Challenges in the microbial production of flavonoids

As flavonoids have beneficial health effects on humans, they are gaining increasing interest from pharmaceutical and health industries. However, current production methods, such as plant extraction and chemical synthesis, are inadequate to meet the demand. Therefore, microbial production might offer a promising alternative. During recent years, microbial strains able to produce flavonoids to a certain extent have been developed. However production titers are limited to the mg l^?1 range, hampering the industrial exploitation of these strains. The latter will not be achieved by simply introducing the heterologous pathway in the production host and optimizing the fermentation process, but will depend on the interaction of different aspects of metabolic engineering and process engineering to overcome the current limitations. Next to engineering the production strain to optimize the availability of precursors, the pathway itself also requires intensive engineering. Currently utilized strategies result in a wide variety of different production strains, requiring high-throughput screening methods to identify optimal performing strains. As more and more organisms are being characterized, each with their own specific properties which might be beneficial for the heterologous production of flavonoids, the choice of the production host is another important aspect. Finally, the use of co-cultures might offer an alternative in which different parts of the process are performed by different organisms. This review aims to provide an overview of the research that has been done on these separate aspects. The work presented here could be used as a framework for further research.     

Diversity of endemic rhizobia on Christmas Island: Implications for agriculture following phosphate mining

Given that phosphate supplies may diminish and become uneconomic to mine after 2020, there is a compelling need to develop alternative industries to support the population on Christmas Island. Former mine sites could be turned into productive agricultural land, however, large-scale commercial agriculture has never been attempted, and, given the uniqueness of the island, the diversity of rhizobia prior to introducing legumes needed evaluation. Therefore, 84 rhizobia isolates were obtained from nine different hosts, both crop and introduced legumes, located at seven sites across the island. Based on 16S rRNA and recA gene sequence analysis, the isolates grouped into 13 clades clustering within the genus Bradyrhizobium, Ensifer, Cupriavidus and Rhizobium. According to the sequences of their symbiosis genes nodC and nifH, the isolates were classified into 12 and 11 clades, respectively, and clustered closest to tropical or crop legume isolates. Moreover, the symbiosis gene phylogeny and Multi Locus Sequence Analysis gene phylogeny suggested vertical transmission in the Alpha-rhizobia but horizontal transmission within the Beta-rhizobia. Furthermore, this study provides evidence of a large diversity of endemic rhizobia associated with both crop and introduced legumes, and highlights the necessity of inoculation for common bean, chickpea and soybean on the Island.     

Toward a Phenomenological Account of Embodied Subjectivity in Autism

Sensorimotor research is currently challenging the dominant understanding of autism as a deficit in the cognitive ability to ‘mindread’. This marks an emerging shift in autism research from a focus on the structure and processes of the mind to a focus on autistic behavior as grounded in the body. Contemporary researchers in sensorimotor differences in autism call for a reconciliation between the scientific understanding of autism and the first-person experience of autistic individuals. I argue that fulfilling this ambition requires a phenomenological understanding of the body as it presents itself in ordinary experience, namely as the subject of experience rather than a physical object. On this basis, I investigate how the phenomenology of Maurice Merleau-Ponty can be employed as a frame of understanding for bodily experience in autism. Through a phenomenological analysis of Tito Mukhopadhyay’s autobiographical work, How can I talk if my lips don’t move (2009), I illustrate the relevance and potential of phenomenological philosophy in autism research, arguing that this approach enables a deeper understanding of bodily and subjective experiences related to autism.     

