Parthenogenetic development of bovine oocytes matured in vitro for 24 hr and activated by ethanol and cycloheximide

This research was undertaken to improve development of parthenogenetic embryos following various combined treatments of ethanol and cycloheximide. In Experiment 1 in vitro matured oocytes (IVM, 24 hr) were treated with 7% ethanol for 5 min followed by incubation in 10 μg/ml cycloheximide in Medium 199 for 0 (control), 5, 10, and 20 hr. Development to 2–8 cells following culture for 3 days was similar among treated groups (32–41%; P > 0.05), which was higher than that of controls (6%; P < 0.05). Experiment 2 compared pre-ethanol exposures for 0, 1, 2.5, and 5 min, followed by 5 hr cycloheximide treatment on activation development. One- to 5-min groups resulted in 42–44% cleavage contrasted to 1–12% for controls (P < 0.05). Experiment 3 examined the effect on oocyte development of ethanol and different concentrations of cycloheximide (0, 1, 5, and 10 μg/ml). Cleavage to 2–8 cells was similar among the 5 and 10 μg/ml cycloheximide groups (36% and 42%, P > 0.05) but lower (P < 0.05) for the 1 μg/ml group (24%) and the controls (2–13%). When 5 μg/ml cycloheximide was used (Experiment 4), pre-exposure to ethanol (1, 2.5, and 5 min) resulted in more oocytes cleaved (38–41%) than in the cycloheximide alone group (0%) or the control (0%, P < 0.05). Experiment 5 tested blastocyst development of the activated oocytes with or without cytochalasin B treatment. Oocytes developed to blastocyts were 0%, 14%, 3%, and 3% (P < 0.05), respectively, for control, treatment with ethanol and cycloheximide in the presence, or absence of cytochalasin B, or electrical pulse plus cycloheximide. In conclusion, the combined ethanol and cycloheximide treatment supported high rates of parthenogenetic development using 24 hr IVM bovine oocytes. Blastocyst rate was significantly higher when cytochalasin B was added to the combined activation regimen. © 1994 Wiley-Liss, Inc.     

Drug Resistance of Staphylococci

According to an epidemiological survey of drug resistance in Staphylococcus aureus, it was found that there are three types of cross-resistance to macrolide antibiotics (EM, erythromycin: OM, oleandomycin: LM, leucomycin: SP, spiramycin), i.e., resistance to EM, OM, LM, and SP, to EM and OM, and inducible resistance to “EM, OM”. In the inducible strains of S. aureus, EM and OM are active inducers, and the optimal concentrations of the inducers are 0.1 μg/ml and 1.0 μg/ml, respectively. The induction of high resistance (800 μg/ml or more) to both EM and OM occurred within 10 min exposure to 0.1 μg/ml of EM, and the resistance of induced cells was lost after overnight growth in the absence of inducer. After 1 to 3 hr exposure to 1.0 μg/ml of OM, the inducible strains acquired high resistance (100 μg/ml or more) to EM and to a lesser extent, resistance to OM, and the acquired resistance was lost when grown in antibiotic free media. When a known concentration of EM was mixed with the induced cells or with a crude extract from induced cells which had acquired high resistance to EM and OM, the antibiotic activity of EM was still retained in the mixture, indicating that the induced mixture or the extract from the induced cells was incapable of antibiotic (EM) inactivation under the test conditions.     

Pentoxifylline Pretreatment Enhances the Oxidative Burst Activity of Human Peripheral Blood Mononuclear Cells

The effect of pentoxifylline pretreatment on the lucigenin-augmented chemiluminescence and dismutase-inhibitable superoxide production of human neutrophils and mononuclear cells (MNCs) was studied. Pentoxifylline at 20–2,000 μg/ml enhanced the lucigenin-augmented chemiluminescence (118–165% of the control, P < 0.01) of phorbol myristate acetate (PMA)-stimulated MNC. Pentoxifylline at 20–2,000 μg/ml increased the MNC superoxide production, i.e., 142–171% of the control (P < 0.05) using PMA stimulation and 145–159% of the control (P < 0.01) using opsonized zymosan stimulation. In contrast, pentoxifylline (up to 2,000 μg/ml) did not influence the lucigenin-augmented chemiluminescence and superoxide production of human neutrophils, stimulated by either PMA or opsonized zymosan. These results suggest that pentoxifylline is an immunomodulator and may have potential usefulness in the enhancement of immune defenses in compromised hosts.     

