[Heterogeneity of Rhodococcus opacus 1CP as a response to stress induced by chlorophenols].

Dissociation of Rhodococcus opacus 1CP during culturing in different media (containing phenol and its monochlorinated derivatives as the sole source of carbon and energy) was studied. Three variants of strain 1CP (S1, S2, and R) differing in the morphology of cells and colonies, lipid composition, and manner of growth on phenol and monochlorophenols were isolated. It was shown that 2- and 4-chlorophenols were most actively degraded by the smooth (S) forms of the culture, and that the rough (R) form predominated when the culture was grown in a rich medium. The S forms differed from the R forms of the strain by an increased content of cardiolipin, fatty acids, and phosphatidylethanolamine.

[The microbial transformation of phenanthrene and anthracene].

The transformation of phenanthrene and anthracene by Rhodococcus rhodnii 135, Pseudomonas fluorescens 26K, and Arthrobacter sp. K3 is studied. Twenty-one intermediates of phenanthrene and anthracene transformation are identified by HPLC, mass spectrometry, and NMR spectroscopy. P. fluorescens 26K and Arthrobacter sp. K3 are found to produce a wide range of intermediates, whereas R. rhodnii 135 oxidizes phenanthrene, resulting in the formation of a sole product, 3-hydroxyphenanthrene. Putative transformation pathways of phenanthrene and anthracene are proposed for the three bacterial strains studied. These strains can be used to obtain valuable compounds (such as hydroxylated polycyclic aromatic hydrocarbons) that are difficult to produce by chemical synthesis.

[Bacterial cold shock response at the level of DNA transcription, translation and chromosome dynamics]. / Reaktsiia bakterial'nykh kletok na kholodovyi shok na urovne dinamiki chromosomy, transkriptsii i translatsii.

The review is an attempt to analyze the data available in the literature concerning the response of mesophilic bacteria to cold shock at the level of DNA transcription, translation, and chromosome dynamics, i.e., in terms of cell biology. The review considers relevant molecular mechanisms and particular regulatory systems within the framework of a general cell response to cold shock. It is suggested that a short-term response to cold shock is necessary for bacteria to transit to a viable but nonculturable state and/or for their physiological and genetic adaptation to psychrotrophic life. It is emphasized that cell responses to cold and heat shocks are different and that DNA dynamics (i.e., its supercoiling, multiple bending, and condensation) and the rearrangement of the protein-synthesizing apparatus of cells (including the induction of alternative translational mechanisms) may play a central role in cell response to cold shock. The role of molecular chaperones in cold shock response is presumably of less importance than it is in the case of heat shock.

[Fluorene degradation by bacteria of the genus Rhodococcus]. / Prevrashchenie fluorena bakteriiami roda Rhodococcus.

Of the four investigated Rhodococcus strains (R. rhodochrous 172, R. opacus 4a and 557, and R. rhodnii 135), the first three strains were found to be able to completely transform fluorene when it was present in the medium as the sole source of carbon at a concentration of 12-25 mg/l. At a fluorene concentration of 50-100 mg/l in the medium, the rhodococci transformed 50% of the substrate in 14 days. The addition of casamino acids and sucrose (1-5 g/l) stimulated fluorene transformation, so that R. rhodochrous 172 could completely transform it in 2-5 days. Nine intermediates of fluorene transformation were isolated, purified, and structurally characterized. It was found that R. rhodnii 135 and R. opacus strains 4a and 557 hydroxylated fluorene with the formation of 2-hydroxyfluorene and 2,7-dihydroxyfluorene. R. rhodochrous 172 transformed fluorene via two independent pathways to a greater degree than did the other rhodococci studied.

[Aspects of cellular biology of the bacterial stationary phase: programmed cell death and regulation by guanosine tetraphosphate]. / Aspekty kletochnoi biologii statsionarnoi fazy bakterii: programmirovannaia gibel' kletok i reguliatsiia guanozintetrafosfatom.

