Development of streptococcal pyrogenic exotoxin C vaccine toxoids that are protective in the rabbit model of toxic shock syndrome

Streptococcal pyrogenic exotoxin C (SPE C) is a superantigen produced by many strains of Streptococcus pyogenes that (along with streptococcal pyrogenic exotoxin A) is highly associated with streptococcal toxic shock syndrome (STSS) and other invasive streptococcal diseases. Based on the three-dimensional structure of SPE C, solvent-exposed residues predicted to be important for binding to the TCR or the MHC class II molecule, or important for dimerization, were generated. Based on decreased mitogenic activity of various single-site mutants, the double-site mutant Y15A/N38D and the triple-site mutant Y15A/H35A/N38D were constructed and analyzed for superantigenicity, toxicity (lethality), immunogenicity, and the ability to protect against wild-type SPE C-induced STSS. The Y15A/N38D and Y15A/H35A/N38D mutants were nonmitogenic for rabbit splenocytes and human PBMCs and nonlethal in two rabbit models of STSS, yet both mutants were highly immunogenic. Animals vaccinated with the Y15A/N38D or Y15A/H35A/N38D toxoids were protected from challenge with wild-type SPE C. Collectively, these data indicate that the Y15A/N38D and Y15A/H35A/N38D mutants may be useful as toxoid vaccine candidates.     

TH(1)- and TH(2)-TYPE cytokine expression by activated t lymphocytes from the lung and spleen during the inflammatory response to respiratory syncytial virus

RSV is an important cause of lower respiratory tract illness in infants and the elderly worldwide. The components involved in immunity and those that contribute to inflammation of RSV-induced disease are not clearly understood. To address the relationship between activation antigen and cytokine expression, intracellular levels of IL-2, IL-4, IL-5 and IFN-gamma were determined for CD3, CD44, CD49d, CD54, CD62L and CD102 lymphocytes from the bronchoalveolar lavage and spleen. To examine activation at the DNA level, lymphocytes expressing IL-2, IL-4, IL-5 or IFN-gamma were analysed for G2+M DNA content or phosphatidylserine expression (apoptosis). Trafficking of lymphocytes to the BAL was detected at day 5 p.i., peaked day 7 p.i., and predominately involved CD54(+)and CD102(+)lymphocytes expressing high levels of IL-2, IL-4, IL-5 and IFN-gamma. Lymphocytes expressing CD44(+), CD49d(+)and CD62L(lo)were also observed, however they expressed these cytokines to a lesser extent. DNA analysis of lymphocytes expressing IL-2 or IFN-gamma revealed higher G2'M levels compared to lymphocytes expressing IL-4 or IL-5, suggesting greater activation of Th(1)-type lymphocytes in the lung. These data demonstrate that RSV-induced pulmonary inflammation involves extensive cellular activation and cytokine expression, particularly by CD54(+)and CD102(+)lymphocytes in the lung.     

Relationship between EGFR overexpression and gene amplification status in central nervous system gliomas

OBJECTIVE: To correlate epidermal growth factor receptor (EGFR) protein overexpression, as assessed by immunohistochemistry, with EGFR gene amplification determined by fluorescence in situ hybridization in a series of gliomas. STUDY DESIGN: Forty-seven central nervous system gliomas, including 34 cases of glioblastoma multiforme, 3 oligodendrogliomas, 4 juvenile pilocytic astrocytomas and 5 low grade astrocytomas, were obtained from the files of the University of Utah Pathology Department. In each case a representative paraffin block was selected, and EGFR protein expression was quantified using immunohistochemistry. EGFR gene amplification status was determined by fluorescence in situ hybridization. RESULTS: EGFR protein overexpression was detected in 9 cases of glioblastoma multiforme. EGFR gene amplification was present in 7 of these cases. Both nonamplified glioblastomas demonstrated only 2+ overexpression of EGFR protein. None of the low grade, pilocytic or anaplastic astrocytomas demonstrated either EGFR protein overexpression or gene amplification. CONCLUSION: EGFR protein overexpression is closely associated with gene amplification. Seventy-eight percent of cases showing protein overexpression demonstrated gene amplification. All cases of central nervous system neoplasms showing protein overexpression but lacking gene amplification were associated with only 2+ protein overexpression. All central nervous system neoplasms demonstrating gene amplification and/or overexpression were high grade neoplasms.     

Sylvatic Plague Vaccine Partially Protects Prairie Dogs (<Emphasis Type="Italic">Cynomys</Emphasis> spp.) in Field Trials

Sylvatic plague, caused by Yersinia pestis, frequently afflicts prairie dogs (Cynomys spp.), causing population declines and local extirpations. We tested the effectiveness of bait-delivered sylvatic plague vaccine (SPV) in prairie dog colonies on 29 paired placebo and treatment plots (1–59 ha in size; average 16.9 ha) in 7 western states from 2013 to 2015. We compared relative abundance (using catch per unit effort (CPUE) as an index) and apparent survival of prairie dogs on 26 of the 29 paired plots, 12 with confirmed or suspected plague (Y. pestis positive carcasses or fleas). Even though plague mortality occurred in prairie dogs on vaccine plots, SPV treatment had an overall positive effect on CPUE in all three years, regardless of plague status. Odds of capturing a unique animal were 1.10 (95% confidence interval [C.I.] 1.02–1.19) times higher per trap day on vaccine-treated plots than placebo plots in 2013, 1.47 (95% C.I. 1.41–1.52) times higher in 2014 and 1.19 (95% C.I. 1.13–1.25) times higher in 2015. On pairs where plague occurred, odds of apparent survival were 1.76 (95% Bayesian credible interval [B.C.I.] 1.28–2.43) times higher on vaccine plots than placebo plots for adults and 2.41 (95% B.C.I. 1.72–3.38) times higher for juveniles. Our results provide evidence that consumption of vaccine-laden baits can protect prairie dogs against plague; however, further evaluation and refinement are needed to optimize SPV use as a management tool.     

