Rapid and extensive vacuolation of the budding yeastsSaccharomyces cerevisiae andCandida albicans by amphotericin B

The effect of amphotericin B (AMPH) on vacuolation in the budding yeastsSaccharomyces cerevisiae andCandida albicans was studied. The minimum inhibitory concentration of AMPH for growth ofS. cerevisiae andC. albicans was 1 µg/ml. In untreated control cultures, mature cells had large central vacuoles in the exponential phase, which hampered the detection of vacuolation effect. Small buds in untreated exponential phase cells, however, only rarely showed vacuoles under the light microscope. Treatment with 0.2 µg/ml of AMPH for 20–30 min induced extensive vacuolation not only in mothers but also buds ofS. cerevisiae. Extensive vacuolation lasted 4 h or more, and growth rate of the cells was much reduced for 8 h or more. Vacuolation itself was not fatal: on removal of the drug most cells gradually recovered from vacuolation and eventually multiplied. A similar effect of AMPH was also observed inC. albicans but at a higher concentration (0.5 µg/ml).     

Effects of organic compounds on the degradation ofp-nitrophenol in lake and industrial wastewater by inoculated bacteria

Many microorganisms fail to degrade pollutants when introduced in different natural environments. This is a problem in selecting inocula for bioremediation of polluted sites. Thus, a study was conducted to determine the success of four inoculants to degradep-nitrophenol (PNP) in lake and industrial wastewater and the effects of organic compounds on the degradation of high and low concentrations of PNP in these environments.Corynebacterium strain Z4 when inoculated into the lake and wastewater samples containing 20 µg/ml of PNP degraded 90% of PNP in one day. Addition of 100 µg/ml of glucose as a second substrate did not enhance the degradation of PNP and the bacterium utilized the two substrates simultaneously. Glucose used at the same concentration (100 µg/ml), inhibited degradation of 20 µg of PNP in wastewater byPseudomonas strain MS. However, glucose increased the extent of degradation of PNP byPseudomonas strain GR. Phenol also enhanced the degradation of PNP in wastewater byPseudomonas strain GR, but had no effect on the degradation of PNP byCorynebacterium strain Z4.Addition of 100 µg/ml of glucose as a second substrate into the lake water samples containing low concentration of PNP (26 ng/ml) enhanced the degradation of PNP and the growth ofCorynebacterium strain Z4. In the presence of glucose, it grew from 2×10⁴ to 4×10⁴ cells/ml in 3 days and degraded 70% of PNP as compared to samples without glucose in which the bacterium declined in cell number from 2×10⁴ to 8×10³ cells/ml and degraded only 30% PNP. The results suggest that in inoculation to enhance biodegradation, depending on the inoculant, second organic substrate many play an important role in controlling the rate and extent of biodegradation of organic compounds.Abbreviations PNP p-nitrophenol     

Synergy of fluconazole with macrophages for antifungal activity againstCandida albicans

The possible synergy between macrophages and fluconazole for antifungal activity against different isolates ofC. albicans was studied. The susceptibility ofC. albicans isolates to fluconazole (FCZ), when incubated in RPMI-1640 with 10% fetal bovine serum (FBS) and 10% fresh mouse serum (test medium, TM) was determined by using a quantative culture methodology. Multiplication of isolate Sh27 was strongly inhibited by FCZ, even at 1.0 µg/ml. However, FCZ even at 100 µg/ml was not fungicidal. Resident murine peritoneal macrophages (MP) incubated for 48 h in RPMI-1640+10% FBS (tissue culture medium, TCM), then challenged with Sh27 in TM for 24 h, were fungistatic (20±9%,n=4). Cultured macrophages synergized with FCZ (10 µg/ml) for fungicidal activity when co-cultured with Sh27 in TM for 24 h (46±8%) and for 48 h (74±5%),n=3. Macrophages and FCZ (10 µg/ml) could not synergize for significant killing of a less FCZ-sensitiveC. albicans isolate 94-164. Multiplication of a FCZ-resistant isolate (94–20) was not inhibited by FCZ at 10 µg/ml TM; however, macrophages and FCZ (10 µg/ml) could synergize for fungistatic (64%), but not fungicidal, activity.     

