Plasmid incidence in bacteria from agricultural and industrial soils

Heavy metal contents of agricultural and industrial soils were determined by atomic absorption spectrophotometry. The analysis of the samples collected from two different locations revealed significantly high levels of Fe, Zn, Cu, Cr and Ni. Certain microbiological parameters (total aerobic heterotrophs, asymbiotic N₂-fixers, total Actinomycetes and fungi) were also monitored from these soils. A total of 70 bacterial isolates from agricultural and industrial soils were examined for plasmid DNA content and resistance to the antibiotics amoxycillin, cloxacillin, chloramphenicol, doxycycline methicillin, nalidixic acid, and tetracycline. Minimum inhibitory concentrations (MICs) of Cu, Cr, Pb, Cd, Hg, Zn, and Ni for each isolate were also determined. Resistance was most frequent to methicillin (48.5%), cloxacillin (45.7%), and nalidixic acid (40%) for all isolates of bacteria. The highest MICs observed were 100 g/ml for mercury, 800 g/ml for Ni and 1600 g/ml for other metals. The incidences of metal resistance and MICs of metals for bacteria from industrial soil were significantly different to those of agricultural soil. On a percentage basis, 91.4% of the total bacterial isolates from industrial soil were found to harbour plasmids whereas 40% of the isolates from agricultural soil contained plasmids.     

PCR amplification of four genes coding for aminoglycoside-modifying enzymes in bacteria of clinical isolates from Jordan University Hospital

Three species of Gram-negative G(–) bacteria were chosen for this study: Escherichia spp. (29 isolates), Pseudomonas spp. (16 isolates) andEnterobacter spp. (17 isolates), utilizing colony PCR to detect genes coding for aminoglycoside-modifying enzymes. The 62 isolates were purified and cultured on nutrient broth media supplemented with 50 g kanamycin/ml. Only 17 out of the 62 isolates were resistant to kanamycin and were subjected to colony PCR protocol using 20 l cell lysate and eight sets of primers with each isolate. Sizing of the amplified fragments was carried out in order to determine the specificity of PCR. The 17 isolates were shown to carry the rrs, aacC2, aacA-aphD and aphA3 genes.     

Bacterial breakdown of benomyl. I. Pure cultures

With different soil and water samples as inoculum and the benzimidazole fungicides benomyl (either as Benlate or as the pure compound) and thiabendazole as selective agents, a large number of, mainly fluorescent, Pseudomonas strains were isolated which nearly all were able to grow in a mineral medium with benomyl as the sole source of carbon. However, no growth occurred with any of a series of other benzimidazole compounds, viz. benzimidazole, 2-aminobenzimidazole (2-AB), thiabendazole and fuberidazole. Although benomyl—or rather its non-enzymatic breakdown product methyl benzimidazol-2-yl carbamate (MBC)—was partially degraded to 2-AB, most probably n-butylamine, which arises after splitting off of the butylcarbamoyl side chain, was the actual carbon source for the Pseudomonas isolates.When incorporated in a lactate medium, 2-AB markedly inhibited the growth of Pseudomonas spp. at a concentration of 250 g/ml, with complete inhibition being attained at 500 g/ml. For Bacillus spp. grown in liquid peptone media benzimidazole compounds were inhibitory at concentrations of 500–1000 g/ml, with a toxicity increasing in the order: benzimidazole 相似文献   

Antibiotic resistance and detection of β-lactamase in bacterial strains of Staphylococci and Escherichia coli isolated from foodstuffs

Seventy strains of Staphylococcus spp. and Escherichia coli (35 each) were isolated from various foodstuffs and identified on the basis of cultural, morphological and biochemical characteristics and were further tested for their antibiotic susceptibility with commonly used antibiotics/drugs. 69.2% of the strains of Staphylococci were resistant to co-trimazine and 34.6% were resistant to penicillin-G. 19.2% of the staphylococcal isolates exhibited resistance to cloxacillin, nalidixic acid, methicillin and tetracycline whereas 15.3% of the staphylococcal isolates were resistant to amoxycillin and nitrofurantoin. The isolated E. coli strains exhibited sharp peaks of resistance to antimicrobial agents such as tetracycline (72%), doxycycline (60%) and nalidixic acid (48%). Forty-four percent of the E. coli strains were resistant to nitrofurantoin and penicillin-G respectively. Among the 13 antibiotics/drugs tested for resistance, six different resistance patterns were observed in staphylococcal isolates and seven different resistance patterns were observed in the E. coli isolates from various foodstuffs. Bacterial strains exhibiting MIC values 100 g/ml for ampicillin and cloxacillin were screened for -lactamase activity and out of 10 staphylococcal isolates, seven were found to be positive for -lactamase, whereas out of 13 E. coli isolates tested for -lactamase production, only three were found to be positive.     

