Light‐Dependent and Aeration‐Independent Gram‐Scale Hydroxylation of Cyclohexane to Cyclohexanol by CYP450 Harboring Synechocystis sp. PCC 6803

Oxygenase‐containing cyanobacteria constitute promising whole‐cell biocatalysts for oxyfunctionalization reactions. Photosynthetic water oxidation thereby delivers the required cosubstrates, that is activated reduction equivalents and O₂, sustainably. A recombinant Synechocystis sp. PCC 6803 strain showing unprecedentedly high photosynthesis‐driven oxyfunctionalization activities is developed, and its technical applicability is evaluated. The cells functionally synthesize a heterologous cytochrome P450 monooxygenase enabling cyclohexane hydroxylation. The biocatalyst‐specific reaction rate is found to be light‐dependent, reaching 26.3 ± 0.6 U g_CDW^?1 (U = μmol min^?1 and cell dry weight [CDW]) at a light intensity of 150 µmol_photons m^?2 s^?1. In situ substrate supply via a two‐liquid phase system increases the initial specific activity to 39.2 ± 0.7 U g_CDW^?1 and stabilizes the biotransformation by preventing cell toxification. This results in a tenfold increased specific product yield of 4.5 g_cyclohexanol g_CDW^?1 as compared to the single aqueous phase system. Subsequently, the biotransformation is scaled from a shake flask to a 3 L stirred‐tank photobioreactor setup. In situ O₂ generation via photosynthetic water oxidation allows a nonaerated process operation, thus circumventing substrate evaporation as the most critical factor limiting the process performance and stability. This study for the first time exemplifies the technical applicability of cyanobacteria for aeration‐independent light‐driven oxyfunctionalization reactions involving highly toxic and volatile substrates.     

Genetic diversity and morphological stasis in the Ceylon Snakehead,Channa orientalis (Teleostei: Channidae)

Ichthyological Research - As the type species of the genus Channa, the identity of the pelvic-finless snakehead Channa orientalis Bloch is important to channid systematics. Although this name has...     

Challenges of Large Carnivore Conservation: Sloth Bear Attacks in Sri Lanka

Sloth bear (Melursus ursinus) attacks in Sri Lanka present a major challenge to their conservation as human populations grow and compete with bears for space and resources. Bear attacks on humans predominantly affect the rural poor and seem to be increasing in frequency. In 2004, we conducted the first island-wide survey of bear attacks in Sri Lanka. Our main goals were to 1) describe where and when attacks occurred, 2) describe the characteristics and outcome of bear attacks, and 3) evaluate the influence of human group size on the severity of bear attacks. We interviewed 271 individuals attacked by sloth bears between 1938 and 2004. Sloth bears rarely entered village compounds, damaged property, or raided crops; conflicts predominantly involved attacks on people in forests. Most sloth bear attacks took place during daylight hours (0900-1600 h) in the dry season, when humans were likely to engage in forest activities such as honey gathering. Most attacks (80 %) occurred when humans and bears surprised each other at close proximity (<10 m). Most (84 %) victims carried a weapon but 55 % said the attack was too sudden to effectively use the weapon in self-defense. Human injuries included lacerations and puncture wounds, broken limbs, skull fractures, and the loss of scalps, eyes, or other parts of the face. Almost all major injuries resulted from bites. In 61 attacks bears were wounded and in 39 attacks bears were killed. The severity of human injuries depended on human group size; solitary humans experienced more severe injuries than expected, suggesting that the presence of other humans may moderate the bear’s aggression and motivate it to flee. Widespread fear of sloth bears poses a significant challenge to modifying ineffective human responses to bear encounters (e.g., fleeing). Conservation outreach could emphasize that greater human safety may be achieved from remaining in groups and using extreme caution around thickets and rock outcrops likely to be used by bears.     

Evaluating scaling of capillary photo-biofilm reactors for high cell density cultivation of mixed trophies artificial microbial consortia

Capillary biofilm reactors (CBRs) are attractive for growing photoautotrophic bacteria as they allow high cell-density cultivation. Here, we evaluated the CBR system's suitability to grow an artificial consortium composed of Synechocystis sp. PCC 6803 and Pseudomonas sp. VBL120. The impact of reactor material, flow rate, pH, O₂, and medium composition on biomass development and long-term biofilm stability at different reactor scales was studied. Silicone was superior over other materials like glass or PVC due to its excellent O₂ permeability. High flow rates of 520 μL min⁻¹ prevented biofilm sloughing in 1 m capillary reactors, leading to a 54% higher biomass dry weight combined with the lowest O₂ concentration inside the reactor compared to standard operating conditions. Further increase in reactor length to 5 m revealed a limitation in trace elements. Increasing trace elements by a factor of five allowed for complete surface coverage with a biomass dry weight of 36.8 g m⁻² and, thus, a successful CBR scale-up by a factor of 25. Practical application : Cyanobacteria use light energy to upgrade CO₂, thereby holding the potential for carbon-neutral production processes. One of the persisting challenges is low cell density due to light limitations and O₂ accumulation often occurring in established flat panel or tubular photobioreactors. Compared to planktonic cultures, much higher cell densities (factor 10 to 100) can be obtained in cyanobacterial biofilms. The capillary biofilm reactor (CBR) offers good growth conditions for cyanobacterial biofilms, but its applicability has been shown only on the laboratory scale. Here, a first scale-up study based on sizing up was performed, testing the feasibility of this system for large-scale applications. We demonstrate that by optimizing nutrient supply and flow conditions, the system could be enlarged by factor 25 by enhancing the length of the reactor. This reactor concept, combined with cyanobacterial biofilms and numbering up, holds the potential to be applied as a flexible, carbon-neutral production platform for value-added compounds.     