Influence of the mycotoxins aflatoxin B1, rubratoxin B,patulin and diacetoxyscirpenol on the fermentation activity of baker's yeast

The fermentation activity of baker's yeast (measured by the amount of produced CO₂) is inhibited by 100µg/ml and 10µg/ml aflatoxin B₁, and by 100µg/ml and 10µg/ml diacetoxyscirpenol. Lower concentrations of these mycotoxins as well as of rubratoxin B enhance the fermentation. Only 0.001µg/ml aflatoxin B₁, 0.00001µg/ml diacetoxyscirpenol and 0.01µg/ml rubratoxin B are without effect or slightly inhibitory. Patulin in all concentrations tested does not influence the CO₂ production significantly. Cytochemical studies show that the enzyme alcohol dehydrogenase is inhibited by 100µg/ml and enhanced by 1µg/ml and 0.1µg/ml aflatoxin B₁. It is suggested that the influence of at least aflatoxin B₁ on the fermentation activity of the yeast cells is due to an interaction with alcohol dehydrogenase. It is possible that the activity of other enzymes of yeast is also influenced by mycotoxins.     

Efficacy of 6-amino-2-n-pentylthiobenzothiazole onTrichophyton in vitro and in vivo

In vitro and in vivo antifungal activities of synthetically parepared 6-animo-2-n-pentylthiobenzothiazole (APB) againstTrichophyton strains were studied. APB inhibited the growth of 3Trichophyton strains at 65 µg/ml. 2-Mercaptobenzothiazole was not effective at 125 µg/ml and ketoconazole inhibited the growth at 20–30 µg/ml. Treatment of experimental dermatophytosis in guinea pigs using 2.5% APB cream was studied in comparison to Canesten cream (1% clotrimazole). Dermatophytosis was considerably reduced after both APB and Canesten therapies.     

Effect of some antibiotics on Sclerotium cepivorum Berk., the cause of white rot of onion

The effect of some antibiotics onSclerotium cepivorum, the cause of white rot of onion was studied in agar culture and soil. The growth ofS. cepivorum was inhibited in Czapek Dox yeast agar containing 50µg of gliotoxin, viridin, actidione and 100µg of patulin per ml of the medium. Lower concentrations of the antibiotics retarded the growth of the fungus. In soil, patulin had no effect in the control ofS. cepivorum infection of onion seedlings. Concentration of actidione of 5µg/g of soil completely controlled white rot infection but severely stunted the growth of onion seedlings; 40µg/g of actidione killed the seedlings. Despite the importance of actidione as a fungistatic agent its use on onion is limited by its phytotoxicity.     

Production,partial purification and bioassay of toxic metabolites of three plant pathogenic species ofColletotrichum in Nigeria

Colletotrichum lindemuthianum, C. truncatum andC. graminicola, produced metabolites in culture which induced necrotic lesions on susceptible hosts. The highest production was obtained from Richard's medium under shake incubation. The toxic metabolites of these pathogens fluoresce at 254 nm and 366 nm under ultraviolet light. The crude metabolites of theseColletotrichum species inhibited seed germination at the concentration of 100 µg/ml while the potency of the metabolites decreases with increase in dilution to 0.1 µg/ml. Similarly, the metabolites also inhibited growth of seedlings of hosts to the pathogen at 100 µg/ml and the potency again reduced with increasing dilution to 0.10 µg/ml.     

The phytotoxicity of selected mycotoxins on mature,germinatingZea mays embryos

Mature maize (Zea mays) embryos were exposed to 5, 10 and 25 µg ml^–1 of deoxynivalenol (DON), zearalenone (ZEA), ochratoxin A (OA) and a mixture of zearalenone and deoxynivalenol (ZEA/DON) for 9 days. DON and the ZEA/DON combination were consistently more inhibitory of the measured parameters than either ZEA or OA. Based on the predicted additive values, it would appear that, in combination, ZEA and DON act synergistically to inhibit root and shoot growth. For ZEA alone, a concentration of 5 µg ml^–1 ZEA was generally inhibitory of root and shoot elongation and fresh mass accumulation, while at 10 and 25 µg ml^–1, this toxin had a stimulatory effect on these parameters. For OA, the measured effects on root and shoot growth at 5 and 25 µg ml^–1 were stimulatory, while at 10 µg ml^–1 OA, an inhibitory effect was observed. For all toxins, inhibitory/stimulatory effects were generally more marked for root parameters than for shoot elongation or mass.Abbreviations ADON acetyldeoxynivalenol - AFB₁ aflatoxin B₁ - DAS diacetyoxyscirpenol - DON deoxynivalenol - FB₁ fumonisin B₁ - FHB Fusaium head blight - MON moniliformin - NIV nivalenol - OA ochratoxin A - ZEA zearalenone     

