Disturbance of primary producer communities disrupts the thermal limits of the associated aquatic fauna.

Environmental fluctuation forms a framework of variability within which species have evolved. Environmental fluctuation includes predictability, such as diel cycles of aquatic oxygen fluctuation driven by primary producers. Oxygen availability and fluctuation shape the physiological responses of aquatic animals to warming, so that, in theory, oxygen fluctuation could influence their thermal ecology. We describe annual oxygen variability in agricultural drainage channels and show that disruption of oxygen fluctuation through dredging of plants reduces the thermal tolerance of freshwater animals. We compared the temperature responses of snails, amphipods, leeches and mussels exposed to either natural oxygen fluctuation or constant oxygen in situ under different acclimation periods. Oxygen saturation in channel water ranged from c. 0 % saturation at night to >300 % during the day. Temperature showed normal seasonal variation and was almost synchronous with daily oxygen fluctuation. A dredging event in 2020 dramatically reduced dissolved oxygen variability and the correlation between oxygen and temperature was lost. The tolerance of invertebrates to thermal stress was significantly lower when natural fluctuation in oxygen availability was reduced and decoupled from temperature. This highlights the importance of natural cycles of variability and the need to include finer scale effects, including indirect biological effects, in modelling the ecosystem-level consequences of climate change. Furthermore, restoration and management of primary producers in aquatic habitats could be important to improve the thermal protection of aquatic invertebrates and their resistance to environmental variation imposed by climate change.

Modified niche optima and breadths explain the historical contingency of bacterial community responses to eutrophication in coastal sediments.

Previous studies have shown that the response of bacterial communities to disturbances depends on their environmental history. Historically fluctuating habitats host communities that respond better to disturbance than communities of historically stable habitats. However, the exact ecological mechanism that drives this dependency remains unknown. Here, we experimentally demonstrate that modifications of niche optima and niche breadths of the community members are driving this dependency of bacterial responses to past environmental conditions. First, we develop a novel, simple method to calculate the niche optima and breadths of bacterial taxa regarding single environmental gradients. Then, we test this method on sediment bacterial communities of three habitats, one historically stable and less loaded and two historically more variable and more loaded habitats in terms of historical chlorophyll-α water concentration, that we subject to hypoxia via organic matter addition ex situ. We find that communities containing bacterial taxa differently adapted to hypoxia show different structural and functional responses, depending on the sediment's environmental history. Specifically, in the historically less fluctuating and loaded sediments where we find more taxa poorly adapted to hypoxic conditions, communities change a lot over time and organic matter is not degraded efficiently. The opposite is true for the historically more fluctuating and loaded sediments where we find more taxa well adapted to hypoxia. Based on the community responses observed here, we also propose an alternative calculation of community resistance that takes into account how rapidly the communities respond to disturbances and not just the initial and final states of the community.

Allochthonous bioaugmentation in ex situ treatment of crude oil-polluted sediments in the presence of an effective degrading indigenous microbiome.

Oil-polluted sediment bioremediation depends on both physicochemical and biological parameters, but the effect of the latter cannot be evaluated without the optimization of the former. We aimed in optimizing the physicochemical parameters related to biodegradation by applying an ex-situ landfarming set-up combined with biostimulation to oil-polluted sediment, in order to determine the added effect of bioaugmentation by four allochthonous oil-degrading bacterial consortia in relation to the degradation efficiency of the indigenous community. We monitored hydrocarbon degradation, sediment ecotoxicity and hydrolytic activity, bacterial population sizes and bacterial community dynamics, characterizing the dominant taxa through time and at each treatment. We observed no significant differences in total degradation, but increased ecotoxicity between the different treatments receiving both biostimulation and bioaugmentation and the biostimulated-only control. Moreover, the added allochthonous bacteria quickly perished and were rarely detected, their addition inducing minimal shifts in community structure although it altered the distribution of the residual hydrocarbons in two treatments. Therefore, we concluded that biodegradation was mostly performed by the autochthonous populations while bioaugmentation, in contrast to biostimulation, did not enhance the remediation process. Our results indicate that when environmental conditions are optimized, the indigenous microbiome at a polluted site will likely outperform any allochthonous consortium.

