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Burbot (Lota lota) are an ideal candidate for cool or cold-water aquaculture and are gaining interest because of their high economic value, low temperature requirements, and fast growth rate. Limited information exists on the innate and adaptive immune systems of this species. This is partly due to the lack of species-specific tools to determine antibody responses following disease or vaccination or to characterize the immune response in general. An anti-IgM monoclonal antibody (mAb 27C) was developed and characterized via enzyme-linked immunosorbent assay (ELISA) and Western blot for species specificity, affinity to the heavy chain of burbot IgM, and cross-reactivity to other reagents used in the analysis. The 27C monoclonal antibody was further utilized to develop an ELISA protocol to measure the specific antibody response of burbot following exposure to two pathogenic strains of Aeromonas sp. (A141 and IR004). This ELISA confirmed that vaccinated burbot that survived the challenge with either strain developed statistically higher titers of anti-Aeromonas antibodies specific for the relative strain when compared to fish that were not vaccinated or challenged. Western blot analysis further demonstrated that burbot surviving challenge had serum IgM that recognized distinct antigens specific to the strain they were challenged with, A141 bound to antigens in the 50-250Kda range and IR004 bound to a distinct 150Kda antigen. Western blots further indicated that each strain shared antigenic regions regardless of experimental Aeromonas strain exposure. Finally, immunofluorescent staining confirmed that mAb 27C binds to membrane-bound IgM (presumably B cells) on burbot head kidney cells. Taken together, results from this study demonstrate that mAb 27C specifically recognized burbot IgM and will be an important tool to further characterize the adaptive and cellular immune responses of this fish species.
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Aeromonas , Gadiformes , Animales , Anticuerpos Monoclonales , Peces , Ensayo de Inmunoadsorción Enzimática/veterinariaRESUMEN
Infectious hematopoietic necrosis (IHN) is a significant viral disease affecting salmonids, whereas Flavobacterium psychrophilum (Fp), the causative agent of bacterial coldwater disease (BCWD), remains one of the most significant bacterial pathogens of salmonids. We explored maternal immunity in the context of IHN and BCWD management in rainbow trout (Oncorhynchus mykiss) aquaculture. Two experimental trials were conducted where different groups of female broodstock were immunized prior to spawning with an IHNV DNA vaccine or a live attenuated F. psychrophilum (Fp B.17-ILM) vaccine alone, or in combination. Progeny were challenged with either a low or high dose of IHNV at 13 days post hatch (dph) and 32 dph or challenged with F. psychrophilum at 13 dph. Mortality following a low-dose IHNV challenge at 13 dph was significantly lower in progeny from vaccinated broodstock vs. unvaccinated broodstock, but no significant differences were observed at 32 dph. Mortality due to BCWD was also significantly reduced in 13 dph fry that originated from broodstock immunized with the Fp B.17-ILM vaccine. After vaccination broodstock developed specific or neutralizing antibodies respectively to F. psychrophilum and IHNV; however, antibody titers in eggs and fry were undetectable. In the eggs and fry mRNA transcripts of the complement components C3 and C5 were detected at much higher levels in progeny from vaccinated broodstock and showed a significantly increased and rapid response post-challenge compared with unvaccinated broodstock. After challenges pro-inflammatory cytokine expression was immediately and considerably elevated in the fry from vaccinated broodstock vs. unvaccinated broodstock, whereas adaptive immune genes were elevated to a lesser degree. Results suggest that maternal transfer of innate and adaptive factors at the transcript level occurred because development of lymphomyeloid organs is not complete in such young fry. In addition to documenting maternally derived immunity in teleosts, this study demonstrates that broodstock vaccination can confer some degree of protection to progeny against viral and bacterial pathogens.
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Enfermedades de los Peces , Infecciones por Flavobacteriaceae , Virus de la Necrosis Hematopoyética Infecciosa , Oncorhynchus mykiss , Infecciones por Rhabdoviridae , Vacunas de ADN , Femenino , Animales , Infecciones por Flavobacteriaceae/prevención & control , Infecciones por Flavobacteriaceae/veterinaria , Flavobacterium , Vacunación/veterinariaRESUMEN
The interplay of transcription factors and cis-regulatory elements (CREs) orchestrates the dynamic and diverse genetic programs that assemble the human central nervous system (CNS) during development and maintain its function throughout life. Genetic variation within CREs plays a central role in phenotypic variation in complex traits including the risk of developing disease. We took advantage of the retina, a well-characterized region of the CNS known to be affected by pathogenic variants in CREs, to establish a roadmap for characterizing regulatory variation in the human CNS. This comprehensive analysis of tissue-specific regulatory elements, transcription factor binding, and gene expression programs in three regions of the human visual system (retina, macula, and retinal pigment epithelium/choroid) reveals features of regulatory element evolution that shape tissue-specific gene expression programs and defines regulatory elements with the potential to contribute to Mendelian and complex disorders of human vision.
