Search

William Gowers' classic single-authored two-volume A manual of diseases of the nervous system appeared in a first edition in 1886 and 1888, and in a second edition in 1892 and 1893, with a third edition of Volume 1 in 1899. No third edition of Volume 2 ever appeared. However, in 1949 Critchley stated that he had seen part of a revision of this volume. Subsequent writers could not find this material, but it recently came to light at Gowers' old hospital at Queen Square, London. The present paper describes the rediscovered material, containing Gowers' handwritten alterations for a further edition of Volume 2, and substantial new material, at least in relation to nystagmus and myasthenia. Gowers' declining health, or a conflict between his planned new text and his contributions to the neurology segments (1899) of Allbutt's System of medicine, may explain why a third edition of Volume 2 of the Manual of diseases of the nervous system never appeared. [ABSTRACT FROM AUTHOR]

It is commonly assumed that elderly patients in long-term care facilities require less total care and less professional care than patients in active 'treatment' settings. However, no objective data have been available to support or refute these assumptions. This study examined patient classification data obtained from eight acute care medical and surgical units and four long-term care geriatric units over a 1-year period. Results indicated that (1) geriatric patients in long-term care units had differing (as opposed to similar) nursing care needs, (2) nursing care requirements of geriatric patients in several long-term care units differed from those of patients in another long-term care unit, (3) geriatric patients in the selected long-term care units required at least as much nursing care as those in the medical surgical units, and (4) the long-term care units had many patients with extensive nursing care requirements but no patients with very complex requirements when compared with the medical/surgical units. The potential impact of these findings on staffing of long-term care units is considerable. Issues relating to reliability, validity and comparability are discussed. There is a great need to utilize standardized, state-of-the-art methods for nursing workload measurement to facilitate comparisons of nursing care requirements of geriatric long-term care patients with those of patients in other areas. [ABSTRACT FROM AUTHOR]

Abstract: Timely diagnosis of systemic mastocytosis (SM) remains challenging because of care heterogeneity. We implemented a standardized approach for SM screening and diagnosis using a novel health care system-wide international screening registry. A retrospective analysis assessed rates of SM, cutaneous mastocytosis (CM), and molecular diagnoses before and 2 years after care standardization. The accuracy of individual and combined SM screening tests, basal serum tryptase (BST) ≥11.5 and ≥20.0 ng/mL, REMA ≥2, monomorphic maculopapular CM (MPCM), and elevated BST based upon tryptase genotype, was analyzed. Tryptase genotyping and high-sensitivity KIT p.D816V testing increased substantially 2 years after care standardization. SM diagnoses doubled from 47 to 94, and KIT p.D816V molecular diagnoses increased from 24 to 79. Mean BST and KIT p.D816V variant allele frequency values were significantly lower in patients diagnosed after standardization. Hereditary-alpha tryptasemia prevalence was increased in SM before care standardization (4/30 [13.3%]) but reflected the general population prevalence 2 years later at (5/76 [6.6%]). Elevated BST based upon genotype and BST ≥11.5 ng/mL had the highest sensitivities at 84.2% and 88.3%, respectively. The presence of monomorphic MPCM, elevated BST based upon tryptase genotype, and the combination of REMA ≥2 with elevated BST based upon tryptase genotype had specificities >90%. BST >20.0 ng/mL had low sensitivity and specificity and was not required to establish any indolent SM (ISM) diagnosis. Care standardization increased SM diagnosis rates, particularly in patients with low BSTs. Stratifying BST based upon genotype had the best overall sensitivity and specificity of any ISM screening test and improved the REMA score specificity.

Reminders such as calendar entries and digital alerts are highly effective at helping people remember what they need to do. Recent work has explored the triggers of reminder setting, also known as intention offloading, finding that low confidence is associated with more offloading in laboratory tasks. This supports a metacognitive model of cognitive offloading. Here, we investigated whether this model generalizes to real-world intentions. We asked 112 participants about their upcoming plans and subsequent fulfillment of those plans in two online surveys conducted in 2022. Participants were more likely to set reminders for plans that (a) they were less confident they would remember and (b) they judged to be more important. Participants with lower confidence that they would remember to return for the second survey were more likely to set a reminder for this plan. This, in turn, predicted greater fulfillment. Therefore, ironically, lower confidence predicted greater success. These findings show that intention offloading predicts fulfillment of real-world intentions. They also point toward metacognition as a target for interventions to facilitate this. (PsycInfo Database Record (c) 2024 APA, all rights reserved).

Burkholderia pseudomallei is the causative agent of melioidosis, which is endemic primarily in Southeast Asia and northern Australia but is increasingly being seen in other tropical and subtropical regions of the world. Melioidosis is associated with high morbidity and mortality rates, which is mediated by the wide range of virulence factors encoded by B. pseudomallei. These virulence determinants include surface polysaccharides such as lipopolysaccharide (LPS) and capsular polysaccharides (CPS). Here, we investigated a predicted arabinose-5-phosphate isomerase (API) similar to KdsD in B. pseudomallei strain K96243. KdsD is required for the production of the highly conserved 3-deoxy-d-manno-octulosonic acid (Kdo), a key sugar in the core region of LPS. Recombinant KdsD was expressed and purified, and API activity was determined. Although a putative API paralogue (KpsF) is also predicted to be encoded, the deletion of kdsD resulted in growth defects, loss of motility, reduced survival in RAW 264.7 murine macrophages, and attenuation in a BALB/c mouse model of melioidosis. Suppressor mutations were observed during a phenotypic screen for motility, revealing single nucleotide polymorphisms or indels located in the poorly understood CPS type IV cluster. Crucially, suppressor mutations did not result in reversion of attenuation in vivo . This study demonstrates the importance of KdsD for B. pseudomallei virulence and highlights further the complex nature of the polysaccharides it produces. IMPORTANCE The intrinsic resistance of B. pseudomallei to many antibiotics complicates treatment. This opportunistic pathogen possesses a wide range of virulence factors, resulting in severe and potentially fatal disease. Virulence factors as targets for drug development offer an alternative approach to combat pathogenic bacteria. Prior to initiating early drug discovery approaches, it is important to demonstrate that disruption of the target gene will prevent the development of disease. This study highlights the fact that KdsD is crucial for virulence of B. pseudomallei in an animal model of infection and provides supportive phenotypic characterization that builds a foundation for future therapeutic development., Competing Interests: The authors declare no conflict of interest.