Do mosquito-associated bacteria of the genus Asaia circulate in humans?

Symbiotic bacteria of the genus Asaia have been proposed as tools for control of mosquito-borne diseases, specifically malaria. However, safety issues are a major concern for paratransgenesis strategies. The aim of this study is to investigate, with immunofluorescence assays and quantitative PCR experiments, whether Asaia spp. is circulating among humans. All human sera and whole blood samples analyzed were negative for Asaia spp., thus suggesting that this organism could be utilized, in the future, as a malaria control tool.

Esterase as an enzymatic signature of Geodermatophilaceae adaptability to Sahara desert stones and monuments.

AIM: To assess esterase profiling of members of Geodermatophilaceae isolated from desert stones and monuments in Tunisia and Egypt. METHODS AND RESULTS: Members of Geodermatophilaceae family isolated from desert stones and monuments in Tunisia and Egypt were characterized by partial 16S rRNA sequences. Twenty-five strains were clustered in three dissimilar groups of the genera Geodermatophilus (12 strains), Blastococcus (5 strains) and Modestobacter (3 strains). Isolates were also screened and typed based on major groups of esterase hydrolytic activity. Their esterase patterns were determined and compared to those of ten reference strains belonging to Geodermatophilaceae family. Strains exhibited a diverse and complex pattern of electrophoretic esterase bands, and 31 haplotypes were obtained for the 35 investigated strains. Esterases produced by members of Geodermatophilaceae family have an optimal activity around 40 degrees C and at pH 8. Esterases from Geodermatophilus strains display a high resistance to thermal inactivation and alkaline pH and retaining 30 and 20% of activity after heating for 20 min at 120 degrees C and at pH 12, respectively, and were completely inactivated after 30 min at 120 degrees C. Enzyme activity has been strongly activated in the presence of Ca(2+)and Mg(2+) ions and moderately by Zn(2+) and was markedly inhibited by Cu(2+) and Co(2+) ions. CONCLUSIONS: Geodermatophilaceae isolates share a rich and particular pool of esterase activities that could be directly linked to harsh conditions characterizing their ecological habitat including high level of aridity, temperature, ionic strength and low nutrient availability. SIGNIFICANCE AND IMPACT OF THE STUDY: Esterase could be considered as enzymatic signature that outlines adaptability of Geodermatophilaceae in arid area.

Multiple symbiosis in the leafhopper Scaphoideus titanus (Hemiptera: Cicadellidae): details of transovarial transmission of Cardinium sp. and yeast-like endosymbionts.

Scaphoideus titanus is the insect vector of flavescence dorée (FD), a yellow disease of grapevines. Observations on adult females and nymphs of S. titanus showed that this insect is associated with a complex microbial community. Ultrastructural analysis showed that the fat body, salivary glands and ovary of the insect harbour microorganisms showing the brush-like structure typically observed in the genus Cardinium. In particular, it has been shown that these symbiotic bacteria are present both in the follicular cells and in the eggs. In addition, cells resembling bacteriocytes, harbouring numerous Cardinium symbionts in the cytoplasm, were observed in the apical portion of the ovary in adult females. These cells are likely responsible for bacterial transmission to the ovary. Optical microscopy showed that the fat body harbours an enormous population of yeast-like symbionts (YLSs). Ultrastructural observations showed that these symbionts are enclosed within specialized cells of the fat body and are also present in the ovary, where they are found in both the follicular cells and the eggs. There is thus evidence that both Cardinium and the YLSs are transovarially transmitted to the offspring. To our knowledge, S. titanus is the sole insect known to transmit two different kinds of symbionts to the eggs, a prokaryote and an eukaryote. Gene sequence analysis and in situ hybridization led to the identification of YLSs as members of the class Sordariomycetes (=Pyrenomycetes). Finally, ultrastructural observation of the midgut content revealed the presence, in both adult females and nymphs, of a complex microbial community, which include a phytoplasma-like microorganism, likely the agent of FD.

Heteroduplex structures in 16S-23S rRNA intergenic transcribed spacer PCR products reveal ribosomal interoperonic polymorphisms within single Frankia strains.