Exercise intervention for unilateral amputees with low back pain: study protocol for a randomised,controlled trial

Background

Atraumatic lower limb amputation is a life-changing event for approximately 185,000 persons in the United States each year. A unilateral amputation is associated with rapid changes to the musculoskeletal system including leg and back muscle atrophy, strength loss, gait asymmetries, differential mechanical joint loading and leg length discrepancies. Even with high-quality medical care and prostheses, amputees still develop secondary musculoskeletal conditions such as chronic low back pain (LBP). Resistance training interventions that focus on core stabilization, lumbar strength and dynamic stability during loading have strong potential to reduce LBP and address amputation-related changes to the musculoskeletal system. Home-based resistance exercise programs may be attractive to patients to minimize travel and financial burdens.

Methods/design

This study will be a single-assessor-blinded, pre-post-test randomised controlled trial involving 40 men and women aged 18–60 years with traumatic, unilateral transtibial amputation. Participants will be randomised to a home-based, resistance exercise group (HBRX) or a wait-list control group (CON). The HBRX will consist of 12 weeks of elastic resistance band and bodyweight training to improve core and lumbopelvic strength. Participants will be monitored via Skype or Facetime on a weekly basis. The primary outcome will be pain severity (11-point Numerical Pain Rating Scale; NRS_pain). Secondary outcomes will include pain impact on quality of life (Medical Outcomes Short Form 36, Oswestry Disability Index and Roland Morris Disability Questionnaire), kinematics and kinetics of walking gait on an instrumented treadmill, muscle morphology (muscle thickness of multifidus, transversus abdominis, internal oblique), maximal muscle strength of key lumbar and core muscles, and daily step count.

Discussion

The study findings will determine whether a HBRX program can decrease pain severity and positively impact several physiological and mechanical factors that contribute to back pain in unilateral transtibial amputees with chronic LBP. We will determine the relative contribution of the exercise-induced changes in these factors on pain responsiveness in this population.

Trial registration

ClinicalTrials.gov, ID: NCT03300375. Registered on 2 October 2017.

     

Factors Influencing Uptake of Sylvatic Plague Vaccine Baits by Prairie Dogs

Sylvatic plague vaccine (SPV) is a virally vectored bait-delivered vaccine expressing Yersinia pestis antigens that can protect prairie dogs (Cynomys spp.) from plague and has potential utility as a management tool. In a large-scale 3-year field trial, SPV-laden baits containing the biomarker rhodamine B (used to determine bait consumption) were distributed annually at a rate of approximately 100–125 baits/hectare along transects at 58 plots encompassing the geographic ranges of four species of prairie dogs. We assessed site- and individual-level factors related to bait uptake in prairie dogs to determine which were associated with bait uptake rates. Overall bait uptake for 7820 prairie dogs sampled was 70% (95% C.I. 69.9–72.0). Factors influencing bait uptake rates by prairie dogs varied by species, however, in general, heavier animals had greater bait uptake rates. Vegetation quality and day of baiting influenced this relationship for black-tailed, Gunnison’s, and Utah prairie dogs. For these species, baiting later in the season, when normalized difference vegetation indices (a measure of green vegetation density) are lower, improves bait uptake by smaller animals. Consideration of these factors can aid in the development of species-specific SPV baiting strategies that maximize bait uptake and subsequent immunization of prairie dogs against plague.     

Enhancing the stability and folding rate of a repeat protein through the addition of consensus repeats

Repeat proteins are constructed from a linear array of modular units, giving rise to an overall topology lacking long-range interactions. This suggests that stabilizing repeat modules based on consensus information might be added to a repeat protein domain, allowing it to be extended without altering its overall topology. Here we add consensus modules the ankyrin repeat domain from the Drosophila Notch receptor to investigate the structural tolerance to these modules, the relative thermodynamic stability of these hybrid proteins, and how alterations in the energy landscape influence folding kinetics. Insertions of consensus modules between repeats five and six of the Notch ankyrin domain have little effect on the far and near-UV CD spectra, indicating that neither secondary nor tertiary structure is dramatically altered. Furthermore, stable structure is maintained at increased denaturant concentrations in the polypeptides containing the consensus repeats, indicating that the consensus modules are capable of stabilizing much of the domain. However, insertion of the consensus repeats appears to disrupt cooperativity, producing a two-stage (three-state) unfolding transition in which the C-terminal repeats unfold at moderate urea concentrations. Removing the C-terminal repeats (Notch ankyrin repeats six and seven) restores equilibrium two-state folding and demonstrates that the high stability of the consensus repeats is propagated into the N-terminal, naturally occurring Notch ankyrin repeats. This stability increase greatly increases the folding rate, and suggests that the transition state ensemble may be repositioned in the chimeric consensus-stabilized proteins in response to local stability.     

A synopsis of Costa Rican Ruellia (Acanthaceae), with descriptions of four new species

We describe four new species of Ruellia from Costa Rica; three from the Osa Peninsula, Puntarenas province (R. exilis, R. mira, R. odorata), and one from the southwestern corner of San José province (R. norvegigratiosa). Ruellia matagalpae is reported as a new record for the country. New combinations are made in Ruellia for the two Costa Rican species of Blechum (R. blechum and R. costaricensis). Three species are reduced to synonymy, and another species is considered doubtfully worthy of recognition. Notable range extensions are reported for two species. Corollas and fruits of all 22 Costa Rican species of Ruellia are shown and compared as easily used identification aids. An informal guide to species groups and a dichotomous key to the 22 species are presented.