Azospirillum strains isolated from roots and rhizosphere soil of wheat (Triticum aestivum L.) grown under different soil moisture conditions

The present study deals with the isolation and characterization of Azospirillum strains isolated from roots and rhizosphere soil of wheat (at tillering and anthesis stages) plants grown under different moisture regimes in the field and in pots. The survival of Azospirillum isolates from plants of irrigated field and those from well-watered pots was higher than that of Azospirillum strains isolated from roots and rhizosphere soils of plants grown under arid and semiarid (14–8% soil moisture) field conditions and under water-stressed (8% soil moisture) conditions in pots. On the basis of carbon/nitrogen source utilization, the Azospirillum strains isolated from wheat under field and pot conditions were grouped in three groups. The unweighted pair group method with arithmetic means cluster analysis based on random amplification of polymorphic DNA showed that two groups of Azospirillum were similar. The strains isolated from plants (at tillering stage) grown under low moisture conditions either in pots or in field were genetically similar to strains isolated from plants grown under well-watered conditions in both pots and field. Inoculation of wheat with isolates from water-stressed plants induced tolerance to water stress in inoculated plants. Isolates from water-stressed conditions exhibited lower production of indole acetic acid, gibberellic acid, and trans zeatin riboside but a higher production of abscisic acid.     

Fatty acid composition and squalene content of the marine microalga Schizochytrium mangrovei

Certain species of thraustochytrids are being explored as potential producer of polyunsaturated fatty acids for nutritional enrichment of food products and use as feed additives in aquaculture. The fatty acid composition and squalene content were determined in the thraustochytrid, Schizochytrium mangrovei that was newly isolated from decaying Kandelia candel leaves in Hong Kong mangrove habitat. The major fatty acid constituents identified in all three S. mangrovei strains were tetradeanoic acid (C14:0), hexadecanoic acid (C16:0), docosapentaenoic acid (C22:5 n-6, DPA) and docosahexaenoic acid (C22:6 n-3, DHA). DHA was the most predominant polyunsaturated fatty acid, and the percentage of DHA (of total fatty acids) in all these strains varied from 32.29 to 39.14%. Only slight changes were observed in fatty acid composition of the S. mangrovei strains harvested at their early (day 3) and late stationary (day 5) phases. In contrast, the cellular squalene content was affected significantly by the culture time; the largest decrease of squalene content from 0.162 mg/g to 0.035 mg/g was found in S. mangrovei FB1 as the culture aged.     

石灰性褐土中磷锌镉相互作用对其有效性的影响

采用室内培养的方法,研究了石灰性褐土中磷、锌、镉相互作用对土壤中磷、锌、镉有效性的影响。结果表明：（1）磷锌共同培养时,施锌提高了土壤速效磷含量,且随培养时间的延长而降低。在相同锌浓度处理下,土壤中的有效锌含量随施磷量的增加而增加,不同锌浓度处理下,有效锌含量随土壤培养时间的延长而显著降低。（2）磷镉共同培养时,施镉对土壤速效磷含量影响不明显;施磷降低了有效镉含量,但效果不显著;且都随时间的延长而降低。（3）锌镉共同培养时,在培养的前30d,土壤中有效锌含量随施镉浓度增加而降低,但在30d后,有效锌含量有增加的趋势。土壤中有效镉含量在不同锌-镉处理下随培养时间变化有较大差异：在Cd3处理下,加入高浓度锌后显著降低土壤有效镉含量;Cd30处理下,在培养前30d,锌的施入对土壤中有效镉含量影响不明显,但30d以后,土壤有效镉含量随施入锌浓度的增加而显著降低。说明两者的竞争机制随时间的延长发生变化,且施锌能明显降低镉的毒性。     

Soil acidity and the composition of an indigenous population of Rhizobium tripolii in nodules of different cultivars of Trifolium subterraneum L.

