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Single-strand annealing (SSA) is initiated when a double strand break (DSB) occurs between two flanking repeated sequences, resulting in a deletion that leaves a single copy of the repeat. We studied budding yeast strains carrying two 200-bp URA3 sequences separated by 2.6 kb of spacer DNA (phage lambda) in which a site-specific DSB can be created by HO or Cas9 endonucleases. Repeat-mediated deletion requires removal of long 3'-ended single-stranded tails (flaps) by Rad1-Rad10 with the assistance of Msh2-Msh3, Saw1 and Slx4. A natural 3% divergence of unequally spaced heterologies between these repeats (designated F and A) causes a significant reduction in the frequency of SSA repair. This decrease is caused by heteroduplex rejection in which mismatches (MMs) in the annealed intermediate are recognized by the MutS (Msh2 and Msh6) components of the MM repair (MMR) pathway coupled to unwinding of the duplex by the Sgs1-Rmi1-Top3 helicase. MutL homologs, Mlh1-Pms1 (MutL), are not required for rejection but play their expected role in mismatch correction. Remarkably, heteroduplex rejection is very low in strains where the divergent repeats were immediately adjacent (Tailless strains) and the DSB was induced by Cas9. These results suggest that the presence of nonhomologous tails strongly stimulates heteroduplex rejection in SSA. DNA sequencing analysis of SSA products from the FA Tailed strain showed a gradient of correction favoring the sequence opposite each 3' end of the annealed strand. Mismatches located in the center of the repair intermediate were corrected by Msh2-Msh6 mediated mismatch correction, while correction of MMs at the extremity of the SSA intermediate often appears to use a different mechanism, possibly by 3' nonhomologous tail removal that includes part of the homologous sequence. In contrast, in FA Tailless strains there was a uniform repair of the MMs across the repeat. A distinctive pattern of correction was found in the absence of MSH2, in both Tailed and Tailless strains, different from the spectrum seen in a msh3Δ msh6Δ double mutant. Previous work has shown that SSA is Rad51-independent but dependent on the strand annealing activity of Rad52. However Rad52 becomes dispensable in a Tailless construct where the DSB is induced by Cas9 or in transformation of a plasmid where SSA occurs in the absence of nonhomologous tails.

Abstract The Tomonaga–Luttinger liquid (TLL) theory describes the low-energy excitations of strongly correlated one-dimensional (1D) fermions. In the past years, a number of studies have provided a detailed understanding of this universality class. More recently, theoretical investigations that go beyond the standard low-temperature, linear-response TLL regime have been developed. While these provide a basis for understanding the dynamics of the spin-incoherent Luttinger liquid, there are few experimental investigations in this regime. Here we report the observation of a thermally induced, spin-incoherent Luttinger liquid in a 6Li atomic Fermi gas confined to 1D. We use Bragg spectroscopy to measure the suppression of spin-charge separation and the decay of correlations as the temperature is increased. Our results probe the crossover between the coherent and incoherent regimes of the Luttinger liquid and elucidate the roles of the charge and the spin degrees of freedom in this regime.

Abstract Selective serotonin reuptake inhibitors (SSRIs) have anti‐inflammatory properties that may have clinical utility in treating severe pulmonary manifestations of COVID‐19. SSRIs exert anti‐inflammatory effects at three mechanistic levels: (a) inhibition of proinflammatory transcription factor activity, including NF‐κB and STAT3; (b) downregulation of lung tissue damage and proinflammatory cell recruitment via inhibition of cytokines, including IL‐6, IL‐8, TNF‐α, and IL‐1β; and (c) direct suppression inflammatory cells, including T cells, macrophages, and platelets. These pathways are implicated in the pathogenesis of COVID‐19. In this review, we will compare the pathogenesis of lung inflammation in pulmonary diseases including COVID‐19, ARDS, and chronic obstructive pulmonary disease (COPD), describe the anti‐inflammatory properties of SSRIs, and discuss the applications of SSRIS in treating COVID‐19‐associated inflammatory lung disease.

