Metabolic capabilities of microorganisms involved in and associated with the anaerobic oxidation of methane

In marine sediments the anaerobic oxidation of methane with sulfate as electron acceptor (AOM) is responsible for the removal of a major part of the greenhouse gas methane. AOM is performed by consortia of anaerobic methane-oxidizing archaea (ANME) and their specific partner bacteria. The physiology of these organisms is poorly understood, which is due to their slow growth with doubling times in the order of months and the phylogenetic diversity in natural and in vitro AOM enrichments. Here we study sediment-free long-term AOM enrichments that were cultivated from seep sediments sampled off the Italian Island Elba (20◦C; hereon called E20) and from hot vents of the Guaymas Basin, Gulf of California, cultivated at 37◦C (G37) or at 50◦C (G50). These enrichments were dominated by consortia of ANME-2 archaea and Seep-SRB2 partner bacteria (E20) or by ANME-1, forming consortia with Seep-SRB2 bacteria (G37) or with bacteria of the HotSeep-1 cluster (G50). We investigate lipid membrane compositions as possible factors for the different temperature affinities of the different ANME clades and show autotrophy as characteristic feature for both ANME clades and their partner bacteria. Although in the absence of additional substrates methane formation was not observed, methanogenesis from methylated substrates (methanol and methylamine) could be quickly stimulated in the E20 and the G37 enrichment. Responsible for methanogenesis are archaea from the genus Methanohalophilus and Methanococcoides, which are minor community members during AOM (1–7h of archaeal 16S rRNA gene amplicons). In the same two cultures also sulfur disproportionation could be quickly stimulated by addition of zero-valent colloidal sulfur. The isolated partner bacteria are likewise minor community members (1–9h of bacterial 16S rRNA gene amplicons), whereas the dominant partner bacteria (Seep-SRB1a, Seep-SRB2, or HotSeep-1) did not grow on elemental sulfur. Our results support a functioning of AOM as syntrophic interaction of obligate methanotrophic archaea that transfer non-molecular reducing equivalents (i.e., via direct interspecies electron transfer) to obligate sulfate-reducing partner bacteria. Additional katabolic processes in these enrichments but also in sulfate methane interfaces are likely performed by minor community members