AIMS: Detection of polymorphisms in intergenic transcribed spacer (ITS) 16S-23S rRNA within single Frankia strains. METHODS AND RESULTS: Polymorphisms in the 16S-23S rRNA ITS were investigated in single-colony subcultures of seven Frankia isolates. Multiple ITS-polymerase chain reaction (PCR) bands were detected solely in isolates BMG5.5 and BMG5.11. The slow-migrating bands in the ITS-PCR agarose gel electrophoresis profiles of the isolates were revealed to be heteroduplexes on the basis of their migration shift in different electrophoretic matrices, southern hybridization and the single-strand DNA mung bean endonuclease digestion. Laser-scanned capillary electrophoresis detected two ITS-PCR fragments differing in length by three and six nucleotide insertions/deletions in strains BMG5.5 and BMG5.11, respectively. Sequence analysis of the cloned ITS showed that in strain BMG5.5 the two ITS differed by the presence of three to four copies of the 3-bp tandem repeat 5'-TGG-3'. In strain BMG5.11, the two ITS differed by the presence of two to three copies of the 6-bp tandem repeat 5'-CTTGGG-3'. CONCLUSIONS: We demonstrate the occurrence of ITS 16S-23S rRNa polymorphisms within single Frankia strains. SIGNIFICANCE AND IMPACT OF THE STUDY: We reported the occurrence of ITS 16S-23S rRNA polymorphisms within single Frankia strains from Elaeagnus host group recognized as the more flexible strains within Frankia genus. Furthermore, we underscored the applied interest of strains BMG5.11 and BMG5.5 in future ecological studies using ITS 16S-23S rRNA as molecular marker.

A novel phenotype based on esterase electrophoretic polymorphism for the differentiation of Lactococcus lactis ssp. lactis and cremoris.

AIMS: To evaluate the esterase phenotype in Lactococcus lactis strains isolated from traditional Tunisian dairy products. METHODS AND RESULTS: A collection of 55 L. lactis strains isolated from traditional fermented milk products and three reference strains were identified at species and subspecies level using molecular methods targeted to the 16S rRNA gene and to the histidine operon. The genotypic data obtained allowed the identification of the strains as L. lactis ssp. lactis and L. lactis ssp. cremoris with the prevalence of the ssp. lactis. The phenotypic identification based on arginine hydrolysis, the growth at 40 degrees C and in presence of 4% NaCl showed several discrepancy with the identification data based on genotypic analysis. Additional experiments carried out evaluating the esterase electrophoretic patterns revealed four classes of esterases identified on the basis of their electrophoretic mobility and specific activity on alpha- and beta-naphthyl ester of acetate and propionate. Esterase profiles discriminated the strains in two main groups corresponding to the subspecies cremoris and lactis according to a DNA-based identification. CONCLUSIONS: The evaluation of esterase activity represents a novel phenotype for the taxonomic discrimination of the L. lactis ssp. lactis and cremoris. SIGNIFICANCE AND IMPACT OF THE STUDY: Besides the DNA-based techniques that allow the rapid and accurate species/subspecies identification, the electrophoretic esterase profiles of L. lactis strains represents: (i) a new phenotypic tool to understand the physiology and the ecology of this species; and (ii) a new test for the potential selection of flavour producing strains.

Ultrastructure of a novel Cardinium sp. symbiont in Scaphoideus titanus (Hemiptera: Cicadellidae).

An ultrastructural study of the novel symbiont Cardinium sp. was performed with particular attention to the description of the structure and organization of highly elaborated cytoplasmic complexes containing microtubule-like elements (MLC). Three major components were observed. The first was a system of microtubule-like elements (ML) arranged in parallel array extending from the plasma membrane into the cytosol of the bacterium. The second, an fibrous electrondense plaque (FEP), approximately 8 nm thick, located 7.5 nm away from the plasma membrane and parallel to it. The third component, not previously reported, was described for the first time in this paper. This consisted of a set of regularly distributed 8 nm electron-dense structures (ES), with a center-to-center spacing of about 12 nm, adhering to the outer leaflet of the plasma membrane. Often, the ES created a close connection between the plasma membrane and the outer membrane, so that in this area they became straight and stiff. The first and second component of these structures are compared to previously described microtubules and microfilaments.

The autolytic phenotype of the Bacillus cereus group.

