root surface-dependent behaviours can be verified by growing seedlings at different gel pitches: plants growing on vertical plates present roots with slight waving and skewing when compared with seedlings grown on plates held at minor angles of < 90 degrees . However, other factors are thought to modulate root growth on agar; for instance, it has been demonstrated that the presence and concentration of certain compounds in the medium (such as sucrose) and of drugs able to modify the plant cell cytoskeleton also affect skewing and waving. The recent discovery of an active role of ethylene on surface-dependent root behaviour, and the finding of new mutants showing anomalous growth, pave the way for a more detailed description of these phenomena.

A supplemented soft agar chemotaxis assay demonstrates the Helicobacter pylori chemotactic response to zinc and nickel

PubMed Central

Sanders, Lisa; Andermann, Tessa M.

2013-01-01

Directed motility, or chemotaxis, is required for Helicobacter pylori to establish infection in the stomach, although the full repertoire of this bacterium’s chemotactic responses is not yet known. Here we report that H. pylori responds to zinc as an attractant and nickel as a repellent. To reach this conclusion, we employed both a temporal chemotaxis assay based on bacterial reversals and a supplemented soft agar spatial assay. We refined the temporal assay using a previously described chemorepellent, acid, and found that H. pylori requires rich media with serum to maintain optimal swimming motility. Surprisingly, we found that some strains respond to acid as an attractant, and that the TlpC chemoreceptor correlated with whether acid was sensed as an attractant or repellent. Using this same assay, we detected weak repellent responses to nickel and copper, and a varied response to zinc. We thus developed an alternative spatial chemotactic assay called the supplemented soft agar assay, which utilizes soft agar medium supplemented with the test compound. With Escherichia coli, the attractant serine slowed overall bacterial migration, while the repellent nickel increased the speed of overall migration. In H. pylori we detected slowed migration with doubled tryptone media, as well as zinc, consistent with an attractant response. In contrast, nickel increased migration, consistent with repulsion. PMID:23139399

Uropathogenic Escherichia coli Metabolite-Dependent Quiescence and Persistence May Explain Antibiotic Tolerance during Urinary Tract Infection

PubMed Central

Leatham-Jensen, Mary P.; Mokszycki, Matthew E.; Rowley, David C.; Deering, Robert; Camberg, Jodi L.; Sokurenko, Evgeni V.; Tchesnokova, Veronika L.; Frimodt-Møller, Jakob; Leth Nielsen, Karen; Sun, Gongqin

2016-01-01

ABSTRACT In the present study, it is shown that although Escherichia coli CFT073, a human uropathogenic (UPEC) strain, grows in liquid glucose M9 minimal medium, it fails to grow on glucose M9 minimal medium agar plates seeded with ≤106 CFU. The cells on glucose plates appear to be in a “quiescent” state that can be prevented by various combinations of lysine, methionine, and tyrosine. Moreover, the quiescent state is characteristic of ~80% of E. coli phylogenetic group B2 multilocus sequence type 73 strains, as well as 22.5% of randomly selected UPEC strains isolated from community-acquired urinary tract infections in Denmark. In addition, E. coli CFT073 quiescence is not limited to glucose but occurs on agar plates containing a number of other sugars and acetate as sole carbon sources. It is also shown that a number of E. coli CFT073 mini-Tn5 metabolic mutants (gnd, gdhA, pykF, sdhA, and zwf) are nonquiescent on glucose M9 minimal agar plates and that quiescence requires a complete oxidative tricarboxylic acid (TCA) cycle. In addition, evidence is presented that, although E. coli CFT073 quiescence and persistence in the presence of ampicillin are alike in that both require a complete oxidative TCA cycle and each can be prevented by amino acids, E. coli CFT073 quiescence occurs in the presence or absence of a functional rpoS gene, whereas maximal persistence requires a nonfunctional rpoS. Our results suggest that interventions targeting specific central metabolic pathways may mitigate UPEC infections by interfering with quiescence and persistence. IMPORTANCE Recurrent urinary tract infections (UTIs) affect 10 to 40% of women. In up to 77% of those cases, the recurrent infections are caused by the same uropathogenic E. coli (UPEC) strain that caused the initial infection. Upon infection of urothelial transitional cells in the bladder, UPEC appear to enter a nongrowing quiescent intracellular state that is thought to serve as a reservoir responsible for recurrent UTIs. Here, we report that many UPEC strains enter a quiescent state when ≤106 CFU are seeded on glucose M9 minimal medium agar plates and show that mutations in several genes involved in central carbon metabolism prevent quiescence, as well as persistence, possibly identifying metabolic pathways involved in UPEC quiescence and persistence in vivo. PMID:27303698

Rapid spot test for the determination of esculin hydrolysis.

PubMed

Edberg, S C; Gam, K; Bottenbley, C J; Singer, J M

1976-08-01

Esculin hydrolysis is a useful test in the differentiation of both gram-positive and gram-negative bacteria covering a wide spectrum of aerobes, facultative anaerobes, and anaerobes. Commonly utilized methods require a minimum of 18 h of incubation in broth or agar medium and utilize the production of a brown-black compound, due to the combination of ferric ions with the hydrolysis product esculetin, as indicator. A procedure is presented that requires 15 to 30 min for completion and utilizes fluorescence loss as the indicator of hydrolysis. Esculin fluoresces at 366 nm, whereas the hydrolysis product esculetin does not. Over 1,400 strains of gram-positive and gram-negative bacteria were tested. There was 98.4% of correlation between the spot test and esculin broth and 97% correlation with the bile-esculin agar.

Implementation of the Thin Layer Agar Method for Diagnosis of Smear-Negative Pulmonary Tuberculosis in a Setting with a High Prevalence of Human Immunodeficiency Virus Infection in Homa Bay, Kenya▿ †

PubMed Central

Martin, Anandi; Munga Waweru, Peter; Babu Okatch, Fred; Amondi Ouma, Naureen; Bonte, Laurence; Varaine, Francis; Portaels, Françoise

2009-01-01

The objective of this study was to evaluate the performance of a low-cost method, the thin layer agar (TLA) method, for the diagnosis of smear-negative patients. This prospective study was performed in Homa Bay District Hospital in Kenya. Out of 1,584 smear-negative sputum samples, 212 (13.5%) were positive by culture in Löwenstein-Jensen medium (LJ) and 220 (14%) were positive by the TLA method. The sensitivities of LJ and TLA were 71% and 74%, respectively. TLA could become an affordable method for the diagnosis of smear-negative tuberculosis in resource-limited settings, with results available within 2 weeks. PMID:19494065

Carboxydocella sporoproducens sp. nov., a novel anaerobic CO-utilizing/H2-producing thermophilic bacterium from a Kamchatka hot spring.

PubMed

Slepova, Tatiana V; Sokolova, Tatyana G; Lysenko, Anatoly M; Tourova, Tatyana P; Kolganova, Tatyana V; Kamzolkina, Olga V; Karpov, Genady A; Bonch-Osmolovskaya, Elizaveta A

2006-04-01

A novel anaerobic, thermophilic, CO-utilizing bacterium, strain KarT, was isolated from a hot spring of Karymskoe Lake, Kamchatka Peninsula. The cells of the novel isolate were Gram-positive, spore-forming, short rods. The bacterium grew chemolithoautotrophically on CO, producing equimolar quantities of H2 and CO2 (according to the equation CO + H2O --> CO2 + H2), and in the absence of CO, under N2 in the gas phase, chemoorganoheterotrophically with yeast extract, sucrose or pyruvate. Growth was observed in the temperature range 50-70 degrees C, with an optimum at 60 degrees C, and in the pH range 6.2-8.0, with an optimum at pH 6.8. The micro-organism did not grow on solid media; it was able to grow only in semi-solid medium containing 0.5 % agar. The generation time under optimal conditions for chemolithoautotrophic growth was 1 h. The G+C content of the DNA was 46.5+/-1 mol%. Growth was completely inhibited by penicillin, novobiocin, streptomycin, kanamycin and neomycin. Analysis of the 16S rRNA gene sequence showed that the isolate should be assigned to the genus Carboxydocella. On the basis of the results of DNA-DNA hybridization and morphological and physiological analyses, strain KarT represents a novel species of the genus Carboxydocella, for which the name Carboxydocella sporoproducens sp. nov. is proposed. The type strain is KarT (=DSM 16521T = VKM B-2358T).

[Inhibitory effects of butyl alcohol extract of Baitouweng decoction on yeast-to-hyphae transition of Candida albicans isolates from VVC in alkaline pH environment].

PubMed

Zhang, Meng-xiang; Xia, Dan; Shi, Gao-xiang; Shao, Jing; Wang, Tian-ming; Tang, Chuan-chao; Wang, Chang-zhong

2015-02-01

To investigate the effects of butyl alcohol extract of Baitouweng decoction ( BAEB) on yeast-to-hyphae transition of Candida albicans isolates from vulvovaginal candidiasis (VVC) in alkaline pH. Serial 2-fold dilution assay was used to determine the minimal inhibitory concentrations (MICs) of Baitouweng decoction extracts against C. albicans isolates from VVC, XTT assay was applied to determine the metabolic activity of C. albicans hypha treated by BAEB for 6 h. The morphological change of C. albicans treated by BAEB was inspected at different pH by inverted microscope, fluorescence microscope, scanning electron microscopy (SEM). Solid agar plate and semi-solid agar were utilized to evaluate colony morphology and invasive growth of C. albicans, respectively. Quantitative Real-time PCR (qRT-PCR) was adopted to observe the expressions of hyphae-specific genes including HWP1, ALS3, CSH1, SUN41 and CaPDE2. The MIC of BAEB against C. albicans is less than that of other extracts; hyphae grow best at pH 8. 0; 512 mg · L(-1) and 1,024 mg · L(-1) BAEB could inhibit formation of hyphae and influence colony morphology. When treated by 512 mg · L(-1) and 1,024 mg · L(-1) BAEB, the colonies became smooth; while by 0 and 256 mg · L(-1) BAEB, the colonies became wrinkled. In semi-solid agar, the length of hyphae decreased steadily as the concentration of BAEB lowered. The expression of HWP1, ALS3, CSHl, SUN41 were downregulated by 5.12, 4.26, 3.2 and 2.74 folds, and CaPDE2 was upregulated by 2.38 fold. BAEB could inhibit yeast-to-hyphae transition of C. albicans isolates from VVC in alkaline pH.

Regulation of coal polymer degradation by fungi. Eighth quarterly report, [April--June 1996

DOE Office of Scientific and Technical Information (OSTI.GOV)

Irvine, R.L.; Bumpus, J.A.

1996-07-28

This project addresses the solubilization of low-rank coal (leonardite) by lignin degrading fungi. During this reporting period efforts were focused on determining the effect of pH on coal solubilization by oxalate ion and other biologically important compounds that might function as metal chelators, on the role of laccase in coal solubilization and metabolism, on decolorization of soluble coal macromolecule by Phanerochaete chrysosporium and T. versicolor in solid agar media, and on solubilization of coal in slurry cultures and solid phase reactors.

Effect of nitrogen source concentration on curdlan production by Agrobacterium sp. ATCC 31749 grown on prairie cordgrass hydrolysates.

PubMed

West, Thomas P

2016-01-01

The effect of nitrogen source concentration on the production of the polysaccharide curdlan by the bacterium Agrobacterium sp. ATCC 31749 from hydrolysates of prairie cordgrass was examined. The highest curdlan concentrations were produced by ATCC 31749 when grown on a medium containing a solids-only hydrolysate and the nitrogen source ammonium phosphate (2.2 mM) or on a medium containing a complete hydrolysate and 3.3 mM ammonium phosphate. The latter medium sustained a higher level of bacterial curdlan production than the former medium after 144 hr. Biomass production by ATCC 31749 was highest after 144 hr when grown on a medium containing a solids-only hydrolysate and 2.2 or 8.7 mM ammonium phosphate. On the medium containing the complete hydrolysate, biomass production by ATCC 31749 was highest after 144 hr when 3.3 mM ammonium phosphate was present. Bacterial biomass production after 144 hr was greater on the complete hydrolysate medium compared to the solids-only hydrolysate medium. Curdlan yield produced by ATCC 31749 after 144 hr from the complete hydrolysate medium containing 3.3 mM ammonium phosphate was higher than from the solids-only hydrolysate medium containing 2.2 mM ammonium phosphate.

Use of an acidophilic yeast strain to enable the growth of leaching bacteria on solid media.

PubMed

Ngom, Baba; Liang, Yili; Liu, Yi; Yin, Huaqun; Liu, Xueduan

2015-03-01

In this study, a Candida digboiensis strain was isolated from a heap leaching plant in Zambia and used in double-layer agar plate to efficiently isolate and purify leaching bacteria. Unlike Acidiphilium sp., the yeast strain was tetrathionate tolerant and could metabolize a great range of organic compounds including organic acids. These properties allowed the yeast strain to enable and fasten the growth of iron and sulfur oxidizers on double-layer agar plate. The isolates were identified as Acidithiobacillus ferrooxidans FOX1, Leptospirillun ferriphilum BN, and Acidithiobacillus thiooxidans ZMB. These three leaching bacteria were inhibited by organic acids such as acetic and propionic acids; however, their activities were enhanced by Candida digboiensis NB under dissolved organic matter stress.

Comparison of bacteriophage and enteric virus removal in pilot scale activated sludge plants.

PubMed

Arraj, A; Bohatier, J; Laveran, H; Traore, O

2005-01-01

The aim of this experimental study was to determine comparatively the removal of two types of bacteriophages, a somatic coliphage and an F-specific RNA phage and of three types of enteric viruses, hepatitis A virus (HAV), poliovirus and rotavirus during sewage treatment by activated sludge using laboratory pilot plants. The cultivable simian rotavirus SA11, the HAV HM 175/18f cytopathic strain and poliovirus were quantified by cell culture. The bacteriophages were quantified by plaque formation on the host bacterium in agar medium. In each experiment, two pilots simulating full-scale activated sludge plants were inoculated with viruses at known concentrations, and mixed liquor and effluent samples were analysed regularly. In the mixed liquor, liquid and solid fractions were analysed separately. The viral behaviour in both the liquid and solid phases was similar between pilots of each experiment. Viral concentrations decreased rapidly following viral injection in the pilots. Ten minutes after the injections, viral concentrations in the liquid phase had decreased from 1.0 +/- 0.4 log to 2.2 +/- 0.3 log. Poliovirus and HAV were predominantly adsorbed on the solid matters of the mixed liquor while rotavirus was not detectable in the solid phase. In our model, the estimated mean log viral reductions after 3-day experiment were 9.2 +/- 0.4 for rotavirus, 6.6 +/- 2.4 for poliovirus, 5.9 +/- 3.5 for HAV, 3.2 +/- 1.2 for MS2 and 2.3 +/- 0.5 for PhiX174. This study demonstrates that the pilots are useful models to assess the removal of infectious enteric viruses and bacteriophages by activated sludge treatment. Our results show the efficacy of the activated sludge treatment on the five viruses and suggest that coliphages could be an acceptable indicator of viral removal in this treatment system.

Multiplex detection of nine food-borne pathogens by mPCR and capillary electrophoresis after using a universal pre-enrichment medium.

PubMed

Villamizar-Rodríguez, Germán; Fernández, Javier; Marín, Laura; Muñiz, Juan; González, Isabel; Lombó, Felipe

2015-01-01

Routine microbiological quality analyses in food samples require, in some cases, an initial incubation in pre-enrichment medium. This is necessary in order to ensure that small amounts of pathogenic strains are going to be detected. In this work, a universal pre-enrichment medium has been developed for the simultaneous growth of Bacillus cereus, Campylobacter jejuni, Clostridium perfringens, Cronobacter sakazakii, Escherichia coli, Enterobacteriaceae family (38 species, 27 genera), Listeria monocytogenes, Staphylococcus aureus, Salmonella spp. (two species, 13 strains). Growth confirmation for all these species was achieved in all cases, with excellent enrichments. This was confirmed by plating on the corresponding selective agar media for each bacterium. This GVUM universal pre-enrichment medium could be useful in food microbiological analyses, where different pathogenic bacteria must be detected after a pre-enrichment step. Following, a mPCR reaction for detection of all these pathogens was developed, after designing a set of nine oligonucleotide pairs from specific genetic targets on gDNA from each of these bacteria, covering all available strains already sequenced in GenBank for each pathogen type. The detection limits have been 1 Genome Equivalent (GE), with the exception of the Fam. Enterobacteriaceae (5 GEs). We obtained amplification for all targets (from 70 to 251 bp, depending on the bacteria type), showing the capability of this method to detect the most important industrial and sanitary food-borne pathogens from a universal pre-enrichment medium. This method includes an initial pre-enrichment step (18 h), followed by a mPCR (2 h) and a capillary electrophoresis (30 min); avoiding the tedious and long lasting growing on solid media required in traditional analysis (1-4 days, depending on the specific pathogen and verification procedure). An external testing of this method was conducted in order to compare classical and mPCR methods. This evaluation was carried out on five types of food matrices (meat, dairy products, prepared foods, canned fish, and pastry products), which were artificially contaminated with each one of the microorganisms, demonstrating the equivalence between both methods (coincidence percentages between both methods ranged from 78 to 92%).

