Influence of whey permeate and millet as substrates on thermotolerance of Beauveria bassiana and Metarhizium anisopliae conidia during storage

Conidia from Beauveria bassiana and Metarhizium anisopliae, produced in quarter-strength Sabouraud dextrose agar amended with yeast extract, whey permeate, and millet agar, were exposed to 45°C for 90 min prior to and at 30 d post-storage at 25°C. B. bassiana produced in whey permeate or millet had better thermotolerance than the other treatments after the storage.     

Nosocomial Invasive Infection Caused by Cunninghamella bertholletiae: Case Report

Human infection by Cunninghamella bertholletiae occurs almost exclusively in immunocompromised patients. Infections due to this microorganism have been most frequently diagnosed in patients with hematological malignancies, with neutropenia and in diabetes mellitus patients. This work reports a case of fungal infection by Cunninghamella bertholletiae isolated from blood in a man with a complex clinical picture, involving diabetes and pharmacological immunosupression. Blood culture at room temperature and at 37 °C on Sabouraud agar grew a single mold with characteristic properties of Cunninghamella. In the microscopic morphology, were found wide, non-septate, branching hyphae with erect sporangiophores terminated in swollen vesicles and sporangioles borne off the vesicles. C. bertholetiae was identified after subculture on Sabouraud dextrose agar at 45 °C. The patient died 15 days after the beginning of amphotericin B therapy.     

Effects of Temperature and Culture Media on Vegetative Growth of an Entomopathogenic Fungus <Emphasis Type="Italic">Isaria</Emphasis> sp. (Hypocreales: Clavicipitaceae) Naturally Affecting the Whitefly, <Emphasis Type="Italic">Bemisia tabaci</Emphasis> in Texas

The effects of temperature and mycological media on mycelial growth and estimates of spore production of an indigenous entomopathogenic fungus, Isaria sp., found during natural epizootics on whiteflies in the Lower Rio Grande Valley of Texas, were investigated. The radial growth (mm/day) of Isaria sp. as a function of temperature fits a linear model; with faster growth on Sabouraud dextrose agar with yeast extract, SDAY slopes (0.23) than on Sabouraud maltose agar, SMA slopes (0.14) from 20 to 30°C, with an optimal temperature of 30°C (SDAY: 4.1 mm, SMA: 3.1 mm). Moderate growth occurred at 25°C (SDAY: 3.4 mm, SMA: 2.7 mm). Growth was lowest at 20°C (SDAY: 1.9 mm, SMA: 1.8 mm). No fungal growth was observed at 35°C and 40°C. However, when Isaria sp. was exposed to 35°C for the first 7 days, it could recover and grow when transferred to 25°C (SDAY: 3.5 mm, SMA: 2.8 mm). No recovery or growth occurred after transfer from 40°C to 25°C. The average conidial production on SDAY after 20 days incubation at 25°C and a photoperiod of 14:10 h light: dark was 1.2 × 10⁸ conidia/cm² with 100% spore viability. When compared on SDAY at 25°C, the radial growth rate of I. javanica ex type CBS 134.22 (5.1 mm/day) was greater than seven Isaria isolates including Isaria sp.; but maximum growth rates were similar among all related Isaria isolates (90–97%). The Isaria sp. fungus tolerates high temperatures (35°C), suggesting that it is naturally selected for the subtropical semi-arid environment, where it could serve as an important natural control agent of the sweet potato whitefly, Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) biotype B, one of the most invasive and economically damaging insects to agriculture. Mention of trade names or commercial products in this publication is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the U. S. Department of Agriculture.     

Mycobiota and molecular detection of Aspergillus flavus and A. parasiticus aflatoxin contamination of Strawberry (Fragaria ananassa Duch.) fruits

Strawberry fungi were isolated from fresh fruits and juice on the two types of media (Sabouraud dextrose agar, SDA and potato-dextrose agar, PDA) at 28 °C. Nineteen fungal species belong to 12 genera were isolated from fruits and juice on both isolation media. The most common fungal genera and species were Aspergillus flavus, A. niger, Mucor racemosus, Neurospora crassa, Penicillium chrysogenum, Rhizopus stolonifer and Trichoderma harzianum. Twenty A. flavus and A. parasitics isolates were assayed for their abilities to produce aflatoxins. The concentration of aflatoxins ranged between 25.8–75.2 and 23.6–71.1 ng/ml at 350 and 365 nm, respectively. Among A. flavus and A. parasiticus strains tested, aflatoxin B contributed 30–60% of total isolates. However, G type contributed 85–90%. The R_f values of B₁, B₂, G₁ and G₂ were 0.79, 0.61, 0.44 and 0.32, respectively. High-performance liquid chromatography analysis of extracts revealed the presence of aflatoxins with variable levels.     