The effect of cycloheximide on Euglena gracilis phenylalanyl-tRNA synthetases

The effect of cycloheximide on the chloroplastic, cytoplasmic and mitochondrial phenylalanyltransferRNA synthetases of Euglena gracilis was studied by growing both logarithmic and stationary phase cultures in the presence of the antibiotic. Enzyme activity was measured relative to untreated control cultures. At very low concentrations of cycloheximide (1 g/ml), all three log phase enzymes showed an increase in activity of 40–50%. At slightly higher concentrations (2.5 g/ml), the phenylalanyl-tRNA synthetase activities were comparable to those of the control cultures. At a cycloheximide concentration of 5g/ml the enzyme activities from stationary phase cultures showed only very slight decreases (5–20%). The cytoplasmic and mitochondrial enzymes behaved similarly in log phase cultures at this concentration. However, the chloroplastic phenylalanyl-tRNA synthetase from log phase cultures treated with 5g/ml cycloheximide showed a marked decrease in activity (70%). A further increase in antibiotic concentration to 10g/ml resulted in significant losses of activity of all three enzymes, from both growth stages. The implications of the data with regard to identification of the site(s) of chloroplast enzyme synthesis are discussed.     

Effect of α-Tocopherol Succinate on Free Radical and Lipid Peroxidation Levels in BL6 Melanoma Cells

Numerous studies have proposed a radical or oxidant involvement in a number of degenerative diseases such as cancer. This has led to suggestions that the supplementation of antioxidants such as α-tocopherol (vitamin E) may function to reduce the growth of cancer. In this study, a nonmalignant Monkey kidney (LLCMK) and a malignant Murine melanoma (BL6-F10) cell line were supplemented with varying levels of α-Tocopherol acid succinate (vitamin E succinate) ranging from 1 to 10 μg/ml. BL6-F10 cells supplemented with 5, 7, and 10 μg/ml vitamin E succinate, showed significant decreases in cell proliferation, and this decrease was accompanied by a concomitant increase rather than a decrease in the levels of free radicals and lipid peroxidation. LLCMK cells supplemented with 1–10 μg/ml vitamin E succinate showed no significant increase or decrease in growth, while the levels of lipid peroxidation were shown to be insignificantly elevated at 5, 7, and 10 μg/ml vitamin E succinate. Free radical levels in LLCMK cells were significantly decreased at 1 μg/ml vitamin E succinate, while at 3, 5, 7, and 10 μg/ml supplementary vitamin E succinate, free radical levels increased compared to the 1 μg/ml group, but not compared to control cultures. These results suggest that the inhibitory effects of vitamin E succinate on BL6-F10 cell growth in vitro is not a consequence of its antioxidant properties, but may, in fact, be due to one or more of its other potential roles within the cells, such as the regulation of cellular enzyme activities involved in growth. © 1997 Elsevier Science Inc.     

Antimicrobial Butyrolactone I Derivatives from the Ecuadorian Soil Fungus Aspergillus terreus Thorn. var terreus

Summary The fungus Aspergillus terreus Thorn var. terreus isolated from an Ecuador soil sample was cultured in liquid and solid media and yielded three main metabolites identified as terreic acid (1), butyrolactone I (2) and lovastatin (3). The natural products as well as three synthetic butyrolactone I derivatives were assessed for antimicrobial activity against Gram-positive and Gram-negative bacteria and fungi as well as for seed germination and seedling growth. Furthermore, the compounds were assessed as inhibitors towards the enzymes acetylcholinesterase, β-glucosidase, and β-glucuronidase. Terreic acid, butyrolactone I, butyrolactone 4′,4′′-diacetate (2.1), and 3′-(3-methylbutyl)-butyrolactone II (2.2) were active towards the phytopathogenic bacteria Erwinia carotovora with IC₅₀ of 5 and 4–18 μg/ml, respectively. Under the same experimental conditions, the IC₅₀ of streptomycin was 1.9 μg/ml. 3′-(3-Methylbutyl)-butyrolactone II was moderately active against Pseudomonas syringae and Botrytis cinerea with IC₅₀ of 21μg/ml and MIC of 15.6 μg/ml, respectively. Butyrolactone I also inhibited germination of the dicot Lactuca sativa with an IC₅₀ of 5 × 10⁻⁵ M. The IC₅₀ of reference herbicide acetochlor was 1 × 10⁻⁵ M. The effect of 2.2 and 2.3, known as butyrolactone III on Panicum millaceum germination and growth was stronger than that of 2 and 2.1. Reduction of the double bond in the isoprenyl side chain of butyrolactone I increased the antibacterial effect against E. carotovora as well as acetylation. To our best knowledge, this is the first report on the antibacterial effect of butyrolactone derivatives towards Erwinia carotovora and the phytopathogenic fungus Botrytis cinerea. The butyrolactone I derivative 2.2 presented a moderate inhibitory effect against the enzyme acetylcholinesterase with an IC₅₀ of 47 μg/ml. Under the same experimental conditions, the reference inhibitor galanthamine had an IC₅₀ of 3 μg/ml.     