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Evolución Molecular , Regulación del Desarrollo de la Expresión Génica , Secuencias Reguladoras de Ácidos Nucleicos/genética , Retina/patología , Enfermedades de la Retina/genética , Adulto , Animales , Análisis Mutacional de ADN , Epigenómica , Femenino , Variación Genética , Humanos , Masculino , Ratones , Persona de Mediana Edad , Mutación , RNA-Seq , Retina/crecimiento & desarrollo , Enfermedades de la Retina/patología , Especificidad de la EspecieRESUMEN
Sablefish (Anoplopoma fimbria) are an emerging aquaculture species native to the continental shelf of the northern Pacific Ocean. There is limited information on both innate and adaptive immunity for this species and new tools are needed to determine antibody response following vaccination or disease outbreaks. In this paper, a monoclonal antibody, UI-25A, specific to sablefish IgM was produced in mice. Western blotting confirmed UI-25A recognizes the heavy chain of IgM and does not cross react to proteins or carbohydrates in serum of four other teleost species. An ELISA was developed to measure Aeromonas salmonicida specific IgM in the plasma of sablefish from a previous experiment where fish were immunized with a proprietary A. salmonicida vaccine. UI-25A was used in Western blot analyses to identify immunogenic regions of A. salmonicida recognized by this specific IgM from vaccinated sablefish. Immunofluorescent staining also demonstrated the ability of UI-25A to recognize membrane-bound IgM and identify IgM + cells in the head kidney. These results demonstrate the usefulness of UI-25A as a tool to improve the understanding of antibody-mediated immunity in sablefish as well as to provide valuable information for vaccine development and expansion of aquaculture efforts for this fish species.
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Anticuerpos Monoclonales , Perciformes , Animales , Anticuerpos Monoclonales/metabolismo , Western Blotting , Carbohidratos , Ensayo de Inmunoadsorción Enzimática/veterinaria , Peces/metabolismo , Inmunoglobulina M/metabolismo , Ratones , Coloración y EtiquetadoRESUMEN
Salmonid diseases caused by infections of Flavobacterium psychrophilum, the causative agent of bacterial coldwater disease, remain difficult to manage as novel, pathogenic strains continue to emerge in aquaculture settings globally. To date, much of the research regarding treatment options and vaccine development has focused on rainbow trout (Oncorhynchus mykiss), but other inland-reared salmonids are also impacted by this Gram-negative bacterium. As such, Atlantic salmon (Salmo salar) and brook trout (Salvelinus fontinalis) were injection-challenged with a variety of previously reported F. psychrophilum strains isolated from disease diagnostic cases in salmonids, as well as a standard and well-studied F. psychrophilum strain (CSF 259-93) known to be virulent in rainbow trout. In three separate virulence assessments (Trials A, B and C), strains US063 (isolated from lake trout; Salvelinus namaycush) and US149 (isolated from Atlantic salmon) caused a significantly higher cumulative per cent mortality (CPM) relative to other strains in Atlantic salmon (p <.001 for all trials), with US149 causing significantly greater mortality than US063 in Trials A (CPM 97% vs. 65%, p =.008) and B (CPM 96% ± 2.3% vs. 81.33% ± 4.8%, p =.014). Trial C used a lower dose (1.86 × 108 CFU/mL) for US149, resulting in a lower mortality (78.67% ± 9.33%) relative to Trials A and B. CSF259-93 did not cause significant mortality in any trials. In brook trout, the strain 03-179 (originally isolated from steelhead trout; Oncorhynchus mykiss) was significantly more virulent than any other (CPM 100% ± 0%, p <.001), followed by US063 (73% ± 3.8%) and US149 (40% ± 6.1%,) respectively. Again, CSF259-93 did not cause significant mortality relative to a mock challenge treatment. Results provide information about the applicability of strain selection in F. psychrophilum virulence testing in Atlantic salmon and brook trout, demonstrating the high virulence of US063 and US149 for these salmonid species. This information is applicable for the development of therapeutics and vaccines against F. psychrophilum infections and demonstrates the reproducibility of the experimental challenge model.
