A. Ramos-Cormenzana

Numerical taxonomy of moderately halophilic Gram-negative rods

Summary: A study was made of 516 randomly selected isolates of moderately halophilic bacteria from solar salterns showing salinities between 8.8 and 40.0% (w/v) total salts, located in S.E. Spain. After purification, many cytological, physiological, biochemical, nutritional and antibiotic sensitivity characters were determined for 106 selected saltern isolates and two reference strains. Data were coded and analysed by numerical techniques using the Jaccard coefficient (S j), and clusters of strains were obtained by average linkage (UPGMA) analysis. Nine major phenons were found at the 72.5% similarity level. The properties of each phenon are given, their taxonomic affinities are discussed, and typical reference strains are suggested. Almost all the strains were related to genera known to contain marine species. A large number of the strains could be tentatively assigned to the genus Vibrio, suggesting that this may be an abundant taxon of moderately halophilic Gram-negative rods in solar salterns.

Deleya halophila, a New Species of Moderately Halophilic Bacteria

Deleya halophila sp. nov., which was isolated from hypersaline soils, is proposed. Each of 38 isolates, together with 8 reference strains, was examined in 97 phenotypic tests, and the data were analyzed by numerical taxonomy procedures. The 38 isolates formed a distinct group at a 76% similarity value, as determined by an analysis performed with the simple matching coefficient and unweighted average linkage clustering. Characteristically, the isolates were aerobic, gram-negative rods that were motile by one to eight peritrichous flagella. They grew optimally in the presence of 7.5% (wt/vol) marine salts. The distinguishing features of the new species are its salt requirement, biochemical features, and nutritional versatility. The guanine-plus-cytosine content of the deoxyribonucleic acid is 66.7 mol%. The type strain of this species is strain F5–7 (= CCM 3662).

Characteristics of the heterotrophic bacterial populations in hypersaline environments of different salt concentrations.

Solar salterns, based on a multi-pond system, give a discontinuous gradient of salt concentrations. The heterotrophic bacterial populations of ponds containing from 10% salt to saturation have been studied. Saltern samples were spread on agar plates containing different media for halophilic bacteria and one medium made with water of the pond plus nutrients. Replica plating was done to determine the salt range for growth of the colonies. We studied 150 strains to determine the salt spectra of growth, the morphology, and nutrient requirements. The following conclusions were reached: (a) In salt concentrations above 10% (total salts), most bacteria are halophilic and few are halotolerant; (b) the two types of halophilic bacteria, moderate and extreme, show different distributions; in these ponds a narrow overlap exists between 25% and 32% salts with moderate halophiles predominating below this interval and extreme halophiles above it; (c) the populations of moderate halophiles are highly heterogeneous, and the salt concentration of their habitat affects their taxonomic composition, salt range for growth, and nutrient requirements. The population composition of extreme halophiles is less affected by the salt concentrations at which these bacteria are found.

Bioremediation and biovalorisation of olive-mill wastes

Olive-mill wastes are produced by the industry of olive oil production, which is a very important economic activity, particularly for Spain, Italy and Greece, leading to a large environmental problem of current concern in the Mediterranean basin. There is as yet no accepted treatment method for all the wastes generated during olive oil production, mainly due to technical and economical limitations but also the scattered nature of olive mills across the Mediterranean basin. The production of virgin olive oil is expanding worldwide, which will lead to even larger amounts of olive-mill waste, unless new treatment and valorisation technologies are devised. These are encouraged by the trend of current environmental policies, which favour protocols that include valorisation of the waste. This makes biological treatments of particular interest. Thus, research into different biodegradation options for olive-mill wastes and the development of new bioremediation technologies and/or strategies, as well as the valorisation of microbial biotechnology, are all currently needed. This review, whilst presenting a general overview, focusses critically on the most significant recent advances in the various types of biological treatments, the bioremediation technology most commonly applied and the valorisation options, which together will form the pillar for future developments within this field.

Studies on antibacterial activity of waste waters from olive oil mills (alpechin): Inhibitory activity of phenolic and fatty acids

Abstract In order to have a clearer understanding of the nature of the substances responsible for the antibacterial activity of alpechin, the inhibitory effect of the phenolic acids found in the alpechin and of the fatty acids present in olive oil were investigated. The antibacterial activity of the phenolic acids (tested separately or in mixtures, at the stated concentrations) when they are tested against Bacillus megaterium ATCC 33085 and against a collection of bacteria isolated from unpolluted soil and alpechin polluted soil, does not coincide with the inhibitory effect of the alpechin. On the other hand, although antibacterial activity has not been detected in olive oil, linoleic, oleic, linolenic, lauric and myristic acids are capable of inhibiting the growth of Bacillus megaterium ATCC 33085.