Maintaining hand hygiene has been an essential preventive measure for reducing disease transmission in public facilities, particularly during the COVID-19 pandemic. The large number of sanitizer stations deployed within public facilities, such as on university campuses, brings challenges for effective facility management. This paper proposes an IoT sensor network for tracking sanitizer usage in public facilities and supporting facility management using a data-driven approach. Specifically, the system integrates low-cost wireless sensors, LoRaWAN, and cloud-based computing techniques to realize data capture, communication, and analysis. The proposed approach was validated through field experiments in a large building on a university campus to assess the network signal coverage and effectiveness of sensor operation for facility monitoring. The results show that a LoRaWAN created from a single gateway can successfully connect to sensors distributed throughout the entire building, with the sensor nodes recording and transmitting events across the network for further analysis. Overall, this paper demonstrates the potential of leveraging the IoT-based Sanitizer Station Network to track public health mitigation methods in a large facility, which ultimately contributes to reducing the burden of maintaining public health during and post-pandemic.

Culled whole grapefruit (WG) and grapefruit juice processing residues (GP) are currently incorporated into low-cost animal feed. If individual chemical components found within these side streams could be recovered as high-value coproducts, this would improve the overall value of the grapefruit crop. In this study, pectic hydrocolloids, sugars, volatiles, phenolics, and flavonoids were extracted from Star Ruby, Rio Red, and Ruby Red GP and WG using a continuous pilot scale steam explosion system. Up to 97% of grapefruit juice oils and peel oils could be volatilized and contained 87–94% d-limonene. The recovery of pectin, as determined by galacturonic acid content, was between 2.06 and 2.72 g 100 g−1. Of the phenolics and flavonoids analyzed in this study, narirutin and naringin were extracted in the amounts of up to 10,000 and 67,000 μg g−1, respectively.

Florida juice processors amassed 51,000 metric tons of grapefruit juice processing residues (GFPR) in the 2018-19 season. GFPR contain valuable compounds such as pectic hydrocolloids, peel oil, sugars, phenolics and flavonoids that are ultimately lost or destroyed when they are converted to low value animal feed and molasses. If these compounds could be isolated from GFPR rather than destroyed, they could be converted to high value products. In this work, red and white GFPR were subjected to steam explosion in a batch system at 130, 150 and 170°C and 1, 2, 4 and 8 min holding time to identify the conditions for maximum recovery of pectic hydrocolloids, peel oil, sugars, phenolics and flavonoids. Steam explosion at 170°C and an 8 min hold time resulted in the recovery of 56-85% of total available glucose and fructose and the volatilization of 68-98% of the available peel oil. Of the phenolics and flavonoids analyzed, 61-62% could be extracted using steam explosion. Maximum recovery of pectic hydrocolloids, estimated by galacturonic acid content, was 1.6446 g • 100 g−1 for red GFPR at 130°C – 8 min and 1.557 g • 100 g−1 for white GFPR at 150°C – 2 min.

Abstract Background Platelets are widely recognized for their role in maintaining hemostasis. Recently, platelets have become appreciated for their varying roles in immunity, neuroprotection, and other physiological processes. While there are currently excellent methods to transiently deplete platelets and models of thrombocytopenia, studying the roles of platelets in chronic processes can be challenging. Objective Phenotypic characterization of the PF4‐DTR mouse model of conditional platelet depletion compared to antibody depletion. Methods We describe the ability of the PF4‐DTR mouse to maintain chronic platelet depletion, along with examining the bleeding phenotype compared to antibody‐mediated platelet depletion. Results Systemic administration of diphtheria toxin resulted in >99% platelet depletion that can be maintained for >2 weeks. When compared to an antibody depletion model, PF4‐DTR mice showed similar phenotypes when challenged with tail bleed and saphenous vein measurements of hemostasis. Mice depleted with diphtheria toxin were also able to undergo adoptive transfer of platelets. If the frequency and amount of diphtheria toxin is reduced, mice can be maintained at >40% depletion for >28 days, showing that this model is tunable. Conclusions When compared to the gold standard of antibody‐mediated depletion, PF4‐DTR mice showed similar phenotypes and should be considered an important tool for examining the impact of thrombocytopenia over longer periods of time.

