Comparative evaluation of four commercial tests for presumptive identification of Candida albicans

Four commercially available tests (Albicans ID2, Chromalbicans Agar, CHROMagar Candida, and BactiCard Candida) and the germ tube (GT) test for presumptive identification of Candida albicans were evaluated using clinical isolates of C. albicans (n=89) and of non-albicans yeasts (n=107). Sensitivities and specificities of all tests regarding the identification of C. albicans were greater than 92%, except for Chromalbicans Agar plates (88.7% after 48 h) and their specificity was 86%. Overall, the four commercial systems were easy to use and are good systems for the routine identification of C. albicans.     

Observations on the use of a medium detecting β-glucuronidase activity and lactose fermentation for the simultaneous detection of Escherichia coli and coliforms

Three hundred and forty clinical isolates of Candida species and Saccharomyces cerevisiae were tested in order to evaluate different methods for identification of Candida albicans using fluorogenic or chromogenic substrates. Detection of N -acetyl-β-D-galactosaminidase (NAGase) was performed with ready-to-use agars such as Fluoroplate Candida Agar (Merck, Germany), MUAG Candida Agar and MUAG Sabouraud Agar (Biolife, Italy) which contained 4-methylumbelliferyl- N -acetyl-β-D-galactosaminide (4-MUAG). MUAG Candi Kit and RAP-ID-ALBICANS (Biolife, Italy) and Albicans ID Agar (bioMérieux, France) were also used. The Vitek AMS System was used as a reference identification method for all isolates. NAGase activity could be detected for C albicans with Fluoroplate Candida Agar (98.8%), MUAG Sabouraud Agar (98.4%), Albicans ID (99.6%). MUAG Candi Kit (97.5%) and RAP-ID-ALBICANS (96.2%). Proline aminopeptidase examined with RAP-ID-ALBICANS was present in 98.7% of C. albicans. There was one false-positive result for C. tropicalis (9.1%) on Fluoroplate Candida Agar, one false-positive result for C. glabrata (2.2%) on Albicans ID Agar: five false-negative results for C. albicans (3.1%), but no false-positive results for the other tested species were observed with RAP-ID-ALBICANS.     

Evaluation of a new chromogenic medium (Candida ID) for the isolation and presumptive identification of Candida albicans and other medically important yeasts

Candidiasis is a frequent human infection caused mainly by Candida albicans. However, other species are emerging as important pathogens, as Candida glabrata, Candida parapsilosis, Candida tropicalis, Candida krusei or Candida guilliermondii. Rapid identification of clinical isolates could facilitate diagnosis and treatment. Candida ID (bioMerieux, Spain) is a new medium for the isolation and presumptive identification of yeasts: C. albicans grows as blue colonies, and C. tropicalis, C. guilliermondii, Candida kefyr and Candida lusitaniae as pink ones. The utility of Candida ID was evaluated with more than 700 clinical isolates and type culture collection strains from different genera including Candida, Cryptococcus, Saccharomyces, and Rhodotorula. Presumptive identification was confirmed by germ tube test, microscopic morphology and chlamydoconidia production on corn meal agar and carbohydrate assimilation on API-ATB ID 32C or Vitek (bioMerieux). Growth on Candida ID was rapid (18-24 h) for most of the yeast strains tested. Sensitivity and specificity of identification of C. albicans was significantly high (>98%), since a very low number of isolates were found to be false negative or false positive. A better result was obtained for species growing as pink colonies (>99.5%). Detection of different species of medical important yeasts was easy with Candida ID, as perfectly distinct colors and textures of colonies were observed on this medium. Candida ID allowed the discrimination between C. glabrata (creamy and smooth) and C. krusei (rough and white) colonies. Other species showed different colony textures and colours, white being the predominant colour. Candida ID was very useful for the presumptive identification C. albicans isolates.     