Background and Objective: Plaque-induced gingival inflammation (gingivitis) is ubiquitous in humans. The epithelial barrier reacts to the presence of oral bacteria and induces inflammatory cascades. The objective of this study was to investigate the mechanism by which the small molecule micronutrient curcumin could decrease inflammatory response in vitro to oral bacterium heat-killed Fusobacterium nucleatum as curcumin could be a useful compound for combatting gingivitis already consumed by humans., Methods: H400 oral epithelial cell line was pre-conditioned with curcumin and the production of cytokines was measured by enzyme-linked immunosorbent assay (ELISA) and translocation of transcription factors was used to monitor inflammatory responses. Haem oxygenase (HO-1) expression and molecules that HO-1 releases were evaluated for their potential to reduce the quantity of cytokine production. Immunofluorescence microscopy and Western blotting were used to evaluate changes in transcription factor and enzyme location., Results: Pre-conditioning of H400 cells with a sub-apoptotic concentration of curcumin (20 μM) attenuated secretion of Granulocyte-Macrophage - Colony-Stimulating Factor (GM-CSF) and reduced NFkB nuclear translocation. This pre-conditioning caused an increase in nuclear Nrf2; an initial drop (at 8 h) followed by an adaptive increase (at 24 h) in glutathione; and an increase in haem oxygenase (HO-1) expression. Inhibition of HO-1 by SnPPIX prevented the curcumin-induced attenuation of GM-CSF production. HO-1 catalyses the breakdown of haem to carbon monoxide, free iron and biliverdin: the HO-1/CO anti-inflammatory pathway. Elevations in carbon monoxide, achieved using carbon monoxide releasing molecule-2 (CORM2) treatment alone abrogated F. nucleatum-induced cytokine production. Biliverdin is converted to bilirubin by biliverdin reductase (BVR). This pleiotropic protein was found to increase in cell membrane expression upon curcumin treatment., Conclusion: Curcumin decreased inflammatory cytokine production induced by Fusobacterium nucleatum in H400 oral epithelial cells. The mechanism of action appears to be driven by the increase of haem oxygenase and the production of carbon monoxide., (© 2023 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: The macrophage infectivity potentiator (Mip) protein, which belongs to the immunophilin superfamily, is a peptidyl-prolyl cis/trans isomerase (PPIase) enzyme. Mip has been shown to be important for virulence in a wide range of pathogenic microorganisms. It has previously been demonstrated that small-molecule compounds designed to target Mip from the Gram-negative bacterium Burkholderia pseudomallei bind at the site of enzymatic activity of the protein, inhibiting the in vitro activity of Mip., Objectives: In this study, co-crystallography experiments with recombinant B. pseudomallei Mip (BpMip) protein and Mip inhibitors, biochemical analysis and computational modelling were used to predict the efficacy of lead compounds for broad-spectrum activity against other pathogens., Methods: Binding activity of three lead compounds targeting BpMip was verified using surface plasmon resonance spectroscopy. The determination of crystal structures of BpMip in complex with these compounds, together with molecular modelling and in vitro assays, was used to determine whether the compounds have broad-spectrum antimicrobial activity against pathogens., Results: Of the three lead small-molecule compounds, two were effective in inhibiting the PPIase activity of Mip proteins from Neisseria meningitidis, Klebsiella pneumoniae and Leishmania major. The compounds also reduced the intracellular burden of these pathogens using in vitro cell infection assays., Conclusions: These results indicate that Mip is a novel antivirulence target that can be inhibited using small-molecule compounds that prove to be promising broad-spectrum drug candidates in vitro. Further optimization of compounds is required for in vivo evaluation and future clinical applications., (© The Author(s) 2022. Published by Oxford University Press on behalf of British Society for Antimicrobial Chemotherapy.)

Background: Rett syndrome (RTT) is a neurodevelopmental disorder most often related to a pathogenic variant in the X-linked MECP2 gene. Internalizing behaviors appear to be common, but standard methods of diagnosing anxiety are not readily applied in this population which typically has cognitive impairment and limited expressive language. This study aims to describe the frequency of anxiety-like behavior and anxiolytic treatments along with associated clinical features in individuals with RTT., Methods: Parental reports and medication logs provided data from 1380 females with RTT participating in two iterations of the multicenter U.S. RTT Natural History Study (RNHS) from 2006 to 2019., Results: Most participants with RTT (77.5%) had at least occasional anxious or nervous behavior. Anxiety was reported to be the most troublesome concern for 2.6%, and within the top 3 concerns for 10.0%, of participants in the second iteration. Parents directly reported treatment for anxious or nervous behavior in 16.6% of participants in the second iteration with most reporting good control of the behavior (71.6%). In the medication logs of both RNHS iterations, the indication of anxiety was listed for a similar number of participants (15% and 14.5%, respectively). Increased use of anxiolytics and selective serotonin reuptake inhibitors (SSRIs) was related to more frequent anxiety-like behaviors (P < 0.001), older age (P < 0.001), and mild MECP2 variants (P = 0.002)., Conclusion: Anxiety-like behavior is frequent at all ages and is a significant parental concern in RTT. Older individuals and those with mild MECP2 variants are more likely to be treated with medications. Better diagnosis and treatment of anxiety in RTT should be a goal of both future studies and clinical care., Trial Registration: NCT00299312 and NCT02738281., (© 2022. The Author(s).)

Burkholderia pseudomallei is the causative agent of melioidosis, a disease endemic to Southeast Asia and northern Australia. Mortality rates in these areas are high even with antimicrobial treatment, and there are few options for effective therapy. Therefore, there is a need to identify antibacterial targets for the development of novel treatments. Cyclophilins are a family of highly conserved enzymes important in multiple cellular processes. Cyclophilins catalyze the cis-trans isomerization of xaa -proline bonds, a rate-limiting step in protein folding which has been shown to be important for bacterial virulence. B. pseudomallei carries a putative cyclophilin B gene, ppiB , the role of which was investigated. A B. pseudomallei ΔppiB ( BpsΔppiB ) mutant strain demonstrates impaired biofilm formation and reduced motility. Macrophage invasion and survival assays showed that although the BpsΔppiB strain retained the ability to infect macrophages, it had reduced survival and lacked the ability to spread cell to cell, indicating ppiB is essential for B. pseudomallei virulence. This is reflected in the BALB/c mouse infection model, demonstrating the requirement of ppiB for in vivo disease dissemination and progression. Proteomic analysis demonstrates that the loss of PpiB leads to pleiotropic effects, supporting the role of PpiB in maintaining proteome homeostasis. The loss of PpiB leads to decreased abundance of multiple virulence determinants, including flagellar machinery and alterations in type VI secretion system proteins. In addition, the loss of ppiB leads to increased sensitivity toward multiple antibiotics, including meropenem and doxycycline, highlighting ppiB inhibition as a promising antivirulence target to both treat B. pseudomallei infections and increase antibiotic efficacy., (Copyright © 2019 American Society for Microbiology.)

Objective: To characterize hand stereotypies (HS) in a large cohort of participants with Rett syndrome (RTT)., Methods: Data from 1,123 girls and women enrolled in the RTT Natural History Study were gathered. Standard tests for continuous and categorical variables were used at baseline. For longitudinal data, we used repeated-measures linear and logistic regression models and nonparametric tests., Results: HS were reported in 922 participants with classic RTT (100%), 73 with atypical severe RTT (97.3%), 74 with atypical mild RTT (96.1%), and 17 females with MECP2 mutations without RTT (34.7%). Individuals with RTT who had classic presentation or severe MECP2 mutations had higher frequency and earlier onset of HS. Heterogeneity of HS types was confirmed, but variety decreased over time. At baseline, almost half of the participants with RTT had hand mouthing, which like clapping/tapping, decreased over time. These 2 HS types were more frequently reported than wringing/washing. Increased HS severity (prevalence and frequency) was associated with worsened measures of hand function. Number and type of HS were not related to hand function. Overall clinical severity was worse with decreased hand function but only weakly related to any HS characteristic. While hand function decreased over time, prevalence and frequency of HS remained relatively unchanged and high., Conclusions: Nearly all individuals with RTT have severe and multiple types of HS, with mouthing and clapping/tapping decreasing over time. Interaction between HS frequency and hand function is complex. Understanding the natural history of HS in RTT could assist in clinical care and evaluation of new interventions., (© 2019 American Academy of Neurology.)

