One-step detection of pathogens and cancer biomarkers by the naked eye based on aggregation of immunomagnetic beads.

This report shows that immunomagnetic beads (IMBs) can act as the optical readout for assays, in addition to serving as the carrier for purification/separation. Under the influence of an external magnet, IMBs are attracted to coat one side of a test tube. IMBs specifically bound to targets can form a narrow brown stripe, whereas free IMBs will form a diffuse, yellow coating on the side of the test tube. Target analytes can aggregate initially dispersed IMBs in a sample concentration-dependent manner, yielding a color change from yellow to brown that can be seen with the naked eye. This assay combines the convenience of a lateral flow assay, allowing a one-step assay to finish within 15 min, with the sensitivity of an enzyme-linked immonosorbent assay.

Azospirillum irakense sp. nov., a nitrogen-fixing bacterium associated with rice roots and rhizosphere soil.

A new species of nitrogen-fixing bacteria, Azospirillum irakense, was found associated with roots and the rhizosphere of rice in the region of Diwaniyah (Qadisya), Iraq. The seven isolates, on which the species description is based, have vibrioid to S-shaped cells with one polar flagellum in liquid medium. Additional lateral flagella are seen on cells grown on nutrient agar. Poly-beta-hydroxybutyrate granules are present in cells. Nitrogen fixation occurs in microaerobic conditions. The phenotypic characters were found to be very close to those of A. amazonense with the following differences: growth occurred in the presence of 3% NaCl, and at pH 5.5 and 8.5, myo-inositol was not utilized as sole source of carbon and energy and pectin was slowly (6 to 9 days) hydrolysed. The seven studied strains formed a DNA-relatedness group distinct from other Azospirillum and Herbaspirillum species. The G + C content of the DNA was 64 to 67 mol %. The type strain is KBC1 (CIP 103311).

Anaerobiospirillum species isolated from humans with diarrhoea.

Flagellated anaerobic motile spiral bacteria were isolated from the faeces of two patients with diarrhoea. They were recovered by the microaerophilic culture method used to detect campylobacters but demanded anaerobic conditions for subculture. Electron microscopy and other investigations showed them to be closely related to Anaerobiospirillum succiniproducens first described in beagle dogs and subsequently in three humans with bacteraemia.

Experimental infection and abortion of pregnant guinea pigs with a unique spirillum-like bacterium isolated from aborted ovine fetuses.

Study was made of the pathogenicity of a spirillum-like, anaerobic, gram-negative bacterium, originally isolated from aborted lambs, for pregnant guinea pigs. Reproducible conditions for propagation and preservation of the bacterium were determined as requisite for the preparation of cultures for animal inoculation. A preliminary experiment was done with 10 pregnant guinea pigs to test for an infective dose of organisms that would produce abortion. High-passage cultures (n = 50) were used to inoculate these guinea pigs intraperitoneally. Six of 10 guinea pigs aborted, and the organism was cultured from fetal tissues of 5 guinea pigs. Isolates from 3 of the 6 guinea pigs were propagated through 4 passages on blood agar and used to infect 3 groups, each of 5 guinea pigs. A 4th group of 5 guinea pigs was inoculated with the original culture. Three of 5 animals in the first 3 groups, which had been given the low-passage cultures from the preliminary trial, and 2 of 5 guinea pigs in the 4th group, which had been given the original culture, aborted. Antibody against the spirillum was detected in 19 of 30 inoculated guinea pigs. The major microscopic lesions were acute suppurative placentitis and splenitis. This bacterium retained pathogenic properties sufficient to cause infection, abortion, and microscopic lesions in two-thirds of the guinea pigs, in spite of high in vitro passage. The organism has unique ultrastructures, and its genus and species are yet to be determined.

Spiral bacterium associated with gastric, ileal and caecal mucosa of mice.

A spiral shaped bacterium was seen in smears and histological sections (stained by carbolfuchsin) of gastric, ileal and caecal mucosa as well as in stool smears from mice. A significant correlation between the presence of the spiral bacterium and the occurrence of gastritis was observed but the ileal and caecal mucosa seemed unaffected. The bacterium was Gram negative and grew on BHM and Skirrow's medium, under microaerophilic conditions, at 37 degrees C. Its major biochemical characteristics included positive catalase and oxidase reactions and a rapidly positive urease test. There were 2 or 3 spiral turns per cell and a tuft of up to 12 sheathed flagella on each pointed end. Entwined, braided periplasmic fibrils covered the surface of the cell. This spiral bacterium seemed to be part of the normal intestinal flora but was associated with gastritis.

