Equilibrium analysis of binding of Candida albicans to human buccal epithelial cells

The effect of growth temperature on the binding of Candida albicans to human buccal epithelial cells (BECs) was examined using an equilibrium of binding analysis. Candida albicans was cultured in M9 medium either for 12 h at 25 degrees C or for 9 h at 25 degrees C and then shifted to 37 degrees C for 3 h. The temperature shift did not result in germ tube formation; however, the adherence of C. albicans to BECs was altered. Shifting temperature increased the yeast's ability to bind to BECs. A Langmuir adsorption isotherm was used to calculate the maximum number of available binding sites (N) and the apparent association constants of binding (Ka) for all resolvable adhesin-receptor interactions. Three classes of adhesin-receptor interactions were resolved when the yeast was cultured at 25 degrees C and included a low copy number site (N = 3.0 cfu/BEC; Ka = 2.11 X 10(-6) mL/cfu), a medium copy number site (N = 23.6 cfu/BEC, Ka = 8.21 X 10(-7) mL/cfu), and a high copy number site (N = 91.7 cfu/BEC, Ka = 3.35 X 10(-8) mL/cfu). Two classes of adhesin-receptor interactions were resolved when the incubation temperature was shifted to 37 degrees C: a low copy number site (N = 4.5 cfu/BEC, Ka = 3.98 X 10(-6) mL/cfu) and a high copy number site (N = 150.5 cfu/BEC, Ka = 8.47 X 10(-8) mL/cfu). Augmented C. albicans adherence to BECs due to the elevated growth temperatures appears to result from a temperature-regulated alteration in the C. albicans adhesin that recognizes a high copy number receptor site with relatively low affinity.     

The lactoperoxidase system: the influence of iodide and the chemical and antimicrobial stability over the period of about 18 months

The lactoperoxidase (LP) system is a natural antimicrobial system, the use of which has been suggested as a preservative in foods and pharmaceuticals. The effect of adding iodide to the LP system, the chemical stability and the change in antimicrobial effectiveness during storage was studied. Addition of iodide with thiocyanate increased the fungicidal and bactericidal effect against Candida albicans, Escherichia coli and Staphylococcus aureus. Pseudomonas aeruginosa showed the same inhibition in the LP system with iodide and without iodide. Storage of the LP system in completely filled airtight containers for 18 months caused a 35% loss of the initial thiocyanate concentration. The antimicrobial activity of this LP system was strong enough to kill inocula of 106 cfu ml-1 of the four test organisms within 2 h of contact time. During storage of the air-containing LP system, the concentration of thiocyanate was reduced below detection limit within 7 d, the concentrations of hypothiocyanite and hypoiodite within 350 d. After 516 d the antimicrobial activity of air-containing LP system was strong enough to kill inocula of 106 cfu ml-1 Ps. aeruginosa within 2 h, Staph. aureus within 4 h and Candida albicans and E. coli within 1 week of contact time.     

Conductivity and pH dual detection of growth profile of healthy and stressed Listeria monocytogenes

In this study, growth of Listeria monocytogenes in a low conductivity growth medium (LCGM) was simultaneously monitored by conductivity and pH measurements. Detection times obtained from the conductivity and pH growth curves were inversely related to the initial concentration of L. monocytogenes in the medium. Linear responses were found by plotting detection times obtained from both conductivity and pH growth curves as a function of initial cell concentration in the range of 10(2) to 10(7) cfu/mL. The detection time was approximately 12 and 2 h for 10(2) and 10(7) cfu/mL of viable L. monocytogenes, respectively, using the conductivity growth curves, whereas it was approximately 1 h less using the pH growth curves. This dual detection system was used for evaluating the growth of acid-, temperature-, and salt-treated L. monocytogenes in the medium. Acid stress at pH 2 and 3 for 3 h caused approximately 12 and 4 h delay in the detection time on pH growth curves, while stress at pH 5 for 3 h did not cause a significant delay in detection time. Delay in detection times was also observed for L. monocytogenes cells exposed to 45 degrees C for more than 1 h (2 and 6 h). Exposure to 10% NaCl for 3 h did not cause visible delay in the detection time. These observations on detection times for stressed L. monocytogenes had a consistent trend with the cell number decrease determined by surface plating method.     

