Research note : Unsuitability of the MRS medium for the screening of hydrogen peroxide-producing lactic acid bacteria

The effect of MRS broth on the stability of hydrogen peroxide (H₂O₂) has been studied. Known concentrations (1–100 μg ml⁻¹) of H₂O₂ were prepared in distilled water, phosphate buffer (pH 7·0) and MRS broth (pH 6·2 and 3·9). H₂O₂ was very stable in aqueous and buffer solutions but it was rapidly degraded in MRS broth (pH 3·9). The presence of H₂O₂ in MRS broth (pH 6·2) could not be detected.     

Glucose-induced acid tolerance appearing at neutral pH in log-phase Escherichia coli and its reversal by cyclic AMP

Escherichia coli shifted from broth at external pH (pH₀) 7·0 to pH₀ 7·0 broth plus glucose rapidly induced marked acid tolerance which also appeared, albeit to a lesser extent, plus maltose, sucrose or lactose. Tolerance appeared without the medium pH becoming acidic. Tolerance was most substantial when glucose was added at pH₀ 7·0 but was also appreciable at pH₀ 7·5, 8·0 and 8·5. Induction of tolerance by glucose was markedly reduced by cyclic AMP and essentially abolished plus NaCl or sucrose ; the induction process was also reduced but not fully inhibited by chloramphenicol, tetracycline and nalidixic acid. Glucose-induced organisms showed less acid damage to DNA and β-galactosidase and it is likely that this is because glucose induces a new pH homeostatic mechanism which keeps internal pH close to neutrality at acidic pH₀. In conclusion, it is clear that glucose induces a novel acid tolerance response in log-phase E. coli at pH₀ 7·0 ; it is now known that induction of this response involves the functioning of extracellular induction components including an extracellular induction protein.     

Enhanced degradation of petrol (Slovene diesel) in an aqueous system by immobilized Pseudomonas fluorescens

The rate of degradation of n -alkanes C₁₂-C₁₈, in petrol (Slovene diesel) in an aqueous system, by free and immobilized Pseudomonas fluorescens in shaking flasks was investigated. Cells were immobilized to a biosupport, Biofix, and a biosorbant, Drizit. Analysis of cellular growth of the free and immobilized bacteria over 8 d of incubation with diesel as the sole carbon source, showed a reduction in the lag phase in the immobilized cultures in comparison to the free system. The free system degraded 52·3% of C₁₂ and 11·6% of C₁₃, but C₁₄-C₁₈ were not degraded. In comparison to the free system and diesel which had not been exposed to experimental conditions (unexposed), the immobilized systems degraded significantly more of C₁₃-C₁₈. Biofix-immobilized cells degraded 14·8% of C₁₂ and an average of 53·5% of C₁₃-C₁₈. Drizit-immobilized cells degraded 24·5% of C₁₂, 52·4% of C₁₃ and an average of 91·2% of C₁₄-C₁₈. This study shows the successful use of immobilized bacteria technology to enhance the degradation of diesel in an aqueous system.     

Anion exchange in the giant erythrocytes of African lungfish

Carbon dioxide transport in African lungfish Protopterus aethiopicus blood conformed to the typical vertebrate scheme, implying a crucial and rate-limiting role of erythrocyte Cl^–/HCO₃^– exchange. The rate coefficient for unidirectional Cl^– efflux via the anion exchanger ( k , s⁻¹) increased with temperature in African lungfish, but values were well below those reported in other species. The erythrocytes of African lungfish were, however, very large (mean cellular volume = 6940 µm³), and the ratio of cell water volume to membrane surface area was high ( V _w A _m⁻¹ = 1·89). Hence, the apparent Cl^– permeability ( P _Cl =  kV _w A _m⁻¹, µm s⁻¹) was close to that in other vertebrates. The plot of ln P _Cl against the inverse absolute temperature was left-shifted in the tropical African lungfish compared to the temperate rainbow trout Oncorhynchus mykiss , which supports the idea that P _Cl is similar among animals when compared at their preferred temperatures. Also, Q ₁₀ for anion exchange calculated from P _Cl values in African lungfish was 2·0, supporting the idea that the temperature sensitivity of erythrocyte anion exchange matches the temperature sensitivity of CO₂ production and transport in ectothermic vertebrates.     

