The wild ancestors of domestic animals as a neglected and threatened component of biodiversity.

Domestic animals have immense economic, cultural, and practical value and have played pivotal roles in the development of human civilization. Many domesticates have, among their wild relatives, undomesticated forms representative of their ancestors. Resurgent interest in these ancestral forms has highlighted the unclear genetic status of many, and some are threatened with extinction by hybridization with domestic conspecifics. We considered the contemporary status of these ancestral forms relative to their scientific, practical, and ecological importance; the varied impacts of wild-domestic hybridization; and the challenges and potential resolutions involved in conservation efforts. Identifying and conserving ancestral forms, particularly with respect to disentangling patterns of gene flow from domesticates, is complex because of the lack of available genomic and phenotypic baselines. Comparative behavioral, ecological, and genetic studies of ancestral-type, feral, and domestic animals should be prioritized to establish the contemporary status of the former. Such baseline information will be fundamental in ensuring successful conservation efforts.

Los Ancestros de Animales Domésticos como un Componente Descuidado y Amenazado de la Biodiversidad Resumen Los animales domésticos tienen un inmenso valor económico, cultural y práctico y han jugado un papel muy importante en el desarrollo de la civilización humana. Muchos animales domesticados tienen, entre sus parientes silvestres, formas no domesticadas representativas de sus ancestros. Un interés renovado en estas formas ancestarles ha destacado el estatus genético poco claro de muchas, y algunas están amenazadas de extinción por hibridación con conespecíficos domesticados. Consideramos el estatus contempóraneo de estas formas ancestrales en relación con su importancia científica, práctica y ecológica; los impactos diversos de la hibridación silvestre-domesticado; y los retos y soluciones potenciales involucrados en los esfuerzos de conservación. La identificación y conservación de formas ancestrales, particularmente en relación con desenredar patrones de flujo génicos, es compleja debido a la carencia de líneas de base genómicas y fenotípicas. Se deben priorizar estudios conductuales, ecológicos y genéticos comparativos de los animales ancestrales, ferales y domésticos para establecer el estatus contemporáno de los primeros. Tal información de base será fundamental para asegurar esfuerzos de conservación exitosos.

A phylogenetic approach for weighting genetic sequences.

BACKGROUND: Many important applications in bioinformatics, including sequence alignment and protein family profiling, employ sequence weighting schemes to mitigate the effects of non-independence of homologous sequences and under- or over-representation of certain taxa in a dataset. These schemes aim to assign high weights to sequences that are 'novel' compared to the others in the same dataset, and low weights to sequences that are over-represented. RESULTS: We formalise this principle by rigorously defining the evolutionary 'novelty' of a sequence within an alignment. This results in new sequence weights that we call 'phylogenetic novelty scores'. These scores have various desirable properties, and we showcase their use by considering, as an example application, the inference of character frequencies at an alignment column-important, for example, in protein family profiling. We give computationally efficient algorithms for calculating our scores and, using simulations, show that they are versatile and can improve the accuracy of character frequency estimation compared to existing sequence weighting schemes. CONCLUSIONS: Our phylogenetic novelty scores can be useful when an evolutionarily meaningful system for adjusting for uneven taxon sampling is desired. They have numerous possible applications, including estimation of evolutionary conservation scores and sequence logos, identification of targets in conservation biology, and improving and measuring sequence alignment accuracy.

Modulation of microglial activation states by spinal cord stimulation in an animal model of neuropathic pain: Comparing high rate, low rate, and differential target multiplexed programming.

While numerous studies and patient experiences have demonstrated the efficacy of spinal cord stimulation as a treatment for chronic neuropathic pain, the exact mechanism underlying this therapy is still uncertain. Recent studies highlighting the importance of microglial cells in chronic pain and characterizing microglial activation transcriptomes have created a focus on microglia in pain research. Our group has investigated the modulation of gene expression in neurons and glial cells after spinal cord stimulation (SCS), specifically focusing on transcriptomic changes induced by varying SCS stimulation parameters. Previous work showed that, in rodents subjected to the spared nerve injury (SNI) model of neuropathic pain, a differential target multiplexed programming (DTMP) approach provided significantly better relief of pain-like behavior compared to high rate (HRP) and low rate programming (LRP). While these studies demonstrated the importance of transcriptomic changes in SCS mechanism of action, they did not specifically address the role of SCS in microglial activation. The data presented herein utilizes microglia-specific activation transcriptomes to further understand how an SNI model of chronic pain and subsequent continuous SCS treatment with either DTMP, HRP, or LRP affects microglial activation. Genes for each activation transcriptome were identified within our dataset and gene expression levels were compared with that of healthy animals, naïve to injury and interventional procedures. Pearson correlations indicated that DTMP yields the highest significant correlations to expression levels found in the healthy animals across all microglial activation transcriptomes. In contrast, HRP or LRP yielded weak or very weak correlations for these transcriptomes. This work demonstrates that chronic pain and subsequent SCS treatments can modulate microglial activation transcriptomes, supporting previous research on microglia in chronic pain. Furthermore, this study provides evidence that DTMP is more effective than HRP and LRP at modulating microglial transcriptomes, offering potential insight into the therapeutic efficacy of DTMP.

