Showing papers on "Agar plate published in 2001"

partners transport proteins from the ER into the cytosol

Abstract: 6colony-forming units per 10 ml -1 in SFM supplemented with 0.3 mM calcium chloride, and inoculated onto the surface of the tissue. After inoculation, we incubated tissue samples at 37 8C with 5% CO2 and no supplemental humidity. Transwells containing the inoculated tissue samples were transferred to fresh blood agar every 2 h. The blood agar plates were then incubated overnight at 37 8C for enumeration of colony-forming units representing the number of organisms emerging from the basal surface of the tissue.

Improved method for detection of Vibrio parahaemolyticus in seafood.

Abstract: We have developed a new, effective procedure for detecting Vibrio parahaemolyticus in seafoods using enrichment and plating onto a chromogenic agar medium. Samples were cultured in salt Trypticase soy broth, which is a nonselective medium, and then a portion of the culture was cultured with salt polymyxin broth, which is a selective medium for V. parahaemolyticus. This two-step enrichment was more effective than the one-step enrichment in salt polymyxin broth alone. The enrichment cultures were then plated onto a new chromogenic agar containing substrates for beta-galactosidase. The V. parahaemolyticus colonies developed a purple color on this growth medium that distinguished them from other related bacterial strains. V. parahaemolyticus was isolated more frequently from naturally contaminated seafood samples using the chromogenic agar than thiosulfate citrate bile salts sucrose agar medium, which is currently used for the isolation of V. parahaemolyticus. Our findings suggest that this new enrichment and isolation scheme is more sensitive and accurate for identifying V. parahaemolyticus in seafood samples than previously used methods.

Improved, Low-Cost Selective Culture Medium for Actinobacillus actinomycetemcomitans

Abstract: Actinobacillus actinomycetemcomitans is considered to be one of the major oral putative pathogens, especially in cases of juvenile periodontitis. This microorganism requires nutritionally complex media for growth, and therefore the media for its primary isolation usually include blood agar or serum in their base. In this study we present a new medium, Dentaid-1, which improves the detection of A. actinomycetemcomitans in periodontal samples. In its composition, blood and serum have been omitted, hence reducing its cost and making it a more restrictive medium against the growth of other microorganisms with high nutritional requirements. The growth yields of pure cultures of the bacteria on Dentaid-1 were comparable to those on nonselective blood agar. Moreover, clinical efficacy was evaluated in subgingival samples from 77 subjects with adult periodontitis. Dentaid-1 detected A. actinomycetemcomitans in 24 subjects, while a previously described tryptic soy-serum-bacitracin-vancomycin agar detected the microorganism in only 19 subjects (79.1%). Dentaid-1 is a low-cost, noninhibitory formula for the improved diagnosis and monitoring of patients subgingivally infected by this important oral putative pathogen.

Host range of a deleterious rhizobacterium for biological control of downy brome

Abstract: Pseudomonas fluorescens strain D7 (P. f. D7; NRRL B-18293) is a root-colonizing bacterium that inhibits downy brome (Bromus tectorum L. BROTE) growth. Before commercialization as a biological control agent, strain D7 must be tested for host plant specificity. Agar plate bioassays in the laboratory and plant–soil bioassays in a growth chamber were used to determine the influence of P. f. D7 on germination and root growth of 42 selected weed, cultivated or native plant species common in the western and midwestern United States. In the agar plate bioassay, all accessions of downy brome were inhibited by P. f. D7. Root growth of seven Bromus spp. was inhibited an average of 87% compared with that of controls in the agar plate bioassay. Root growth of non-Bromus monocots was reduced by 0 to 86%, and only 6 out of 17 plant species were inhibited 40% or greater. Among all plant species, only downy brome root growth from two accessions was significantly inhibited by P. f. D7 in plant–soil bioassays (42 a...

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

Abstract: 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.

Survey of yeast mastitis in dairy herds of small-type farms in the Lublin region, Poland.

Abstract: The purpose of this study was to isolate fungi from the quarter milk of cow udders in 9 dairy farms. The samples were collected between May 1996 and April 2000 in the Lublin region. Six hundred and four milk samples collected from the quarters of 172 cows with clinical and subclinical mastitis were examined. Milk samples were plated as soon as possible on blood agar (BA), Mac Conkey agar, aesculin-tallium acetate-crystal violet blood agar, and Sabouraud agar with chloramphenicol and gentamicin. Fifty eight (9.6%) of the samples were positive for fungi. All of the fungal isolates were yeasts of the genera Candida, Rhodotorula and Trichosporon. We also isolated Streptococcus agalactiae (4.9%), Str. spp.(6.4%), Staphylococcus aureus (10.4%), coagulase-negative staphylococci-CNS (36.6%), Escherichia coli (3.5%), other microorganisms (2.6%) and no growth (25.8%).

