Passive Polarized Vision for Autonomous Vehicles: A Review.

This review article aims to address common research questions in passive polarized vision for robotics. What kind of polarization sensing can we embed into robots? Can we find our geolocation and true north heading by detecting light scattering from the sky as animals do? How should polarization images be related to the physical properties of reflecting surfaces in the context of scene understanding? This review article is divided into three main sections to address these questions, as well as to assist roboticists in identifying future directions in passive polarized vision for robotics. After an introduction, three key interconnected areas will be covered in the following sections: embedded polarization imaging; polarized vision for robotics navigation; and polarized vision for scene understanding. We will then discuss how polarized vision, a type of vision commonly used in the animal kingdom, should be implemented in robotics; this type of vision has not yet been exploited in robotics service. Passive polarized vision could be a supplemental perceptive modality of localization techniques to complement and reinforce more conventional ones.

Shaping the energy curves of a servomotor-based hexapod robot.

The advantageous versatility of hexapod robots is often accompanied by high power consumption, while animals have evolved an energy efficient locomotion. However, there are a lack of methods able to compare and apply animals' energetic optimizations to robots. In this study, we applied our method to a full servomotor-based hexapod robot to evaluate its energetic performance. Using an existing framework based on the laws of thermodynamics, we estimated four metrics using a dedicated test bench and a simulated robotic leg. We analyzed the characteristics of a single leg to shape the energetic profile of the full robot to a given task. Energy saving is improved by 10% through continuous duty factor adjustment with a 192% increase in power maximization. Moreover, adjusting the robot's velocity by the step length and associating this with gait switching, reduces the power loss by a further 10% at low-speed locomotion. However, unlike in animals, only one unique optimal operating point has been revealed, which is a disadvantage caused by the low energetic efficiency of servomotor-based hexapods. Thus, these legged robots are severely limited in their capacity to optimally adjust their locomotion to various tasks-a counter-intuitive conclusion for a supposedly versatile robot.

Evaluation of Ki-67, goblet cell and MUC2 mucin RNA expression in dogs with lymphoplasmacytic and granulomatous colitis.

Intestinal mucus barrier disruption may occur with chronic inflammatory enteropathies. The lack of studies evaluating mucus health in dogs with chronic colitis arises from inherent challenges with assessment of the intestinal mucus layer. It is therefore unknown if reduced goblet cell (GBC) numbers and/or mucin 2 (MUC2) expression, which are responsible for mucus production and secretion, correlate with inflammation severity in dogs with granulomatous colitis (GC) or lymphocytic-plasmacytic colitis (LPC). It is undetermined if Ki-67 immunoreactivity, which has been evaluated in dogs with small intestinal inflammation, similarly correlates to histologic severity in GC and LPC. Study objectives included comparing Ki-67 immunoreactivity, GBC population and MUC2 expression in dogs with GC, LPC and non-inflamed colon; and exploring the use of ribonucleic acid (RNAscope®) in-situ hybridization (ISH) to evaluate MUC2 expression in canine colon. Formalin-fixed endoscopic colonic biopsies were obtained from 48 dogs over an eight-year period. A blinded pathologist reviewed all biopsies. Dogs were classified into the GC (n=19), LPC (n=19) or no colitis (NC) (n=10) group based on final histopathological diagnosis. Ki-67 immunohistochemistry, Alcian-Blue/PAS staining to highlight GBCs, and RNAscope® ISH using customized canine MUC2-targeted probes were performed. At least five microscopic fields per dog were selected to measure Ki-67 labelling index (KI67%), GBC staining percentage (GBC%) and MUC2 expression (MUC2%) using image analysis software. Spearman's correlation coefficients were used to determine associations between World Small Animal Veterinary Association histologic score (WHS) and measured variables. Linear regression models were used to compare relationships between WHS with KI67%, GBC%, and MUC2%; and between GBC% and MUC2%. Median WHS was highest in dogs with GC. Median KI67% normalised to WHS was highest in the NC group (6.69%; range, 1.70-23.60%). Median GBC% did not correlate with colonic inflammation overall. Median MUC2% normalised to WHS in the NC group (10.02%; range, 3.05-39.09%) was two- and three-fold higher than in the GC and LPC groups respectively. With increased colonic inflammation, despite minimal changes in GBC% overall, MUC2 expression markedly declined in the LPC group (-27.4%; 95%-CI, -49.8, 5.9%) and mildly declined in the GC and NC groups. Granulomatous colitis and LPC likely involve different pathways regulating MUC2 expression. Decreased MUC2 gene expression is observed in dogs with chronic colitis compared to dogs without colonic signs. Changes in MUC2 expression appear influenced by GBC activity rather than quantity in GC and LPC.

