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Abstract Unbiased population density estimates are critical for ecological research and wildlife management but are often difficult to obtain. Researchers use a variety of sampling and statistical methods to generate estimates of density, but few studies have compared estimates across methods. During 2016–2017, we surveyed Canada lynx (Lynx canadensis) in southwestern Yukon Territory, Canada, using track transect counts, hair snares, camera traps, live traps, and Global Positioning System (GPS) collars. From these data, we estimated lynx density with two linearly scaled count methods, one spatial mark–recapture method, three spatial mark–resight methods, and one cumulative‐time method. We found up to fivefold variation in point density estimates despite adhering to method requirements and assumptions in a manner consistent with other studies. Our results highlight the dependency of density estimates on sampling process and model assumptions and demonstrate the value of careful and unbiased sampling design. Further research is needed to fully assess the accuracy and limitations of the many wildlife density estimation methods that are currently in use so that techniques can be appropriately applied to typical study systems and species.

Animals exhibit dynamic movement and activity in response to environmental variation including changes in reproductive opportunities, predation risk, or food availability. Yet, it remains unclear which factors are primary in affecting animal movement, and whether the relative importance of these factors are consistent through time. We tracked snowshoe hares (Lepus americanus) using GPS telemetry during eight summers spanning a hare population cycle (2015–2022) in southwestern Yukon, Canada, to determine associations between environmental variation and hare movement and home range size. Hare density varied 25-fold during the study and home range size increased markedly during low hare density, especially for males. Both sexes retained similar core space use and linearity of movements, but at low densities males had greater and more variable movement rates and time spent travelling. Trail cameras revealed that annual changes in hare movement were also correlated with relative abundance of lynx (Lynx canadensis) and coyotes (Canis latrans). However, hare detection rates within a season were not closely associated with seasonal variation in predator detection. Observed differences between male and female hares in some metrics highlighted that different life histories and reproductive behavior are likely the main drivers of hare movement dynamics. Therefore, fitness rewards associated with successful mate search and reproduction appear to outweigh risks associated with increased movement, even in highly variable environments where costs of prioritizing reproduction-related activities are notably high and variable.

Host-pathogen interactions play a critical role in infectious diseases, and understanding the underlying mechanisms is vital for developing effective therapeutic strategies. The visualization and characterization of bacterial proteins within host cells is key to unraveling the dynamics of these interactions. Various protein labeling strategies have emerged as powerful tools for studying host-pathogen interactions, enabling the tracking, localization, and functional analysis of bacterial proteins in real-time. However, the labeling and localization of Salmonella secreted type III secretion system (T3SS) effectors in host cells poses technical challenges. Conventional methods disrupt effector stoichiometry and often result in non-specific staining. Bulky fluorescent protein fusions interfere with effector secretion, while other tagging systems such as 4Cys-FLaSH/Split-GFP suffer from low labeling specificity and a poor signal-to-noise ratio. Recent advances in state-of-the-art techniques have augmented the existing toolkit for monitoring the translocation and dynamics of bacterial effectors. This comprehensive review delves into the bacterial protein labeling strategies and their application in imaging host-pathogen interactions. Lastly, we explore the obstacles faced and potential pathways forward in the realm of protein labeling strategies for visualizing interactions between hosts and pathogens.

The transcriptional regulator SsrB acts as a switch between virulent and biofilm lifestyles of non-typhoidal Salmonella enterica serovar Typhimurium. During infection, phosphorylated SsrB activates genes on Salmonella Pathogenicity Island-2 (SPI-2) essential for survival and replication within the macrophage. Low pH inside the vacuole is a key inducer of expression and SsrB activation. Previous studies demonstrated an increase in SsrB protein levels and DNA-binding affinity at low pH; the molecular basis was unknown (Liew et al., 2019). This study elucidates its underlying mechanism and in vivo significance. Employing single-molecule and transcriptional assays, we report that the SsrB DNA-binding domain alone (SsrBc) is insufficient to induce acid pH-sensitivity. Instead, His12, a conserved residue in the receiver domain confers pH sensitivity to SsrB allosterically. Acid-dependent DNA binding was highly cooperative, suggesting a new configuration of SsrB oligomers at SPI-2-dependent promoters. His12 also plays a role in SsrB phosphorylation; substituting His12 reduced phosphorylation at neutral pH and abolished pH-dependent differences. Failure to flip the switch in SsrB renders Salmonella avirulent and represents a potential means of controlling virulence.