Evaluation of antiprotozoal and antimycobacterial activities of the resin glycosides and the other metabolites of Scrophularia cryptophila

Resin glycosides are secondary metabolites exclusive to the convolvulaceous plants. In this study, crypthophilic acids A–C (1–3), the first resin glycosides occurring in another family (Scrophulariaceae), and the other constituents of Scrophularia cryptophila were examined for in vitro antiprotozoal and antimycobacterial potentials. Except for crypthophilic acid B (2), all tested compounds exhibited growth-inhibitory effect against Trypanosoma brucei rhodesiense, with l-tryptophan (6) and buddlejasaponin III (7) being the most potent ones (IC₅₀'s 4.1 and 9.7 μg/ml). In contrast, the activity towards Trypanosoma cruzi was poor, and only crypthophilic acid C (3), 6 and 7 were trypanocidal at concentrations above 40 μg/ml. With the exception of 2 and 6, all compounds were active against Leishmania donovani. Harpagide (4) and 3 emerged as the best leishmanicidal agents (IC₅₀'s 2.0 and 5.8 μg/ml). Only compounds 3, 6 and 7 showed antimalarial activity against Plasmodium falciparum with IC₅₀ values of 4.2, 16.6 and 22.4 μg/ml. Overall the best and broadest spectrum activity was presented by compounds 3 and 7, as they inhibited all four parasitic protozoa. None of the isolates had significant activity against Mycobacterium tuberculosis (MICs >100 μg/ml) or were toxic towards mammalian (L6) cells. This is the first report of antiprotozoal activity for natural resin glycosides, as well as for harpagide (4), acetylharpagide (5), tryptophan (6) and buddlejasaponin III (7).     

Control of blast and sheath blight diseases of rice using antifungal metabolites produced by Streptomyces sp. PM5

Two antifungal aliphatic compounds, SPM5C-1 and SPM5C-2 with a lactone and ketone carbonyl unit, respectively obtained from Streptomyces sp. PM5 were evaluated under in vitro and in vivo conditions against major rice pathogens, Pyricularia oryzae and Rhizoctonia solani. These compounds were dissolved in distilled water/medium to get the required concentrations. The well diffusion bioassay indicated that the of SPM5C-1 remarkably inhibited the mycelial growth of P. oryzae and R. solani in comparison to SPM5C-2. Though SPM5C-2 showed low antifungal activity against P. oryzae, it was not active against R. solani. Further, SPM5C-1 completely inhibited the growth of P. oryzae and R. solani at concentrations of 25, 50, 75 and 100 μg/ml. Greenhouse experiments revealed that spraying of SPM5C-1 at 500 μg/ml on rice significantly decreased blast and sheath blight development by 76.1% and 82.3%, respectively, as compared to the control with a corresponding increase in rice grain yield.     

In vitro susceptibilities of clinical yeast isolates to three antifungal agents determined by the microdilution method