Economic models of democratic decision-making tend to assume that voters have preferences and politicians adjust their platforms to conform to voter preferences. However, the direction of causation (mostly) goes the other way. Political elites offer policy platforms and voters adopt the policy preferences of their political anchors. Because the choices of individual voters do not affect aggregate political outcomes, voters tend to vote expressively, and might vote for outcomes they would not choose if the choice were theirs alone. The concept of expressive preferences is well-established. This paper takes the next step by explaining how voters form their expressive preferences. Expressive preferences tend to be anchored in a political identity associated with a candidate, party, or ideology, and people's political preferences on most issues are derived from their anchor preferences, which are defined by political elites.

Medical student wellness is of great concern in the health care field. A growing number of studies point to increases in suicide, depression, anxiety, mood disorders, and burnout related to physician lifestyles. Mental health issues commencing in medical school have been suggested to have a significant impact on future physician lifestyle and burnout. Tracking the mental health of medical students at the University of Toledo College of Medicine and Life Sciences (UTCOMLS) with standardized indices will help elucidate triggers of poor mental health. Anonymous surveys were developed and distributed to preclinical medical students at five strategic time points throughout the 2018 2019 academic year. Surveys collected basic demographic information as well as inventories measuring perceived stress, burnout, resilience, and mindfulness. 172 M1s (83 males and 89 females) were included in the study and average response rate for the first 4 (out of 5) surveys averaged 74.8%. M1 males and females had on average increased personal burnout over time with females consistently scoring higher. Both males and females had an increase in stress from August to each subsequent month (p

Platelet-leukocyte aggregates (PLAs) are associated with increased thrombosis risk. The influence of PLA formation is especially important for cancer patients, since thrombosis accounts for approximately 10% of cancer-associated deaths. Our objective was to characterize and quantify PLAs in whole blood samples from lung cancer patients compared to healthy volunteers with the intent to analyze PLA formation in the context of lung cancer-associated thrombosis. Consenting lung cancer patients (57) and healthy volunteers (56) were enrolled at the Dana Cancer Center at the University of Toledo Health Science Campus. Peripheral blood samples were analyzed by flow cytometry. Patient medical history was reviewed through electronic medical records. Most importantly, we found lung cancer patients to have higher percentages of platelet-T cell aggregates (PTCAs) than healthy volunteers among both CD4+ T lymphocyte and CD8+ T lymphocyte populations. Our findings demonstrate that characterization of PTCAs may have clinical utility in differentiating lung cancer patients from healthy volunteers and stratifying lung cancer patients by history of thrombosis.

Florida citrus production has declined 75% due to Huanglongbing (HLB), a disease caused by the pathogenic bacterium Candidatus Liberibacter asiaticus (CLas). Methods to combat CLas are costly and only partially effective. The cross-compatible species Poncirus trifoliata and some of its hybrids are known to be highly tolerant to CLas, and thus can potentially serve as an alternative feedstock for many citrus products. To further investigate the commercial potential of citrus hybrids, three citrus hybrids, US-802, US-897, and US-942, were studied for their potential as feedstocks for citrus co-products using steam explosion (STEX) followed by water extraction. Up to 93% of sugars were recovered. US-897 and US-942 have similar volatile profiles to that of the commercial citrus fruit types and as much as 85% of these volatiles could be recovered. Approximately 80% of the pectic hydrocolloids present in all three hybrids could be obtained in water washes of STEX material. Of the phenolics identified, the flavanone glycosides, i.e., naringin, neohesperidin, and poncirin were the most abundant quantitatively in these hybrids. The ability to extract a large percentage of these compounds, along with their inherent values, make US-802, US-897, and US-942 potentially viable feedstock sources for citrus co-products in the current HLB-blighted environment.

The geoPebble system is a network of wirelessly interconnected seismic and GPS sensor nodes with geophysical sensing capabilities for the study of ice sheets in Antarctica and Greenland, as well as mountain glaciers. We describe our design methodology, which has enabled us to develop these state-of-the art units using commercial-off-the-shelf hardware combined with custom-designed hardware and software. Each geoPebble node is a self-contained, wirelessly connected sensor for collecting seismic activity and position information. Each node is built around a three-component seismic recorder, which includes an amplifier, filter, and 24-bit analog-to-digital converter that can sample incoming seismic signals up to 10 kHz. The timing for each node is available from GPS measurements and a local precision oscillator that is conditioned by the GPS timing pulses. In addition, we record the carrier-phase measurement of the L1 GPS signal in order to determine location at sub-decimeter accuracy (relative to other geoPebble nodes within a radius of a few kilometers). Each geoPebble includes 32 GB of solid-state storage, wireless communications capability to a central supervisory unit, and auxiliary measurements capability (including tilt from accelerometers, absolute orientation from magnetometers, and temperature). The geoPebble system has been successfully validated in the field in Antarctica and Greenland.