Mercury‐induced genes in Arabidopsis thaliana: identification of induced genes upon long‐term mercuric ion exposure

Abstract: Mercuric-ion-induced gene expression was studied in Arabidopsis thaliana Columbia wild type. Rosettes of plants grown for 21 d on agar medium supplemented with 20, 30 and 40 μM HgCl 2 were pooled and used to isolate cDNAs of induced genes by suppression subtractive hybridization. Of the 576 clones isolated initially, 31 turned out to be mercury-induced by Northern hybridization. However, kinetic studies using cDNA arrays clearly showed that seven genes were exclusively mercuric-ion-induced, 14 were induced by mercury but also affected by a diurnal rhythm, and 10 clones were only modulated by the day-night cycle. The expression levels of the metal-induced genes increased from 1.5-fold to 10-fold. Functional classification resulted in genes encoding proteins for the photosynthetic apparatus and for the antioxidative system. In addition, unexpected genes, whose connection to mercury ion stress is not evident, were identified.

Use of GBS media for rapid detection of group B streptococci in vaginal and rectal swabs from women in labor

Abstract: In order to evaluate the differences in efficacy, three methods were used to detect group B streptococci (GBS) in women in labor. The recommended method for detecting GBS carriage in pregnant women is to culture vaginal and anorectal swabs in a selective broth medium and to subculture them onto blood agar. This method was compared with the use of GBS agar and GBS broth, both of which produce an orange pigment in response to GBS strains. A total of 319 women in labor were screened. Among the 638 specimens tested, 134 (21%) were positive in the selective Todd-Hewitt broth subcultured onto sheep blood agar, 133 (20.8%) were positive on the GBS agar and 126 (19.7%) were positive in the GBS broth. Altogether, 89 (27.9%) women in labor were found to be colonized with GBS; 87 (97.8%) of them were identified as carriers using the Todd-Hewitt broth, 87 (97.8%) with the GBS agar and 86 (96.6%) with the GBS broth. These results indicate that both GBS agar and GBS broth are reliable methods that can be used to screen for maternal and neonatal GBS colonization.

Fluconazole Disk Diffusion Test with Methylene Blue- and Glucose-Enriched Mueller-Hinton Agar for Determining Susceptibility of Candida Species

Abstract: A 25-microg fluconazole disk diffusion test using a Mueller-Hinton agar plate containing 2% glucose and 5 microg of methylene blue/ml (GM-MH) was compared to the macrodilution reference method for 210 Candida species. The GM-MH agar plate was read at 24 h. The predictive values of disks with susceptible, intermediate, and resistant results on the GM-MH agar plate at 24 h were 97.1, 56.3, and 76.5%, respectively.

Xbp1-mediated repression of CLB gene expression contributes to the modifications of yeast cell morphology and cell cycle seen during nitrogen-limited growth.

Abstract: Yeast cells undergo morphological transformations in response to diverse environmental signals. One such event, called pseudohyphal differentiation, occurs when diploid yeast cells are partially starved for nitrogen on a solid agar medium. The nitrogen-starved cells elongate, and a small fraction form filaments that penetrate the agar surface. The molecular basis for the changes in cell morphology and cell cycle in response to nitrogen limitation are poorly defined, in part because the heterogeneous growth states of partially starved cells on agar media are not amenable to biochemical analysis. In this work, we used chemostat cultures to study the role of cell cycle regulators with respect to yeast differentiation in response to nitrogen limitation under controlled, homogeneous culture conditions. We found that Clb1, Clb2, and Clb5 cyclin levels are reduced in nitrogen-limited chemostat cultures compared to levels in rich-medium cultures, whereas the Xbp1 transcriptional repressor is highly induced under these conditions. Furthermore, the deletion of XBP1 prevents the drop in Clb2 levels and inhibits cellular elongation in nitrogen-limited chemostat cultures as well as inhibiting pseudohyphal growth on nitrogen-limited agar media. Deletion of the CLB2 gene restores an elongated morphology and filamentation to the xbp1Δ mutant in response to nitrogen limitation. Transcriptional activation of the XBP1 gene and the subsequent repression of CLB gene expression are thus key responses of yeast cells to nitrogen limitation.