Occurrence of itraconazole-tolerant micromycetes in the soil and food products

Unexpected pathogens from the environment represent considerable risk for humans with impaired health. We examined the occurrence of itraconazole tolerant micromycetes in soil and in maize products. Five concentrations of itraconazole (2.5–12.5 μg/mL) selected according to known treatment schedules for human patients were incorporated into Sabouraud agar with chloramphenicol and Rose Bengal and diluted samples were inoculated onto the agar surface. After 7-d growth at 22°C colonies ofAlternaria sp.,Aspergillus clavatus. A. glaucus group,A. flavus. A, fumigatus, A. niger group,A. ochraceus group,A. ochraceus, Chœtomium sp.,Cladosporium cladosporioides. Cylindrocarpon sp.Doratomyces sp.,Fusarium sp.,F. moniliforme. F. oxysporum. F. solani, F. subglutinans. Marianaea elegans, Mortierella sp.,Mucor sp.,Myrothecium sp.,Penicillium sp.,Rhizopus sp.,Scopulariopsis brevicaulis. Sepedonium sp.,Stachybotrys chartarum. Stemphylium sp.,Torula humicola andTrichoderma viride were isolated.     

Thermal ecotypes of amphi-Atlantic algae. II. Cold-temperate species (Furcellaria lumbricalis andPolyides rotundus)

Two species of cold-temperate algae from the North Atlantic Ocean,Polyides rotundus andFurcellaria lumbricalis, were tested for growth and survival over a temperature range of −5 to 30 °C. In comparisons of eastern and western isolates, bothF. lumbricalis, a North Atlantic endemic, andP. rotundus, a species having related populations in the North Pacific, were quite homogeneous.F. lumbricalis tolerated −5 to 25°C and grew well from 0 to 25°C, with optimal growth at 10–15 °C.P. rotundus tolerated −5 to 27°C, grew well from 5 to 25°C, and had a broad optimal range of 10–25°C. Both species tolerated 3 months in darkness at 0°C. In neither case could any geographic boundary be explained in terms of lethal seasonal temperatures, suggesting that these species are restricted in distribution by strict thermal and/or daylength requirements for reproduction. The hypothesis that northern species are more homogeneous than southern taxa in terms of thermal tolerance was supported. A second hypothesis, that disjunct cold-temperate species should be more variable than pan-Arctic species, was not supported.     

Aflatoxin and Ochratoxin Production by <Emphasis Type="Italic">Aspergillus</Emphasis> Species Under Ex Vivo Conditions

Aspergillus species are increasingly important human pathogens. It is not known whether toxic metabolites of many of these pathogenic species can act as virulence factors in aspergillosis. We examined isolates of aflatoxin and ochratoxin-producing species for toxin production in ex vivo conditions. Seven of the 21 aflatoxin-producing isolates screened produced aflatoxin at 35 and 37°C on the general medium yeast extract sucrose agar (YES). However, none of them produced toxin at these temperatures on brain heart infusion agar (BHA), a medium that mimics human tissue, or on BHA with modified pH or sugar levels. Six of the 12 ochratoxin-producing isolates examined produced toxin at 35°C on YES. All three isolates of A. alliaceus produced ochratoxin on BHA or modified BHA at 37°C. One strain of A. pseudoelegans produced a minute amount of ochratoxin on modified BHA at 37°C. These data indicate that aflatoxin is an unlikely virulence, factor but that ochratoxin may be a potential virulence factor in aspergillosis.     

Impact of some environmental factors on growth and production of ochratoxin A of/by <Emphasis Type="Italic">Aspergillus tubingensis,A. niger</Emphasis>, and <Emphasis Type="Italic">A. carbonarius</Emphasis> isolated from moroccan grapes

The effects of temperature, water activity (a_w), incubation time, and their combinations on radial growth and ochratoxin A (OTA) production of/by eight Aspergillus niger aggregate strains (six A. tubingensis and two A. niger) and four A. carbonarius isolated from Moroccan grapes were studied. Optimal conditions for the growth of most studied strains were shown to be at 25°C and 0.95 a_w. No growth was observed at 10°C regardless of the water activity and isolates. The optimal temperature for OTA production was in the range of 25°C∼30°C for A. carbonarius and 30°C∼37°C for A. niger aggregate. The optimal a_w for toxin production was 0.95∼0.99 for A. carbonarius and 0.90∼0.95 for A. niger aggregate. Mean OTA concentration produced by all the isolates of A. niger aggregate tested at all sampling times shows that maximum amount of OTA (0.24 μg/g) was produced at 37°C and 0.90 a_w. However, for A. carbonarius, mean maximum amounts of OTA (0.22 μg/g) were observed at 25°C and 0.99 a_w. Analysis of variance showed that the effects of all single factors (a_w, isolate, temperature and incubation time) and their interactions on growth and OTA production were highly significant.     