The use of impedance for preservative efficacy testing of pharmaceuticals and cosmetic products

Impedance was investigated for its applicability to preservative efficacy testing of pharmaceuticals and cosmetics. A good correlation between impedance detection time ( Td ) and total colony counts (colony-forming units (cfu) was obtained for untreated suspensions of Staphylococcus aureus, Candida albicans, Aspergillus niger and Pseudomonas aeruginosa in phosphate-buffered saline (PBS). A good correlation between Td and the number of cfu was also obtained for suspensions of test organisms treated for varying contact periods with selected concentrations of chlorhexidine, methyl paraben and phenoxyethanol in PBS, and methyl paraben in cetomacrogol cream, but these correlations were significantly different from those for untreated suspensions. It was found that for any given number of cfu the Td for preservative treated cells was extended. It is concluded that impedance represents a valid method for preservative efficacy testing of pharmaceuticals and cosmetics which could be used to achieve more comprehensive but economic screening of formulations against a wider range of preservative systems and concentrations than is the current approach where only a limited range of systems are tested because of the workload involved.     

Cellular elements in the resistance to candida infection in mice. I. Contribution of T lymphocytes and phagocytes at various stages of infection.

Live organisms (cfu) of Candida albicans per organ were counted 1 hr and 1 to 20 days after an intravenous inoculation into various groups of mice which had distinct levels of immunologic or non-immunologic defense mechanisms. a) The number of cfu in the liver decreased progressively in normal mice, but those in the kidney maintained a constant level during the observation period. b) The number of cfu in the liver decreased progressively also in nude mice. In their kidneys, however, cfu increased progressively at a late stage of infection. c) In lethally irradiated AKR of nude mice in which phagocyte functions were severely depressed, the number of cfu increased progressively in both liver and kidney from the initial stage of infection. d) In immunized AKR mice, growth of C. albicans was suppressed at late stages of infection. Such protective immunity could be transferred partly with immune lymphoid cells but not with hyperimmune serum in the experimental system employed. In protection against candida infection, non-immune phagocytosis and T cell-mediated immunity appear to be required at the early and late stages of infection, respectively.     

A medium for the detection of Lancefield Group D cocci in skimmed milk powder by measurement of conductivity changes

A selective medium for the detection of Lancefield Group D cocci in skimmed milk powder by conductivity measurements was developed and evaluated using the Bactometer M123 and Malthus 128H systems. This medium promoted large changes in conductance and capacitance. The calibration curve of detection times vs concentration of Lancefield Group D cocci showed a linear correlation coefficient of 0.93 and the method gave comparable results in both conductivity instruments. Naturally contaminated samples containing c. 10(3) cfu/g of Lancefield Group D cocci gave detection times within 16-18 h which was sufficiently rapid for the medium to be used for the routine screening of skimmed milk powder.     

A medium for the detection of Lancefield Group D cocci in skimmed milk powder by measurement of conductivity changes

A selective medium for the detection of Lancefield Group D cocci in skimmed milk powder by conductivity measurements was developed and evaluated using the Bactometer M123 and Malthus 128H systems. This medium promoted large changes in conductance and capacitance. The calibration curve of detection times vs concentration of Lancefield Group D cocci showed a linear correlation coefficient of 0.93 and the method gave comparable results in both conductivity instruments. Naturally contaminated samples containing c. 10³ cfu/g of Lancefield Group D cocci gave detection times within 16–18 h which was sufficiently rapid for the medium to be used for the routine screening of skimmed milk powder.     