Genetic diversity and symbiotic effectiveness of Phaseolus vulgaris-nodulating rhizobia in Kenya

Phaseolus vulgaris (common bean) was introduced to Kenya several centuries ago but the rhizobia that nodulate it in the country remain poorly characterised. To address this gap in knowledge, 178 isolates recovered from the root nodules of P. vulgaris cultivated in Kenya were genotyped stepwise by the analysis of genomic DNA fingerprints, PCR-RFLP and 16S rRNA, atpD, recA and nodC gene sequences. Results indicated that P. vulgaris in Kenya is nodulated by at least six Rhizobium genospecies, with most of the isolates belonging to Rhizobium phaseoli and a possibly novel Rhizobium species. Infrequently, isolates belonged to Rhizobium paranaense, Rhizobium leucaenae, Rhizobium sophoriradicis and Rhizobium aegyptiacum. Despite considerable core-gene heterogeneity among the isolates, only four nodC gene alleles were observed indicating conservation within this gene. Testing of the capacity of the isolates to fix nitrogen (N₂) in symbiosis with P. vulgaris revealed wide variations in effectiveness, with ten isolates comparable to Rhizobium tropici CIAT 899, a commercial inoculant strain for P. vulgaris. In addition to unveiling effective native rhizobial strains with potential as inoculants in Kenya, this study demonstrated that Kenyan soils harbour diverse P. vulgaris-nodulating rhizobia, some of which formed phylogenetic clusters distinct from known lineages. The native rhizobia differed by site, suggesting that field inoculation of P. vulgaris may need to be locally optimised.     

Spatio-temporal variability and environmental controls of methane fluxes at the forest–tundra ecotone in the Fennoscandian mountains

We report on temporal and spatial variability in net methane (CH₄) fluxes measured during the thaw period of 1999 and 2000 at three study sites along a c. 8° latitudinal gradient in the Fennoscandian mountain range and across the mountain birch‐tundra ecotone. All of the sites studied here were underlain by well‐drained mesic soils. In addition, we conducted warming experiments in the field to simulate future climate change. Our results show significant CH₄ uptake at mesic sites spanning the forest‐tundra ecotone: on average 0.031 and 0.0065 mg CH₄ m^?2 h^?1 during the 1999 and 2000 thaw periods, respectively, in Abisko (Sweden), and 0.019 and 0.032 mg CH₄ m^?2 h^?1 during 2000 in Dovrefjell and Joatka (Norway), respectively. These values were both temporally and spatially highly variable, and multiple regression analysis of data from Abisko showed no consistent relationship with soil‐moisture status and temperature. Also, there was no consistent difference in CH₄ fluxes between forest and tundra plots; our data, therefore, provide no support for the hypothesis that conversion of tundra to mountain birch forest, or vice versa, would result in a systematic change in the magnitude or direction of net CH₄ fluxes in this region. Experimental warming treatments were associated with a 2.4 °C increase in soil temperatures (5 cm depth) in 1999 in Abisko, but no consistent soil warming was noted at any of the three field locations during 2000. In spite of this, there were significant treatment effects, principally early during the thaw period, with increased CH₄ uptake compared with control (ambient) plots. These results suggest that direct effects of air warming on vegetation processes (e.g. transpiration, root exudation and nutrient assimilation) can influence CH₄ fluxes even in predominantly methanotrophic environments. We conclude that net CH₄ oxidation is significant in these cold, mesic soils and could be strengthened in an environmental change scenario involving a combination of (i) an increase in the length of the thaw period and (ii) increased mean temperatures during this period in combination with decreased soil‐moisture content.     

Met23Lys mutation in subunit gamma of FOF1-ATP synthase from Rhodobacter capsulatus impairs the activation of ATP hydrolysis by protonmotive force

H⁺-F_OF₁-ATP synthase couples proton flow through its membrane portion, F_O, to the synthesis of ATP in its headpiece, F₁. Upon reversal of the reaction the enzyme functions as a proton pumping ATPase. Even in the simplest bacterial enzyme the ATPase activity is regulated by several mechanisms, involving inhibition by MgADP, conformational transitions of the ε subunit, and activation by protonmotive force. Here we report that the Met23Lys mutation in the γ subunit of the Rhodobacter capsulatus ATP synthase significantly impaired the activation of ATP hydrolysis by protonmotive force. The impairment in the mutant was due to faster enzyme deactivation that was particularly evident at low ATP/ADP ratio. We suggest that the electrostatic interaction of the introduced γLys23 with the DELSEED region of subunit β stabilized the ADP-inhibited state of the enzyme by hindering the rotation of subunit γ rotation which is necessary for the activation.