Three methods of strain identification were used to determine the composition of the Rhizobium trifolii population in nodules formed on four cultivars of Trifolium subterraneum L. grown in a soil unamended (pH 4.8) or amended with CaCO₃ (pH 6.4). Whole cell somatic tube agglutination placed 68–90% of the nodule occupants from each of the cultivars Mt Barker, Nangeela and Howard into four identifiable serogroups. Only in the case of cultivar Woogeneliup were the majority of the isolates unidentifiable with the antisera at our disposal. In unamended soil different serogroups dominated each cultivar whereas in limed soil serogroup 27 was the dominant identifiable serogroup on all of the cultivars. The latter serogroup was also the most promiscuous nodule occupant being found in seven of eight treatments. Gel-immune-diffusion analysis revealed three serotypes within serogroup 27; serotype 27-A was dominant in the nodules on cultivar Nangeela alone whereas serotype 27-B was dominant on both cultivars Mt Barker and Woogenellup. Separation of cellular proteins by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) revealed four and five different strains within serotypes 27-A and 27-B respectively. Only one strain from serotype 27-B could be considered a common nodule occupant on cultivars Mt Barker and Woogenellup. Two of three strains, representing 82% of the total isolates of serotype 27-B found on cultivar Mt Barker, were nodule occupants of that cultivar in both soil treatments. In contrast, only one of four strains, representing 38.5% of the total isolates of serotype 27-A found on cultivar Nangeela, could be considered a common nodule occupant in both soil treatments.     

Emerging resistant microbiota from an acidic soil exposed to toxicity of Cr, Cd and Pb is mainly influenced by the bioavailability of these metals

Purpose

The effect of pollutants in soil microorganisms is an important issue in order to understand their toxic effects in the environment, as well as for developing adequate bioremediation strategies. In this sense, the main objective of this study was to assess the involvement of the indigenous microbiota of an acidic forest Mediterranean soil by artificial pollution with heavy metals, and to detect and isolate resistant microorganisms that could be useful for bioremediation.

Materials and methods

Samples from a previously unpolluted acidic forest soil were amended with Cr(VI), Cd(II) or Pb(II) at total amounts ranging from 0.1 to 5,000 mg?kg^?1. These soil microcosms were incubated under controlled laboratory conditions for 28 days. Soluble fractions of metals were determined from aqueous extracts. Both activity and composition of the microbial community were assessed, respectively, by respirometric assays and molecular analysis (polymerase chain reaction denaturing gradient gel electrophoresis). The isolation of metal-resistant microorganisms was attempted by culture plating from microcosms incubated with high concentrations of metals. Isolated strains were tested in cultures with minimal medium to check for their metal resistance and their capacity to reduce the presence of toxic Cr(VI).

Results and discussion

A decrease in the soil respirometric activity and changes in the microbial community composition were detected from 10/100 mg?kg^?1 Cr and 1,000 mg?kg^?1 Cd and Pb. Presumably resistant bacterial and fungal populations developed in most of these polluted microcosms; however, the microbiota was severely impaired at the highest additions of Cr. Even though Cr was the most damaging metal in soil microcosms, if the soluble fractions of metals are considered instead of their total added amounts, the comparison among their toxic effects suggests a similar potential toxicity of Cr and Pb. Isolated multiresistant microorganisms were related mainly to Actinobacteria, Firmicutes and Ascomycota. Some of them showed the capacity to reduce Cr(VI) concentrations between 54 % and 70 % of the initial value. These strains were affiliated to several species of Streptomyces and Bacillus.

Conclusions

The combination of respirometric assays with molecular methods has been useful to assess the effect of metals on the soil microbial community, which can greatly be explained by their differential bioavailability. Cultivation-dependent and -independent approaches have proved the presence and development of multiresistant microorganisms in a previously unpolluted soil. Due to their properties, some of the isolated strains are potentially useful for soil bioremediation.     

Effects of humic acids on the growth of bacteria

The influence of humic acids of different origins on the growth of bacterial cultures of different taxa isolated from the soil and the digestive tracts of earthworms (Aporrectodea caliginosa)—habitats with contrasting conditions—was studied. More than half of the soil and intestinal isolates from the 170 tested strains grew on the humic acid of brown coal as the only carbon source. The specific growth rate of the bacteria isolated from the intestines of the earthworms was higher than that of the soil bacteria. The use of humic acids by intestinal bacteria confirms the possibility of symbiotic digestion by earthworms with the participation of bacterial symbionts. Humic acids at a concentration of 0.1 g/l stimulated the growth of the soil and intestinal bacteria strains (66 strains out of 161) on Czapek’s medium with glucose (1 g/l), probably, acting as a regulator of the cell metabolism. On the medium with the humic acid, the intestinal bacteria grew faster than the soil isolates did. The most active growth of the intestinal isolates was observed by Paenibacillus sp., Pseudomonas putida, Delftia acidovorans, Microbacterium terregens, and Aeromonas sp.; among the soil ones were the representatives of the Pseudomonas genus. A response of the bacteria to the influence of humic acids was shown at the strain level using the example of Pseudomonas representatives. The Flexom humin preparation stimulated the growth of the hydrocarbon-oxidizing Acinetobacter sp. bacteria. This effect can be used for creating a new compound with the elevated activity of bacteria that are destroyers of oil and oil products.     