Metal resistance in Pseudomonas strains isolated from soil treated with industrial wastewater

Heavy metal concentrations in soil treated with industrial wastewater of Aligarh City (U.P.), India were determined. The analysis of test samples revealed high levels of Fe, Zn, Ni and Cu. A total of 45 Pseudomonas spp. were isolated from soil and were characterized on the basis of morphological, cultural and biochemical characteristics. MICs of Hg²⁺, Cd²⁺, Cu²⁺, Cr³⁺, and Zn²⁺ for each isolate were determined. Eighty percent of the strains isolated from soil harboured resistance to copper, whereas 73.3% of the isolates exhibited resistance to cadmium, 71.1% to chromium and zinc and 48.8% to mercury. A maximum MIC of 200 g/ml for mercury and 1600 g/ml for other metals was observed. Metal resistance was found to be plasmid mediated as evidenced by transformation studies. Further, the transmissible nature of chromium resistance was confirmed by conjugation. Agarose gel electrophoresis using the miniprep method for plasmid isolation revealed that these isolates harboured plasmids of molecular weights (45 & 47 kb) using EcoRI and HindIII digests of DNA and undigested DNA as standard markers.     

Toxicity of carbofuran to soil isolates of Chlorella vulgaris,Nostoc linckia and N. muscorum

A microalga, Chlorella vulgaris, and two diazotrophic cyanobacteria, Nostoc linckia and N. muscorum, all isolated from a rice soil, were compared for their response in terms of growth and metabolic activities to the application of carbofuran. The toxicity criteria included cell constituents (chlorophyll a, total protein, carbohydrate), ¹⁴CO₂ uptake and nitrate reductase, besides nitrogenase activity (acetylene reduction) in the cyanobacteria. C. vulgaris and N. muscorum were more sensitive to carbofuran than was N. linckia. The significant toxicity of the insecticide, observed with higher concentrations of 20 and 50 g ml^–1, to nitrogenase activity in N. linckia was reversed by the addition of ATP at 10 M. Transmission electron microscopy of the cultures, exposed to 50 g carbofuran ml^–1 showed certain cellular abnormalities, indicating interference of the insecticide with membrane properties. Correspondence to: K. Venkateswarlu     

Effect of malathion on nucleic acid synthesis in phytohemagglutinin-stimulated human lymphocytes

Summary The effect of malathion, an organophosphorus insecticide, on DNA and RNA synthesis was investigated by measuring the rate of incoporation of ³H thymidine and ³H uridine, respectively, into human lymphocytes stimulated by phytohemagglutinin (PHA). Increasing concentrations of malathion, from 10 to 70 g/ml, were added to human lymphocyte cultures at different times in relation to PHA introduction. The lowest applied dose of malathion (10 g/ml) in most cases led to increased incorporation of both ³H thymidine and ³H uridine. Higher concentrations of malathion (30, 50, 70 g/ml) caused a time- and dosedependent decrease of radioisotope incorporation.     