BUILDING EFFICIENT AND COST-EFFECTIVE CLOUD-BASED BIG DATA MANAGEMENT SYSTEMS

In today’s big data world, data is being produced in massive volumes, at great velocity and from a variety of different sources such as mobile devices, sensors, a plethora of small devices hooked to the internet (Internet of Things), social networks, communication networks and many others. Interactive querying and large-scale analytics are being increasingly used to derive value out of this big data. A large portion of this data is being stored and processed in the Cloud due the several advantages provided by the Cloud such as scalability, elasticity, availability, low cost of ownership and the overall economies of scale. There is thus, a growing need for large-scale cloud-based data management systems that can support real-time ingest, storage and processing of large volumes of heterogeneous data. However, in the pay-as-you-go Cloud environment, the cost of analytics can grow linearly with the time and resources required. Reducing the cost of data analytics in the Cloud thus remains a primary challenge. In my dissertation research, I have focused on building efficient and cost-effective cloud-based data management systems for different application domains that are predominant in cloud computing environments. In the first part of my dissertation, I address the problem of reducing the cost of transactional workloads on relational databases to support database-as-a-service in the Cloud. The primary challenges in supporting such workloads include choosing how to partition the data across a large number of machines, minimizing the number of distributed transactions, providing high data availability, and tolerating failures gracefully. I have designed, built and evaluated SWORD, an end-to-end scalable online transaction processing system, that utilizes workload-aware data placement and replication to minimize the number of distributed transactions that incorporates a suite of novel techniques to significantly reduce the overheads incurred both during the initial placement of data, and during query execution at runtime. In the second part of my dissertation, I focus on sampling-based progressive analytics as a means to reduce the cost of data analytics in the relational domain. Sampling has been traditionally used by data scientists to get progressive answers to complex analytical tasks over large volumes of data. Typically, this involves manually extracting samples of increasing data size (progressive samples) for exploratory querying. This provides the data scientists with user control, repeatable semantics, and result provenance. However, such solutions result in tedious workflows that preclude the reuse of work across samples. On the other hand, existing approximate query processing systems report early results, but do not offer the above benefits for complex ad-hoc queries. I propose a new progressive data-parallel computation framework, NOW!, that provides support for progressive analytics over big data. In particular, NOW! enables progressive relational (SQL) query support in the Cloud using unique progress semantics that allow efficient and deterministic query processing over samples providing meaningful early results and provenance to data scientists. NOW! enables the provision of early results using significantly fewer resources thereby enabling a substantial reduction in the cost incurred during such analytics. Finally, I propose NSCALE, a system for efficient and cost-effective complex analytics on large-scale graph-structured data in the Cloud. The system is based on the key observation that a wide range of complex analysis tasks over graph data require processing and reasoning about a large number of multi-hop neighborhoods or subgraphs in the graph; examples include ego network analysis, motif counting in biological networks, finding social circles in social networks, personalized recommendations, link prediction, etc. These tasks are not well served by existing vertex-centric graph processing frameworks whose computation and execution models limit the user program to directly access the state of a single vertex, resulting in high execution overheads. Further, the lack of support for extracting the relevant portions of the graph that are of interest to an analysis task and loading it onto distributed memory leads to poor scalability. NSCALE allows users to write programs at the level of neighborhoods or subgraphs rather than at the level of vertices, and to declaratively specify the subgraphs of interest. It enables the efficient distributed execution of these neighborhood-centric complex analysis tasks over largescale graphs, while minimizing resource consumption and communication cost, thereby substantially reducing the overall cost of graph data analytics in the Cloud. The results of our extensive experimental evaluation of these prototypes with several real-world data sets and applications validate the effectiveness of our techniques which provide orders-of-magnitude reductions in the overheads of distributed data querying and analysis in the Cloud

Physiological and genomic characterization of thermophilic methanotrophic archaea and their partner-bacteria

Methane is a potent greenhouse gas and its atmospheric concentration is strongly influenced by microbial processes. In anoxic marine environments 80% of the methane is oxidized by anaerobic microorganisms leading to reduced oceanic methane emissions. This anaerobic oxidation of methane (AOM) is coupled to sulfate reduction and is mediated by microbial consortia of anaerobic methane-oxidizing archaea and partner bacteria. The physiology of the consortia is incompletely understood but is thought to base on a metabolic interdependency of the partners, a syntrophy. The research presented in this PhD thesis focused on the physiology and genomic profile of AOM consortia, in particular on the microorganisms that are active at elevated temperatures (thermophiles). The thermophilic AOM is performed by a unique consortium of ANME-1 archaea and HotSeep-1 bacteria. In Chapter II we describe physiological studies and gene expression experiments with thermophilic AOM consortia and propose a syntrophy of AOM via direct exchange of reducing equivalents. In support of this hypothesis we visualized cell-to-cell connections in these consortia that we suggest to function as conductive nanowires in interspecies electron transfer. For the thermophilic bacterial partner, HotSeep-1 we obtained an ANME-1-free enrichment culture using hydrogen as alternative energy source, and by physiological and genomic investigation we show in Chapter III that this bacterial partner grows as chemolithoautotrophic sulfate reducer. Based on phylogenetic analysis we propose that HotSeep-1 presents a novel species, Candidatus Desulfofervidus auxilii. ANME-1, the archaeon participating in thermophilic AOM, belongs to a large group of uncultured organisms, which are known to have reversed the methanogenesis pathway to metabolize methane. The metabolic diversity among members of the ANME-1 group is still widely unexplored. In a comparative genome analysis of different ANME-1 in Chapter IV we show central aspects of their metabolism including a modified reverse methanogenesis pathway and abundant cytochromes possibly relevant for electron transfer. Environments of AOM activity and in vitro AOM enrichments are dominated by AOM consortia, but other microorganisms sustain as low abundant community whose function is not well understood. In Chapter V we show the cultivation of methanogens and sulfur-disproportionating bacteria from AOM enrichments. In conclusion the work of this PhD thesis has advanced our understanding of the functioning of thermophilic AOM, while further detailed comparative approaches are necessary to comprehend AOM syntrophy in all its detail and diversity