Multiplex detection of nine food-borne pathogens by mPCR and capillary electrophoresis after using a universal pre-enrichment medium

PubMed Central

Villamizar-Rodríguez, Germán; Fernández, Javier; Marín, Laura; Muñiz, Juan; González, Isabel; Lombó, Felipe

2015-01-01

Routine microbiological quality analyses in food samples require, in some cases, an initial incubation in pre-enrichment medium. This is necessary in order to ensure that small amounts of pathogenic strains are going to be detected. In this work, a universal pre-enrichment medium has been developed for the simultaneous growth of Bacillus cereus, Campylobacter jejuni, Clostridium perfringens, Cronobacter sakazakii, Escherichia coli, Enterobacteriaceae family (38 species, 27 genera), Listeria monocytogenes, Staphylococcus aureus, Salmonella spp. (two species, 13 strains). Growth confirmation for all these species was achieved in all cases, with excellent enrichments. This was confirmed by plating on the corresponding selective agar media for each bacterium. This GVUM universal pre-enrichment medium could be useful in food microbiological analyses, where different pathogenic bacteria must be detected after a pre-enrichment step. Following, a mPCR reaction for detection of all these pathogens was developed, after designing a set of nine oligonucleotide pairs from specific genetic targets on gDNA from each of these bacteria, covering all available strains already sequenced in GenBank for each pathogen type. The detection limits have been 1 Genome Equivalent (GE), with the exception of the Fam. Enterobacteriaceae (5 GEs). We obtained amplification for all targets (from 70 to 251 bp, depending on the bacteria type), showing the capability of this method to detect the most important industrial and sanitary food-borne pathogens from a universal pre-enrichment medium. This method includes an initial pre-enrichment step (18 h), followed by a mPCR (2 h) and a capillary electrophoresis (30 min); avoiding the tedious and long lasting growing on solid media required in traditional analysis (1–4 days, depending on the specific pathogen and verification procedure). An external testing of this method was conducted in order to compare classical and mPCR methods. This evaluation was carried out on five types of food matrices (meat, dairy products, prepared foods, canned fish, and pastry products), which were artificially contaminated with each one of the microorganisms, demonstrating the equivalence between both methods (coincidence percentages between both methods ranged from 78 to 92%). PMID:26579100

Efficient callus formation and plant regeneration of goosegrass [Eleusine indica (L.) Gaertn.].

PubMed

Yemets, A I; Klimkina, L A; Tarassenko, L V; Blume, Y B

2003-02-01

Efficient methods in totipotent callus formation, cell suspension culture establishment and whole-plant regeneration have been developed for the goosegrass [ Eleusine indica (L.) Gaertn.] and its dinitroaniline-resistant biotypes. The optimum medium for inducing morphogenic calli consisted of N6 basal salts and B5 vitamins supplemented with 1-2 mg l(-1) 2,4-dichlorophenoxyacetic acid (2,4-D), 2 mg l(-1) glycine, 100 mg l(-1) asparagine, 100 mg l(-1) casein hydrolysate, 30 g l(-1) sucrose and 0.6% agar, pH 5.7. The presence of organogenic and embryogenic structures in these calli was histologically documented. Cell suspension cultures derived from young calli were established in a liquid medium with the same composition. Morphogenic structures of direct shoots and somatic embryos were grown into rooted plantlets on medium containing MS basal salts, B5 vitamins, 1 mg l(-1) kinetin (Kn) and 0.1 mg l(-1) indole-3-acetic acid (IAA), 3% sucrose, 0.6% agar, pH 5.7. Calli derived from the R-biotype of E. indica possessed a high resistance to trifluralin (dinitroaniline herbicide) and cross-resistance to a structurally non-related herbicide, amiprophosmethyl (phosphorothioamidate herbicide), as did the original resistant plants. Embryogenic cell suspension culture was a better source of E. indica protoplasts than callus or mesophyll tissue. The enzyme solution containing 1.5% cellulase Onozuka R-10, 0.5% driselase, 1% pectolyase Y-23, 0.5% hemicellulase and N(6) mineral salts with an additional 0.2 M KCl and 0.1 M CaCl(2) (pH 5.4-5.5) was used for protoplast isolation. The purified protoplasts were cultivated in KM8p liquid medium supplemented with 2 mg l(-1) 2,4-D and 0.2 mg l(-1) Kn.

[Isolation and characterization of vaginal lactobacilli producing hydrogen peroxide].

PubMed

Pashaian, M M; Oganesian, G G

2011-01-01

Isolation and characteristics of vaginal lactobacilli that actively generate H2O2 and have high antagonistic activity. Staphylococcus aureus 8956, Escherichia coli 8852, Klebsiella pneumoniae 8795 and Candida albicans 5646 were used as target-strains. Skim milk and MRS medium were used for lactobacilli isolation and cultivation. Antagonism was studied in complete agar and Saburo medium. Merckoquant peroxide test (Merck) stripes were used for the determination of H2O2. Antibacterial activity was determined by diffusion into agar. Specific culture growth rate was determined by conventional method, acidification of the culture medium--by pH-meter. 12 strains were isolated from vaginal smears of healthy women. These strains have an ability to ferment milk among which a highly active H2O2 producer was isolated and attributed to Lactobacillus delbrueckii by the results of 16S rRNA and alpha-subunit RNA polymerase gene sequence analysis (16S rDNA and rpoA genes are registered in GenBank, numbers HQ379171 and HQ379180 respectively). L. delbrueckii MH-10 bacterial cells were characterized by specific growth speed 1.26 per hour, reaching a maximum titer of 2 x 10(9) PFU/ml with lowering medium pH to 4.0. Under aerated conditions H2O2 concentration reached 100 microg/ml or more. L. delbrueckii MH-10 has high antibacterial activity against S. aureus, E. coli, K. pneumoniae. L. delbrueckii MH-10 isolate is an active H2O2 producer, has high growth speed and broad antibacterial activity spectrum, is a perspective candidate for the development of probiotic preparation for the prophylaxis and therapy of vaginoses.

Characterization of rhizosphere bacteria for control of phytopathogenic fungi of tomato.

PubMed

Pastor, Nicolás; Carlier, Evelin; Andrés, Javier; Rosas, Susana B; Rovera, Marisa

2012-03-01

Fluorescent Pseudomonas spp., isolated from rhizosphere soil of tomato and pepper plants, were evaluated in vitro as potential antagonists of fungal pathogens. Strains were characterized using the API 20NE biochemical system, and tested against the causal agents of stem canker and leaf blight (Alternaria alternata f. sp. lycopersici), southern blight (Sclerotium rolfsii Sacc.), and root rot (Fusarium solani). To this end, dual culture antagonism assays were carried out on 25% Tryptic Soy Agar, King B medium, and Potato Dextrose Agar to determine the effect of the strains on mycelial growth of the pathogens. The effect of two concentrations of FeCl(3) on antagonism against Alternaria alternata f. sp. lycopersici was also tested. In addition, strains were screened for ability to produce exoenzymes and siderophores. Finally, the selected Pseudomonas strain, PCI2, was evaluated for effect on tomato seedling development and as a potential candidate for controlling tomato damping-off caused by Sclerotium rolfsii Sacc., under growth chamber conditions. All strains significantly inhibited Alternaria alternata f. sp. lycopersici, particularly in 25% TSA medium. Antagonistic effect on Sclerotium rolfsii Sacc. and Fusarium solani was greater on King B medium. Protease was produced by 30% of the strains, but no strains produced cellulase or chitinase. Growth chamber studies resulted in significant increases in plant stand as well as in root dry weight. PCI2 was able to establish and survive in tomato plants rhizosphere after 40 days following planting of bacterized seeds. Copyright © 2011 Elsevier Ltd. All rights reserved.

Health significance and occurrence of injured bacteria in drinking water

NASA Technical Reports Server (NTRS)

McFeters, G. A.; LeChevallier, M. W.; Singh, A.; Kippin, J. S.

1986-01-01

Enteropathogenic and indicator bacteria become injured in drinking water with exposure to sublethal levels of various biological, chemical and physical factors. One manifestation of this injury is the inability to grow and form colonies on selective media containing surfactants. The resulting underestimation of indicator bacteria can lead to a false estimation of water potability. m-T7 medium was developed specifically for the recovery of injured coliforms (both "total" and fecal) in drinking water. The m-T7 method was used to survey operating drinking water treatment and distribution systems for the presence of injured coliforms that were undetected with currently used media. The mean recovery with m-Endo LES medium was less than 1/100 ml while it ranged between 6 and 68/100ml with m-T7 agar. The majority of samples giving positive results with m-T7 medium yielded no detectable coliforms with m-Endo LES agar. Over 95% of the coliform bacteria in these samples were injured. Laboratory experiments were also done to ascribe the virulence of injured waterborne pathogens. Enteropathogens including Salmonella typhimurium, Yersinia enterocolitica and Shigella spp. required up to 20 times the chlorine levels to produce the same injury in enterotoxigenic Escherichia coli (ETEC) and nonpathogenic coliforms. Similar results were seen with Y. enterocolitica exposed to copper. The recovery of ETEC was followed by delayed enterotoxin production, both in vitro and in the gut of experimental animals. This indicates that injured waterborne enteropathogenic bacteria can be virulent.

Single-cell analysis of S. cerevisiae growth recovery after a sublethal heat-stress applied during an alcoholic fermentation.

PubMed

Tibayrenc, Pierre; Preziosi-Belloy, Laurence; Ghommidh, Charles

2011-06-01

Interest in bioethanol production has experienced a resurgence in the last few years. Poor temperature control in industrial fermentation tanks exposes the yeast cells used for this production to intermittent heat stress which impairs fermentation efficiency. Therefore, there is a need for yeast strains with improved tolerance, able to recover from such temperature variations. Accordingly, this paper reports the development of methods for the characterization of Saccharomyces cerevisiae growth recovery after a sublethal heat stress. Single-cell measurements were carried out in order to detect cell-to-cell variability. Alcoholic batch fermentations were performed on a defined medium in a 2 l instrumented bioreactor. A rapid temperature shift from 33 to 43 °C was applied when ethanol concentration reached 50 g l⁻¹. Samples were collected at different times after the temperature shift. Single cell growth capability, lag-time and initial growth rate were determined by monitoring the growth of a statistically significant number of cells after agar medium plating. The rapid temperature shift resulted in an immediate arrest of growth and triggered a progressive loss of cultivability from 100 to 0.0001% within 8 h. Heat-injured cells were able to recover their growth capability on agar medium after a lag phase. Lag-time was longer and more widely distributed as the time of heat exposure increased. Thus, lag-time distribution gives an insight into strain sensitivity to heat-stress, and could be helpful for the selection of yeast strains of technological interest.

Effect of minerals on accumulation of Cs by fungus Saccaromyces cerevisiae.

PubMed

Ohnuki, Toshihiko; Sakamoto, Fuminori; Yamasaki, Shinya; Kozai, Naofumi; Shiotsu, Hiroyuki; Utsunomiya, Satoshi; Watanabe, Naoko; Kozaki, Tamotsu

2015-06-01

The accumulation of Cs by unicellular fungus of Saccharomyces cerevisiae in the presence of minerals has been studied to elucidate the role of microorganisms in the migration of radioactive Cs in the environment. Two different types of experiments were employed: experiments using stable Cs to examine the effect of a carbon source on the accumulation of Cs, and accumulation experiments of radioactive Cs from agar medium containing (137)Cs and zeolite, vermiculite, phlogopite, smectite, mica, or illite as mineral supplements. In the former type of experiments, the Cs-accumulated cells were analyzed by scanning electron microscopy equipped with energy dispersive X-ray analysis (SEM-EDS). In the latter type, the radioactivity in the yeast cells was measured by an autoradiography technique. When a carbon source was present, higher amounts of Cs accumulated in the cells than in the resting condition without a carbon source. Analyses with SEM-EDS showed that no mineral formed on the cell surface. These results indicate that the yeast cells accumulate Cs by adsorption on the cell surface and intracellular accumulation. In the presence of minerals in the agar medium, the radioactivity in the yeast cells was in the order of mica > smectite, illite > vermiculite, phlogopite, zeolite. This order is inversely correlated to the ratio of the concentration of radioactive Cs between the minerals and the medium solution. These results strongly suggest that the yeast accumulates radioactive Cs competitively with minerals. Copyright © 2015 Elsevier Ltd. All rights reserved.

Esculin-based medium for isolation and identification of Cryptococcus neoformans.

PubMed Central

Edberg, S C; Chaskes, S J; Alture-Werber, E; Singer, J M

1980-01-01

A simple medium was developed, using esculin as the substrate, for the isolation and identification of Cryptococcus neoformans. C. neoformans produced a brown-black pigment on the medium; all other yeasts produced no pigment or were light yellow. Esculin is beta-glucose-6,7-dihydroxycoumarin. C. neoformans produced pigment because the 6,7-dihydroxycoumarin component of the esculin molecule was converted to a melanin-like pigment. We think the reaction was similar to the conversion of diphenols, aminophenols, and diaminobenzenes to melanin. Laboratory studies with isolates of C. neoformans, C. albidus, C. luteolus, and C. terreus and representatives of the genera Candida, Torulopsis, Geotrichum, and Rhodotorula, plus environmental field studies, demonstrated that over 95% of C. neoformans isolates were correctly identified, whereas all other fungi were excluded. Esculin agar was a sensitive, specific medium for the isolation and identification of C. neoformans. It was inexpensive and had a long storage life. Images PMID:7012169

The growth of Treponema hyodysenteriae and other porcine intestinal spirochaetes in a liquid medium.

PubMed

Lemcke, R M; Bew, J; Burrows, M R; Lysons, R J

1979-05-01

A new simple method for the preparation of a liquid medium containing rabbit serum for the propagation of Treponema hyodysenteriae and other porcine intestinal spirochaetes is described. The medium, when dispensed in shallow layers and sealed under 10 per cent CO2 in nitrogen, had a redox potential not greater than -125mV and an initial pH of about 6.9 when buffered with bicarbonate. Growth of T hyodysenteriae developed more rapidly and viable counts reached higher levels at 42 degrees C than at 37 degrees C. Viable counts increased at least 10,000-fold after two to five days' incubation, depending on the temperature. Growth could be initiated from small inocula that failed to produce colonies on blood agar. Using a 1 per cent inoculum, the medium supported the growth of two strains of T hyodysenteriae through 10 serial passages.

Antagonistic potential of Pseudomonas graminis 49M against Erwinia amylovora, the causal agent of fire blight.

PubMed

Mikiciński, Artur; Sobiczewski, Piotr; Puławska, Joanna; Malusa, Eligio

2016-08-01

In a previous study (Mikiciński et al. in Eur J Plant Pathol, doi: 10.1007/s10658-015-0837-y , 2015), we described the characterization of novel strain 49M of Pseudomonas graminis, isolated from the phyllosphere of apple trees in Poland showing a good protective activity against fire blight on different organs of host plants. We now report investigations to clarify the basis for this activity. Strain 49M was found to produce siderophores on a medium containing complex CAS-Fe(3+) and HDTMA, but was not able to produce N-acyl homoserine lactones (AHLs). Moreover, it formed a biofilm on polystyrene and polyvinyl chloride (PVC) surfaces. Strain 49M gave a positive reaction in PCR with primers complementary to gacA, the regulatory gene influencing the production of several secondary metabolites including antibiotics. The genes prnD (encoding pyrrolnitrin), pltC, pltB (pyoluteorin), phlD (2,4-diacetyl-phloroglucinol) and phzC as well as phzD (and their homologs phzF and phzA encoding phenazine), described for antagonistic fluorescent pseudomonads, however, were not detected. Research into the biotic relationship between strain 49M and Erwinia amylovora strain Ea659 on five microbiological media showed that this strain clearly inhibited the growth of the pathogen on King's B and nutrient agar with glycerol media, to a very small extent on nutrient agar with sucrose, and not at all on Luria-Bertani agar. On medium 925, strain 49M even stimulated E. amylovora growth. The addition of ferric chloride to King's B resulted in the loss of its inhibitory ability. Testing the survival of 49M in vitro showed its resistance to drought, greater than that of E. amylovora.