念珠菌血症时甘露聚糖抗原及其IgG抗体检测诊断价值的临床研究

目的评价念珠菌甘露聚糖抗原（M抗原）及甘露聚糖IgG抗体（M-IgG抗体）检测诊断念珠菌血症的价值。方法收集2013年5月～2014年1月我院住院患者及健康体检人群共107例,包括念珠菌血症组（念珠菌血培养阳性患者）13例、危险因素组（临床诊断侵袭性念珠菌病或接受化疗恶性疾病、留置深静脉置管等侵袭性念珠菌病感染高危患者）63例和对照组（健康体检人群）31例。通过ELISA方法检测甘露聚糖抗原及甘露聚糖IgG抗体,比较3组人群检测阳性情况及持续时间,计算两种方法的灵敏度、特异度、阴性预测值、阳性预测值、ROC曲线下面积及Kappa值。结果白念珠菌和光滑念珠菌为念珠菌血症的主要念珠菌病原,均为5例。念珠菌血症的7／14菌株（1例患者合并2种念珠菌感染）来自重症医学科,其次为感染内科（2／14株）。除1例死亡病例外,余12例患者进行了M抗原和M—IgG抗体监测。首次M抗原检测中,4例阳性,1例可疑阳性;首次M-IgG抗体检测中,11例阳性,1例可疑阳性。经抗真菌治疗,监测14d,M-IgG抗体持续阳性时间长于M抗原。甘露聚糖抗原在诊断念珠菌血症的敏感度41．7％,特异度98．8％,阴性预测值92．4％,阳性预测值100％。甘露聚糖IgG抗体在诊断念珠菌血症的敏感度91．7％,特异度52．8％,阴性预测值100％,阳性预测值27．5％。M抗原、M抗原并M—IgG抗体作为念珠菌血症时诊断实验的ROC曲线下面积均为0．708（95％CI：0．517—0．900）,两者的Kappa值分别为0．520和0．559。结论甘露聚糖抗原在诊断念珠菌血症时的特异度较高,甘露聚糖IgG抗体在诊断念珠菌血症的敏感性较高,两者的联合检测可以适当提高检测的敏感度及特异度,有助于念珠菌血症的诊断。     

Comparative study of two culture media for the detection of phospholipase activity of Candida albicans and Cryptococcus neoformans strains

Phospholipase activity (PHA) is considered a virulence factor related to pathogenicity of Candida albicans and Cryptococcus neoformans. The aim of this work was to compare the ability of two culture media: malt egg-yolk agar (MEA) and Sabouraud-egg yolk agar (SEA), for the detection of phospholipase activity. Forty four strains of C. neoformans and 54 of C. albicans isolated from different clinical specimens of human origin were studied. The phospholipase production was determined as a ratio between the diameter of each colony and the corresponding lysis halo. The values ranged between 0 and 1, and the highest level of enzymatic activity was the nearest to 0. Enzymatic activity was observed in 34 C. neoformans strains, grown either in MEA or SEA media; 59% of enzyme producers were detected in SEA only, while five strains (15% of producers) were detected just in MEA medium. Phospholipase activity was observed in both media only in nine of 34 enzyme producer strains. Forty two out of 54 strains of C. albicans were detected as enzyme producers; 31 of them (73.8%) were detected in MEA medium only. On the other hand 10 strains (23.8% of the enzyme producers) showed phospholipase activity just in SEA medium. Detection of PHA could be done by both media in one case only. In order to evaluate the time needed to detect PHA, 41 C. albicans strains were incubated 72 h. They were read at 24 h intervals. No enzyme activity was detected at 24 h, 15 enzyme producer strains remain negative at 48 h and the halos of all strains with PHA were better distinguished after 72 h. It was possible to conclude that neither MEA nor SEA media were good enough as the unique medium to detect phospholipase activity. Nevertheless, MEA was better than SEA to detect PHA of C. albicans after 72 h incubation. The opposite situation was seen when we studied PHA in C. neoformans strains. In this case, greater sensibility was observed with SEA medium compared with MEA medium. Six days incubation, but not longer incubation times, were necessary to detect phospholipase activity in C. neoformans strains.     