The study of entrepreneurship often focuses on the activities of the entrepreneur. While entrepreneurship is undertaken by individuals, the degree to which individuals are entrepreneurial, and the directions in which their entrepreneurial actions take, are the result of the institutional environment. Some economies are more entrepreneurial than others. After discussing the institutional factors that encourage entrepreneurship, the ways in which economic models depict the economy are discussed, pointing toward the different policy conclusions regarding entrepreneurship that emanate from different assumptions in economic conditions.

The objective of this research was to evaluate the use of a controlled-release carvacrol powder to delay storage decay and maintain the safety of blueberries. The controlled-release carvacrol powder was a microcapsule of carvacrol (11% (w/w) active carvacrol) surrounded by a pectin/sodium alginate matrix. The microcapsules were packed in an air-permeable pouch, and then attached to the top of a clamshell filled with blueberries. The blueberries, inoculated with Escherichia coli or Colletotrichum acutatum, or non-inoculated control, were monitored for microbial growth and quality for 10 days at 10 °C and 5 days at 20 °C. Three treatments were compared: controlled-release microencapsulated carvacrol, non-encapsulated carvacrol, and control. The results showed that both the microencapsulated carvacrol and the non-encapsulated carvacrol treatments significantly reduced the populations of yeast and mold, and of E. coli and mesophilic aerobic bacteria. The microencapsulated carvacrol treated berries retained better quality due to significantly lower weight loss than control after 10 days at 10 °C. Sensory panelists found that the microencapsulated carvacrol berries had significantly higher overall blueberry flavor and lower discernible off-flavor in comparison with the non-encapsulated treatment after 3 days at 20 °C. The fruit internal quality, including total soluble solids content (SSC), and titratable acidity (TA), was not significantly affected by any treatment. These results indicate that pectin/sodium alginate controlled-release microencapsulated carvacrol can be used for the preservation of blueberries or other small fruit.

Following injury, the mammalian spinal cord forms a glial scar and fails to regenerate. In contrast, vertebrate fish spinal cord tissue regenerates significantly to restore function. Cord transection in zebrafish (Danio rerio) initially causes paralysis and neural cell death. Subsequently, ependymal glia proliferate, bipolar glia extend across the lesion, and new neurons are born; axons from spared and nascent neurons extend along trans-lesional glial bridges to restore functional connectivity. Here we report that glucocorticoids, used in the clinical management of spinal cord injury, directly inhibit neural repair by targeting ependymal glia independently of hematogenous cells and microglia. After transecting injury, the glucocorticoid receptor in ependymal glia is regulated differentially in zebrafish (becoming inactive) vs. the rat (becoming active). Glucocorticoid blockade of neural regeneration via a direct effect on ependymal glia has important therapeutic implications for the putative benefit of corticosteroids in early management of spinal cord injury.

Quantum simulators are a promising technology on the spectrum of quantum devices from specialized quantum experiments to universal quantum computers. These quantum devices utilize entanglement and many-particle behavior to explore and solve hard scientific, engineering, and computational problems. Rapid development over the last two decades has produced more than 300 quantum simulators in operation worldwide using a wide variety of experimental platforms. Recent advances in several physical architectures promise a golden age of quantum simulators ranging from highly optimized special purpose simulators to flexible programmable devices. These developments have enabled a convergence of ideas drawn from fundamental physics, computer science, and device engineering. They have strong potential to address problems of societal importance, ranging from understanding vital chemical processes, to enabling the design of new materials with enhanced performance, to solving complex computational problems. It is the position of the community, as represented by participants of the National Science Foundation workshop on “Programmable Quantum Simulators,” that investment in a national quantum simulator program is a high priority in order to accelerate the progress in this field and to result in the first practical applications of quantum machines. Such a program should address two areas of emphasis: (1) support for creating quantum simulator prototypes usable by the broader scientific community, complementary to the present universal quantum computer effort in industry; and (2) support for fundamental research carried out by a blend of multi-investigator, multidisciplinary collaborations with resources for quantum simulator software, hardware, and education.This document is a summary from a U.S. National Science Foundation supported workshop held on 16–17 September 2019 in Alexandria, VA. Attendees were charged to identify the scientific and community needs, opportunities, and significant challenges for quantum simulators over the next 2–5 years.

