Aquabacter spiritensis,gen. nov., sp. nov. an aerobic,gas-vacuolate aquatic bacterium

A gram-negative, rod-shaped, gas-vacuolate, aerobic, heterotrophic bacterium was isolated from Spirit Lake, Washington. It was further characterized by phase-contrast and electron microscopy and phenotypic tests. The isolate does not utilize carbohydrates. Deoxyribonucleic acid-ribosomal ribonucleic acid (rRNA) hybridizations showed that this bacterium is a member of rRNA superfamily IV where it occupies a separate position on the Azorhizobium caulinodans rRNA sub-branch which, is part of an rRNA cluster containing also the Beijerinckia, the Bradyrhizobium japonicum and the Rhodopseudomonas palustris sub-branches. Its relationship to other gas-vacuolated genera within rRNA superfamily IV is discussed. A new genus Aquabacter with one species Aquabacter spiritensis is proposed. The type strain is strain SPL-1 (=ATCC 43981=LMG 8611).     

Purification and Properties of Lytic β-Glucanase from an Arthrobacter Bacterium

A glucanase was isolated from a culture fluid of an Arthrobacter bacterium. The purified enzyme preparations consisted of the glucanase components having the same enzymatic activity. The enzyme was stable in a broad pH range, but lost its activity rapidly at above 60°C. Optimum pH values were found to be 5.5~6.5.

The glucanase attacked the following glucan preparations and liberated a relatively small amount of reducing power: Saccharomyces cerevisiae glucan, Candida albicans glucan, Saccharomyces fragilis glucan, pachyman, curdlan and laminaran. The most prominent sugar spot on the chromatogram of the digest from yeast glucan was identified with laminan-pentaose, and the other faint spots with a series of laminaridextrins. The β-1,6 glucosidic bonds in yeast glucan were not hydrolyzed and concentrated in a soluble fraction which was found near the origin of the chromatogram.     

10(R)-Hydroxystearic acid production by a novel microbe,NRRL B-14797, isolated from compost

Summary A bacterium, NRRL B-14797, isolated from composted manure, converted oleic acid exclusively to 10(R)-hydroxystearic acid in 3-day batch cultures. 9(Z)-Unsaturated fatty acids in a lipid extract from soybean soapstock were also hydrated effectively. Aerobic bioconversions by isolate B-14797 were compared with those byPseudomonas B-2994 andNocardia 5767, which produce mixtures of 10-hydroxy- and 10-ketostearic acids. The results of studies with resting cells and cell-free extracts were consistent with action of a hydratase and absence of secondary alcohol dehydrogenase in strain B-14797.The mention of firm names or trade products does not imply that they are endorsed or recommended by the US Department of Agriculture over other firms or similar products not mentioned.     

Identification of Potent Bacterium for Production of Chillproofing Enzyme

A bacterium was isolated which produced an extracellular proteolytic enzyme preventing chill haze of beer. This strain was named B–103.

Taxonomical studies on strain B–103 indicated that it was Gram-negative, aerobic, motile, and catalase-positive. Cells were single rods (0.2~0.6×1.0~1.8 μ) with a peritrichous flagella. The bacterium produced nitrite from nitrate, extracellular DNase and 5′-nucleotide from nucleoside. It was capable of nitrate respiration and of oxidative or fermentative cleavage of carbohydrates.

From these characteristics, it was identified as a strain of Serratia marcescens.     

