Identification of a Single Strand Origin of Replication in the Integrative and Conjugative Element ICEBs1 of Bacillus subtilis

We identified a functional single strand origin of replication (sso) in the integrative and conjugative element ICEBs1 of Bacillus subtilis. Integrative and conjugative elements (ICEs, also known as conjugative transposons) are DNA elements typically found integrated into a bacterial chromosome where they are transmitted to daughter cells by chromosomal replication and cell division. Under certain conditions, ICEs become activated and excise from the host chromosome and can transfer to neighboring cells via the element-encoded conjugation machinery. Activated ICEBs1 undergoes autonomous rolling circle replication that is needed for the maintenance of the excised element in growing and dividing cells. Rolling circle replication, used by many plasmids and phages, generates single-stranded DNA (ssDNA). In many cases, the presence of an sso enhances the conversion of the ssDNA to double-stranded DNA (dsDNA) by enabling priming of synthesis of the second DNA strand. We initially identified sso1 in ICEBs1 based on sequence similarity to the sso of an RCR plasmid. Several functional assays confirmed Sso activity. Genetic analyses indicated that ICEBs1 uses sso1 and at least one other region for second strand DNA synthesis. We found that Sso activity was important for two key aspects of the ICEBs1 lifecycle: 1) maintenance of the plasmid form of ICEBs1 in cells after excision from the chromosome, and 2) stable acquisition of ICEBs1 following transfer to a new host. We identified sequences similar to known plasmid sso's in several other ICEs. Together, our results indicate that many other ICEs contain at least one single strand origin of replication, that these ICEs likely undergo autonomous replication, and that replication contributes to the stability and spread of these elements.National Institute of General Medical Sciences (U.S.) (Award R01GM050895)National Institute of General Medical Sciences (U.S.) (Pre-Doctoral Training Grant T32GM007287

Strategies for Wildlife Disease Surveillance

Epidemiologic surveillance is defined by the Centers for Disease Control and Prevention (CDC) as the ongoing systematic and continuous collection, analysis, and interpretation of health data\u27: The objective of surveillance is to generate data for rapid response to the detection of a disease of concern to apply prevention, control, or eradication measures as well as to evaluate such interventions. This is distinct from disease monitoring, which usually does not involve a particular response to disease detection. Surveillance for wildlife diseases has increased in importance due to the emergence and re-emergence of wildlife diseases that are threats to human, animal, and ecosystem health, or could potentially have a negative economic impact. It has been estimated that 75% of emerging human diseases are zoonotic in origin, of which the majority originate from wildlife (Taylor et al. 2001). However, there are unique challenges concerning wildlife disease surveillance such that disease and pathogens can be very difficult to detect and measure in wild animals. These challenges have been described previously (Wobeser 2006), but one of the primary issues is that disease in wildlife often goes unrecognized, especially in remote locations. Furthermore, sick and dead animals are very difficult to detect, as animals will disguise the signs of illness or hide when diseased. Carcasses from diseased animals are also rapidly removed by scavengers or will rapidly decompose, rendering them suboptimal for diagnostic purposes. There is also a lack of validated diagnostic tests for most wildlife disease agents as well as baseline data. The paucity of laboratory capacity with expertise in wildlife disease diagnostic investigation is also an impediment. Finally, surveillance networks for wildlife diseases that perform field investigations and report disease events are under-developed in most regions of the world. Despite these challenges, a number of very important epidemiological surveillance projects have been ongoing or recently developed, and some examples are described in this chapter. The examples are mostly drawn from the experiences of the U.S. Geological Survey National Wildlife Health Center (NWHC) and are provided to illustrate the different surveillance strategies and sampling techniques that can be used and have proven successful. Some future directions for wildlife disease surveillance are also suggested

A new approach to varietal identification in plants by microsatellite high resolution melting analysis: application to the verification of grapevine and olive cultivars

<p>Abstract</p> <p>Background</p> <p>Microsatellites are popular molecular markers in many plant species due to their stable and highly polymorphic nature. A number of analysis methods have been described but analyses of these markers are typically performed on cumbersome polyacrylamide gels or more conveniently by capillary electrophoresis on automated sequencers. However post-PCR handling steps are still required. High resolution melting can now combine detailed sequence analysis with the closed-tube benefits of real-time PCR and is described here as a novel way to verify the identity of plant varieties such as grapevine and olive.</p> <p>Results</p> <p>DNA melting profiles for various plant variety and rootstock samples were compared to profiles for certified reference samples. Two closely related grapevine rootstocks differing by as little as a single di-nucleotide repeat could be rapidly differentiated while there was high reproducibility of melting profiles for identical cultivars.</p> <p>Conclusion</p> <p>This novel microsatellite analysis method allows high sample throughput with greatly reduced time to results for varietal certification and is amenable to other microsatellite analyses.</p

Srs2 promotes synthesis-dependent strand annealing by disrupting DNA polymerase δ-extending D-loops.