Background: Burkholderia pseudomallei is a human pathogen causing severe infections in tropical and subtropical regions and is classified as a bio-threat agent. B. thailandensis strain E264 has been proposed as less pathogenic surrogate for understanding the interactions of B. pseudomallei with host cells., Results: We show that, unlike B. thailandensis strain E264, the pattern of growth of B. thailandensis strain E555 in macrophages is similar to that of B. pseudomallei. We have genome sequenced B. thailandensis strain E555 and using the annotated sequence identified genes and proteins up-regulated during infection. Changes in gene expression identified more of the known B. pseudomallei virulence factors than changes in protein levels and used together we identified 16% of the currently known B. pseudomallei virulence factors. These findings demonstrate the utility of B. thailandensis strain E555 to study virulence of B. pseudomallei., Conclusions: A weakness of studies using B. thailandensis as a surrogate for B. pseudomallei is that the strains used replicate at a slower rate in infected cells. We show that the pattern of growth of B. thailandensis strain E555 in macrophages closely mirrors that of B. pseudomallei. Using this infection model we have shown that virulence factors of B. pseudomallei can be identified as genes or proteins whose expression is elevated on the infection of macrophages. This finding confirms the utility of B. thailandensis strain E555 as a surrogate for B. pseudomallei and this strain should be used for future studies on virulence mechanisms.

The highly virulent intracellular pathogen Francisella tularensis is a Gram-negative bacterium that has a wide host range, including humans, and is the causative agent of tularemia. To identify new therapeutic drug targets and vaccine candidates and investigate the genetic basis of Francisella virulence in the Fischer 344 rat, we have constructed an F. tularensis Schu S4 transposon library. This library consists of more than 300,000 unique transposon mutants and represents a transposon insertion for every 6 bp of the genome. A transposon-directed insertion site sequencing (TraDIS) approach was used to identify 453 genes essential for growth in vitro Many of these essential genes were mapped to key metabolic pathways, including glycolysis/gluconeogenesis, peptidoglycan synthesis, fatty acid biosynthesis, and the tricarboxylic acid (TCA) cycle. Additionally, 163 genes were identified as required for fitness during colonization of the Fischer 344 rat spleen. This in vivo selection screen was validated through the generation of marked deletion mutants that were individually assessed within a competitive index study against the wild-type F. tularensis Schu S4 strain. IMPORTANCE The intracellular bacterial pathogen Francisella tularensis causes a disease in humans characterized by the rapid onset of nonspecific symptoms such as swollen lymph glands, fever, and headaches. F. tularensis is one of the most infectious bacteria known and following pulmonary exposure can have a mortality rate exceeding 50% if left untreated. The low infectious dose of this organism and concerns surrounding its potential as a biological weapon have heightened the need for effective and safe therapies. To expand the repertoire of targets for therapeutic development, we initiated a genome-wide analysis. This study has identified genes that are important for F. tularensis under in vitro and in vivo conditions, providing candidates that can be evaluated for vaccine or antibacterial development., (© Crown copyright 2019.)

We have resequenced the genomes of four Burkholderia pseudomallei K96243 laboratory cultures and compared them to the reported genome sequence that was published in 2004. Compared with the reference genome, these laboratory cultures harbored up to 42 single-nucleotide variants and up to 11 indels, including a 31.7-kb deletion in one culture.

Introduction: Rett syndrome (RTT) is a complex neurodevelopmental disorder with known behavioral abnormalities, both internalizing (e.g., anxiety, social withdrawal) and externalizing (e.g., aggression, self-abuse). However, a broad evaluation of behavioral abnormalities in a large cohort is lacking., Objective: In this report, we describe profiles of internalizing and externalizing behaviors in individuals evaluated in the multi-center U.S. Rett Natural History Study., Methods: Cross-sectional and longitudinal data were collected from 861 females with RTT and from 48 females who have MECP2 mutations without meeting criteria for RTT. Standard statistical methods including linear regression evaluated internalizing behavioral components from the Child Health Questionnaire (CHQ-PF50) and externalizing components from the Motor Behavioral Assessment (MBA)., Results: We found mildly to moderately severe internalizing behaviors in nearly all individuals with RTT, while externalizing behaviors were mild and uncommon. Internalizing behavior in RTT was comparable to groups with psychiatric disorders. Participants with mixed (internalizing and externalizing) behaviors were younger and less affected overall, but showed prominent self-injury and worsening internalizing behaviors over time., Conclusions: This study revealed that internalizing behaviors are common at a clinically significant level in RTT. Understanding clinical features associated with behavioral profiles could guide treatment strategies., (Copyright © 2018 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.)

Objectives: We evaluated the association of health literacy and attention to the pictorial imagery of 9 health warning labels (HWLs) in a tobacco advertisement among rural US smokers. We hypothesized that lower health literacy would be associated with greater visual attention to pictorial portions of HWLs and evaluated the association between health literacy and recall of advertisement elements., Methods: Adult smokers from Appalachian Ohio (N = 180) viewed a pictorial HWL encompassing 20% or 33% of a cigarette advertisement while eye tracking software recorded eye movements toward the advertisement. Health literacy was measured with The Short Test of Functional Health Literacy in Adults., Results: Generalized linear regression revealed that every one-unit decrease in health literacy increased viewing time of the pictorial portion of the health warning label by 1.3 percentage points. Logistic regression revealed that the odds of recalling elements of the pictorial portion of the health warning label increased 20% for every one-unit increase in health literacy., Conclusions: Rural smokers with lower health literacy view pictorial portions of health warning labels longer than those with greater health literacy supporting that health literacy is an important consideration in health communications, including future cigarette warning labels.

Objective: To describe maternal characteristics related to early screening for diabetes in obese women and evaluate the benefits of early diabetes screening and diagnosis., Study Design: Retrospective cohort of obese women (BMI ≥30 kg/m 2 ) without pregestational diabetes who delivered a singleton gestation between 2011 and 2012. Maternal characteristics/demographics and maternal and neonatal outcomes were compared between women with early diabetes screening (<20 weeks) versus traditional screening. We additionally compared maternal and neonatal outcomes for women with an early versus traditional diabetes diagnosis., Results: Of the 504 eligible women, 135 (26.8%) had early diabetes screening. Obese women with early screening were older, had a higher BMI, were more likely to have hypertension and neonates admitted to the NICU. Of women with early screening, 31 (23%) were diagnosed early. Women with an early diagnosis of diabetes were more likely to require treatment with insulin (36% vs. 23%, p = 0.003). Women with an early diagnosis of diabetes were more likely to have neonates in the NICU (48% vs. 26%, p = 0.03)., Conclusions: Early screening for diabetes was more common in older women with additional comorbidities. Obese women diagnosed via early screening were more likely to require medical treatment for diabetes, suggesting a value to early screening.