Toxicity of Sodium and Chloride Ions to Rhizobium spp. in Broth and Peat Culture

The growth of a strain of Rhizobium trifolii and of R. meliloti was studied in broth and peat cultures to determine the relative toxicity of Na⁺ and Cl^-. The following salts were added in a range of concentrations: Na₂HPO₄ as a source of Na⁺, CaCl₂.2H₂O as a source of Cl^-, and NaCl. Disodium hydrogen orthophosphate affected the growth rate of both strains in broth culture but not in peat culture. Unexpectedly, calcium chloride was more toxic than NaCl in broth and peat culture. The toxicity of NaCl can be ascribed to the Cl^-. Rhizobium meliloti strains grew on 3·5% NaCl after adaptation during a long period. Rhizobia for soya bean and cowpea grew at 0·5% NaCl and those for clover and pea, at 1·0% NaCl.     

An effective method for isolating alginate lyase-producing Bacillus sp. ATB-1015 strain and purification and characterization of the lyase

A new alginate lyase-producing micro-organism, designated as Bacillus sp. strain ATB-1015, was effectively isolated from soil samples pretreated for 3 months with a substrate of the enzyme, sodium alginate. Alginate lyase activity was assayed by the degrading activity of biofilm on Teflon sheet discs, which was formed by a mucoid strain of Pseudomonas aeruginosa PAM3 selected from clinical isolates. The extracellular alginate lyase was precipitated with ammonium sulphate from the culture broth, and purified by gel filtration and anion exchange chromatography. The molecular weight of the lyase was estimated to be 41 kDa by SDS polyacrylamide gel electrophoresis and Sephacryl S-200 HR column chromatography. The optimum pH and temperature for the enzyme activity were around 7·5 and 37 °C, respectively, and the Km value was 0·17% with the substrate, sodium alginate. The lyase activity was completely inhibited by treatment with 1 mmol l⁻¹ of EDTA and the decreased activity was almost completely recovered by the addition of 2 mmol l⁻¹ of CaCl₂. The activity was not affected by treatment with the protein denaturants, 0·01 mol l⁻¹ of SDS or 1 mmol l⁻¹ of urea. The lyase had substrate specificity for both the poly-guluronate and poly-mannuronate units in the alginate molecule.     

Effects of hypercapnia on the buffer capacity and haematological values in Salmo salar (L.)

The effects of high environmental carbon dioxide tension (PCO₂) on buffering status, pH, haematocrit, red blood cell (RBC) count, mean corpuscle volume (MCV), and chloride (Cl^–) distribution in the blood of young salmon were investigated after 30 min and 24 h. An increase in blood buffering capacity was observed after 24 h. As compared to the preacclimation situation, haematocrit, RBC, MCV and Cl^– content of the erythrocytes increased after 30 min but decreased if hypercapnia was sustained for 24 h. However the plasma bicarbonate (HCO₃) content increased over the whole experimental period. Plasma Cl^– content was decreased after 24 h, but the ratio of Cl^– between erythrocytes and plasma increased at the onset of hypercapnia, then remained unchanged during the rest of the investigation period.     

Soil microbial carbon uptake characteristics in relation to soil management

Abstract The kinetics of glucose uptake by soil microbial communities in 16 different soild (7 under monocultures and 9 under crop rotations) differing in microbial biomass content, % C_org, pH and clay content were investigated at 22°C. The V _max value of microbial bimasses under monoculture, was o.27 μg C_gluc · μg⁻¹ C_mic · h⁻¹ (range 0.18–0.44), twice as high as the mean value of V _max of microbial biomasses under rotations (0.13 μg C_gluc, range 0.07–0.19). Mean values of K _m were 714 μg C_gluc and 290 μg C_gluc · g⁻¹ soil, respectively.
These differences were highly significant ( P =0.001, based on SE) and could not be relate to particle size distribution of the soils, pH or C_org. A Michaelis-Menten type uptake response was apparent over the total range of glucose concentrations used (45.4–1453.3 μg C_gluc · g⁻¹ soil) for microbial biomasses under rotation while the majority of microbial biomasses under monocultures showed a similar response only at low glucose concentrations. A different uptake mechanism appeared to be involved at higher glucose concentrations (similar to diffusion) in monoculture soils.     

Application of bioluminescence-based microbial biosensors to the ecotoxicity assessment of organotins

The toxicity of two common organotin pollutants and their initial breakdown products (tributyltin, dibutyltin, triphenyltin and diphenyltin) were assessed using two different bioluminescent microbial biosensors: Microtox and lux -modified Pseudomonas fluorescens pUCD 607. The organotins were made up as standards, and tested both in double-deionized water and in extracted soil solution, the latter representing a realistic matrix for terrestrial contamination. Microtox was especially sensitive to the organotins, with 50% effective concentration (EC₅₀) values (15 min) for tributyltin as low as 21·9 μg l⁻¹ in pure water, and 0·118 μg l⁻¹ in soil extract. The sensitivity of Microtox was increased by an order of magnitude in soil extract. The Ps. fluorescens was less sensitive, with EC₅₀ values (30 min) of around 800 μg l⁻¹ in pure water. The toxicity to Ps. fluorescens was decreased by around an order of magnitude in soil extract. The two biosensors showed different response patterns, with Microtox being more sensitive to the triorganotins and Ps. fluorescens being more sensitive to the diorganotins.     