Evaluation of the API ZYM system and RPMI agar plates supplemented with Tween 40 for detection of lipolytic enzymes of Candida spp

Lipolytic activity of 40 strains of Candida spp. was tested on API ZYM system and on RPMI agar plates supplemented with 1% Tween 40. Lipolytic activity was indicated by opaque zones around the inoculum cylindrical holes were punched in the medium. Clearing of the medium around the bacterial colonies indicated that an isolate produce lipase. Only 4 (21.1%) strains of C. albicans, and 3 (14.1%) strains of non-C. albicans which hydrolyzed 2-naftylomirystylan by use of the API ZYM system was observed. In contrast, 16 (78.9%) strains of C. albicans and 17 (80.7%) strains of non-C. albicans produced lipases on the agar plate using RPMI agar plates supplemented with 1.0% Tween 40. Determination oflipase activities with the API ZYM system were in no agreement with lipase tests in RPMI supplemented with Tween 40. Our study verify greater usefulness of RPMI supplemented with Tween 40 for detection of lipolytic enzymes of Candida species in comparison to the API ZYM.     

Improved detection of Listeria monocytogenes in soft mould-ripened cheese

In comparison with standard methods, enrichment in half-Fraser broth for 24 h at 30 degrees C, followed by plating out onto Listeria monocytogenes blood agar (LMBA) and PALCAM medium combined with an additional streak proved to be the most rapid and specific method for the detection of indigenous L. monocytogenes populations from soft mould-ripened cheese. This procedure, with a high sensitivity (93%) and a low detection limit (1-10 cfu 25 g-1), provided negative and presumptive positive results within 2-3 d. Differences between LMBA, PALCAM and Oxford medium turned out to be highly significant (at 99% significance level); plating on LMBA after standard enrichment protocols giving the best overall results. An improvement in detection was also obtained by modifying the confirmation procedure. A loopful of culture (an additional streak) from PALCAM or Oxford medium was streaked on non-selective medium in addition to streaking only separate colonies as specified in the standards.     

Effect of temperature on siderophore production by Candida albicans

The purpose of this study was to examine the effect of elevated temperature on growth and siderophore production by Candida albicans. The results showed that an increase in incubation temperature from 37 degrees C to 41 degrees C produced a marked decrease in both the rate and quantity of siderophore production. Elevated temperature was unable to suppress growth of C. albicans in either a control culture medium or a deferrated culture medium. A significant suppression of growth compared to the controls was observed in the deferrated media at both 37 degrees C and 41 degrees C. However with time, the growth of cells in the deferrated media showed partial recovery which was followed by an increase in siderophore production. Thus, elevation of temperature to suppress growth and siderophore production by C. albicans appears to be an ineffective host defense mechanism.     

Membrane attack complex formation on yeast as trigger of selective release of terminal complement proteins from human polymorphonuclear leukocytes

It has recently been shown that measurable amounts of complement proteins, C6 and in particular C7, are released from human polymorphonuclear leukocytes (PMNs). The aim of the present study was to investigate the impact of opsonized Candida albicans on this release. Stimulation with opsonized C. albicans led to a rapid and sustained increase of C6 and C7 in the cell culture supernatant beginning within 5 min of placing in co-culture, whereas co-culture with unopsonized C. albicans or C. albicans mock-opsonized with inactivated human serum did not affect the release. In contrast, even after stimulation employing opsonized C. albicans, no release of the complement component C8 and only trace amounts of C9 were detected. The presence of the membrane attack complex (MAC) on C. albicans after opsonization was demonstrated by indirect immunofluorescence. Opsonization of C. albicans with human serum deficient in or depleted of a terminal complement component resulted in only minor stimulation of C6 and C7 release, although C3 deposition on the surface of C. albicans was not affected as determined by direct immunofluorescence. Detailed analyses with inactivated or deficient sera showed that detection of C6 and C7 was not due to insufficient washing of the opsonized yeast prior to co-culture and suggest that only a small proportion of these proteins was derived from the membrane bound and then cleaved off MAC. Thus, these findings imply that MAC on the fungal surface may represent an additional trigger for the release of C6 and C7 from PMNs, suggesting a new role for the terminal complement complex (TCC) on target membranes as modulator of PMN functions locally at the site of inflammation.     