Biomineralization of carbonates by Halomonas eurihalina in solid and liquid media with different salinities : crystal formation sequence

Carbonate precipitation by 20 strains of the moderately halophilic species Halomonas eurihalina in both solid and liquid media was studied. The influence of salinity and temperature on the quantity and type of crystals precipitated was also investigated. Some strains of H. eurihalina formed crystals in all conditions tested. The mineral phases precipitated were magnesium calcite, aragonite and monohydrocalcite in variable proportions depending on various factors such as the type of growth medium employed and its salinity. Scanning electron microscopy and X-ray dispersive energy microanalysis were used to investigate the crystal formation sequence. The process of biolith formation was sequential. It started with chains or filaments of bacteria, giving way to discs which finally produced spherical forms of approximately 50 microns in diameter. We suggest a mechanism of carbonate crystal formation by H. eurihalina.

Biomineralization of carbonates by Halobacillus trueperi in solid and liquid media with different salinities.

We investigated the precipitation of carbonates by Halobacillus trueperi in both solid and liquid media at different salt concentrations and different magnesium/calcium ratios. H. trueperi precipitated at all assayed salt concentrations. When salt concentration increased, the quantity and the size of bioliths decreased and the time required increased. The precipitated minerals (determined by X-ray diffraction) were calcite, magnesium calcite and monohydrocalcite in variable proportions depending on the salinity and the physical state of the medium; the magnesium content of the magnesium calcites also varied with regard to the culture type. According to the saturation indices other minerals could also precipitate. Scanning electron microscopy showed that dominant morphologies of the bioliths were spherulitic with fibrous radiated interiors. We show that H. trueperi plays an active role in the precipitation of carbonates and we hypothesize about this process of biomineralization.

Characteristics of soil after pollution with waste waters from olive oil extraction plants

Abstract Higher levels of soil salinity due to potassium and sodium replacement of soil cations were detected in an alkaline soil after pollution with waste waters from olive oil extraction plants. The pH was practically unchanged and soil C/N ratio was increased. A reversible decrease in the count of sporulated bacteria was noted.

Numerical Taxonomy of Moderately Halophilic Gram-negative Bacteria from Hypersaline Soils

SUMMARY: A total of 132 moderately halophilic bacteria were isolated from hypersaline soils with a Cl- content between 2.36 and 12.72% (w/v) located near Alicante (S.E. Spain) and examined for 98 phenotypic characteristics including their response to cytological, physiological, biochemical and nutritional tests. They were submitted to a numerical analysis together with six reference strains using both simple matching (SSM ) and Jaccard (SJ ) coefficients, and cluster analysis was carried out by the unweighted pair group method of association (UPGMA), single linkage and complete linkage. With the SJ coefficient and UPGMA clustering, eight phenons were obtained at the 65% similarity level. From each phenon representative strains were chosen for the determination of DNA base composition and for electron microscopy. Bacteria belonging to phenons D, E, and F were assigned to the genus Alcaligenes. Phenon G included 27 strains assigned to Acinetobacter, but the high G + C composition (58.9 mol %) of a representative strain of this phenon suggests that it may represent a new taxon. Phenons A, B, and C were designated Flavobacterium and phenon H was Pseudomonas. The bacteria found in these environments are not related to those from hypersaline waters or normal soils.

Volcaniella eurihalina gen. nov., sp. nov., a moderately halophilic nonmotile gram-negative rod

A comparison of 16 gram-negative moderately halophilic aerobic rod-shaped bacteria with other halophilic and nonhalophilic gram-negative bacteria supported the establishment of Volcaniella eurihalina gen. nov., sp. nov. This comparison included phenotypic properties, salt requirements, and guanine-plus-cytosine contents of the DNAs, as well as DNA-DNA homology studies. The distinguishing features of this new bacterial genus are as follows: The organisms are nonmotile short rods that are oxidase negative; they are aerobic with a strictly respiratory type of metabolism; they are moderate halophiles, optimal growth occurs at a total salt concentration of 7.5% (wt/vol), and they exhibit a strongly euryhaline character; and they have a specific requirement for Na+ ions (sodium can be supplied as NaCl, Na2SO4, or NaBr). The minimum NaCl concentration required is 1.5% (wt/vol). The guanine-plus-cytosine content of the DNA is 59.1 to 65.7 mol%. This organism was isolated from hypersaline habitats, including saline soils and salt ponds, and from seawater. The type strain is strain F9-6 (= ATCC 49336).