Year Of Incubation Research Articles

Comparison of soil phosphorus extractants as affected by fertilizer phosphorus sources, lime recommendation and time among four Arkansas soils

Phosphorus (P) sources were evaluated for their usefulness as P fertilizers in four acid Arkansas soils. Soils were amended with acidulated rock phosphorous, Florida rock phosphate, liquid ammonium phosphate, North Carolina rock phosphate, and triple superphosphorous, at 50 mg P kg‐1. After application, the soils were incubated for two years at 33 kPa soil moisture potential, with samples taken periodically. At each sampling Mehlich III (M‐III) and Bray‐Kurtz PI (BK1) extractants were used to measure available P levels. Acidulated rock phosphate was consistently the best P source for all soils, although P levels were significantly raised above the check by all P sources. Mehlich III extracted significantly more P than did BK1 for all soils. Phosphorus release varied among amendments during the incubation period. Acidulated rock phosphate, liquid ammonium phosphate, and triple superphosphate exhibited rapid initial release. The Florida and North Carolina rock phosphates did not achieve their maximum P release until one year of incubation. The correlation between extractants for P was M‐III = 1.2 + 1.06 (BK1) with r2 = 0.94.

Metal mobility and potential bioavailability in organic matter-rich soil-sludge mixtures: effect of soil type and contact time

The potential bioavailability of metals in two 2:1 (w:w) soil:sludge mixtures prepared by adding sludge to two different soils was studied. The samples were incubated for 1 year at ambient temperature and humidity close to field capacity. The distribution of Cr, Cu, Mn, Pb and Zn in sequential extractions was periodically determined using a five-step chemical fractionation procedure (MgCl 2, NaOAc/HOAc, (NH 4) 2C 2O 4+H 2C 2O 4, Na 4P 2O 7, HF+HNO 3). The potential availability (DTPA extraction) of Cu, Mn, and Zn, organic matter content, and the pH were also determined. The pH of the soils was 5.9 and 8.5 but the pH of both mixtures was similar to the sludge pH (6.3) due to the buffering capacity of sludge organic matter. A decrease in the amount of the organic matter with time and a modification in its composition was observed in the mixtures. The ratio between the quantities of humic and fulvic acids (IA/SA) varied from start to end of the experiment from 0.17 to 0.49 for Soil I-sludge mixture and from 0.22 to 0.42 for Soil II-sludge mixture. All metals, except Mn, became more easily extractable after 1 year of incubation. A positive significant correlation between the metal amounts extracted with DTPA and the sum of the first two sequential extracted fractions (F 1+F 2) was found. With the sludge rate used the behaviour of cations in the mixtures was fundamentally controlled by the sludge, but the type of soil still had an influence for Cr, Mn and Pb. In spite of the increase of the potential mobility of metals with time, the size of the available amounts declined at the end of the experiment.

Thymidine and leucine incorporation into bacteria from soils experimentally contaminated with heavy metals

Bacterial activities, estimated using the thymidine and leucine incorporation techniques, were measured in soil artificially contaminated with five different metals (Cd, Cu, Ni, Pb, Zn) at two different concentrations and in an uncontaminated control. The activity of the bacteria was followed during 14 months of incubation under laboratory conditions. After an initial decrease, which was correlated to the level of metals added, the bacterial activity recovered. Peak activities, much higher than those of the uncontaminated control, were detected after different incubation times for the different metal treatments. Cu contaminated soils showed peak activities after 1 month of incubation, while for Zn, Pb, and especially Ni, peak activities were not found until up to after 1 year of incubation. The leucine/thymidine incorporation ratio (mol/mol) did not differ over time, but significant differences were found between different metal treatments. Thus, while the uncontaminated control, and Cd, Zn and Ni treated soils had ratios around 20, the Cu contaminated soils had high ratios, around 50. The Pb contaminated soils, on the other hand, had low ratios (around 7). This was interpreted as an indication of different bacterial communities in the different treatments, where the Cu treated soils differed most from the other soils, while Cd, Zn or Ni application gave rise to similar communities. Other techniques to characterize soil microorganisms have earlier indicated the same effect of the metals on the bacterial community.