Cultural,morphological and bio-chemical variability of different isolates of Colletotrichum truncatum causing anthracnose of greengram

A laboratory experiment was conducted to study the variability among the eight isolates of Colletotrichum truncatum of greengram collected from different locations on the basis of cultural, morphological and biochemical characteristics by using nine different culture media viz., Potato Dextrose Agar (PDA), Potato Carrot Agar (PCA), Oat Meal Agar (OMA), Corn Meal Agar (CMA), Carrot Agar (CA), Sabouraud’s Agar (SA), Czapek’s Dox Agar (CDA), Richard’s Agar (RA) and V₈ Juice Agar. Colony colour varied in different media from white or white with light brown centres which later changed to black or dark to light brown with increase in the age of the fungal cultures. Mostly, the colonies had fluffy or cottony mycelial growth with slight variations and regular to irregular white margin. PDA, PCA, CA and RA produced maximum mycelial growth (90?mm) at 10 DAI (days after inoculation). Minimum growth was observed on SA (69.56?mm) and V₈ juice agar media (55.42?mm) and their difference was statistically significant. Morphological variability among the isolates was studied by comparing their conidial length, breadth and length and breadth of setae and their differences were statistically significant. Biochemical variability among the isolates was based on α- and β-esterase and peroxidise profiling. Positive activity was observed for both α- and β-esterase. α-Esterase enzyme showed the highest enzyme activity in terms of maximum numbers of banding loci among the three isozymes tested. The findings of the present study clearly revealed that cultural, morphological and biochemical variability did exist among the different isolates of C. truncatum.     

Application of Replica Plating and Computer Analysis for Rapid Identification of Bacteria in Some Foods: I. Identification Scheme

A method was devised and tested for a quantitative identification of microbial flora in foods. The colonies developing on the initial isolation plates were picked with sterile toothpicks and inoculated on a master plate in prearranged spacing and order. The growth on the master plates was then replicated on a series of solid-agar plates containing differential or selective agents. The characteristic growth and physiological responses of microbial isolates to penicillin, tylosin, vancomycin, streptomycin, chloramphenicol, neomycin, colistin, and to S S Agar, Staphylococcus Medium No. 110, and Potato Dextrose Agar were recorded, together with Gram reaction and cell morphology. This information was then fed into an IBM 1410 digital computer which grouped and analyzed each isolate into 10 microbial genera, or groups, according to the identification key. The identification scheme was established by use of reference culture studies and from the literature. This system was used to analyze the microbial flora in dover sole (Microstomus pacificus) and ground beef. The method described in this article enables one to examine large numbers of microbial isolates with simplicity.     

Phenotypic evaluation to differentiate Candida albicans from Candida dubliniensis

This study evaluated the phenotypic tests used to differentiate Candida albicans from Candida dubliniensis. A total of 55 isolates from vaginal secretions, oral cavity and hemoculture were studied. They were originally identified as C. albicans, based on their morphological and physiological characteristics. These isolates were tested for colony color development on CHROMagar Candida medium, growth at 45 degrees C on Sabouraud Dextrose agar, lipolytic activity on Tween 80 Agar medium and colony morphology and chlamydoconidia formation on Staib agar medium. Of the 55 isolates studied, seven yielded one or more phenotypic characteristics suggestive of Candida dubliniensis. These isolates were tested by PCR with specific primers for Candida dubliniensis and API ID 32. The seven isolates were confirmed as Candida albicans. All of these finding indicate that DNA based tests should be used for definitive identification of Candida dubliniensis.     

An improved medium for the detection of Aspergillus flavus and A. parasiticus

An effective selective medium for the enumeration of Aspergillus flavus and Aspergillus parasiticus has been developed by modification of Bothast and Fennell's Aspergillus Differential Medium. Results can be obtained with the new medium, Aspergillus flavus and parasiticus Agar (AFPA), after 42 h incubation at 30 degrees C. The medium is thus suitable for use in quality control as a guide to the presence of A. flavus and, potentially, of aflatoxins. AFPA has been extensively tested on peanuts and soils. Results were reproducible and comparable with those on standard fungal enumeration media incubated for much longer periods. A very low percentage of false positive or negatives was found.     