Thermoanaerobium lactoethylicum spec. nov. a new anaerobic bacterium from a hot spring of Kamchatka

A new anaerobic thermophilic Gram-positive, nonsporeforming bacterium strain ZE-1 was isolated from a hot spring of Kamchatka (USSR). The cells are rod-shaped, (0.5–0.8 · 2.0–20 m), non-motile. The bacterium can grow between 42 and 75°C; the optimal temperature is 65°C. The growth is possible between pH values 5.0 and 8.5; optimal pH is 7.0. The cultures grow on the media containing peptone, yeast extract, or casein hydrolysate as nitrogen sources in the presence of glucose or some other sugars, mannitol or starch. The main fermentation products of glucose are ethanol, acetate, lactate, H₂, CO₂; byproducts are propionic, butyric and isovaleric acids. Glucose is metabolized via Embden-Meyerhoff-Parnas pathway. Molecular hydrogen does not inhibit growth. The bacterium does not reduce aceton to isopropanol, but is able to form H₂S from elemental sulfur. The bacterium contains a soluble hydrogenase. This enzyme catalyzes both evolution and uptake of H₂ and is active in the presence of methyl viologen. The DNA-base composition is 34.6 mol%; the genome size 2.08x10⁹ D. The name proposed for the isolated bacterium strain ZE-1 is Thermoanaerobium lactoethylicum spec. nov.     

Zymobacter palmae gen. nov., sp. nov., a new ethanol-fermenting peritrichous bacterium isolated from palm sap

Zymobacter palmae gen. nov., sp. nov. was proposed for a new ethanol-fermenting bacterium that was isolated from palm sap in Okinawa Prefecture, Japan. The bacterium is gram-negative, facultatively anaerobic, catalase-positive, oxidase-negative, nonspore-forming and peritrichously flagellated. It requires nicotinic acid for growth. It ferments hexoses, -linked di- and tri-saccarides, and sugar alcohols (fructose, galactose, glucose, mannose, maltose, melibiose, saccharose, raffinose, mannitol and sorbitol). Fifteen percent of maltose in broth medium is effectively fermented, whereas glucose with a concentration higher than 10% delayed growth initiation and decreased growth rates. Maltose is fermented to produce ethanol and CO₂ with a trace amount of acids. Approximately 2 mol of ethanol are produced from 1 mol moiety of hexose of maltose. The organism possesses ubiquinone-9. The G+C content of the DNA is 55.8+-0.4 mol%. Major cellular fatty acids were palmitic and oleic acids and cyclopropanic acid of C_19:0. Characteristic hydroxylated acid was 3-hydroxy dodecanoic acid. The bacterium is distinct from other ethanol-fermenting bacteria belonging to the genera Zymomonas Kluyver and van Niel 1936 and Saccharobacter Yaping et al. 1990 with respect to chemotaxonomic and other phenotypic characters to warrant to compose a new genus and a new species. The type strain is strain T109 (= IAM 14233).Abbreviation IAM IAM Culture Collection, Institute of Applied Microbiology. The University of Tokyo     

Infection of potato tubers with soft rot bacteria

Stolons attached to developing potato tubers were inoculated with the soft rot bacterium Erwinia carotovora var. atroseptica. Almost all the stolons rotted, but soft rots developed in less than 10% of new tubers; the bacterium was isolated later from these tubers. No rots developed in the other tubers but the bacterium was later isolated from about half of them. It could not be isolated from tubers attached to inoculated stolons where the rot on them did not extend to the tuber or from tubers attached to stolons that were not inoculated though many of these rotted. The bacterium was reisolated from almost all arrested lesions in tubers inoculated 8 month earlier with E. carotovora var. atroseptica. Blackleg did not develop from plants grown fom these tubers under various soil conditions. It did develop in a large proportion of plants from tubers inoculated shortly before planting and grown in cool, wet soil. Less than 1% blackleg developed in plants grown from tubers from plants with blackleg or from plants immediately adjacent. The presence of pectolytic bacteria and E. caratovora var. atroseptica in seed and new tubers was investigated during June, July and August. Although E. caratovora var. atroseptica was obtained from c. 40% tubers, only c. 0·3% of c. 8400 plants developed blackleg. The bacterium was isolated from only three of 160 new tubers sampled during the summer.     