Kinetically-Controlled Ni-Catalyzed Direct Carboxylation of Unactivated Secondary Alkyl Bromides without Chain Walking.

Herein, we report the direct carboxylation of unactivated secondary alkyl bromides enabled by the merger of photoredox and nickel catalysis, a previously inaccessible endeavor in the carboxylation arena. Site-selectivity is dictated by a kinetically controlled insertion of CO2 at the initial C(sp3)-Br site by the rapid formation of Ni(I)-alkyl species, thus avoiding undesired ß-hydride elimination and chain-walking processes. Preliminary mechanistic experiments reveal the subtleties of stereoelectronic effects for guiding the reactivity and site-selectivity.

Nodular hyperplasia of lymphoglandular complexes in dogs: A potential diagnostic pitfall for rectal masses.

Lymphoglandular complexes are components of the gut-associated lymphoid tissue that are characterized by submucosal lymphoid aggregates invested by projections of mucosal epithelium. Reports of pathology involving these structures are rare in both human and veterinary literature. Here, the authors report 2 cases of rectal masses excised from dogs following a period of tenesmus and hematochezia. In both animals, the masses were composed of lymphoid tissue closely encompassing tubuloacinar structures. Immunohistochemistry and polymerase chain reaction antigen receptor rearrangement testing demonstrated that the lymphoid population was polyclonal, comprising T and B cells arranged in loosely follicular aggregates centered on the epithelial foci. In light of these findings, a diagnosis of lymphoglandular complex nodular hyperplasia was reported. To the authors' knowledge, this is the first report of this condition in dogs.

SkyPole-A method for locating the north celestial pole from skylight polarization patterns.

True north can be determined on Earth by three means: magnetic compasses, stars, and via the global navigation satellite systems (GNSS), each of which has its own drawbacks. GNSS are sensitive to jamming and spoofing, magnetic compasses are vulnerable to magnetic interferences, and the stars can be used only at night with a clear sky. As an alternative to these methods, nature-inspired navigational cues are of particular interest. Celestial polarization, which is used by insects such as Cataglyphis ants, can provide useful directional cues. Migrating birds calibrate their magnetic compasses by observing the celestial rotation at night. By combining these cues, we have developed a bioinspired optical method for finding the celestial pole during the daytime. This method, which we have named SkyPole, is based on the rotation of the skylight polarization pattern. A polarimetric camera was used to measure the degree of skylight polarization rotating with the Sun. Image difference processes were then applied to the time-varying measurements in order to determine the north celestial pole's position and thus the observer's latitude and bearing with respect to the true north.

Biomarkers of endothelial activation and inflammation in dogs with organ dysfunction secondary to sepsis.

Introduction: Alteration in endothelial function during sepsis is thought to play a key role in the progression of organ failure. We herein compared plasma concentrations of endothelial activation biomarkers vascular endothelial growth factor (VEGF), hyaluronan (HA), plasminogen activator inhibitor-1 (PAI-1) and von Willebrand factor (vWF), as well as inflammatory mediator concentrations (IL-6, IL-8, IL-10, C-reactive protein and monocyte chemoattractant protein-1) in dogs with sepsis to healthy dogs. Methods: This study was a multicenter observational clinical trial conducted at two university teaching hospitals from February 2016 until July 2017. The study included 18 client-owned dogs hospitalized with sepsis and at least one distant organ dysfunction, as well as 20 healthy dogs. Plasma biomarker concentrations were measured using ELISA. Severity of illness in dogs with sepsis was calculated using the 5-variable acute physiologic and laboratory evaluation (APPLEFAST) score. Biomarker concentrations were compared between septic and healthy dogs using linear models. Results: Septic peritonitis was the most frequent source of sepsis (11/18; 61%), followed by pneumonia (4/18; 22%). Ten dogs (56%) had only 1 organ dysfunction, whereas 3 dogs (17%) had 2, 3 (17%) had 3, 1 (6%) had 4 and 1 (6%) had 5 organ dysfunctions. The median APPLEFAST score in the septic dogs was 28.5 (Q1-Q3, 24-31). Mean plasma concentrations of all endothelial and inflammatory biomarkers, except vWF, were higher in the sepsis cohort than in controls. The mean endothelial biomarker concentrations in the septic cohort ranged from ~2.7-fold higher for HA (difference in means; 118.2 ng/mL, 95% credible limit; 44.5-221.7) to ~150-fold for VEGF (difference in means; 76.6 pg./mL, 95% credible limit; 33.0-143.4), compared to the healthy cohort. Fifteen dogs with sepsis (83%) died; 7 (46%) were euthanized and 8 (53%) died during hospitalization. Conclusion: Dogs with naturally occurring sepsis and organ dysfunction had higher mean concentrations of biomarkers of endothelial activation and inflammation compared to healthy dogs, broadening our understanding of the pathophysiology of sepsis secondary to endothelial dysfunction.