Sequence deeper without sequencing more: Bayesian resolution of ambiguously mapped reads

Next-generation sequencing (NGS) has transformed molecular biology and contributed to many seminal insights into genomic regulation and function. Apart from whole-genome sequencing, an NGS workflow involves alignment of the sequencing reads to the genome of study, after which the resulting alignments can be used for downstream analyses. However, alignment is complicated by the repetitive sequences; many reads align to more than one genomic locus, with 15–30% of the genome not being uniquely mappable by short-read NGS. This problem is typically addressed by discarding reads that do not uniquely map to the genome, but this practice can lead to systematic distortion of the data. Previous studies that developed methods for handling ambiguously mapped reads were often of limited applicability or were computationally intensive, hindering their broader usage. In this work, we present SmartMap: an algorithm that augments industry-standard aligners to enable usage of ambiguously mapped reads by assigning weights to each alignment with Bayesian analysis of the read distribution and alignment quality. SmartMap is computationally efficient, utilizing far fewer weighting iterations than previously thought necessary to process alignments and, as such, analyzing more than a billion alignments of NGS reads in approximately one hour on a desktop PC. By applying SmartMap to peak-type NGS data, including MNase-seq, ChIP-seq, and ATAC-seq in three organisms, we can increase read depth by up to 53% and increase the mapped proportion of the genome by up to 18% compared to analyses utilizing only uniquely mapped reads. We further show that SmartMap enables the analysis of more than 140,000 repetitive elements that could not be analyzed by traditional ChIP-seq workflows, and we utilize this method to gain insight into the epigenetic regulation of different classes of repetitive elements. These data emphasize both the dangers of discarding ambiguously mapped reads and their power for driving biological discovery.     

Gamma irradiating chromatography columns enables bioburden-free integrated continuous biomanufacturing

An important consideration for integrated continuous biomanufacturing is that the downstream chromatography steps integrated with the bioreactor should maintain a low bioburden state throughout the entire duration of the operation. One potential strategy to achieve this is to start bioburden-free and functionally close the chromatography system. While chromatography skids themselves can be rendered bioburden-free, limitations exist in applying these methods to chromatography columns. The small column sizes used in continuous multicolumn chromatography enable gamma irradiation of disposable columns to render them bioburden-free. However, this approach has not been widely implemented, likely because gamma irradiation can negatively impact resin performance. Here, several protective mobile-phase modifiers were screened and shown to help chromatography resins retain naïve-like performance. Gamma irradiated columns were then integrated into perfusion bioreactors for continuous capture. Successful integrated continuous capture downstream of perfusion bioreactors for greater than 40 days using protein A, custom affinity, and non-affinity capture resins for multiple biologic modalities is demonstrated in development and commercial settings. No indications of time-based performance decline or bioburden growth have been observed. This strategy enables bioburden-free integrated continuous biomanufacturing operations and could allow full process closure and decreased environmental control requirements for facilities; thus, permitting simultaneous multi-product operations in a ballroom arrangement.     

Integration of evidence across human and model organism studies: A meeting report

The National Institute on Drug Abuse and Joint Institute for Biological Sciences at the Oak Ridge National Laboratory hosted a meeting attended by a diverse group of scientists with expertise in substance use disorders (SUDs), computational biology, and FAIR (Findability, Accessibility, Interoperability, and Reusability) data sharing. The meeting's objective was to discuss and evaluate better strategies to integrate genetic, epigenetic, and 'omics data across human and model organisms to achieve deeper mechanistic insight into SUDs. Specific topics were to (a) evaluate the current state of substance use genetics and genomics research and fundamental gaps, (b) identify opportunities and challenges of integration and sharing across species and data types, (c) identify current tools and resources for integration of genetic, epigenetic, and phenotypic data, (d) discuss steps and impediment related to data integration, and (e) outline future steps to support more effective collaboration—particularly between animal model research communities and human genetics and clinical research teams. This review summarizes key facets of this catalytic discussion with a focus on new opportunities and gaps in resources and knowledge on SUDs.     

Development of persistent lymphocytosis in cattle is closely associated with DRB2

Correspondence to: H. A. Lewin.