The paper discusses (1) programmed cell death, the phenomenon typical of the stationary phase of bacteria occurring under unfavorable conditions, (2) its pleiotropic regulation by guanosine tetraphosphate, and (3) the conception of "addiction module," a specific genetic system responsible for the cell choice between survival and death under unfavorable conditions. The shortcomings of the proposed interpretation of the problem at hand are considered and the necessity of their further investigation is substantiated.

[The mechanism of formation of Pseudomonas aeruginosa biofilm, a type of structured population]. / Mekhanizm formirovaniia bioplenki--strukturirovannoi populiatsii Pseudomonas aeruginosa.

The paper is an attempt to analyze and generalize molecular and cell biology data on the formation of polysaccharide matrix--based biofilms. The conception of biofilms as structured populations sharing the characteristics of uni- and multicellular organisms and population is proposed.

Purification and properties of p-hydroxybenzoate hydroxylases from Rhodococcus strains.

Gram-positive bacteria of the genus Rhodococcus catabolize p-hydroxybenzoate (PHB) through the initial formation of 3,4-dihydroxybenzoate. High levels of p-hydroxybenzoate hydroxylase (PHBH) activity are induced in six different Rhodococcus species when these strains are grown on PHB as sole carbon source. The PHBH enzymes were purified to apparent homogeneity and appeared to be homodimers of about 95 kD with each subunit containing a relatively weakly bound FAD. In contrast to their counterparts from gram-negative microorganisms, the Rhodococcus PHBH enzymes prefer NADH to NADPH as external electron donor. All purified enzymes were inhibited by Cl- and for five of six enzymes more pronounced substrate inhibition was observed in the presence of chloride ions.

[Ecological strategy of bacteria: specific nature of the problem]. / Ekologicheskaia strategiia bakterii: spetsifika problemy.

An attempt is made to sum up the results of the many years of using the conception of ecological strategy in bacterial ecology. Taking into account the specificities of microorganisms and their natural selection and the coevolution of microorganisms within evolving microbial communities, an inference is derived that the ecological strategy of most bacteria is the sum of a number of particular canonical strategies, some of which are common to higher organisms. It is proposed to term these particular strategies ecological tactics. The author considers this review as a basis for discussion.

Identification of fluoropyrogallols as new intermediates in biotransformation of monofluorophenols in Rhodococcus opacus 1cp.

The transformation of monofluorophenols by whole cells of Rhodococcus opacus 1cp was investigated, with special emphasis on the nature of hydroxylated intermediates formed. Thin-layer chromatography, mass spectrum analysis, and (19)F nuclear magnetic resonance demonstrated the formation of fluorocatechol and trihydroxyfluorobenzene derivatives from each of three monofluorophenols. The (19)F chemical shifts and proton-coupled splitting patterns of the fluorine resonances of the trihydroxyfluorobenzene products established that the trihydroxylated aromatic metabolites contained hydroxyl substituents on three adjacent carbon atoms. Thus, formation of 1,2, 3-trihydroxy-4-fluorobenzene (4-fluoropyrogallol) from 2-fluorophenol and formation of 1,2,3-trihydroxy-5-fluorobenzene (5-fluoropyrogallol) from 3-fluorophenol and 4-fluorophenol were observed. These results indicate the involvement of fluoropyrogallols as previously unidentified metabolites in the biotransformation of monofluorophenols in R. opacus 1cp.

[General and molecular ecology of Legionella]. / Obshchaia i molekuliarnaia ékologiia legionell.

The review is devoted to the general and molecular ecology of bacteria of the genus Legionella in natural and anthropogenic environments. Invasion of amoebae and infusoria by legionellae and their replication in these protozoa can be considered to be a pre-adaptation for invasion of the human immune system. Symbiosis of bacteria and protozoa as a promising model of cellular microbiology and the conception of bacterial ecological niches are discussed in relation to the low fidelity of most bacterial species to their habitats (biotopes). The necessity of elaboration of a similar conception for microbial consortia and associations is emphasized.

[Dependence of transformation of chlorophenols by Rhodococci on position and number of chlorine atoms in the aromatic ring]. / Zavisimost' prevrashcheniia khlorfenolov rodokokkami ot polozheniia i chisla atomov khlora v aromaticheskom kol'tse.