Spinal cord stimulation using differential target multiplexed programming modulates neural cell-specific transcriptomes in an animal model of neuropathic pain.

Spinal cord stimulation is a proven effective therapy for treating chronic neuropathic pain. Previous work in our laboratory demonstrated that spinal cord stimulation based on a differential target multiplexed programming approach provided significant relief of pain-like behavior in rodents subjected to the spared nerve injury model of neuropathic pain. The relief was significantly better than obtained using high rate and low rate programming. Furthermore, transcriptomics-based results implied that differential target multiplexed programming modulates neuronal-glial interactions that have been perturbed by the pain process. Although differential target multiplexed programming was developed to differentially target neurons and glial cells, our previous work did not address this. This work presents transcriptomes, specific to each of the main neural cell populations (neurons, microglia, astrocytes, and oligodendrocytes), obtained from spinal cord subjected to continuous spinal cord stimulation treatment with differential target multiplexed programming, high rate programming, or low rate programming compared with no spinal cord stimulation treatment, using the spared nerve injury model. To assess the effect of each spinal cord stimulation treatment on these cell-specific transcriptomes, gene expression levels were compared with that of healthy animals, naïve to injury and interventional procedures. Pearson correlations and cell population analysis indicate that differential target multiplexed programming yielded strong and significant correlations to expression levels found in the healthy animals across every evaluated cell-specific transcriptome. In contrast, high rate programming only yielded a strong correlation for the microglia-specific transcriptome, while low rate programming did not yield strong correlations with any cell types. This work provides evidence that differential target multiplexed programming distinctively targeted and modulated the expression of cell-specific genes in the direction of the healthy state thus supporting its previously established action on regulating neuronal-glial interaction processes in a pain model.

Clinical Effectiveness and Mechanism of Action of Spinal Cord Stimulation for Treating Chronic Low Back and Lower Extremity Pain: a Systematic Review.

PURPOSE OF REVIEW: The purpose of the present systematic review is to provide a current understanding of the mechanism of action and the evidence available to support clinical decision-making. The focus is to summarize randomized controlled trials (RCTs) and nonrandomized or observational studies of spinal cord stimulation in chronic pain to understand clinical effectiveness and the mechanism of action. RECENT FINDINGS: Several recent studies have demonstrated the benefit of spinal cord stimulation in managing chronic pain. Until recently, the mechanism of action was founded on a central paradigm derived from gate control theory, which is the need to stimulate the dorsal column of the spinal cord to generate paresthesia. The recent development of new therapies that do not rely on paresthesia has left the field without a clear mechanism of action that could serve as a strong foundation to further improve clinical outcomes. Consequently, multiple theories have emerged to explain how electrical pulse applied to the spinal cord could alleviate pain, including activation of specific supraspinal pathways, and segmental modulation of the neurological interaction. Recent systematic reviews also have shown the clinical effectiveness of spinal cord stimulation in managing chronic spinal pain, phantom limb pain, complex regional pain syndrome, and other chronic painful conditions. Spinal cord stimulation for the treatment of chronic pain is rapidly evolving with technology at its forefront. This comprehensive focused review evaluated 11 RCTs and 7 nonrandomized/observational studies which provided levels of evidence ranging from I to II.

Effects of Phase Polarity and Charge Balance Spinal Cord Stimulation on Behavior and Gene Expression in a Rat Model of Neuropathic Pain.

