Search

The objective of the current study was to examine cutinolytic esterase (i.e., cutinase) activity by pseudomonads and bacteria isolated from mixed-plant compost. Approximately 400 isolates representing 52 taxa recovered from mixed-plant compost using cuticle baits, along with 117 pseudomonad isolates obtained from a culture collection (i.e., non-compost habitats), were evaluated. The ability of isolates to degrade the synthetic cutin polycaprolactone (PCL) was initially measured. Isolates from 23 taxa recovered from the compost degraded PCL. As well, isolates from 13 taxa of pseudomonads cleared PCL. Secondary screening measured esterase activity induced by the presence of apple cuticle using the chromogenic substrate p-nitrophenyl butyrate. Eighteen isolates representing four taxa ( Alcaligenes faecalis , Bacillus licheniformis , Bacillus pumilus , and Pseudomonas pseudoalcaligenes ) recovered from compost exhibited substantial esterase activity when grown with cuticle. In contrast, none of the pseudomonad isolates from the culture collection produced appreciable esterase activity. Although degradation of PCL was not correlated with esterase activity, isolates that were unable to degrade PCL failed to produce measureable esterase activities. Zymogram analysis indicated that the esterases produced by bacteria from compost ranged in size from 29 to 47 kDa. A gene from P. pseudoalcaligenes (cutA) was found to code for a cutin-induced esterase consisting of 302 amino acids and a theoretical protein size of 32 kDa. The enzyme was unique and was most closely related to other bacterial lipases (≤48% similarity).

Three bacteria, Alcaligenes faecalis , Flavobacterium sp., and Providencia vermicola , were isolated from dauer juveniles of Rhabditis blumi . The pathogenic effects of the bacteria against 4th instar larvae of Galleria mellonella were investigated. Providencia vermicola and Flavobacterium sp. showed 100% mortality at 48 h after haemocoelic injection, whereas A. faecalis showed less than 30% mortality. Dauer juveniles showed 100% mortality against G. mellonella larvae, whereas axenic juveniles, which do not harbor associated bacteria, exhibited little mortality. All of the associated bacteria were used as a food source for nematode growth, and nematode yield differed with bacterial species. Among the bacterial species, P. vermicola was most valued for nematode yield, showing the highest yield of 5.2 × 10(4) nematodes/mL in the plate. In bacterial cocultures using two of the three associated bacteria, one kind stimulated the other. The highest total bacterial yield of 12.6 g/L was obtained when the inoculum ratio of P. vermicola to A. faecalis was 10:1. In air-lift bioreactors, the nematode growth rate increased with an increasing level of dissolved oxygen. The maximum nematode yield of 1.75 × 10(5) nematodes/mL was obtained at 192 h with an aeration rate of 6 vvm.

Pseudomonas testosteroni B-356 was able to degrade approximately 50% of the Aroclor 1242 mixture in shaken culture. The aims of the present study were to evaluate the capabilities of this bacterial strain to degrade PCBs in soil microcosms and to identify some of the factors likely to favor the degradative performance of the implanted bacteria. The presence of biphenyl as cosubstrate was the most important factor affecting PCB degradation in soil. However, because biphenyl was rapidly depleted in soil microcosms, repeated addition of small amounts of biphenyl to maintain a constant level of the cosubstrate allowed the achievement of a higher degree of degradation of the tetrachlorinated components of Aroclor 1242 than was achieved with a single addition at the time of inoculation. Degradation of di- and tri-chlorinated PCB congeners was less affected by repeated addition of biphenyl because these congeners were degraded very fast and complete degradation was achieved before biphenyl was depleted in the soil. Biodegradation was also related to bioavailability of the substrate. We observed that the proportion of each congener degraded was higher in the microcosms receiving both the producer of the surface-active agent, Alcaligenes faecalis B-556, and strain B-356. Under the best conditions (presence of a constant level of biphenyl and of strain B-556) P. testosteroni B-356 was able to degrade approximately 30% of the Aroclor 1242 added to soil microcosms; some other factors reducing the PCB degradation capabilities of the implanted bacteria are also discussed.Key words: Aroclor 1242, biodegradation, bioremediation, Pseudomonas, soil.

As a guide to both strain and process improvement and based on certain assumptions concerning both glucose and energy metabolism of the process organism, Alcaligenes faecalis var. myxogenes, the theoretical "maximum" carbon (glucose) substrate to product conversion efficiency (i.e., product yield) has been estimated for "curdlan-type" β(1 → 3)-glucan exopolysaccharide production in batch fermentations. Under nitrogen limitation, which promotes curdlan biosynthesis (μ = 0), the rate of glucose consumption for cellular maintenance energy (grams of glucose per gram of cells per hour) was approximately five times higher than under carbon limitation. The decrease in the theoretical "maximum" curdlan conversion efficiency of 74% to the average value of 50–56% was due primarily to the high maintenance coefficient of the nitrogen-starved culture.

Diazotrophic Alcaligenes faecalis strain A15 isolated from rice roots is associated with rice roots and resides in the mucilage layer. On examination by light and electron microscopy, about 10% of the bacteria that accumulated on the surface entered the root cells. Some bacteria were observed to penetrate the cell wall. By using a 10B α-track technique, the presence of bacteria inside the root was also demonstrated after the rice plants were inoculated with A. faecalis. This was further supported by results of an immunofluorescence study. Both single cells and protoplasts were isolated from rice root and its callus. Transmission and scanning electron microscopy showed that A. faecalis grew inside the root cell. Callus, developed from rice root cells inoculated with this bacterium, grew well in nitrogen-free culture medium, while it died within 30 days without inoculation. Nitrogen fixation in the callus was confirmed by 15N tracer methodology and the rate ranged from 18.5 to 35.5 μg N fixed/g dry weight callus per day.Key words: Alcaligenes faecalis, endorhizospheric nitrogen fixation, rice–bacteria symbiont.