Relationship between denitrification and redox potential in two sediment-water systems

The process of denitrification was observed in vitro for two sediment-water systems. Overlying water was adjusted to a pH of 7.0, a temperature of 20�C and a nitrate concentration of 10.0 mg L-1 N-NO3-. Dissolved oxygen was maintained at a concentration above 6.0 mg L-1 O2 and incubations were carried out in the dark. These conditions insured that reduction of nitrate would result only from denitrification occurring in the sediments. The average rate of the denitrification, calculated from the amount of nitrate removed during about one year of incubation, was 50 mg N-NO3- m-2 day-1 for lake sediments and 171 mg N-NO3- m-2 day-1 for river sediments. Continuous measurements of the redox potential at various depths in the sediments allowed determination of the zone of denitrification and estimation of its migration into the sediment. Denitrification in the sediments occurred in close proximity to the oxygenated surface of sediments. Study of the kinetics of the denitrification process showed that the reactions for both sediment types were first-order. The exponential character of the reactions was presumably due to depletion of organic matter and could in part be a consequence of the diffusion-limited transport of nitrate into sediments. Diffusion of nitrate into the sediments was a major factor controlling denitrification rate.

Microbial responses to large concentrations of herbicides in soil

To determine the effects of herbicide spills on microbial activity, soil was treated with excessive amounts (10,000 mg/kg) of formulated alachlor alone or as a mixture with formulated atrazine, metolachlor, and trifluralin. Soil was also treated with the same combinations of herbicides at a rate of 10 mg/kg, which is typical of normal farming practices. Simulated spills initially inhibited bacteria, but after 7 d, bacterial numbers had recovered to levels similar to those in untreated controls. Fungal populations were drastically reduced after 1 d and became undetectable after 7 and 21 d of incubation in the mixed herbicide and alachlor-only treatments, respectively. Dehydrogenase activities were severely depressed after 1 d in both high-level herbicide treatments and did not recover after 125 d. Esterase activities were depressed in soils receiving the mixture of herbicides but were generally unaffected in the alachlor-only treatment. Normally applied concentrations did not have any pronounced effect on bacterial and fungal populations or on the activities of soil dehydrogenase and esterase. At 10 mg/kg, all the herbicides degraded by 80 to 90% within one year; there was no difference between degradation of alachlor applied alone or as a herbicide mixture. With high concentrations of herbicides, degradation was negligible after one year of incubation.

The Effect of Simulated Acid Rain on Pine Needle and Birch Leaf Litter Decomposition

(1) The effects of simulated acid rain at pH3 and pH4 (each made from H2SO4 and HNO3) on the decomposition of Scots pine needle litter and mountain birch leaf litter were studied in a field experiment in northern Finland. The effects of irrigationper se were tested by having 'dry' control plots receiving only ambient rain, and irrigated controls receiving similar amounts of water to the acid treatments. (2) Treatments and the vegetation type in the plots had significant main effects on mass loss of pine needle litter. Differences between dry and irrigated controls, and the lower mass loss in plots with drier vegetation types showed the importance of moisture conditions in regulating pine litter decomposition. (3) The effects of acid rain treatments on pine litter decomposition were more subtle than those of irrigation per se. In the moistest vegetation type there was no decrease in mass loss due to acid treatments, but mass loss rates at late stages of decomposition were slightly reduced by acid treatments on the drier plots. (4) Birch litter decomposition was significantly retarded in acid treated plots compared to the irrigated controls during the second year of incubation in one experiment. In the other experiment there was a similar trend, although not significant. At the end of the experiments the difference in mass loss between acid treatments and irrigated controls was 2-5% units. The two acid treatments did not differ significantly from each other.

Effect of the addition of municipal solid-waste compost on microbial biomass and enzyme activities in soil

Changes in the content of C, N, P, and S in the soil biomass and in phosphatase, urease, protease, deaminase, and arylsulphatase activity, induced by amendment with municipal solid-waste compost, were determined in a clay loam soil during 1 year of incubation at 25° and 35°C.

Bioremediation Potential of Terrestrial Fuel Spills

A bioremediation treatment that consisted of liming, fertilization, and tilling was evaluated on the laboratory scale for its effectiveness in cleaning up a sand, a loam, and a clay loam contaminated at 50 to 135 mg g of soil by gasoline, jet fuel, heating oil, diesel oil, or bunker C. Experimental variables included incubation temperatures of 17, 27, and 37 degrees C; no treatment; bioremediation treatment; and poisoned evaporation controls. Hydrocarbon residues were determined by quantitative gas chromatography or, in the case of bunker C, by residual weight determination. Four-point depletion curves were obtained for the described experimental variables. In all cases, the disappearance of hydrocarbons was maximal at 27 degrees C and in response to bioremediation treatment. Poisoned evaporation controls underestimated the true biodegradation contribution, but nevertheless, they showed that biodegradation makes only a modest contribution to gasoline disappearance from soil. Bunker C was found to be structurally recalcitrant, with close to 80% persisting after 1 year of incubation. The three medium distillates, jet fuel, heating oil, and diesel oil, increased in persistence in the listed order but responded well to bioremediation treatment under all test conditions. With bioremediation treatment, it should be possible to reduce hydrocarbons to insignificant levels in contaminated soils within one growing season.