OBJECTIVE: To investigate the effect of phase polarity and charge balance of spinal cord stimulation (SCS) waveforms on pain behavior and gene expression in a neuropathic pain rodent model. We hypothesized that differing waveforms will result in diverse behavioral and transcriptomics expression due to unique mechanisms of action. MATERIALS AND METHODS: Rats were implanted with a four-contact cylindrical mini-lead and randomly assigned to two control (no-pain and pain model) and five test groups featuring monophasic, as well as charge-unbalanced and charge-balanced biphasic SCS waveforms. Mechanical and cold allodynia were assessed to measure efficacy. The ipsilateral dorsal quadrant of spinal cord adjacent to the lead was harvested post-stimulation and processed to determine gene expression via real-time reverse-transcriptase polymerase chain reaction (RT-PCR). Gene expression, SCS intensity (mA), and behavioral score as percent of baseline (BSPB) were statistically analyzed and used to generate correlograms using R-Studio. Statistical analysis was performed using SPSS22.0, and p < 0.05 was considered significant. RESULTS: As expected, BSPB was significantly lower for the pain model group compared to the no-pain group. BSPB was significantly improved post-stim compared to pre-stim using cathodic, anodic, symmetric biphasic, or asymmetric biphasic 1:2 waveforms; however, BSPB was not restored to Sham levels. RT-PCR analysis showed that eight genes demonstrated a significant difference between the pain model and SCS waveforms and between waveforms. Correlograms reveal a linear correlation between regulation of expression of a given gene in relation to mA, BSPB, or other genes. CONCLUSIONS: Our results exhibit that specific SCS waveforms differentially modulate several key transcriptional pathways that are relevant in chronic pain conditions. These results have significant implications for SCS: whether to move beyond traditional paradigm of neuronal activation to focus also on modulating immune-driven processes.

Development and validation of a high-speed video system for measuring saccadic eye movement.

Laboratory-based retroreflective and magnetic scleral search-coil technologies are the current standards for collecting saccadometric data, but such equipment is costly and cumbersome. We have validated a novel, portable, high-speed video camera-based system (Exilim EX-FH20, Casio, Tokyo, Japan) for measuring saccade reaction time (RT) and error rate in a well-lit environment. This system would enable measurements of pro- and antisaccades in athletes, which is important because antisaccade metrics provide a valid tool for concussion diagnosis and determining an athlete's safe return to play. A total of 529 trials collected from 15 participants were used to compare saccade RT and error rate measurements of the high-speed camera system to a retroreflective video-based eye tracker (Eye-Trac 6: Applied Sciences Laboratories, Bedford, MA). Bland-Altman analysis revealed that the RT measurements made by the high-speed video system were 11 ms slower than those made by the retroreflective system. Error rate measurements were identical between the two systems. An excellent degree of reliability was found between the system measurements and in the ratings of independent researchers examining the video data. A strong association (r = .97) between the RTs determined via the retroreflective and high-speed camera systems was observed across all trials. Our high-speed camera system is portable and easily set up, does not require extensive equipment calibration, and can be used in a well-lit environment. Accordingly, the camera-based capture of saccadometric data may provide a valuable tool for neurological assessment following a concussive event and for the continued monitoring of recovery.

Endovascular Applications for the Management of High-Grade Gliomas in the Modern Era.

High-grade gliomas (HGGs) have a poor prognosis and are difficult to treat. This review examines the evolving landscape of endovascular therapies for HGGs. Recent advances in endovascular catheter technology and delivery methods allow for super-selective intra-arterial cerebral infusion (SSIACI) with increasing precision. This treatment modality may offer the ability to deliver anti-tumoral therapies directly to tumor regions while minimizing systemic toxicity. However, challenges persist, including blood-brain barrier (BBB) penetration, hemodynamic complexities, and drug-tumor residence time. Innovative adjunct techniques, such as focused ultrasound (FUS) and hyperosmotic disruption, may facilitate BBB disruption and enhance drug penetration. However, hemodynamic factors that limit drug residence time remain a limitation. Expanding therapeutic options beyond chemotherapy, including radiotherapy and immunobiologics, may motivate future investigations. While preclinical and clinical studies demonstrate moderate efficacy, larger randomized trials are needed to validate the clinical benefits. Additionally, future directions may involve endovascular sampling for peri-tumoral surveillance; changes in drug formulations to prolong residence time; and the exploration of non-pharmaceutical therapies, like radioembolization and photodynamic therapy. Endovascular strategies hold immense potential in reshaping HGG treatment paradigms, offering targeted and minimally invasive approaches. However, overcoming technical challenges and validating clinical efficacy remain paramount for translating these advancements into clinical care.