[Human chronic gastritis associated with non-Helicobacter pylori spiral organisms (Gastrospirillum hominis). Four cases and review of the literature]. / Gastrite chronique associée chez l'homme à la présence de germes spiralés non-Helicobacter pylori (Gastrospirillum hominis). Présentation de 4 cas avec revue de la littérature.

Four cases of human active chronic gastritis associated with Gastrospirillum hominis, a recently described spiral shaped organism are presented. These 4 cases originated from a series of 1976 consecutive gastric biopsies, i.e. a prevalence of 0.25 percent in our material, are compared with Helicobacter pylori prevalence of 45 percent. Histopathological findings were chronic active gastritis with mild or no atrophy. Electron microscopy showed spiral bacteria with terminal flagellae, identical to those previously described in the literature. These bacteria have not yet been cultured; similar organisms are found in many animal species, and it seems that they do not provoke gastric inflammation. Gastrospirillum hominis could be responsible for cases of Helicobacter pylori negative chronic gastritis in man, but its pathogenicity remains to be demonstrated.

[Sodoku--a case report]. / Sodoku--descrição de um caso.

Rat-bite fever results from an infection with the organism Spirillum minus. The authors report a symptomatic patient with fever, malaise, and ulcer in forehead after a rodent exposure. The organism was identified in darkfield examination of the ulcer exudate. Penicillin was the drug used with clinical improvement of symptoms.

[Isolation of atmospheric-nitrogen-fixing spirilla from the waters of the Paraná delta and other rivers]. / Aislamiento de espirilos fijadores de nitrógeno atmósferico de las aguas del delta del Paraná y otros ríos.

Seventeen strains of Spirillum-like organisms (2) were isolated from the Parana Delta and other rivers (Table 1), using the following medium: 0.5% malic acid; 0.4% KOH; 0.5% K2HPO4; 0.005% yeast extract; pH was adjusted to 7 with KOH and 0.15% agar. The organisms produced a white, dense and sub-superficial pellicle in this medium, and streaking them on plates, single colonies could be isolated. They were easily recognized thanks to the Congo red added to the medium, because the bacteria, as it happens with Azospirillum species (6) (Spirillum lipoferum) (1), concentrated the strain. Using a colorimetric determination for ethylene (5), nitrogenase activity was detected in all the strains and in the NFb and potato-agar media the colonies were typical of Azospirillum (3). In PSS-semisolid and solid media, the growth was similar to that observed with Azospirillum strains. The isolated organisms developed in this medium better at 28 degrees C than at 37 degrees C, and the same was observed with 8 strains of Azospirillum isolated from plan roots of different species obtained from temperate regions. Some physiological tests were performed to identify the isolated organisms (Table 2). None of the isolated strains could be classified as a known species of Spirillum (4, 7), according to these tests. As thirteen strains showed properties in common with the terrestrial species A. brasilense and the others with A. lipoferum (8) they were tentatively identified as members of these species. Further studies are needed to ensure the classification of these strains and to determine the importance of these bacteria in the nitrogen balance of the waters from where they were isolated.

Isolation and characterization of a marine magnetotactic spirillum axenic culture QH-2 from an intertidal zone of the China Sea.

Magnetotactic bacteria (MTB) are ubiquitous in aquatic habitats. Because of their fastidious requirements for growth conditions, only very few axenic MTB cultures have been obtained worldwide. In this study, we report a novel marine magnetotactic spirillum axenic culture, designated as QH-2, isolated from the China Sea. It was able to grow in semi-solid or liquid chemically defined medium. The cells were amphitrichously flagellated and contained one single magnetosome chain with an average number of 16 magnetosomes per cell. Phosphate and lipid granules were also observed in the cells. Both rock magnetism and energy-dispersive X-ray spectroscopy characterizations indicated that the magnetosomes in QH-2 were single-domain magnetites (Fe(3)O(4)). QH-2 cells swam mostly in a straight line at a velocity of 20-50 microm/s and occasionally changed to a helical motion. Unlike other magnetotactic spirilla, QH-2 cells responded to light illumination. As a consequence of illumination, the cells changed the direction in which they swam from parallel to the magnetic field to antiparallel. This response appears to be similar to the effect of an increase in [O(2)]. Analysis of the QH-2 16S rRNA sequence showed that it had greater than 11% sequence divergence from freshwater magnetotactic spirilla. Thus, the marine QH-2 strain seems to be both phylogenetically and magnetotactically distinct from the freshwater Magnetospirillum spp. studied previously.