The Effect of Freezing on the Radiation Sensitivity of Vegetative Bacteria

S ummary : Five strains of bacteria were irradiated, suspended in heart infusion broth or in phosphate buffer, in aerated or anoxic conditions, at temperatures of 10–13° or -79°. Survivors under the different conditions were enumerated by plate counts on heart infusion agar.
Exponential survivor-dose curves were obtained with a Pseudomonas strain and with Escherichia coli B/r when irradiated at room temperature with aeration, whereas an Alcaligenes strain and 2 strains of Streptococcus faecium gave sigmoid curves. The decreased radiosensitivity in the frozen state was measured by comparing the D₁₀ values for exponential curves, or for the exponential portion of sigmoid curves, with that observed for irradiation at room temperature with aeration. This 'D₁₀ ratio' svaried between 2°5 and 8·5. For the Alcaligenes strain it was about 4, whether the frozen irradiation took place in the presence or absence of oxygen. With the Pseudomonas irradiated in the frozen state in the absence of oxygen the 'D₁₀ ratio' was usually about 1·5 times higher than when oxygen was present. The highest ratio (8·5) was obtained for anoxic irradiation of the Pseudomonas strain.
In general, the shapes of survival curves for frozen irradiation differed from those obtained at room temperature. The sigmoid curves for the Alcaligenes strain irradiated under aerobic conditions when frozen showed a marked decrease in extrapolation numbers. E. coli B/r when frozen in heart infusion broth gave a double exponential curve with a shallow slope initially, followed by a steeper slope. The most radiation resistant strain, Strep. faecium R53, gave sigmoid curves with a D₁₀ value of 300 Krads when frozen, and was then of similar resistance to Clostridium botulinum spores.     

Plasmid transfer between strains of Pseudomonas putida, and their survival, within a pilot scale percolating-filter sewage treatment system

Abstract: The effect of Pseudomonas aeruginosa UG2 biosurfactants or UG2 inocula on phenanthrene mineralization in uninoculated nonsterile soil slurries and slurries inoculated with the phenanthrene-mineralizing Pseudomonas sp. UG14r was investigated. In sandy loam and silt loam slurries amended with phenanthrene, inoculation with UG14r alone or in co-culture with UG2Lr reduced the lag period before onset of phenanthrene mineralization by 1 week. The total amount mineralized after 5 weeks was lower or not significantly different from the uninoculated control slurries. Inoculation with P. aeruginosa UG2Lr alone did not improve phenanthrene mineralization. In creosote-contaminated soil slurries, no lag period in phenanthrene mineralization was observed in any treatment. After 4 weeks, the greatest extent of mineralization was observed in creosote-contaminated soil slurries inoculated with the UG14r-UG2Lr co-culture and UG14r alone. In sandy loam and silt loam soil slurries inoculated with Pseudomonas sp. UG14r, addition of UG2 rhamnolipid biosurfactants (100 to 400 mg rhamnose equivalents (RE) · l⁻¹ slurry) inhibited phenanthrene mineralization by 10 to 15%. Mineralization was also inhibited in uninoculated sandy loam slurries. In creosote-contaminated soil slurries inoculated with Pseudomonas sp. UG14r, biosurfactants at 250 mg RE · l⁻¹ slurry enhanced mineralization whereas 400 mg RE · l⁻¹ had no effect, compared to unamended slurries. In uninoculated creosote-contaminated soil slurries, UG2 biosurfactants at 250 and 400 mg RE · l⁻¹ slurry enhanced mineralization, compared to unamended slurries.     

Use of impedance measurements to estimate numbers of antibiotic resistant Salmonella strains

Impedance detection times were compared with traditional plating methods for enumerating antibiotic-resistant strains of Salmonella stanley, Salm. thompson and Salm. infantis grown in laboratory medium and pork slurry. The correleation of log₁₀ counts of salmonellas with detection times was highly significant ( r = -0·96 for broth and r = -0·94 for slurries. The confidence limits (± og₁₀ 1·0 for broth and ± log₁₀ 1·65 for slurry) indicated that detection times could reliably be used as a rapid means of enumerating salmonellas when large numbers of counts of known strains are required for growth studies. Use of antibiotic-resistant strains also permitted their selective detection by impedance from the natural spoilage flora of pork slurry when the same antibiotics were incorporated in the detection medium.     