Survival of Escherichia coli in soil with modified microbial community composition

Understanding the survival and persistence of Escherichia coli in soil with different microbial composition is essential for the accuracy of water quality assessment and microbial source tracking. This microcosm experiment was conducted to investigate the survival pattern of three E. coli strains (originated from soil, dog feces and human feces, separately) in soil with modified microbial community composition. Bile salt No. 3 (BS3) of progressively increased density (0.05%, 0.15%, 0.30% and 0.50%) was added into sandy loam soils and incubated for 90 days. Laboratory cultured E. coli were then inoculated into soil and incubated for another 150 days to monitor their survival pattern. Change of bacterial community diversity by BS3 was detected by both cultivation based and cultivation independent (PCR-Denaturing Gradient Gel Electrophoresis) methods. In general, progressively increased BS3 concentration resulted in decreased CFU counts both at 10 days and 90 days incubation. DGGE analysis indicated only a slight change in bacterial community composition at 10 days but a significant change at 90 days. Cluster analysis suggested that BS3 treatment grouped separately from controls. Survival of E. coli in soil was significantly influenced by the complexity of the microbial community, as die-off rate of E. coli progressively declined with the reduction of microbial community diversity. Differential survival of E. coli under different soil microbial stress highlights the importance of incorporating biotic factors in predictive models for water quality management and microbial source tracking study.     

Studies on the bacteria isolated anaerobically from paddy field soil IV. Model experiments on the production of branched-chain fatty acids

Based on the fact that the clostridia isolated from paddy field soil were mostlyproteolytic and that they produced some branched-chain volatile fatty acids on peptone media, the occurrence and the substrates for the production of such acids in paddy soil were investigated. The results obtained are as follows:

• 1) It was revealed that two branched-chain volatile fatty acids, iso-valeric and iso-butyric acids, were detected In fresh paddy field soil.

• 2) The paddy soil supplemented with peptone solution produced significant amounts of branched-chain volatile fatty acids-iso-butyric and iso-valeric acidsin addition to straight-chain volatile fatty acids-acetic, propionic, and butyric acids. When glucose was added instead of peptone, not branched-chain fatty and propionic acids were detected but only acetic and butyric acids. These facts indicate that the branched-chain fatty acids may originate from amino acids.

• 3) When a pair of amino acids, such as proline and leucine or proline-and valine, were supplemented in addition to peptone, the production of iso-valeric or iso-butyric acids was increased, respectively. Even In the case of glucose, the supplementation of a pair of amino acids supported the production of branched-chain fatty acids. These facts indicate that the Stickland reaction may take place in paddy field soil.

• 4) All the strains of strict anaerobes isolated from the soil and supplemented with peptone formed iso-valeric and iso-butyric acids in addition to two or three kinds of volatile fatty acids when they were cultured in medium containing peptone. All of them were classified as belonging to genus Clostridium. No strains of facultative anaerobes isolated from the same soil produced any branched-chain fatty acids, but produced only acetic acid.

• 5) In conclusion, these results reveal that the branched-chain fatty acid in paddy field soil is produced only by clostridia through the Stickland reaction.

     