The boreal forest, the world’s largest terrestrial biome, is undergoing dramatic changes owing to anthropogenic stressors, including those of climate change. To track terrestrial ecosystem changes through space and time, robust monitoring programs are needed that survey a variety of ecosystem constituents. We monitored white spruce (Picea glauca) cone crops, berry (Empetrum nigrum, Shepherdia canadensis) production, above-ground mushroom abundance, and the abundance of small mammals (Clethrionomys rutilus, Peromyscus maniculatus), North American red squirrels (Tamiascirus hudsonicus), snowshoe hares (Lepus americanus), and carnivores (Lynx canadensis, Canis latrans, Vulpes vulpes, Martes americana, Mustela erminea) across 5 sites in the Yukon, Canada. Monitoring began in 1973 at Lhù’ààn Mân’ (Kluane Lake) and additional protocols were added until a complete sequence was fixed in 2005 at all 5 sites and continued until 2022. White spruce cone counts show mast years at 3–7-year intervals. Ground berries and soapberry counts were highly variable among sites and counts did not correlate among sites or between years for different species. Red-backed voles showed clear 3–4-year cycles at Kluane and probably at the Mayo and Watson Lake sites, but showed only annual cycles in Whitehorse and Faro. Snowshoe hares fluctuated in 9–10-year cycles in a travelling wave, peaking one year earlier at Watson Lake but in synchrony at all other sites, with no clear sign of peak density changing or cyclic attenuation over the last 50 years. Red squirrel numbers at Kluane exhibit marked inter-year variability, driven mainly by episodic white spruce cone crops and predation from Canada lynx and coyotes as hare densities undergo cyclic decline. Snow track counts to index mammalian predators have been conducted on our Kluane and Mayo sites, indicating that lynx numbers rise and fall with a 1–2-year lag at these two sites, tracking the hare cycle. Coyotes and lynx at Kluane peak together following the hare cycle, but coyote counts are also depressed during deep snow years. To summarize, we noted considerable inter-site variability in the population dynamics of many boreal forest ecosystem constituents, but the keystone species (snowshoe hare, Canada lynx) exhibit remarkably similar population trends across the region. We continue to monitor wildlife abundance, cone crops, berry production, and mushroom biomass to determine changes associated with increasing temperature and fluctuating rainfall. The Yukon boreal forest is changing as climate shifts, but the changes are slow, variable across sites, taxa specific, and of uncertain predictability.

The integrator complex is required for the synthesis of protein coding and non-coding RNA and contains the protein INTS13. Here, the authors find germline mutations in INTS13 in two families with oral facial digital syndrome and show that the mutation affects the c-terminal domain of the protein and disrupts cilliogenesis.

Summary: A simple and robust cell culture system is essential for generating authentic SARS-CoV-2 stocks for evaluation of viral pathogenicity, screening of antiviral compounds, and preparation of inactivated vaccines. Evidence suggests that Vero E6, a cell line commonly used in the field to grow SARS-CoV-2, does not support efficient propagation of new viral variants and triggers rapid cell culture adaptation of the virus. We generated a panel of 17 human cell lines overexpressing SARS-CoV-2 entry factors and tested their ability to support viral infection. Two cell lines, Caco-2/AT and HuH-6/AT, demonstrated exceptional susceptibility, yielding highly concentrated virus stocks. Notably, these cell lines were more sensitive than Vero E6 cells in recovering SARS-CoV-2 from clinical specimens. Further, Caco-2/AT cells provided a robust platform for producing genetically reliable recombinant SARS-CoV-2 through a reverse genetics system. These cellular models are a valuable tool for the study of SARS-CoV-2 and its continuously emerging variants.