Isolation and characterization of a Clostridium sp. with cinnamoyl esterase activity and unusual cell envelope ultrastructure

Microorganisms that hydrolyse the ester linkages between phenolic acids and polysaccharides in plant cell walls are potential sources of enzymes for the degradation of lignocellulosic waste. An anaerobic, mesophilic, spore-forming, xylanolytic bacterium with high hydroxy cinnamic acid esterase activity was isolated from the gut of the grass-eating termite Tumilitermes pastinator. The bacterium was motile and rod-shaped, stained gram-positive, had an eight-layered cell envelope, and formed endospores. Phylogenetic analysis based on 16S rRNA indicated that the bacterium is closely related to Clostridium xylanolyticum and is grouped with polysaccharolytic strains of clostridia. A wide range of carbohydrates were fermented, and growth was stimulated by either xylan or cellobiose as substrates. The bacterium hydrolysed and then hydrogenated the hydroxy cinnamic acids (ferulic and p-coumaric acids), which are esterified to arabinoxylan in plant cell walls. Three cytoplasmic enzymes with hydroxy cinnamic acid esterase activity were identified using non-denaturing gel electrophoresis. This bacterium possesses an unusual multilayered cell envelope in which both leaflets of the cytoplasmic membrane, the peptidoglycan layer and the S layer are clearly discernible. The fate of all these components was easily followed throughout the endospore formation process. The peptidoglycan component persisted during the entire morphogenesis. It was seen to enter the septum and to pass with the engulfing membranes to surround the prespore. It eventually expanded to form the cortex, verification for the peptidoglycan origin of the cortex. Sporogenic vesicles, which are derived from the cell wall peptidoglycan, were associated with the engulfment process. Spore coat fragments appeared early, in stage II, though spore coat formation was not complete until after cortex formation. Received: 11 February 1999 / Accepted: 28 May 1999     

Plasmodium paranucleophilum n. sp. from a South American Tanager*

SYNOPSIS. The avian malaria parasite described in this paper was isolated from a South American tanager of uncertain species, somewhat tentatively identified as belonging to the genus Tachyphonus. It is believed to have come from northern Brazil. Because the erythrocytic stages are, except for minor differences, similar to those of Plasmodium nucleophilum it is proposed to call it P. paranucleophilum. The chief difference separating the 2 species is the appearance of phanerozoites of the new species in lymphocytes of the circulating blood; these are characteristically vacuolated and give rise to moderate numbers of merozoites, probably in most cases 50 or less. Strains of P. nucleophilum isolated from other passerine birds also produce phanerozoites parasitizing lymphocytes, but such cells have never been observed in the blood stream. A subspecies, P. nucleophilum toucani, which we isolated from a Swainson's toucan Ramphastos swainsonii, also produces phanerozoites invading lymphocytes but it is less selective; endothelial cells of the brain are heavily attacked. It is also very lethal to canaries, whereas passerine strains of P. nucleophilum are usually quite benign. P. paranucleophilum, if the strain we have isolated is typical, lies between the 2 in pathogenicity, causing marked anemia but seldom death.     

Biochemical and physiological characteristics of Xenorhabdus species,symbiotically associated with entomopathogenic nematodes including Steinernema kushidai and their pathogenicity against Spodoptera litura (Lepidoptera: Noctuidae)

The symbiotic bacterium strain, SK-1 isolated from Steinernema kushidai, a new species of entomopathogenic nematode, was compared with other strains of Xenorhabdus species. Like other Xenorhabdus nematophilus strains, this new strain is gram-negative, facultatively anaerobic, peritrichously flagellated rod and negative for catalase and nitrate reduction. It can be distinguished from the other Xenorhabdus spp. by differences in reactions to phenylalanine deaminase, no acid production from myo-inositol and utilizations of inosine, dl-malate, formate and methanol. Intra-haemocoelic injection of actual cells or liquid culture supernatant into sixth instar larvae of Spodoptera litura for either Phase I or II variants were not pathogenic. Other strains of X. nematophilus showed pathogenicity for whole cell injections. The supernatants of strain D-1 and ATCC 19061, which are symbionts of Steinernema carpocapsae were pathogenic, however pathogenicity decreased and then terminated by increases in temperature.     