Platelets have long been recognized for their role in maintaining the balance between hemostasis and thrombosis. While their contributions to blood clotting have been well established, it has been increasingly evident that their roles extend to both innate and adaptive immune functions during infection and inflammation. In this comprehensive review, we describe the various ways in which platelets interact with different microbes and elicit immune responses either directly, or through modulation of leukocyte behaviors.

The objectives of this research were to microencapsulate tangeretin, and to evaluate the basic characteristics of the microcapsule products. Tangeretin is a polymethoxyflavone (PMF) which has been revealed to possess various health benefits and is abundant in tangerine and other citrus peels. Microencapsulation technology is widely employed in the food and pharmaceutical industries to exploit functional ingredients, cells, and enzymes. Spray drying is a frequently applied microencapsulation method because of its low cost and technical requirements. In this research, tangeretin dissolved at different concentrations in bergamot oil was microencapsulated in a citrus pectin/sodium alginate matrix. The resulting microcapsule powder showed promising physical and structural properties. The retention efficiency of tangeretin was greater at a concentration of 2.0% (98.92%) than at 0.2% (71.05%), probably due to the higher temperature of the emulsion during the homogenizing and spray-drying processes. Encapsulation efficiency was reduced with increased concentration of tangeretin. Our results indicate that tangeretin could be successfully encapsulated within a citrus pectin/sodium alginate matrix using bergamot oil as a carrier.

The ability to orchestrate appropriate physiological and behavioral responses to stress is important for survival, and is often dysfunctional in neuropsychiatric disorders that account for leading causes of global disability burden. Numerous studies have shown that the endocannabinoid neurotransmitter system is able to regulate stress responses and could serve as a therapeutic target for the management of these disorders. We used quantitative reverse transcriptase-polymerase chain reactions to show that genes encoding enzymes that synthesize (abhd4, gde1, napepld), enzymes that degrade (faah, faah2a, faah2b), and receptors that bind (cnr1, cnr2, gpr55-like) endocannabinoids are expressed in zebrafish (Danio rerio). These genes are conserved in many other vertebrates, including humans, but fatty acid amide hydrolase 2 has been lost in mice and rats. We engineered transcription activator-like effector nucleases to create zebrafish with mutations in cnr1 and faah2a to test the role of these genes in modulating stress-associated behavior. We showed that disruption of cnr1 potentiated locomotor responses to hyperosmotic stress. The increased response to stress was consistent with rodent literature and served to validate the use of zebrafish in this field. Moreover, we showed for the first time that disruption of faah2a attenuated the locomotor responses to hyperosmotic stress. This later finding suggests that FAAH2 may be an important mediator of stress responses in non-rodent vertebrates. Accordingly, FAAH and FAAH2 modulators could provide distinct therapeutic options for stress-aggravated disorders.

Murray Rothbard's For a New Liberty, originally published in 1973, remains one of the most significant books on libertarianism, in large part because he explains how market institutions can replace [...]

Attention deficit hyperactivity disorder (ADHD) is the most common neuropsychiatric disorder in children and is increasing in prevalence. There has also been a related increase in prescribing stimulant medication despite some controversy whether ADHD medication makes a lasting difference in school performance or achievement. Families who are apprehensive about side effects and with concerns for efficacy of medication pursue integrative medicine as an alternative or adjunct to pharmacologic and cognitive behavioral treatment approaches. Integrative medicine incorporates evidence-based medicine, both conventional and complementary and alternative therapies, to deliver personalized care to the patient, emphasizing diet, nutrients, gut health, and environmental influences as a means to decrease symptoms associated with chronic disorders. Pediatric integrative medicine practitioners are increasing in number throughout the United States because of improvement in patient health outcomes. However, limited funding and poor research design interfere with generalizable treatment approaches utilizing integrative medicine. The use of research designs originally intended for drugs and procedures are not suitable for many integrative medicine approaches. This article serves to highlight integrative medicine approaches in use today for children with ADHD, including dietary therapies, nutritional supplements, environmental hygiene, and neurofeedback.