Burkholderia pseudomallei and its close relative B. mallei are human pathogens that are classified as Tier 1 bio-threat agents. Both organisms have previously been shown to constitutively produce a capsular polysaccharide (CPS) that is both a virulence determinant and protective antigen. Extraction and purification of CPS for use as a potential vaccine candidate requires containment level 3 laboratories which is expensive and time-consuming. B. thailandensis strain E555 is closely related to B. pseudomallei and B. mallei, but is non-pathogenic to humans and based on immunological cross-reactivity has previously been shown to express a B. pseudomallei-like CPS. In this study, capsular polysaccharide isolated from an O-antigen deficient strain of B. thailandensis E555 was identified by 1 H and 13 C NMR spectroscopy as -3-)-2-O-acetyl-6-deoxy-β-d-manno-heptopyranose-(-1, and identical to that produced by B. pseudomallei. This was further substantiated by anti-CPS monoclonal antibody binding. In connection with the production of CPS fragments for use in glycoconjugate vaccines, we set out to assess the importance or otherwise of the CPS 2-OAc groups in immune protection. To this end conjugates of the native and de-O-acetylated CPS with the Hc fragment of tetanus toxin (TetH c ) were used as vaccines in a mouse model of melioidosis. The level of protection provided by deacetylated CPS was significantly lower than that from native, acetylated CPS. In addition, sera from mice vaccinated with the deacetylated CPS conjugate did not recognise native CPS. This suggests that CPS extracted from B. thailandensis can be used as antigen and that the acetyl group is essential for protection., (Crown Copyright © 2017. Published by Elsevier Ltd. All rights reserved.)

Background: The World Health Organization has categorized plague as a re-emerging disease and the potential for Yersinia pestis to also be used as a bioweapon makes the identification of new drug targets against this pathogen a priority. Environmental temperature is a key signal which regulates virulence of the bacterium. The bacterium normally grows outside the human host at 28 °C. Therefore, understanding the mechanisms that the bacterium used to adapt to a mammalian host at 37 °C is central to the development of vaccines or drugs for the prevention or treatment of human disease., Results: Using a library of over 1 million Y. pestis CO92 random mutants and transposon-directed insertion site sequencing, we identified 530 essential genes when the bacteria were cultured at 28 °C. When the library of mutants was subsequently cultured at 37 °C we identified 19 genes that were essential at 37 °C but not at 28 °C, including genes which encode proteins that play a role in enabling functioning of the type III secretion and in DNA replication and maintenance. Using genome-scale metabolic network reconstruction we showed that growth conditions profoundly influence the physiology of the bacterium, and by combining computational and experimental approaches we were able to identify 54 genes that are essential under a broad range of conditions., Conclusions: Using an integrated computational-experimental approach we identify genes which are required for growth at 37 °C and under a broad range of environments may be the best targets for the development of new interventions to prevent or treat plague in humans.

Sphingosine-1-phosphate (S1P), a bioactive sphingolipid metabolite, plays a critical role in the orchestration of immune responses. S1P levels within the mammalian host are tightly regulated, in part through the activity of S1P lyase (S1PL) which catalyses its irreversible degradation. Herein, we describe the identification and characterization of secreted S1PL orthologues encoded by the facultative intracellular bacteria Burkholderia pseudomallei and Burkholderia thailandensis. These bacterial orthologues exhibited S1PL enzymatic activity, functionally complemented an S1PL-deficient yeast strain and conferred resistance to the antimicrobial sphingolipid D-erythro-sphingosine. We report that secretion of these bacterial S1PLs is pH-dependent, and is observed during intracellular infection. S1PL-deficient mutants displayed impaired intracellular replication in murine macrophages (associated with an inability to evade the maturing phagosome) and were significantly attenuated in murine and larval infection models. Furthermore, treatment of Burkholderia-infected macrophages with either S1P or a selective agonist of S1P receptor 1 enhanced bacterial colocalisation with LAMP-1 and reduced their intracellular survival. In summary, our studies confirm bacterial-encoded S1PL as a critical virulence determinant of B. pseudomallei and B. thailandensis, further highlighting the pivotal role of S1P in host-pathogen interactions. In addition, our data suggest that S1P pathway modulators have potential for the treatment of intracellular infection., (© 2016 John Wiley & Sons Ltd.)

Melioidosis is an emerging infectious disease caused by Burkholderia pseudomallei and is associated with high morbidity and mortality rates in endemic areas. Antibiotic treatment is protracted and not always successful; even with appropriate therapy, up to 40% of individuals presenting with melioidosis in Thailand succumb to infection. In these circumstances, an effective vaccine has the potential to have a dramatic impact on both the scale and the severity of disease. Currently, no vaccines are licensed for human use. A leading vaccine candidate is the capsular polysaccharide consisting of a homopolymer of unbranched 1→3 linked 2-O-acetyl-6-deoxy-β-d-manno-heptopyranose. Here, we present the chemical synthesis of this challenging antigen using a novel modular disaccharide assembly approach. The resulting hexasaccharide was coupled to the nontoxic Hc domain of tetanus toxin as a carrier protein to promote recruitment of T-cell help and provide a scaffold for antigen display. Mice immunized with the glycoconjugate developed IgM and IgG responses capable of recognizing native capsule, and were protected against infection with over 120 × LD50 of B. pseudomallei strain K96243. This is the first report of the chemical synthesis of an immunologically relevant and protective hexasaccharide fragment of the capsular polysaccharide of B. pseudomallei and serves as the rational starting point for the development of an effective licensed vaccine for this emerging infectious disease.

There is an urgent need for an effective vaccine against human disease caused by Burkholderia pseudomallei, and although a wide range of candidates have been tested in mice none provide high level protection. We considered this might reflect the inability of these vaccine candidates to protect against chronic disease. Using Q-RT PCR we have identified 6 genes which are expressed in bacteria colonising spleens and lungs of chronically infected mice. Three of the genes (BPSL1897, BPSL3369 and BPSL2287) have been expressed in Escherichia coli and the encoded proteins purified. We have also included BPSL2765, a protein known to induce immune responses associated with a reduced incidence of chronic/recurrent disease in humans. Immunisation of mice with a combination of these antigens resulted in the induction of antibody responses against all of the proteins. Compared with mice immunised with capsular polysaccharide or LolC protein, mice immunised with the combination of chronic stage antigens showed enhanced protection against experimental disease in mice., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Plague is a highly pathogenic disease caused by the bacterium Yersinia pestis. There is currently no vaccine available for prophylaxis and antibiotic resistant strains have been isolated, thus there is a need for the development of new countermeasures to treat this disease. Survival protein A (SurA) is a chaperone that has been linked to virulence in several species of bacteria, including the close relative Yersinia pseudotuberculosis. In this study, we aimed to evaluate the role of SurA in virulence of the highly pathogenic Y. pestis by creating an unmarked surA deletion mutant. The Y. pestis ΔsurA mutant was found to be more susceptible to membrane perturbing agents and was completely avirulent in a mouse infection model when delivered up to 2.1 × 10(5) CFU by the subcutaneous route. This provides strong evidence that SurA would make a promising antimicrobial target., (Crown Copyright © 2016. Published by Elsevier Ltd. All rights reserved.)