Endophytic fungi from Musa acuminata leaves and roots in South China

One hundred and sixty-three endophytic fungal cultures were isolated from 200 leaf samples of Musa acuminata trees, which were soaked in 36% formaldehyde solution for surface sterilization. They belonged to the genera of Gloeosporium musae (45%), Myxosporium spp. (11%), Deightoniella torulosa (8.5%), Alternaria tenuis (7.9%), Sphaceloma spp. (7.4%), Aureobasidium spp. (4.3%), Melida spp. (1.8%), Uncinula spp. (1.8%), Penicillium spp. (1.8%), Aspergillus spp. (1.2%), Sarcinella spp. (1.2%), Cladosporium sp. (0.6%), Cephalosporium sp. (0.6%) and sterile mycelium (6.7%). Sixty-eight endophytic fungal cultures were isolated from 100 root samples. They respectively belonged to the genera of Aspergillus spp. (31%), Paecilomyces spp. (16%), Penicillium spp. (15%), Fusarium spp. (10%), Gloeosporium musae (6%), yeast (3%), Deightoniella torulosa (3%), Spicaria sp. (1.4%), Cephalosporrium sp. (1.4%), Meliola sp. (1.4%) and sterile mycelium (10%). Water agar (containing 50 g chloramphenicol ml^–1 and 50 g streptomycin ml^–1) seemed to be a better medium for isolation of endophytic fungi than potato-dextrose agar (PDA, containing 50 g chloramphenicol ml^–1 and 50 g streptomycin ml^–1).     

Removal of uranium by an exopolysaccharide from Pseudomonas sp.

Summary Accumulation of uranium (U) is reported for isolated exopolysaccharide produced by Pseudomonas sp. EPS-5028. A maximum uptake of 96 g U/mg polymer was observed. In contrast, the maximum accumulation of uranium by deacylated polysaccharide was 46 g/mg. This metal-complexing capacity observed suggests that the anionic reactive sites on the structure could be responsible for this activity. Metal uptake was affected by pH and was not affected by temperature. Expolysaccharide from Pseudomonas sp. EPS-5028 obeyed the Freundlich isotherm indicating single layer adsorption. Offprint requests to: F. Congregado     

Effect of minerals on the production of the delta endotoxin byBacillus thuringiensis subsp.israelensis

Summary A study has been made of the mineral requirements ofBacillus thuringiensis subsp.israelensis for production of the mosquitocide delta endotoxin. The optimum concentrations of K₂HOP₄, MgSO₄.7H₂O and CaCO₃ for toxin production are 1g/l, 0.3g/l and 1g/l respectively while the elements Fe, Mn, Cu are required at levels of 2 g/ml, 5 g/ml and 0.25 g/ml respectively.     

In vitro comparison of ceftazidime,aztreonam, cefpirome,gentamicin, imipenem,enoxacin, and ticarcillin plus clavulanic acid against 108 strains ofPseudomonas maltophilia

Pseudomonas maltophilia is an uncommon cause of hospital-acquired infection and is resistant to most of the antimicrobial agents used in the treatment of gram-negative infections. Susceptibility of 108 isolates ofP. maltophilia to ceftazidime, aztreonam, defpirome, gentamicin, imipenem, enoxacin, and ticarcillin plus clavulanic acid was determined by an agar dilution method. The isolates were in general resistant to the antibiotics. Imipenem and cefpirome were not active at clinically achievable levels. Of the isolates, 20% were susceptible to 16 g/ml ceftazidime, 53% were susceptible to 4 g/ml enoxacin, 10% were susceptible to 4 g/ml gentamicin, and 25% were susceptible to 64 g/ml ticarcillin plus 2 g/ml clavulanic acid.     

Untersuchungen zur Regulation der Bacteriochlorophyll-Synthese bei Rhodospirillum rubrum

Zusammenfassung Wenn Rhodospirillum rubrum aus aeroben Dunkelkulturen in ein synthetisches Medium übertragen und anaerob im Licht bebrütet wird, beginnt die Bacteriochlorophyllbildung bereits nach 1 Std, das Wachstum erst nach 5–8 Std Bebrütung. — Puromycin (10 g/ml) und Chloramphenicol (20 g/ml) hemmen in diesen Kulturen Protein- und Pigmentsynthese vollständig. Eine Hemmung wird auch beobachtet, wenn die Antibiotica erst mehrere Stunden nach Beginn der Lichtbebrütung zugesetzt werden. Synthese von Thylakoidprotein und Bacteriochlorophyll scheinen regulatorisch gekoppelt zu sein.Actinomycin C (D) (40 g/ml) und Mitomycin (1 g/ml) hemmen die Bacteriochlorophyllbildung enbenfalls. Die Thylakoidbildung ist vom Vorhandensein einer funktionsfähigen DNS und RNS abhängig.Die spezifische Aktivität der -Aminolaevulinsäuresynthetase nimmt in anaeroben Lichtkulturen gegenüber aeroben Dunkelkulturen um etwa das Vierfache zu. Sie wird durch die verwendeten antibiotica nicht beeinflußt. Die Biosynthese des Enzyms wird durch Mitomycin und Puromycin, nicht aber durch Actinomycin gehemmt.