Radical surgical resection with molecular margins is associated with improved survival in IDH wildtype GBM.

BACKGROUND: Survival is variable in patients with glioblastoma IDH wild-type (GBM), even after comparable surgical resection of radiographically-detectable disease, highlighting the limitations of radiographic assessment of infiltrative tumor anatomy. The majority of post-surgical progressive events are failures within 2cm of the resection margin, motivating supramaximal resection strategies to improve local control. However, which patients benefit from such radical resections remains unknown. METHODS: We developed a predictive model to identify which IDH wild-type GBM are amenable to radiographic gross total resection (GTR). We then investigated whether GBM survival heterogeneity following GTR is correlated with microscopic tumor burden a by analyzing tumor cell content at the surgical margin with a rapid qPCR-based method for detection of TERT promoter mutation. RESULTS: Our predictive model for achievable GTR, developed on retrospective radiographic and molecular data of GBM patients undergoing resection, had an AUC of 0.83, sensitivity of 62%, and specificity of 90%. Prospective analysis of this model in 44 patients found 89% of patients were correctly predicted to achieve a RV<4.9cc. Of the 44 prospective patients undergoing rapid qPCR TERT promoter mutation analysis at the surgical margin, 7 had undetectable TERT mutation, of which 5 also had a gross total resection (RV<1cc). In these 5 patients at 30 months follow up, 75% showed no progression, compared to 0% in the group with TERT mutations detected at the surgical margin (p=0.02). CONCLUSIONS: These findings identify a subset of patients with GBM that may derive local control benefit from radical resection to undetectable molecular margins.

Assembly and architecture of biogenesis of lysosome-related organelles complex-1 (BLOC-1).

BLOC-1 (biogenesis of lysosome-related organelles complex-1) is critical for melanosome biogenesis and has also been implicated in neurological function and disease. We show that BLOC-1 is an elongated complex that contains one copy each of the eight subunits pallidin, Cappuccino, dysbindin, Snapin, Muted, BLOS1, BLOS2, and BLOS3. The complex appears as a linear chain of eight globular domains, ∼300 Å long and ∼30 Å in diameter. The individual domains are flexibly connected such that the linear chain undergoes bending by as much as 45°. Two stable subcomplexes were defined, pallidin-Cappuccino-BLOS1 and dysbindin-Snapin-BLOS2. Both subcomplexes are 1:1:1 heterotrimers that form extended structures as indicated by their hydrodynamic properties. The two subcomplexes appear to constitute flexible units within the larger BLOC-1 chain, an arrangement conducive to simultaneous interactions with multiple BLOC-1 partners in the course of tubular endosome biogenesis and sorting.

Postexposure application of Fas receptor small-interfering RNA to suppress sulfur mustard-induced apoptosis in human airway epithelial cells: implication for a therapeutic approach.

Sulfur mustard (SM) is a vesicant chemical warfare and terrorism agent. Besides skin and eye injury, respiratory damage has been mainly responsible for morbidity and mortality after SM exposure. Previously, it was shown that suppressing the death receptor (DR) response by the dominant-negative Fas-associated death domain protein prior to SM exposure blocked apoptosis and microvesication in skin. Here, we studied whether antagonizing the Fas receptor (FasR) pathway by small-interfering RNA (siRNA) applied after SM exposure would prevent apoptosis and, thus, airway injury. Normal human bronchial/tracheal epithelial (NHBE) cells were used as an in vitro model with FasR siRNA, FasR agonistic antibody CH11, and FasR antagonistic antibody ZB4 as investigative tools. In NHBE cells, both SM (300 µM) and CH11 (100 ng/ml) induced caspase-3 activation, which was inhibited by FasR siRNA and ZB4, indicating that SM-induced apoptosis was via the Fas response. FasR siRNA inhibited SM-induced caspase-3 activation when added to NHBE cultures up to 8 hours after SM. Results using annexin V/propidium iodide-stained cells showed that both apoptosis and necrosis were involved in cell death due to SM; FasR siRNA decreased both apoptotic and necrotic cell populations. Bronchoalveolar lavage fluid (BALF) of rats exposed to SM (1 mg/kg, 50 minutes) revealed a significant (P < 0.05) increase in soluble Fas ligand and active caspase-3 in BALF cells. These findings suggest an intervention of Fas-mediated apoptosis as a postexposure therapeutic strategy with a therapeutic window for SM inhalation injury and possibly other respiratory diseases involving the Fas response.