Association of electrophoretic karyotype of Candida stellatoidea with virulence for mice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kwon-Chung, K.J.; Wickes, B.L.; Merz, W.G.

1988-07-01

Seven isolates of Candida stellatoidea were studied for their electrophoretic karyotype, virulence for mice, sensitivity to UV radiation, growth rate in vitro, reaction on cycloheximide-indicator medium, and proteinase activity. The isolates exhibited one of two distinct electrophoretic karyotypes as determined by orthogonal field alternating gel electrophoresis (OFAGE). Four isolates, including the type culture of C. stellatoidea, belonged to electrophoretic karyotype type I by OFAGE, showing eight to nine bands of which at least two bands were less than 1,000 kilobases in size as estimated by comparison with the DNA bands of Saccharomyces cerevisiae. These isolates failed to produce fatal infectionmore » in mice within 20 days when 5 X 10(5) cells were injected intravenously. The yeasts were cleared from the kidneys of two of three mice tested by day 30. Type I showed proteinase activity on bovine serum albumin agar at pH 3.8 and produced a negative reaction on cycloheximide-bromcresol green medium within 48 h. The three grouped in type II by OFAGE showed banding patterns similar to those of a well-characterized isolate of Candida albicans. The isolates of type II had an electrophoretic karyotype of six to seven bands approximately 1,200 kilobases or greater in size. All three type II isolates were highly virulent for mice, producing fatality curves similar to those of a previously studied C. albicans isolate. From 80 to 90% of the mice injected with 5 X 10(5) cells intravenously died within 20 days. The type II isolates produced a positive reaction on cycloheximide-bromcresol green agar and showed no proteinase activity on bovine serum albumin agar at the low pH. In addition, the type II isolates grew faster and were significantly more resistant to UV irradiation than the type I isolates.« less

Oral candidal species among smokers and non-smokers.

PubMed

Rasool, S; Siar, C H; Ng, K P

2005-11-01

To determine the various oral Candidal species among healthy Malaysian adults. Case-control study. This study was collaborated between the Department of Medical Microbiology, Faculty of Medicine and Department of Oral Pathology, Oral Medicine and Periodontology, Faculty of Dentistry, University of Malaya, Kuala Lumpur, Malaysia, between September 2002 till January 2004. One hundred adults (50 smokers and 50 non-smokers), aged between 40 and 70 years were studied. Swabs and carbohydrate assimilation (Saboraud Dextrose Agar, Corn Meal Agar, API 20C AUX System) were performed. Specimens were collected from dorsum of the tongue, buccal mucosa and commissures (right and left each). Colony forms were established by positive colony forming units, on SDA medium (24-48 hours). Germ tube test for (true/pseudohyphae) growth was done on Corn Meal Agar Medium. Candida biotypes were evaluated by API 20C AUX system, which had a numerical 7 digit profile, added to evaluate a definite Candida species. Thirty-five percent of Malaysian adults harbored Candida intraorally. Candidal species identified among 100 subjects had C. albicans (27) 77%, C. glabrata (3) 8%, C. famata, C. tropicalis, C. krusei, C. lusitaniae and C. guilliermondii (1) 3% each. Thirty-three positive cases comprised of 35 species i.e. two cases had two species each. Fifty seven percent of these were smokers and 43% non-smokers. These included 40% Chinese, 36% Malays and 24% Indians. Species were, however, not specified according to intra-oral sites i.e. buccal, commissural mucosa and dorsum of tongue. On this series C. albicans is the most common species found in the oral cavity of Malaysian adults. It is equally frequent in smokers and non-smokers, but showed a predilection for the ethnic Chinese group.

Evaluation of chromogenic medium and direct latex agglutination test for detection of group B streptococcus in vaginal specimens from pregnant women in Lebanon and Kuwait.

PubMed

Ghaddar, Nahed; Alfouzan, Wadha; Anastasiadis, Elie; Al Jiser, Tamima; Itani, Saad Eddine; Dernaika, Racha; Eid, Toufic; Ghaddar, Ali; Charafeddine, Adib; Dhar, Rita; El Hajj, Hiba

2014-10-01

This study was undertaken to evaluate chromogenic medium and a direct latex agglutination test (DLA) for detection of Group B Streptococcus (GBS) in the vaginal specimens of pregnant women, and to ascertain the prevalence of GBS in this population in Kuwait and Lebanon. Vaginal swabs, collected from women at 35-37 weeks of gestation, were cultured on 5 % sheep blood agar (SBA), colistin nalidixic acid agar (CNA), Strept B Select chromogenic agar (SBS) as well as Lim enrichment broth in 168 cases in Lebanon while only SBA was used for 1391 samples in Kuwait. In addition, vaginal samples from 102 GBS-positive and 20 GBS-negative women near the time of delivery were collected in Kuwait for evaluation of the DLA test. During the study period, the prevalence of GBS colonization was determined to be 20.7 % (288/1391) in Kuwait while 18.4 % (31) of 168 pregnant women in Lebanon had vaginal cultures positive for GBS. By direct plating of vaginal swabs on the three media used, the isolation rates of GBS were 51.6, 64.5 and 77.4 % on SBA, CNA and SBS, respectively, which increased to 90.35, 93.1 and 96.8 %, respectively, following subculture in Lim broth after 18 h of incubation. The sensitivity of the DLA test was found to be dependent on the density of GBS colonization, resulting in 100 % sensitivity and 100 % specificity for heavy (>10(2) c.f.u. per swab) and moderately heavy (50-100 c.f.u. per swab) growth of GBS. However, for vaginal specimens yielding <50 c.f.u. per swab, the sensitivity, specificity, positive and negative predictive values of the DLA test were 100, 55.5, 63.6 and 100 %, respectively. In conclusion, a chromogenic agar, such as SBS, and a DLA test can be used for rapid detection of GBS in pregnant women. The DLA test, in particular, could prove to be a useful tool for immediate detection of GBS in women near delivery so that intrapartum antibiotic prophylaxis can be initiated. © 2014 The Authors.

Evaluation of different selective media and culturing techniques for the quantification of Campylobacter ssp. from broiler litter.

PubMed

Kiess, A S; Parker, H M; McDaniel, C D

2010-08-01

Poultry is a major reservoir for Campylobacter, the leading cause of foodborne illness in the United States, but how broilers become initially colonized is still under debate. Broiler litter is a potential source, but the best technique for quantifying Campylobacter from litter is still unknown. Therefore, our objectives were to determine if certain media are more selective for quantifying Campylobacter and if enrichment allows for the detection of stressed or viable but nonculturable cells from broiler litter samples. In this trial, 5 media and 2 culturing techniques were used to enumerate Campylobacter from broiler litter. The media used were campy-Line agar (CLA), campy-cefex agar (CCA), modified CCA, Campylobacter agar plates (CAP), and modified charcoal cefoperazone deoxycholate agar. Litter samples were obtained from a commercial broiler house. Each sample was equally divided and diluted 10-fold into peptone, for direct plating, or 4-fold into Campylobacter enrichment broth. Samples diluted in peptone were direct-plated onto each media and incubated under microaerophilic conditions for 48 h at 42 degrees C. Samples diluted in enrichment broth were incubated under the same conditions for 24 h, then further diluted to 10-fold before plating. Plates from enriched samples were incubated for an additional 24 h after plating. After incubation, all plates (direct and enriched) were counted and presumptive positive colonies were confirmed using a Campylobacter latex agglutination kit. Results indicated that there was no difference in the ability of any of the selective media tested to grow Campylobacter. Direct-plated samples had a higher Campylobacter isolation rate compared with enriched samples. The CLA and CAP were able to suppress total bacterial growth better than modified charcoal cefoperazone deoxycholate, modified CCA, and CCA. The CLA and CAP were the only media able to detect total bacterial population shifts over time. In conclusion, it is important before making a final decision on a selective medium to consider the medium's ability to suppress total bacterial growth as well as isolate Campylobacter.

Lippia dulcis shoot cultures as a source of the sweet sesquiterpene hernandulcin.

PubMed

Sauerwein, M; Flores, H E; Yamazaki, T; Shimomura, K

1991-04-01

The axenic shoot culture of Lippia dulcis Trev., Verbenaceae, was established on hormone-free Murashige-Skoog solid medium containing 3% sucrose. Shoots were cultured in various liquid or solid media. Woody Plant liquid medium was best for shoot multiplication, but the production of hernandulcin was relatively low. The highest hernandulcin content (2.9% dry wt) was obtained after 28 days of culture on Murashige-Skoog solid medium containing 2% sucrose. The addition of chitosan to the culture media enhanced the growth of shoots as well as the production of hernandulcin, especially with the liquid medium.

Light scattering sensor for real-time identification of Vibrio parahaemolyticus, V. vulnificus and V. cholera colonies on solid agar plates

USDA-ARS?s Scientific Manuscript database

The three most common pathogenic species of Vibrio, V. cholerae, V. parahemolyticus and V. vulnificus, are of major concern as water- and food-borne pathogens because of an increasing incidence of water and seafood related outbreaks and illnesses worldwide. Current methods are time-consuming and req...

Diversity and dynamics of antibiotic-resistant bacteria in cheese as determined by PCR denaturing gradient gel electrophoresis.

PubMed

Flórez, Ana Belén; Mayo, Baltasar

2015-12-02

This work reports the composition and succession of tetracycline- and erythromycin-resistant bacterial communities in a model cheese, monitored by polymerase chain reaction denaturing gradient gel electrophoresis (PCR-DGGE). Bacterial 16S rRNA genes were examined using this technique to detect structural changes in the cheese microbiota over manufacturing and ripening. Total bacterial genomic DNA, used as a template, was extracted from cultivable bacteria grown without and with tetracycline or erythromycin (both at 25 μg ml(-1)) on a non-selective medium used for enumeration of total and viable cells (Plate Count agar with Milk; PCA-M), and from those grown on selective and/or differential agar media used for counting various bacterial groups; i.e., lactic acid bacteria (de Man, Rogosa and Sharpe agar; MRSA), micrococci and staphylococci (Baird-Parker agar; BPA), and enterobacteria (Violet Red Bile Glucose agar; VRBGA). Large numbers of tetracycline- and erythromycin-resistant bacteria were detected in cheese samples at all stages of ripening. Counts of antibiotic-resistant bacteria varied widely depending on the microbial group and the point of sampling. In general, resistant bacteria were 0.5-1.0 Log10 units fewer in number than the corresponding susceptible bacteria. The PCR-DGGE profiles obtained with DNA isolated from the plates for total bacteria and the different bacterial groups suggested Escherichia coli, Lactococcus lactis, Enterococcus faecalis and Staphylococcus spp. as the microbial types resistant to both antibiotics tested. This study shows the suitability of the PCR-DGGE technique for rapidly identifying and tracking antibiotic resistant populations in cheese and, by extension, in other foods. Copyright © 2015 Elsevier B.V. All rights reserved.

Production of secondary metabolites by some terverticillate penicillia on carbohydrate-rich and meat substrates.

PubMed

Núñez, Félix; Westphal, Carmen D; Bermúdez, Elena; Asensio, Miguel A

2007-12-01

Most terverticillate penicillia isolated from dry-cured meat products are toxigenic, but their ability to produce hazardous metabolites on meat-based substrates is not well known. The production of extrolites by selected terverticillate penicillia isolated from dry-cured ham has been studied on carbohydrate-rich media (malt extract agar, Czapek yeast autolysate agar, rice extract agar, and rice), meat extract triolein salt agar, and ham slices. Chloroform extracts from the selected strains grown on malt extract agar were toxic for the brine shrimp (Artemia salina) larvae and VERO cells at a concentration of 2 mg/ml, but 0.02 mg/ml produced no toxic effect. Analysis by high-pressure liquid chromatography (HPLC) coupled with photodiode array detection (DAD) or with mass spectrometry (MS) and an atmospheric pressure chemical ionization (APCI) source revealed different biologically active metabolites: cyclopiazonic acid and rugulovasine A from Penicillium commune; verrucosidin, anacine, puberuline, verrucofortine, and viridicatols from Penicillium polonicum; arisugacin and viridicatols from Penicillium echinulatum; and compactin and viridicatols from Penicillium solitum. Most of these metabolites, including the amino acid-derived compounds, were produced in the media containing high levels of carbohydrates. High concentrations of nitrogen compounds in the medium does not imply a greater production of the metabolites studied, not even those derived from the amino acids. However, molds growing on dry-cured ham are able to synthesize limited amounts of some secondary metabolites, a fact not previously reported. The combination of HPLC coupled with DAD and MS-APCI was useful for identification of closely related terverticillate Penicillium species from dry-cured ham. These techniques could be used to characterize the risk associated with the potential production of secondary metabolites in cured meats.

Assessing the growth and recovery of Salmonella Enteritidis SE86 after sodium dichloroisocyanurate exposure

PubMed Central

Ferreira, Fernanda Stoduto; Horvath, Mariana Bandeira; Tondo, Eduardo Cesar

2013-01-01

The objective of the present study was to assess the growth and the recovery of Salmonella (S.) Enteritidis SE86 in different diluents, culture media and using different plating methods after the exposure to 200 mg/kg sodium dichloroisocyanurate (NaDCC). Before and after NaDCC exposure, SE86 was cultured at 30 °C and 7 °C in the following diluents: Peptone water (P), Saline solution (SaS), Peptone water+Saline solution (P+SaS), Peptone water+Tween 80+Lecithin+Sodium thiosulfate (P+N) and Saline solution+Tween 80+Lecithin+Sodium thiosulfate (SaS+N). The SaS diluent was chosen because it was able to maintain cells viable without growth and was further used for plating SE86 on non selective medium (Tryptic Soy Agar-TSA) and on selective media (Mannitol Lysine Crystal Violet Brilliant Green Agar-MLCB; Brilliant Green Agar-BGA; Salmonella Shigella Agar-SS and Xylose Lysine Dextrose–XLD). The Thin Agar Layer method (TAL) i.e., selective media overlayed with non selective TSA was also evaluated. Results indicated that SE86 not exposed to NaDCC was able to grow in P, P+N, SaS+N and P+SaS, but not in SaS, that was able to maintain cells viable. SE86 exposed to NaDCC demonstrated similar counts after dilution in SaS and the plating on non selective TSA, selective media MLCB, BGA, SS and XLD and on TAL media. SE86, S. Typhimurium and S. Bredeney, exposed or not exposed to NaDCC, showed no significant differences in counts on TSA, XLD and XLD overlayed with TSA, suggesting that all those media may be used to quantify NaDCC-exposed Salmonella by plating method. PMID:24516446

Mycotoxin production by different ochratoxigenic Aspergillus and Penicillium species on coffee- and wheat-based media.

PubMed

Muñoz, Katherine; Vega, Mario; Rios, Gisela; Geisen, Rolf; Degen, Gisela H

2011-11-01

Ochratoxin A (OTA) is one of the most widespread mycotoxins, and is produced by several Aspergillus or Penicillium species. Human exposure to OTA is mainly by intake of contaminated food, with cereal products, followed by coffee and red wine as the main sources of OTA. In this study, the OTA production of four ochratoxigenic fungi (two Aspergillus and two Penicillium species) was investigated in four different media, i.e. wheat and coffee model media as food-based media and two standard laboratory media (malt extract glucose agar, MEA and yeast extract sucrose agar, YES). Colony growth was documented and OTA concentrations in cultures were determined at day 2, 4 and 8 of incubation at 25°C by high-performance thin-layer chromatography (HPTLC) and high-performance liquid chromatography (HPLC). OTA production clearly depended upon time of incubation, fungal species, and medium composition. On coffee based medium, moderate OTA levels were produced by A. ochraceus BFE635 (9.8 μg/g) and by A. niger BFE632 (10.6 μg/g) on day 8 of incubation. In wheat-based medium, these strains produced much more OTA than in coffee. The highest OTA concentration (83.8 μg/g on day 8) was formed by A. ochraceus BFE635 followed by the other Aspergillus niger BFE632 (49 μg/g). Lower OTA levels were produced by P. verrucosum BFE550 and P. nordicum BFE487, in both wheat and in YES medium, whilst OTA was hardly detectable in coffee and in MEA in case of P. nordicum. Colony growth of the tested strains on different media was not indicative of OTA production. Guttation droplets developed on wheat-based medium with the Aspergillus strains within a week, and this phenomenon coincided with the high OTA amounts formed by these species. Results from this study add to our knowledge on the behaviour of ochratoxigenic fungal species when cultured on food based media.