Synthesis-dependent strand annealing (SDSA) is the preferred mode of homologous recombination in somatic cells leading to an obligatory non-crossover outcome, thus avoiding the potential for chromosomal rearrangements and loss of heterozygosity. Genetic analysis identified the Srs2 helicase as a prime candidate to promote SDSA. Here, we demonstrate that Srs2 disrupts D-loops in an ATP-dependent fashion and with a distinct polarity. Specifically, we partly reconstitute the SDSA pathway using Rad51, Rad54, RPA, RFC, DNA Polymerase δ with different forms of PCNA. Consistent with genetic data showing the requirement for SUMO and PCNA binding for the SDSA role of Srs2, Srs2 displays a slight but significant preference to disrupt extending D-loops over unextended D-loops when SUMOylated PCNA is present, compared to unmodified PCNA or monoubiquitinated PCNA. Our data establish a biochemical mechanism for the role of Srs2 in crossover suppression by promoting SDSA through disruption of extended D-loops

Femoroacetabular Impingement as a Complication of Acetabular Fracture Fixation

Case We present the case of a thirteen-year-old female who sustained a posterior wall acetabular fracture dislocation. She underwent urgent closed reduction and subsequent uncomplicated open reduction and internal fixation. Post reduction computed tomography demonstrated a concentrically reduced hip joint with no evidence of femoroacetabular impingement (FAI). She subsequently healed her fracture and returned to running activities; however, one year later presented with aching pain in her thigh. Radiographs demonstrated the development of a large osseous prominence on her anterolateral femoral neck consistent with femoroacetabular impingement. Based on these findings she was evaluated by a hip preservation specialist. She subsequently underwent successful hip arthroscopy for labral repair and femoral osteochondroplasty. She was eventually able to return to running sports with little pain. Summary We present a case of FAI presenting as a complication of acetabular fracture fixation. This should be discussed with patients presenting with traumatic hip dislocations as a possible complication of surgical fixation or possibly of the injury itself

Spatial Patterns and Sequential Sampling Plans for Estimating Densities of Stink Bugs (Hemiptera: Pentatomidae) in Soybean in the North Central Region of the United States

Stink bugs are an emerging threat to soybean (Fabales: Fabaceae) in the North Central Region of the United States. Consequently, region-specific scouting recommendations for stink bugs are needed. The aim of this study was to characterize the spatial pattern and to develop sampling plans to estimate stink bug population density in soybean fields. In 2016 and 2017, 125 fields distributed across nine states were sampled using sweep nets. Regression analyses were used to determine the effects of stink bug species [Chinavia hilaris (Say) (Hemiptera: Pentatomidae) and Euschistus spp. (Hemiptera: Pentatomidae)], life stages (nymphs and adults), and field locations (edge and interior) on spatial pattern as represented by variance–mean relationships. Results showed that stink bugs were aggregated. Sequential sampling plans were developed for each combination of species, life stage, and location and for all the data combined. Results for required sample size showed that an average of 40–42 sample units (sets of 25 sweeps) would be necessary to achieve a precision of 0.25 for stink bug densities commonly encountered across the region. However, based on the observed geographic gradient of stink bug densities, more practical sample sizes (5–10 sample units) may be sufficient in states in the southeastern part of the region, whereas impractical sample sizes (\u3e100 sample units) may be required in the northwestern part of the region. Our findings provide research-based sampling recommendations for estimating densities of these emerging pests in soybean

A Pulsational Model for the Orthogonal Polarization Modes in Radio Pulsars

In an earlier paper, we introduced a model for pulsars in which non-radial oscillations of high spherical degree (\el) aligned to the magnetic axis of a spinning neutron star were able to reproduce subpulses like those observed in single-pulse measurements of pulsar intensity. The model did not address polarization, which is an integral part of pulsar emission. Observations show that many pulsars emit radio waves that appear to be the superposition of two linearly polarized emission modes with orthogonal polarization angles. In this paper, we extend our model to incorporate linear polarization. As before, we propose that pulsational displacements of stellar material modulate the pulsar emission, but now we apply this modulation to a linearly-polarized mode of emission, as might be produced by curvature radiation. We further introduce a second polarization mode, orthogonal to the first, that is modulated by pulsational velocities. We combine these modes in superposition to model the observed Stokes parameters in radio pulsars.Comment: 19 pages, 4 figures accepted Ap

Testosterone disrupts human collaboration by increasing egocentric choices

Collaboration can provide benefits to the individual and the group across a variety of contexts. Even in simple perceptual tasks, the aggregation of individuals' personal information can enable enhanced group decision-making. However, in certain circumstances such collaboration can worsen performance, or even expose an individual to exploitation in economic tasks, and therefore a balance needs to be struck between a collaborative and a more egocentric disposition. Neurohumoral agents such as oxytocin are known to promote collaborative behaviours in economic tasks, but whether there are opponent agents, and whether these might even affect information aggregation without an economic component, is unknown. Here, we show that an androgen hormone, testosterone, acts as such an agent. Testosterone causally disrupted collaborative decision-making in a perceptual decision task, markedly reducing performance benefit individuals accrued from collaboration while leaving individual decision-making ability unaffected. This effect emerged because testosterone engendered more egocentric choices, manifest in an overweighting of one's own relative to others' judgements during joint decision-making. Our findings show that the biological control of social behaviour is dynamically regulated not only by modulators promoting, but also by those diminishing a propensity to collaborate