Mosquito transmission of dengue viruses to humans starts with infection of skin resident cells at the biting site. There is great interest in identifying transmission-enhancing factors in mosquito saliva in order to counteract them. Here we report the discovery of high levels of the anti-immune subgenomic flaviviral RNA (sfRNA) in dengue virus 2-infected mosquito saliva. We established that sfRNA is present in saliva using three different methods: northern blot, RT-qPCR and RNA sequencing. We next show that salivary sfRNA is protected in detergent-sensitive compartments, likely extracellular vesicles. In support of this hypothesis, we visualized viral RNAs in vesicles in mosquito saliva and noted a marked enrichment of signal from 3’UTR sequences, which is consistent with the presence of sfRNA. Furthermore, we show that incubation with mosquito saliva containing higher sfRNA levels results in higher virus infectivity in a human hepatoma cell line and human primary dermal fibroblasts. Transfection of 3’UTR RNA prior to DENV2 infection inhibited type I and III interferon induction and signaling, and enhanced viral replication. Therefore, we posit that sfRNA present in salivary extracellular vesicles is delivered to cells at the biting site to inhibit innate immunity and enhance dengue virus transmission. Author summary Mosquitoes transmit many viruses, such as those that cause dengue, introducing them in our bodies with the saliva that is deposited in our skins when they bite us. Scientists who study biting behavior and properties of mosquito saliva have concluded that there may be components in saliva that promote virus infection. In this study we show that when dengue virus infects mosquitoes one can find in their saliva not only the expected dengue viruses but also a dengue virus product that can reduce our anti-virus defense systems. It happens that this product is what is known as a non-coding RNA, a class of molecules that have recently been shown to mediate important biological regulation. We propose that by introducing this RNA at the biting site dengue infected saliva prepares the terrain for an efficient infection and gives the virus an advantage in the first battle between it and our immune defenses.

Purpose: To quantify surgeon intuition, determine whether a surgeon’s prediction of outcomes after hip arthroscopy correlates with actual patient-reported outcomes (PRO), and identify differences in clinical judgment between expert and novice examiners. Methods: This prospective, longitudinal study was conducted at an academic medical center on adults undergoing primary hip arthroscopy for treatment of femoroacetabular impingement. A Surgeon Intuition and Prediction (SIP) score was completed preoperatively by an attending surgeon (expert) and physician assistant (novice). Baseline and postoperative outcome measures included legacy hip scores (e.g., Modified Harris Hip score) and Patient-Reported Outcomes Information System tools. Mean differences were assessed using t-tests. Generalized estimating equations assessed longitudinal changes. Pearson correlation coefficients (r) evaluated associations between SIP score and PRO scores. Results: Data from 98 patients (mean age 36 years, 67% female) with complete data sets at 12-month follow-up were analyzed. Weak-to-moderate strength correlations were seen between SIP score and PRO scores (r = 0.36 to r = 0.53) for pain, activity and physical function. Significant improvements were seen in all primary outcome measures at 6 and 12 months postoperatively when compared to baseline scores (P < .05), with about 50% to 80% of patients achieving the minimum clinically important difference and patient acceptable symptomatic state thresholds postoperatively. Conclusions: An experienced, high-volume hip arthroscopist had only weak-to-moderate ability to intuitively predict PRO. Surgical intuition and judgment were not superior in an expert examiner compared to a novice. Level of Evidence: Level III, retrospective comparative prognostic trial.

ABSTRACT Recently, highly transmissible severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants B.1.617.1 (Kappa), B.1.617.2 (Delta), and B.1.618 with mutations within the spike proteins were identified in India. The spike protein of Kappa contains the four mutations E154K, L452R, E484Q, and P681R, and Delta contains L452R, T478K, and P681R, while B.1.618 spike harbors mutations Δ145–146 and E484K. However, it remains unknown whether these variants have alterations in their entry efficiency, host tropism, and sensitivity to neutralizing antibodies as well as entry inhibitors. In this study, we found that Kappa, Delta, or B.1.618 spike uses human angiotensin-converting enzyme 2 (ACE2) with no or slightly increased efficiency, while it gains a significantly increased binding affinity with mouse, marmoset, and koala ACE2 orthologs, which exhibit limited binding with wild-type (WT) spike. Furthermore, the P681R mutation leads to enhanced spike cleavage, which could facilitate viral entry. In addition, Kappa, Delta, and B.1.618 exhibit a reduced sensitivity to neutralization by convalescent-phase sera due to the mutation E484Q, T478K, Δ145–146, or E484K, but remain sensitive to entry inhibitors such as ACE2-Ig decoy receptor. Collectively, our study revealed that enhanced human and mouse ACE2 receptor engagement, increased spike cleavage, and reduced sensitivity to neutralization antibodies of Kappa, Delta and B.1.618 may contribute to the rapid spread of these variants. Furthermore, our results also highlight that ACE2-Ig could be developed as a broad-spectrum antiviral strategy against SARS-CoV-2 variants. IMPORTANCE SARS-CoV-2, the causative agent of pandemic COVID-19, is rapidly evolving to be more transmissible and to exhibit evasive immune properties, compromising neutralization by antibodies from vaccinated individuals or convalescent-phase sera. Recently, SARS-CoV-2 variants B.1.617.1 (Kappa), B.1.617.2 (Delta), and B.1.618 with mutations within the spike proteins were identified in India. In this study, we examined cell entry efficiencies of Kappa, Delta, and B.1.618. In addition, the variants, especially the Delta variant, exhibited expanded capabilities to use mouse, marmoset, and koala ACE2 for entry. Convalescent sera from patients infected with nonvariants showed reduced neutralization titers among the Kappa, Delta, and B.1.618 variants. Furthermore, the variants remain sensitive to ACE2-Ig decoy receptor. Our study thus could facilitate understanding how variants have increased transmissibility and evasion of established immunity and also could highlight the use of an ACE2 decoy receptor as a broad-spectrum antiviral strategy against SARS-CoV-2 variants.