Study of the conversion of chlorophenols by Rhodococcus opacus 1G, R. rhodnii 135, R. rhodochrous 89, and R. opacus 1cp disclosed the dependence of the conversion rate and pathway on the number and position of chlorine atoms in the aromatic ring. The most active chlorophenol converter, strain R. opacus 1cp, grew on each of the three isomeric monochlorophenols and on 2,4-dichlorophenol; the rate of growth decreased from 4-chlorophenol to 3-chlorophenol and then to 2-chlorophenol. The parameters of growth on 2,4-dichlorophenol were the same as on 3-chlorophenol. None of the strains studied utilized trichlorophenols. A detailed study of the pathway of chlorophenol transformation showed that 3-chloro-, 4-chloro-, and 2,4-dichlorophenol were utilized by the strains via a modified ortho-pathway. 2-Chlorophenol and 2,3-dichlorophenol were transformed by strains R. opacus 1cp and R. rhodochrous 89 via corresponding 3-chloro- and 3,4-dichloropyrocatechols, which were then hydroxylated with the formation of 4-chloropyrogallol and 4,5-dichloropyrogallol; this route had not previously been described in bacteria. Phenol hydroxylase of R. opacus 1G exhibited a previously undescribed catalytic pattern, catalyzing oxidative dehalogenation of 2,3,5-trichlorophenol with the formation of 3,5-dichloropyrocatechol but not hydroxylation of the nonsubstituted position 6.

[Physiology of microbial cell and metabolic engineering]. / Fiziologiia mikrobnoi kletki i metabolicheskaia inzheneriia.

This review is devoted to the problems of the physiology and cell biology of microorganisms in relation to metabolic engineering. The latter is considered as a branch of fundamental and applied biotechnology aimed at controlling microbial metabolism by methods of genetic engineering and classical genetics and based on intimate knowledge of cell metabolism. Attention is also given to the problems associated with the metabolic limitation of microbial biosyntheses, analysis and control of metabolic fluxes, rigidity of metabolic pathways, the role of pleiotropic (global) regulatory systems in the control of metabolic fluxes, and prospects of physiological and evolutionary approaches in metabolic engineering.

Preferential oxidative dehalogenation upon conversion of 2-halophenols by Rhodococcus opacus 1G.

The regiospecificity of hydroxylation of C2-halogenated phenols by Rhodococcus opacus 1G was investigated. Oxidative defluorination at the C2 position ortho with respect to the hydroxyl moiety was preferred over hydroxylation at the non-fluorinated C6 position for all 2-fluorophenol compounds studied. Initial hydroxylation of 2,3, 5-trichlorophenol resulted in the exclusive formation of 3, 5-dichlorocatechol. These results indicate that, in contrast to all other phenol ortho-hydroxylases studied so far, phenol hydroxylase from R. opacus 1G is capable of catalyzing preferential oxidative defluorination but also oxidative dechlorination.

Isolation and characterization of catechol 1,2-dioxygenases from Rhodococcus rhodnii strain 135 and Rhodococcus rhodochrous strain 89: comparison with analogous enzymes of the ordinary and modified ortho-cleavage pathways.

Catechol 1,2-dioxygenases of the ordinary ortho-cleavage pathway have been isolated from strains Rhodococcus rhodnii 135 and Rhodococcus rhodochrous 89 grown on phenol as the sole source of carbon and energy. The activities of the catechol 1,2-dioxygenases with 3- and 4-methylpyrocatechols were 1.3-1.5 times higher than those with pyrocatechol. The rate of oxidation of 3-chloropyrocatechol catalyzed by both enzymes was 20% of the rate of oxidation of unsubstituted pyrocatechol. The enzymes are homodimers composed of 37-kD subunits.

[Introduction to the biology of the bacterial stationary phase: mechanism of general response to stress]. / Vvedenie v biologiiu statsionarnoi fazy bakterii: mekhanizm obshchegootveta na stressy.