Proposal of Spirillum winogradskyi sp. nov., a novel microaerophilic species, an emended description of the genus Spirillum and Request for an Opinion regarding the status of the species Spirillum volutans Ehrenberg 1832.

A novel obligately organotrophic, facultatively microaerophilic spirillum, designated strain D-427(T), was isolated from sulfidic sludge of a municipal wastewater-treatment plant. Cells were Gram-negative, large and highly motile due to bipolar tufts of flagella covered with mucous sheaths. Coccoid cells were sometimes formed. Strain D-427(T) grew optimally at pH 7.5-7.8 and 28 degrees C in the presence of 2 % O(2) in the gas phase. The organism showed oxidase and very low catalase activity. The isolate grew chemo-organotrophically with a limited number of organic acids as substrates. The DNA G+C content was 38.0 mol% (T(m)). Phylogenetic analysis of the 16S rRNA gene sequence placed strain D-427(T) in the genus Spirillum within the class Betaproteobacteria. The 16S rRNA gene sequence similarity between strain D-427(T) and Spirillum volutans ATCC 19554(T), the type strain of the single species of the genus, was 98.6 %. The low level of DNA-DNA hybridization and different phenotypic properties indicate that strain D-427(T) is clearly distinguishable from Spirillum volutans. No strain of S. volutans is available from any established culture collection or from the authors who described this species. Therefore, on the basis of phenotypic and genotypic data and the fact that the type and single species of the genus Spirillum cannot be included in any scientific study, since the type strain has been lost, we propose to assign strain D-427(T) as a novel species of the genus Spirillum, Spirillum winogradskyi sp. nov. (type strain D-427(T) =DSM 12756(T) =VKM B-2518(T)), and we request that the Judicial Commission place the name Spirillum volutans on the list of rejected names if a suitable type strain is not found or a neotype is not proposed within 2 years following the publication of this paper. An emended description of the genus Spirillum is also provided.

[Two new species of microaerophilic sulfur spirilla, Spirillum winogradskii sp. nov. and Spirillum kriegii sp. nov].

New microaerophilic sulfur-oxidizing spirilla were isolated from hydrogen sulfide sludge of wastewater treatment plants. Strains D-427 and D-430 have spiral cells that are highly motile due to bipolar flagellum bundles covered with mucous sheaths. Under a phase-contrast microscope, these bundles are visible as single polar flagella. Spheroplasts are formed in the stationary growth phase. Both strains are obligate organotrophs able to oxidize a number of reduced sulfur compounds. The oxidation of sulfide and polysulfide leads to the formation of intracellular globules of elemental sulfur; thiosulfate oxidation results in tetrathionate accumulation in the medium. The cells are unable to utilize reduced sulfur compounds in the energy metabolism; their oxidation is caused by a chemical interaction with H2O2 and O2, synthesized in the electron transport chain. Both strains are obligate microaerophiles with an optimal oxygen concentration in the gas phase of 2 and 0.8% for strains D-427 and D-430, respectively. The strains utilize a limited number of organic acids as growth substrates, mainly tricarboxylic-acid-cycle intermediates. The DNA G+C content is 38.0 mol % (T(m)) for strain D-427 and 38.9 mol % for strain D-430. Phylogenetic analysis, based on the comparison of 16S rRNA gene sequences, revealed that the new isolates of sulfur spirilla are the most closely related to Spirillum volutans, the type species of the genus (97.4% similarity). They were assigned to the genus Spirillum within the class Beta-proteobacteria as two new species, S. winogradskii sp. nov. (D-427T = DSM 12756T) and S. kriegii sp. nov. (B-430T = BKM B-2372T). The emended description of the genus Spirillum is provided.