Early-maturing and full-season Bt and non-Bt cottons were exposed to high densities of tobacco budworm (Heliothis virescens (F.)) and bollworm (Helicoverpa zea Boddie) in 0.04 ha field cages during the summers of 2011 and 2012 to measure the possible need for supplemental use of insecticides on Bt cotton. Fruit survival within-season and at-harvest was carefully mapped on individual plants within comparative plots of all cottons untreated and sprayed with lambda-cyhalothin (0.0448 kg a.i./ha) or chlorantraniliprole (0.1009 kg a.i./ha) following insect infestations. Differences in lint yields among cotton maturity groups were not always detected, but early-maturing Bt cottons were among the higher yielding experimental plots for both years. Depending on the insecticide treatment, average harvested fruit ranged from 0.3 to 7.1 open bolls per plant for non-Bt cotton plots, while Bt cotton plots ranged from 1.8 to 7.5 open bolls per plant during the two-year study. Bt cottons generally protected fruit from insect damage and resulted in final yields comparable to those of insecticide sprayed Bt and non-Bt cottons. Unsprayed non-Bt cottons were significantly damaged by insects in these high-infestation environments.

Neutron beta decay is one of the most fundamental processes in nuclear physics and provides sensitive means to uncover the details of the weak interaction. Neutron beta decay can evaluate the ratio of axial-vector to vector coupling constants in the standard model, λ = gA/gV, through multiple decay correlations. The Nab experiment will carry out measurements of the electron-neutrino correlation parameter a with a precision of δa/a = 10−3 and the Fierz interference term b to δb = 3 × 10−3 in unpolarized free neutron beta decay. These results, along with a more precise measurement of the neutron lifetime, aim to deliver an independent determination of the ratio λ with a precision of δλ/λ = 0.03% that will allow an evaluation of Vud and sensitively test CKM unitarity, independent of nuclear models. Nab utilizes a novel, long asymmetric spectrometer that guides the decay electron and proton to two large area silicon detectors in order to precisely determine the electron energy and an estimation of the proton momentum from the proton time of flight. The Nab spectrometer is being commissioned at the Fundamental Neutron Physics Beamline at the Spallation Neutron Source at Oak Ridge National Lab. We present an overview of the Nab experiment and recent updates on the spectrometer, analysis, and systematic effects.

Purpose. In this study, we evaluated our experience with salvage brachytherapy after discovery of biochemical recurrence after a prior brachytherapy procedure. Methods and Materials. From 2001 through 2012 twenty-one patients treated by brachytherapy within University of Kentucky or from outside centers developed biochemical failure and had no evidence of metastases. Computed tomography (CT) scans were evaluated; patients who had an underseeded portion of their prostate were considered for reimplantation. Results. The majority of the patients in this study (61.9%) were low risk and median presalvage PSA was 3.49 (range 17.41–1.68). Mean follow-up was 61 months. At last follow-up after reseeding, 11/21 (52.4%) were free of biochemical recurrence. There was a trend towards decreased freedom from biochemical recurrence in low risk patients (p=0.12). International Prostate Symptom Scores (IPSS) increased at 3-month follow-up visits but decreased and were equivalent to baseline scores at 18 months. Conclusions. Salvage brachytherapy after primary brachytherapy is possible; however, in our experience the side-effect profile after the second brachytherapy procedure was higher than after the first brachytherapy procedure. In this cohort of patients we demonstrate that approximately 50% oncologic control, low risk patients appear to have better outcomes than others.

Constitutional political economy has focused heavily on designing constitutional rules sufficient to constrain governmental power. More attention has been devoted to designing rules that are effective constraints than on the institutions that would be required to enforce them. One problem is that rules are interpreted and enforced by the political elite, who tend to interpret and enforce them in ways that favor their interests over those of the masses. Democratic oversight is ineffective because voters realize they have no influence over public policy, and are therefore rationally ignorant. A system of checks and balances within government is necessary for enforcing constitutional constraints because it divides power among elites with competing interests and enables one group of elites to check the power of others. Checks and balances within governmental institutions are necessary to constrain the government from abusing its power.