People who either use an upper limb prosthesis and/or have used services provided by a prosthetic rehabilitation centre, experience limitations of currently available prosthetic devices. Collaboration between academia and a broad range of stakeholders, can lead to the development of solutions that address peoples' needs. By doing so, the rate of prosthetic device abandonment can decrease. Co-creation is an approach that can enable collaboration of this nature to occur throughout the research process. We present findings of a co-creation project that gained user perspectives from a user survey, and a subsequent workshop involving: people who use an upper limb prosthesis and/or have experienced care services (users), academics, industry experts, charity executives, and clinicians. The survey invited users to prioritise six themes, which academia, clinicians, and industry should focus on over the next decade. The prioritisation of the themes concluded in the following order, with the first as the most important: function, psychology, aesthetics, clinical service, collaboration, and media. Within five multi-stakeholder groups, the workshop participants discussed challenges and collaborative opportunities for each theme. Workshop groups prioritised the themes based on their discussions, to highlight opportunities for further development. Two groups chose function, one group chose clinical service, one group chose collaboration, and another group chose media. The identified opportunities are presented within the context of the prioritised themes, including the importance of transparent information flow between all stakeholders; user involvement throughout research studies; and routes to informing healthcare policy through collaboration. As the field of upper limb prosthetics moves toward in-home research, we present co-creation as an approach that can facilitate user involvement throughout the duration of such studies.

Type three secretion systems enable bacterial pathogens to inject effectors into the cytosol of eukaryotic hosts to reprogram cellular functions. It is technically challenging to label effectors and the secretion machinery without disrupting their structure/function. Herein, we present a new approach for labeling and visualization of previously intractable targets. Using genetic code expansion, we site-specifically labeled SsaP, the substrate specificity switch, and SifA, a here-to-fore unlabeled secreted effector. SsaP was secreted at later infection times; SsaP labeling demonstrated the stochasticity of injectisome and effector expression. SifA was labeled after secretion into host cells via fluorescent unnatural amino acids or non-fluorescent labels and a subsequent click reaction. We demonstrate the superiority of imaging after genetic code expansion compared to small molecule tags. It provides an alternative for labeling proteins that do not tolerate N- or C-terminal tags or fluorophores and thus is widely applicable to other secreted effectors and small proteins.

Coronavirus interaction with its viral receptor is a primary genetic determinant of host range and tissue tropism. SARS-CoV-2 utilizes ACE2 as the receptor to enter host cell in a species-specific manner. We and others have previously shown that ACE2 orthologs from New World monkey, koala and mouse cannot interact with SARS-CoV-2 to mediate viral entry, and this defect can be restored by humanization of the restrictive residues in New World monkey ACE2. To better understand the genetic determinants behind the ability of ACE2 orthologs to support viral entry, we compared koala and mouse ACE2 sequences with that of human and identified the key residues in koala and mouse ACE2 that restrict viral receptor activity. Humanization of these critical residues rendered both koala and mouse ACE2 capable of binding the spike protein and facilitating viral entry. Our study shed more lights into the genetic determinants of ACE2 as the functional receptor of SARS-CoV-2, which facilitates our understanding of viral entry.