An improved medium for the detection of Aspergillus flavus and A. parasiticus

An effective selective medium for the enumeration of Aspergillus flavus and Aspergillus parasiticus has been developed by modification of Bothast and Fennell's Aspergillus Differential Medium. Results can be obtained with the new medium, Aspergillus flavus and parasiticus Agar (AFPA), after 42 h incubation at 30°C. The medium is thus suitable for use in quality control as a guide to the presence of A. flavus and, potentially, of aflatoxins. AFPA has been extensively tested on peanuts and soils. Results were reproducible and comparable with those on standard fungal enumeration media incubated for much longer periods. A very low percentage of false positives or negatives was found.     

A screen for Clostridium difficile in the vagina: an out-patient study using and comparing selective media

Cycloserine-Cefoxitin-Fructose Agar (CCFA) gives good presumptive identification of Clostridium difficile after 1- or 2-day incubation whereas Reinforced Clostridial Medium (RCM)/p-cresol is not very selective for the organism from the vagina. The identification of 91.5% of the isolates from an initial screen subjected to biochemically based tests was achieved. Conventional screening of vaginal swabs failed to confirm any significant occurrence of Cl. difficile in the vagina of pregnant or non-pregnant women. The incorporation of an enrichment stage in the isolation procedure, however, did reveal a significant presence of the organism in the vagina of both pregnant and non-pregnant women.     

临床分离161株念珠菌菌种鉴定及氟康唑药敏试验分析

目的调查临床分离的念珠菌种类及其对氟康唑的敏感性。方法采用科玛嘉念珠菌显色培养基和YBC平板对山东大学齐鲁医院细菌室分离到的161株念珠菌进行鉴定,并对分离出的白念珠菌分别采用NCCLS推荐的微量稀释法和ROSCO药敏纸片法对氟康唑进行药物敏感性试验。结果在161株念珠菌中白念珠菌为69．57％,热带念珠菌为19．88％,近平滑念珠菌为4．97％,克柔念珠菌为2．48％,光滑念珠菌为1．86％,其他念珠菌为1．24％。两种方法药敏结果显示：112株白念珠菌对氟康唑的敏感率分别为96．43％和97．32％,仅有2株耐药。结论白念珠菌仍然是我院分离率最高的念珠菌,其次是热带念珠菌和近平滑念珠菌。白念珠菌对氟康唑仍敏感,耐药菌株极少。     

A Modification of Brilliant Green Agar for Improved Isolation of Salmonella

Five organisms commonly found to be capable of growth on commercial Brilliant Green Agar (BGA) after enrichment in Muller-Kauffman Tetrathionate broth (MKT) were tested for sensitivity to 18 antimicrobial agents. The sensitivities of two Salmonella serotypes to these agents were also tested. A combination of sulphacetamide (at 1.0 mg/ml) and mandelic acid (at 0.25 mg/ml) incorporated into BGA was found to give maximum recovery of salmonellas from MKT broth enrichment whilst giving maximum suppression of contaminating organisms. More importantly, this Antibiotic-enriched Brilliant Green Agar (ABG) gave a lower incidence of false positive results when compared with commercial BGA. Increasing the incubation temperature from 35 to 43°C was found to accentuate the selectivity of ABG without inhibiting the growth of salmonellas. A total of 31 Salmonella serotypes were tested for their ability to grow on ABG at 43°C; all produced typical colonies.     