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Summary If dark-aerobically grown Rhodospirillum rubrum is transferred to anaerobic conditions in the light, the synthesis of photosynthetic pigments starts after 1 or 2 hours. The growth of the culture begins in the synthetic medium not before 5 hours incubation. In these cultures Puromycin (10 g/ml) and Chloramphenicol (20 g/ml) inhibit synthesis of protein and bacteriochlorophyll both. An inhibition is also observed when the antibiotics are added some hours after the beginning of anaerobic light incubation.The synthesis of chromatophore protein and bacteriochlorophyll are likely connected by gen-regulation.Actinomycin C (D) (40 g/ml) and Mitomycin C (1 g/ml) inhibit the bacteriochlorophyll synthesis likewise. The effect of actinomycin is increased by preincubation with the antibiotic in the dark. Mitomycin C stops synthesis of bacteriochlorophyll and protein even if it added after preincubation in the light.The level of -aminolevulinic acid-synthetase increased fourfold in anaerobic light-cultures compared to dark-aerobically grown cells. The activity of the enzyme is not influenced by the antibiotics. But the rate of biosynthesis is inhibited by Puromycin and Mitomycin, but not by Actinomycin.

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Abkürzungen im Text ALS -Aminolaevulinsäure - B-Chlorophyll Bacteriochlorophyll - DNS Desoxyribonucleinsäure - RNS Ribonucleinsäure     

Factors affecting the survival and growth of bacteria introduced into lake water

The populations of Pseudomonas sp. B4, Escherichia coli, Klebsiella pneumoniae, Micrococcus flavus, and Rhizobium leguminosarum biovar phaseoli declined rapidly in lake water. The initially rapid decline of the two pseudomonads and R. phaseoli was followed by a period of slow loss of viability, but viable cells of the other species were not found after 10 days. The rapid initial phase of decline was not a result of Bdellovibrio spp., bacteriophages, or toxins in the water since Bdellovibrio spp. were not present and passage of the lake water through filters that should not have removed bacteriophages or soluble toxins led to the elimination of the rapid phase of decline. The addition of 250 g of cycloheximide and 30 g of nystatin per ml eliminated viable protozoa form the lake water, and the population of Pseudomonas sp. B4 did not fall and the decline of E. coli and K. pneumoniae was delayed or slowed under these conditions. Pseudomonas sp. L2 proliferated rapidly in lake water amended with glucose, phosphate, and NH₄NO₃, but its numbers subsequently fell abruptly; however, in water amended with cycloheximide and nystatin, which killed indigenous protozoa, the population density was higher and the fall in numbers was delayed. Of the nutrients, the chief response was to carbon, but when glucose was added, phosphorus and nitrogen stimulated growth further. Removing other bacteria by filtering the lake water before inoculation with Pseudomonas sp. L2 suggested that competition reduced the extent of response of the pseudomonad to added nutrients. We suggest that the decline in lake water of bacteria that are resistant to starvation may be a result of protozoan grazing and that the extent of growth of introduced species may be limited by the supply of available carbon and sometimes of nitrogen and phosphorus, and by predation by indigenous protozoa.     

Production of trichothecene and non-trichothecene mycotoxins by Fusarium species isolated from maize in Minnesota

Eighty-two cultures of Fusarium species isolated in 1986 from moldy maize in Minnesota were each cultured on rice for 4 weeks and found to produce the following mycotoxins: F. graminearum isolates, deoxynivalenol (DON, 4–225 g/g), 3-acetyldeoxynivalenol (3-ADON, 2–4g/g), 15-acetyldeoxynivalenol (15-ADON, 1–35 g/g) and zearalenone (ZEA, 5–4350 g/g); F. moniliforme, fusarin C (detectable amounts to 1000 g/g); F. mòniliforme, F. oxysporum, F. proliferatum and F. subglutinans isolates, moniliformin (15–6775 g/g); F. moniliforme, F. proliferatum, and F. subglutinans isolates, fusaric acid (detectable amounts). Other mycotoxins screened for in each rice sample and not detected were T-2 toxin, HT-2 toxin, neosolaniol, T-2 tetraol, nivalenol, fusarenon-X, scirpenols, alpha and beta trans-zearalenols, wortmannin, and fusarochromanone. The rat feeding bioassay indicated that other, unidentified toxins may be present.     