Pseudomonas syringae pv. helianthi (Kawamura 1934) Dye, Wilkie et Young 1978, a Pathogen of Sunflower (Helianthus annuus L.)

Among the bacterial strains isolated from diseased sunflower leaves, eight were studied in some detail. A fluorescent pseudomonad isolated from necrotic tissues and its reisolates belong to group Ia of phytopathogenic pseudomonads which includes Pseudomonas syringae bacterium. A study of host range indicated that the pathogen infects only sunflower but not the other plant species. Based on the pathogenicity study and biochemical and physiological tests, it was concluded that the pathogen belongs to the bacterium Pseudomonas syringae pv. helianthi.     

Pseudomonas syringae pv. helianthi (Kawamura 1934) Dye,Wilkie et Young 1978, a Pathogen of Sunflower (Helianthus annuus L.)

Among the bacterial strains isolated from diseased sunflower leaves, eight were studied in some detail. A fluorescent pseudomonad isolated from necrotic tissues and its reisolates belong to group Ia of phytopathogenic pseudomonads which includes Pseudomonas syringae bacterium. A study of host range indicated that the pathogen infects only sunflower but not the other plant species. Based on the pathogenicity study and biochemical and physiological tests, it was concluded that the pathogen belongs to the bacterium Pseudomonas syringae pv. helianthi.     

Isolation and characterization of a novel lactic acid bacterium for the production of lactic acid

We isolated a novel lactic acid bacterium from a Korean traditional fermented food, soybean paste. The newly isolated strain, dubbed RKY2, grew well on glucose, sucrose, galactose, and fructose, but it could not utilize xylose, starch, or glycerol. When the partially amplified 16S rDNA sequence (772 bp) of the strain RKY2 was compared with 10 reference strains, it was found to be most similar toLactobacillus pentosus JCM 1588^T, with 99.74% similarity. Therefore, the strain RKY2 was renamedLactobacillus sp. RKY2, which has been deposited in the Korean Collection for Type Cultures as KCTC 10353BP.Lactobacillus sp. RKY2 was found to be a homofermentative lactic acid bacterium, because its end-product from glucose metabolism was found to be mainly lactic acid. It could produce more than 90 g/L of lactic acid from MRS medium supplemented with 100 g/L of glucose, with 5.2 g L⁻¹ h⁻¹ of productivity and 0.95 g/g of lactic acid yield.     

Streptomyces neyagawaensis as a control for the hazardous biomass of Microcystis aeruginosa (Cyanobacteria) in eutrophic freshwaters

An aquatic bacterium capable of eliminating the cyanobacterium Microcystis aeruginosa was isolated from the sediment of an eutrophic lake (Lake Juam, Korea). On the basis of 16S rDNA sequences and biochemical and morphological characteristics, the isolate was determined to be Streptomyces neyagawaensis. It grew optimally at 40 °C and pH 7. In the presence of this bacterium, the biomass of cyanobacterium M. aeruginosa NIES-298 was strongly suppressed, by up to 84.5% in abundance compared to the control. The antialgal activity of S. neyagawaensis depended on the growth phase of the cyanobacterium, but not of the antialgal bacterium. The antialgal activity of S. neyagawaensis was effective against a wide range of algae, including the green alga Chlorella sp., the diatoms Aulacoseira granulata and Stephanodiscus hantzschii, and four cyanobacteria, M. aeruginosa NIES-44, Anabaena cylindrica, Anabaena flos-aquae, and Oscillatoria sancta. S. neyagawaensis indirectly attacked M. aeruginosa by secretion of extracellular antialgal substances that were localized in the bacterial periplasm and had a specific activity of 7.7 U/μg. These results suggest that indigenous bacteria isolated from sediments may have potential application in controlling harmful cyanobacterial blooms in freshwaters.     

