Stable-Isotope Probing of Microorganisms Thriving at Thermodynamic Limits: Syntrophic Propionate Oxidation in Flooded Soil

Propionate is an important intermediate of the degradation of organic matter in many anoxic environments. In methanogenic environments, due to thermodynamic constraints, the oxidation of propionate requires syntrophic cooperation of propionate-fermenting proton-reducing bacteria and H₂-consuming methanogens. We have identified here microorganisms that were active in syntrophic propionate oxidation in anoxic paddy soil by rRNA-based stable-isotope probing (SIP). After 7 weeks of incubation with [¹³C]propionate (<10 mM) and the oxidation of ~30 μmol of ¹³C-labeled substrate per g dry weight of soil, we found that archaeal nucleic acids were ¹³C labeled to a larger extent than those of the bacterial partners. Nevertheless, both terminal restriction fragment length polymorphism and cloning analyses revealed Syntrophobacter spp., Smithella spp., and the novel Pelotomaculum spp. to predominate in “heavy” ¹³C-labeled bacterial rRNA, clearly showing that these were active in situ in syntrophic propionate oxidation. Among the Archaea, mostly Methanobacterium and Methanosarcina spp. and also members of the yet-uncultured “rice cluster I” lineage had incorporated substantial amounts of ¹³C label, suggesting that these methanogens were directly involved in syntrophic associations and/or thriving on the [¹³C]acetate released by the syntrophs. With this first application of SIP in an anoxic soil environment, we were able to clearly demonstrate that even guilds of microorganisms growing under thermodynamic constraints, as well as phylogenetically diverse syntrophic associations, can be identified by using SIP. This approach holds great promise for determining the structure and function relationships of further syntrophic or other nutritional associations in natural environments and for defining metabolic functions of yet-uncultivated microorganisms.     

Economic importance ofRhagoletis cerasi L., the feasibility of genetic control and resulting research problems

The authors — all members of the recently established O.I.L.B. working group — give in this first report a brief introduction into the pest problem and the biological, economic and environmental aspects which are favourable for the application of genetic control methods againstR. cerasi.

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Résumé En vue du développement des recherches sur la Mouche des cerises en Europe, un groupe de travail de l'O.I.L.B. pour la lutte génétique contreRhagoletis cerasi a été formé par les 4 auteurs. Des contributions du groupe seront désormais publiées sur le problème de la lutte autocide sous ce titre. Les hautes exigences imposées par le marché à la qualité intérieure et extérieure des cerises de table et de conserves d'une part, les pertes financières importantes causées par la disqualification des fruits infestés pour la distillation d'autre part, exigent des moyens de lutte efficaces. L'influence des mesures de lutte sur la biocénose doit cependant être minime. Les possibilités et avantages ainsi que les limites de l'application de la lutte génétique sont discutés en fonction de la situation favorable des points de vue topographique, écologique et biologique. En conclusion, la situation pour l'application de la lutte génétique peut être jugée favorable dans des cas choisis. Les taches de recherche qui en résultent sont commentées et complétées par un exposé sur la situation actuelle des travaux en cours.

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First Report of the O.I.L.B. working group on genetic control of the European cherry fruit fly,Rhagoletis cerasi L.     

Genetic control of invasive fish: technological options and its role in integrated pest management

Genetic options for the control of invasive fishes were recently reviewed and synthesized at a 2010 international symposium, held in Minneapolis/St. Paul, MN, USA. The only option currently available “off-the-shelf” is triploidy, which can be used to produce sterile males for a release program analogous to those widely and successfully used for biological control of insect pests. However, the Trojan Y and several recombinant options that heritably distort pest population sex ratios are technologically feasible, are at or are close to proof-of-concept stage and are potentially much more effective than sterile male release programs. All genetic options at this stage require prolonged stocking programs to be effective, though gene drive systems are a potential for recombinant approaches. They are also likely to differ in their current degree of social acceptability, with chromosomal approaches (triploidy and Trojan Y) likely to be the most readily acceptable to the public and least likely to require changes in legislative or policy settings to be implemented. Modelling also suggests that the efficacy of any of these genetic techniques is enhanced by, and in turn non-additively enhance, conventional methods of pest fish control.     