Development of a high-throughput screening method for recombinant Escherichia coli with intracellular dextransucrase activity

To efficiently engineer intracellular dextransucrase (DSase) expression in Escherichia coli, a high-throughput screening method was developed based on the polymer-forming activity of the enzyme. Recombinant E. coli containing the Leuconostoc citreum DSase (LcDS) gene was grown on Luria-Bertani agar plates, containing 2% sucrose, at 37°C for 8 h. The plates were then evenly overlaid with 0.6% soft agar, containing 1.2 mg/ml D-cycloserine, and incubated at 30°C to allow gradual cell disruption until a dextran polymer grew through the overlaid layer. A significant correlation between dextran size and enzyme activity was established and applied for screening truncated mutants with LcDS activity.     

First Isolation of <Emphasis Type="Italic">Candida dubliniensis</Emphasis> from Oral Cavities of Dermatological Patients in Nanjing,China

Candida dubliniensis is an emerging pathogen capable of causing both superficial and systemic infections. Although C. dubliniensis and C. albicans are phenotypically similar, the two species differ in terms of epidemiology and the ability to rapidly develop resistance to fluconazole. C. dubliniensis is primarily associated with oral candidiasis of human immunodeficiency virus (HIV)-infected individuals. In this study, we describe the first recovery of C. dubliniensis from oral cavities of non-HIV-infected patients with dermatological diseases in Nanjing, China. The isolates were phenotypically characterized as C. dubliniensis by their production of brown rough colonies and chlamydospores on tobacco agar and their inability to grow on hypertonic Sabouraud dextrose agar or to assimilate xylose or α-methyl-d-glycoside. The species identification was subsequently confirmed by amplification and sequencing of the internal transcribed spacer region (ITS). Three C. dubliniensis isolates out of 128 (2.3%) presumptive C. albicans/C. dubliniensis ones were finally identified. Further sequence analysis separated the three isolates into two of the four reported ITS genotypes. Antifungal susceptibility testing showed that they were susceptible to fluconazole, itraconazole, voriconazole, micafungin, and amphotericin B. This study adds to the accumulating evidence that C. dubliniensis is widely distributed in non-HIV-infected populations worldwide.     

Simple rapid identification of Candida albicans with emphasis on the differentiation between Candida albicans and Candida stellatoidea

Summary Subcultures ofC. albicans, made from Sabouraud agar, grown at room temperature for 48 hours, were inoculated into a 10 times saline dilution of Sabouraud liquid medium and left in the incubator for 45–60 minutes at 37° C, transferred to corn meal agar plates and incubated at 37° C for 18–24 hours.Small portions of the surface agar containing the yeasts from these plates were pressed under cover glasses and examined under the oil immersion lens.Under these conditions,C. albicans cultures were observed to produce only yeast-like cells, whereasC. stellatoidea cultures contained predominantly abundant, long, thin mycelia.     

Cultural and physiological characteristics of Stemphylium lycopersici causing leaf blight disease on vegetable crops

During 2011–2012, an extensive leaf spot disease caused by Stemphylium lycopersici was observed on vegetable crops including, tomato, eggplant, pepper and lettuce in major vegetable-growing regions of Malaysia. Four isolates of S. lycopersici obtained from each vegetable crop were used to determine cultural and physiological characteristics. The variations were found in colony colour (pale to light grey or light as well as the brown), texture (cottony or mycelium flat), shape (regular with concentric growth rings or irregular) and pigmentation (yellow or deep red) of the cultures. The optimum temperature for the conidial germination and mean radial growth of the isolates was 25?°C, and the radial growth of the isolates was maximal on V-8 juice agar followed by potato carrot agar. The maximum sporulation of S. lycopersici isolates was observed on V-8 juice agar media under 12/12 h light/darkness photoperiod at 25?°C.     