Aim: The aim of this study was to investigate the association of serum vitamin B12 with the progression of periodontitis and risk of tooth loss in a prospective cohort study., Materials and Methods: In the Study of Health in Pomerania, 1648 participants were followed from 2002-2006 to 2008-2012 (mean duration 5.9 years). Serum vitamin B12 was measured by chemiluminescent enzyme immunoassay. Probing pocket depth (PD) and clinical attachment loss (CAL) were measured to reflect periodontal status on a half-mouth basis at each survey cycle. Tooth numbers are based upon a full-mouth tooth count., Results and Conclusions: In multivariate regression models, baseline vitamin B12 was inversely associated with changes in mean PD (Ptrend = 0.06) and mean CAL (Ptrend = 0.01), and risk ratios of tooth loss (TL; Ptrend = 0.006) over time. Compared to participants in the highest vitamin B12 quartile, those in the lowest quartile had 0.10 mm (95%CI: 0.03, 0.17; Pdifference = 0.007) greater increase in mean PD, 0.23 mm (95%CI: 0.09, 0.36; Pdifference = 0.001) greater increase in mean CAL and a relative risk of 1.57 (95%CI: 1.22, 2.03; Pdifference < 0.001) for TL. Stratified analyses showed stronger associations between vitamin B12 and changes in mean CAL among never smokers (Pinteraction = 0.058). Further studies are needed to understand the potential mechanisms of these findings., (© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Gram-negative bacteria utilize heptoses as part of their repertoire of extracellular polysaccharide virulence determinants. Disruption of heptose biosynthesis offers an attractive target for novel antimicrobials. A critical step in the synthesis of heptoses is their 1-O phosphorylation, mediated by kinases such as HldE or WcbL. Here, we present the structure of WcbL from Burkholderia pseudomallei. We report that WcbL operates through a sequential ordered Bi-Bi mechanism, loading the heptose first and then ATP. We show that dimeric WcbL binds ATP anti-cooperatively in the absence of heptose, and cooperatively in its presence. Modeling of WcbL suggests that heptose binding causes an elegant switch in the hydrogen-bonding network, facilitating the binding of a second ATP molecule. Finally, we screened a library of drug-like fragments, identifying hits that potently inhibit WcbL. Our results provide a novel mechanism for control of substrate binding and emphasize WcbL as an attractive anti-microbial target for Gram-negative bacteria., (Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Understanding the three-dimensional (3-D) micro-architecture of lung tissue can provide insights into the pathology of lung disease. Micro computed tomography (µCT) has previously been used to elucidate lung 3D histology and morphometry in fixed samples that have been stained with contrast agents or air inflated and dried. However, non-destructive microstructural 3D imaging of formalin-fixed paraffin embedded (FFPE) tissues would facilitate retrospective analysis of extensive tissue archives of lung FFPE lung samples with linked clinical data., Methods: FFPE human lung tissue samples (n = 4) were scanned using a Nikon metrology µCT scanner. Semi-automatic techniques were used to segment the 3D structure of airways and blood vessels. Airspace size (mean linear intercept, Lm) was measured on µCT images and on matched histological sections from the same FFPE samples imaged by light microscopy to validate µCT imaging., Results: The µCT imaging protocol provided contrast between tissue and paraffin in FFPE samples (15 mm x 7 mm). Resolution (voxel size 6.7 µm) in the reconstructed images was sufficient for semi-automatic image segmentation of airways and blood vessels as well as quantitative airspace analysis. The scans were also used to scout for regions of interest, enabling time-efficient preparation of conventional histological sections. The Lm measurements from µCT images were not significantly different to those from matched histological sections., Conclusion: We demonstrated how non-destructive imaging of routinely prepared FFPE samples by laboratory µCT can be used to visualize and assess the 3D morphology of the lung including by morphometric analysis.

Background: Previous experimental models suggest that vitamin E may ameliorate periodontitis. However, epidemiologic studies show inconsistent evidence in supporting this plausible association., Objective: We investigated the association between serum α-tocopherol (αT) and γ-tocopherol (γT) and periodontitis in a large cross-sectional US population., Methods: This study included 4708 participants in the 1999-2001 NHANES. Serum tocopherols were measured by HPLC and values were adjusted by total cholesterol (TC). Periodontal status was assessed by mean clinical attachment loss (CAL) and probing pocket depth (PPD). Total periodontitis (TPD) was defined as the sum of mild, moderate, and severe periodontitis. All measurements were performed by NHANES., Results: Means ± SDs of serum αT:TC ratio from low to high quartiles were 4.0 ± 0.4, 4.8 ± 0.2, 5.7 ± 0.4, and 9.1 ± 2.7 μmol/mmol. In multivariate regression models, αT:TC quartiles were inversely associated with mean CAL (P-trend = 0.06), mean PPD (P-trend < 0.001), and TPD (P-trend < 0.001) overall. Adjusted mean differences (95% CIs) between the first and fourth quartile of αT:TC were 0.12 mm (0.03, 0.20; P-difference = 0.005) for mean CAL and 0.12 mm (0.06, 0.17; P-difference < 0.001) for mean PPD, whereas the corresponding OR for TPD was 1.65 (95% CI: 1.26, 2.16; P-difference = 0.001). In a dose-response analysis, a clear inverse association between αT:TC and mean CAL, mean PPD, and TPD was observed among participants with relatively low αT:TC. No differences were seen in participants with higher αT:TC ratios. Participants with γT:TC ratio in the interquartile range showed a significantly lower mean PPD than those in the highest quartile., Conclusions: A nonlinear inverse association was observed between serum αT and severity of periodontitis, which was restricted to adults with normal but relatively low αT status. These findings warrant further confirmation in longitudinal or intervention studies., (© 2015 American Society for Nutrition.)

Burkholderia pseudomallei, the etiologic agent of melioidosis, is a CDC tier 1 select agent that causes severe disease in both humans and animals. Diagnosis and treatment of melioidosis can be challenging, and in the absence of optimal chemotherapeutic intervention, acute disease is frequently fatal. Melioidosis is an emerging infectious disease for which there are currently no licensed vaccines. Due to the potential malicious use of B. pseudomallei as well as its impact on public health in regions where the disease is endemic, there is significant interest in developing vaccines for immunization against this disease. In the present study, type A O-polysaccharide (OPS) and manno-heptose capsular polysaccharide (CPS) antigens were isolated from nonpathogenic, select-agent-excluded strains of B. pseudomallei and covalently linked to carrier proteins. By using these conjugates (OPS2B1 and CPS2B1, respectively), it was shown that although high-titer IgG responses against the OPS or CPS component of the glycoconjugates could be raised in BALB/c mice, only those animals immunized with CPS2B1 were protected against intraperitoneal challenge with B. pseudomallei. Extending upon these studies, it was also demonstrated that when the mice were immunized with a combination of CPS2B1 and recombinant B. pseudomallei LolC, rather than with CPS2B1 or LolC individually, they exhibited higher survival rates when challenged with a lethal dose of B. pseudomallei. Collectively, these results suggest that CPS-based glycoconjugates are promising candidates for the development of subunit vaccines for immunization against melioidosis., (Copyright © 2014, American Society for Microbiology. All Rights Reserved.)

The United States (U.S.) response to viral hemorrhagic septicemia virus (VHSV) IVb emergence in the Laurentian Great Lakes (GL) included risk-based surveillance for cost-effective decision support regarding the health of fish populations in open systems. All U.S. VHSV IVb isolations to date derive from free-ranging fish from GL States. Most originate in the region designated by US Geological Survey hydrologic unit code (HUC) 04, with the exception of two detections in neighboring Upper Mississippi (HUC 05) and Ohio (HUC 07) regions. For States outside the GL system, disease probability was assessed using multiple evidence sources. None substantiated VHSV IVb absence using surveillance alone, in part due to the limited temporal relevance of data in open systems. However, Bayesian odds risk-based analysis of surveillance and population context, coupled with exclusions where water temperatures likely preclude viral replication, achieved VHSV IVb freedom assurance for 14 non-GL States by the end of 2012, with partial evidence obtained for another 17 States. The non-GL region (defined as the aggregate of 4-digit HUCs located outside of GL States) met disease freedom targets for 2012 and is projected to maintain this status through 2016 without additional active surveillance. Projections hinge on continued basic biosecurity conditions such as movement restrictions and passive surveillance. Areas with navigable waterway connections to VHSV IVb-affected HUCs (and conducive water temperatures) should receive priority for resources in future surveillance or capacity building efforts. However, 6 years of absence of detections in non-GL States suggests that existing controls limit pathogen spread, and that even spread via natural pathways (e.g., water movement or migratory fish) appears contained to the Great Lakes system. This report exemplifies the cost-effective use of risk-based surveillance in decision support to assess and manage aquatic animal population health in open systems., (Published by Elsevier B.V.)

