Cas9+ conditionally-immortalized macrophages as a tool for bacterial pathogenesis and beyond.

Macrophages play critical roles in immunity, development, tissue repair, and cancer, but studies of their function have been hampered by poorly-differentiated tumor cell lines and genetically-intractable primary cells. Here we report a facile system for genome editing in non-transformed macrophages by differentiating ER-Hoxb8 myeloid progenitors from Cas9-expressing transgenic mice. These conditionally immortalized macrophages (CIMs) retain characteristics of primary macrophages derived from the bone marrow yet allow for easy genetic manipulation and a virtually unlimited supply of cells. We demonstrate the utility of this system for dissection of host genetics during intracellular bacterial infection using two important human pathogens: Listeria monocytogenes and Mycobacterium tuberculosis

Functional evolution of the feeding system in rodents

The masticatory musculature of rodents has evolved to enable both gnawing at the incisors and chewing at the molars. In particular, the masseter muscle is highly specialised, having extended anteriorly to originate from the rostrum. All living rodents have achieved this masseteric expansion in one of three ways, known as the sciuromorph, hystricomorph and myomorph conditions. Here, we used finite element analysis (FEA) to investigate the biomechanical implications of these three morphologies, in a squirrel, guinea pig and rat. In particular, we wished to determine whether each of the three morphologies is better adapted for either gnawing or chewing. Results show that squirrels are more efficient at muscle-bite force transmission during incisor gnawing than guinea pigs, and that guinea pigs are more efficient at molar chewing than squirrels. This matches the known diet of nuts and seeds that squirrels gnaw, and of grasses that guinea pigs grind down with their molars. Surprisingly, results also indicate that rats are more efficient as well as more versatile feeders than both the squirrel and guinea pig. There seems to be no compromise in biting efficiency to accommodate the wider range of foodstuffs and the more general feeding behaviour adopted by rats. Our results show that the morphology of the skull and masticatory muscles have allowed squirrels to specialise as gnawers and guinea pigs as chewers, but that rats are high-performance generalists, which helps explain their overwhelming success as a group

Evasion of autophagy mediated by Rickettsia surface protein OmpB is critical for virulence.

Rickettsia are obligate intracellular bacteria that evade antimicrobial autophagy in the host cell cytosol by unknown mechanisms. Other cytosolic pathogens block different steps of autophagy targeting, including the initial step of polyubiquitin-coat formation. One mechanism of evasion is to mobilize actin to the bacterial surface. Here, we show that actin mobilization is insufficient to block autophagy recognition of the pathogen Rickettsia parkeri. Instead, R. parkeri employs outer membrane protein B (OmpB) to block ubiquitylation of the bacterial surface proteins, including OmpA, and subsequent recognition by autophagy receptors. OmpB is also required for the formation of a capsule-like layer. Although OmpB is dispensable for bacterial growth in endothelial cells, it is essential for R. parkeri to block autophagy in macrophages and to colonize mice because of its ability to promote autophagy evasion in immune cells. Our results indicate that OmpB acts as a protective shield to obstruct autophagy recognition, thereby revealing a distinctive bacterial mechanism to evade antimicrobial autophagy

Archaeological Investigations at Four San Antonio Missions: Mission Trails Underground Conversion Project