Variation in the sensitivity to metalaxyl of Pythium spp. isolated from carrot and other sources

Over a 3-yr period 261 isolates of 17 species of Pythium were tested for sensitivity to metalaxyl at concentrations of 5, 50 or 100 μ/ml. A wide range of responses was observed, from isolates where growth ceased at 5 μg/ml to those where growth at 100 μg/ml was similar to that of the untreated controls. In further tests isolates of 11 different species had ED50's < 1 μg/ml. A lower sensitivity was detected in isolates of six Pythium spp. where values in the range 1–10 μg/ml were obtained. This lower sensitivity was not related to previous known use of metalaxyl. Three isolates of Pythium dissotocum from sites where the fungicide had been used repeatedly had ED50's > 100 μg/ml and were considered resistant. The resistance was stable over a 2-yr period and isolates were cross-resistant to furalaxyl, benalaxyl, ofurace, cyprofuram and oxadixyl. Increasing concentrations of metalaxyl reduced or prevented the production of zoospores by four species of Pythium, although when zoospores were produced, this was followed by the normal processes of encystment and germination. Culturing P. dissotocum on different sub-lethal concentrations of metalaxyl for 18 wk did not induce a high level of resistance to the fungicide.     

Hyaluronidase activity of macaque sperm assessed by an in vitro cumulus penetration assay

A model system consisting of cynomolgus macaque sperm and ovulated hamster ova-cumulus complexes (OCCs) was utilized to study the role of the sperm protein PH-20 in cumulus penetration. The hyaluronidase activity of solubilized macaque sperm PH-20 was evaluated using an ELISA-like microplate assay prior to and following the addition of the hyaluronidase inhibitors heparin (0–100 μg/ml) and apigenin (250 μM), as well as the Ig fraction of a polyclonal antibody raised against purified recombinant macaque PH-20 (R10; 10–400 μg/ml). Sperm motility following exposure to enzyme inhibitors was evaluated using computer-aided sperm motility analysis. Macaque sperm were labeled with the permeant fluorescent nuclear dye, Hoechst 33342, and were coincubated with ovulated hamster OCCs for 30 min at 37°C. The addition of heparin, apigenin, or R10 antibody to solubilized sperm extracts resulted in a linear dose-dependent decrease in hyaluronidase activity (P < .01). In the heterologous cumulus penetration assay, fluorescently labeled macaque sperm that were pretreated with heparin (1–100 μg/ml), apigenin (250 μM), or R10 antibody (Ig fraction, 10–400 μg/ml) demonstrated a dose-dependent decrease in the ability to penetrate hamster OCCs (P < 0.01), in the absence of effects on sperm motility. In the homologous assay, experiments using macaque OCCs and fluorescently labeled macaque sperm confirmed that the same concentrations of heparin and R10 antibody similarly suppressed spermatozoal cumulus penetration (P < .01). These results suggest that macaque sperm PH-20-derived hyaluronidase participates in cumulus penetration in this species, and that this model system is useful for further studies into primate gamete interaction. Mol. Reprod. Dev. 46:392–400, 1997. © 1997 Wiley-Liss, Inc.     