Objective: We aimed to establish the feasibility of single-heartbeat 320-multidetector computed tomographic coronary angiography (CTCA) and assess variables affecting image quality., Methods: Consecutive patients (n = 249, 38% male) underwent CTCA. Two observers assessed image quality using a 4-point scale (1, excellent; 4, poor)., Results: Mean heart rate was 60 beats per minute (95% confidence interval, 59-62); body mass index, 29 kg/m (28-30); and dose-length product, 283 mGy·cm (266-301). During scanning, 133 (51%) received sublingual glyceryl trinitrate (GTN), 9 (4%) had ectopics, and 12 (5%) had atrial fibrillation. Diagnostic image quality was obtained in 99% with mean image quality of 1.4 (1.3, 1.5). Age, sex, atrial fibrillation, ectopics, diabetes mellitus (12%), and obstructive disease were not related to image quality. A lower heart rate and GTN were associated with improved image quality (P ≤ 0.001)., Conclusions: Optimal image quality in single-heartbeat 320-multidetector CTCA is achievable in 99% of unselected patients. Image quality is improved by lower heart rate and GTN.

Objective: To determine the prevalence of feeding dysfunction in children with single ventricle defects and identify associated risk factors., Study Design: Patients aged 2-6 years with single ventricle physiology presenting for routine cardiology follow-up at the Children's Hospital of Wisconsin were prospectively identified. Parents of the patients completed 2 validated instruments for assessment of feeding dysfunction. Chart review was performed to retrospectively obtain demographic and diagnostic data., Results: Instruments were completed for 56 patients; median age was 39 months. Overall, 28 (50%) patients had some form of feeding dysfunction. Compared with a normal reference population, patients with single ventricle had statistically significant differences in dysfunctional food manipulation (P < .001), mealtime aggression (P = .002), choking/gagging/vomiting (P < .001), resistance to eating (P < .001), and parental aversion to mealtime (P < .001). Weight and height for age z-scores were significantly lower in subjects with feeding dysfunction (-0.84 vs -0.33; P < .05 and -1.46 vs -0.56; P = .001, respectively). Multivariable analysis identified current gastrostomy tube use (P = .02) and a single parent household (P = .01) as risk factors for feeding dysfunction., Conclusion: Feeding dysfunction is common in children with single ventricle defects, occurring in 50% of our cohort. Feeding dysfunction is associated with worse growth measures. Current gastrostomy tube use and a single parent household were identified as independent risk factors for feeding dysfunction., (Copyright © 2014 Mosby, Inc. All rights reserved.)

Melioidosis is a severe infectious disease caused by Burkholderia pseudomallei. It is refractory to antibiotic treatment and there is currently no licensed vaccine. In this report we detail the construction and protective efficacy of a polysaccharide-protein conjugate composed of B. pseudomallei lipopolysaccharide and the Hc fragment of tetanus toxin. Immunisation of mice with the lipopolysaccharide-conjugate led to significantly reduced bacterial burdens in the spleen 48 hours after challenge and afforded significant protection against a lethal challenge with B. pseudomallei. The conjugate generated significantly higher levels of antigen-specific IgG1 and IgG2a than in lipopolysaccharide-immunised mice. Immunisation with the conjugate also demonstrated a bias towards Th1 type responses, evidenced by high levels of IgG2a. In contrast, immunisation with unconjugated lipopolysaccharide evoked almost no IgG2a demonstrating a bias towards Th2 type responses. This study demonstrates the effectiveness of this approach in the development of an efficacious and protective vaccine against melioidosis.

Aims: To examine the relative and combined value of late gadolinium enhancement (LGE) and low-dose dobutamine (LDD) cardiac magnetic resonance (CMR) to predict 'adverse remodelling' (AR) following acute myocardial infarction (AMI)., Methods and Results: Forty-five patients with AMI were recruited. CMR was performed 2-4 days after presentation and at 6 months. Ventricular wall motion and volume were recorded at rest and following dobutamine infusion. Measures of first pass perfusion, persistent microvascular obstruction (PMO), and LGE were obtained following contrast administration. Quantitation was performed using the MEDIS 6.2 software. Regression analysis was employed to determine the univariables and multivariate models most predictive of AR at 6 months. The incremental and relative value of LDD over LGE was investigated. The most predictive univariable was 'volume of PMO' (r = 0.51, r2 = 0.26, P < 0.001). The optimal 'combined' multivariate model, utilizing data from all components, was highly predictive of AR (r = 0.82, r2 = 0.67, P < 0.001). The optimal model using parameters only from the LGE component also predicted remodelling (r = 0.65, r2 = 42.0, P = 0.001) but with less accuracy. In contrast, the optimal model using variables from the LDD component alone predicted remodelling with a similar accuracy to the optimal combined model (r = 0.82, r2 = 0.67, P < 0.001)., Conclusion: A comprehensive CMR examination accurately predicts AR following AMI. LDD is superior to LGE CMR in this respect. These data suggest that LDD not only adds incremental value to LGE in the prediction of remodelling post-AMI but also may be utilized alone with the same predictive power.

Melioidosis is a severe infectious disease caused by Burkholderia pseudomallei. It is highly resistant to antibiotic treatment, and there is currently no licensed vaccine. Burkholderia thailandensis is a close relative of Burkholderia pseudomallei but is essentially avirulent in mammals. In this report, we detail the protective efficacy of immunization with live B. thailandensis E555, a strain which has been shown to express an antigenic capsule similar to that of B. pseudomallei. Immunization with E555 induced significant protection against a lethal intraperitoneal B. pseudomallei challenge in a mouse model of infection, with no mice succumbing to infection over the course of the study, even with challenges of up to 6,000 median lethal doses. By comparison, mice immunized with B. thailandensis not expressing a B. pseudomallei-like capsule had significantly decreased levels of protection. E555-immunized mice had significantly higher levels of IgG than mice immunized with noncapsulated B. thailandensis, and these antibody responses were primarily directed against the capsule.

Periodontitis, a ubiquitous chronic inflammatory disease, is associated with reduced antioxidant defences and neutrophil hyperactivity in terms of reactive oxygen species (ROS) generation. Its phenotype is thus characterized by oxidative stress. We have determined the effect of antioxidant micronutrients ascorbate and α-tocopherol on neutrophil ROS generation. Peripheral neutrophils from periodontally-healthy individuals (n = 20) were challenged with phorbol myristate acetate, IgG-opsonised Staphylococcus aureus, Fusobacterium nucleatum or PBS in the presence and absence of micronutrients (50 µM). Total and extracellular ROS were measured by luminol and isoluminol chemiluminescence respectively. Total and extracellular unstimulated, baseline ROS generation was unaffected by α-tocopherol, but inhibited by ascorbate and a combination of both micronutrients. Fcγ-receptor (Fcγ-R)-stimulated total or extracellular ROS generation was not affected by the presence of individual micronutrients. However, the combination significantly reduced extracellular FcγR-stimulated ROS release. Neither micronutrient inhibited TLR-stimulated total ROS, but the combination caused inhibition. Ascorbate and the micronutrient combination, but not α-tocopherol, inhibited extracellular ROS release by TLR-stimulated cells. Such micronutrient effects in vivo could be beneficial in reducing collateral tissue damage in chronic inflammatory diseases, such as periodontitis, while retaining immune-mediated neutrophil function.