Based on biochemical and physiological behavior,where isAspergillus egyptiacus better placed?

Physiological and biochemical properties were tested in 45 isolates ofAspergillus egyptiacus (16 isolates),Emericella nidulans (16) andAspergillus versicolor (13). The three fungal species exhibited common and similar features. The big similarity betweenA. egyptiacus andE. nidulans was greater than betweenA. egyptiacus andA. versicolor. It included the inability to produce base either from sodium citrate or lactic acid media, growth at 45 °C (thermophilicity), and production of very similar pigmentations onAspergillus flavus andparasiticus agar.A. egyptiacus is therefore better placed in theAspergillus nidulans-Emericella assemblage.     

Thermal ecotypes of amphi-Atlantic algae. I. Algae of Arctic to cold-temperate distribution (Chaetomorpha melagonium,Devaleraea ramentacea andPhycodrys rubens)

Three species of Arctic to cold-temperate amphi-Atlantic algae, all occurring also in the North Pacific, were tested for growth and/or survival at temperatures of −20 to 30°C. When isolates from both western and eastern Atlantic shores were tested side-by-side, it was found that thermal ecotypes may occur in such Arctic algae.Chaetomorpha melagonium was the most eurythermal of the 3 species. Isolates of this alga were alike in temperature tolerance and growth rate but Icelandic plants were more sensitive to the lethal temperature of 25°C than were more southerly isolates from both east and west. With regard toDevaleraea ramentacea, one Canadian isolate grew extraordinarily well at −2 and 0°C, and all tolerated temperatures 2–3°C higher than the lethal limit (18–20°C) of isolates from Europe. ConcerningPhycodrys rubens, both eastern and western isolates died at 20°C but European plants tolerated the lethal high temperature longer, were more sensitive to freezing, and attained more rapid growth at optimal temperatures. The intertidal species,C. melagonium andD. ramentacea, both survived freezing at −5 and −20°C, at least for short time periods.C. melagonium was more susceptible thanD. ramentacea to desiccation. Patterns of thermal tolerance may provide insight into the evolutionary history of seaweed species.     

Diversity in biofilm formation and production of curli fimbriae and cellulose of Salmonella Typhimurium strains of different origin in high and low nutrient medium

The biofilm forming behavior of 51 Salmonella Typhimurium strains was determined in Tryptone Soya Broth (TSB) and 20 times diluted TSB (1/20TSB) at 25°C and 37°C. The results indicated that biofilm forming behavior is influenced by environmental conditions and associated with the origin of the strains. Clinical, outbreak-associated and retail product isolates showed dense biofilm formation in both media at 25°C, and in TSB also at 37°C. However, industrial isolates only showed dense biofilm formation in 1/20TSB at 25°C. By enumeration of biofilm cells, LIVE/DEAD staining and SEM analysis of biofilms it was found that the ratio of cells and extracellular matrix is affected by environmental conditions. Indeed, the genes involved in curli fimbriae and cellulose production are highly induced during biofilm formation at 25°C in 1/20TSB. This indicates that these are important matrix components during biofilm formation in 1/20TSB at 25°C and that other factors contribute to biofilm formation of clinical, outbreak-associated and retail product isolates at 37°C and/or nutrient-rich conditions.     

Diagnostic properties of three conventional selective plating media for selection of <Emphasis Type="Italic">Bacillus cereus</Emphasis>, <Emphasis Type="Italic">B. thuringiensis</Emphasis> and <Emphasis Type="Italic">B. weihenstephanensis</Emphasis>

The aim of this study was to assess the diagnostic properties of the two selective plating media and a chromogenic medium for identification of Bacillus cereus. The 324 isolates were B. cereus (37%), Bacillus weihenstephanensis (45%) or Bacillus thuringiensis (18%), as identified by a new combination of techniques. All isolates were growing on mannitol–egg yolk–polymyxin agar (MYP), and they did not form acid from mannitol. However, a significant lower number of B. thuringiensis isolates did not show lecithinase activity. All isolates were also growing on polymyxin–egg yolk–mannitol–bromothymol blue agar (PEMBA); however, 11% isolates indicated that they did produce acid from mannitol, and 15% isolates did not show any lecithinase activity. Five of the isolates did not grow at all on the chromogenic agar, and 14 of the growing isolates were β-glucosidase negative. It is concluded that the two recommended selective plating media MYP and PEMBA for detection of B. cereus group bacteria both have their limitations for identification of some B. cereus, B. weihenstephanensis or B. thuringiensis. However, MYP is preferable compared to PEMBA. The chromogenic medium has its own advantages and limitations, and some of the limitations seem to be solved by incubation at 30°C instead of the recommended 37°C.     