Size and persistence of the microbial biomass formed during the humification of glucose, hemicellulose, cellulose, and straw in soils containing different amounts of clay

14C-labelled substrates were incubated at 20°C in 4 soils with clay contents ranging from 6 to 34%. Glucose was most readily decomposed, followed in order by hemicellulose, cellulose, maize straw, and barley straw. After the first 10 days of incubation, about 60% of the glucose-C had left the soils as CO2, compared with only 23% of the barley-C. The humified matter that remained in the soils after 3 months decayed at almost the same rate whether the origin of the matter was glucose, hemicellulose, cellulose or straw; this rate was, on the whole, independent of the caly content of the soils. Half-life values for the labelled C in the soils during the second and third year of incubation ranged from 5 to 7 years. The amino acid-C percentages of the humified matter tended in all four soils to be largest in matter originating from glucose and least in that originating from straw. The amino acid-C percentages increased with the clay content of the soils. The biomass was determined by fumigation with CHCl3 according to Jenkinson. After 3 months an average of 17% of the residual labelled C was in biomass; the values ranged from 37% when the labelled C was added as glucose to 2–9% when added as barley straw. The half-life of labelled C in biomass during the second year of incubation ranged from 2 to 3 years. Native C in biomass ranged from 0.5 to 1.4% of the total C in native soil organic matter, the highest values occurring in the clay-rich soils. The half-life of native soil C, estimated from CO2 evolution during 3-month periods, ranged from 13 to 29 years.

Isolation of Bacillus licheniformis var. endoparasiticus from the blood of rheumatoid arthritis patients and normal subjects.

Sixty-five samples of blood from patients with rheumatoid arthritis and 94 from control subjects were examined by multiple culture and monthly subculture for periods of up to 33 months to detect the revision stages of Bacillus licheniformis var. endoparasiticus (BLE), which exists as L-forms in blood. Isolation of BLE were obtained more often from the blood of control subjects than from rheumatoid patients during the first 6 months of incubation, when there was clustering of positive cultures within samples. Thereafter, the isolation rate was similar for the two groups and positive cultures were distributed randomly between samples. Isolation of diphtheroid intermediate reversion stages (phase A and B) occurred mainly during the first year of incubation, but isolations of the fully reverted sporing bacillus (phase C) increased in frequency with incubation time, particularly from cultures with a high rate of desiccation during prolonged incubation. The proportion of different phases of BLE amongst the isolates and the distribution of the phases with incubation time were similar for the rheumatoid-arthritis patients and the normal subjects.

Carbon-nitrogen relationships during the humification of cellulose in soils containing different amounts of clay

14C-labelled cellulose and 15N-labelled (NH 4) 2SO 4 were added to four soils with clay contents of 4, 11, 18 and 34%, respectively. Labelled cellulose was added to each soil in amounts corresponding to 1, 2 and 4 mg C g −1 soil, respectively, and labelled NH 4 + at the rate of 1 mg N per 25 mg labelled C. After the first month of incubation at temperatures of 10, 20 and 30°C, respectively, from 38 to 65% of the labelled C added in cellulose had disappeared from the soils as CO 2, and from 60 to nearly 100% of the labelled N added as NH 4 + were incorporated into organic forms. The ratio of labelled C remaining in the soils to labelled N in organic forms was close to 25 after 10 days of incubation, decreasing to about 15 after 1 month and about 10 after 4 yr. The retention of total labelled C was largest in the soil with the highest content of clay where after 4 yr it was 25% of that added, compared to 12 in the soil with the lowest content of clay. The incorporation of labelled N in organic forms and its retention in these forms was not directly related to the content of clay in the soils, presumably because the two soils with the high content of clay had a relatively high content of available unlabelled soil-N which was used for synthesis of metabolic material. The proportionate retention of labelled C for a given soil was largely independent of the size of the amendments, whereas the proportionate amount of labelled N incorporated into organic forms increased in the clay-rich soils with increasing size of amendments. Presumably this is because the dilution with unlabelled soil-N was less with the large amendments. From 50 to 70% of the total labelled C remaining in the soils after the first month of incubation was acid hydrolyzable, as compared to 80–100% of the total remaining labelled organic N. This relationship held throughout the incubation and was independent of the size of the amendment and of the temperature of incubation. During the second, third and fourth year of incubation the half-life of labelled amino acid-N in the soils was longer than the half-life of labelled amino acid-C, presumably due to immobilization reactions. Some of the labelled organic N when mineralized was re-incorporated into organic compounds containing increasing proportions of native soil-C. whereas labelled C when mineralized as CO 2 disappeared from the soils. In general, native C and native organic N were less acid hydrolyzable and were accounted for less in amino acid form than labelled C and N. The amount of labelled amino acid-C, formed during decomposition of the labelled cellulose, and retained in the soil, was proportional to the clay content. This amount was about three times as large in the soil with the highest content of clay as in the soil with the lowest content. This difference between the soils was established during the first 10 days of incubation when biological activity was most intense, and it held throughout the 4 yr of incubation; proportionally it was independent of the amount of cellulose added and the temperature. In contrast, the labelled amino acid-N content was not directly related to the amount of clay in the soil, presumably because more unlabelled soil-N was available for synthesis of metabolic material in the two clay-rich soils than in those soils with less clay. The wider ratio between labelled amino acid-C and labelled amino acid-N in the two clay-rich soils as compared with those obtained with the soils with less clay indicates this. The effect of clay in increasing the content of organic matter in soil is possibly caused by newly synthesized matter, extracellular metabolites, as well as cellular material, forming biostable complexes and aggregates with clay. The higher the concentration of clay the more readily the interactions take place. The presence of clay may also increase the efficiency of using substrate for synthesis.