Parental provisioning behaviour in Pied Flycatchers Ficedula hypoleuca is well adjusted to local conditions in a mosaic of deciduous and coniferous habitat

Capsule Pied Flycatchers are better able than Great Tits to adjust their feeding behaviour to varying conditions in the same area.

Aims Great Tits breeding in a mosaic of deciduous and coniferous forests in the northern temperate region exhibit consistently lower breeding success in their preferred deciduous habitat than in coniferous habitat. This was explained by the unexpectedly poor nestling feeding conditions in deciduous forests of this region. We studied whether the same paradox applies to Pied Flycatchers that occupy the same habitats in the same area.

Methods Parental provisioning behaviour was studied using video‐recording and experimental manipulation. Caterpillar abundance and basic breeding parameters were measured in different habitat types.

Results Parental provisioning frequency and the proportion of caterpillars in nestling diet was lower, while food objects were on average larger, in coniferous compared with deciduous habitat. However, the total volume of caterpillars and adult Lepidoptera delivered to nestlings did not differ between habitats. In contrast to Great Tits, offspring body parameters in Pied Flycatchers did not differ between habitat types.

Conclusions These results demonstrate how the relative suitability of particular habitat types varies between species and is dependent upon geographical location.     

Risk prediction for clinical phenotype in myotonic dystrophy type 1: data from 2,650 patients

Myotonic dystrophy type 1 (DM1) is a multisystem disorder that affects skeletal and smooth muscle as well as the eye, heart, endocrine system, and central nervous system. DM1 is caused by expansion of a CTG trinucleotidedaggerrepeat in the gene DMPK. Clinical findings in DM1 span a continuum from mild to severe. Although the CTG repeat correlates with the disease phenotype, caution is used in predicting disease severity on the basis of CTG repeat number. This study reports an extensive genotype-phenotype study to evaluate the clinical validity and clinical utility of the molecular genetic test. Data were analyzed by multiple logistic regression, used to estimate the odds ratio (OR) and correlation coefficients for patients phenotype in respect to the categorical variables expansion class, gender, familiarity, and the continuous variables age and disease duration. We assessed disease expression by clinical evaluation and the molecular genetic test in 2,650 patients identified by accurate clinical diagnosis and family segregation. We were able to estimate OR and correlation coefficients for patients phenotype according to CTG number. A genotype-phenotype correlation was established to derivate a clinical predictive risk on the basis of molecular data. This study demonstrates that measurement of triplet expansions in patients' DNA can be considered as a useful tool for DM1 phenotype assessment and presymptomatic testing.     

Identification of acetate-utilizing Bacteria and Archaea in methanogenic profundal sediments of Lake Kinneret (Israel) by stable isotope probing of rRNA

Acetate is an important intermediate in the decomposition of organic matter in anoxic freshwater sediments. Here, we identified distinct microorganisms active in its oxidation and transformation to methane in the anoxic methanogenic layers of Lake Kinneret (Israel) profundal sediment by rRNA-based stable isotope probing (RNA-SIP). After 18 days of incubation with amended [U-(13)C]acetate we found that archaeal 16S rRNA was (13)C-labelled to a far greater extent than bacterial rRNA. We identified acetoclastic methanogens related to Methanosaeta concilii as being most active in the degradation and assimilation of acetate. Oxidation of the acetate-methyl group played only a minor role, but nevertheless 'heavy'(13)C-labelled bacterial rRNA templates were identified. 'Heavy' bacteria were mainly affiliated with the Betaproteobacteria (mostly Rhodocyclales and Nitrosomonadales), the Nitrospira phylum (related to 'Magnetobacterium bavaricum' and Thermodesulfovibrio yellowstonii), and also with the candidate phylum 'Endomicrobia'. However, the mode of energy gain that allowed for the assimilation of (13)C-acetate by these bacterial groups remains unknown. It may have involved syntrophic oxidation of acetate, reduction of chlorinated compounds, reduction of humic substances, fermentation of organic compounds, or even predation of (13)C-labelled Methanosaeta spp. In summary, this SIP experiment shows that acetate carbon was predominantly consumed by acetoclastic methanogens in profundal Lake Kinneret sediment, while it was also utilized by a small and heterogeneous community of bacteria.     