Morphological engineering of Streptomyces hygroscopicus var. geldanus: regulation of pellet morphology through manipulation of broth viscosity

Actinomycetes, especially members of the genus Streptomyces, are responsible for producing the majority of known antibiotics. The production of antibiotics by filamentous organisms is often dependent on the morphology and size distribution of the pellet population within the culture. Particle interaction and subsequent pellet formation are primarily dependent on the rate of collision of particles in culture, which is in turn, a function of fluid turbulence. The microbial polysaccharide xanthan gum was used to artificially regulate the apparent viscosity (_a) of S. hygroscopicus fermentation broths with the aim of controlling particle interaction, aggregation and hence pellet formation. An increase in both pellet count and biomass concentration from approximately 2,000 to 8,000 pellets ml^–1 and 0.9–2.1 g l^–1 dry weight of biomass, as well a decrease in the mean pellet volume from 0.014 to 0.004 mm³ was observed in cultures supplemented with 3 g l^–1 xanthan gum. The addition of xanthan gum significantly alters fluid rheology by increasing the _a. Counter-intuitively, an increase in the _a within the experimental range examined resulted in an increase in the rate of gas–liquid mass transfer. This was attributed to the predominantly diffusive nature of oxygen transfer in shake flask cultures.     

Untraditional N2-fixing bacteria as biofertilizers for wheat and barley

The screening of 27 isolates grown on nitrogen-free medium for nitrogen-fixing ability resulted in the isolation of five organisms belonging toBacillaceae, Enterobacteriaceae andPseudomonadaceae. Estimates of N₂-fixation efficiencies of these isolates indicated that they may be responsible for low rates of N₂-fixation in soil. The possible association of these isolates as well as ofAzotobacter andAzospirillum with wheat and barley was investigated in a greenhouse experiment. The highest values of nitrogenase activity on plant root were recorded in treatments inoculated with composite inocula of the isolated N₂-fixers, particularly whenAzotobacter and/orAzospirillum were added in combination. Inoculation with single inoculum of each of the N₂-fixing isolates had no significant influence on plant growth, except withPseudomonas andBacillus for wheat and barley, respectively. Highly significant increases in growth of both plants were recorded in all cases of multistrain inoculation.     

Toxicity and moniliformin production by four recently described species of Fusarium and two uncertain taxa

Four recently described species, Fusarium nygamai, F. dlamini, F. beomiforme and F. napiforme and two uncertain taxa, F. nygamai from millet in Africa and Fusarium species from rice with Bakanae disease, were tested for toxicity and moniliformin production. Cultures grown on autoclaved corn were fed to groups of four one-day-old ducklings for 14 days. Isolates that caused the death of 3 or 4 out of 4 ducklings were considered to be toxic and analyzed for moniliformin. All 15 isolates of F. dlamini tested were nontoxic. The other taxa contained some isolates that were toxic to ducklings and produced moniliformin in corn cultures. This is the first report of moniliformin production by F. beomiforme (200–890 g/g), and F. napiforme (16–388 g/g), and by F. nygamai not obtained from millet in Africa (15–874 g/g). The highest production of moniliformin was obtained from the 19 isolates of F. nygamai from millet in Africa (4300–18200g/g) and the 15 isolates from rice with Bakanae disease (2300–19300 g/g). The taxonomic position of these two uncertain taxa should be re-evaluated.     