This report contains the results of archaeological work performed by the Center for Archaeological Research (CAR) at The University of Texas San Antonio (UTSA) for City Public Service (CPS). The archaeological investigation and monitoring for the Mission Trails Underground Conversion Project was carried out under Texas Historical Commission (THC) Permit Number: 2020, and the work was conducted at each of the four historical San Antonio missions which make up San Antonio Missions National Historical Park. The Mission Trails Underground Conversion Project was created to enhance and upgrade electrical and utility connections by replacing aboveground connections with underground connections at each ofthe four missions. Mission San Juan (41BX5)-In October of 1998, personnel from CAR began archaeological investigations at Mission San Juan Capistrano in advance of the proposed installation of underground utility lines for the Mission Trails Underground Conversion Project. The purpose of the investigation was to test for intact, buried cultural features in advance of trenching activities involved in the relocation of utility lines and connections. CAR archaeologists developed a plan regarding the area to be impacted which included ten 1 x 1-m excavations units and a series of twelve shovel tests along the proposed path of the utility trench. Excavation units uncovered evidence of existing buildings, wall foundations and Colonial-period flooring episodes, revealing new data west of the known structures and not previously accounted for. Further excavations are warranted to determine the nature of features such as a Colonial-period wall revealed near one of the excavation units. This material discovered by the archaeological investigations resulted in the altering of the projected utility trench alignment to avoid disturbing deposits. Further archaeological investigations are recommended for the newly revealed features. Mission San Jose (41BX3)- In January of 1999, personnel from CAR began archaeological investigations at Mission San Jose y San Miguel de Aguayo in advance of the proposed installation of underground utility lines for the Mission Trails Underground Conversion Project. The purpose of the investigation was to test for intact, buried cultural features in advance of trenching activities involved in the relocation of utility lines and connections. CAR archaeologists conducted a series of shovel tests along the center-lines of the proposed utility trenches to identifY areas of possible impact to intact Spanish Colonial cultural materials. In addition to shovel testing, monitoring of the trenching was conducted in areas where the possibility of impacting cultural material was high. An early-twentieth century trash midden was encountered during monitoring of the trench where it paralleled the Service Drive in the northwest comer of the Mission compound. This material did not warrant altering the course of the trench, and no further archaeological work is recommended at this time. Mission Concepcion (41BX12)- In January of 1999, personnel from CAR began archaeological investigations at Mission Nuestra Senora de la Purisima Concepcion as part of the Mission Trails Underground Conversion Project. A construction crew, while excavating a manhole designed to provide access to underground electrical utilities, had unearthed a large number of animal bones. After concluding a large feature had been unexpectedly impacted, CAR conducted test excavations to ascertain the nature of this feature. It was determined to be a section of an acequia which had been refilled during the Colonial period with construction debris and trash, including a large number of animal bones. Test units and the original manhole excavations allowed for a profile of the acequia to be constructed. This information combined with previous archaeological work and historical research, allowed a tentative proposal on the sequence of acequia construction and re-routing for the Concepcion mission. Monitoring of other underground work conducted in the area resulted in the discovery of a location of a Colonial-period wall, southeast of the Visitors\u27 Center, which may be a portion of the original south wall of the mission. It is recommended that if any further work is required, consideration should be given to testing and/or monitoring to reduce the risk of negative impact to historic remains. Mission Espada (41BX4)- In July of 1999, personnel from CAR began archaeological investigations at Mission San Francisco de la Espada in advance of the proposed installation of underground utility lines for the Mission Trails Underground Conversion Project. From initial discussions, it was noted that the proximity of the proposed route of the underground utility trench could have an impact on the old compound walls of the mission and the path was altered to avoid any possible impact. A series of thirteen shovel tests were conducted in advance along the altered path of the proposed utility trench as it progressed from the southwest comer of the mission compound across the dry irrigation ditch or acequia to the northwest. Monitoring of the trenching activities followed with no significant deposits being exposed. No further work is recommended within the project area, but if future construction is to occur in the area between the utility trench and the Church at Mission Espada, monitoring should then be considered

On the robustness of H-deficient post-AGB tracks

We analyze the robustness of H--deficient post--AGB tracks regarding previous evolution of their progenitor stars and the constitutive physics of the remnants. Our motivation is a recent suggestion of Werner & Herwig (2006) that previous evolution should be important in shaping the final post--AGB track and the persisting discrepancy between asteroseismological and spectroscopical mass determinations. This work is thus complementary to our previous work (Miller Bertolami & Althaus 2006) and intends to shed some light on the uncertainty behind the evolutionary tracks presented there. We compute full evolutionary models for PG1159 stars taking into account different extramixing (overshooting) efficiencies and lifetimes on the TP-AGB during the progenitor evolution. We also assess the effect of possible differences in the opacities and equation of state by artificially changing them before the PG1159 stage. Also comparisons are made with the few H-deficient post--AGB tracks available in the literature. Contrary to our expectations, we found that previous evolution is not a main factor in shaping H--deficient post--AGB tracks. Interestingly enough, we find that only an increase of $\sim50$ in the intershell opacities at high effective temperatures may affect the tracks as to reconcile spectroscopic and asteroseismologic mass determinations. This forces us to conclude that our previous tracks (Miller Bertolami & Althaus 2006) are robust enough as to be used for spectroscopic mass determinations, unless opacities in the intershell region are substantially different. Our results, then, call for an analysis of possible systematics in the usually adopted asteroseismological mass determination methods.Comment: Accepted for publication in Astronomy and Astrophysc

Digital dissection of the model organism Xenopus laevis using contrast-enhanced computed tomography

The African clawed frog, Xenopus laevis, is one of the most widely used model organisms in biological research. However, the most recent anatomical description of X. laevis was produced nearly a century ago. Compared with other anurans, pipid frogs – including X. laevis – exhibit numerous unusual morphological features; thus, anatomical descriptions of more ‘typical’ frogs do not detail many aspects of X. laevis skeletal and soft‐tissue morphology. The relatively new method of using iodine‐based agents to stain soft tissues prior to high‐resolution X‐ray imaging has several advantages over gross dissection, such as enabling dissection of very small and fragile specimens, and preserving the three‐dimensional topology of anatomical structures. Here, we use contrast‐enhanced computed tomography to produce a high‐resolution three‐dimensional digital dissection of a post‐metamorphic X. laevis to successfully visualize: skeletal and muscular anatomy; the nervous, respiratory, digestive, excretory and reproductive systems; and the major sense organs. Our digital dissection updates and supplements previous anatomical descriptions of this key model organism, and we present the three‐dimensional data as interactive portable document format (PDF) files that are easily accessible and freely available for research and educational purposes. The data presented here hold enormous potential for applications beyond descriptive purposes, particularly for biological researchers using this taxon as a model organism, comparative anatomy and biomechanical modelling