Manipulation of laying effort reveals habitat-specific variation in egg production constraints in Great Tits (Parus major)

It has been suggested that a bird's clutch size is not limited by the amount of resources available at the time of laying but that differences in the availability of food for nestlings is the ultimate underlying factor determining spatio-temporal variations in clutch size. However, habitat-related variations in egg production ability has yet to be investigated explicitly. We studied the breeding of Great Tits Parus major in deciduous and coniferous forests in the same area. The sizes of both the clutches and the eggs were, on average, larger in the former habitat than in the latter. A number of females were induced to lay more eggs than usual by removing four eggs from designated experimental clutches early in the laying period. These manipulated females laid approximately one egg more than control females, with the number of additional eggs laid not differing between the habitats. However, in both study years the relative size of the extra eggs – relative to the mean size of earlier laid eggs of the same clutch – was smaller in the coniferous habitat than in the deciduous habitat, while there was no habitat-related difference in the relative size of the last-laid eggs of control clutches. This result indicates that some form of proximate limitation during egg-laying period can contribute to the relatively small clutches and eggs in the coniferous habitat. Our results emphasize the need to take egg production costs into account when attempting to account for spatial variation in the reproductive behaviour of birds.     

Gonadal differentiation and hormonal sex reversal in Arctic charr (Salvelinus alpinus)

The purpose of this study was to develop a practical protocol for the production of female populations of Arctic charr (Salvelinus alpinus). Achieving this required knowledge of the timing of gonadal differentiation. Undifferentiated gonads were observed microscopically to be present by 194 degrees C-days post-hatch and definitive germ cells by 346 degrees C-days post-hatch, where " degrees C-days" denote acquired thermal units calculated as the product of temperature and days. Some of the gonads had developed a lumen by 510 degrees C-days post-hatch, and by 681 degrees C-days post-hatch anatomical divergence into two types of gonads was clear. Two protocols (immersion and feeding) were tested for hormonal sex reversal of genotypic females using the synthetic androgen 17alpha-methyldihydrotestosterone (MDHT). Six-hour MDHT immersions (0.5, 1, 3, 5 and 10 mg/L) were carried out weekly from hatch to first feeding (140 degrees C-days post-hatch), whereas daily feeding treatments (0.5 mg/kg) went from 140 to 600 degrees C-days post-hatch. The sex ratios of all immersion experimental groups were significantly different from the control, with the proportion of presumptive males increasing as MDHT concentration increased. The highest immersion treatment, 10 mg/L, yielded a population of 90% presumptive males and 10% with atypical gonads. However, the most effective treatment, yielding a population of 90% presumptive males and no fish with atypical gonads, was the feeding treatment. Given that female salmonid fishes are homogametic, sex-reversed (masculinized) genotypic females produced in this way can serve as broodstock for the creation of all-female charr populations for aquaculture.     

Sensitive commercial NASBA assay for the detection of respiratory syncytial virus in clinical specimen

The aim of the study was to evaluate the usability of three diagnostic procedures for the detection of respiratory syncytial virus in clinical samples. Therefore, the FDA cleared CE marked NOW(R) RSV ELISA, the NucliSENS EasyQ RSV A+B NASBA, and a literature based inhouse RT-PCR protocol were compared for their relative sensitivities. Thereby, NASBA turned out to be the most sensitive method with a total number of 80 RSV positive samples out of a cohort of 251 nasopharyngeal washings from patients suffering from clinical symptoms, followed by the inhouse RT-PCR (62/251) and ELISA (52/251). Thus, NASBA may serve as a rapid and highly sensitive alternative for RSV diagnostics.