AIM: To determine the autolytic phenotype of five species in the Bacillus cereus group. METHODS AND RESULTS: The autolytic rate of 96 strains belonging to five species in the B. cereus group was examined under starvation conditions at pH 6, 6.5 and 8.5 in different buffers. The autolytic rate was strain-dependent with a wide variability at pH 6, but higher and more uniform at pH 6.5. At pH 8.5, and respect to the extent of autolysis at pH 6.5, it was relatively low for most of the strains with the lowest values between 13 and 52% in Bacillus mycoides and Bacillus pseudomycoides. Peptidoglycan hydrolase patterns evaluated by renaturing sodium dodecyl sulfate-polyacrylamide gel electrophoresis using cells of Bacillus thuringiensis ssp. tolworthi HD125 as an indicator, revealed complex profiles with lytic bands of about 90, 63, 46, 41, 38, 32, 28 and 25 kDa in B. cereus, B. thuringiensis and Bacillus weihenstephanensis. Bacillus mycoides and B. pseudomycoides had simpler profiles with lytic bands of 63, 46 and 38 kDa. Changes in the autolytic pattern were observed for cells harvested at the stationary phase of growth (72 h) showing an increase in the intensity of the 25 kDa band in the case of B. cereus, B. thuringiensis and B. weihenstephanensis, while no changes were observed for B. mycoides. Using Micrococcus lysodeicticus and Listeria monocytogenes as indicators lytic activity was retained by proteins of 63, 46, 38, 32 and 25 kDa and a new one of about 20 kDa in B. mycoides. Growth in the different media did not affect the autolytic pattern. NaCl abolished the activity of all the peptidoglycan hydrolases except for those of B. mycoides and B. weihenstephanensis. Lytic activity was retained in the presence of MgCl(2), MnCl(2) and EDTA and increased at basic pH. CONCLUSIONS: Bacillus cereus/B. thuringiensis/B. weihenstephanensis showed a high extent of autolysis around neutral pH, even though they presented relatively complex autolysin profiles at alkaline pH. Bacillus mycoides/B. pseudomycoides had a higher extent of autolysis at acidic pH and a simpler autolysin pattern. SIGNIFICANCE AND IMPACT OF THE STUDY: Information on the autolytic phenotype expand the phenotypic characterization of the different species in the B. cereus group.

The autolytic phenotype of Bacillus thuringiensis.

AIMS: To evaluate the autolytic phenotype of Bacillus thuringiensis. METHODS AND RESULTS: The autolytic rate of 87 strains belonging to different subsp. of B. thuringiensis was examined at pH 6, 6.5 and 8.5 in different buffers under starvation conditions. At pH 6 the extent of autolysis (average in the strain collection 38.3 +/- 21.1) was strain-dependent with wide variability, while at pH 6.5 and 8.5 (averages 72.0 +/- 9.0 and 63.1 +/- 8.2, respectively) it was much more uniform with only a few strains showing low autolytic rates. Forty-one per cent of the strains showed high resistance (>/=80%) to mutanolysin, a commercial muramidase from Streptomyces. The peptidoglycan hydrolase pattern was evaluated by renaturing SDS-PAGE using cells of B. thuringiensis subsp. tolworthi HD125 as indicator. The strain collection showed seven major lytic bands of about 90, 63, 46, 38, 32, 28 and 25 kDa, and in the stationary growth phase (72 h) there was a more intense 25 kDa band in the autolytic pattern. Using Micrococcus lysodeicticus and Listeria monocytogenes as the indicators lytic activity was retained, as seen by the bands of 63, 46, 38, 32 and 25 kDa. Growth in the different media did not affect the autolytic pattern. NaCl abolished the activity of all the peptidoglycan hydrolases in the gel, but in the presence of KCl, MgCl(2), MnCl(2) and EDTA some activity was retained. At basic pH the lytic activity increased. CONCLUSIONS: The autolytic phenotype of B. thuringiensis was found to be strain-dependent, and different proteins exibited peptidoglycan hydrolase activity, particularly at alkaline pH. Several of these proteins retained lytic activity against other bacterial species. SIGNIFICANCE AND IMPACT OF THE STUDY: The characterisation of the autolytic phenotype of B. thuringiensis should expand the prospects of using this species in bacterial bio-control and field applications.