Automation of anatomic torsion monitor for evaluation and improvement of low back dysfunction

The existing Anatomical Torsion Monitor (ATM) to evaluate mechanical stiffness and viscoelasticity of the low back suffers from various inherent defects. This has to be replaced by an improved device. Also the existing ATM cannot provide oscillations to the low back. The main objective is to automate the existing ATM for evaluating the low back immediately using objective methods. The specific objective is to provide oscillations for improving the low back dysfunction. The laser platform and the target chart for recording the readings are dispensed with the existing ATM. Instead, the ultrasound transducers are attached to the pads to record the readings for loading and unloading the low back. The voltage readings are directly recorded in the computer through a DAQ card and the Hysteresis Loop Areas (HLAs) are evaluated using MATLAB. In addition to automation of the ATM for evaluating the lows back, a technique is developed for improving the low back dysfunction by imparting oscillations to the low back. These oscillations can be delivered to the subject using a cam mechanism and a DC motor fitted to the automated ATM (A- ATM). The cam mechanism is used with pneumatic cylinders in order to give the oscillation alternately to both contact pads. The frequency of the oscillations can be controlled by using a speed controller switch. Ten control subjects (nine males and one female) in the age group of (24-77) were given oscillations to the low back for five minutes duration. HLAs were evaluated before and after the treatment in the form of oscillations. The frequency for each oscillation was 20 cycles per minute with amplitude of 2 inches. The percentage change in HLA as well as Range of motion were obtained and summarized. The existing ATM is successfully automated which results in objectively evaluating the passive low back and obtaining the results quicker compared to unautomated ATM. The automated ATM can also deliver quantifiable oscillations to the passive low back. It is observed that providing oscillations to the low back results in improved viscoelasticity of the low back for those subjects whose BMI is 25 or less and an insignificant change in range of motion for all the subjects. It is further observed that based on our tests, the optimal duration of oscillations is 5 minutes. However, the correct displacement amplitude, frequency, and duration of treatment will have to be determined from individual medical and physical conditions

Sutureless repair of transected nerves using photochemically bonded collagen membranes

Peripheral nerve injuries are relatively common with broad-ranging aetiologies that often produce debilitating functional consequences. In its most severe form, nerve trauma involves complete transection of the nerve causing denervation of the target tissue with corresponding functional deficit. While the body possesses an ability to regenerate the severed axons through the mechanism of axon budding, such process is generally incomplete and fails to fully restore sensory and/or motor functions. The outcomes are, among other things, influenced by the surgical repair technique used. For example, standard surgical approach involves suturing the approximated nerve ends, which does not always ensure good alignment and leads to retention of permanent non-absorbable suture material, which often leads to intraneural scarring. Here I have tested a novel sutureless nerve repair technique using a biodegradable collagen membrane bonded with a photochemically activated dye. This process avoids the tissue tension/compression and foreign material retention commonly associated with non-absorbable sutures. In a transected rat sciatic nerve model, this technique has demonstrated superior histological and functional recovery when compared to a standard suturing approach. In future it may form a viable and, potentially, better alternative for surgical treatment of nerve injuries in clinical practice. Additional research will be required to further quantify functional sensory and motor recovery process, as well as histological changes and outcomes in regard to inflammation and regeneration