OprF Impacts Pseudomonas aeruginosa Biofilm Matrix eDNA Levels in a Nutrient-Dependent Manner.

The biofilm matrix is composed of exopolysaccharides, eDNA, membrane vesicles, and proteins. While proteomic analyses have identified numerous matrix proteins, their functions in the biofilm remain understudied compared to the other biofilm components. In the Pseudomonas aeruginosa biofilm, several studies have identified OprF as an abundant matrix protein and, more specifically, as a component of biofilm membrane vesicles. OprF is a major outer membrane porin of P. aeruginosa cells. However, current data describing the effects of OprF in the P. aeruginosa biofilm are limited. Here, we identify a nutrient-dependent effect of OprF in static biofilms, whereby ΔoprF cells form significantly less biofilm than wild type when grown in media containing glucose or low sodium chloride concentrations. Interestingly, this biofilm defect occurs during late static biofilm formation and is not dependent on the production of PQS, which is responsible for outer membrane vesicle production. Furthermore, while biofilms lacking OprF contain approximately 60% less total biomass than those of wild type, the number of cells in these two biofilms is equivalent. We demonstrate that P. aeruginosa ΔoprF biofilms with reduced biofilm biomass contain less eDNA than wild-type biofilms. These results suggest that the nutrient-dependent effect of OprF is involved in the maintenance of P. aeruginosa biofilms by retaining eDNA in the matrix. IMPORTANCE Many pathogens form biofilms, which are bacterial communities encased in an extracellular matrix that protects them against antibacterial treatments. The roles of several matrix components of the opportunistic pathogen Pseudomonas aeruginosa have been characterized. However, the effects of P. aeruginosa matrix proteins remain understudied and are untapped potential targets for antibiofilm treatments. Here, we describe a conditional effect of the abundant matrix protein OprF on late-stage P. aeruginosa biofilms. A ΔoprF strain formed significantly less biofilm in low sodium chloride or with glucose. Interestingly, the defective ΔoprF biofilms did not exhibit fewer resident cells but contained significantly less extracellular DNA (eDNA) than wild type. These results suggest that OprF is involved in matrix eDNA retention in biofilms.

Identification of novel p-cresol inhibitors that reduce Clostridioides difficile's ability to compete with species of the gut microbiome.

Treatment of Clostridioides difficile infection (CDI) is expensive and complex, with a high proportion of patients suffering infection relapse (20-35%), and some having multiple relapses. A healthy, unperturbed gut microbiome provides colonisation resistance against CDI through competition for nutrients and space. However, antibiotic consumption can disturb the gut microbiota (dysbiosis) resulting in the loss of colonisation resistance allowing C. difficile to colonise and establish infection. A unique feature of C. difficile is the production of high concentrations of the antimicrobial compound para-cresol, which provides the bacterium with a competitive advantage over other bacteria found in the gut. p-cresol is produced by the conversion of para-Hydroxyphenylacetic acid (p-HPA) by the HpdBCA enzyme complex. In this study, we have identified several promising inhibitors of HpdBCA decarboxylase, which reduce p-cresol production and render C. difficile less able to compete with a gut dwelling Escherichia coli strain. We demonstrate that the lead compound, 4-Hydroxyphenylacetonitrile, reduced p-cresol production by 99.0 ± 0.4%, whereas 4-Hydroxyphenylacetamide, a previously identified inhibitor of HpdBCA decarboxylase, only reduced p-cresol production by 54.9 ± 13.5%. To interpret efficacy of these first-generation inhibitors, we undertook molecular docking studies that predict the binding mode for these compounds. Notably, the predicted binding energy correlated well with the experimentally determined level of inhibition, providing a molecular basis for the differences in efficacy between the compounds. This study has identified promising p-cresol production inhibitors whose development could lead to beneficial therapeutics that help to restore colonisation resistance and therefore reduce the likelihood of CDI relapse.