This review is devoted to the biology of stationary-phase bacteria, occurring in a specific physiological state at which they arrive in the process of complex response to various kinds of stresses accompanying the retardation and cessation of growth and reproduction. A general account of the problem is presented. Special emphasis is placed on one of the metabolic mechanisms involved in the formation of the physiological state of stationary-phase bacteria and performing primarily protective functions (the so-called general response of cells to stresses). The relationship between this and other regulatory mechanisms involved in the transition of bacteria to the stationary phase and the maintenance of this phase is discussed.

[Metastable phenotype of bacteria]. / Metastabil'nost' fenotipa u bakterii.

This review analyzes data available in the literature and the author's own data on the phenotypic variability of bacteria that occurs within the framework of a genotype unchanging in terms of the genetic information stored. This variability is a form of bacterial adaptation to an unstable environment and results from a specific form of natural selection. This phenomenon arose evolutionarily not as a mechanism to provide genetic diversity for the divergence process but as a mechanism of species stabilization; therefore, it was termed phenotype metastability. It includes, as specific variants, processes known as phase and antigenic variations, R-S-M dissociation, phenotype conversion, etc. The mechanisms of phenotype metastability are extremely diverse. They include alternative expression (of the switch on-switch off type) of individual genes or small groups of genes; variation in the composition of synthesized proteins controlled at the level of transcription; expression of complex phenotypes adapted to different environmental conditions that involves phage transposition, reading-frame-shift mutations, etc. The phenomenon of phenotype metastability is widespread among bacteria.

[Old problems of new bacterial systematics]. / O starykh problemakh novoi sistematiki bakterii.

This review analyzes the situation that arose after the existence of adaptive mutations had been proven. Manifold acceleration of the mutation process and intragenome rearrangements in nondividing cells demonstrates the inapplicability of the theory of neutral evolution to bacteria, the lack of isochronism between the evolution of individual macromolecules and the organism as a whole, and thus, the invalidity of using rRNA homologies as markers of the position of the organism in the phylogenetic tree and in the taxonomic system.

19F NMR study on the biodegradation of fluorophenols by various Rhodococcus species.

Of all NMR observable isotopes 19F is the one perhaps most convenient for studies on biodegradation of environmental pollutants. The reasons underlying this potential of 19F NMR are discussed and illustrated on the basis of a study on the biodegradation of fluorophenols by four Rhodococcus strains. The results indicate marked differences between the biodegradation pathways of fluorophenols among the various Rhodococcus species. This holds not only for the level and nature of the fluorinated biodegradation pathway intermediates that accumulate, but also for the regioselectivity of the initial hydroxylation step. Several of the Rhodococcus species contain a phenol hydroxylase that catalyses the oxidative defluorination of ortho-fluorinated di- and trifluorophenols. Furthermore, it is illustrated how the 19F NMR technique can be used as a tool in the process of identification of an accumulated unknown metabolite, in this case most likely 5-fluoromaleylacetate. Altogether, the 19F NMR technique proved valid to obtain detailed information on the microbial biodegradation pathways of fluorinated organics, but also to provide information on the specificity of enzymes generally considered unstable and, for this reason, not much studied so far.

[Kinetics of Pseudomonas fluorescens growth under different conditions of culturing]. / Kinetika rosta Pseudomonas fluorescens pri razlichnykh usloviiakh kul'tivirovaniia.

The dynamics of Pseudomonas fluorescens VKM-1472 growth was studied under the conditions of batch and continuous cultivation, and the behaviour of the organism was investigated upon nutrition shifts and starvation. At D greater than 0.375 h-1, just as in the batch culture with a substrate excess, the strain realised excessive metabolism [15]: the yield in terms of the substrate fell down (38% for the batch culture and 40% for the continuous culture), the titre of viable cells decreased to a considerable extent, and maintenance expenditures increased (3 times for qgluc and 7.5 times for qO2). When the culture was incubated under the oligotrophous conditions, the biomass yield decreased fourfold within 8 days and the titre of viable cells dropped down twofold within the same period of time. However, the organism was still capable of oxidising a wide range of substrates (17 substrates were studied). As soon as a substrate was added, it was oxidised at a high rate and changes in the macromolecular composition were characteristic of an "up-jump" in the growth rate [11]. It is possible that the growth and behaviour of this organism are associated with its ecological niche occupied in natural systems.