Purification and Amino Acid Sequence of MP-III 4R D49 Phospholipase A2 from Bothrops pirajai Snake Venom, a Toxin with Moderate PLA2 and Anticoagulant Activities and High Myotoxic Activity

MP-III 4R PLA₂ was purified from the venom of Bothrops pirajai venom (Bahia's jararacussu) after three chromatographic steps which started with RP-HPLC. The complete amino acid sequence of MP-III 4R PLA₂ from Bothrops pirajai was determined by amino acid sequencing of reduced and carboxymethylated MP-III 4R and the isolated peptides from clostripain and protease V8 digestion. MP-III 4R is a D49 PLA₂ with 121 amino acid residues and has a molecular weight estimated at 13,800 Da, with 14 half-cysteines. This protein showed moderate PLA₂ and anticoagulant activity. This PLA₂ does not have a high degree of homology with other bothropic PLA₂-like myotoxins (~75%) and nonbothropic myotoxins (~60%). MP-III 4R is a new PLA₂, which was isolated using exclusively analytical and preparative HPLC methods. Based on the N-terminal sequence and biological activities, MP-III 4R was identified as similar to piratoxin-III (PrTX-III), which was isolated by conventional chromatography based on molecular exclusion ion exchange chromatography. Clinical manifestations indicate that at the site of toxin injection, there may be pain of variable intensity, because animals continue to lick the limb. No clinical sign indicating general toxicity was noticed. Myotoxicity was observed in gastrocnemius muscle cells after exposure to MP-III 4R, with a high frequency (70%) of affected muscle fibers.     

Lysis of Aphanizomenon flos-aquae (Cyanobacterium) by a bacterium Bacillus cereus

A phytoplankton-lytic (PL) bacterium, Bacillus cereus, capable of lysing the bloom-forming cyanobacterium Aphanizomenon flos-aquae was isolated from Lake Dianchi of Yunnan province, China. This bacterium showed lytic activities against a wide range of cyanobacteria/algae, including A. flos-aquae, Microcystis viridis, Microcystis wesenbergi, Microcystis aeruginosa, Chlorella ellipsoidea, Oscillatoria tenuis, Nostoc punctiforme, Anabaena flos-aquae, Spirulina maxima, and Selenastrum capricornutum. Chlorophyll a contents, phycocyanin contents, and photosynthetic activities of the A. flos-aquae decreased evidently in an infected culture for a period. Bacterium B. cereus attacked rapidly A. flos-aquae cells by cell-to-cell contact mechanism. It was shown that the lysis of A. flos-aquae began with the breach of the cyanobacterial cell wall, and the cyanobacterial cell appeared abnormal in the presence of the PL bacterium. Moreover, transmission electron microscope examinations revealed that a close contact between the bacterium and the cyanobacterium was necessary for lysis. Some slime extrusions produced from B. cereus assisted the bacterial cells to be in close association with and lyse the cyanobacterial cells. These findings suggested that this bacterium could play an important role in controlling the Aphanizomenon blooms in freshwaters.     

What happens in the pith stays in the pith: tissue‐localized defense responses facilitate chemical niche differentiation between two spatially separated herbivores

Herbivore attack is known to elicit systemic defense responses that spread throughout the host plant and influence the performance of other herbivores. While these plant‐mediated indirect competitive interactions are well described, and the co‐existence of herbivores from different feeding guilds is common, the mechanisms of co‐existence are poorly understood. In both field and glasshouse experiments with a native tobacco, Nicotiana attenuata, we found no evidence of negative interactions when plants were simultaneously attacked by two spatially separated herbivores: a leaf chewer Manduca sexta and a stem borer Trichobaris mucorea. T. mucorea attack elicited jasmonic acid (JA) and jasmonoyl‐l ‐isoleucine bursts in the pith of attacked stems similar to those that occur in leaves when M. sexta attacks N. attenuata leaves. Pith chlorogenic acid (CGA) levels increased 1000‐fold to levels 6‐fold higher than leaf levels after T. mucorea attack; these increases in pith CGA levels, which did not occur in M. sexta‐attacked leaves, required JA signaling. With plants silenced in CGA biosynthesis (irHQT plants), CGA, as well as other caffeic acid conjugates, was demonstrated in both glasshouse and field experiments to function as a direct defense protecting piths against T. mucorea attack, but not against leaf chewers or sucking insects. T. mucorea attack does not systemically activate JA signaling in leaves, while M. sexta leaf‐attack transiently induces detectable but minor pith JA levels that are dwarfed by local responses. We conclude that tissue‐localized defense responses allow tissue‐specialized herbivores to share the same host and occupy different chemical defense niches in the same hostplant.     