Parasite exchange and hybridisation at a wild-feral-domestic interface.

Interactions between wild, feral, and domestic animals are of economic and conservation significance. The pigeon Columba livia is a synanthropic species in a feral form, but it also includes the rare Rock Dove. Columba livia is an important player at the wild-domestic interface, acting as a carrier of avian diseases, and the feral form threatens Rock Doves with extinction via hybridisation. Despite its abundance, little is known about drivers of disease prevalence in C. livia, or how disease and hybridisation represent synergistic threats to Rock Doves. We focused on infection by the parasite Trichomonas, first collating prevalence estimates in domestic and free-living populations from relevant studies of C. livia. Second, we characterised variation in the diversity and prevalence of Trichomonas among three C. livia populations in the United Kingdom: a feral, a Rock Dove, and a feral-wild hybrid population. Across multiple continents, free-living pigeons had lower Trichomonas infection than captive conspecifics, but the effect was weak. Environmental factors which could impact Trichomonas infection status did not explain variation in infection among populations. Among the British populations, strain diversity varied, and there was lower parasite prevalence in Rock Doves than feral pigeons. Individual infection status was not explained by the available covariates, including hybrid score and site. The drivers of Trichomonas prevalence are unclear, perhaps due to idiosyncratic local-scale drivers. However, given the population-level variation in both infection prevalence and introgressive hybridisation, the potential combined effects could accelerate the extinction of the Rock Dove. Further study of the synergistic effects of multiple types of biotic interactions at the wild-feral-domestic interface is warranted, especially where vagile, globally distributed and superabundant animals are involved.

Impact of aluminum adjuvants on the stability of pneumococcal polysaccharide-protein conjugate vaccines.

Development of a vaccine drug product requires formulation optimization to ensure that the vaccine's effectiveness is preserved upon storage throughout the shelf-life of the product. Although aluminum adjuvants have been widely used in vaccine formulations to safely and effectively potentiate an immune response, careful attention must be directed towards ensuring that the type of aluminum adjuvant does not impact the stability of the antigenic composition. PCV15 is a polysaccharide-protein conjugate vaccine comprising the pneumococcal polysaccharide (PnPs) serotypes (1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 18C, 19A, 19F, 22F, 23F and 33F), each individually conjugated to the protein carrier CRM197. PCV15 was formulated with either amorphous aluminum hydroxyphosphate sulfate adjuvant (AAHS) or aluminum phosphate adjuvant (AP) and examined for both stability and immunogenicity. Using a collection of methods to evaluate vaccine stability, it was discovered that certain PCV15 serotypes (e.g., 6A, 19A, 19F) formulated with AAHS resulted in a reduction of immunogenicity in vivo and a reduction in recoverable dose as tested by an in vitro potency assay. The same polysaccharide-protein conjugates formulated with AP were stable regarding all measures tested. Moreover, the reduction in potency of certain serotypes correlated with chemical degradation of the polysaccharide antigen caused by the aluminum adjuvant as measured by reducing polyacrylamide gel electrophoresis (SDS-PAGE), High-Pressure Size Exclusion Chromatography coupled with UV detection (HPSEC-UV) and ELISA immunoassay. This study suggests a formulation, which includes AAHS, may negatively impact the stability of a pneumococcal polysaccharide-protein conjugate vaccine that contains phosphodiester groups. This decrease in stability would likely result in a decrease in the "active" concentration of antigen dose, and herein, it is shown that such instability directly compromised vaccine immunogenicity in an animal model. The results presented in this study help to explain critical degradation mechanisms of pneumococcal polysaccharide-protein conjugate vaccines.

Preclinical evaluation of an investigational 21-valent pneumococcal conjugate vaccine, V116, in adult-rhesus monkey, rabbit, and mouse models.