Gut microbiota fermentation profiles of pre-digested mycoprotein (Quorn) using faecal batch cultures in vitro: a preliminary study.

High-fibre diets are beneficial for many health outcomes via a wide range of mechanisms including gut microbiota fermentation-derived short-chain fatty acid (SCFAs) production. Mycoprotein (marketed as Quorn) is a food high in fibre (>6 g/100 g wet weight (ww)) and protein (13 g/100 g ww) which has been shown to have positive effects on glycemic control and appetite in humans. Nevertheless, the mechanisms underpinning this are poorly understood. Here, we investigate the changes in gut microbiota α- and ß-diversity, pH and SCFAs production in faecal batch cultures supplemented with pre-digested mycoprotein (Quorn), soy, chicken and control (unsupplemented) using eight fresh stools from healthy donors. The results showed that pre-digested mycoprotein did not alter pH (p = .896), α- or ß-diversity of the gut microbiota when compared to the control, soy, and chicken. Nevertheless, chicken led to a significant increase in total SCFAs post-24 h vs. control (+57.07 mmol/L, p = .01). In particular, propionate increased when compared to soy (+19.59 mmol/L, p = .03) and the control (+23.19 mmol/L, p < .01). No other differences in SCFAs were detected. In conclusion, pre-digested mycoprotein was not fermented in vitro by healthy gut microbiota in the settings of this experiment.

Single-particle cryo-EM analysis of the shell architecture and internal organization of an intact α-carboxysome.

Carboxysomes are proteinaceous bacterial microcompartments that sequester the key enzymes for carbon fixation in cyanobacteria and some proteobacteria. They consist of a virus-like icosahedral shell, encapsulating several enzymes, including ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCO), responsible for the first step of the Calvin-Benson-Bassham cycle. Despite their significance in carbon fixation and great bioengineering potentials, the structural understanding of native carboxysomes is currently limited to low-resolution studies. Here, we report the characterization of a native α-carboxysome from a marine cyanobacterium by single-particle cryoelectron microscopy (cryo-EM). We have determined the structure of its RuBisCO enzyme, and obtained low-resolution maps of its icosahedral shell, and of its concentric interior organization. Using integrative modeling approaches, we have proposed a complete atomic model of an intact carboxysome, providing insight into its organization and assembly. This is critical for a better understanding of the carbon fixation mechanism and toward repurposing carboxysomes in synthetic biology for biotechnological applications.

Honeybees Use Multiple Invariants to Control Their Altitude.

How do bees perceive altitude changes so as to produce safe displacements within their environment? It has been proved that humans use invariants, but this concept remains little-known within the entomology community. The use of a single invariant, the optical speed rate of change, has been extensively demonstrated in bees in a ground-following task. Recently, it has been demonstrated that another invariant, the splay angle rate of change, could also be used by bees to adjust their altitude. This study aims to understand how bees use these invariants when they are available simultaneously. This issue has been addressed using an experimental setup providing discordant information to bees. We have shown that when the two invariants were available, bees performed ground-following tasks relying primarily on optical speed rate of change. Conversely, when optical speed rate of change was less easily accessible, splay angle rate of change was prioritized, unless the bees perceive danger. Taken together, these results illustrate how the joint use of several invariants allows bees to produce adaptive behaviors.

Visual augmentation of deck-landing-ability improves helicopter ship landing decisions.

When attempting to land on a ship deck tossed by the sea, helicopter pilots must make sure that the helicopter can develop sufficient lift to be able to safely touchdown. This reminder of affordance theory led us to model and study the affordance of deck-landing-ability, which defines whether it is possible to land safely on a ship deck depending on the helicopter's available lift and the ship's deck heave movements. Two groups of participants with no piloting experience using a laptop helicopter simulator attempted to land either a low-lifter or a heavy-lifter helicopter on a virtual ship deck by either triggering a pre-programmed lift serving as the descent law if it was deemed possible to land, or aborting the deck-landing maneuver. The deck-landing-ability was manipulated by varying the helicopter's initial altitude and the ship's heave phase between trials. We designed a visual augmentation making visible the deck-landing-ability, and thus enabling participants to maximize the safety of their deck-landing attempts and reduce the number of unsafe deck-landing. The visual augmentation presented here was perceived by participants as a means of facilitating this decision-making process. The benefits were found to have originated from the clear-cut distinction it helped them to make between safe and unsafe deck-landing windows and the display of the optimal time for initiating the landing.