A new medium for Caenorhabditis elegans toxicology and nanotoxicology studies designed to better reflect natural soil solution conditions.

PubMed

Tyne, William; Lofts, Stephen; Spurgeon, David J; Jurkschat, Kerstin; Svendsen, Claus

2013-08-01

A new toxicity test medium for Caenorhabditis elegans is presented. The test solution is designed to provide a better representation of natural soil pore water conditions than currently available test media. The medium has a composition that can readily be modified to allow for studies of the influences of a range of environmentally relevant parameters on nematode biology and toxicology. Tests conducted in the new medium confirmed that nematodes' reproduction was possible at a range of solution pH levels, offering the potential to conduct toxicity studies under a variety of conditions. A test to establish silver nanoparticle and dissolved silver nitrate toxicity, a study type not feasible in M9 or agar media due to precipitation and nanoparticle agglomeration, indicated lower silver nanoparticle (median effective concentration [EC50] of 6.5 mg Ag/L) than silver nitrate (EC50 0.28 mg Ag/L) toxicity. Characterization identified stable nanoparticle behavior in the new test medium. Copyright © 2013 SETAC.

Physiological and Molecular Characterization of Cephaleuros virescens Occurring in Mango Trees.

PubMed

Vasconcelos, Camila Vilela; Pereira, Fabíola Teodoro; Duarte, Elizabeth Amélia Alves; de Oliveira, Thiago Alves Santos; Peixoto, Nei; Carvalho, Daniel Diego Costa

2018-06-01

The objective of this work was to accomplish the isolation, molecular identification and characterizing the physiology of the causal agent of the algal spot in mango trees. For this purpose, the pathogen growth was assessed in different culture media, with subsequent observation and measurements of the filamentous cells. The molecular identification was made using mycelium obtained from leaf lesions and pure algae colonies grown in culture medium. Descriptions based on DNA sequencing indicated that the algae is Cephaleuros virescens . The algae must be isolated primarily in liquid medium for further pricking into agar medium. The highest mycelial growth average in Petri dishes occurred when the algae were grown in Trebouxia and BBM. Trebouxia enabled larger cells in the filamentous cells when compared to other culture media.

Influence of Culture Media on Microbial Fingerprints Using Raman Spectroscopy.

PubMed

Mlynáriková, Katarína; Samek, Ota; Bernatová, Silvie; Růžička, Filip; Ježek, Jan; Hároniková, Andrea; Šiler, Martin; Zemánek, Pavel; Holá, Veronika

2015-11-24

Raman spectroscopy has a broad range of applications across numerous scientific fields, including microbiology. Our work here monitors the influence of culture media on the Raman spectra of clinically important microorganisms (Escherichia coli, Staphylococcus aureus, Staphylococcus epidermidis and Candida albicans). Choosing an adequate medium may enhance the reproducibility of the method as well as simplifying the data processing and the evaluation. We tested four different media per organism depending on the nutritional requirements and clinical usage directly on a Petri dish. Some of the media have a significant influence on the microbial fingerprint (Roosvelt-Park Institute Medium, CHROMagar) and should not be used for the acquisition of Raman spectra. It was found that the most suitable medium for microbiological experiments regarding these organisms was Mueller-Hinton agar.

Influence of Culture Media on Microbial Fingerprints Using Raman Spectroscopy

PubMed Central

Mlynáriková, Katarína; Samek, Ota; Bernatová, Silvie; Růžička, Filip; Ježek, Jan; Hároniková, Andrea; Šiler, Martin; Zemánek, Pavel; Holá, Veronika

2015-01-01

Raman spectroscopy has a broad range of applications across numerous scientific fields, including microbiology. Our work here monitors the influence of culture media on the Raman spectra of clinically important microorganisms (Escherichia coli, Staphylococcus aureus, Staphylococcus epidermidis and Candida albicans). Choosing an adequate medium may enhance the reproducibility of the method as well as simplifying the data processing and the evaluation. We tested four different media per organism depending on the nutritional requirements and clinical usage directly on a Petri dish. Some of the media have a significant influence on the microbial fingerprint (Roosvelt-Park Institute Medium, CHROMagar) and should not be used for the acquisition of Raman spectra. It was found that the most suitable medium for microbiological experiments regarding these organisms was Mueller-Hinton agar. PMID:26610516

Evaluation of media for simultaneous enumeration of total coliform and Escherichia coli in drinking water supplies by membrane filtration techniques.

PubMed

Wang, Dunling; Fiessel, Wanda

2008-01-01

This study evaluated three different dehydrated media for simultaneous detection and enumeration of total coliform (TC) and Escherichia coli in drinking water samples with a standard membrane filtration procedure. The experiment indicated that the differential coliform agar (DCA) medium was the most effective among the tested media in enumerating TC and E. coli, without the need for extensive accompanying confirmation tests. The results for DCA medium were highly reproducible for both TC and E. coli with standard deviation of 6.0 and 6.1, respectively. A high agreement (82%) was found between DCA and m-Endo media on 152 drinking water samples in terms of TC positive. The DCA medium also reduced concealment of background bacteria.

Preliminary Evaluation of the Control of Microbial Fouling by Laboratory and Pilot-Scale Air-Stripping Columns

DTIC Science & Technology

1985-03-01

used to remove trichloroethylene (TCE) from contaminated well water. 7 MATERIALS Chemicals: Trichloroethylene (Aldrich chemical, Milwaukee, WI), sodium ...Cleveland, OH), sodium hydrcxide (J.T. Baker, Phillipsburg, NJ), potassium dichloroisocyanurate (Dorex Inc., Frankfort, IL), potassium iodide starch...NJ). Media and Reagents: Plate count agar (Difco Laboratories, Detroit, MI), lauryl tryptose broth (Difco Laboratories, Detroit, MI), motility medium

Bacterial Chemotaxis to Naphthalene and Nitroarene Compounds

DTIC Science & Technology

2008-07-31

Qualitative capillary assays showing chemotaxis of succinate-grown 17 (uninduced) and induced (succinate plus salicylate -grown) Acidovorax sp. JS42...succinate plus 2NT- or succinate plus salicylate -grown) wild-type Acidovorax sp. JS42 cells List of Tables Table 1. Summary of chemotaxis...mM salicylate , or naphthalene crystals. Noble agar (1.8%; Difco) was used to solidify MSB medium for plates. For plasmid selection and maintenance

Genome Sequence of Halomonas sp. Strain KO116, an Ionic Liquid- Tolerant Marine Bacterium Isolated from a Lignin-Enriched Seawater Microcosm

DOE PAGES

O'Dell, Kaela; Woo, Hannah L.; Utturkar, Sagar M.; ...

2015-05-07

Halomonas sp. strain KO116 was isolated from Nile Delta Mediterranean Sea surface water enriched with insoluble organosolv lignin. It was further screened for growth on alkali lignin minimal salts medium agar. The strain tolerates the ionic liquid 1-ethyl-3-methylimidazolium acetate. Its complete genome sequence is presented in this report.

Modelling the effect of essential oil of betel leaf (Piper betle L.) on germination, growth, and apparent lag time of Penicillium expansum on semi-synthetic media.

PubMed

Basak, Suradeep; Guha, Proshanta

2015-12-23

The current study aimed at characterizing the chemical components of betel leaf (Piper betle L. var. Tamluk Mitha) essential oil (BLEO) and modelling its effect on growth of Penicillium expansum on semi-synthetic medium. Gas chromatography-mass spectrophotometry (GC-MS) analysis of BLEO revealed the presence of different bioactive phenolic compounds in significant amounts. Among 46 different components identified, chavibetol (22.0%), estragole (15.8%), β-cubebene (13.6%), chavicol (11.8%), and caryophyllene (11.3%) were found to be the major compounds of BLEO. A disc diffusion and disc volatilization method were used to evaluate antifungal activity of the oil against a selected food spoilage mould. The logistic model was used to study the kinetics of spore germination. Prediction and validation of antifungal effect of BLEO was performed on semi-synthetic medium (apple juice agar) using predictive microbiological tools. The Baranyi and Roberts model was used to estimate maximum growth rate (μmax in mm/day) and apparent lag time (λ in days) of the mould. Secondary modelling was performed using a re-parameterized Monod-type equation based on cardinal values to study the effect of different BLEO concentration on estimated growth parameters. Emax (minimum concentration of oil at which mould growth was inhibited) and MIC (minimum inhibitory concentration of BLEO at which lag time is infinite) value of BLEO against P. expansum was estimated to be 0.56 and 0.74 μl/ml, respectively, which was found to be similar on potato dextrose agar (PDA) as well as apple juice agar (AJA) medium. The correlation between estimated growth parameters of the mould on both the media was obtained with satisfactory statistical indices (R(2) and RMSE). This study revealed inhibitory efficacy of BLEO on spore germination, mycelial growth and apparent lag time of P. expansum in a dose-dependent manner. Hence, BLEO has potential to be used as a natural food preservative. Copyright © 2015 Elsevier B.V. All rights reserved.

Solid hydrogen coated graphite particles in the interstellar medium. I.

NASA Technical Reports Server (NTRS)

Swamy, K. S. K.; Wickramasinghe, N. C.

1969-01-01

Solid para hydrogen coated graphite particles expulsion into interstellar medium from star formation regions, considering mantles stability and particles extinction efficiency, albedo and phase function

Optimization of single plate-serial dilution spotting (SP-SDS) with sample anchoring as an assured method for bacterial and yeast cfu enumeration and single colony isolation from diverse samples.

PubMed

Thomas, Pious; Sekhar, Aparna C; Upreti, Reshmi; Mujawar, Mohammad M; Pasha, Sadiq S

2015-12-01

We propose a simple technique for bacterial and yeast cfu estimations from diverse samples with no prior idea of viable counts, designated as single plate-serial dilution spotting (SP-SDS) with the prime recommendation of sample anchoring (10 0 stocks). For pure cultures, serial dilutions were prepared from 0.1 OD (10 0 ) stock and 20 μl aliquots of six dilutions (10 1 -10 6 ) were applied as 10-15 micro-drops in six sectors over agar-gelled medium in 9-cm plates. For liquid samples 10 0 -10 5 dilutions, and for colloidal suspensions and solid samples (10% w/v), 10 1 -10 6 dilutions were used. Following incubation, at least one dilution level yielded 6-60 cfu per sector comparable to the standard method involving 100 μl samples. Tested on diverse bacteria, composite samples and Saccharomyces cerevisiae , SP-SDS offered wider applicability over alternative methods like drop-plating and track-dilution for cfu estimation, single colony isolation and culture purity testing, particularly suiting low resource settings.

Bayesian Integrated Microbial Forensics

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jarman, Kristin H.; Kreuzer-Martin, Helen W.; Wunschel, David S.

2008-06-01

In the aftermath of the 2001 anthrax letters, researchers have been exploring ways to predict the production environment of unknown source microorganisms. Different mass spectral techniques are being developed to characterize components of a microbe’s culture medium including water, carbon and nitrogen sources, metal ions added, and the presence of agar. Individually, each technique has the potential to identify one or two ingredients in a culture medium recipe. However, by integrating data from multiple mass spectral techniques, a more complete characterization is possible. We present a Bayesian statistical approach to integrated microbial forensics and illustrate its application on spores grownmore » in different culture media.« less

A minimal cost micropropagation protocol for Dianthus caryophyllus L.-- a commercially significant venture.

PubMed

Pant, Manu

2016-03-01

In tissue culture, high production cost of the products restricts their reach. Though tissue culture is a major strength in floriculture it is marred by pricing issues. Hence, we developed a complete regeneration low cost micropropagation protocol for an economically important floriculture crop, carnation (Dianthus caryophyllus L.). Successful regeneration of carnation from nodal explants on cost-efficient medium indicates that psyllium husk, sugar and RO water can effectively replace the conventional medium comprising agar, sucrose and distilled water. The protocol can contribute to increased carnation production at comparatively reduced cost, and there by encourage wide scale adoption by the common growers.

Electrical response of culture media during bacterial growth on a paper-based device

NASA Astrophysics Data System (ADS)

Srimongkon, Tithimanan; Buerkle, Marius; Nakamura, Akira; Enomae, Toshiharu; Ushijima, Hirobumi; Fukuda, Nobuko

2017-05-01

In this work, we evaluated the feasibility of a paper-based bacterial detection system. The paper served as a substrate for the measurement electrodes and the culture medium. Using a printing technique, we patterned gold electrodes onto the paper substrate and applied Luria broth (LB) agar gel as a culture medium on top of the electrodes. As the first step towards the development of a bacterial detection system, we determined changes in the surface potential during bacterial growth and monitored these changes over 24 h. This allowed us to correlate changes in the surface potential with the different growth phases of the bacteria.

Inactivation of Salmonella and Listeria in ground chicken breast meat during thermal processing.

PubMed

Murphy, R Y; Marks, B P; Johnson, E R; Johnson, M G

1999-09-01

Thermal inactivation of six Salmonella spp. and Listeria innocua was evaluated in ground chicken breast and liquid medium. Survival of Salmonella and Listeria was affected by the medium composition. Under the same thermal process condition, significantly more Salmonella and Listeria survived in chicken breast meat than in 0.1% peptone-agar solution. The thermal lethality of six tested Salmonella spp. was additive in chicken meat. Survival of Listeria in chicken meat during thermal processing was not affected by the presence of the six Salmonella spp. Sample size and shape affected the inactivation of Salmonella and Listeria in chicken meat during thermal processing.

Method and apparatus for adapting steady flow with cyclic thermodynamics

DOEpatents

Swift, Gregory W.; Reid, Robert S.; Ward, William C.

2000-01-01

Energy transfer apparatus has a resonator for supporting standing acoustic waves at a selected frequency with a steady flow process fluid thermodynamic medium and a solid medium having heat capacity. The fluid medium and the solid medium are disposed within the resonator for thermal contact therebetween and for relative motion therebetween. The relative motion is produced by a first means for producing a steady velocity component and second means for producing an oscillating velocity component at the selected frequency and concomitant wavelength of the standing acoustic wave. The oscillating velocity and associated oscillating pressure component provide energy transfer between the steady flow process fluid and the solid medium as the steady flow process fluid moves through the resonator.

Rapid spot test for the determination of esculin hydrolysis.

PubMed Central

Edberg, S C; Gam, K; Bottenbley, C J; Singer, J M

1976-01-01

Esculin hydrolysis is a useful test in the differentiation of both gram-positive and gram-negative bacteria covering a wide spectrum of aerobes, facultative anaerobes, and anaerobes. Commonly utilized methods require a minimum of 18 h of incubation in broth or agar medium and utilize the production of a brown-black compound, due to the combination of ferric ions with the hydrolysis product esculetin, as indicator. A procedure is presented that requires 15 to 30 min for completion and utilizes fluorescence loss as the indicator of hydrolysis. Esculin fluoresces at 366 nm, whereas the hydrolysis product esculetin does not. Over 1,400 strains of gram-positive and gram-negative bacteria were tested. There was 98.4% of correlation between the spot test and esculin broth and 97% correlation with the bile-esculin agar. Images PMID:787006

Bacillus subtilis UBTn7, a potential producer of L - Methioninase isolated from mangrove, Rhizophora mucronata

NASA Astrophysics Data System (ADS)

Prihanto, A. A.

2018-04-01

L-methioninase is an enzyme that degrades sulfur-containing amino acids to α-keto acids, ammonia, and thiols. L-methioninase could be found in plants, bacteria, and fungi. The aims of this study was to obtain L-methioninase-producing endophytic bacteria isolated from mangrove Rhizophora mucronata. The mangrove was collected from Jenu Beach, Tuban, East Java, Indonesia. The samples were roots, stems, and leaves of Rhizophora mucronata. Endophytic bacteria were pure isolated using LB agar medium. Each bacteria were screened its capability to produce L-methioninase using selective media namely modified Czapek Dox agar. The best producer of enzyme was further identified with morphological and biochemical analysis. The result showed that three bacteria produced L-methioninase. Based on the result of morphological and biochemical analysis, the best producer was Bacillus subtilis UBTn7.