Changes in the bovine zona pellucida induced by plasmin or embryonic plasminogen activator

Effects of bovine plasmin and plasminogen activator recovered from bovine embryo-conditioned medium (bePA) on the polypeptide profile and solubility of bovine zonae pellucidae (ZP) were evaluated. ZP were isolated from bovine ovarian oocytes and incubated at 39°C with 0, 100, or 200 μg/ml plasmin for 0, 24, or 48 hr or bePA with 0 or 100 μg/ml human plasminogen for 0 or 48 hr. ZP were evaluated either by SDS-PAGE or for changes in solubility using a zona pellucida dissolution time (ZPDT) assay. Two prominent polypeptides, molecular weight (MW) 76,000 and 65,000, and two minor polypeptides, MW 23,000 and 22,000, were resolved by SDS-PAGE. No changes occurred in the polypeptide profile for ZP incubated with 0 μg/ml plasmin for 0, 24, or 48 hr, and ZPDT did not differ (P > 0.10). Treatment with 100 or 200 μg/ml plasmin induced reductions in the MW 76,000, 23,000, and 22,000 polypeptides and the appearance of MW 45,000 and <10,000 polypeptides. ZPDT were less (P < 0.05) in 100 and 200 μg/ml compared with 0 μg/ml plasmin. Polypeptide profiles and ZPDT for ZP incubated with bePA were similar (P > 0.10) to ZP incubated with unconditioned medium. Addition of human plasminogen to ZP incubated with bePA reduced the MW 76,000, 23,000, and 22,000 polypeptides, caused the appearance of MW 45,000 and 20,000 polypeptides, and decreased ZPDT (P < 0.05). These results demonstrate that bovine plasmin is capable of proteolytically degrading the bovine ZP and that bePA can indirectly affect the ZP by converting plasminogen to plasmin. Mol. Reprod. Dev. 51:330–338, 1998. © 1998 Wiley-Liss, Inc.     

In vitro inhibitory activity of sertraline against clinical isolates of Sporothrix schenckii

BackgroundSertraline (SRT) is an antidepressant that has proven its activity in vitro against Cryptococcus, Coccidioides, Trichosporon and other fungi. Disseminated sporotrichosis, although rare, has a high mortality and its treatment is difficult and prolonged, often relying in combining two or more antifungals.AimsIn our study we evaluate the antifungal activity of SRT, alone and in combination with itraconazole (ITC), voriconazole (VRC) and amphotericin B (AMB), against 15 clinical isolates of Sporothrix schenckii.MethodsWe used the broth microdilution method as described by the CLSI to test the susceptibility to antifungals, and the checkerboard microdilution method to evaluate drug interactions.ResultsThe minimum inhibitory concentration (MIC) with SRT was in the range of 4–8 μg/ml, while for AMB, VRC and ITC were 0.5–4 μg/ml, 0.5–8 μg/ml and 0.125–2 μg/ml, respectively. In addition, SRT showed synergy with ITC in one strain, mainly additivity with VRC, and indifference with AMB in others.ConclusionsThe MIC values with SRT for the isolates studied show the potential role of this drug as an adjuvant in the treatment of sporotrichosis, especially in disseminated or complicated cases.     

Double resistance by citrus green mould Penicillium digitatum to the fungicides guazatine and benomyl

In vitro dosage response data with different isolates of Penicillium digitatum and the fungicide guazatine indicated an approximate 10-fold shift in tolerance when compared with wild type strains. ED₅₀ values for resistant strains were approximately 0.5 μg/ml compared to approximately 0.05, μg/ml for the wild type strains. Colony growth of guazatine resistant isolates on selective media containing carbendazim showed that they were also resistant to the benzimidazole group of fungicides. In vivo tests in inoculated oranges with strains previously characterised by in vitro tests confirmed resistance to guazatine and benomyl. A combined treatment of these fungicides at 400 /μ/ml and 500 μg/ml respectively, which normally gives protection against decay, also failed to provide adequate mould control. Growth and pathogenicity of the resistant strains in these tests in oranges were indistinguishable from that of wild type strains.     