A comparative evaluation of the in vitro susceptibilities of 597 clinical yeast isolates to amphotericin B, fluconazole, and 5-fluorocytosine (5FC) was conducted. The broth macrodilution reference method of the National Committee for Clinical Laboratory Standards (NCCLS, M27-P) was adapted to the microdilution method. Microdilution endpoints for amphotericin B were scored as the lowest concentration in which a score of 0 (complete absence of growth) was observed and for 5FC and fluconazole as the lowest concentration in which a score of 2 (prominent decrease in turbidity; MIC-2) was observed compared to the growth control. The MIC values were read after 24 and 48 h incubation. A broad range of MIC values was observed with each antifungal agent. Amphotericin B was very active (MIC₉₀1.0 µg/ml) against all of the yeast isolates with the exception ofC. lusitaniae (MIC₉₀2.0 µg/ml). Fluconazole was most active againstC. parapsilosis (MIC₉₀ of 1.0 µg/ml) and least active againstC. krusei (MIC₉₀ of 32 µg/ml). 5FC was most active againstC. albicans, C. parapsilosis, C. tropicalis, andT. glabrata (MIC₉₀1.0 µg/ml) and was least active againstC. krusei andC. lusitaniae (MIC₉₀16 µg/ml). These data indicate that the microdilution method, performed in accordance with M27-P, provides a means of testing larger numbers of yeast isolates against an array of antifungal agents and allows this to be accomplished in a reproducible and standardized manner. Given these results, it appears that the microdilution method may be a useful alternative to the macrodilution reference method for susceptibility testing of yeasts.     

Production of type A trichothecenes and enniatin B byFusarium sambucinum Fuckel sensu lato

Twenty-nineFusarium isolates, representing three new taxa originated by Nirenberg fromF. sambucinum Fuckel sensu lato, namely:F. sambucinum Fuckel sensu stricto,F. venenotum Nirenb., andF. torulosum (Berk. & Curt.) Nirenb., were tested for in vitro production of toxic secondary metabolites on autoclaved corn kernels.F. sambucinum sensu stricto was able to produce type A trichothecenes and enniatin B (EB). In particular, amongst the 14 isolates tested, 5 produced only diacetoxyscirpenol (DAS) (up to 700 µg/g); 1 produced only neosolaniol (NEOS) (250 µg/g); 2 produced T-2 toxin (T-2) + NEOS (up to 175 and 150 µg/g, respectively); 1 produced NEOS + DAS (300 and 100 µg/g, respectively); and 5 produced DAS + EB (up to 500 and 140 µg/g, respectively). All six isolates ofF. venenotum were able to produce only DAS (up to 100 µg/g).F. torulosum produced no trichothecenes, but four out of nine tested isolates were able to produce EB (up to 140 µg/g). Zearalenones and type B trichothecenes were not found. The toxicity of the culture extracts towardsArtemia salina L. was correlated in general with the occurrence of the above toxins, except for someF. torulosum strains. However, the lack of correlation between the amounts of toxins recovered and toxic activity observed in theGeotrichum candidum Link ex Pers. andA. salina assays suggested the presence of unknown toxic compounds.     

Promoting effect of wood vinegar compounds on fruit-body formation ofPleurotus ostreatus

The promoting effect of wood vinegar compounds on the fruiting ofPleurotus ostreatus (Japanese name, Hiratake) was investigated. Not only crude wood vinegar but its components, 3,5-dimethylphenol, 2-methoxyphenol, butanoic acid and 1-pentanol, had the ability to promote fruit-body formation on liquid medium. For use of these promoters industrially, a test for practical cultivation was carried out using a commercial sawdust medium. The addition of 100 µg/ml butanoic acid and 100 µg/ml 2-methoxyphenol into the sawdust medium after removal of the surface mycelial layer (kinkaki in Japanese) produced 29 and 23% higher yields of fruit-bodies than the control cultures (137.2 g/bottle), respectively. The addition of the crude wood vinegar as a medium component into sawdust substrates in the concentration range of 0.1–6% increased yields of fruit-bodies by 21–42% over the control.     

Suppressive effect of streptomycin on the phagocytic activity of mouse peritoneal macrophages for Histoplasma capsulatum

Streptomycin inhibited the phagocytic activity of mouse peritoneal macrophages forHistoplasma capsulatum. The inhibitory effect was demonstrable following both in vitro and in vivo administration of drug. The observations from examination by direct smear were confirmed by culturing for viable phagocytized organisms. A simple and reproducible technique for the counting of viable phagocytized organisms was developed. Forty-eight hours in vitro treatment of macrophage cultures with 10 to 200 µg/ml of streptomycin produced a graded inhibition of phagocytic activity, minimal at 10 µg/ml and maximal at 200 µg/ml of streptomycin. The parenteral administration of streptomycin significantly reduced phagocytic activity of mouse peritoneal macrophages forH. capsulatum. Mice were treated daily with the subcutaneous injections of 5, 2.5 or 1 mg streptomycin or saline. At 7, 14, 21 and 28 days post-treatment phagocytic activity of macrophages obtained from these mice was tested. There was a progressive, dose-dependent decrease in the phagocytic activity of macrophages derived from streptomycin-treated mice.     