Microbial susceptibility to calcium hydroxide pastes and their vehicles.

PubMed

Gomes, Brenda Paula Figueiredo de Almeida; Ferraz, Caio Cezar Randi; Garrido, Fabio Devora; Rosalen, Pedro Luiz; Zaia, Alexandre Augusto; Teixeira, Fabricio Batista; de Souza-Filho, Francisco José

2002-11-01

The aim of this study was to investigate the susceptibility of some microorganisms commonly isolated from root canals to calcium hydroxide in combination with several vehicles by the agar diffusion method. Stainless-steel cylinders were placed on each inoculated agar medium. The test medications and their controls were placed inside the cylinders. The zones of growth inhibition were measured and recorded after the incubation period for each plate, and the results were analyzed statistically. Enterococcus faecalis was most resistant, whereas the anaerobic Porphyromonas endodontalis was more susceptible to all medications, followed by P. gingivalis and Prevotella intermedial intermedia. Ca(OH)2 + CMCP + glycerin showed significantly larger mean zones of inhibition when compared with the other medications. We conclude that anaerobic Gram-negative bacteria are more susceptible to calcium hydroxide pastes than facultative Gram-positive microorganisms.

CHROMagar Orientation Medium Reduces Urine Culture Workload

PubMed Central

Manickam, Kanchana; Karlowsky, James A.; Adam, Heather; Lagacé-Wiens, Philippe R. S.; Rendina, Assunta; Pang, Paulette; Murray, Brenda-Lee

2013-01-01

Microbiology laboratories continually strive to streamline and improve their urine culture algorithms because of the high volumes of urine specimens they receive and the modest numbers of those specimens that are ultimately considered clinically significant. In the current study, we quantitatively measured the impact of the introduction of CHROMagar Orientation (CO) medium into routine use in two hospital laboratories and compared it to conventional culture on blood and MacConkey agars. Based on data extracted from our Laboratory Information System from 2006 to 2011, the use of CO medium resulted in a 28% reduction in workload for additional procedures such as Gram stains, subcultures, identification panels, agglutination tests, and biochemical tests. The average number of workload units (one workload unit equals 1 min of hands-on labor) per urine specimen was significantly reduced (P < 0.0001; 95% confidence interval [CI], 0.5326 to 1.047) from 2.67 in 2006 (preimplementation of CO medium) to 1.88 in 2011 (postimplementation of CO medium). We conclude that the use of CO medium streamlined the urine culture process and increased bench throughput by reducing both workload and turnaround time in our laboratories. PMID:23363839

[Isolation of Aspergillus tritici from internal environment (Chile): Ecological and clinical scope].

PubMed

Vieille Oyarzo, Peggy; Cruz Choappa, Rodrigo; Piontelli Laforet, Eduardo

2018-03-29

Indoor environments provide important protective habitats for humans, who live or work in them most of the time. Many of these environments lack ventilation, which affects the composition of microbial communities, especially that of the fungal community. The aim of this study is to report the isolation of Aspergillus section Candidi from indoor environments of the School of Medicine at Universidad de Valparaiso, Chile, and identification through morpho-physiological and molecular approaches. Their ecological and clinical features were highlighted. An environmental non-volumetric sampling was performed on PDA medium; 2 petri dishes were exposed in 10 different places to select the Aspergillus samples. Subcultures were performed on agar Czapek with yeast extract (CYA), malt extract agar (MEA) and creatin sacarose agar (CREA) media only for the morpho-physiological and later the molecular identification of white spore species. Of the 20 samples analyzed, one Aspergillus belonging to Candidi section was isolated. Based on its morphology and molecular features, it was classified as Aspergillustritici Mehrotra & Basu. Its ecology and medical relevance are reviewed and discussed. Copyright © 2017 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.

Quantifying Mold Biomass on Gypsum Board: Comparison of Ergosterol and Beta-N-Acetylhexosaminidase as Mold Biomass Parameters

PubMed Central

Reeslev, M.; Miller, M.; Nielsen, K. F.

2003-01-01

Two mold species, Stachybotrys chartarum and Aspergillus versicolor, were inoculated onto agar overlaid with cellophane, allowing determination of a direct measurement of biomass density by weighing. Biomass density, ergosterol content, and beta-N-acetylhexosaminidase (3.2.1.52) activity were monitored from inoculation to stationary phase. Regression analysis showed a good linear correlation to biomass density for both ergosterol content and beta-N-acetylhexosaminidase activity. The same two mold species were inoculated onto wallpapered gypsum board, from which a direct biomass measurement was not possible. Growth was measured as an increase in ergosterol content and beta-N-acetylhexosaminidase activity. A good linear correlation was seen between ergosterol content and beta-N-acetylhexosaminidase activity. From the experiments performed on agar medium, conversion factors (CFs) for estimating biomass density from ergosterol content and beta-N-acetylhexosaminidase activity were determined. The CFs were used to estimate the biomass density of the molds grown on gypsum board. The biomass densities estimated from ergosterol content and beta-N-acetylhexosaminidase activity data gave similar results, showing significantly slower growth and lower stationary-phase biomass density on gypsum board than on agar. PMID:12839773

Congo red agar, a differential medium for Aeromonas salmonicida, detects the presence of the cell surface protein array involved in virulence.

PubMed Central

Ishiguro, E E; Ainsworth, T; Trust, T J; Kay, W W

1985-01-01

Strains of the fish pathogen Aeromonas salmonicida which possess the cell surface protein array known as the A-layer (A+) involved in virulence formed deep red colonies on tryptic soy agar containing 30 micrograms of Congo red per ml. These were readily distinguished from colorless or light orange colonies of avirulent mutants lacking A-layer (A-). The utility of Congo red agar for quantifying A+ and A- cells in the routine assessment of culture virulence was demonstrated. Intact A+ cells adsorbed Congo red, whereas A- mutants did not bind Congo red unless first permeabilized with EDTA. The dye-binding component of A+ cells was shown to be the 50,000-Mr A-protein component of the surface array. Purified A-protein avidly bound Congo red at a dye-to-protein molar ratio of about 30 by a nonspecific hydrophobic mechanism enhanced by high salt concentrations. Neither A+ nor A- cells adsorbed to Congo red-Sepharose columns at low salt concentrations. On the other hand, A+ (but not A-) cells were avidly bound at high salt concentrations. Images PMID:3934141

Clinical use of photodynamic antimicrobial chemotherapy for the treatment of deep carious lesions

NASA Astrophysics Data System (ADS)

Guglielmi, Camila De Almeida B.; Simionato, Maria Regina L.; Ramalho, Karen Müller; Imparato, José Carlos P.; Pinheiro, Sérgio Luiz; Luz, Maria A. A. C.

2011-08-01

The purpose of this study was to assess photodynamic antimicrobial chemotherapy (PACT) via irradiation, using a low power laser associated with a photosensitization dye, as an alternative to remove cariogenic microorganisms by drilling. Remaining dentinal samples in deep carious lesions on permanent molars (n = 26) were treated with 0.01% methylene blue dye and irradiated with a low power laser (InGaAIP - indium gallium aluminum phosphide; λ = 660 nm; 100 mW; 320 Jcm-2 90 s; 9J). Samples of dentin from the pulpal wall region were collected with a micropunch before and immediately after PACT and kept in a transport medium for microbiological analysis. Samples were cultured in plates of Brucella blood agar, Mitis Salivarius Bacitracin agar and Rogosa SL agar to determine the total viable bacteria, mutans streptococci and Lactobacillus spp. counts, respectively. After incubation, colony-forming units were counted and microbial reduction was calculated for each group of bacteria. PACT led to statistically significant reductions in mutans streptococci (1.38 log), Lactobacillus spp. (0.93 log), and total viable bacteria (0.91 log). This therapy may be an appropriate approach for the treatment of deep carious lesions using minimally invasive procedures.

Isolation of Cryptococcus neoformans and other opportunistic fungi from pigeon droppings.

PubMed

Soltani, Maryam; Bayat, Mansour; Hashemi, Seyed J; Zia, Mohammadali; Pestechian, Nader

2013-01-01

Invasive fungal infections cause considerable morbidity and mortality in immunocompromised hosts. Pigeon droppings could especially be a potential carrier in the spread of pathogenic yeasts and mold fungi into the environment. The objective of this study was to isolation of Cryptococcus neoformans and other opportunistic fungi from pigeon droppings. One hundred twenty samples of pigeon droppings were suspended 1:10 in saline solution and then cultured. Identification of C. neoformans was performed on bird seed agar, presence of a capsule on India ink preparation, urease production on urea agar medium and RapID yeast plus system. The identification of candida species was based on micro-morphological analysis on corn meal-Tween 80 agar, RapID yeast plus system and growth in CHROMagar candida. The identification of other fungi was based on macromorphologic, microscopic, biochemical and physiological characteristics. The highest frequency of yeasts and mold fungi were observed in Candida albicans 6.6% and Penicillium spp. 25%. The frequency rate of C. neoformans isolation was 2.5%. Several types of fungi are present in pigeon droppings that can spread in environment and transmit to children and elderly as well as immunocompromised patients who are at increased risk of contracting opportunistic diseases.

A double medium model for diffusion in fluid-bearing rock

NASA Astrophysics Data System (ADS)

Wang, H. F.

1993-09-01

The concept of a double porosity medium to model fluid flow in fractured rock has been applied to model diffusion in rock containing a small amount of a continuous fluid phase that surrounds small volume elements of the solid matrix. The model quantifies the relative role of diffusion in the fluid and solid phases of the rock. The fluid is the fast diffusion path, but the solid contains the volumetrically significant amount of the diffusing species. The double medium model consists of two coupled differential equations. One equation is the diffusion equation for the fluid concentration; it contains a source term for change in the average concentration of the diffusing species in the solid matrix. The second equation represents the assumption that the change in average concentration in a solid element is proportional to the difference between the average concentration in the solid and the concentration in the fluid times the solid-fluid partition coefficient. The double medium model is shown to apply to laboratory data on iron diffusion in fluid-bearing dunite and to measured oxygen isotope ratios at marble-metagranite contacts. In both examples, concentration profiles are calculated for diffusion taking place at constant temperature, where a boundary value changes suddenly and is subsequently held constant. Knowledge of solid diffusivities can set a lower bound to the length of time over which diffusion occurs, but only the product of effective fluid diffusivity and time is constrained for times longer than the characteristic solid diffusion time. The double medium results approach a local, grain-scale equilibrium model for times that are large relative to the time constant for solid diffusion.

Generic model of morphological changes in growing colonies of fungi

NASA Astrophysics Data System (ADS)

López, Juan M.; Jensen, Henrik J.

2002-02-01

Fungal colonies are able to exhibit different morphologies depending on the environmental conditions. This allows them to cope with and adapt to external changes. When grown in solid or semisolid media the bulk of the colony is compact and several morphological transitions have been reported to occur as the external conditions are varied. Here we show how a unified simple mathematical model, which includes the effect of the accumulation of toxic metabolites, can account for the morphological changes observed. Our numerical results are in excellent agreement with experiments carried out with the fungus Aspergillus oryzae on solid agar.

Passage of Campylobacter jejuni and Campylobacter coli subtypes through 0.45 and 0.65 µm pore size nitro-cellulose filters

USDA-ARS?s Scientific Manuscript database

Campylobacter can be difficult to recover from complex samples due to overgrowth by background bacteria. A 0.45 or 0.65 µm pore size filter overlaid on agar plates can be used as a means to separate Campylobacter from confounding non-Campylobacter cells, facilitating detection on solid plating medi...

Adaptation response of Pseudomonas fragi on refrigerated solid matrix to a moderate electric field.

PubMed

Chen, Wenbo; Hu, Honghai; Zhang, Chunjiang; Huang, Feng; Zhang, Dequan; Zhang, Hong

2017-02-10

Moderate electric field (MEF) technology is a promising food preservation strategy since it relies on physical properties-rather than chemical additives-to preserve solid cellular foods during storage. However, the effectiveness of long-term MEF exposure on the psychrotrophic microorganisms responsible for the food spoilage at cool temperatures remains unclear. The spoilage-associated psychrotroph Pseudomonas fragi MC16 was obtained from pork samples stored at 7 °C. Continuous MEF treatment attenuated growth and resulted in subsequent adaptation of M16 cultured on nutrient agar plates at 7 °C, compared to the control cultures, as determined by biomass analysis and plating procedures. Moreover, intracellular dehydrogenase activity and ATP levels also indicated an initial effect of MEF treatment followed by cellular recovery, and extracellular β-galactosidase activity assays indicated no obvious changes in cell membrane permeability. Furthermore, microscopic observations using scanning and transmission electron microscopy revealed that MEF induced sublethal cellular injury during early treatment stages, but no notable changes in morphology or cytology on subsequent days. Our study provides direct evidence that psychrotrophic P. fragi MC16 cultured on nutrient agar plates at 7 °C are capable of adapting to MEF treatment.

Improvement of Aconitum napellus micropropagation by liquid culture on floating membrane rafts.

PubMed

Watad, A A; Kochba, M; Nissim, A; Gaba, V

1995-03-01

An efficient method was developed using floating membrane rafts (Liferaft(™)) for the micropropagation of Aconitum napellus (Ranunculaceae), a cut flower crop with a low natural propagation rate. This was achieved by introducing shoot tips into culture on Murashige and Skoog's (1962) solid medium, or liquid medium-supported rafts, supplemented by different levels of benzyl adenine (BA). Optimum shoot proliferation on solid medium required 4mg/l BA, whereas for expiants supported on rafts optimal proliferation was achieved at 0.25mg/l BA. Maximum shoot proliferation was found using the floating rafts (propagation ratio of 4.2 per month), 45% higher than the maximum value on solid medium. A similar value could be obtained on solid medium after a period of 2 months. The optimal response to BA was similar for fresh weight gain and shoot length. Growth in a shallow layer of liquid in shake flasks gives a similar shoot multiplication rate to that on floating rafts; however, submerged leaves brown and die.

Quantitative analysis of morphological changes in yeast colonies growing on solid medium: the eccentricity and Fourier indices.

PubMed

Gil de Prado, Elena; Rivas, Eva-María; de Silóniz, María-Isabel; Diezma, Belén; Barreiro, Pilar; Peinado, José M

2014-11-01

The colony shape of four yeast species growing on agar medium was measured for 116 days by image analysis. Initially, all the colonies are circular, with regular edges. The loss of circularity can be quantitatively estimated by the eccentricity index, Ei , calculated as the ratio between their orthogonal vertical and horizontal diameters. Ei can increase from 1 (complete circularity) to a maximum of 1.17-1.30, depending on the species. One colony inhibits its neighbour only when it has reached a threshold area. Then, Ei of the inhibited colony increases proportionally to the area of the inhibitory colony. The initial distance between colonies affects those threshold values but not the proportionality, Ei /area; this inhibition affects the shape but not the total surface of the colony. The appearance of irregularities in the edges is associated, in all the species, not with age but with nutrient exhaustion. The edge irregularity can be quantified by the Fourier index, Fi , calculated by the minimum number of Fourier coefficients that are needed to describe the colony contour with 99% fitness. An ad hoc function has been developed in Matlab v. 7.0 to automate the computation of the Fourier coefficients. In young colonies, Fi has a value between 2 (circumference) and 3 (ellipse). These values are maintained in mature colonies of Debaryomyces, but can reach values up to 14 in Saccharomyces. All the species studied showed the inhibition of growth in facing colony edges, but only three species showed edge irregularities associated with substrate exhaustion. Copyright © 2014 John Wiley & Sons, Ltd.

Optimization of laccase production from Marasmiellus palmivorus LA1 by Taguchi method of Design of experiments.