Bacterial degradation of 3-chloroacrylic acid and the characterization of cis- and trans-specific dehalogenases

A coryneform bacterium that is able to utilize cis- and trans-3-chloroacrylic acid as sole carbon source for growth was isolated from freshwater sediment. The organism was found to produce two inducible dehalogenases, one specific for the cis- and the other for trans-3-chloroacrylic acid. Both dehalogenases were purified to homogeneity from cells induced for dehalogenase synthesis with 3-chlorocrotonic acid. The enzymes produced muconic acid semialdehyde (3-oxopropionic acid) from their respective 3-chloroacrylic acid substrate. No other substrates were found. The cis-3-chloroacrylic acid dehalogenase consisted of two polypeptide chains of a molecular weight 16.2 kDa. Trans-3-chloroacrylic acid dehalogenase was a protein with subunits of 7.4 and 8.7 kDa. The subunit and amino acid compositions and the N-terminal amino acid sequences of the enzymes indicate that they are not closely related.     

Production of a new compound, 7,10-dihydroxy-8-(E)-octadecenoic acid from oleic acid byPseudomonas sp. PR3

Summary Sixty-two cultures from the ARS Culture Collection and 10 cultures isolated from soil and water samples collected in Illinois were screened for their ability to convert agricultural oils to value-added industrial chemicals. A new compound, 7,10-dihydroxy-8-(E)-octadecenoic acid (DOD) was produced from oleic acid by a new strain,Pseudomonas sp. PR3 isolated from a water sample in Morton, IL. Strain PR3 is a motile, small rod-shaped, Gram-negative bacterium. It has multiple polar flagellae and is oxidase-positive. Strain PR3 grows aerobically and cannot grow anaerobically. The strain produces white, smooth colonies on agar plate and no water-soluble pigment. The yield of the product was greater than 60%. The optimum time, pH and temperature for the production of DOD were: 2 days, 7.0, and 30°C, respectively. Glycerol and dextrose support the growth of strain PR3, but the cells grown from the former failed to catalyse the conversion of oleic acid to DOD. The production of DOD is unique in that it involves a hydroxylation at two positions and a rearrangement of the double bond of the substrate molecule.The mention of firm names or trade products does not imply that they are endorsed or recommended by the U.S. Department of Agriculture over other firms or similar products not mentioned.     

Rhodospira trueperi gen. nov., spec. nov., a new phototrophic Proteobacterium of the alpha group

A new phototrophic purple bacterium was isolated from a flat, laminated microbial mat in a salt marsh near Woods Hole, Mass., USA. The spiral-shaped bacterium was highly motile and had bipolar tufts of flagella and intracytoplasmic membranes of the vesicular type. The major photosynthetic pigments were identified as the carotenoid tetrahydrospirilloxanthin and bacteriochlorophyll b. The long wavelength in vivo absorption maximum of the bacteriochlorophyll was at 986 nm. The marine bacterium showed optimal growth in the presence of 2% NaCl. It utilized a number of organic substrates as carbon and energy sources and required vitamins and sulfide as a reduced sulfur source for growth. In the presence of sulfide, elemental sulfur globules were formed outside the cells. Elemental sulfur was not further oxidized to sulfate. The new isolate had a unique lipid and fatty acid composition, and according to the 16S rRNA gene sequence, it is most similar to Rhodospirillum rubrum. It is described as a new species and assigned to a new genus with the proposed name Rhodospira trueperi. Received: 16 September 1996 / Accepted: 24 January 1997