Characterization of urease genes cluster of Streptococcus thermophilus.

AIMS: The milk acidification rate of Streptococcus thermophilus strains can be affected by several factors, one of which is the hydrolysis of urea by the urease complex. To evaluate the technological suitability of S. thermophilus strains deprived of urease activity in milk fermentation, the genetic cluster related to urease enzymatic activity has been characterized in the type strain DSM 20167T. METHODS AND RESULTS: Amplification of the urease genes of S. thermophilus DSM 20167T was developed on the basis of the urease gene cluster of the phylogenetically related S. salivarius. Nucleotide sequencing revealed the presence of eight open reading frames, which were most homologous to ureABC (structural genes) and ureI, ureEFGD (accessory genes) of S. salivarius and other ureolytic bacteria. Reverse transcriptase PCR experiments were in agreement with an operon organization for the eight genes (ureIABCEFGD). A food grade mutant A16 (DeltaureC3) with a 693 bp in-frame deletion in ureC gene exhibited a urease negative (Ure-) phenotype. Unlike the wild-type strain, the acidification rate of the mutant in reconstituted skimmed milk was not affected by the presence of urea or nickel ions. A small-scale yoghurt fermentation trials were carried out using the wild-type or the Ure- mutant A16 (DeltaureC3) in co-culture with Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842 in presence of urea. The result obtained underlines that when the Ure- mutant was used as a co-starter the acidification rate was higher than that obtained using the wild-type strain. CONCLUSIONS: The study provides the first genetic characterization and the technological implication of S. thermophilus DSM 20617T urease activity. SIGNIFICANCE AND IMPACT OF THE STUDY: The detrimental effect of ureolytic activity on the rate of milk acidification was evaluated and superseded using a food-grade Ure- recombinant strain. Small-scale yoghurt production trials highlighted the positive role of a Ure-S. thermophilus mutant as a co-starter in milk fermentations. Moreover, the vector pMI108 developed for the construction of the Ure- strain, should be considered as a potential tool for the generation of Ure- dairy S. thermophilus strains selected for other relevant technological properties but characterized by the undesirable ureolytic phenotype.

Detection and characterization of the novel bacteriocin entomocin 9, and safety evaluation of its producer, Bacillus thuringiensis ssp. entomocidus HD9.

AIMS: To identify and characterize new bacteriocins from a collection of 41 strains belonging to 27 subspecies of Bacillus thuringiensis, and to evaluate the safety of the producers. METHODS AND RESULTS: Bacillus thuringiensis ssp. entomocidus HD9 produced in the culture supernatant an antimicrobial activity against Gram-positive bacteria including Listeria monocytogenes, one of four pathogenic Pseudomonas aeruginosa and several fungi. Production of the antibacterial activity, named entomocin 9, started during mid-logarithmic growth reaching its maximum at the early stationary phase. Entomocin 9 retained more than 72% of activity after incubation for 20 min at 121 degrees C. Activity was lost after proteinase K treatment, it was stable in a pH range between 3 and 9, and resistant to lyophilization. After partial purification with ammonium sulphate precipitation followed by gel-filtration and anion-exchange chromatography, an active protein of ca 12.4 kDa was isolated. The mode of action of entomocin 9 was bactericidal and caused cell lysis of growing cells. Despite the presence of a range of virulence related genes, including haemolysin BL, nonhaemolytic enterotoxin, cytotoxin K and several hydrolytic activities, B. thuringiensis HD9 was not toxic against Vero cells. CONCLUSIONS: Entomocin 9 is a novel heat-stable, bacteriocin produced by B. thuringiensis HD9. The absence of toxicity against Vero cells suggests the suitability of strain HD9 for a safe application in antimicrobial treatments. SIGNIFICANCE AND IMPACT OF THE STUDY: New finding on entomocin 9 would make B. thuringiensis attractive in biotechnological applications as an antimicrobial agent in agriculture and food industry.

Genetic relationship in the 'Bacillus cereus group' by rep-PCR fingerprinting and sequencing of a Bacillus anthracis-specific rep-PCR fragment.