PubMed

Chenthamarakshan, Aiswarya; Parambayil, Nayana; Miziriya, Nafeesathul; Soumya, P S; Lakshmi, M S Kiran; Ramgopal, Anala; Dileep, Anuja; Nambisan, Padma

2017-02-13

Fungal laccase has profound applications in different fields of biotechnology due to its broad specificity and high redox potential. Any successful application of the enzyme requires large scale production. As laccase production is highly dependent on medium components and cultural conditions, optimization of the same is essential for efficient product production. Production of laccase by fungal strain Marasmiellus palmivorus LA1 under solid state fermentation was optimized by the Taguchi design of experiments (DOE) methodology. An orthogonal array (L8) was designed using Qualitek-4 software to study the interactions and relative influence of the seven selected factors by one factor at a time approach. The optimum condition formulated was temperature (28 °C), pH (5), galactose (0.8%w/v), cupric sulphate (3 mM), inoculum concentration (number of mycelial agar pieces) (6Nos.) and substrate length (0.05 m). Overall yield increase of 17.6 fold was obtained after optimization. Statistical optimization leads to the elimination of an insignificant medium component ammonium dihydrogen phosphate from the process and contributes to a 1.06 fold increase in enzyme production. A final production of 667.4 ± 13 IU/mL laccase activity paves way for the application of this strain for industrial applications. Study optimized lignin degrading laccases from Marasmiellus palmivorus LA1. This laccases can thus be used for further applications in different scales of production after analyzing the properties of the enzyme. Study also confirmed the use of taguchi method for optimizations of product production.

Suppressive Effect of Trichoderma spp. on toxigenic Fusarium species.

PubMed

Błaszczyk, Lidia; Basińska-Barczak, Aneta; Ćwiek-Kupczyńska, Hanna; Gromadzka, Karolina; Popiel, Delfina; Stępień, Łukasz

2017-03-30

The aim of the present study was to examine the abilities of twenty-four isolates belonging to ten different Trichoderma species (i.e., Trichoderma atroviride, Trichoderma citrinoviride, Trichoderma cremeum, Trichoderma hamatum, Trichoderma harzianum, Trichoderma koningiopsis, Trichoderma longibrachiatum, Trichoderma longipile, Trichoderma viride and Trichoderma viridescens) to inhibit the mycelial growth and mycotoxin production by five Fusarium strains (i.e., Fusarium avenaceum, Fusarium cerealis, Fusarium culmorum, Fusarium graminearum and Fusarium temperatum). Dual-culture bioassay on potato dextrose agar (PDA) medium clearly documented that all of the Trichoderma strains used in the study were capable of influencing the mycelial growth of at least four of all five Fusarium species on the fourth day after co-inoculation, when there was the first apparent physical contact between antagonist and pathogen. The qualitative evaluation of the interaction between the colonies after 14 days of co-culturing on PDA medium showed that ten Trichoderma strains completely overgrew and sporulated on the colony at least one of the tested Fusarium species. Whereas, the microscopic assay provided evidence that only T. atroviride AN240 and T. viride AN255 formed dense coils around the hyphae of the pathogen from where penetration took place. Of all screened Trichoderma strains, T. atroviride AN240 was also found to be the most efficient (69-100% toxin reduction) suppressors of mycotoxins (deoxynivalenol, 3-acetyl-deoxynivalenol, 15-acetyl-deoxynivalenol, nivalenol, zearalenone, beauvericin, moniliformin) production by all five Fusarium species on solid substrates. This research suggests that T. atroviride AN240 can be a promising candidate for the biological control of toxigenic Fusarium species.

MALDI-TOF mass spectrometry for early identification of bacteria grown in blood culture bottles.

PubMed

Zabbe, Jean-Benoît; Zanardo, Laura; Mégraud, Francis; Bessède, Emilie

2015-08-01

This note reports an interesting way to rapidly identify bacteria grown from blood culture bottles. Chocolate agar plates were inoculated with 1 drop of the positive blood bottle medium. After a 3-hour incubation, the growth veil was submitted to MALDI-TOF mass spectrometry: 77% of the bacteria present have been correctly identified. Copyright © 2015 Elsevier B.V. All rights reserved.

Chromogenic agar medium for detection and isolation of Escherichia coli serogroups O26,O45,O103,O111,O121, and O145 from fresh beef and cattle feces

USDA-ARS?s Scientific Manuscript database

Non-O157 Shiga toxin–producing Escherichia coli (STEC) strains are clinically important foodborne pathogens. Unlike E. coli O157:H7, these foodborne pathogens have no unique biochemical characteristics to readily distinguish them from other E. coli strains growing on plating media. In this study, a ...

[1st environmental isolation of Cryptococcus neoformans var. gattii in Argentina].

PubMed

Davel, G; Abrantes, R; Brudny, M; Córdoba, S; Rodero, L; Canteros, C E; Perrotta, D

2003-01-01

Two strains of C. neoformans var. gattii serotype B were isolated from a park in Buenos Aires city, Argentina. Samples were collected in spring, rubbing with swabs on the inner hollow of Eucalyptus spp. and other trees, which were not identified at the collection moment. Samples were swabbed directly onto cafeic acid with antibiotic agar, then incubated at 28 degrees C and observed daily for a week. Isolates were recovered from cafeic acid medium showing characteristic colonies of the species that allowed differentiation from other microorganisms of the sample. Identification was performed using carbohydrate fermentation, assimilation of nitrogen and carbon sources, and urease and phenoloxidase detection. Strain variety was determined with canavanine-glicine-bromotimol blue agar medium (CGB). Genotypic characterization using AP-PCR with (GACA)4 aleatory primer showed that both band profiles could be differentiated by only one band. These two strains were isolated from tree species different from Eucalyptus spp. These isolates from natural sources demonstrate that C. neoformans var. gattii efosts in Buenos Aires city environment and that it is not necessarily associated with Eucalyptus spp. The presence of C. neoformansvar. gattii in the environment, detected for the first time in Argentina, may explain its sporadic association with human infections in the local population.

Broad-spectrum inhibition of Phytophthora infestans by fungal endophytes

PubMed Central

von Dahlen, Janina K; Schnake, Anika; Ginschel, Sarah; Schulz, Barbara; Rose, Laura E

2018-01-01

ABSTRACT Phytophthora infestans is a devastating pathogen of tomato and potato. It readily overcomes resistance genes and applied agrochemicals and hence even today causes large yield losses. Fungal endophytes provide a largely unexplored avenue of control of Phy. infestans. Not only do endophytes produce a wide array of bioactive metabolites, they may also directly compete with and defeat pathogens in planta. Here, we tested 12 fungal endophytes isolated from different plant species in vitro for their production of metabolites with anti- Phytophthora activity. Four well-performing isolates were evaluated for their ability to suppress nine isolates of Phy. infestans on agar medium and in planta. Two endophytes reliably inhibited all Phy. infestans isolates on agar medium, of which Phoma eupatorii isolate 8082 was the most promising. It nearly abolished infection by Phy. infestans in planta. Our data indicate a role for the production of anti-Phytophthora compounds by the fungus and/or an enhanced plant defense response, as evident by an enhanced anthocyanin production. Here, we present a potential biocontrol agent, which can inhibit a broad-spectrum of Phy. infestans isolates. Such broadly acting inhibition is ideal, because it allows for effective control of genetically diverse isolates and may slow the adaptation of Phy. infestans. PMID:29528408

Effect of media composition, including gelling agents, on isolation of previously uncultured rumen bacteria.

PubMed

Nyonyo, T; Shinkai, T; Tajima, A; Mitsumori, M

2013-01-01

The aim of this study was to develop novel anaerobic media using gellan gum for the isolation of previously uncultured rumen bacteria. Four anaerobic media, a basal liquid medium (BM) with agar (A-BM), a modified BM (MBM) with agar (A-MBM), an MBM with phytagel (P-MBM) and an MBM with gelrite (G-MBM) were used for the isolation of rumen bacteria and evaluated for the growth of previously uncultured rumen bacteria. Of the 214 isolates composed of 144 OTUs, 103 isolates (83 OTUs) were previously uncultured rumen bacteria. Most of the previously uncultured strains were obtained from A-MBM, G-MBM and P-MBM, but the predominant cultural members, isolated from each medium, differed. A-MBM and G-MBM showed significantly higher numbers of different OTUs derived from isolates than A-BM (P < 0·05). The Shannon index indicated that the isolates of A-MBM showed the highest diversity (H' = 3·89) compared with those of G-MBM, P-MBM and A-BM (H' = 3·59, 3·23 and 3·39, respectively). Although previously uncultured rumen bacteria were isolated from all media used, the ratio of previously uncultured bacteria to total isolates was increased in A-MBM, P-MBM and G-MBM. © 2012 The Society for Applied Microbiology.

Isolation, characterization, and complementation of a motility mutant of Spiroplasma citri.

PubMed Central

Jacob, C; Nouzières, F; Duret, S; Bové, J M; Renaudin, J

1997-01-01

The helical mollicute Spiroplasma citri, when growing on low-agar medium, forms fuzzy colonies with occasional surrounding satellite colonies due to the ability of the spiroplasmal cells to move through the agar matrix. In liquid medium, these helical organisms flex, twist, and rotate rapidly. By using Tn4001 insertion mutagenesis, a motility mutant was isolated on the basis of its nondiffuse, sharp-edged colonies. Dark-field microscopy observations revealed that the organism flexed at a low frequency and had lost the ability to rotate about the helix axis. In this mutant, the transposon was shown to be inserted into an open reading frame encoding a putative polypeptide of 409 amino acids for which no significant homology with known proteins was found. The corresponding gene, named scm1, was recovered from the wild-type strain and introduced into the motility mutant by using the S. citri oriC plasmid pBOT1 as the vector. The appearance of fuzzy colonies and the observation that spiroplasma cells displayed rotatory and flexional movements showed the motile phenotype to be restored in the spiroplasmal transformants. The functional complementation of the motility mutant proves the scm1 gene product to be involved in the motility mechanism of S. citri. PMID:9244268

Mathematical modeling of ethanol production in solid-state fermentation based on solid medium' dry weight variation.

PubMed

Mazaheri, Davood; Shojaosadati, Seyed Abbas; Zamir, Seyed Morteza; Mousavi, Seyyed Mohammad

2018-04-21

In this work, mathematical modeling of ethanol production in solid-state fermentation (SSF) has been done based on the variation in the dry weight of solid medium. This method was previously used for mathematical modeling of enzyme production; however, the model should be modified to predict the production of a volatile compound like ethanol. The experimental results of bioethanol production from the mixture of carob pods and wheat bran by Zymomonas mobilis in SSF were used for the model validation. Exponential and logistic kinetic models were used for modeling the growth of microorganism. In both cases, the model predictions matched well with the experimental results during the exponential growth phase, indicating the good ability of solid medium weight variation method for modeling a volatile product formation in solid-state fermentation. In addition, using logistic model, better predictions were obtained.

An axenic culture system for fruiting body formation by an edible bolete phylogenetically related to culinary-medicinal penny bun mushroom, Boletus edulis Bull.:Fr. strains from China.

PubMed

Fu, Shao Chun; Zhang, Mei Yan; Shang, Xiao Dong; Chen, Ming Jie; Tan, Qi

2011-01-01

The ability of two freshly isolated Boletus stains to fruit under axenic conditions was tested using different solid and liquid nutrient media. One strain (YNCX04) produced numerous primordia from which fruiting bodies, 12 mm and 10 mm in length, with grey, convex pilei, and yellow-white, clavate stipes developed between 15 and 30 d after inoculation of fungal mycelium onto a solid medium consisting of mineral salts, thiamine, glucose, potato, an extract of Cunninghamia lanceolata root, and agar. The other strain (YNB200) produced numerous primordia but no sporophores. Strain YNCX04 lost the ability to form fruiting bodies in axenic culture 6 mo after initial isolation but retained the ability to form primordia for up to 18 mo. Based on internal transcribed spacer sequencing data, strains YNB200 and YNCX04 formed a sub-cluster together with four previously designated Boletus edulis strains from China. Phylogenetic analysis placed the Chinese strains closer to B. aestivalis than to European and North American strains of B. edulis, although a 29-bp fragment specific to all the B. aestivalis strains was absent from all the Chinese strains. Furthermore, partial 18S rDNA sequences from strains YNB200 and YNCX04 exhibited 98% similarity with an 18S rDNA sequence from B. edulis strain Be3. Further molecular studies are indicated to more accurately establish the taxonomic positions ofF3 and F4-3, as well as the Chinese strains designated as B. edulis.

[Embolic complications by ink clots removed from syringes during cerebral angiography].

PubMed

Kohyama, Shinya; Ishihara, Shoichiro; Yamane, Fumitaka; Ishihara, Hideaki; Kanazawa, Ryuzaburo; Suzuki, Masanori; Neki, Hiroaki; Ohkawara, Mai

2009-01-01

We noted, during cerebral angiography, that the contrast medium was contaminated with numerous small black ink clots from gradation marks on syringes. In this report, we show that ink can be removed from syringes in solid form, and that they may result in embolic complications during cerebral angiography. To demonstrate that the ink from gradation marks on syringes can come off in a solid form and attach itself to the gloves during cerebral angiography, syringes were gripped many times (just as in an angiographic procedure) after immersion in contrast medium or 0.9% saline for 10 minutes. To see if difference of contrast medium and syringes could affect the removing of ink, five types of nonangiographic syringes and one type of angiographic syringe were rubbed with gauze after certain time periods after immersing them in four kinds of contrast medium or 0.9% saline. Ink attached itself to the gloves in a solid form by repeated gripping due to adherence of contrast medium. Ink was removed from all nonangiographic syringes by rubbing after immersion in any type of contrast medium for two hours. Gradation marks on angiographic syringes were stable with all types of contrast medium. Thus, ink for gradation marks on nonangiographic syringes, which is easily removed in a solid form due to contrast medium, can be the source of embolic complication during cerebral angiography.

The use of bile - esculin agar for the taxonomic classification of the family Enterobacteriaceae.

PubMed

Edberg, S C; Pittman, S; Singer, J M

1977-01-01

Bile-esculin medium has been used for many years for the presumptive identification of group D Streptococcus. The test is based on the ability of a bacterium to grow in the presence of 40% bile and produce esculinase. 2935 strains of Enterobacteriaceae were inoculated onto bile-esculin agar slants and incubated at 35 C. Esculin hydrolysis was determined after 24 and 48 hours. At 24 hours of incubation esculin hydrolysis was limited to the genera Klebsiella, Enterobacter, Serratia, and the species P. vulgaris, P. rettgeri, and C. diversus. Not all strains of these species were positive, however. All other members of the family were negative. At 48 hours of incubation 37% of E. coli gave a positive reaction; all other Enterobacteriaceae which were negative at 24 hours remained negative. Esculin hydrolysis is a valuable test for the taxonomic classification of the family Enterobacteriaceae.

Evaluation of antimicrobial efficacy of Aloe vera and its effectiveness in decontaminating gutta percha cones.

PubMed

Athiban, Prakash P; Borthakur, Bikash Jyoti; Ganesan, S; Swathika, B

2012-07-01

The aim of this study was to evaluate the antimicrobial efficacy of Aloe vera and to determine its effectiveness in decontaminating gutta percha cones. A concentrated extract of Aloe vera was used to check for the antimicrobial efficacy using the agar well diffusion method. Presence of zones' of diffusion was identified against three common GP contaminants namely, E.coli, E.faecalis and Staph. aureus. New GP Cones, freshly taken out of the packet were then decontaminated for 1minute using Aloe vera gel and then placed in thioglycolate broth to check for the presence of turbidity. The zones of inhibition on the agar plate were measured as 24mm,21mm and 24mm respectively. The broth remained clear even after 48 hours of incubation. We conclude that Aloe vera is indeed effective as a GP decontaminant and it holds a promising future as a medium for storage of GP cones.

Discrimination of selected species of pathogenic bacteria using near-infrared Raman spectroscopy and principal components analysis

NASA Astrophysics Data System (ADS)

de Siqueira e Oliveira, Fernanda S.; Giana, Hector E.; Silveira, Landulfo, Jr.

2012-03-01

It has been proposed a method based on Raman spectroscopy for identification of different microorganisms involved in bacterial urinary tract infections. Spectra were collected from different bacterial colonies (Gram negative: E. coli, K. pneumoniae, P. mirabilis, P. aeruginosa, E. cloacae and Gram positive: S. aureus and Enterococcus sp.), grown in culture medium (Agar), using a Raman spectrometer with a fiber Raman probe (830 nm). Colonies were scraped from Agar surface placed in an aluminum foil for Raman measurements. After pre-processing, spectra were submitted to a Principal Component Analysis and Mahalanobis distance (PCA/MD) discrimination algorithm. It has been found that the mean Raman spectra of different bacterial species show similar bands, being the S. aureus well characterized by strong bands related to carotenoids. PCA/MD could discriminate Gram positive bacteria with sensitivity and specificity of 100% and Gram negative bacteria with good sensitivity and high specificity.