New high-performance liquid chromatographic method for amphotericin B analysis using an internal standard

A simple and reproducible HPLC method for the analysis of amphotericin B (AmB) in serum, lung and liver using natamycin as the internal standard was developed. AmB and natamycin were extracted from serum, lung and liver and were separated using an isocratic elution from a C₁₈ reversed-phase column. The mobile phase consisted of acetonitrile-10 mM acetate buffer pH 4.0 (37:63, v/v). The HPLC system had two detectors in series. One was set at 303 nm and the other at 383 nm for the detection of natamycin and AmB, respectively. The retention times of AmB and natamycin were 15 and 6 min, respectively. The recovery efficiency was 96-70%. The limit of quantification was 0.1 μg/ml. The assay was reproducible, the within-day coefficient of variation (n=6) was <8% for serum, lungs and liver. The between-day variability (n=6) was <7.7% for serum, liver and lungs at 1 μg/ml or 1 μg/g tissue concentration. The assay was linear within the range 1–40 μg/ml (r²=0.999).     

High-performance liquid chromatographic assay for the novel antitumor drug,bryostatin-1, incorporating a serum extraction technique

An HPLC assay incorporating a solid-phase extraction technique has been devised for bryostatin-1. Quantitation of bryostatin was found to be linear over the concentration range 0.012–25 μg/ml (0.2–25 ng on column) and was found to have a limit of detection of 0.2 ng on column, with a correlation coefficient of 0.9999. Following extraction of bryostatin over a range of concentrations from horse serum (0.012–25 μg/ml) and human serum (0.01–0.32 μg/ml) using a 100-mg C₁₈ solid-phase extraction cartridge, extraction efficiencies consistently greater than 90% were obtained for extraction from horse serum and varied between 57 and 85% from human serum. However, on extending this work to blood samples from patients undergoing therapy with bryostatin-1, the drug was not detectable even at the maximum dose given, demonstrating the rapid loss of this agent from peripheral circulation.     

Quantification of Cyanamide Contents in Herbaceous Plants

Cyanamide (NH₂CN) is found in nature, although it has long been recognized as an industrial product. Distribution of cyanamide in the plant kingdom was investigated using a direct quantitative determination method to detect and measure cyanamide by stable isotope dilution gas chromatography–mass spectrometry (the SID–GC–MS method). The SID–GC–MS method proved to be a robust way to quantify cyanamide contents in the extracts of 101 species of herbaceous plants. The average recovery of cyanamide from all plants tested was 55.6±20.3%. Vicia villosa and V. cracca contained cyanamide at 369–498 μg/gFW and 3,460–3,579 μg/gFW respectively, while the other 99 species contained no detectable cyanamide (<1 μg/gFW). This result suggests that distribution of cyanamide in the plant kingdom is limited and uneven.     

Determination of protoberberine alkaloids in medicinal plants based on acidic potassium permanganate chemiluminescence system

A simple method was established to determine protoberberine alkaloids in Cortex Phellodendri and Rhizoma Coptidis based on an acidic potassium permanganate chemiluminescence (CL) system. The optimum conditions for the CL reaction between protoberberine alkaloids and potassium permanganate were studied in detail. Under the optimum conditions, the linear response ranges for berberine, palmatine and jatrorrhizine were 0.038–7.27, 0.031–18.1 and 0.012–3.61 μg/mL with detection limits of 0.005, 0.004 and 0.0007 μg/mL, respectively. This method was successfully applied to determine the content of protoberberine alkaloids (calculated using berberine as an index) in Cortex Phellodendri and Rhizoma Coptidis. In addition, a possible mechanism of this CL reaction was proposed on the basis of the investigation of CL, UV and fluorescent spectra of protoberberine alkaloids in acidic solution containing potassium permanganate. Copyright © 2009 John Wiley & Sons, Ltd.     