Physiological and anatomical disturbances induced by chloride salts in sensitive and tolerant citrus: beneficial and detrimental effects of cations

In citrus, the relative contributions of chloride and cations to growth disturbances induced by salinity are a matter of controversy. Chloride salts (15 mol m^–3 CaCl₂, 30 mol m^–3 CaCl and 30 mol m^–3 KCl) reduced growth and gas exchange parameters, increased leaf damage and abscission and produced anatomical disarrangements and mineral imbalances in seedlings of sensitive Carrizo citrange ( Citrus sinensis x Poncirus trifoliata ) and tolerant Cleopatra mandarin ( Citrus reshni ). In both cultivars, Ca²⁺ was more beneficial, and K⁺ more detrimental, for growth than sodium. Photosynthesis and growth disturbances were highly correlated ( P ≤ 0·001) with leaf Cl^– build-up. In the sensitive genotype, Cl^– was also significantly correlated with several leaf anatomical disarrangements, such as increase in succulence. In comparison with sodium, both calcium and potassium increased leaf Cl^– content (up to 25 and 69%, respectively). Protective calcium effects were not linked to improvement of photosynthesis, reduction of leaf anatomical disarrangements, or prevention of Cl^– and Na⁺ increases. It is proposed that the ameliorative effects of calcium on citrus grown under salinity are mostly related to reduction of leaf abscission. Collectively, the data suggest a cause–effect relationship between Cl^– build-up and reduced growth, whereas chloride correlations with declines in photosynthesis or increases in succulence appear to be indirect.     

Effect of pH, temperature and culture medium composition on the production of an extracellular cysteine proteinase by Micrococcus sp. INIA 528

Growth and proteinase production by Micrococcus sp. INIA 528 in a batch-operated laboratory fermentor were investigated, with trypticase soy broth as the basal medium for studies on optimum temperature, pH and medium composition. Maximum growth was recorded at 34°C and pH 715, whereas optimum temperature and pH for proteinase production were 31°C and pH 6.25. Maximum rate of enzyme production occurred during the late log and early stationary phases of growth. Addition of 5.0 g 1^-1 yeast extract, 1.0 g 1^-1 glucose, 1.0 g 1^-1 MgSO₄ or 1.0 g 1^-1 K₂HPO₄ to basal medium resulted in a lower enzyme yield, but supplementation of basal medium with 2.5 g 1^-1 (NH₄)₂SO₄ increased enzyme production by 45%. A high initial biomass added to fresh broth supplemented with 2.5 g 1^-1 (NH₄)₂SO₄ only increased enzyme activity by 19%, compared to the maximum enzyme activity achieved with the standard inoculum.     

KINETICS OF GROWTH OF HAEMOPOIETIC COLONY CELLS IN AGAR

Cell kinetic parameters of mouse granulocytic and mononuclear cells growing in colonies in agar cultures have been measured. Analysis of flash and continuous labelling studies with ³H-thymidine together with determinations of colony size, growth fraction and mitotic indices, gave the following values for the phases of the cell cycle: G₁= 6·3 1·6 hr, S = 5·8 ± 1·4 hr, G₂= 1·7 ± 0·1 hr and M = 0·7 ± 0·1 hr (42 ± 8 min). No difference in the cell cycle parameters of granulocytic and mononuclear cells were found in this study.
Colonies of different size from cultures of the same age group had similar labelling indices, indicating that the size of a colony is not a function of the rate of proliferation of cells in the colony. Rather, variation in colony size is probably representative of an initial delay in the onset of colony development.     

The linamarase of Mucor circinelloides LU M40 and its detoxifying activity on cassava

Mucor circinelloides LU M40 produced 12·2 mU ml⁻¹ of linamarase activity when grown in a 3 l fermenter in the following optimized medium (g l⁻¹ deionized water): pectin, 10·0; (NH₄)₂SO₄,
1·0; KH₂PO₄, 2·0; Na₂HPO₄, 0·7; MgSO₄.7H₂O, 0·5; yeast extract, 1·0; Tween-80,
1·0, added after 48 h of fermentation. The purified linamarase was a dimeric protein with a molecular mass of 210 kDa; the enzyme showed optimum catalytic activity at pH 5·5 and 40 °C and had a wide range (3·0–7·0) of pH stability. The enzyme substrate specificity on plant cyanogenic glycosides was wide; the K_m value for linamarin was 2·93 mmol l⁻¹. The addition, before processing, of the fungal crude enzyme to cassava roots facilitated and shortened detoxification; after 24 h of fermentation, all cyanogenic glycosides were hydrolysed.     