Bacterial diversity in the bacterioneuston (sea surface microlayer): the bacterioneuston through the looking glass.

The bacterioneuston is defined as the community of bacteria present within the neuston or sea surface microlayer. Bacteria within this layer were sampled using a membrane filter technique and bacterial diversity was compared with that in the underlying pelagic coastal seawater using molecular ecological techniques. 16S rRNA gene libraries of approximately 500 clones were constructed from both bacterioneuston and the pelagic water samples and representative clones from each library were sequenced for comparison of bacterial diversity. The bacterioneuston was found to have a significantly lower bacterial diversity than the pelagic seawater, with only nine clone types (ecotaxa) as opposed to 46 ecotaxa in the pelagic seawater library. Surprisingly, the bacterioneuston clone library was dominated by 16S rRNA gene sequences affiliated to two groups of organisms, Vibrio spp. which accounted for over 68% of clones and Pseudoalteromonas spp. accounting for 21% of the library. The dominance of these two 16S rRNA gene sequence types within the bacterioneuston clone library was confirmed in a subsequent gene probing experiment. 16S rRNA gene probes specific for these groups of bacteria were designed and used to probe new libraries of 1000 clones from both the bacterioneuston and pelagic seawater DNA samples. This revealed that 57% of clones from the bacterioneuston library hybridized to a Vibrio sp.-specific 16S rRNA gene probe and 32% hybridized to a Pseudoalteromonas sp.-specific 16S rRNA gene probe. In contrast, the pelagic seawater library resulted in only 13% and 8% of 16S rRNA gene clones hybridizing to the Vibrio sp. and Pseudoalteromonas sp. probes respectively. Results from this study suggest that the bacterioneuston contains a distinct population of bacteria and warrants further detailed study at the molecular level.

Occurrence of Azospirillum in Polish soils.

N2-fixing Azospirillum strains were isolated from garden soils and roots of wheat, rye, barley, oat and maize. The strains most active in C2H2 reduction were those isolated from garden soils, wheat and barley roots which produced 304 to 351 nmoles of C2H4 X h-1 per culture. The results demonstrate the presence of Azospirillum in Polish soils.

Titanospirillum velox: a huge, speedy, sulfur-storing spirillum from Ebro Delta microbial mats.

A long (20-30 micrometer), wide (3-5 micrometer) microbial-mat bacterium from the Ebro Delta (Tarragona, Spain) was grown in mixed culture and videographed live. Intracellular elemental sulfur globules and unique cell termini were observed in scanning-electron-microprobe and transmission-electron micrographs. A polar organelle underlies bundles of greater than 60 flagella at each indented terminus. These Gram-negative bacteria bend, flex, and swim in a spiral fashion; they translate at speeds greater than 10 body lengths per second. The large size of the spirillum permits direct observation of cell motility in single individual bacteria. After desiccation (i.e., absence of standing water for at least 24 h), large populations developed in mat samples remoistened with sea water. Ultrastructural observations reveal abundant large sulfur globules irregularly distributed in the cytoplasm. A multilayered cell wall, pliable and elastic yet rigid, distends around the sulfur globules. Details of the wall, multiflagellated termini, and large cytoplasmic sulfur globules indicate that these fast-moving spirilla are distinctive enough to warrant a genus and species designation: Titanospirillum velox genus nov., sp. nov. The same collection techniques at a similar habitat in the United States (Plum Island, northeast Essex County, Massachusetts) also yielded large populations of the bacterium among purple phototrophic and other inhabitants of sulfurous microbial-mat muds. The months-long survival of T. velox from Spain and from the United States in closed jars filled with mud taken from both localities leads us to infer that this large spirillum has a cosmopolitan distribution.

Isolation of Azospirullum from diverse geographic regions.

We have isolated Azospirillum (Spirullum lipoferum) from roots of grasses of several genera collected from a number of tropical and subtropical-temperate locations. Pure cultures were obtained from a small percentage of samples; no higher percentage was secured from tropical than from other grasses. Acetylene reduction and distinctive growth in N-free soft agar deeps were inadequate to identify this genus, although helpful in initial screening. Fluorescent antibody tests with antiserum against characterized strains were helpful. There is some evidence that this genus of bacteria may be favored in the rhizoplane.