AIMS: To evaluate the genetic relationship in the Bacillus cereus group by rep-PCR fingerprinting. METHODS AND RESULTS: A collection of 112 strains of the six species of the B. cereus group was analysed by rep-PCR fingerprinting using the BOX-A1R primer. A relative genetic distinctness was found among the species. Cluster analysis of the rep-PCR patterns showed clusters of B. thuringiensis strains quite separate from those of B. cereus strains. The B. anthracis strains represented an independent lineage in a B. cereus cluster. The B. mycoides, B. pseudomycoides and B. weihenstephanensis strains were clustered into three groups at some distance from the other species. Comparison of sequences of AC-390, a typical B. anthracis rep-PCR fragment, from 27 strains of B. anthracis, B. cereus, B. thuringiensis and B. weihenstephanensis, representative of different clusters identified by rep-PCR fingerprinting, confirmed that B. anthracis diverges from its related species. CONCLUSIONS: The genetic relationship deduced from the rep-PCR patterns indicates a relatively clear separation of the six species, suggesting that they can indeed be considered as separate units. SIGNIFICANCE AND IMPACT OF THE STUDY: rep-PCR fingerprinting can make a contribution in the clarification of the genetic relationships between the species of the B. cereus group.

Response of bacterial community during bioremediation of an oil-polluted soil.

AIM: To study the response of the bacterial community to bioremediation of a soil with an aged contamination of crude oil. METHODS AND RESULTS: The bacterial community in laboratory soil columns during a 72-day biostimulation treatment was followed by analysing the number of total cultivable hydrocarbon-degrading bacteria, soil respiratory activity and the 16S-23S rDNA internal transcribed spacer homoduplex heteroduplex polymorphisms (ITS-HHP) of total soil bacterial DNA. ITS-HHP permits an estimate of both length and sequence polymorphism in a 16S-23S rDNA spacer population, using to advantage the homoduplex and heteroduplex fragments that are generated during PCR. The treatment, made by air sparging and biostimulation with a mineral nutrient and surfactant solution, resulted in a 39.5% decrease of the total hydrocarbon content. Within 4 days of treatment onset the bacterial community underwent a first phase of activation that led to a substantial increase in the observable diversity. Subsequently, after a 12-day period of stability, another activation phase was observed with further shifts of the community structure and an increase in the abundance and diversity of catechol-2,3-dioxygenase (C23O) genes. CONCLUSIONS: The overall data suggest an important contribution of uncultivable bacteria to the soil bioremediation, since, during the second activation phase, the increases of the respiratory activity, bacterial diversity and C23O gene abundance and diversity were not accompanied by a corresponding increase of the cultivable bacteria number. SIGNIFICANCE AND IMPACT OF THE STUDY: This study shows that successive phases of activation of bacterial populations occur during a bioremediation treatment of oil-polluted soil.

Characterization of a repetitive element polymorphism-polymerase chain reaction chromosomal marker that discriminates Bacillus anthracis from related species.

AIMS: To identify a chromosomal marker with signature nucleotides specific for Bacillus anthracis. METHODS AND RESULTS: Repetitive element polymorphism-polymerase chain reaction with BOX-A1R primer was used to discriminate 52 strains of all six species of the 'B. cereus group'. A B. anthracis signature fragment, named AC-390, was cloned and sequenced. The deduced amino acid sequence was homologous to that of YwfK of B. subtilis. Using two internal primers, the AC-390 fragment was sequenced from two other B. anthracis strains as well as from strains of B. cereus and B.thuringiensis which have an AC-390 fragment homologous to that of B. anthracis as shown by Southern hybridization experiments. CONCLUSIONS: Two new signature sequences specific for B. anthracis were identified on a chromosomal fragment homologous to YwfK, a transcriptional regulator of B. subtilis. SIGNIFICANCE AND IMPACT OF THE STUDY: These results show a new chromosomal DNA trait useful for distinguishing B. anthracis from the related species of the B. cereus group, regardless of the presence of the virulence plasmids pXO1 and pXO2.

Biodiversity of Geodermatophilaceae isolated from altered stones and monuments in the Mediterranean basin.