Effect of feeding level and feeding frequency on specific dynamic action in Silurus meridionalis

The effect of feeding level ( F _L; 0·5 to 4% dry diet mass per wet fish body mass) and feeding frequency (once every 4 days to twice per day) on postprandial metabolic response was investigated in southern catfish Silurus meridionalis at 27·5° C. The results showed that there was no significant difference in the specific dynamic action (SDA) coefficient among the groups of different feeding levels ( P  > 0·05). The duration increased from 26·0 to 40·0 h and the peak metabolic rate increased from 207·8 to 378·8 mg O₂ kg⁻¹ h⁻¹ when the feeding level was increased from 0·5 to 4%. The relationship between the peak metabolic rate ( R _P, mg O₂ kg⁻¹ h⁻¹) and F _L could be described as: R _P = 175·4 + 47·3 F _L( r ² = 0·943, n  = 40, P  < 0·001). The relationship between the SDA duration ( D , h) and F _L could be described as D =30·97 F _L^0·248 ( r ²=0·729, n =40, P  < 0·001).     

CMC-liquefying enzyme, a low molecular mass initial cellulose-decomposing cellulase responsible for fragmentation from Streptomyces sp. LX

Streptomyces sp. LX, newly isolated from soil, was shown to secrete a carboxylmethylcellulose (CMC)-liquefying enzyme that cleaves the CMC chains, releasing negligible reducing terminals. The new enzyme, named component C₂, was purified to homogeneity by dialysation. It has a molecular mass of 9·8 kDa. The pH optimum of the enzyme activity is 6·4 and its temperature optimum is 50°C. It retains full activity at pH 4–6·4 upon incubation at 50°C for 30 min. The enzyme has significant fragmentation activity on filter paper despite the absence of weight loss, release of reducing sugars and depolymerization during incubation with filter paper. The one-electron oxidative reaction is shown not to participate in the fragmentation of filter paper by enzyme C₂.     

Sibling cannibalism among juvenile vundu under controlled conditions. II. Effect of body weight and environmental variables on the periodicity and intensity of type II cannibalism

Cannibalism among starved groups of juvenile (19–48 days old) vundu catfish Heterobranchus longifilis was 66·5% nocturnal, and its impact under modified day length was proportional to the duration of the dark phase. Shallow depth and high population density decreased the intensity of cannibalism, whereas low density and deeper environments had an opposite effect. The presence of refuges had no significant effect on cannibalism. The maintenance ( R _maint) and maximum ( R _max) daily food rations (% day⁻¹) of cannibals feeding on live prey were modelled as R _maint=3·899 W _C^0·327 ( r ²=0·684; d.f.=31), and R _max=49.545 W _C^0·321 ( r ²=0·999; d.f.=5), where W _C was the body weight of the cannibal (g). The latter model indicated that the impact of a cannibal on a population decreased by a 20% margin each time the cannibal doubled its body weight, and suggested that cannibalism among vundu would become insignificant for cannibals heavier than 30 g. The significance of these findings is discussed within the contexts of vundu aquaculture and of general, conceptual models of the dynamics of cannibalism among fishes.     

Effects of temperature, body size and feeding on rates of metabolism in young‐of‐the‐year haddock

The mean rate of oxygen consumption (routine respiration rate, R _R, mg O₂ fish⁻¹ h⁻¹), measured for individual or small groups of haddock Melanogrammus aeglefinus (3–12 cm standard length, L _S) maintained for 5 days within flow‐through respiratory chambers at four different temperatures, increased with increasing dry mass ( M _D). The relationship between R _R and M _D was allometric ( R _R = α  M ^b ) with b values of 0·631, 0·606, 0·655 and 0·650 at 5·0, 8·0, 12·0 and 15·0° C, respectively. The effect of temperature ( T ) and M _D on mean R _R was described by     indicating a Q ₁₀ of 2·27 between 5 and 15° C. Juvenile haddock routine metabolic scope, calculated as the ratio of the mean of highest and lowest deciles of R _R measured in each chamber, significantly decreased with temperature such that the routine scope at 15° C was half that at 5° C. The cost of feeding ( R _SDA) was c . 3% of consumed food energy, a value half that found for larger gadoid juveniles and adults.