On 10 May 1893, William Gowers began a series of weekly clinical demonstrations at the National Hospital for the Relief and Cure of the Paralysed and Epileptic at Queen Square, London. The contents of some of these demonstrations were published as 'Post-graduate Clinical Lectures' in the Clinical Journal, and in other learned periodicals. Some were also later included in his book Clinical Lectures on Diseases of the Nervous System. Recently, the manuscripts of what appear to be verbatim transcripts of two further but unpublished demonstrations from Gowers' course in 1895 came to light, one containing alterations made in Gowers' handwriting. The first concerned a case of disseminated sclerosis and its differentiation from hysterical paraplegia, the second transverse myelitis and its consequences for bladder function. Why these lectures were never published remains uncertain, but their relatively unedited contents reveal something of the neurological knowledge, diagnostic reasoning, clinical examination and teaching methods employed by one of the great pioneers of clinical neurology.

A weighted benefit-cost analysis (BCA) supports prioritization of animal health surveillance activities to safeguard animal agriculture industries and reduce the impact of disease on the national economy. We propose to determine the value of investment in surveillance by assessing benefits from: avoiding disease incursion and expansion modified by the probability of occurrence of the disease event, the sensitivity of systems to detect it, and the degree to which we can mitigate disease impact when detected. The weighted benefit-cost ratio is the modified value of surveillance as laid out above divided by the cost of surveillance. We propose flexible, stream-based surveillance that capitalizes on combining multiple streams of information from both specific pathogen based and non-pathogen based surveillance. This stream-based type of system provides high value with lower costs and will provide a high return for the funds invested in animal health surveillance., (Published by Elsevier B.V.)

Burkholderia pseudomallei is the causative agent of melioidosis, a disease endemic to regions of Southeast Asia and Northern Australia. Both humans and a range of other animal species are susceptible to melioidosis, and the production of a group 3 polysaccharide capsule in B. pseudomallei is essential for virulence. B. pseudomallei capsular polysaccharide (CPS) I comprises unbranched manno-heptopyranose residues and is encoded by a 34.5-kb locus on chromosome 1. Despite the importance of this locus, the role of all of the genes within this region is unclear. We inactivated 18 of these genes and analyzed their phenotype using Western blotting and immunofluorescence staining. Furthermore, by combining this approach with bioinformatic analysis, we were able to develop a model for CPS I biosynthesis and export. We report that inactivating gmhA, wcbJ, and wcbN in B. pseudomallei K96243 retains the immunogenic integrity of the polysaccharide despite causing attenuation in the BALB/c murine infection model. Mice immunized with the B. pseudomallei K96243 mutants lacking a functional copy of either gmhA or wcbJ were afforded significant levels of protection against a wild-type B. pseudomallei K96243 challenge.

Aim: To characterize and map temporal changes in the biological and clinical phenotype during a 21-day experimental gingivitis study., Materials and Methods: Experimental gingivitis was induced over 21 days in healthy human volunteers (n = 56), after which normal brushing was resumed (resolution phase). Gingival and plaque indices were assessed. Gingival crevicular fluid was collected from four paired test and contra-lateral control sites in each volunteer during induction (Days 0, 7, 14 and 21) and resolution (Days 28 and 42) of experimental gingivitis. Fluid volumes were measured and a single analyte was quantified from each site-specific, 30s sample. Data were evaluated by analysis of repeated measurements and paired sample tests., Results: Clinical indices and gingival crevicular fluid volumes at test sites increased from Day 0, peaking at Day 21 (test/control differences all p < 0.0001) and decreased back to control levels by Day 28. Levels of four inflammatory markers showed similar patterns, with significant differences between test and control apparent at Day 7 (substance P, cathepsin G, interleukin-1β, elastase: all p < 0.03) and peaking at Day 21 (all p < 0.002). Levels of α-1-antitrypsin showed no pattern., Conclusions: Levels of substance P, cathepsin G, interleukin-1β and neutrophil elastase act as objective biomarkers of gingival inflammation induction and resolution that typically precede phenotypical changes., (© 2011 John Wiley & Sons A/S.)

The goal of this study was to evaluate the test sensitivity (SE) and specificity (SP) of the gamma interferon (G-IFN) assay used for the detection of bovine tuberculosis (bTB) in U.S. cattle herds. In addition, the study assessed the association between G-IFN test results and bTB status of cattle, and explored different cut off values for classification of test results in adult cattle using receiver operating characteristics (ROC) curve analysis. Test SE was estimated using a population of 87 confirmed infected cattle from 14 herds distributed in 6 states. Test SP was estimated using a population of 4123 cattle representing 3000 premises in 3 states. These animals were from bTB free areas, accredited bTB free herds, or herds that were historically bTB free based on the absence of lesions found at slaughter and historical records of negative tests performed for bTB surveillance. The distribution of G-IFN results and its association with bTB infection status was also explored in a group of 914 exposed cattle in which infection was not confirmed. The results showed that the SE of the G-IFN for a cut-off value ≥0.1 was 83.9% (76.1, 91.6). The SP of the G-IFN was 90.7% (95% CI: 89.8, 91.6), 97% (95% CI: 96.5, 97.5), and 98.6%(95% CI: 98.2, 98.9), for cut off values of 0.1, 0.3, and 0.5, respectively. For a cut off value ≥0.1, the likelihood ratio of a positive G-IFN test was 9.03 (95% CI: 7.90, 10.31), and the likelihood ratio of a negative G-IFN test was 0.18 (95% CI: 0.11, 0.29). The area under the ROC curve was 0.976 (95% CI: 0.97, 0.98), characteristic of a highly accurate test. ROC analysis also showed that lower cut-off values, such as 0.1, have high SE with suitable SP for use in parallel testing, while cut-off values ranging between 0.3 and 0.6 provide the high SP desired in series-testing protocols with lower SE values. Findings from this study indicated that the G-IFN performs with high accuracy in the field, yielding SE and SP estimates comparable to those reported in previous evaluations (Ryan et al., 2000; Ameni et al., 2000; de la Rua-Domenech et al., 2006; Gormley et al., 2006)., (Published by Elsevier B.V.)

Glycosylation of proteins is known to impart novel physical properties and biological roles to proteins from both eukaryotes and prokaryotes. In this study, gel-based glycoproteomics were used to identify glycoproteins of the potential biothreat agent Burkholderia pseudomallei and the closely related but nonpathogenic B. thailandensis. Top-down and bottom-up mass spectrometry (MS) analyses identified that the flagellin proteins of both species were posttranslationally modified by novel glycans. Analysis of proteins from two strains of each species demonstrated that B. pseudomallei flagellin proteins were modified with a glycan with a mass of 291 Da, while B. thailandensis flagellin protein was modified with related glycans with a mass of 300 or 342 Da. Structural characterization of the B. thailandensis carbohydrate moiety suggests that it is an acetylated hexuronic acid. In addition, we have identified through mutagenesis a gene from the lipopolysaccharide (LPS) O-antigen biosynthetic cluster which is involved in flagellar glycosylation, and inactivation of this gene eliminates flagellar glycosylation and motility in B. pseudomallei. This is the first report to conclusively demonstrate the presence of a carbohydrate covalently linked to a protein in B. pseudomallei and B. thailandensis, and it suggests new avenues to explore in order to examine the marked differences in virulence between these two species.