Mitomycin‐supplemented washed blood agar for the isolation of Shiga toxin‐producing Escherichia coli other than O157:H7

Abstract: Aims: Isolation and recognition of the prominent Shiga toxin (Stx)-producing strains of Escherichia coli (STEC) serovar O157:H7 can be confirmed easily by their late fermentation of sorbitol and lack of β-glucuronidase activity, but there has been no culture method of choice for detecting non-O157 STEC strains because of their biochemical diversity. Apart from Stx, many STEC strains produce enterohaemolysin (Ehly) regardless of their serovars. Methods and Results: Although washed blood agar media, with or without the addition of antibiotics (vancomycin, cefixime, and cefsulodin) (WBA and WBVCCA), have been used to detect Ehly, a proportion of STEC strains consistently failed to produce haemolysin on these media. Washed blood agar medium was therefore studied further in order to increase the yield of strains producing Ehly. Conclusions: It was found that the addition of 0·5 μg ml–1 of mitomycin C to the agar medium (WBMA) markedly increased the number of such strains. Thus, of 185 STEC strains comprising 95 O157 and 90 non-O157 STEC consisting of 34 serovars. Ninety-seven per cent of these strains produced haemolysis on WBMA, compared with only 76% and 83%, respectively, on WBA and WBVCCA. Significance and Impact of the Study: The appearance of the Ehly zone of haemolysis that was easily distinguishable from that of α-haemolysin was enhanced by the incorporation of mitimycin C into washed-blood medium.

Bacterial flora of Hirudo medicinalis and their antibiotic sensitivities in the Middle Black Sea Region, Turkey.

Abstract: The rate of infectious complications of leech therapy is almost 20% because Hirudo medicinalis has endosymbiotic bacteria. The aim of this study was to determine the bacterial flora of H. medicinalis and their antibiotic sensitivities in a region in Turkey. Sixteen adult leeches were collected in Middle Black Sea region, Turkey. They were rubbed onto blood agar plates directly under ether anesthesia to obtain surface cultures. They were then killed to obtain mouth and gut cultures. Culture swabs were applied to blood agar, eosin methylene blue agar, and ampicillin blood agar plates. Gut contents were applied to blood culture medium as well. Bacteria were isolated in 15 of 16 leech surfaces, in 7 of 16 mouths, and in 15 of 16 guts. Isolated bacteria were identified with Analytical Profile Index 32 E and Analytical Profile Index 20 NE (fermentative and nonfermentative respectively). Most common types of cultured bacteria were Aeromonas hydrophila (N = 25), Ochrobacter anthropi (N = 23), nonfermenting Gram-negative rods (N = 12), Acinetobacter lwoffi (N = 3), and A. sobria (N = 2) in 73 isolates. A standard disk diffusion test was performed on isolated bacteria. All isolates were 100% susceptible to ciprofloxacin, cefotaxime, ceftazidime, gentamicin, and trimethoprim/sulfamethoxazole. Because leeches are carriers of Aeromonas and other bacteria, appropriate antibiotic prophylaxis should be administrated to the patient who needs leech therapy. Antibacterial agents can be determined by the resistance pattern of the bacterial flora of regional H. medicinalis.

Effect of garlic bulb extract on the growth and enzymatic activities of rhizosphere and rhizoplane fungi.

Abstract: Eighteen fungal species were isolated from rhizospheric soil and rhizoplane samples of three plant crops in southern Iraq. The fungal isolates were examined for the activities of four enzymes (amylase, cellulase, phenoloxidase, and protease), as well as their growth, against crude garlic extract added to the culture agar medium. A high reduction or inhibition of enzymatic activities was observed for the fungi treated with garlic extract compared with untreated fungal cultures. However, most of the species showed inhibition of enzymes due to the effect of garlic extract. The growth of the fungal species was also remarkably reduced by the garlic extract.