CARDIAC RECONSTRUCTION WITH ORGAN SPECIFIC EXTRACELLULAR MATRIX

Surgical reconstruction of congenital heart defects is often limited by the non-resorbable material used to approximate normal anatomy. In contrast, non-crosslinked extracellular matrix (ECM) biologic scaffold materials have been used for tissue reconstruction of multiple organs and are replaced by host tissue. Preparation of whole organ ECM by vascular perfusion can maintain much of the native three-dimensional (3D) structure, strength, and tissue specific composition. A 3D Cardiac-ECM (C-ECM) biologic scaffold material would logically have structural and functional advantages over materials such as Dacron™ for myocardial repair, but the in vivo remodeling characteristics of C-ECM have not been investigated to date. Intact porcine and rat hearts were decellularized through retrograde aortic perfusion to create a 3D C-ECM biologic scaffold material. C-ECM biochemical and structural composition were evaluated. C-ECM was not different in cell survival assays from a standard ECM material, urinary bladder matrix (UBM), and supported cardiomyocytes in both 2D and 3D culture. Finally, a porcine C-ECM or Dacron™ patch was used to reconstruct a full thickness right ventricular outflow tract (RVOT) defect in a rat model with a primary endpoint of 16 wk The Dacron patch was encapsulated by dense fibrous tissue and showed little cellular infiltration. Echocardiographic analysis showed that the Dacron patched heart had dilated right ventricular minimum and maximum dimensions at 16 wk compared to pre-surgery baseline values. The C-ECM patch remodeled into dense, cellular connective tissue including: collagen, endothelium, smooth muscle, and small islands of cardiomyocytes. The C-ECM patch showed no ventricular dimensional or functional differences to baseline values at either the 4 or 16 wk time point. The porcine and rat heart can be efficiently decellularized using perfusion in less than 10 hours. The potential benefit of the 2D and 3D C-ECM was shown to support cardiomyocytes with an organized sarcomere structure. The C-ECM patch was associated with better function and histomorphology compared to the Dacron™ patch in this rat model of RVOT reconstruction. While there is much work to be done, the methodology described herein provides a useful step to fully realizing a functional cardiac patch

The 15th Aerospace Mechanisms Symposium

Technological areas covered include: aerospace propulsion; aerodynamic devices; crew safety; space vehicle control; spacecraft deployment, positioning, and pointing; deployable antennas/reflectors; and large space structures. Devices for payload deployment, payload retention, and crew extravehicular activities on the space shuttle orbiter are also described

Physiological and genomic characterization of thermophilic methanotrophic archaea and partner bacteria

Methane is a potent greenhouse gas and its atmospheric concentration is strongly influenced by microbial processes. In anoxic marine environments 80% of the methane is oxidized by anaerobic microorganisms leading to reduced oceanic methane emissions. This anaerobic oxidation of methane (AOM) is coupled to sulfate reduction and is mediated by microbial consortia of anaerobic methane-oxidizing archaea and partner bacteria. The physiology of the consortia is incompletely understood but is thought to base on a metabolic interdependency of the partners, a syntrophy. The research presented in this PhD thesis focused on the physiology and genomic profile of AOM consortia, in particular on the microorganisms that are active at elevated temperatures (thermophiles). The thermophilic AOM is performed by a unique consortium of ANME-1 archaea and HotSeep-1 bacteria. In Chapter II we describe physiological studies and gene expression experiments with thermophilic AOM consortia and propose a syntrophy of AOM via direct exchange of reducing equivalents. In support of this hypothesis we visualized cell-to-cell connections in these consortia that we suggest to function as conductive nanowires in interspecies electron transfer. For the thermophilic bacterial partner, HotSeep-1 we obtained an ANME-1-free enrichment culture using hydrogen as alternative energy source, and by physiological and genomic investigation we show in Chapter III that this bacterial partner grows as chemolithoautotrophic sulfate reducer. Based on phylogenetic analysis we propose that HotSeep-1 presents a novel species, Candidatus Desulfofervidus auxilii. ANME-1, the archaeon participating in thermophilic AOM, belongs to a large group of uncultured organisms, which are known to have reversed the methanogenesis pathway to metabolize methane. The metabolic diversity among members of the ANME-1 group is still widely unexplored. In a comparative genome analysis of different ANME-1 in Chapter IV we show central aspects of their metabolism including a modified methanogenesis pathway and abundant cytochromes possibly relevant for electron transfer. Environments of AOM activity and in vitro AOM enrichments are dominated by AOM consortia, but other microorganisms sustain as low abundant community whose function is not well understood. In Chapter V we show the cultivation of methanogens and sulfur disproportionating bacteria from AOM enrichments. In conclusion the work of this PhD thesis has advanced our understanding of the functioning of thermophilic AOM, while further detailed comparative approaches are necessary to comprehend AOM syntrophy in all its detail and diversity