Effect of Naloxone on the Activity of Cl–-Activated Mg2+-ATPase from Plasma Membrane Fraction of Bream Brain (Abramis brama L.) in the Presence of GABAa-ergic Substances

We studied the effect of naloxone—an antagonist of the opioid receptors—on sensitivity of Cl^–-activated Mg²⁺-ATPase from the plasma membrane fraction of bream brain (Abramis brama L.) to GABA_a-ergic substances. Preincubation of the plasma membranes with 1–100 M naloxone increased the basal Mg²⁺-ATPase activity and suppressed its activation by chloride ions. The same effects were observed in the presence of the agonists of GABA_a/benzodiazepine receptors: 0.1–100 M GABA, 1–500 M pentobarbital, and 0.1–100 M phenazepam. Naloxone (10 M) inhibited activation of the basal Mg²⁺-ATPase by the studied ligands and restored the enzyme sensitivity to Cl^–. However, the effect of naloxone was not observed in the presence of high concentrations of pentobarbital (500 M) and phenazepam (100 M). The obtained data show that naloxone modulates the activity of Cl^–-activated Mg²⁺-ATPase from the plasma membranes of bream brain and antagonizes the GABA_a receptor ligands.     

In vitro susceptibility ofTrichophyton rubrum isolates to griseofulvin and tioconazole. Induction and isolation of a resistant mutant to both antimycotic drugs

The in vitro susceptibility of three clinicalTrichophyton rubrum isolates to griseofulvin and tioconazole, determined by the minimal inhibitory concentration (MIC), was 2 and 0.5 to 1.0g/ml, respectively. One mutant (gril) obtained after mutagenic treatment of one of these isolates was selected and showed simultaneous resistance to griseofulvin (MIC > 2000g/ml) and tioconazole (MIC=1.0g/ml). The clinical importance and the possibility of a multidrug resistance (MDR)-type mechanism being involved in this event is discussed.     

Survey of petroleum-degrading bacteria in coastal waters of Sunderban Biosphere Reserve

A survey of petroleum-degrading bacteria was carried out in the Indian part of deltaic Sunderbans to evaluate the distribution of the naturally occurring petroleum-degrading aerobic bacteria. Bacteriological analysis of surface water samples collected from five different locations in the Hooghly–Matla river mouth showed that, depending on the location, 0.08–2.0% of the heterotrophic bacteria culturable in marine agar medium could degrade crude petroleum hydrocarbons as the sole source of carbon. In the entire study area, the number of heterotrophic bacteria ranged from 1 × 10³ to 3.8 × 10⁵ c.f.u/ml, amongst which 2.7 × 10¹ to 6 × 10³ c.f.u/ml were petroleum degraders. There was a maximum number of petroleum-degrading bacteria in the waters of Haldia Port and its surrounding areas, where the water is highly polluted by hydrocarbon discharges from a nearby oil refinery and from the ships docking at the port. Among the isolates, identified on the basis of their Gram reaction, morphological and biochemical tests including the use of API20E strips, Pseudomonas, Mycobacterium, Klebsiella, Acinetobacter, Micrococcus, and Nocardia were the most common petroleum degraders. Other heterotrophic bacteria included several species of Escherichia, Klebsiella, non-oil-degrading Pseudomonas, Vibrio, Streptococcus, Staphylococcus and Bacillus. Following preliminary selection, five strains, showing best growth in medium with oil fraction as sole carbon source, were chosen for estimation of the efficiency of crude oil biodegradation. The selected strains belonged to Pseudomonas (two strains), Mycobacterium (two strains), and Nocardia (one strain). These strains degraded 47–78% of Arab-Mix crude oil over a period of 20 days. The best oil-degrading isolate, a strain of Pseudomonas aeruginosa, (BBW1), was found to degrade and multiply more rapidly in crude oil than the rest. BBW1 showed profuse growth in Bushnell Haas medium containing crude oil (as sole source of carbon) at high concentrations ranging from 0.2 to 20% (v/v), with optimum at 10%. As much as 75% of the oil was degraded within 72 h of incubation with the bacteria. Physicochemical analysis showed considerable decrease in initial boiling point and carbon residue of the degraded oil. The ability to degrade crude oil was found to be associated with a single 70-kb plasmid, pBN70. Resistance to the metals Mn²⁺ (50 mM), Mg²⁺ (200 mM), Zn²⁺ (6 mM), Ni²⁺ (10 mM) and antibiotics like ampicillin (10 g/ml), cephalexin (30 g/ml), nitrofurantoin (300 g/ml) and penicillin (10 U/ml) were plasmid-mediated.