Peroxynitrite-Induced Apoptosis in Epithelial (T84) and Macrophage (RAW 264.7) Cell Lines: Effect of Legume-Derived Polyphenols (Phytolens)

Peroxynitrite (ONOO⁻) has been proposed as a mediator of gut inflammation and as an inducer of cell death by apoptosis. Phytolens (PHY), a water-soluble extract of polyphenolic antioxidants from nonsoy legumes (Biotics Research Corp, patent pending), was evaluated as a cytoprotective agent in human colonic (T84) and murine macrophage (RAW 264.7) cell lines. In the antioxidant testing, PHY showed a significant free radical scavenging ability against 1,1-diphenyl-2-picrylhydrazyl radical (DPPH) and superoxide (O₂^•) radicals with an IC₅₀of 4.44 and 5.87 μg/ml against DPPH and O₂^•, respectively. Apoptosis (DNA fragmentation) was measured by an ELISA technique. Cells were exposed to oxidative stress by treating them with peroxynitrite (100–300 μM) for 4 h in the presence and absence of PHY. Peroxynitrite elicited a dose-dependent increase in DNA fragmentation in both cell lines compared to the control group receiving decomposed ONOO⁻. PHY (10, 30, or 50 μg/ml) significantly attenuated the degree of apoptosis in T84 cells induced by ONOO⁻(P< 0.05). PHY (10–100 μg/ml) did not directly affect T84 cell viability or induce apoptosis after 4 h or overnight exposure. RAW 264.7 cells exposed to PHY alone (>30 μg/ml) for 4 h displayed decreased cell viability (P< 0.05) and increased apoptosis (P< 0.05). Phytolens may have beneficial effects on inflammation by attenuating peroxynitrite-induced apoptosis. The sparing of epithelial cells while compromising the viability of macrophages suggests that PHY may be beneficial in autoimmune disorders.     

In vitro Activity of Gatifloxacin Against 238 Strains of Anaerobic Bacteria

The activity of gatifloxacin, a new 8-methoxy-fluoroquinolone, was tested against 208 pulmonary pathogens and against an additional 30 isolates of the Bacteroides fragilis group. Pulmonary isolates were from patients with documented anaerobic pleuropulmonary infections and were obtained by appropriate sampling methods. MICs were determined using the NCCLS-approved Wadsworth brucella laked blood agar method and compared to those of clindamycin, imipenem, metronidazole and trovafloxacin. Breakpoints used to define susceptible and [resistant] categories were (in μg/ml): Clindamycin-2, imipenem-4, metronidazole 8 and trovafloxacin. No breakpoint has been defined for gatifloxacin. Gatifloxacin inhibited 99% of all anaerobes tested at 4 μg/ml and 97% of all strains at 2 μg/ml. One strain of B. fragilis was resistant to gatifloxacin at 4 μg/ml; all strains of other B. fragilis group species were susceptible. One strain of Peptostreptococcus sp. was resistant to both gatifloxacin and trovafloxacin (MIC >4 μg/ml). All other strains were susceptible to all agents at ≤μg/ml. All of the non-sporeforming Gram-positive rods were susceptible to gatifloxacin at ≤μg/ml (three strains had an MIC of 4 μg/ml). Trovafloxacin had MICs of 4 μg/ml for two strains, and an MIC of 8 μg/ml for one strain. Five percent of B. fragilis, 21% of other B. fragilis group species and 20% of Clostridium species (other than C. difficile, C. perfringens or C. ramosum) were resistant to clindamycin. No imipenem resistant isolates were found in this study. Gatifloxacin appears to have excellentin vitro activity against pulmonary isolates of anaerobes and very good activity against strains of the B. fragilis group.     