An investigation was made into the occurrence and biodiversity of Geodermatophilaceae on 78 samples of altered stone surfaces from 24 monuments and natural stones in the Mediterranean basin; it was found that the total microbial counts ranged between 0 and 10(7) cfu g(-1) dry weight. Members of the Geodermatophilaceae family were isolated from 22 of the 78 samples examined, with the incidence of Geodermatophilaceae colonies in the cultivable population ranging from 1% to 100%. The highest percentage was found in six samples of markedly deteriorated stone. Sixty-five strains randomly isolated from the plates were clustered in six different groups by amplified 16S rDNA restriction analysis (ARDRA) using five different restriction enzymes. Twenty-five strains, representing all the ARDRA haplotypes, were characterized further by partial sequencing (350-550 bp) of the 16S rDNA and by analysing 76 morphological, metabolic and physiological properties. The strains were associated with three well-separated clusters of the genera Geodermatophilus, Blastococcus and Modestobacter. On the basis of 16S rDNA sequence and ARDRA analysis, only two strains were found to be related to the two reference strains of Geodermatophilus. All the others could be grouped with Blastococcus aggregatus (19 strains) or the Antarctic species Modestobacter multiseptatus (44 strains), suggesting that it is these two groups, rather than Geodermatophilus, that tend to colonize the stone surfaces, and that Modestobacter-like strains are also found in temperate/Mediterranean climates. From the BOX-polymerase chain reaction (PCR) data, it can be seen that the Modestobacter-like strains, belonging to the most represented ARDRA haplotype (haplotype B, 34 strains), are very polymorphic and that, over a stone surface, there is a wide genetic diversity at the microsite level.

Aromatic hydrocarbon degradation patterns and catechol 2,3-dioxygenase genes in microbial cultures from deep anoxic hypersaline lakes in the eastern Mediterranean sea.

Several mixed cultures able to grow on different aromatic hydrocarbons were obtained from different depths (between 3500 and 3660 m under the sea surface) of water/brine interfaces (1 to 5 m over the estimated brine surface) of three deep hypersaline anoxic basins (Urania, Discovery and Atalante) in the eastern Mediterranean sea. Eight strains which completely removed toluene from the medium in six to 10 days were isolated from one of the mixed cultures obtained from the Urania basin. The strains grew on toluene and yeast extract in the presence of NaCl concentrations of up to 50 and 100 g l(-1), respectively, indicating that they are halotolerant rather than halophilic. Even though DNA fingerprinting methods showed that the strains were strictly related, two groups could be found on the basis of the plasmid profile. Metabolic profiling and partial sequencing (350 bp) of the 16S rDNA showed that the strains were related to Pseudomonas mendocina. A 320 bp fragment of the catechol 2,3-dioxygenase gene from all the strains was aimplified by PCR. The sequence of the fragment showed 100% identity with xylE from pWW53 of Pseudomonas putida MT53 isolated from soil. Southern hybridisation experiments showed that catechol 2,3-dioxygenase is plasmid encoded.

Thuricin 7: a novel bacteriocin produced by Bacillus thuringiensis BMG1.7, a new strain isolated from soil.

AIMS: Detection and identification of new antagonistic activities towards Bacillus cereus and relatives. METHODS AND RESULTS: Twenty Bacillus thuringiensis strains were screened for their capacity to express bacteriocin-like agents. Strain BMG1.7, isolated from soil, showed an antagonistic activity called thuricin 7. Thuricin 7 was active against several species of the genus Bacillus, including three of the four known B. thuringiensis/B. cereus bacteriocin producers, as well as against Streptococcus pyogenes and Listeria monocytogenes strains. Antimicrobial activity was lost after treatment with proteinase K. The active protein had an apparent molecular weight of 11.6 kDa, and was secreted at the end of the exponential growth phase. Thuricin 7 retained 55% of the activity after incubation at 98 degrees C for 30 min. The mode of action of thuricin 7 was shown to be bactericidal and bacteriolytic. CONCLUSION: Thuricin 7 is a novel bacteriocin produced by a newly isolated Bacillus thuringiensis strain BMG1.7. SIGNIFICANCE AND IMPACT OF THE STUDY: The characteristics of thuricin 7 indicate that it is a new bacteriocin which may have interesting biotechnological applications due to its relatively large activity spectrum.