Use of immunoperoxidase for the rapid identification of human myxoviruses and paramyxoviruses in tissue culture

The immunoperoxidase method, using commercially available antisera, was compared with standard virological methods for the identification and typing of 77 isolates of human myxoviruses and paramyxoviruses. Results of typing using neutralization tests and the immunoperoxidase technique were identical for 76 of the 77 isolates. With the immunoperoxidase method there was one false negative reaction, but no false positive reactions. Cross-reactivity between influenza A (soluble) and A₂/HK antisera with influenza A isolates was noted, but did not interfere with the interpretation of results. It is concluded that the immunoperoxidase method is ideally suited for the rapid identification and typing of common human respiratory viruses on a routine basis. It offers a number of decided advantages over both immunofluorescence and standard virological methods.     

A medium detecting β-glucuronidase for the simultaneous membrane filtration enumeration of Escherichia coli and coliforms from drinking water

A medium for the single membrane enumeration of Escherichia coli and coliforms from potable water was developed by the modification of the standard UK membrane filtration medium. The medium, membrane-Lactose Glucuronide Agar (m-LGA), employs a chromogenic substrate for the detection of β-glucuronidase activity and sodium pyruvate to enhance recovery of chlorine-stressed coliforms. Escherichia coli identification was significantly improved on m-LGA with 98.6% of presumptive isolates confirming. Recovery of coliforms from drinking water samples was also significantly improved.     

Antibiotic Response and Plasmid Profile of Bacteria Isolated from a Landfill

Bacteria isolates belonging to the genera Bacillus, Corynebacterium, Aeromonas, and Enterobacter were isolated from a municipal waste landfill in Durham, NC. Bacterial counts obtained with three general purpose media were log₁₀ colony-forming units (cfu)/g of 9.30, 9.26, and 9.20 respectively for Plate Count Agar, Brain Heart Infusion Agar, and Nutrient Agar. Coliform count from MacConkey agar was log₁₀ 7.28/g sample. Isolates were generally sensitive to tetracycline and chloramphenicol (13 of 14 isolates) and generally resistant to ampicillin (9 of 9), erythromycin (10 of 14), streptomycin (8 of 14), with 3 of 14 isolates having multiple resistance to the last three antibiotics. A dose-independent growth response to ampicillin was observed for two isolates. The detection of a 22,000-bp plasmid in one but not in the second ampicillin-resistant isolate suggests more than one mechanism of antibiotic resistance. Received: 23 March 1999 / Accepted: 6 July 1999     

Serological identification of group D streptococci using commercial antisera

Three commercial group D streptococcal antisera were tested for the serological identification of 100 group D enterococci; 20 Streptococcus bovis; 5 isolated from each of the following streptococcal groups: A, B, C and G; and 3 isolates from serological group F. Antisera from Difco Laboratories, BBL, and Wellcome Reagents Limited were used in the classic capillary tube precipitin test on extracts prepared using the Rantz and Randall procedure. No false positive precipitin reactions were observed. Of the enterococcal isolates, all 100 reacted with the Wellcome, 99 reacted with the BBL, and 96 reacted with the Difco group D antisera. However, of the 20 S. bovis isolates, only 2 reacted with the BBL, and 1 reacted with both the Difco and the Wellcome antisera. Each antiserum was then used to prepare staphylococcal coagglutination (CoA) reagents and each isolate was subsequently tested. A simple extraction procedure was performed by suspending colonies of an isolate in a loopful of salin on a microscope slide and gently heating the slide directly in the flame of a Bunsen burner. All 100 enterococci and all 20 S. bovis gave positive results with the BBL and the Wellcome CoA reagents. Using the Difco reagent, 2 S. bovis isolates failed to produce postitive results. No false positive results were observed with the non-Group D isolates. Our results indicate that the CoA technique using commercial group D antisera may provide faster and sometimes more sensitive serological identification than the classic capillary tube precipitin test.     