Chiral Assay of Atenolol Present in Microdialysis and Plasma Samples of Rats Using Chiral CBH as Stationary Phase

Two different enantioselective chiral chromatographic methods were developed and validated to investigate the disposition of the β₁-receptor antagonist atenolol in blood and in brain extracellular fluid of rats (tissue dialysates). System A for the plasma samples was a one-column chromatographic system with a Chiral CBH column with an aqueous buffer as mobile phase into which cellobiose was added for selective regulation of the retention of the internal standard, (S)-metoprolol. The plasma samples were analysed after a simple extraction procedure. The limit of quantitation was 0.2 μg/ml for the atenolol enantiomers. The repeatability of the medium concentration quality control plasma sample (6.0 μg rac-atenolol/ml) was 11–18% for the enantiomers. The dynamic linear range of the plasma samples was 0.5–20 μg/ml. For system B, since atenolol is an extremely hydrophilic drug, the tissue dialysate sample required a much more sensitive system as compared to the plasma samples. A coupled column system was used for peak compression of the enantiomers in the eluate after the separation on the Chiral CBH column, hence increasing the detection sensitivity. The limit of quantification was 0.045 μg/ml for the atenolol enantiomers in artificial CSF. The repeatability of the medium concentration quality control samples (0.1 and 4.0 μg rac-atenolol/ml in artificial CSF and Hepes Ringer, respectively) was 2.8–9.3% for the two enantiomers. The dynamic linear range of the brain samples was 0.05–1.0 and 0.5–20 μg/ml in artificial CSF and Hepes Ringer, respectively. Chirality 9:329–334, 1997. © 1997 Wiley-Liss, Inc.     

Clinical isolates of <Emphasis Type="Italic">Nocardia brasiliensis</Emphasis> from Japan exhibit variable susceptibility to the antibiotic imipenem

Clinical isolates of Nocardia brasiliensis from Japan were classified into two groups based on their susceptibility to the carbapenem antibiotic, imipenem (IPM). Of 33 strains tested, 10 belonged to an IPM susceptible group, with MIC of from 0.25 to 2 εg/ml and a MIC₈₀ value of 1.5 εg/ml for this antibiotic. The remaining 23 strains belonged to an IPMresistant group with MIC and MIC₈₀ values of 8–16 εg/ml and >16 εg/ml, respectively. The type strain of N. brasiliensis belonged to this resistant group. Analysis of 16S rDNA genes sequences showed that the IPM susceptible group had characteristic single nucleotide substitutions at positions 103 (T), 381 (A), and 456 (A), in contrast to the IPM resistant group. This grouping, however, was not associated with their clinical manifestation.     

Detection and molecular characterization of carbendazim-resistant Colletotrichum truncatum Isolates causing anthracnose of soybean in Thailand

A loss of fungicide efficacy, particularly for carbendazim, was noted in soybean fields in Thailand and was considered to be due to the development of Colletotrichum truncatum resistance. The carbendazim sensitivity of C. truncatum populations isolated from various soybean fields in Thailand was thus evaluated with in vitro sensitivity assays and molecular characterization of mutations in the sequences of the ß₂-tubulin (TUB2) gene that confer carbendazim resistance in the pathogen. Among 52 isolates, 46 isolates were classified as highly resistant (HR) to carbendazim (EC₅₀ > 1,000 µg/ml). All HR isolates grew on PDA amended with carbendazim at 1,000 µg/ml. Six isolates were classified as carbendazim sensitive (S) (EC₅₀ < 1 µg/ml). Mycelial growth on PDA amended with 1 µg/ml carbendazim was inhibited by over 50% compared with growth on PDA alone. When a partial TUB2 gene from the isolates was amplified and analysed using predicted amino acid sequences, an alteration from glutamic acid to alanine at codon 198 (E198A) was found in 45 HR isolates for which the EC₅₀ was higher than 2000 µg/ml. This mutation resulted from a nucleotide substitution from adenine to cytosine (GA G → GC G). The other HR isolate, CtPhS_1, with EC₅₀ of 1,127 µg/ml, had an alteration at codon 200 (F200Y) (TT C → TA C).     