Fate and impact on microorganisms of rice allelochemicals in paddy soil

Allelopathic rice can release allelochemicals from roots to inhibit neighboring plant species, but little is currently known about their fate and impact on microorganisms in paddy soil. This study showed that allelopathic rice PI312777 released much higher concentrations of allelochemical (5,7,4'-trihydroxy-3',5'-dimethoxyflavone) than non-allelopathic rice Liaojing-9 in field. When quantitative 5,7,4'-trihydroxy-3',5'-dimethoxyflavone was added into soil, flavone gave a short half-life of 18.27 +/- 2.32 h (r(2) = 0.94) and could easily be degraded into benzoic acid. Benzoic acid with a half-life of 29.99 +/- 2.19 h (r(2) = 0.96) was more resistant toward degradation in paddy soil. Furthermore, both the culturable microbial population and the entire microbial community structure of soil incubated with flavone and benzoic acid were evaluated using the soil dilution plate method and phospholipid fatty acid (PLFA) analysis, respectively. It appeared from the results that flavone could reduce microorganisms especially for fungi present in paddy soil, while benzoic acid could induce a higher response for soil microorganisms especially for bacteria. Consequently, flavone would be responsible for the dynamics of soil microorganisms during the early period, and any observed effect during the late period would be very likely due to its degradation product benzoic acid rather than flavone itself. These results suggested that allelopathic rice varieties could modify soil microorganisms to their advantage through the release of allelochemicals. The concentration and fate of discriminating 5,7,4'-trihydroxy-3',5'-dimethoxyflavone between allelopathic and non-alleloparhic varieties tested in rice soil would result in the different patterns of microbial population and community structure in paddy ecosystems.     

The effect of earthworms and liming on soil microbial communities

The effect of liming and earthworms on the composition and function of soil microbial communities was investigated in an upland soil from the UK in order to understand interactions between the biotic and abiotic components of soil systems. A factorial experiment was established using soils from the Sourhope Farm, near Kelso, with lime or no lime added, with or without earthworms added and a combined treatment of both lime and earthworm additions. The soils were incubated and destructively sampled after 180 days. Measurements of soil microbial biomass, dehydrogenase activity, phenotypic structure (by phospholipid fatty acid analysis (PLFA) and responses to four carbon substrates (d-glucose, l-arginine, α-ketoglutaric acid, α-cyclodextrin) were determined. Statistically significant results were limited to the litter layers, with no significant observations in either the H or Ah horizons. There were significant decreases in the soil microbial biomass and microbial activity in the litter layers caused by the addition of earthworms; liming reduced microbial biomass only. The addition of earthworms caused a significant difference in the PLFA principle component analysis (PCA) profile, as did liming. For the PLFA PCA profile, earthworm plus lime treatment was indistinguishable from the liming result. Addition of earthworms significantly suppressed the response to glucose; this effect was removed by liming. This indicates that liming may significantly alter the ecological interactions between earthworms and the microbial community.     

Evaluation of genetic diversity and plant growth promoting activities of nitrogen-fixing bacilli isolated from rice fields in South Brazil

Several strains of bacilli, mainly species of the genera Bacillus and Paenibacillus, displaying important plant growth promoting (PGP) characteristics were isolated from seven distinct rice production zones of the Rio Grande do Sul State, Brazil. Of 296 isolates, 155 were from rhizospheric soil and 141 from bulk soil. In order to evaluate the diversity among the isolates of each bacterial population the Shannon index was used on a 70% similarity basis. Diversity indices varying from 2.27 to 5.51 were obtained. Using principal coordinate analysis (PCA) to correlate bacterial diversity with soil parameters, it was found that soil pH was the characteristic most closely related to bacilli diversity. The bacilli isolated were also analyzed for some PGP activities. Of those 296 isolates, 94 and 148 produced between 0.1 and 30 μg of indole-3-acetic acid (IAA) ml⁻¹ in vitro after 72 and 144 h of incubation, respectively. Twenty-two isolates were able to solubilize phosphate and 32 isolates produced siderophores. Paenibacillus and Bacillus genera were the most prominent groups in the rhizosphere and soil populations analyzed. Paenibacillus borealis was the most frequent species in both locations. The isolate SVPR30, identified by 16S rRNA gene sequence analysis as a strain of Bacillus sp., was chosen for in vivo greenhouse experiments and proved to be very efficient in promoting a significant increase in the roots and shoot parts of rice plants. The identification and isolation of PGP bacilli from temperate and subtropical soils, which combine the ability to fix nitrogen with the production of substances capable to promote the plant growth, could significantly increase productivity of grain crops in Brazil.     