Abstract Purpose: Research demonstrates that children receive twice as much medical radiation from Computed Tomography (CT) scans performed at non-pediatric facilities as equivalent CTs performed at pediatric trauma centers (PTCs). In 2014, AFMC outreach staff educated Emergency Department (ED) staff on appropriate CT imaging utilization to reduce unnecessary medical radiation exposure. We set out to determine the educational campaign’s impact on injured children received radiation dose. Methods: All injured children who underwent CT imaging and were transferred to a Level I PTC during 2010 to 2013 (pre-campaign) and 2015 (post-campaign) were reviewed. Patient demographics, mode of transportation, ED length of stay, scanned body region, injury severity score, and trauma center level were analyzed. Median effective radiation dose (ERD) controlled for each variable, pre-campaign and post-campaign, was compared using Wilcoxon rank sum test. Results: Three hundred eighty-five children under 17 years were transferred from 45 and 48 hospitals, pre- and post-campaign. Most (43%) transferring hospitals were urban or critical access hospitals (30%). Pre- and post-campaign patient demographics were similar. We analyzed 482 and 398 CT scans pre- and post-campaign. Overall, median ERD significantly decreased from 3.80 to 2.80. Abdominal CT scan ERD declined significantly from 7.2 to 4.13 (P-value 0.03). Head CT scan ERD declined from 3.27 to 2.45 (P-value < 0.0001). Conclusion: A statewide, CT scan educational campaign contributed to ERD decline (lower dose scans and fewer repeat scans) among transferred injured children seen at PTCs. State-level interventions are feasible and can be effective in changing radiology provider practices.

Gram-positive and Gram-negative pathogens exist as planktonic cells only at limited times during their life cycle. In response to environmental signals such as temperature, pH, osmolality, and nutrient availability, pathogenic bacteria can adopt varied cellular fates, which involves the activation of virulence gene programs and/or the induction of a sessile lifestyle to form multicellular surface-attached communities. In Salmonella, SsrB is the response regulator which governs the lifestyle switch from an intracellular virulent state to form dormant biofilms in chronically infected hosts. Using the Salmonella lifestyle switch as a paradigm, we herein compare how other pathogens alter their lifestyles to enable survival, colonization and persistence in response to different environmental cues. It is evident that lifestyle switching often involves transcriptional regulators and their modification as highlighted here. Phenotypic heterogeneity resulting from stochastic cellular processes can also drive lifestyle variation among members of a population, although this subject is not considered in the present review.

After Salmonella is phagocytosed, it resides in an acidic vacuole. Its cytoplasm acidifies to pH 5.6; acidification activates pathogenicity island 2 (SPI-2). SPI-2 encodes a type three secretion system whose effectors modify the vacuole, driving endosomal tubulation. Using super-resolution imaging in single bacterial cells, we show that low pH induces expression of the SPI-2 SsrA/B signaling system. Single particle tracking, atomic force microscopy, and single molecule unzipping assays identified pH-dependent stimulation of DNA binding by SsrB. A so-called phosphomimetic form (D56E) was unable to bind to DNA in live cells. Acid-dependent DNA binding was not intrinsic to regulators, as PhoP and OmpR binding was not pH-sensitive. The low level of SPI-2 injectisomes observed in single cells is not due to fluctuating SsrB levels. This work highlights the surprising role that acid pH plays in virulence and intracellular lifestyles of Salmonella; modifying acid survival pathways represents a target for inhibiting Salmonella.

Walking aids are widely used by older adults, however, alarmingly, their use has been linked to increased falls-risk, yet clinicians have no objective way of assessing user stability. This work aims to demonstrate the application of a novel methodology to investigate how the type of walking task, the amount of body weight supported by the device (i.e., device loading), and task performance strategy affect stability of rollator users. In this context, ten users performed six walking tasks with an instrumented rollator. The combined stability margin "SM" was calculated, which considers user and rollator as a combined system. A Friedman Test was used to investigate the effects of task on SM and a least-squares regression model was applied to investigate the relationship between device loading and SM. In addition, the effects of task performance strategy on SM were explored. As a result, it was found that: the minimum SM for straight line walking was higher than for more complex tasks (p