Alternating Metal-Ligand Coordination Improves Electrocatalytic CO2 Reduction by a Mononuclear Ru Catalyst.

Molecular electrocatalysts for CO2 -to-CO conversion often operate at large overpotentials, due to the large barrier for C-O bond cleavage. Illustrated with ruthenium polypyridyl catalysts, we herein propose a mechanistic route that involves one metal center that acts as both Lewis base and Lewis acid at different stages of the catalytic cycle, by density functional theory in corroboration with experimental FTIR. The nucleophilic character of the Ru center manifests itself in the initial attack on CO2 to form [Ru-CO2 ]0 , while its electrophilic character allows for the formation of a 5-membered metallacyclic intermediate, [Ru-CO2 CO2 ]0,c , by addition of a second CO2 molecule and intramolecular cyclization. The calculated activation barrier for C-O bond cleavage via the metallacycle is decreased by 34.9 kcal mol-1 as compared to the non-cyclic adduct in the two electron reduced state of complex 1. Such metallacyclic intermediates in electrocatalytic CO2 reduction offer a new design feature that can be implemented consciously in future catalyst designs.

A single parameter can predict surfactant impairment of superhydrophobic drag reduction.

Recent experimental and computational investigations have shown that trace amounts of surfactants, unavoidable in practice, can critically impair the drag reduction of superhydrophobic surfaces (SHSs), by inducing Marangoni stresses at the air-liquid interface. However, predictive models for realistic SHS geometries do not yet exist, which has limited the understanding and mitigation of these adverse surfactant effects. To address this issue, we derive a model for laminar, three-dimensional flow over SHS gratings as a function of geometry and soluble surfactant properties, which together encompass 10 dimensionless groups. We establish that the grating length g is the key geometric parameter and predict that the ratio between actual and surfactant-free slip increases with g2. Guided by our model, we perform synergistic numerical simulations and microfluidic experiments, finding good agreement with the theory as we vary surfactant type and SHS geometry. Our model also enables the estimation, based on velocity measurements, of a priori unknown properties of surfactants inherently present in microfluidic systems. For SHSs, we show that surfactant effects can be predicted by a single parameter, representing the ratio between the grating length and the interface length scale beyond which the flow mobilizes the air-water interface. This mobilization length is more sensitive to the surfactant chemistry than to its concentration, such that even trace-level contaminants may significantly increase drag if they are highly surface active. These findings advance the fundamental understanding of realistic interfacial flows and provide practical strategies to maximize superhydrophobic drag reduction.

3D Printable Hybrid Gel Made of Polymer Surface-Modified Cellulose Nanofibrils Prepared by Surface-Initiated Controlled Radical Polymerization (SI-SET-LRP) and Upconversion Luminescent Nanoparticles.

A cellulose nanofibril-based hybrid gel material was developed by grafting the polymerized stearyl acrylate (PSA) and upconversion nanoparticles (UCNPs) onto cellulose nanofibrils (CNFs) via Cu0-mediated radical polymerization (SET-LRP) to create a highly cross-linked CNF system. A two-step strategy was exploited to surface-exchange the ligand of the UCNPs from a hydrophobic ligand (oleic acid) to a hydrophilic small-molecule ligand (2-acrylamido-2-methyl-1-propanesulfonic acid, AMPS) and therefore be suitable for SET-LRP. The characteristics and properties of the hybrid material (UCNP-PSA-CNF) were monitored by Fourier transform infrared (FTIR) spectroscopy, thermogravimetric analysis (TGA), rheology, X-ray diffraction (XRD), and microscopic analysis. Those characterization techniques prove the efficient modification of the CNF, with the presence of 1.8% UCNPs. The luminescence measurement was carried out using a homebuilt confocal microscope with a 980 nm laser source. The nanostructure of UCNPs and their incorporated CNF species were measured by small-angle X-ray scattering (SAXS). In addition, this CNF-based hybrid gel has decisive rheological properties, such as good viscoelasticity (loss tangent was below 0.35 for the UCNP-PSA-CNF gel, while the PSA-CNF gel reached the highest value of 0.42), shear-thinning behavior, and shape retention, and was successfully applied to three-dimensional (3D) gel printing throughout various 3D print models.