Despite the widespread effectiveness of pneumococcal conjugate vaccines on the overall incidence of invasive pneumococcal disease, the global epidemiological landscape continues to be transformed by residual disease from non-vaccine serotypes, thus highlighting the need for vaccines with expanded disease coverage. To address these needs, we have developed V116,an investigational 21-valent non-adjuvanted pneumococcal conjugate vaccine (PCV),containingpneumococcal polysaccharides (PnPs) 3, 6A, 7F, 8, 9N, 10A, 11A,12F, 15A, 16F, 17F, 19A, 20, 22F, 23A, 23B, 24F, 31, 33F, 35B, anda de-O-acetylated 15B(deOAc15B) individually conjugated to the nontoxic diphtheria toxoid CRM197 carrier protein. Preclinical studies evaluated the immunogenicity of V116 inadult monkeys, rabbits, and mice. Following one dose, V116 was found to be immunogenic in preclinical animal species and induced functional antibodies for all serotypes included in the vaccine, in addition to cross-reactive functional antibodies to serotypes 6C and 15B. In these preclinical animal studies, the increased valency of V116 did not result in serotype-specific antibody suppression when compared to lower valent vaccines V114 or PCV13. In addition, when compared with naïve controls, splenocytes from V116 to immunized animals demonstrated significant induction of CRM197-specific T cells in both IFN-γ and IL-4 ELISPOT assays, as well as Th1 and Th2 cytokine induction through in vitro stimulation assays, thus suggesting the ability of V116 to engage T cell dependent immune response pathways to aid in development of memory B cells. V116 also demonstrated significant protection in mice from intratracheal challenge with serotype 24F, a novel serotype not contained in any currently licensed vaccine.

Variables associated with cortical motor mapping thresholds: A retrospective data review with a unique case of interlimb motor facilitation.

Introduction: Intraoperative neuromonitoring (IONM) is crucial to preserve eloquent neurological functions during brain tumor resections. We observed a rare interlimb cortical motor facilitation phenomenon in a patient with recurrent high-grade glioma undergoing craniotomy for tumor resection; the patient's upper arm motor evoked potentials (MEPs) increased in amplitude significantly (up to 44.52 times larger, p < 0.001) following stimulation of the ipsilateral posterior tibial nerve at 2.79 Hz. With the facilitation effect, the cortical MEP stimulation threshold was reduced by 6 mA to maintain appropriate continuous motor monitoring. It likely has the benefit of reducing the occurrence of stimulation-induced seizures and other adverse events associated with excessive stimulation. Methods: We conducted a retrospective data review including 120 patients who underwent brain tumor resection with IONM at our center from 2018 to 2022. A broad range of variables collected pre-and intraoperatively were reviewed. The review aimed to determine: (1) whether we overlooked this facilitation phenomenon in the past, (2) whether this unique finding is related to any specific demographic information, clinical presentation, stimulation parameter (s) or anesthesia management, and (3) whether it is necessary to develop new techniques (such as facilitation methods) to reduce cortical stimulation intensity during intraoperative functional mapping. Results: There is no evidence suggesting that clinical presentation, stimulation configuration, or intraoperative anesthesia management of the patient with the facilitation effect were significantly different from our general patient cohort. Even though we did not identify the same facilitation effect in any of these patients, we were able to determine that stimulation thresholds for motor mapping are significantly associated with the location of stimulation (p = 0.003) and the burst suppression ratio (BSR) (p < 0.001). Stimulation-induced seizures, although infrequent (4.05%), could occur unexpectedly even when the BSR was 70%. Discussion: We postulated that functional reorganization and neuronal hyperexcitability induced by glioma progression and repeated surgeries were probable underlying mechanisms of the interlimb facilitation phenomenon. Our retrospective review also provided a practical guide to cortical motor mapping in brain tumor patients under general anesthesia. We also underscored the need for developing new techniques to reduce the stimulation intensity and, hence, seizure occurrence.

Analytical technology development to monitor the stability of Polysaccharide-Protein conjugate vaccines.