ABSTRACT Nontyphoidal Salmonella (NTS), particularly Salmonella enterica serovar Typhimurium, is among the leading etiologic agents of bacterial enterocolitis globally and a well-characterized cause of invasive disease (iNTS) in sub-Saharan Africa. In contrast, S. Typhimurium is poorly defined in Southeast Asia, a known hot spot for zoonotic disease with a recently described burden of iNTS disease. Here, we aimed to add insight into the epidemiology and potential impact of zoonotic transfer and antimicrobial resistance (AMR) in S. Typhimurium associated with iNTS and enterocolitis in Vietnam. We performed whole-genome sequencing and phylogenetic reconstruction on 85 human (enterocolitis, carriage, and iNTS) and 113 animal S. Typhimurium isolates isolated in Vietnam. We found limited evidence for the zoonotic transmission of S. Typhimurium. However, we describe a chain of events where a pandemic monophasic variant of S. Typhimurium (serovar I:4,[5],12:i:− sequence type 34 [ST34]) has been introduced into Vietnam, reacquired a phase 2 flagellum, and acquired an IncHI2 multidrug-resistant plasmid. Notably, these novel biphasic ST34 S. Typhimurium variants were significantly associated with iNTS in Vietnamese HIV-infected patients. Our study represents the first characterization of novel iNTS organisms isolated outside sub-Saharan Africa and outlines a new pathway for the emergence of alternative Salmonella variants into susceptible human populations. IMPORTANCE Salmonella Typhimurium is a major diarrheal pathogen and associated with invasive nontyphoid Salmonella (iNTS) disease in vulnerable populations. We present the first characterization of iNTS organisms in Southeast Asia and describe a different evolutionary trajectory from that of organisms causing iNTS in sub-Saharan Africa. In Vietnam, the globally distributed monophasic variant of Salmonella Typhimurium, the serovar I:4,[5],12:i:− ST34 clone, has reacquired a phase 2 flagellum and gained a multidrug-resistant plasmid to become associated with iNTS disease in HIV-infected patients. We document distinct communities of S. Typhimurium and I:4,[5],12:i:− in animals and humans in Vietnam, despite the greater mixing of these host populations here. These data highlight the importance of whole-genome sequencing surveillance in a One Health context in understanding the evolution and spread of resistant bacterial infections.

Bacteria survive and respond to diverse environmental conditions and during infection inside the host by systematic regulation of stress response genes. E. coli and S. Typhimurium can undergo large changes in intracellular osmolality (up to 1.8 Osmol/kg) and can tolerate cytoplasmic acidification to at least pHi 5.6. Recent analyses of single cells challenged a long held view that bacteria respond to extracellular acid stress by rapid acidification followed by a rapid recovery. It is now appreciated that both S. Typhimurium and E. coli maintain an acidic cytoplasm through the actions of the outer membrane protein regulator OmpR via its regulation of distinct signaling pathways. However, a comprehensive comparison of OmpR regulons between S. Typhimurium and E. coli is lacking. In this study, we examined the expression profiles of wild-type and ompR null strains of the intracellular pathogen S. Typhimurium and a commensal E. coli in response to acid and osmotic stress. Herein, we classify distinct OmpR regulons and also identify shared OmpR regulatory pathways between S. Typhimurium and E. coli in response to acid and osmotic stress. Our study establishes OmpR as a key regulator of bacterial virulence, growth and metabolism, in addition to its role in regulating outer membrane proteins.

ABSTRACT Bacterial pathogens exploit eukaryotic pathways for their own end. Upon ingestion, Salmonella enterica serovar Typhimurium passes through the stomach and then catalyzes its uptake across the intestinal epithelium. It survives and replicates in an acidic vacuole through the action of virulence factors secreted by a type three secretion system located on Salmonella pathogenicity island 2 (SPI-2). Two secreted effectors, SifA and SseJ, are sufficient for endosomal tubule formation, which modifies the vacuole and enables Salmonella to replicate within it. Two-color, superresolution imaging of the secreted virulence factor SseJ and tubulin revealed that SseJ formed clusters of conserved size at regular, periodic intervals in the host cytoplasm. Analysis of SseJ clustering indicated the presence of a pearling effect, which is a force-driven, osmotically sensitive process. The pearling transition is an instability driven by membranes under tension; it is induced by hypotonic or hypertonic buffer exchange and leads to the formation of beadlike structures of similar size and regular spacing. Reducing the osmolality of the fixation conditions using glutaraldehyde enabled visualization of continuous and intact tubules. Correlation analysis revealed that SseJ was colocalized with the motor protein kinesin. Tubulation of the endoplasmic reticulum is driven by microtubule motors, and in the present work, we describe how Salmonella has coopted the microtubule motor kinesin to drive the force-dependent process of endosomal tubulation. Thus, endosomal tubule formation is a force-driven process catalyzed by Salmonella virulence factors secreted into the host cytoplasm during infection. IMPORTANCE This study represents the first example of using two-color, superresolution imaging to analyze the secretion of Salmonella virulence factors as they are secreted from the SPI-2 type three secretion system. Previous studies imaged effectors that were overexpressed in the host cytoplasm. The present work reveals an unusual force-driven process, the pearling transition, which indicates that Salmonella-induced filaments are under force through the interactions of effector molecules with the motor protein kinesin. This work provides a caution by highlighting how fixation conditions can influence the images observed.