Bacteriological study of juvenile periodontitis in China.

PubMed

Han, N M; Xiao, X R; Zhang, L S; Ri, X Q; Zhang, J Z; Tong, Y H; Yang, M R; Xiao, Z R

1991-09-01

The predominant cultivable bacteria associated with juvenile periodontitis (JP) in China were studied for the first time. Subgingival plaque samples were taken on paper points from 23 diseased sites in 15 JP patients and from 7 healthy sites in 7 control subjects. Serially diluted plaque samples were plated on nonselective blood agar and on MGB agar, a selective medium for the isolation of Actinobacillus actinomycetemcomitans. Fifteen or more isolated colonies from each sample (in sequence without selection) were purified for identification. The results indicated that the microflora in healthy sulci of the 7 control subjects was significantly different from that in diseased sites of JP patients. The predominant species in healthy sulci were Streptococcus spp. and Capnocytophaga gingivalis. In JP patients, Eubacterium sp. was found in significantly higher frequency and proportion. Actinobacillus actinomycetemcomitans was not detected in any samples. It appears that this species is not associated with juvenile periodontitis in China.

Validation of an automated colony counting system for group A Streptococcus.

PubMed

Frost, H R; Tsoi, S K; Baker, C A; Laho, D; Sanderson-Smith, M L; Steer, A C; Smeesters, P R

2016-02-08

The practice of counting bacterial colony forming units on agar plates has long been used as a method to estimate the concentration of live bacteria in culture. However, due to the laborious and potentially error prone nature of this measurement technique, an alternative method is desirable. Recent technologic advancements have facilitated the development of automated colony counting systems, which reduce errors introduced during the manual counting process and recording of information. An additional benefit is the significant reduction in time taken to analyse colony counting data. Whilst automated counting procedures have been validated for a number of microorganisms, the process has not been successful for all bacteria due to the requirement for a relatively high contrast between bacterial colonies and growth medium. The purpose of this study was to validate an automated counting system for use with group A Streptococcus (GAS). Twenty-one different GAS strains, representative of major emm-types, were selected for assessment. In order to introduce the required contrast for automated counting, 2,3,5-triphenyl-2H-tetrazolium chloride (TTC) dye was added to Todd-Hewitt broth with yeast extract (THY) agar. Growth on THY agar with TTC was compared with growth on blood agar and THY agar to ensure the dye was not detrimental to bacterial growth. Automated colony counts using a ProtoCOL 3 instrument were compared with manual counting to confirm accuracy over the stages of the growth cycle (latent, mid-log and stationary phases) and in a number of different assays. The average percentage differences between plating and counting methods were analysed using the Bland-Altman method. A percentage difference of ±10 % was determined as the cut-off for a critical difference between plating and counting methods. All strains measured had an average difference of less than 10 % when plated on THY agar with TTC. This consistency was also observed over all phases of the growth cycle and when plated in blood following bactericidal assays. Agreement between these methods suggest the use of an automated colony counting technique for GAS will significantly reduce time spent counting bacteria to enable a more efficient and accurate measurement of bacteria concentration in culture.

High solid loading aqueous base metal/ceramic feedstock for injection molding

NASA Astrophysics Data System (ADS)

Behi, Mohammad

2001-07-01

Increasing volume fraction of metal powder in feedstock provided lower shrinkage. Reduction of the shrinkage results in better dimensional precision. The rheology of the feedstock material plays an important role to allowing larger volume fractions of the metal powder to be incorporated in the feedstock formulations. The viscosity of the feedstock mainly depends on the binder viscosity, powder volume fraction and characteristics of metal powder. Aqueous polysaccharide agar was used as a baseline binder system for this study. The effect of several gel-strengthening additives on 1.5wt% and 2wt% agar gel was evaluated. A new gel-strengthening additive was found to be the most effective among the others. The effect of other additives such as glucose, sucrose and fructose on viscosity of baseline binder and feedstock was investigated. Two new agar based binder compositions were developed. The use of these new binder formulations significantly improved the volume fraction of the metal powder, the stability of the feedstock, and reduced the final shrinkage of the molded articles. Two types of 17-4PH stainless steel metal powders, one gas atomized and, the other water atomized, were used for this research.

Comparison of Dry Medium Culture Plates for Mesophilic Aerobic Bacteria in Milk, Ice Cream, Ham, and Codfish Fillet Products

PubMed Central

Park, Junghyun; Kim, Myunghee

2013-01-01

This study was performed to compare the performance of Sanita-Kun dry medium culture plate with those of traditional culture medium and Petrifilm dry medium culture plate for the enumeration of the mesophilic aerobic bacteria in milk, ice cream, ham, and codfish fillet. Mesophilic aerobic bacteria were comparatively evaluated in milk, ice cream, ham, and codfish fillet using Sanita-Kun aerobic count (SAC), Petrifilm aerobic count (PAC), and traditional plate count agar (PCA) media. According to the results, all methods showed high correlations of 0.989~1.000 and no significant differences were observed for enumerating the mesophilic aerobic bacteria in the tested food products. SAC method was easier to perform and count colonies efficiently as compared to the PCA and PAC methods. Therefore, we concluded that the SAC method offers an acceptable alternative to the PCA and PAC methods for counting the mesophilic aerobic bacteria in milk, ice cream, ham, and codfish fillet products. PMID:24551829

Media and growth conditions for induction of secondary metabolite production.

PubMed

Frisvad, Jens C

2012-01-01

Growth media and incubation conditions have a very strong influence of secondary metabolite production. There is no consensus on which media are the optimal for metabolite production, but a series of useful and effective media and incubation conditions have been listed here. Chemically well-defined media are suited for biochemical studies, but in order to get chemical diversity expressed in filamentous fungi, sources rich in amino acids, vitamins, and trace metals have to be added, such as yeast extract and oatmeal. A battery of solid agar media is recommended for exploration of chemical diversity as agar plug samples are easily analyzed to get an optimal representation of the qualitative secondary metabolome. Standard incubation for a week at 25°C in darkness is recommended, but optimal conditions have to be modified depending on the ecology and physiology of different filamentous fungi.

Novel Selective Detection Method of Tumor Angiogenesis Factors Using Living Nano-Robots.

PubMed

Al-Fandi, Mohamed; Alshraiedeh, Nida; Owies, Rami; Alshdaifat, Hala; Al-Mahaseneh, Omamah; Al-Tall, Khadijah; Alawneh, Rawan

2017-07-14

This paper reports a novel self-detection method for tumor cells using living nano-robots. These living robots are a nonpathogenic strain of E. coli bacteria equipped with naturally synthesized bio-nano-sensory systems that have an affinity to VEGF, an angiogenic factor overly-expressed by cancer cells. The VEGF-affinity/chemotaxis was assessed using several assays including the capillary chemotaxis assay, chemotaxis assay on soft agar, and chemotaxis assay on solid agar. In addition, a microfluidic device was developed to possibly discover tumor cells through the overexpressed vascular endothelial growth factor (VEGF). Various experiments to study the sensing characteristic of the nano-robots presented a strong response toward the VEGF. Thus, a new paradigm of selective targeting therapies for cancer can be advanced using swimming E. coli as self-navigator miniaturized robots as well as drug-delivery vehicles.

Comparative Study of the New Colorimetric VITEK 2 Yeast Identification Card versus the Older Fluorometric Card and of CHROMagar Candida as a Source Medium with the New Card

PubMed Central

Aubertine, C. L.; Rivera, M.; Rohan, S. M.; Larone, D. H.

2006-01-01

The new VITEK 2 colorimetric card was compared to the previous fluorometric card for identification of yeast. API 20C was considered the “gold standard.” The new card consistently performed better than the older card. Isolates from CHROMagar Candida plates were identified equally as well as those from Sabouraud dextrose agar. PMID:16390976

Evaluation of the Biomic V3 Microbiology System for Identification of Selected Species on BBL CHROMagar Orientation Agar and CHROMagar MRSA Medium ▿

PubMed Central

Baron, Ellen Jo; D'Souza, Holly; Qi Wang, Andrew; Gibbs, David L.

2008-01-01

The Biomic V3 microbiology system identifies bacteria by reading the color of colonies selected by the user. For CHROMagar orientation, Biomic results agreed with conventional methods for 94% of the strains assayed. For CHROMagar MRSA, Biomic correctly identified 100% of the strains tested and did not misidentify two methicillin-susceptible Staphylococcus aureus strains growing on the plates. PMID:18701661

Somatic embryogenesis from flower explants of cocoa (Theobroma cacao L.).

PubMed

Silva, J J; Debergh, P

2001-01-01

Two types of flower explants, staminoides and petals, were used for in vitro induction of somatic embryos in cocoa. After 14 days in culture, we observed globular structures and callus formation on both types of explants. However, the better results were obtained on staminoides: 98.3% formed callus and 86.2% somatic embryos on Murashige and Skoog (1962) medium supplemented with sucrose, coconut water, 2,4-D, kinetin and agar.

The epidemiology of Candida species associated with vulvovaginal candidiasis in an Iranian patient population.

PubMed

Mahmoudi Rad, M; Zafarghandi, S; Abbasabadi, B; Tavallaee, M

2011-04-01

Vulvovaginal candidiasis is a common infection among women worldwide. According to previous epidemiological studies, Candida albicans is the most common species of Candida. The prevalence of non-Candida species, however, is increasing. Identification of Candida species among the population will not only help health professionals to choose suitable antifungal treatments, but also prevent development of drug resistance. The aim of this study was to identify, using chromogenic agar medium, the Candida species associated with vulvovaginal candidiasis among a sample of the Iranian population. In a prospective cohort study during a two year period from March 2006 to March 2008, swab samples of vaginal discharge/secretion were taken from 200 patients admitted to the gynecology clinic of Mahdieh Hospital (Tehran, Iran) with a clinical presentation suggestive of vulvovaginal candidiasis. The isolates obtained were cultured on Sabouraud dextrose agar and chromogenic agar medium. Candida species were also identified by germ tube formation in serum, chlamydospore production on Corn Meal Agar and carbohydrate absorption using the API 20C-AUX kit. Participants were asked to complete a questionnaire investigating the risk factors associated with candidiasis. An assessment of the different species of recurrent and non-recurrent candidiasis was also made. Descriptive statistics, chi-square test, and t-test were used to analyze the data. A total of 191 isolates were obtained from 175 vaginal specimens. Candida albicans accounted for 67% of the strains including single and mixed infections. The other identified species were Candida glabrata (18.3%), Candida tropicalis (6.8%), Candida krusei (5.8%), Candida parapsilosis (1.6%), and Candida guilliermondii (0.5%) respectively. Mixed infection with two or more species of Candida was seen in 10.3% of patients. The most common mixed cause was the combination of Candida albicans and Candida glabrata. Participants who were sexually active and those who had orogenital sex were more likely to suffer recurrent vulvovaginal candidiasis. Candida albicans was the most common cause of recurrent and non-recurrent vulvovaginitis. The second most common species was Candida glabrata. This study suggests CHROMagar method as a convenient and cost effective yet reliable method to isolate the species of Candida especially in cases where more than one species is present. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

Water relations in culture media influence maturation of avocado somatic embryos.

PubMed

Márquez-Martín, Belén; Sesmero, Rafael; Quesada, Miguel A; Pliego-Alfaro, Fernando; Sánchez-Romero, Carolina

2011-11-15

Application of transformation and other biotechnological tools in avocado (Persea americana Mill.) is hampered by difficulties in obtaining mature somatic embryos capable of germination at an acceptable rate. In this work, we evaluated the effect of different compounds affecting medium water relations on maturation of avocado somatic embryos. Culture media were characterized with respect to gel strength, water potential and osmotic potential. Improved production of mature somatic embryos was achieved with gelling agent concentrations higher than those considered standard. The osmotic agents such as sorbitol and PEG did not have positive effects on embryo maturation. The number of w-o mature somatic embryos per culture was positively correlated with medium gel strength. Gel strength was significantly affected by gelling agent type as well as by gelling agent and PEG concentration. Medium water potential was influenced by sorbitol concentration; incorporation of PEG to a culture medium did not affect medium water potential. The highest maturation results were achieved on a medium gelled with 10 gl(-1) agar. Moreover, these somatic embryos had improved germination rates. These results corroborate the role of water restriction as a key factor controlling maturation of somatic embryos. Copyright © 2011 Elsevier GmbH. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fox, Sandra Lynn; Bala, Greg Alan

Surfactin, a lipopeptide biosurfactant, produced by Bacillus subtilis is known to reduce the surface tension of water from 72 to 27 mN/m. Potato substrates were evaluated as a carbon source for surfactant production by B. subtilis ATCC 21332. An established potato medium, simulated liquid and solid potato waste media, and a commercially prepared potato starch in a mineral salts medium were evaluated in shake flask experiments to verify growth, surface tension reduction, and carbohydrate reduction capabilities. Total carbohydrate assays and glucose monitoring indicated that B. subtilis was able to degrade potato substrates to produce surfactant. Surface tensions dropped from 71.3±0.1more » to 28.3±0.3 mN/m (simulated solid potato medium) and to 27.5±0.3 mN/m (mineral salts medium). A critical micelle concentration (CMC) of 0.10 g/l was obtained from a methylene chloride extract of the simulated solid potato medium.« less

High energy bursts from a solid state laser operated in the heat capacity limited regime

DOEpatents

Albrecht, G.; George, E.V.; Krupke, W.F.; Sooy, W.; Sutton, S.B.

1996-06-11

High energy bursts are produced from a solid state laser operated in a heat capacity limited regime. Instead of cooling the laser, the active medium is thermally well isolated. As a result, the active medium will heat up until it reaches some maximum acceptable temperature. The waste heat is stored in the active medium itself. Therefore, the amount of energy the laser can put out during operation is proportional to its mass, the heat capacity of the active medium, and the temperature difference over which it is being operated. The high energy burst capacity of a heat capacity operated solid state laser, together with the absence of a heavy, power consuming steady state cooling system for the active medium, will make a variety of applications possible. Alternately, cooling takes place during a separate sequence when the laser is not operating. Industrial applications include new material working processes. 5 figs.

High energy bursts from a solid state laser operated in the heat capacity limited regime

DOEpatents

Albrecht, Georg; George, E. Victor; Krupke, William F.; Sooy, Walter; Sutton, Steven B.

1996-01-01

High energy bursts are produced from a solid state laser operated in a heat capacity limited regime. Instead of cooling the laser, the active medium is thermally well isolated. As a result, the active medium will heat up until it reaches some maximum acceptable temperature. The waste heat is stored in the active medium itself. Therefore, the amount of energy the laser can put out during operation is proportional to its mass, the heat capacity of the active medium, and the temperature difference over which it is being operated. The high energy burst capacity of a heat capacity operated solid state laser, together with the absence of a heavy, power consuming steady state cooling system for the active medium, will make a variety of applications possible. Alternately, cooling takes place during a separate sequence when the laser is not operating. Industrial applications include new material working processes.

Cultivation characteristics and gene expression profiles of Aspergillus oryzae by membrane-surface liquid culture, shaking-flask culture, and agar-plate culture.