Studies on the significance of positive bacterial semen cultures in male fertility in Nigeria

Abstract: Early warning signals (EWS) of altered reproductive potential may be very important in the prevention and management of male infertility. The presence of bacteria in semen (bacterisemia) may be an EWS. This was evaluated by determining the incidence of bacteria in semen of males with fertility problems in Benin City by culturing their semen. Diluted semen samples were cultured on blood agar chocolate agar MacConkey agar nutrient agar and sabouraud dextrose agar slants for the isolation of microorganisms. Colonies of a single type of microorganism (>1000 cfu/ml) were picked for identification and sensitivity tests using antibacterial agents. Each semen sample was further processed for spermatozoal morphology and motility presence of peroxidase-positive cells and other accompanying cells. Correlative studies on the relationship between bacterisemia and semen/spermatozoa variables such as total number and motility were also done. Pathogenic microorganisms were present in 78/163 (47.1%) semen samples. The microbial isolates were Staphylococcus aureus 35 (43.7%) Klebsiella species 22 (28.2%) Escherichia coli 9 (11.5%) and Candida albicans 6 (7.7%). The bacterial isolates were most sensitive to ceftazidime and pefloxacin and least to amoxycillin and tetracycline. There was a positive correlation (r = 0.9774) between azoospermia in males and presence of Candida albicans in semen as well as between the presence of microorganisms and poor semen quality (r = 0.8563) and the presence of microorganisms and reduced motility (r = 0.8246). Presence of pathogenic microorganisms in semen which may be related to a breach in the integrity of the blood-testes barrier may provide early warning signals of impairment of male fertility. (authors)

Evaluation of the Mastalex latex agglutination test for methicillin resistance in Staphylococcus aureus grown on different screening media

Abstract: The Mastalex MRSA latex agglutination method was evaluated with 52 methicillin-resistant and 27 methicillin-susceptible strains of Staphylococcus aureus grown on various media. All tests were correct with colonies grown on blood agar with or without oxacillin 2 mg/L. Tests on colonies grown on mannitol salt agar were less reliable, although addition of oxacillin 2 mg/L improved performance. One of the 26 MRSA which grew on Baird-Parker medium with 8 mg/L ciprofloxacin gave a false-negative result. Agglutination was faster when strains were grown on media with oxacillin. The method would be particularly useful for urgent confirmation of resistance.

Effects of Salinity Stress on the Seminal Root Tip Ultrastructures of Rice Seedlings (Oryza sativa L.)

Abstract: Growth and structural changes in the seminal root tip of rice seedlings (Oryza sativa L. cv. Nipponbare) in response to NaCl salinity were studied. Seedlings were grown in agar medium with 0, 0.1, 0.3, 1.0, 2.0 and 3.0% NaCl(agar culture), and in water with 0, 0.01, 0.03, 0.06 and 0.1% NaCl (water culture). Seedling growth was significantly suppressed by higher concentrations of NaCl. The effect of NaCl appeared faster in water culture than in agar culture. In both agar and water cultures, root growth was markedly suppressed over shoot growth. Under saline conditions, epidermis, cortex and root cap cells appear to be damaged to a greater extent than the meristem and stelar cells. The most notable ultrastructural change in response to salinity was the development and increment of vacuoles, which seem to provide a space for accumulation of excess ions. Many electron dense deposits were observed in the larger vacuoles of the epidermal and cortical cells. Under saline conditions, cell wall thickening ...

Purification and properties of a beta-1,6-glucanase from Streptomyces sp. EF-14, an actinomycete antagonistic to Phytophthora spp.

Abstract: Extracellular enzymes with glucanase activities are an important component of actinomycete-fungus antagonism. Streptomyces sp. EF-14 has been previously identified as one of the most potent antagonists of Phytophthora spp. A beta-1,6-glucanase (EC 3.2.1.75; glucan endo-1,6-beta-glucosidase) was purified by four chromatographic steps from the culture supernatant of strain EF-14 grown on a medium with lyophilized cells of Candida utilis as main nutrient source. The glucanase level in this medium followed a characteristic pattern in which the rise of beta-1,6-glucanase activity always preceded that of beta-1,3-glucanase. The molecular mass of the enzyme was estimated to be 65 kDa and the pI approximately 5.5. It hydrolyzed pustulan by an endo-mechanism generating gentiobiose and glucose as final products. Laminarin was not hydrolyzed indicating that the enzyme does not recognize beta-1,6-links flanked by beta-1,3-links. No significant clearing of yeast cell walls in liquid suspensions or in agar plates was observed indicating that this beta-1,6-glucanase is a non-lytic enzyme. This is the first beta-1,6-glucanase characterized from an actinomycete.

Phenotypic expression of Vogesella indigofera upon exposure to hexavalent chromium, Cr6+.