Stress-induced reversible cell-cycle arrest requires PRC2/PRC1-mediated control of mitophagy in Drosophila germline stem cells and human iPSCs.

Following acute genotoxic stress, both normal and tumorous stem cells can undergo cell-cycle arrest to avoid apoptosis and later re-enter the cell cycle to regenerate daughter cells. However, the mechanism of protective, reversible proliferative arrest, "quiescence," remains unresolved. Here, we show that mitophagy is a prerequisite for reversible quiescence in both irradiated Drosophila germline stem cells (GSCs) and human induced pluripotent stem cells (hiPSCs). In GSCs, mitofission (Drp1) or mitophagy (Pink1/Parkin) genes are essential to enter quiescence, whereas mitochondrial biogenesis (PGC1α) or fusion (Mfn2) genes are crucial for exiting quiescence. Furthermore, mitophagy-dependent quiescence lies downstream of mTOR- and PRC2-mediated repression and relies on the mitochondrial pool of cyclin E. Mitophagy-dependent reduction of cyclin E in GSCs and in hiPSCs during mTOR inhibition prevents the usual G1/S transition, pushing the cells toward reversible quiescence (G0). This alternative method of G1/S control may present new opportunities for therapeutic purposes.

An experimental setup for decoupling optical invariants in honeybees' altitude control.

Bees outperform pilots in navigational tasks, despite having 100,000 times fewer neurons. It is commonly accepted in the literature that optic flow is a key parameter used by flying insects to control their altitude. The ambition of the present work was to design an innovative experimental setup that would make it possible to determine whether bees could rely simultaneously on several optical invariants, as pilots do. We designed a flight tunnel to enable manipulation of an optical invariant, the Splay Angle Rate of Change (SARC) and the restriction of the Optical Speed Rate of Change (OSRC) in the optic flow. It allows us to determine if bees use the SARC to control their altitude and to identify the integration process combining these two optical invariants. Access to the OSRC can be restricted by using different textures. The SARC can be biased thanks to motorized rods. This device allows to record bees' trajectories in different visual configurations, including impoverished conditions and conditions containing contradictory information. The comparative analysis of the recorded trajectories provides first time evidence of SARC use in a ground-following task by a non-human animal. This new tunnel allows a precise experimental control of the visual environment in ecological experimental conditions. Therefore, it could pave the way for a new type of ecologically based studies examining the simultaneous use of several information sources for navigation by flying insects.

Structure and Assembly of the Bacterial Flagellum.

The bacterial flagellum is a large macromolecular assembly that acts as propeller, providing motility through the rotation of a long extracellular filament. It is composed of over 20 different proteins, many of them highly oligomeric. Accordingly, it has attracted a huge amount of interest amongst researchers and the wider public alike. Nonetheless, most of its molecular details had long remained elusive.This however has changed recently, with the emergence of cryo-EM to determine the structure of protein assemblies at near-atomic resolution. Within a few years, the atomic details of most of the flagellar components have been elucidated, revealing not only its overall architecture but also the molecular details of its rotation mechanism. However, many questions remained unaddressed, notably on the complexity of the assembly of such an intricate machinery.In this chapter, we review the current state of our understanding of the bacterial flagellum structure, focusing on the recent development from cryo-EM. We also highlight the various elements that still remain to be fully characterized. Finally, we summarize the existing model for flagellum assembly and discuss some of the outstanding questions that are still pending in our understanding of the diversity of assembly pathways.

Cryo-EM of the injectisome and type III secretion systems.

Double-membrane-spanning protein complexes, such as the T3SS, had long presented an intractable challenge for structural biology. As a consequence, until a few years ago, our molecular understanding of this fascinating complex was limited to composite models, consisting of structures of isolated domains, positioned within the overall complex. Most of the membrane-embedded components remained completely uncharacterized. In recent years, the emergence of cryo-electron microscopy (cryo-EM) as a method for determining protein structures to high resolution, has be transformative to our capacity to understand the architecture of this complex, and its mechanism of substrate transport. In this review, we summarize the recent structures of the various T3SS components, determined by cryo-EM, and highlight the regions of the complex that remain to be characterized. We also discuss the recent structural insights into the mechanism of effector transport through the T3SS. Finally, we highlight some of the challenges that remain to be tackled.