纳豆杆菌对白假丝酵母菌的拮抗作用

目的探讨纳豆杆菌对白假丝酵母菌的拮抗作用。方法将纳豆杆菌和白假丝酵母菌混合培养24 h后,应用沙保弱平板培养基分离白假丝酵母菌,计数菌落,计算纳豆杆菌对白假丝酵母菌的拮抗率。结果纳豆杆菌对白假丝酵母菌的拮抗作用明显,拮抗率高达91.91%;纳豆杆菌肉汤培养物的除菌滤液对白假丝酵母菌也有明显的拮抗作用,拮抗率为79.05%。结论纳豆杆菌对白假丝酵母菌具有明显拮抗作用,是白假丝酵母菌的理想拮抗菌株。     

Antimicrobial activity of clove oil dispersed in a concentrated sugar solution

Essential oil of clove, dispersed (0.4% v/v) in a concentrated sugar solution, had a marked germicidal effect against various bacteria and Candida albicans. Staphylococcus aureus (five strains), Klebsiella pneumoniae, Pseudomonas aeruginosa, Clostridium perfringens, and Escherichia coli inoculated at a level of 10(7) cfu/ml, and C. albicans (inoculum 4.0 x 10(5) cfu/ml) were killed (greater than 99.999%) after 2-7 min in a laboratory broth supplemented with 63% (v/w) of sugar, and containing 0.4% (v/w) of essential oil of clove. Added organic matter (i.e. human or bovine serum) did not impair its antimicrobial activity. Sugar was not necessary for the antimicrobial activity of clove oil, but the concentrated sugar solution provided a good vehicle for obtaining an oil dispersion that is relatively stable for certain practical applications.     

Comparison of methods for the recovery and detection of low levels of injured Salmonella in ice cream and milk powder

This study compared the ability of four rapid methods and a standard cultural method to detect low levels of heat-injured cells of Salmonella typhimurium in ice cream and skimmed milk powder. The detection of Salmonella in samples contaminated with low levels (< 10 cfu 25 g-1) was significantly greater with the novel broth method than with the other methods (P 10 cfu 25 g-1, there was no significant difference between the methods except for the novel broth method and a dipstick-based immunoassay (P 相似文献   

Detection of low numbers of pathogenic Yersinia enterocolitica in environmental water and sewage samples by nested polymerase chain reaction

Isolation of pathogenic Yersinia enterocolitica from water and sewage by traditional culture techniques is time-consuming and subsequent differentiation between pathogenic and non-pathogenic strains can be difficult and unreliable. A nested polymerase chain reaction (PCR) procedure was used for the detection of low numbers of Y. enterocolitica in spiked samples from natural surface sources with variable background flora ranging from oligotrophic water to sewage. Water and sewage samples were filtered and filters enriched overnight in a non-selective medium. Nested PCR conducted on enriched broth, prepared by use of a rapid and simple preparation step consisting of centrifugation, proteinase K treatment and boiling, enabled the detection of 8-17 cfu 100 ml-1 water with background levels of up to 8700 heterotrophic organisms ml-1 and 10,000 cfu coliform organisms 100 ml-1 water. The analysis can be completed within 2-3 d and should be a significant tool in monitoring environmental waters and drinking water sources for the presence of pathogenic Y. enterocolitica.     