Alcaligenes faecalis, grown on 3-cyclohexenecarboxylic acid supplemented with gluconate, was induced to metabolize cyclohexanecarboxylic acid, 3-cyclohexenecarboxylic acid, 1, 4-cyclohexadienecarboxylic acid, benzoic acid, and catechol. During growth, 1, 4-cyclohexadienecarboxylic acid, 3-heptene-1, 7-dioic acid, and benzoic acid were shown to be present in the culture supernatants. The results suggest that 3-cyclohexenecarboxylic acid may be metabolized by two pathways having 1, 4-cyclohexadienecarboxylic acid as a common intermediate. One pathway involved β-oxidation of coenzyme A intermediates; the other pathway involved benzoic acid as an intermediate. Studies on the consumption of oxygen per mole of substrate by cell suspensions indicated that the major portion of 3-cyclohexenecarboxylic acid was metabolized by the pathway involving benzoic acid. When grown on 3-cyclohexenecarboxylic acid as sole carbon and energy source, the organism was additionally induced to metabolize salicylic acid and gentisic acid, and growth medium contained salicylic acid. Thus, under the latter growth conditions a pathway involving salicylic acid as an intermediate was also induced.

Forty strains of Gram-negative rods which resembled Alcaligenes in failing to attack carbohydrates were studied. Twenty-four strains had lophotrichous flagella, three were monotrichous, three were either lophotrichous or monotrichous, six were peritrichous, and four were nonflagellated. Strains were grouped by their action on gelatin and nitrate, and by motility. Most lophotrichous and monotrichous organisms liquefied gelatin, reduced nitrate, or did both, whereas most peritrichous and nonflagellated rods failed to do either.Serological studies indicated antigenic heterogeneity, but some strains in the more homogeneous physiological groups were related, mainly through flagellar antigens. Three phages were obtained without induction but only one of these was lytic for a single heterologous strain. Inducible substances resembling bacteriocines were elaborated by several strains, and the activity was confined primarily to homogeneous physiological and serological groups. Some lophotrichous strains closely resembled but were not identical with the genus Lophomonas. The validity of the genus Alcaligenes is questioned, since few isolates resemble the original description (peritrichous flagella) and these could be included in the genus Achromobacter.

D-Amino acids have been found to induce protoplast-like forms (crescents) in Alcaligenes fecalis strain LB. This induction is similar in most respects to that by penicillin with the exception of the effects produced in tryptone. D-Amino acids appear to act synergistically with penicillin. The amino acid composition of cell walls of D-amino acid and penicillin-induced crescents as well as of normal cells has been examined by column chromatography. The main difference between normal and crescent cell walls consists of a reduction in the amount of the basic peptide unit of the phenol-insoluble fraction. These results are discussed in relation to the induction of protoplast-like forms.

The ability of exogenous D-methionine to prevent the incorporation of exogenous L-methionine into the protein of Alcaligenes fecalis has been studied. D-Methionine has been found to competitively inhibit the ability of the cell to concentrate medium L-methionine with an efficiency of about 14:1. It does not inhibit the incorporation of endogenous methionine into cell protein. D-Methionine is replaceably concentrated by the cell and is able to specifically displace L-methionine previously concentrated by the cell. The significance of these findings for the mechanism of concentration of amino acids by bacteria is discussed.

The biosynthesis of a thermogelable, extracellular homo-β-(1 → 3)-glucan called "curdlan," has been studied in batch and continuous cultures of Alcaligenes faecalis var. myxogenes. Curdlan production is associated with the poststationary phase of a nitrogen-depleted, aerobic batch culture. Exopolymer is not detected in single-stage, carbon-limited continuous cultures but curdlan can be isolated from the effluent of a nitrogen-limited chemostat operating at a dilution rate (D) of −1. A spontaneous variant of strain ATCC 21680 was isolated and found to be compatible with long-term, nitrogen-limited chemostat culture. The specific rate of curdlan production is approximately four times higher in poststationary batch cultures than in single-stage continuous fermentations. The product yield (Yp/s) associated with batch processing (nongrowing cultures) is approximately 0.5 g curdlan/g glucose, with CO2 being the only detectable by-product.

Aliphatic alcohols induced a state of continuous uncoordinated translational movement in Pseudomonas fluorescens, Citrobacter freundii, Alcaligenes faecalis, and Rhodospirillum rubrum. This behavior was reversible and ceased upon removal of the stimulus. A hypothesis based upon electrostatic mechanisms is presented to explain these observations.

The effect of penicillin on strain LB of Alcaligenes fccalis has been studied. In tryptone, 50 units/ml. of penicillin transforms the bacillary form of this organism into a protoplast-like structure (globular form) which is capable of exponential growth and division. The division rate of this globular form is about one-half that of the bacillary form. Removal of penicillin results in the reversion of the globular to the bacillary form. Multiplication of the globular form has been found to depend on the presence of some principle present in tryptone not required for bacillary growth in the absence of penicillin. The kinetics of the transformation and reversion process have been studied, leading to the hypothesis that penicillin affects some component: within the cell, this component being concerned with cell division and the elaboration of the cell wall.

Experiments have been carried out on the penicillin-controlled induction of protoplast-like forms, crescents, in synchronized cultures of a strain of Alcaligenes fecalis. The results have shown that during the cell-division cycle the cell passes through a stage shortly before division during which it is less easily induced to form crescents than at other periods in the division cycle. The data presented indicate that this is not due to a transitory period in the division cycle during which the cell is refractory to penicillin. The findings are discussed in support of the hypothesis that penicillin exerts its effect by destroying some component within the cell whose functioning is necessary for further cell-wall synthesis.