In vitro antifungal activities of voriconazole and reference agents as determined by NCCLS methods: Review of the literature

Voriconazole (Vfend™) is a new triazole that currently is undergoing phase III clinical trials. This review summarizes the published data obtained by NCCLS methods on the in vitro antifungal activity of voriconazole in comparison to itraconazole, amphotericin B, fluconazole, ketoconazole and flucytosine. Voriconazole had fungistatic activity against most yeasts and yeastlike species (minimum inhibitory concentrations [MICs] <2 μg/ml) that was similar or superior to those of fluconazole, amphotericin B, and itraconazole. Against Candida glabrata and C. krusei, voriconazole MIC ranges were 0.03 to 8 and 0.01 to >4 μg/ml, respectively. For four of the six Aspergillus spp. evaluated, voriconazole MICs (< 0.03 to 2 μg/ml) were lower than amphotericin B (0.25 to 4 μg/ml) and similar to itraconazole MICs. Voriconazole fungistatic activity against Fusarium spp. has been variable. Against F. oxysporum and solani, most studies showed MICs ranging from 0.25 to 8 μg/ml. Voriconazole had excellent fungistatic activity against five of the six species of dimorphic fungi evaluated (MIC₉₀s < 1.0 μg/ml). The exception was Sporothrix schenckii (MIC₉₀s and geometric mean MICs ≥ 8 μg/ml). Only amphotericin B had good fungistatic activity against the Zygomycetes species (voriconazole MICs ranged from 2 to >32 μg/ml). Voriconazole showed excellent in vitro activity (MICs < 0.03 to 1.0 μg/ml) against most of the 50 species of dematiaceous fungi tested, but the activity of all the agents was poor against most isolates of Scedosporium prolificans and Phaeoacremonium parasiticum (Phialophora parasitica). Voriconazole had fungicidal activity against most Aspergillus spp., B. dermatitidis, and some dematiaceous fungi. In vitro/in vivo correlations should aid in the interpretation of these results. This revised version was published online in June 2006 with corrections to the Cover Date.     

Prostaglandin levels in BL6 melanoma cells cultured in vitro: the effect of vitamin E succinate supplementation

Malignant murine melanoma (BL6-F10) cells convert arachidonic acid primarily to PGD₂, PGF_2α, PGE₂, PGI₂ in descending order of magnitude. Supplementation with 1–10 μg/ml vitamin E succinate resulted in a significant (P ≤ 0.05) decrease in PGD₂ levels at vitamin concentrations of 3,5,7 and 10 μg/ml respectively, while PGF_2α levels were significantly decreased at 1,3,5 (P ≤ 0.05), 7 and 10 μg/ml (P ≤ 0.01) vitamin E succinate. BL6-F10 cells supplemented with 7 and 10 μg/ml vitamin E succinate showed a marked increase in PGE₂ levels with a significant increase occurring at 10 μg/ml (P ≤ 0.025). PGI₂ levels followed a similar trend to PGE₂ with a significant increase (P ≤ 0.05) occurring at 10 μg/ml.     

Penicillium marneffei: types and drug susceptibility

The PCR fingerprints of 30 Penicillium marneffei isolates from Chiang Rai in northern Thailand and Bangkok in central Thailand were studied through use of single-nucleotide primers (GACA)₄ and the phage M13 core sequence. Discrimination of fingerprint patterns was based on differences in the number of major bands. The P. marneffei isolates were divided into four types, i.e., A, B, C, and D. Type A was found in two isolates from Chiang Rai (6.7%). Types B and C respectively were found in two (6.7%) and one (3.3%) isolates from Bangkok. The predominate type D (83.3%) was found in isolates obtained from Chiang Rai and Bangkok. The PCR fingerprinting method was found to be useful for the epidemiological study of P. marneffei, a dimorphic opportunistic fungus and an emerging pathogen in the HIV pandemic. In vitro drug susceptibility testing by broth macrodilution to four antifungal agents against the yeast form ofP. marneffei was performed. The MIC ranges for amphotericin B, fluconazole, itraconazole, and ketoconazole were 0.125–0.5, 4.0–8.0, <0.032, and <0.125 μg/ml respectively. This revised version was published online in June 2006 with corrections to the Cover Date.