Growth inhibitory activity of extracts and compounds from Cimicifuga species on human breast cancer cells

The purpose of this report is to explore the growth inhibitory effect of extracts and compounds from black cohosh and related Cimicifuga species on human breast cancer cells and to determine the nature of the active components. Black cohosh fractions enriched for triterpene glycosides and purified components from black cohosh and related Asian species were tested for growth inhibition of the ER⁻ Her2 overexpressing human breast cancer cell line MDA-MB-453. Growth inhibitory activity was assayed using the Coulter Counter, MTT and colony formation assays.Results suggested that the growth inhibitory activity of black cohosh extracts appears to be related to their triterpene glycoside composition. The most potent Cimicifuga component tested was 25-acetyl-7,8-didehydrocimigenol 3-O-β-d-xylopyranoside, which has an acetyl group at position C-25. It had an IC₅₀ of 3.2 μg/ml (5 μM) compared to 7.2 μg/ml (12.1 μM) for the parent compound 7,8-didehydrocimigenol 3-O-β-d-xylopyranoside. Thus, the acetyl group at position C-25 enhances growth inhibitory activity.The purified triterpene glycoside actein (β-d-xylopyranoside), with an IC₅₀ equal to 5.7 μg/ml (8.4 μM), exhibited activity comparable to cimigenol 3-O-β-d-xyloside. MCF7 (ER⁺Her2 low) cells transfected for Her2 are more sensitive than the parental MCF7 cells to the growth inhibitory effects of actein from black cohosh, indicating that Her2 plays a role in the action of actein. The effect of actein on Her2 overexpressing MDA-MB-453 and MCF7 (ER⁺Her2 low) human breast cancer cells was examined by fluorescent microscopy. Treatment with actein altered the distribution of actin filaments and induced apoptosis in these cells.These findings, coupled with our previous evidence that treatment with the triterpene glycoside actein induced a stress response and apoptosis in human breast cancer cells, suggest that compounds from Cimicifuga species may be useful in the prevention and treatment of human breast cancer.     

Optimum growth conditions and light utilization efficiency of Spirulina platensis (= Arthrospira fusiformis) (Cyanophyta) from Lake Chitu, Ethiopia

Spirulina platensis (= Arthrospira fusiformis) was isolated from Lake Chitu, a saline, alkaline lake in Ethiopia, where it forms an almost unialgal population. Optimum growth conditions were studied in a turbidostat. Cultures grown in modified Zarrouk's medium and exposed to a range of light intensities (20–500 µmol photons m^–2s^–1) showed a maximum specific growth rate (µ_max) of 1.78 d^–1. Quantum yield for growth (µ) was 3.8% at the optimum light for growth of 330 µmol photons m^–2s^–1, and ranged from 2.8 to 9.4%. With increase in irradiance, the chlorophyll a concentration decreased, and the carotenoids/chlorophyll a ratio increased by a factor of 2.4. The phosphorus to carbon ratio (P/C) showed some variation, while the nitrogen to carbon ratio (N/C) remained relatively constant, thus causing fluctuations in the N:P ratio (7–11) of cells. An optimum N:P ratio of about 7 was attained in cells growing at the optimum light for growth. Results from the continuous culture experiments agreed well with maximum values of photosynthetic efficiency given in the literature for natural populations of S. platensis in the soda lakes of East Africa, Lake Arenguade (Ethiopia), and Lake Simbi (Kenya).     

Aflatoxin M1 removal from aqueous solutions by Flavobacterium aurantiacum

In liquid cultures growing and stationary phase cells ofFlavobacterium aurantiacum NRRL B-184 eliminated aflatoxin M₁. Toxin concentrations of 15µg/ml and 37.5µg/ml interfered with bacterial growth, and at the higher level 4.4µg M₁ was removed from the growth medium by a milligram (dry weight) of bacteria. Toxin was completely removed from the liquid medium by incubating 5 × 10¹⁰ resting cells per milliliter with 8µg/ml of aflatoxin M₁ for 4 h. Attempted recovery of M₁ from cells following incubation of the bacteria with the toxin demonstrated that the M₁ was essentially nonextractable. Bacterial cells also removed aflatoxin M₁ from toxin-contaminated milk.