The covalent attachment of a bacterial-derived capsular polysaccharide to protein is of critical importance in transforming the polysaccharide from an antigen with limited immunogenicity in infants and older adults to an antigen that can prevent potentially fatal disease. For a polysaccharide-protein conjugate vaccine (PCV) candidate to be successful, it must be sufficiently stable. Chemical breakage of carbohydrate bonds in the polysaccharide may result in the reduction of "conjugate dose" and could negatively impact immunogenicity and the ability of the vaccine to prime for memory responses. Therefore, development of analytical tools to monitor the integrity of a polysaccharide-protein conjugate (glycoconjugate) vaccine is of practical significance. In this work, reducing SDS-PAGE, Intrinsic Protein Fluorescence Spectroscopy (IPFS), Differential Scanning Fluorimetry (DSF) were evaluated methods to study the impact of time, temperature, and formulation composition on the stability of a glycoconjugate vaccine prepared by multisite coupling of polysaccharide to a carrier protein. In addition, an automated capillary Western system was also evaluated to study the impact of storage on glycoconjugate vaccine stability. Two streptococcus pneumoniae polysaccharide-protein conjugates (serotype 3 and serotype 19A) were chosen to examine their physicochemical stability when formulated as a single antigen vaccine. While all methods require only a small amount of test article and can test multiple samples per assay run, automated capillary Western has the additional advantage of being highly sensitive even at low concentrations in complex vaccine formulations that contain aluminum adjuvant and multiple antigens. Results suggest that automated capillary Western is stability-indicating and may be an effective analytical technology tool for the formulation development of a multivalent glycoconjugate vaccine.

Limited domestic introgression in a final refuge of the wild pigeon.

Domesticated animals have been culturally and economically important throughout history. Many of their ancestral lineages are extinct or genetically endangered following hybridization with domesticated relatives. Consequently, they have been understudied compared to the ancestral lineages of domestic plants. The domestic pigeon Columba livia, which was pivotal in Darwin's studies, has maintained outsized cultural significance. Its role as a model organism spans the fields of behavior, genetics, and evolution. Domestic pigeons have hybridized with their progenitor, the Rock Dove, rendering the latter of dubious genetic status. Here, we use genomic and morphological data from the putative Rock Doves of the British Isles to identify relictual undomesticated populations. We reveal that Outer Hebridean Rock Doves have experienced minimal levels of introgression. Our results outline the contemporary status of these wild pigeons, highlighting the role of hybridization in the homogenization of genetic lineages.

Structural basis for ubiquitin recognition and autoubiquitination by Rabex-5.

Rabex-5 is an exchange factor for Rab5, a master regulator of endosomal trafficking. Rabex-5 binds monoubiquitin, undergoes covalent ubiquitination and contains an intrinsic ubiquitin ligase activity, all of which require an N-terminal A20 zinc finger followed immediately by a helix. The structure of the N-terminal portion of Rabex-5 bound to ubiquitin at 2.5-A resolution shows that Rabex-5-ubiquitin interactions occur at two sites. The first site is a new type of ubiquitin-binding domain, an inverted ubiquitin-interacting motif, which binds with approximately 29-microM affinity to the canonical Ile44 hydrophobic patch on ubiquitin. The second is a diaromatic patch on the A20 zinc finger, which binds with approximately 22-microM affinity to a polar region centered on Asp58 of ubiquitin. The A20 zinc-finger diaromatic patch mediates ubiquitin-ligase activity by directly recruiting a ubiquitin-loaded ubiquitin-conjugating enzyme.

Immunogenicity of PCV24, an expanded pneumococcal conjugate vaccine, in adult monkeys and protection in mice.

Invasive pneumococcal disease (IPD) is responsible for serious illnesses such as bacteremia, sepsis, meningitis, and pneumonia in young children, older adults, and persons with immunocompromising conditions and often leads to death. Although the most recent pneumococcal conjugate vaccines (PCVs) have been designed to target serotypes identified as the primary causative agents of IPD, the epidemiological landscape continues to change stressing the need to develop new PCVs. We have developed an investigational 24-valent PCV (PCV24) including serotypes 1, 2, 3, 4, 5, 6A, 6B, 7F, 8, 9N, 9V, 10A, 11A, 12F, 14, 15B, 17F, 18C, 19A, 19F, 20, 22F, 23F, and 33F all conjugated to CRM197 and evaluated this vaccine in adult monkeys. PCV24 was shown to be immunogenic and induced functional antibody for all vaccine serotypes. Of the serotypes common to PCV13 and V114 (PCV15), PCV24 had a similar immunogenic response with the exceptions of 23F which had higher IgG GMCs for PCV13 and V114, and 7F which had higher GMCs for PCV13. Functional antibody responses were similar for the serotypes in common between PCV24, PCV13 and V114 vaccines, with the exception of serotype 7F which was greater for PCV13. Overall, this study shows that PCV24 provided similar immunogenicity as the lower valent vaccines in adult monkeys with no apparent serotype interference. In addition, PCV24 also provided protection against pneumococcal infection in a mouse challenge model.