Multiple Metal Resistant Transferable Phenotypes in Bacteria as Indicators of Soil Contamination with Heavy Metals (6 pp)

Background, Aim and Scope   Environmental contamination by heavy metals affects microbial communities. The number of single and multiple heavy metal resistant bacteria may be an indictor of the level of contamination. This paper details the isolation and characterisation of metal resistant microorganisms isolated from rhizosphere/soil samples obtained from an abandoned zinc, lead and copper mine and a local unaffected site. This data was compared to the level of heavy metal in the soils to establish the effect of metals on the microbial community and to determine the relationship between pollutant levels and resistant strains. This paper outlines the diversity of transferable resistance determinants between both sites and details the levels of heavy metal resistant bacteria and those expressing transferable multiple heavy metal tolerance.Methods   The sample sites were located in Co. Galway, Ireland. The first sample site (site A) was a former lead, zinc and copper mine, which was closed in 1961 due to exhaustion of ore. The second site (site B) was located two and a half kilometres from the mining site and was not affected by the mining operations. Composite soil samples were characterised for general soil matrix composition, organic content, pH and general chemical parameters. The soil was also enumerated for the total viable heterotrophic counts and tested on Pseudomonas selective agar (PSA) for total Pseudomonas counts and Sucrose Asparagine (SA), which is semi-selective for fluorescent Pseudomonas.Results and Discussion   Samples from both site A and site B were analysed by atomic absorption spectrophotometry for the presence of heavy metals. In the case of copper, which has a Dutch list recommended minimum permissible level of 190 µg/Kg dry weight, the levels detected at site A were 1270 µg/Kg dry weight while site B was detected at 36 µg/Kg. The arsenic levels detected at site A were eight times the permissible level (416 µg/Kg) while only half the permissible level was found at site B (13 µg/Kg). Zinc concentrations were also high at site A (4460 µg/Kg) while at site B (553 µg/Kg) they were well below the Dutchlist guidelines (720 µg/Kg). A large number of heavy metal tolerant strains were isolated from both sites. 270 isolates (site A (170) and site B (100)) were screened against 8 metals to examine the extent of multiple resistance. 82% of the strains from site A were found to be resistant to 5 metals. A total of 18% showed resistance to all 8 metals and of those examined only 4% were resistant to only one metal. In contrast isolates from site B showed no multiple resistance to more than 5 metals, while 62% showed resistance to individual metals only. Site A had a higher level of multiple heavy metal resistance strains. Stains isolated from site A had 23 (14%) isolates resistant to zinc, copper, nickel, arsenic and cobalt and site B had no bacteria resistant to all five of these selected metals. The transferability of heavy metal resistance was investigated in the case of 60 multiple heavy metal resistant isolates taken from site A and 50 multiple resistance isolates from site B. Transfer was only detected in isolates from site A, 13% showed transfer and expression of copper, zinc and arsenic resistance determinants. In most cases the transconjugants only expressed resistance to copper, zinc and arsenic, which were the metals, used for selection. This co-transfer of all three determinants suggests a genetic link between these resistance determinants.Conclusion   Heavy metal resistant bacteria are present in both sites, however, the number and presence of multiple transferable resistance phenotypes are confined to the isolates from the heavy metal contaminated site. The presence of high levels of heavy metals selects these multiple resistance phenotypes. Within these communities there seems to be little diversity between the microorganisms, which provides a hugely preferable environment for gene transfer of such metal resistant determinants.Recommendation and Perspective   The experiments have shown a microbes ability to mobilize heavy metal determinants and a relationship between heavy metal resistance and metal contamination has been identified These multiple heavy metal resistant bacteria could eventually be used for detection and qualification of the level of heavy metal-polluted soil/water environments.     

Screening of rhizobacteria isolated from maize (Zea mays L.) in Rio Grande do Sul State (South Brazil) and analysis of their potential to improve plant growth

Plant growth-promoting rhizobacteria (PGPR) are considered to have a beneficial effect on host plants and may facilitate plant growth by different mechanisms. In this work, the influence of different soil types on the bacterial diversity and the stimulatory effects of selected PGPR on two cultivars of maize were investigated. A set of 292 strains was isolated from the roots and rhizosphere soil of maize cultivated in five different areas of the Rio Grande do Sul State in Brazil. 16S rDNA-PCR-RFLP and 16S rDNA partial sequencing were used for identification, and the Shannon–Weaver index was used to evaluate bacterial diversity. We evaluated the ability of each isolate to produce indole acetic acid (IAA), siderophores and solubilize phosphates. On the basis of multiple PGP traits, six isolates were selected to test their potential as plant growth-promoting rhizobacteria on maize plants. In both the roots and the rhizospheric soil of maize, the dominant bacterial genera identified were Klebsiella and Burkholderia. IAA producers were distributed widely among isolates, regardless of the sampling site. Approximately 42% of the isolates exhibited at least two attributes, and 24% showed all three PGP traits. Three strains, identified as Achromobacter, Burkholderia, and Arthrobacter, were effective as PGPR in both of the cultivars evaluated.     

