Pseudomonas putida is able to grow on malonate as a sole source of carbon and energy. Malonate decarboxylase is a key enzyme catabolizing malonate to acetate and CO₂. The enzyme consists of the five different subunits, α (60 kDa), β (33 kDa), γ (28 kDa), δ (13 kDa), and ε (30 kDa). The smallest subunit δ is an acyl-carrier protein (ACP) possessing 2'-(5"-phosphoribosyl)-3'-dephospho-CoA as a prosthetic group. Acylation of ACP is the initial step triggering the decarboxylation of malonate in a cyclic manner, i.e., the δ subunit is activated to form acyl-S-ACP by the ε subunit in the presence of acetyl-CoA or malonyl-CoA as an active acyl donor. The acetyl residue on the δ subunit is replaced with malonate by the α subunit, and the malonyl residue subsequently undergoes decarboxylation by the subunits β and γ, thereby regenerating acetyl-S-ACP. This unique cyclic reaction mechanism that amplifies acetate as its product in proportion to the amount of a given acetyl-CoA and/or malonyl-CoA is allows one to detect pmol levels of the CoA derivatives. Moreover, one can separately measure the three CoA molecular species, acetyl-CoA, malonyl-CoA, and nonesterified CoA (CoASH), by a combination of the elimination of acetyl-CoA with citrate synthetase (EC 4.1.3.7) or acetylation of CoASH with phosphate acetyltransferase (EC 2.3.1.8). The micromethod, named the acyl-CoA cycling method, is useful to define the rapid changes in vivo in the size and composition of the CoA pool. By this micromethod, it has been demonstrated that there is a remarkable difference in the size and composition of intracellular pools of CoASH and CoA thioesters between aerobic bacteria and facultatively anaerobic bacteria. It has also been revealed that the composition of CoA pools in facultatively anaerobic bacteria drastically changes within minuites in response to the quality and quantity of the carbon source in the medium, growth phase, pH, incubation temperature, osmotic stress, or antibiotics to inhibit energy yielding systems and fatty acid biosynthesis.

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Actinobacillus actinomycetemcomitans, a Gram-negative periodontopathic bacterium, produces a leukotoxin belonging to the RTX family. The leukotoxin determinant consists of four contiguous genes designated ltxC, ltxA, ltxB and ltxD. Although the ltx operon appears to be present in all A. actinomycetemcomitans strains, the level of toxin expression varies considerably between the strains. Several molecular genetic studies have indicated that the difference in leukotoxin production among strains was dependent on the structure of the leukotoxin promoter region. In addition, chemostat culture studies revealed that the production of leukotoxin was influenced greatly by various environmental factors, e.g. growth rate, oxygen, bicarbonate, external sugar level, and pH. In this paper, the state of knowledge of leukotoxin expression in A. actinomycetemcomitans is reviewed.

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The germination of bacterial endospores involves the following structural and physiological events: binding of nutrients to specific receptors located in the spore, the release of cations and calcium dipicolinate (CaDPA) from the core, degradation of the spore cortex, and the breakdown of small acid soluble proteins (SASP) protecting the DNA in the core. This minireview focuses specifically on the action of germination-specific lytic enzymes (GSLE) and the role of calcium ions in the degradation and collapse of the spore cortex. Several key questions for future research are discussed.

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The study of fibrolytic enzymes of rumen ciliate protozoa has lagged behind that of rumen bacteria and fungi. However, molecular biology has enhanced the investigation of rumen protozoa, and significant information has been accumulated in the last decade. The cellulolytic activities of cell-free extracts from each species of rumen ciliate protozoa have been summarized. Large Ophyroscolecidae such as Epidinium, Polyplastron, and Eudiplodinium have higher levels of endoglucanase and xylanase activity. On the other hand, Entodinium spp. have only weak activity to degrade plant fibers. The molecular weight of the main endoglucanase appears to be less than 40k Daltons, and the endoglucanases have the ability to bind cellulose. A total of eight fibrolytic enzyme genes have been cloned from cDNA libraries of Epidinium caudatum and Polyplastron multivesuculatum, and have been registered in GenBank. Four of these 8 clones have high homology with glycoside hydrolase family 5 endoglucanase (GHF5) genes. The remaining four have homology with xylanase (GHF10 and GHF11) genes. The guanine (G)+cytocine (C) content (mol%) of protozoal genes is about 30% and the codon usage is specific and different from that of the endoglucanase or xylanase genes originating from other microorganisms. The codon for Lys in the protozoal ORF is mainly (about 90%) AAA, unlike the Lys codons in rumen bacteria (25%) and the fungal gene (27%). The phylogenetic tree for family 5 endoglucanases shows that the endoglucanases of four ciliates are closely related to each other and form a cluster. On the other hand, the xylanases of these ciliates appear more closely related to xylanases from gram-positive bacteria such as Ruminococcus and Clostridium than to other xylanases of eukaryotic origin.

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We analyzed symbiotic protistan engulfment systems using the method of fluorescent imaging. Symbiotic protists can be divided into two protistan groups, Type S (protists that contain uniformly small and spherical vesicles) and Type L (protists that contain vesicles that vary in both size and form). This classification corresponds not to the taxonomic classification, but to the micro-habitats of the protists. Our results suggested that these protists might have adapted to different resources in the hindgut environment of lower termites.