Numerical taxonomy ofKlebsiella pneumoniae strains isolated from clinical and nonclinical sources

A total of 180 clinical and nonclinical isolates ofKlebsiella pneumoniae, for which 99 characteristics were recorded, were subjected to numerical taxonomy analysis. Of these strains, 172 clustered into five major groups, with an overall similarity of 64%. Intragroup similarities ranged from 77 to 82%, with the subgroups corresponding to the speciesK. pneumoniae sensu stricto, K. oxytoca, andKlebsiella spp. 1, 2, and 3. Biochemical tests useful in distinguishing the species included production of indole, degradation of pectate, growth at 10°C, fecal coliform response, production of urease, fermentation of inulin andd-tartrate, utilization ofl-arginine and gentisate andm-hydroxybenzoate, and pigment formation ond-gluconate ferric citrate agar.     

DRBC agar: a new tool for <Emphasis Type="Italic">Candida dubliniensis</Emphasis> identification

Candida dubliniensis pathogenic species, which shares many phenotypic features with C. albicans, may be misidentified in the microbiology laboratory. The growth on DRBC agar at 25 °C was shown to be a new tool for differentiation between C. dubliniensis and C. albicans. All 27 isolates of C. dubliniensis showed in this medium rough colonies (peripheral hyphal fringes) and abundant chlamydospore production, while all 103 isolates of C. albicans showed smooth colonies without fringes or chlamydospores. DRBC agar allowed the differentiation of C. albicans from C. dubliniensis with 100 % sensitivity and specificity.     

In vivo assessment of the effect of an adhesive for complete dentures on colonisation of Candida species

doi: 10.1111/j.1741‐2358.2009.00345.x
In vivo assessment of the effect of an adhesive for complete dentures on colonisation of Candida species Objective: Denture adhesives have long been recognised by denture wearers as a useful adjunct to denture retention and stability. The objective of the present study was to evaluate, in vivo, the effect of a denture adhesive on oral quantities of Candida species by determination of absolute counts of colony‐forming units (CFU) per ml of saliva of individuals who use this denture adhesive for a period of 14 days. Materials and methods: Twenty‐four individuals were randomised in two equal groups of 12 (test and control), with the individuals of the test group using the adhesive for 14 days. Samples of saliva were collected from all individuals on days 0 (initial), 7 and 14. Aliquots of saliva were diluted and plated in duplicate on Sabouraud dextrose agar with chloramphenicol and incubated for 37°C for 48 h, the CFU/ml were counted in the individuals of each group and the data of each group were compared at the different time periods and analysed statistically by the non‐parametric Mann‐Whitney U‐test (α ≤ 5%). Results: There were no statistically significant differences between the test and control groups during the test periods. Conclusion: Within of the limitations of this study, the data suggested that the denture adhesive tested did not significantly alter the oral microbiota during the 14‐day trial period.     

Effectiveness of different cleaning agents on the adherence of Candida albicans to acrylic denture base resin

doi: 10.1111/j.1741‐2358.2010.00379.x
Effectiveness of different cleaning agents on the adherence of Candida albicans to acrylic denture base resin Objective:  To evaluate the ability of three alkaline peroxide‐type (Polident, Efferdent, Fittydent) and two mouth rinse cleaning agents (CloSYSII and Corsodyl) to inhibit Candida albicans on acrylic denture base resin. Background:  Appropriate routine cleaning of dentures is necessary to prevent denture stomatitis and maintenance of healthy supporting tissues. Materials and methods:  A total of 180 acrylic resin specimens (10 × 10 × 2 mm) were prepared and divided into six groups. Candida albicans was incubated on Sabouraud dextrose agar (SDA) at 37°C for 48 h. After dilution, a final yeast suspension of approximately 10 6 C. albicans per millimetre was prepared. Ten acrylic resin specimens for each group were placed in a sterile Petri dish covered with 20 ml of fungal suspension and incubated at 37°C for 90 min. Then, the specimens were immersed in 40 ml of the test solution at 37°C for 15, 30 and 60 min. Fungal cells adhering to acrylic resin surfaces were fixed in formaldehyde and counted microscopically. Results:  Mouth rinses showed the highest removal activity for all the treatment times and completely eliminated the adherence of C. albicans. Conclusions:  The use of mouth rinse may be a suitable method for cleaning dentures.