The influence of clay on the rate of decay of amino acid metabolites synthesized in soils during decomposition of cellulose

14C-labelled cellulose was added to seven different soils containing silt + clay (particles < 0.02 mm) in amounts which varied from 8 to 75 per cent. The cellulose was allowed to decompose, and the amounts of labelled C transformed into metabolites hydrolyzable into amino acids were determined. The amounts of labelled amino acid C in the soils were proportional to their content of silt + clay. After 30 days of incubation labelled amino acid C remaining in the soil with the lowest content of silt + clay constituted 6 per cent of the carbon added in cellulose, as compared with 18 per cent in the soil with the highest content of silt + clay. These values had decreased to 5 and 13 per cent respectively after 2 years of incubation. The order between the soils in the content of labelled amino acid C established during the first month of incubation, was thus roughly maintained throughout the period of incubation. The biological half-life of the labelled C in amino acids varied in the seven soils during the last year of incubation from 3 to 8 years. The variation was, however, not related to the amount of silt + clay. n the soils had been incubated with the labelled material for 2 years, samples of the soils were exposed to “stress” treatments: air drying-rewetting; increased biological activity caused by addition of glucose, and exposure to chloroform vapour. The treatments resulted in an evolution of labelled C in CO, which was 5–10 times larger than the evolution from untreated samples. The increase in the CO 2 evolution caused by the treatments in the different soils was, however, not related to the amount of silt + clay, and a high content of this material did not protect organic material against the effect of the treatments. is concluded that the silt + clay fraction ensures stabilization of amino acid metabolites produced during the period of intense biological activity that follows the addition of decomposable, energy rich material to the soil. The amount of amino acid metabolites stabilized increased with increasing concentration of silt + clay, but the rate of decay of the amino acid material during later stages was largely independent of the concentration of silt + clay.

Modification of the exogenous carbon and nitrogen requirements for chlamydospore germination of Fusarium solani by contact with soil.

After exogenous carbon-independent macroconidium germination by Fusarium solani at 1 × 104 conidia/ml in axenic culture, chlamydospores were formed terminally on germ tubes. Low chlamydospore germination was supported under axenic conditions by 0.004 ng C/spore (calculated value), supplied as ethanol, or by 0.04 ng C/spore, supplied as glucose or several other sugars. Chlamydospores in direct contact with a nonsterile loamy fine sand soil (6.2 μg NH4+-N and 7.1 μg NO3−-N/g soil) had a greater exogenous carbon requirement and did not germinate until 2.0 ng glucose-C/spore was supplied. Supplemental exogenous nitrogen, added as NH4Cl, had little or no influence on percentage of germination at the levels of exogenous carbon supporting low germination, both in soil and in axenic culture. However, increasing the amounts of glucose plus NH4Cl supplied per spore increased percentage of chlamydospore germination more than increasing the amount of glucose alone for both soil and axenic systems. In axenic culture, complete germination was observed at 2.0 ng C plus 0.13 ng N/spore. Greater than 90% germination occurred for chlamydospores in contact with soil at the highest glucose plus NH4Cl level examined (200 ng C plus 13 ng N/spore), but not in glucose alone. Sterile acid-washed sand did not have a similar adverse effect on chlamydospore germination. Chlamydospores were nearly 100% germinable after 1 year of incubation in an inorganic salts medium under axenic conditions.