PubMed

Imanaka, Hiroyuki; Tanaka, Soukichi; Feng, Bin; Imamura, Koreyoshi; Nakanishi, Kazuhiro

2010-03-01

We cultivated a filamentous fungus, Aspergillus oryzae IAM 2706 by three different cultivation methods, i.e., shaking-flask culture (SFC), agar-plate culture (APC), and membrane-surface liquid culture (MSLC), to elucidate the differences of its behaviors by different cultivation methods under the same media, by measuring the growth, secretion of proteases and alpha-amylase, secreted protein level, and gene transcriptional profile by the DNA microarray analysis. The protease activities detected by MSLC and APC were much higher than that by SFC, using both modified Czapek-Dox (mCD) and dextrin-peptone-yeast extract (DPY) media. The alpha-amylase activity was detected in MSLC and APC in a much larger extent than that in SFC when DPY medium was used. On the basis of SDS-PAGE analyses and N-terminal amino acid sequences, 6 proteins were identified in the supernatants of the culture broths using DPY medium, among which oryzin (alkaline protease) and alpha-amylase were detected at a much higher extent for APC and MSLC than those for SFC while only oryzin was detected in mCD medium, in accordance with the activity measurements. A microarray analysis for the fungi cultivated by SFC, APC, and MSLC using mCD medium was carried out to elucidate the differences in the gene transcriptional profile by the cultivation methods. The gene transcriptional profile obtained for the MSLC sample showed a similar tendency to the APC sample while it was quite different from that for the SFC sample. Most of the genes specifically transcribed in the MSLC sample versus those in the SFC sample with a 10-fold up-regulation or higher were unknown or predicted proteins. However, transcription of oryzin gene was only slightly up-regulated in the MSLC sample and that of alpha-amylase gene, slightly down-regulated. Copyright 2009 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Isolation of Cryptococcus neoformans and other opportunistic fungi from pigeon droppings

PubMed Central

Soltani, Maryam; Bayat, Mansour; Hashemi, Seyed J.; Zia, Mohammadali; Pestechian, Nader

2013-01-01

Background: Invasive fungal infections cause considerable morbidity and mortality in immunocompromised hosts. Pigeon droppings could especially be a potential carrier in the spread of pathogenic yeasts and mold fungi into the environment. The objective of this study was to isolation of Cryptococcus neoformans and other opportunistic fungi from pigeon droppings. Materials and Methods: One hundred twenty samples of pigeon droppings were suspended 1:10 in saline solution and then cultured. Identification of C. neoformans was performed on bird seed agar, presence of a capsule on India ink preparation, urease production on urea agar medium and RapID yeast plus system. The identification of candida species was based on micro-morphological analysis on corn meal-Tween 80 agar, RapID yeast plus system and growth in CHROMagar candida. The identification of other fungi was based on macromorphologic, microscopic, biochemical and physiological characteristics. Results: The highest frequency of yeasts and mold fungi were observed in Candida albicans 6.6% and Penicillium spp. 25%. The frequency rate of C. neoformans isolation was 2.5%. Conclusion: Several types of fungi are present in pigeon droppings that can spread in environment and transmit to children and elderly as well as immunocompromised patients who are at increased risk of contracting opportunistic diseases. PMID:23901339

The fungicidal and phytotoxic properties of benomyl and PPM in supplemented agar media supporting transgenic arabidopsis plants for a Space Shuttle flight experiment

NASA Technical Reports Server (NTRS)

Paul, A. L.; Semer, C.; Kucharek, T.; Ferl, R. J.

2001-01-01

Fungal contamination is a significant problem in the use of sucrose-enriched agar-based media for plant culture, especially in closed habitats such as the Space Shuttle. While a variety of fungicides are commercially available, not all are equal in their effectiveness in inhibiting fungal contamination. In addition, fungicide effectiveness must be weighed against its phytotoxicity and in this case, its influence on transgene expression. In a series of experiments designed to optimize media composition for a recent shuttle mission, the fungicide benomyl and the biocide "Plant Preservative Mixture" (PPM) were evaluated for effectiveness in controlling three common fungal contaminants, as well as their impact on the growth and development of arabidopsis seedlings. Benomyl proved to be an effective inhibitor of all three contaminants in concentrations as low as 2 ppm (parts per million) within the agar medium, and no evidence of phytotoxicity was observed until concentrations exceeded 20 ppm. The biocide mix PPM was effective as a fungicide only at concentrations that had deleterious effects on arabidopsis seedlings. As a result of these findings, a concentration of 3 ppm benomyl was used in the media for experiment PGIM-01 which flew on shuttle Columbia mission STS-93 in July 1999.

Nodulation-dependent communities of culturable bacterial endophytes from stems of field-grown soybeans.

PubMed

Okubo, Takashi; Ikeda, Seishi; Kaneko, Takakazu; Eda, Shima; Mitsui, Hisayuki; Sato, Shusei; Tabata, Satoshi; Minamisawa, Kiwamu

2009-01-01

Endophytic bacteria (247 isolates) were randomly isolated from surface-sterilized stems of non-nodulated (Nod(-)), wild-type nodulated (Nod(+)), and hypernodulated (Nod(++)) soybeans (Glycine max [L.] Merr) on three agar media (R2A, nutrient agar, and potato dextrose agar). Their diversity was compared on the basis of 16S rRNA gene sequences. The phylogenetic composition depended on the soybean nodulation phenotype, although diversity indexes were not correlated with nodulation phenotype. The most abundant phylum throughout soybean lines tested was Proteobacteria (58-79%). Gammaproteobacteria was the dominant class (21-72%) with a group of Pseudomonas sp. significantly abundant in Nod(+) soybeans. A high abundance of Alphaproteobacteria was observed in Nod(-) soybeans, which was explained by the increase in bacterial isolates of the families Rhizobiaceae and Sphingomonadaceae. A far greater abundance of Firmicutes was observed in Nod(-) and Nod(++) mutant soybeans than in Nod(+) soybeans. An impact of culture media on the diversity of isolated endophytic bacteria was also observed: The highest diversity indexes were obtained on the R2A medium, which enabled us to access Alphaproteobacteria and other phyla more frequently. The above results indicated that the extent of nodulation changes the phylogenetic composition of culturable bacterial endophytes in soybean stems.

Evaluation of a modified Cefsulodin-Irgasan-Novobiocin agar for isolation of Yersinia spp.

PubMed

Tan, Lai Kuan; Ooi, Peck Toung; Carniel, Elisabeth; Thong, Kwai Lin

2014-01-01

Y. enterocolitica and Y. pseudotuberculosis are important food borne pathogens. However, the presence of competitive microbiota makes the isolation of Y. enterocolitica and Y. pseudotuberculosis from naturally contaminated foods difficult. We attempted to evaluate the performance of a modified Cefsulodin-Irgasan-Novobiocin (CIN) agar in the differentiation of Y. enterocolitica from non-Yersinia species, particularly the natural intestinal microbiota. The modified CIN enabled the growth of Y. enterocolitica colonies with the same efficiency as CIN and Luria-Bertani agar. The detection limits of the modified CIN for Y. enterocolitica in culture medium (10 cfu/ml) and in artificially contaminated pork (10(4) cfu/ml) were also comparable to those of CIN. However, the modified CIN provided a better discrimination of Yersinia colonies from other bacteria exhibiting Yersinia-like colonies on CIN (H2S-producing Citrobacter freundii, C. braakii, Enterobacter cloacae, Aeromonas hydrophila, Providencia rettgeri, and Morganella morganii). The modified CIN exhibited a higher recovery rate of Y. enterocolitica from artificially prepared bacterial cultures and naturally contaminated samples compared with CIN. Our results thus demonstrated that the use of modified CIN may be a valuable means to increase the recovery rate of food borne Yersinia from natural samples, which are usually contaminated by multiple types of bacteria.

Mycoflora of soybeans used for meju fermentation.

PubMed

Kim, Dae-Ho; Kim, Seon-Hwa; Kwon, Soon-Wo; Lee, Jong-Kyu; Hong, Seung-Beom

2013-06-01

Diverse fungi are present in Korean traditional meju and they are known to play an important role in fermented soybean products. To determine the origin of the fungi in meju, we examined the mycoflora of soybeans from 10 traditional meju factories. The samples were untreated or treated with sodium hypochlorite, and placed on malt extract agar (MEA), dichloran 18% glycerol agar (DG18), and dichloran rose bengal chloramphenicol agar (DRBC) medium. A total of 794 fungal strains were isolated and they were identified as 41 genera and 86 species. From sodium hypochlorite untreated soybeans, the genera, Cladosporium (55%), Eurotium (51%), Fusarium (33%), Penicillium (22%), and Aspergillus (exclusion of Eurotium) (20%), were mainly isolated, and Eurotium herbariorum (22%), Eurotium repens (18%), Cladosporium tenuissimum (18%), F. fujikuroi (18%), Aspergillus oryzae/flavus (7%), and Penicillium steckii (6%) were the predominant species. In case of sodium hypochlorite-treated soybeans, Eurotium (31%) and Cladosporium (5%) were frequently isolated, but Aspergillus (excluding Eurotium), Penicillium and Fusarium which were frequently isolated from untreated soybeans, were rarely isolated. Eurotium herbariorum (21%), Eurotium repens (8%), and Cladosporium tenuissimum (3%) were the predominant species. Of the 41 genera and 86 species isolated from soybeans, 13 genera and 33 species were also found in meju. These results suggest that the fungi on soybeans may influence the mycoflora of meju.

Mycoflora of Soybeans Used for Meju Fermentation

PubMed Central

Kim, Dae-Ho; Kim, Seon-Hwa; Kwon, Soon-Wo; Lee, Jong-Kyu

2013-01-01

Diverse fungi are present in Korean traditional meju and they are known to play an important role in fermented soybean products. To determine the origin of the fungi in meju, we examined the mycoflora of soybeans from 10 traditional meju factories. The samples were untreated or treated with sodium hypochlorite, and placed on malt extract agar (MEA), dichloran 18% glycerol agar (DG18), and dichloran rose bengal chloramphenicol agar (DRBC) medium. A total of 794 fungal strains were isolated and they were identified as 41 genera and 86 species. From sodium hypochlorite untreated soybeans, the genera, Cladosporium (55%), Eurotium (51%), Fusarium (33%), Penicillium (22%), and Aspergillus (exclusion of Eurotium) (20%), were mainly isolated, and Eurotium herbariorum (22%), Eurotium repens (18%), Cladosporium tenuissimum (18%), F. fujikuroi (18%), Aspergillus oryzae/flavus (7%), and Penicillium steckii (6%) were the predominant species. In case of sodium hypochlorite-treated soybeans, Eurotium (31%) and Cladosporium (5%) were frequently isolated, but Aspergillus (excluding Eurotium), Penicillium and Fusarium which were frequently isolated from untreated soybeans, were rarely isolated. Eurotium herbariorum (21%), Eurotium repens (8%), and Cladosporium tenuissimum (3%) were the predominant species. Of the 41 genera and 86 species isolated from soybeans, 13 genera and 33 species were also found in meju. These results suggest that the fungi on soybeans may influence the mycoflora of meju. PMID:23874133

Biodegradative potential of fungal isolates from sacral ambient: In vitro study as risk assessment implication for the conservation of wall paintings

PubMed Central

Dimkić, Ivica; Stupar, Miloš; Stanković, Slaviša; Vukojević, Jelena; Ljaljević Grbić, Milica

2018-01-01

The principal purpose of the study was to evaluate in vitro the potential ability of fungal isolates obtained from the painted layer of frescoes and surrounding air to induce symptoms of fresco deterioration, associated with their growth and metabolism, so that the risk of such deterioration can be precisely assessed and appropriate conservation treatments formulated. Biodegradative properties of the tested microfungi were qualitatively characterized through the use of a set of special agar plates: CaCO3 glucose agar (calcite dissolution), casein nutrient agar (casein hydrolysis), Czapek-Dox minimal medium (pigment secretion); and Czapek-Dox minimal broth (acid and alkali production). Most of the tested isolates (71.05%) demonstrated at least one of the degradative properties, with Penicillium bilaiae as the most potent, since it tested positive in all four. The remaining isolates (28.95%) showed no deterioration capabilities and were hence considered unlikely to partake in the complex process of fungal deterioration of murals via the tested mechanisms. The obtained results clearly indicate that utilization of fast and simple plate assays can provide insight into the biodegradative potential of deteriogenic fungi and allow for their separation from allochthonous transients, a prerequisite for precise assessment of the amount of risk posed by a thriving mycobiota to mural paintings. PMID:29309432

l-Asparaginase from Proteus vulgaris1

PubMed Central

Tosa, Tetsuya; Sano, Ryujiro; Yamamoto, Kozo; Nakamura, Masatoshi; Ando, Katsuko; Chibata, Ichiro

1971-01-01

To produce an immunologically and enzymologically new type of l-asparaginase, 108 strains of bacteria were screened for enzyme production. As a result, 13 bacteria belonging to the genera Alcaligenes, Bacterium, and Proteus were found to produce l-asparaginases in high levels. Among these l-asparaginases, partially purified l-asparaginases from B. cadaveris and P. vulgaris showed antitumor activity. A partially purified l-asparaginase preparation of P. vulgaris did not react with the antibody of Escherichia colil-asparaginase on the Ouchterlony agar plate. Culture conditions for the production of l-asparaginase by P. vulgaris were investigated in detail. The enzyme was produced in high yields when cells were grown aerobically in a medium containing sodium fumarate and corn steep liquor. The addition of glucose or ammonium ion to the medium, however, resulted in depressed production of l-asparaginase. Under the optimum conditions, 3,700 international units of l-asparaginase was obtained from 1 liter of culture medium. Images PMID:5000866

The Resazurin-Agar Method - a Quick Test to Determine Water Quality

NASA Astrophysics Data System (ADS)

Huckfeldt, J.; Westphal, B.; Claußen, L.

2015-12-01

Rezasurin has been used as a smart tracer in stream ecosystems to indicate metabolic activity, specifically aerobic respiration by heterotrophic bacteria. Resazurin is a blue compound which is irreversibly reduced to the pink resorufin in the presence of aerobic bacteria. The degree and speed of colour change from blue to pink is a measure of the degree of oxygen consumption and thus an indirect indication of the concentration of aerobic bacteria in a given medium. A high concentration of bacteria in water indicates a bad water quality. In our work a method was developed using resazurin agar plates to find a quick and easy way for testing water quality and comparing concentrations of bacteria in freshwater and seawater samples. The theory was to concentrate bacteria from a defined volume of water sample onto polycarbonate filters (0.2 μm), which are then placed onto the resazurin agar plate. The presence of aerobic bacteria on the filter will reduce the resazurin in the agar and the compound changes its colour. First tests conducted with different dilutions of a pure culture of yoghurt bacteria showed promising results and confirmed the feasibility of the method. In a further assay, we used water samples from different water layers and different temperatures and were also able to observe differences in the concentration of bacteria, depending on these different environmental conditions.The assay was also successfully used with seawater samples, collected from 2 different stations at 3 different depths in the Baltic Sea (salinity=15). The discolouration of the plates showed good correlation with the oxygen concentrations in the water. The resazurin-agar plate method is economical and fast. Several samples could be investigated at the same time without sacrificing the reliability of the results. Thus it is a good pre-screening test for a quantitative evaluation of bacteria in a water sample.

[Antibacterial effect of self-etching adhesive systems on Streptococcus mutans].

PubMed

Zhang, Lu; Yuan, Chong-yang; Tian, Fu-cong; Wang, Xiao-yan; Gao, Xue-jun

2016-02-18

To investigate the antibacterial effect of different self-etching adhesive systems against Streptococcus mutans (S. mutans). Six reagents Clearfil(TM) SE Bond primer (SP), Clearfil(TM) SE Bond adhesive (SA),Clearfil(TM) Protect Bond primer (PP), which contained antibacterial monomer methacryloyloxydodecylpyridinium bromide (MDPB), ClearfilTM Protect Bond adhesive (PA), positive control chlorhexidine acetate [CHX, 1% (mass fraction)], and negative control phosphate buffer solution (PBS) were selected. They were mixed with S. mutans for 30 s respectively, then colony-forming units (CFU) were counted after incubated for 48 h on brain heart infusion (BHI) agar medium. The 6 reagents were applied to the sterile paper discs, and distributed onto the BHI agar medium with S. mutans and incubated for 24 h, then the inhibition zones were observed. CHX, PBS, PP, and SP were added on the dentin with artificial caries induced by S. mutans and kept for 30 s, then confocal laser scanning microscope (CLSM) was used to observe the live and dead bacteria after staining. The ratio of live to dead bacteria was calculated. PP+PA and SP+SA were applied on the dentin according to the manual and light cured. S. mutans were incubated on the samples for 2 h, ultrasonically treated and incubated on BHI agar medium for 48 h, then CFU was counted. The data were analyzed by non-parametric analysis and one-way ANOVA. Compared with PBS, the PP, SP, PA, SA and CHX showed the antibacterial effect on free S. mutans (P<0.05); SP and PP showed stronger antibacterial effect than PA, SA and CHX (P<0.05). CHX, SP and PP presented inhibition zones, while PBS, SA and PA did not. Compared with PBS, the CHX, SP and PP could lower the ratio of the live to dead bacteria significantly (P<0.05). Cured self-etching adhesive systems did not show any antibacterial effect on the free S. mutans. The primer of self-etching adhesives Clearfil(TM) SE Bond and Clearfil(TM) Protect Bond showed significant antibacterial effect on free and attached S. mutans. The adhesive only showed antibacterial effect on free S. mutans before light-cured polymerization. After being cured, the self-etching adhesive systems did not show antibacterial effect anymore.