Polyunsaturated fatty acids (PUFAs) content of the fungus Mortierella alpina isolated from soil

Twenty-five isolates of Mortierella spcies were prepared, which can be used for the production of polyunsaturated fatty acids (PUFAs)-rich oil for nutritional supplements. The fatty acid contents were determined after heterotrophic fermentation. The content of total fatty acids (TFAs) in the cell dry weight of all isolates including two commercially purchased Mortierella alpina strains ranged from 207.51 to 370.11 mg/g, whereas PUFAs were the dominant fatty acid type. The highest PUFA-containing strain, M. alpina SC9, was identified and confirmed as a new strain of M. alpina through comparison analysis of the sequences of internal transcribed spacers 1 and 2 (ITS1 and ITS2) and the 5.8S rDNA region. During a 7-day fermentation, the PUFAs content of M. alpina SC9 varied between 189.83 and 240.00 mg/g, with a remarkable correlation between the oleic acid (C18:1, OA) and arachidonic acid (C20:4n-6, AA) contents and between the linoleic acid (C18:2n-6, LA) and AA contents, suggesting the PUFA content in the fungus is tightly regulated. This study provides a framework of isolation, identification, and characterization of an important PUFA-producing species, M. alpina.     

Impact of sulfadiazine and chlorotetracycline on soil bacterial community structure and respiratory activity

Veterinary medicines enter agricultural soils by the use of animal excrements as fertilizers. To study their impact on soil bacterial communities, microcosms containing orthic luvisol soil were spiked with the antimicrobial agents sulfadiazine (SDZ) and chlorotetracycline (CTC) at three different concentrations (1, 10, 50 mg kg⁻¹ soil) and incubated for 48 days at 20 °C. The impact on the microbial respiratory activity was measured continuously in a respirometer (Sapromat). Changes in bacterial community structure were visualized by means of PCR-denaturing gradient gel electrophoresis (DGGE) of 16S rDNA derived from soil samples after 1, 7, 11 and 48 days. Additionally, growth inhibitory effects of SDZ and CTC on bacteria previously isolated from the same soil were tested in agar diffusion tests. In microcosms with soil and antibiotics only, no effects could be observed, either on respiratory activity or on bacterial population structure. Therefore, further incubations were conducted in the presence of an additional assimilable carbon source (5 g glucose kg⁻¹ soil). In the presence of glucose, SDZ affected soil respiration as well as the bacterial community structure: Additional bands appeared and some bands already visible at the beginning of incubations increased in intensity. A clear relationship between SDZ concentrations and changes in DGGE patterns became visible. During 48 days of incubation, changes in DGGE patterns were minimal in microcosms with 50 mg SDZ kg⁻¹soil indicating an inhibition of strains, which were capable of growing on glucose in the presence of lower SDZ concentrations. Only a few soil bacterial isolates (5 out of 47 strains tested) were weakly inhibited by SDZ in agar diffusion disk tests. Contrastingly, CTC inhibited growth of 12 soil bacterial isolates significantly in disk tests, but no effects on soil respiration and bacterial community structure could be observed. In the presence of the soil matrix the growth inhibitory potential of CTC decreased due to adsorption or complexation. This was confirmed in growth inhibition experiments with soil suspensions and time-dependent sampling.     

Biologically-induced hydrolysis of parathion in soil: Isolation of hydrolyzing bacteria

Fifty bacterial isolates from a parathion-treated soil (Gilat, Israel) were tested for their ability to hydrolyze the organophosphorus insecticide, parathion in peptone-yeast extract medium. After 5 days 33 isolates had hydrolyzed at least a portion of the added parathion. Eight of these isolates hydrolyzed 75% of the added parathion in 5 days and appeared to be Bacillus strains. Ten of these 33 isolates had hydrolyzed all of the parathion after 5 days and appeared to be Arthrohacter strains. One isolate from each group was tested further. During the logarithmic phase of growth, Bacillus sp., isolate 10, hydrolyzed less than 10% of the parathion added to peptone-yeast extract medium and was not active in parathion hydrolysis when inoculated into sterilized, parathion-treated soil. Arthrobacter sp., isolate 6, hydrolyzed parathion rapidly in peptone-yeast extract medium and in sterilized, parathion-treated soil. It used parathion or its hydrolysis product, p-nitrophenol, as sole carbon source. The parathion hydrolyzing enzyme appeared to be constitutive in isolate 6. Single applications of p-nitrophenol at concentrations greater than 1 mM inhibited growth but successive additions of smaller amounts permitted growth to continue.     