Background: Neuroinflammation is a complex process involving cells from the immune system and the central nerve system (CNS). Polymorphonuclear neutrophils (PMN) are the most abundant class of white blood cells, and typically the first type of leukocyte recruited to sites of inflammation. In the CNS, astrocytes are the most abundant glial cell population and participate in the local innate immune response triggered by a variety of insults. In the present study, we investigated the impacts of astrocytes on PMN function., Methods: Primary astrocyte cultures were derived from postnatal C57BL/6 mice and primary neutrophils were isolated from 8 to 12 weeks old C57BL/6 mice. PMNs respiratory burst was analyzed by H2DCFDA assay. For phagocytosis assay, neutrophils were incubated with FITC-labeled E. coli and the phagocytosis of E coli was determined by flow cytometer. PMNs degranulation was determined by myeloperoxidase assay. Cytokine expression was determined by real-time PCR. To determine the involvement of different signaling pathway, protein lysates were prepared and western blots were conducted to assess the activation of Akt, Erk1/2, and p38., Results: Using ex vivo neutrophils and primary astrocyte cultures, our study demonstrated that astrocytes differentially regulate neutrophil functions, depending upon whether the interactions between the two cell types are direct or indirect. Upon direct cell-cell contact, astrocytes attenuate neutrophil apoptosis, respiratory bust, and degranulation, while enhancing neutrophil phagocytic capability and pro-inflammatory cytokine expression. Through indirect interaction with neutrophils, astrocytes attenuate apoptosis and enhance necrosis in neutrophils, augment neutrophil phagocytosis and respiratory burst, and inhibit neutrophil degranulation. In addition, astrocytes could augment Akt, Erk1/2, and p38 activation in neutrophils., Conclusions: Astrocytes differentially regulate neutrophil functions through direct or indirect interactions between the two cell types. The diversified actions of astrocytes on neutrophils might provide protection against potential microbial infections given compromised blood-brain barrier integrity under certain neuropathological conditions. The complex actions of astrocytes on neutrophils could provide further insight to harness the inflammatory response to promote CNS repair.

The 21-day experimental gingivitis model, an established noninvasive model of inflammation in response to increasing bacterial accumulation in humans, is designed to enable the study of both the induction and resolution of inflammation. Here, we have analyzed gingival crevicular fluid, an oral fluid comprising a serum transudate and tissue exudates, by LC-MS/MS using Fourier transform ion cyclotron resonance mass spectrometry and iTRAQ isobaric mass tags, to establish meta-proteomic profiles of inflammation-induced changes in proteins in healthy young volunteers. Across the course of experimentally induced gingivitis, we identified 16 bacterial and 186 human proteins. Although abundances of the bacterial proteins identified did not vary temporally, Fusobacterium outer membrane proteins were detected. Fusobacterium species have previously been associated with periodontal health or disease. The human proteins identified spanned a wide range of compartments (both extracellular and intracellular) and functions, including serum proteins, proteins displaying antibacterial properties, and proteins with functions associated with cellular transcription, DNA binding, the cytoskeleton, cell adhesion, and cilia. PolySNAP3 clustering software was used in a multilayered analytical approach. Clusters of proteins that associated with changes to the clinical parameters included neuronal and synapse associated proteins.

Background: Our understanding of the dynamics of genome stability versus gene flux within bacteriophage lineages is limited. Recently, there has been a renewed interest in the use of bacteriophages as 'therapeutic' agents; a prerequisite for their use in such therapies is a thorough understanding of their genetic complement, genome stability and their ecology to avoid the dissemination or mobilisation of phage or bacterial virulence and toxin genes. Campylobacter, a food-borne pathogen, is one of the organisms for which the use of bacteriophage is being considered to reduce human exposure to this organism., Results: Sequencing and genome analysis was performed for two Campylobacter bacteriophages. The genomes were extremely similar at the nucleotide level (> or = 96%) with most differences accounted for by novel insertion sequences, DNA methylases and an approximately 10 kb contiguous region of metabolic genes that were dissimilar at the sequence level but similar in gene function between the two phages. Both bacteriophages contained a large number of radical S-adenosylmethionine (SAM) genes, presumably involved in boosting host metabolism during infection, as well as evidence that many genes had been acquired from a wide range of bacterial species. Further bacteriophages, from the UK Campylobacter typing set, were screened for the presence of bacteriophage structural genes, DNA methylases, mobile genetic elements and regulatory genes identified from the genome sequences. The results indicate that many of these bacteriophages are related, with 10 out of 15 showing some relationship to the sequenced genomes., Conclusions: Two large virulent Campylobacter bacteriophages were found to show very high levels of sequence conservation despite separation in time and place of isolation. The bacteriophages show adaptations to their host and possess genes that may enhance Campylobacter metabolism, potentially advantaging both the bacteriophage and its host. Genetic conservation has been shown to extend to other Campylobacter bacteriophages, forming a highly conserved lineage of bacteriophages that predate upon campylobacters and indicating that highly adapted bacteriophage genomes can be stable over prolonged periods of time.

Background: Elevated left ventricular filling pressure after acute myocardial infarction (AMI) may be identified using clinical assessment, echocardiography, and B-type natriuretic peptide (BNP) levels. All of these predict outcome in this setting. There are, however, no data assessing their relative prognostic value. The current study addresses this., Methods: Four hundred patients underwent detailed echocardiography and measurement of BNP levels after AMI (median 1 day). The study end points were (1) a composite of death, recurrent AMI, and/or admission to hospital with heart failure within 1 year and (2) all-cause mortality during medium-term follow-up (median 2.9 years)., Results: Both an elevated ratio of early transmitral flow to early mitral annulus velocity (E/e') and higher BNP levels were associated with an increased risk of an adverse event within the first year (odds ratio 6.14 for E/e' >15, P < .001; odds ratio 1.19 per 50-pg/mL increase in BNP, P < .001) and medium-term mortality (hazard ratio 4.67 for E/e' >15, P < .001; hazard ratio 1.10 per 50-pg/mL increase in BNP, P < .001). Among patients with BNP levels higher than the median or in the upper quartile, an E/e' ratio >15 identified a subgroup at greatest risk of mortality (P < .001 for both)., Conclusions: The E/e' ratio and BNP levels play important and complementary roles in the risk stratification of patients after AMI., (Copyright 2010 Mosby, Inc. All rights reserved.)

Epidemiologic information, including animal characteristics (e.g., observable risk factors or clinical signs) predisposing to animal disease, is frequently used for design of targeted surveillance systems, but this information is infrequently used for population inference. In this study, we report the evaluation of use of epidemiologic information for population inference in targeted surveillance in three animal disease scenarios. We adapted sampling theory using Monte Carlo methods to determine target population sample size to detect disease with 95% confidence, using information from the epidemiologic parameters risk ratio and fraction of the population with the characteristic. These parameters and their uncertainties were derived from a reference population. The next step was to use a second (sampled) population to evaluate effects of sampling the targeted population. The focus of the study was on estimation of prevalence. Our results showed that if one is less certain of the epidemiologic parameters, a rational decision is to model the input parameter distributions reflecting this uncertainty, thereby increasing the sample size above the minimum needed for the detection of the disease with a known confidence. Targeted surveillance is appropriate for prevalence estimation when one has representative and justifiable estimates of key epidemiologic parameters.