無細胞半合成半流動寒天培地を用いる人癩菌の培養

One-tenth ml each of bacterial suspensions derived from nodules of 4 different L-type patients was inoculated into 0.1% semiliquid agar media of various semi-synthetic base compositions and incubated at 37°C. The results revealed that rather large colony-like masses of bacteria had appeared, in most cases, after about 20 weeks and persisted even after about one year of incubation at 37°C. These masses varied in size and shape. Generally, they were composed of smaller, round bacterial masses of 10 to 20μ in diameter, confluent with one another. Each mass seemed to have boundary around it. It consisted of strongly acid-fast rods, stainable with malachite green. It was suggested that the bacterial masses might be colonial growths.Bacterial suspensions prepared from nearly one-year-old cultures by centrifugation induced essentially the same intradermal reaction in leprous patients as Dharmendra's antigen. Therefore, it was revealed clearly that the bacterial masses appearing in culture tubes were composed of M. leprae.

The effect of temperature on toxin formation and toxin stability of Clostridium botulinum type E in different environments.

Clostridium botulinum type E was found to produce toxin at temperatures between 3° and 30 °C in Robertson"s chopped meat medium and in fish dialyzate. Toxin was produced more rapidly in the meat medium. No toxin was found after 1 year of incubation at 1 °C. At 3 °C a slight toxin production was noticed after 120 days, when the inoculum consisted of a mixture of vegetative cells and spores.The lowest concentration of NaCl necessary to inhibit toxin production was found to vary with the incubation temperature and duration of storage. The inhibiting effect of NaCl was more pronounced at lower temperature.Type E spores mixed in fish meat medium heated for 110 min at 80 °C or for 10 min at 90 °C produced toxin during prolonged storage at 20 °C but not after they were heated at 90 °C for 20 min.Toxin (type E) proved more thermostable in meat broth and fish dialyzate than in phosphate buffer solution or in buffer supplemented with gelatine. In meat broth the toxin was inactivated after 5 min at 65 °C.

Effect of Sodium Chloride and pH on the Outgrowth of Spores of Type E Clostridium botulinum at Optimal and Suboptimal Temperatures1

The sodium chloride inhibition of spore outgrowth of four strains of type E Clostridium bolulinum was determined in a Trypticase-peptone-glucose (TPG) medium. At 16, 21, and 30 C, spores of three strains required 5.0% and one strain 4.5% salt for complete inhibition during 1 year of incubation. At 8 and 10 C, spores of the four strains required 4.5% salt for definite inhibition. Salt concentrations slightly lower than those providing inhibition tended to extend spore outgrowth time at low temperatures. The minimal pH permitting outgrowth of type E spore inocula was affected by the concentration of reducing compound present in the system. When either 0.02% sodium thioglycolate or 0.05% L-cysteine hydrochloride was used, outgrowth at 30 and 8 C occurred at much lower pH levels than when 0.2% thioglycolate was added. At 30 C, spores of one strain showed outgrowth in TPG medium as low as pH 5.21 with an inoculum of 2 million spores per replicate tube. At a 10-fold higher inoculum, the same strain showed outgrowth at pH 5.03 in one of five replicate tubes. At 8 C, spore outgrowth of the four strains occurred at pH 5.9, but not at pH 5.7, in TPG medium containing L-cysteine hydrochloride.

Effect of sodium chloride and pH on the outgrowth of spores of type E Clostridium botulinum at optimal and suboptimal temperatures.

The sodium chloride inhibition of spore outgrowth of four strains of type E Clostridium bolulinum was determined in a Trypticase-peptone-glucose (TPG) medium. At 16, 21, and 30 C, spores of three strains required 5.0% and one strain 4.5% salt for complete inhibition during 1 year of incubation. At 8 and 10 C, spores of the four strains required 4.5% salt for definite inhibition. Salt concentrations slightly lower than those providing inhibition tended to extend spore outgrowth time at low temperatures. The minimal pH permitting outgrowth of type E spore inocula was affected by the concentration of reducing compound present in the system. When either 0.02% sodium thioglycolate or 0.05% L-cysteine hydrochloride was used, outgrowth at 30 and 8 C occurred at much lower pH levels than when 0.2% thioglycolate was added. At 30 C, spores of one strain showed outgrowth in TPG medium as low as pH 5.21 with an inoculum of 2 million spores per replicate tube. At a 10-fold higher inoculum, the same strain showed outgrowth at pH 5.03 in one of five replicate tubes. At 8 C, spore outgrowth of the four strains occurred at pH 5.9, but not at pH 5.7, in TPG medium containing L-cysteine hydrochloride.