Nitrification and autotrophic nitrifying bacteria in acid tea soils

Pure cultures of ammonium-oxidizing, autotrophic, nitrifying bacteria were isolated from acid soils (pH range, 4.0–4.5) from tea estates in Sri Lanka (8 soils) and Bangladesh (4 soils). All the Bangladesh nitrifiers were Nitrosospira spp but the Sri Lanka isolates were identified as Nitrosolobus spp Nitrosospira spp and one species of Nitrosovibrio. Nitrite-oxidizing nitrifiers were detected in several of the soils but pure cultures were not isolated.Evidence was obtained that Nitrosospira caused nitrification in situ in an acid soil (pH 4.1). Indigenous nitrate was first eliminated from the soil by denitrification. The soil was then incubated aerobically and the nitrate formed was estimated as N₂O by gas chromatography after denitrification.     

Melanogenic actinomycetes (Streptomyces spp.) from Brazilian soils

Summary Melanogenic actinomycetes were isolated from cerrado soils. Starch agar with a neutral pH was the best medium for selecting pigment-producing colonies. A pigmentation screening test selected 52% of these as possible melanin producers. Tests on liquid (organic and inorganic) and solid (peptone and tyrosine) media, and enzymatic tests, confirmed about 90% as melanin producers, 68% as dihydroxyphenylalanine (DOPA)-melanin and 32% as possible other kinds of melanin producers. Melanin production occurred mostly with an organic N, or an inorganic N with an additional organic N source. An exception was observed with three strains, which were able to produce melanins with an entirely inorganic N source in the medium. Instability of melanin production was a common feature in many strains. Further characterization of melanins produced by actinomycetes compared with soil humic acids may clarify the possible role of melanogenic actinomycetes in soil organic matter formation.     

Effect of aqueous and lipophilic mullet (Mugil cephalus) Bottarga extracts on the growth and lipid profile of intestinal Caco-2 cells

The importance of n-3 polyunsaturated fatty acid (n-3 PUFA) intake has long been recognized in human nutrition. Although health benefits, n-3 PUFA are subject to rapid and/or extensive oxidation during processing and storage, resulting in potential alteration in nutritional composition and quality of food. Bottarga, a salted and semi-dried mullet ( Mugil cephalus ) ovary product, is proposed as an important source of n-3 PUFA, having high levels of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). In this work, we investigated the extent of lipid oxidation of grated bottarga samples during 7 months of storage at -20 °C and room temperature under light exposure. Cell viability, lipid composition, and lipid peroxidation were measured in intestinal differentiated Caco-2 cell monolayers after 6-48 h of incubation with lipid and hydrophilic extracts obtained from bottarga samples at different storage conditions. The storage of bottarga did not affect the n-3 PUFA level, but differences were observed in hydroperoxide levels in samples from different storage conditions. All tested bottarga extracts did not show a toxic effect on cell viability of differentiated Caco-2 cells. Epithelial cells incubated with bottarga oil had significant changes in fatty acid composition but not in cholesterol levels with an accumulation of EPA, DHA, and 22:5. Cell hydroperoxides were higher in treated cells, in relation to the oxidative status of bottarga oil. Moreover, the bottarga lipid extract showed an in vitro inhibitory effect on the growth of a colon cancer cell line (undifferentiated Caco-2 cells).     

Gram-positive bacterial flora on the root surface of wheat (Triticum aestivum L.) grown under different soil conditions

We identified 108 Gram-positive bacterial strains isolated from the root surface of wheat grown under different soil conditions. The strains were divided into four groups based on morphological and physiological characteristics, but most appeared to be coryneform. The taxonomic position of the various groups was verified by the guanine+cytosine DNA contents of the strains. In general, the ranges of these values agreed with those described for the respective taxonomic positions in the literature, with a few exceptions. With soil improvement the distribution of the various groups on the root surface changed, with the coryneform group becoming dominant. This group was further divided into five subgroups, according to cell wall components, cellulose-decomposition, and morphological characteristics, and were identified to genus level. The distribution of these subgroups on the root surface of wheat did not alter with soil improvement. The genus Arthrobacter, the dominant subgroup, predominated in every plot.