Concentration-response assays were conducted from 2008 through 2015 to measure the susceptibility of field populations of Lygus lineolaris (Palisot de Beauvois) from the Delta regions of Arkansas, Louisiana, and Mississippi to acephate, imidacloprid, thiamethoxam, permethrin, and sulfoxaflor. A total of 229 field populations were examined for susceptibility to acephate, 145 for susceptibility to imidacloprid, and 208 for susceptibility to thiamethoxam. Permethrin assays were conducted in 2014 and 2015 to measure levels of pyrethroid resistance in 44 field populations, and sulfoxaflor assays were conducted against 24 field populations in 2015. Resistance to acephate and permethrin is as high or higher than that previously reported, although some populations, especially those exposed to permethrin, appear to be susceptible. Variable assay responses were measured for populations exposed to imidacloprid and thiamethoxam. Average response metrics suggest that populations are generally susceptible to the neonicotinoids, but a few populations from cotton fields experiencing control problems exhibited elevated LC50s. Efforts to associate variability in LC50s with recorded use of insecticides and estimated cotton insect losses and control costs suggest that intensive use of insecticides over several decades may have elevated general detoxifying enzymes in L. lineolaris and some field populations may be exhibiting resistance to multiple classes of insecticide. These results suggest that efforts should be made to manage these pests more efficiently with a reduced use of insecticides and alternative controls.

ABSTRACT Haemophilus ducreyi causes chancroid, a sexually transmitted infection. A primary means by which this pathogen causes disease involves eluding phagocytosis; however, the molecular basis for this escape mechanism has been poorly understood. Here, we report that the LspA virulence factors of H. ducreyi inhibit phagocytosis by stimulating the catalytic activity of C-terminal Src kinase (Csk), which itself inhibits Src family protein tyrosine kinases (SFKs) that promote phagocytosis. Inhibitory activity could be localized to a 37-kDa domain (designated YL2) of the 456-kDa LspA1 protein. The YL2 domain impaired ingestion of IgG-opsonized targets and decreased levels of active SFKs when expressed in mammalian cells. YL2 contains tyrosine residues in two EPIYG motifs that are phosphorylated in mammalian cells. These tyrosine residues were essential for YL2-based inhibition of phagocytosis. Csk was identified as the predominant mammalian protein interacting with YL2, and a dominant-negative Csk rescued phagocytosis in the presence of YL2. Purified Csk phosphorylated the tyrosines in the YL2 EPIYG motifs. Phosphorylated YL2 increased Csk catalytic activity, resulting in positive feedback, such that YL2 can be phosphorylated by the same kinase that it activates. Finally, we found that the Helicobacter pylori CagA protein also inhibited phagocytosis in a Csk-dependent manner, raising the possibility that this may be a general mechanism among diverse bacteria. Harnessing Csk to subvert the Fcγ receptor (FcγR)-mediated phagocytic pathway represents a new bacterial mechanism for circumventing a crucial component of the innate immune response and may potentially affect other SFK-involved cellular pathways. IMPORTANCE Phagocytosis is a critical component of the immune system that enables pathogens to be contained and cleared. A number of bacterial pathogens have developed specific strategies to either physically evade phagocytosis or block the intracellular signaling required for phagocytic activity. Haemophilus ducreyi, a sexually transmitted pathogen, secretes a 4,153-amino-acid (aa) protein (LspA1) that effectively inhibits FcγR-mediated phagocytic activity. In this study, we show that a 294-aa domain within this bacterial protein binds to C-terminal Src kinase (Csk) and stimulates its catalytic activity, resulting in a significant attenuation of Src kinase activity and consequent inhibition of phagocytosis. The ability to inhibit phagocytosis via Csk is not unique to H. ducreyi, because we found that the Helicobacter pylori CagA protein also inhibits phagocytosis in a Csk-dependent manner. Harnessing Csk to subvert the FcγR-mediated phagocytic pathway represents a new bacterial effector mechanism for circumventing the innate immune response.