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Termites of most isopteran families and wood-feeding cockroaches of the genus Cryptocercus usually harbor more than one symbiotic flagellate species in their hindgut. To evaluate the similarity of their symbiont faunae, data on symbiont composition at a generic level were examined by cluster analysis and type III quantification method. In both analyses, the symbiont composition recorded from host insects belonging to the same families or monophyletic family groups tended to be similar. This tendency was particularly remarkable in the clade Kalotermitidae and the clade Rhinotermitidae plus Serritermitidae. Two basal host groups, the Cryptocercidae and the Mastotermitidae, exhibited very different symbiont compositions. These findings suggested that the symbiont faunae mainly reflect the host's phylogenetic relationships. Within the Rhinotermitid hosts, the genus Reticulitermes showed a unique symbiont fauna although it is not a basal taxon in the Rhinotermitidae. Horizontal transfers of symbiotic protists might explain such anomalistic fauna.

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Four strains of hydrogenotrophic methanogens showing narrow ranges of utilizable substrates were isolated in pure cultures from the guts of various feeding groups of (phylogenetically) higher termites. An analysis of the 16S rRNA gene sequence revealed that three strains were closely related to Methanobacterium bryantii (>99% nucleotide identity), the other to the genus Methanobrevibacter. The latter was related to a clone identified previously from the gut of a higher termite without cultivation (clone MPn19) and Methanobrevibacter arboriphilicus (99.0 and 97.9% nucleotide identity, respectively) but distinct from the species identified in lower termites. The specific detection of related methanogens in the gut population by nested-PCR indicated that every termite harbored the species of Methanobrevibacter. However, the methanogen related to the Methanobacterium strain was not detected in two termite species from which the Methanobacterium strains were isolated, suggesting that they are less prevalent in the gut community.

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Isolates from two multinucleate Rhizoctonia species, namely Rhizoctonia solani and Rhizoctonia circinata, and binucleate Rhizoctonia spp., were evaluated for the presence of lectins, using human erythrocytes. Isolates from R. solani had similar lectins across the anastomosis groups. Agglutination profiles, however, revealed individual preferences for human blood types but with a general preference for type A over B and O. R. solani lectins had a general affinity for N-acetyl-D-galactosamine and mucin. Also, some isolates from the binucleate Rhizoctonia AG-DII, had lectins that showed strong affinity for glycoproteins, fetuin and mucin. There was no lectin activity in members of R. circinata. The lectins in members of the genus Rhizoctonia varied in content and character across the species sampled but had similar affinity for mucin. Furthermore, 62% of the self-anastomosing and 35% of the non-self anastomosing isolates tested showed lectin activity. The presence of lectins in both self-anastomosing and non-self anastomosing isolates suggests that lectins may not be directly involved in the recognition process in hyphal anastomosis.

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Serratia marcescens produces the red pigment prodigiosin and biosurfactant serrawettin at 30°C but not at 37°C. In an analytic study of the thermoregulation of S. marcescens 274, we noticed another temperature-dependent activity that was bacteriostatic to gram-negative and -positive bacterial species, at 37°C but not at 30°C. The thermo-insensitive isogenic mutant strain N0075 (a producer of prodigiosin and serrawettin even at 37°C) was devoid of such growth inhibitory activity. The revertant of strain N0075 made by transformation again exhibited the temperature-dependent mode of prodigiosin and serrawettin production, and bacteriostatic activity. This novel activity was correlated to the thermo-responsive acidification of the medium by S. marcescens. In response to higher environmental temperatures, S. marcescens seems to suppress its own growth and the growth of other bacteria.

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Trichosporon cutaneum strain KUY-6A, a cyclohexanecarboxylic acid-utilizing yeast, was able to grow on cyclopentanol (CPOL) as a sole source of carbon and energy. Growth experiments revealed the strain, KUY-6A, could utilize up to 42 mM of CPOL with an optimum at 24 mM. Optimal growth was found between pH 4.0 to 9.0. The generation time under optimal growth conditions on CPOL was 3.0 h. Analysis indicated that cyclopentanone (CPON) and glutaric acid were intermediates of CPOL metabolism in strain KUY-6A. The results of growth and enzyme experiments are consistent with the degradation of CPOL via CPON, 5-valerolactone, 5-hydroxyvaleric acid, and glutaric acid.

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The effects of soil moisture conditions and pre-incubation at low temperature on bacterial wilt of tomato caused by Ralstonia solanacearum were evaluated using two soils. Soils were adjusted to different levels of moisture and inoculated with the pathogen. Then, tomato seedlings were transplanted and grown in a climatron at 30°C. In some treatments, pre-incubation at different temperatures was conducted for 3 days before cultivation at 30°C. Wilt incidence was lower under drier soil conditions like 20 to 30% of maximum water-holding capacity. The survival of the pathogen deteriorated under drier conditions, supporting the lower wilt incidence in the drier soils. Wilt incidence decreased when transplanted seedlings were pre-incubated at lower temperatures (20°C to 4°C), especially 4°C. Reduced wilt incidence was accompanied by reduced survival of the pathogen. This study suggests that lower moisture conditions and pre-incubation at lower temperatures reduced bacterial wilt and have a negative effect on the survival of R. solanacearum.

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Microbes and Environments volume 19, No. 2 (June 2004), in the article "Microbial Biomass, Abundance and Community Diversity Determined by Terminal Restriction Fragment Length Polymorphism Analysis in Soil at Varying Periods after Occurrence of Forest Fire" by Jhonamie A. Mabuhay et al., page 154-162.
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