Abstract: A eubacterium producing a blue pigment was isolated from a drinking water filter, and subsequently identified as Vogesella indigofera. This bacterium was further investigated for its morphological and biochemical characteristics after exposure to hexavalent chromium, Cr6+. The threshold Cr6+ concentration inhibiting the pigment production by V. indigofera was 200–300 μg ml−1 in liquid cultures of nutrient broth and 100–150 μg ml−1 on nutrient agar plates. The Cr6+ concentration preventing V. indigofera growth was 300–400 μg ml−1 in liquid cultures, but greater than 150 μg ml−1 on agar plates. Moreover, rugose colonies without the blue pigmentation were observed on agar plates amended with 150 (μg Cr6+) ml−1. The biochemical utilization profiles of the colonies without pigmentation did not differ from the original pigment-producing ones, indicating phenotypic plasticity of this bacterium. The difference of phenotypic expression of V. indigofera under various Cr6+ concentrations might have potential application as a pollution bioindicator for heavy metals.

Simple and rapid method for screening antimicrobial activities of bifidobacterium species of human isolates

Abstract: The objective of this research was to develop a rapid procedure for the screening antimicrobial activities of Bifidobacterium species of human isolates A bifidobacteria selective medium BIM-25 agar was modified by adding of 05 g/L cysteine-hydrochloride, 15 g/L lithium chloride, 10 g/L beef extract, and 5 mL/L Tween 20 This medium was inoculated (45C) with diluted fecal material from 5 subjects and overlaid into 01% Tween 20 BHI agar plates Plates were incubated in anaerobic chamber at 37C for 48 h Plates were then inverted to allow the two layers of agar to fall into the petri lid BHI soft agar (045%) containing Micrococcus luteus (as indicator) was overlaid onto the other layers in the petri dish Plates were incubated at 37C overnight and zone of growth inhibition was observed This method is simple and rapid whereas the original method for screening of antimicrobial activities of bifidobacteria is a more time consuming and cumbersome procedure

Seed germination of Haemaria discolor var. dawsoniana and the use of mycorrhizae

Abstract: Haemaria discolor var. dawsoniana is an orchid with beautiful red-green striped leaves, which can be used as an ornamental or for medical purposes. Seeds of H. discolor in Murashige & Skoog (MS) medium or Hyponex (#1,2 and 3) agar media germinated. In MS medium, the germination rate was optimal in 20 and 30 g/l of sucrose, but the germination rate was only 34%. However, in Hyponex #1 and H. #3 agar media, the germination rate was 60-63% within 60 days at room temperatures. Vegetative growth of the H. discolor seedlings in phytotrons was best with a 30/25°C day/night temperature regimes. Light microscopy and SEM observation showed that the cortex of roots could be infected by the orchid mycorrhizal fungi (OMF), and typical orchid mycorrhizal structures of tolypophagy with young and old pelotons were present in the same root. Of seven Rhizoctonia spp. isolates, only R.01 and R.02 enhanced seed germination of H. discolor, but the germination rate was only 5% and 37% respectively. For the inoculation of Rhizoctonia sp. in vitro, oat meal agar (OMA) medium was a simple effective medium. The growth of the H. discolor seedlings in vitro was enhanced by the inoculation of Rhizoctonia sp. encoded as R.01 and R.02. For practical use, seeds of H. discolor should be asymbiotically germinated in Hyponex # 3 agar medium with 20 g of sucrose, and the protocorms or small seedlings should be inoculated near the roots with R.01 or R.02 isolate after subculturing into OMA medium.

A new isolation method for labyrinthulids using a bacterium, Psychrobacter phenylpyruvicus.

Abstract: A new isolation method for labyrinthulids, marine microbes with spindle-shaped vegetative cells and gliding movement, is presented. The method for isolating labyrinthulids has been found to be more difficult and less reproducible than that for thraustochytrids, classified in the same order. So far serum seawater agar fortified with antibiotics has been proposed to be the best for isolation of labyrinthulids. The method presented here involves placing plant samples on an agar medium on which a marine bacterium, Psychrobacter phenylpyruvicus, has been grown. The new method, which utilizes fallen mangrove leaves as source material, was more than twice as effective as isolation agar medium without the bacterium. The increased effectiveness appears to derive partly from the bacterial colonies' delaying extension of fungal mycelium. The bacterium was more effective for the isolation of labyrinthulids than either the bacterium Shewanella sp. or the yeast Rhodotorula rubra.