Detection of low numbers of Salmonella in environmental water, sewage and food samples by a nested polymerase chain reaction assay

A polymerase chain reaction (PCR) assay with two nested pairs of primers selected from conserved sequences within a 2.3 kb randomly cloned DNA fragment from the Salmonella typhimurium chromosome was developed. The nested PCR assay correctly identified 128 of a total of 129 Salmonella strains belonging to subspecies I, II, IIIb and IV. One strain of Salm. arizona (ssp. IIIa) tested negative. No PCR products were obtained from any of the 31 non-Salmonella strains examined. The sensitivity of the assay was 2 cfu, as determined by analysis of proteinase K-treated boiled lysates of Salm. typhimurium. The performance of the assay was evaluated for environmental water, sewage and food samples spiked with Salm. typhimurium. Water and sewage samples were filtered and filters were enriched overnight in a non-selective medium. Prior to PCR, the broth cultures were subjected to a rapid and simple preparation procedure consisting of centrifugation, proteinase K treatment and boiling. This assay enabled detection of 10 cfu 100 ml(-1) water with background levels of up to 8700 heterotrophic organisms ml(-1) and 10000 cfu of coliform organisms 100 ml(-1) water. Spiked food samples were analysed with and without overnight enrichment in a non-selective medium using the same assay as above. Nested PCR performed on enriched broths enabled detection of <10 cfu g(-1) food. Variable results were obtained for food samples examined without prior enrichment and most results were negative. This rapid and simple assay provides a sensitive and specific means of screening drinking water or environmental water samples, as well as food samples, for the presence of Salmonella spp.     

Probeliatrade mark PCR system for rapid detection of Salmonella in milk powder and ricotta cheese

The Probeliatrade mark Salmonella sp. PCR amplification and detection kits (Sanofi Diagnostics Pasteur, Marnes La Coquette, France) were evaluated for the rapid and specific detection of Salmonella agona artificially inoculated into skim milk powder and ricotta cheese. The Probeliatrade mark results were compared with those obtained using the Australian Standard Method. Using a pure culture of Salm. agona, the detection limit of Probeliatrade mark was between 8 and 79 cfu ml-1, equivalent to 0.2-2 cfu per PCR reaction. Detection of Salm. agona inoculated in skim milk powder (at 5-10 cfu g-1, stored at 5, 15 or 25 degrees C) and ricotta cheese (at 1-2, 10-20 and 100-200 cfu per 25 g) was effected by using non-selective enrichment prior to the PCR determinations. For all of the 40 milk powder samples and 12 ricotta cheese samples, the Probeliatrade mark results were consistent with those using the Australian Standard Method. Using Probeliatrade mark, Salmonella was detected to genus level in the dairy products within 24-28 h, whereas the cultural technique required 3-4 d for presumptive positive isolates and further time for confirmation.     

阴道白色念珠菌致病株与携带株两相性与毒力关系研究

目的探讨阴道白色念珠菌致病株和携带株菌丝相和酵母相分泌型酸性蛋白酶和细胞外磷脂酶活性以及与其毒力的关系。方法分别采用牛奶平板和卵黄培养基法检测白色念珠菌致病株和携带株200株分泌型酸性蛋白酶和细胞外磷脂酶的活力,分别将致病产酶株的菌丝相和孢子相菌悬液（5×10^6CFU／ral）注射小鼠尾静脉,1个月内观察小鼠死亡率及平均存活时间,以平均存活时间评价菌株毒力。结果白色念珠菌致病株和携带株分泌型酸性蛋白酶检出率分别为83．3％和35．7％（P〈0．01）;细胞外磷脂酶阳性率分别为87．5％和39．3％（P〈0．01）。动物实验结果表明,白色念珠菌致病株菌丝相分泌型酸性蛋白酶和细胞外磷脂酶的活力均显著高于孢子相（P〈0．01,P〈0．005）;注射菌丝相白色念珠菌的小鼠死亡率高于注射孢子相的小鼠（P〈0．01）,且平均存活期短于注射孢子相的小鼠（P〈0．01）。结论分泌型酸性蛋白酶和细胞外磷脂酶是白色念珠菌重要毒力因子,致病株毒力高于携带株,菌丝相毒力高于酵母相。