Comparison of disk diffusion test and Etest for voriconazole and fluconazole susceptibility testing

The distribution for voriconazole and fluconazole susceptibility was determined by Etest and disk diffusion test in 143 clinical isolates. The majority of the strains of Aspergillus spp., Candida krusei, C. inconspicua, C norvegensis and Saccharomyces cerevisiae displayed resistance or decreased susceptibility to fluconazole in contrast to voriconazole. The absolute categorical agreement for voriconazole and fluconazole susceptibility results by the disk method and Etest was 90.5 and 74.8 % respectively. The error rate bounding analysis showed only 0.7 % of false susceptible results ( very major error) with voriconazole, but 2.8 % with fluconazole. Fluconazole can be used as a surrogate factor to predict voriconazole susceptibility but with lower reliability for susceptible-dose dependent and resistance category, especially in Candida glabrata isolates. The results of the disk method were not substantially influenced by the composition of media (Mueller-Hinton agar vs antimycotic Sensitivity Test agar), even if with the latter the results had fewer tendencies to produce false susceptibility of C.glabrata isolates to both of the triazole drugs. Disk test as well as Etest were shown to represent suitable methods for routine evaluation of susceptibility of clinical isolates of pathogenic fungi, including aspergilli, to fluconazole and voriconazole.     

金沙江干热河谷地区芒果畸形病的病原菌

从金沙江干热河谷地区采集芒果畸形病组织,运用柯赫氏法则证实分离物MG6为该病的致病菌。菌株MG6在马铃薯蔗糖琼脂培养基（PSA）上菌丝白色,无色素产生,米饭培养基浅粉红色;在康乃馨叶片培养基（CLA）上以单瓶梗或复瓶梗假头状产孢,不产生链状孢子;小型分生孢子卵形或长椭圆形,具0－1个分隔,3.1－10.2×1.5－2.2μm;大型分生孢子呈镰刀形,通常3个分隔,18－38×1.8－2.4μm。EF-1α测序结果在Fusarium数据库中进行同源性分析显示,菌株MG6与F. mangiferae的同源性最高,达99.68%。综合培养性状、形态学特征和EF-1α序列分析,将菌株MG6鉴定为Fusarium mangiferae。     

A comparison of different culture media for the membrane filter quantification of Aeromonas in water

A comparative assessment of culture media for the membrane filter enumeration of Aeromonas spp. in water was performed, testing the effects of different incubation conditions (aerobic and anaerobic), temperatures (30 and 37 degrees C) and times (24 and 48 h). Different water samples seeded with test suspensions of Aeromonas spp., fecal material or raw sewage were examined. Results indicate clearly that plates should be incubated aerobically at 30 degrees C for 24 h. If the bacterial contamination is likely to be low, the use of most sensitive culture media, such as SAA, mA, ADA or PADE Agar, is recommended. By contrast, samples with an expected high level of background microbial flora should be analysed through more selective media, such as MIX Agar. However, the low selectivity of all media tested and the high likelihood of false negatives based upon the macroscopic examination of colonies means that further research directed to the development of more efficient media is needed.     

A rapid urease test for presumptive identification of Cryptococcus neoformans

A rapid method to evidence urease activity is described. Urea hydrolysis and consequent production ammonia are detected by a chemical reaction producing a blue phenol compound (indophenol blue). Three hundred and three yeast were tested. Out of 107 urease-positive organisms detected by Christensen's Urea Agar Test (CUAT) 102 were positive by our method. No false negatives were observed by this method when testing 87 Cryptococcus strains. Ths practical screening test for presumptive identification of Cryptococcus neoformans is simple, unaffected by pH changes and requires 15 minutes to be performed.     

Enumeration of byssochlamys and other heat-resistant molds

Methods for the detection of low numbers of heat-resistant molds on fruits were studied by using cultures of Byssochlamys and a number of unidentified mold isolates. Ascospore dormancy had a marked effect on viable recoveries, and the medium in which ascospores were heated influenced activation rates. Best results were obtained when fruit homogenates were heated for 60 min at 70 C in Concord grape juice, followed by culturing on acidified Potato Dextrose Agar.