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Enfermedades de los Peces/mortalidad , Infecciones por Flavobacteriaceae/veterinaria , Flavobacterium/fisiología , Salmo salar , Trucha , Animales , Enfermedades de los Peces/microbiología , Infecciones por Flavobacteriaceae/microbiología , Infecciones por Flavobacteriaceae/mortalidadRESUMEN
Burbot (Lota lota maculosa) are a potential new species for commercial aquaculture. As burbot culture expands, there is a need to further define pathogen susceptibility and characterize aspects of the burbot immune response in an effort to assess fish health. A recent clinical diagnostic case from juvenile burbot reared at a commercial production facility resulted in the isolation and identification of Flavobacterium columnare along with several Aeromonas spp. The F. columnare isolate was assigned to genetic group 1 via multiplex PCR, a genetic group commonly associated with columnaris disease cases in rainbow trout (Oncorhynchus mykiss). Virulence of the F. columnare isolate was assessed in vivo in both juvenile burbot and rainbow trout. Additionally, several of the Aeromonas sp. case isolates were identified via sequencing (16S rRNA, gyrB and rpoD) and a putative A. sobria isolate (BI-3) was used to challenge burbot, along with a known virulent Aeromonas sp. (A141), but BI-3 was not found to be virulent. Burbot were refractory to F. columnare when challenged by immersion, and it is likely that this is a secondary pathogen for burbot. Although refractory in burbot, the identified F. columnare isolate (BI-1) was found to be virulent in rainbow trout.
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Aeromonas/fisiología , Enfermedades de los Peces/microbiología , Infecciones por Flavobacteriaceae/veterinaria , Flavobacterium/fisiología , Gadiformes , Infecciones por Bacterias Gramnegativas/veterinaria , Animales , Infecciones por Flavobacteriaceae/microbiología , Infecciones por Bacterias Gramnegativas/microbiología , Reacción en Cadena de la Polimerasa Multiplex/veterinaria , Análisis de Secuencia de ARN/veterinariaRESUMEN
Vaccines continue to play an enormous role in the progression of aquaculture industries worldwide. Though preventable diseases cause massive economic losses, injection-based vaccine delivery is cost-prohibitive or otherwise impractical for many producers. Most oral vaccines, which are much cheaper to administer, do not provide adequate protection relative to traditional injection or even immersion formulas. Research has focused on determining why there appears to be a lack of protection afforded by oral vaccines. Here, we review the basic immunological principles associated with oral vaccination before discussing the recent progress and current status of oral vaccine research. This knowledge is critical for the development and advancement of efficacious oral vaccines for the aquaculture industry.
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Throughout the current conflicts in Afghanistan and Iraq, there have been more than 1100 combat-related major limb amputations, with approximately 80% involving the lower extremity. There is, however, a paucity of data regarding the number of amputations below the level of the ankle. Although not as common, partial foot amputations, in the appropriate setting, offer a way to improve function and decrease energy consumption when compared to proximal amputations. Sound surgical tenets are prerequisite for successful outcomes when performing a distal amputation. Maintaining a robust soft tissue envelope allowing for tension-free wound closure is paramount in determining the feasibility of a partial foot amputation. Careful consideration of tendon balancing is also of utmost importance in avoiding common complications of contracture and deformity. Partial foot amputations present a viable surgical option for successful outcomes and maximization of patient function in the combat injured when certain criteria are met.
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Amputación Traumática/cirugía , Traumatismos por Explosión/cirugía , Traumatismos de los Pies/cirugía , Recuperación del Miembro/métodos , Personal Militar , Campaña Afgana 2001- , Amputación Traumática/rehabilitación , Traumatismos de los Pies/rehabilitación , Humanos , Guerra de Irak 2003-2011 , Recuperación del Miembro/rehabilitación , Masculino , Aparatos Ortopédicos , Complicaciones Posoperatorias/terapia , Estados Unidos , Adulto JovenRESUMEN
A web-based survey was conducted of ergonomics practitioners holding certifications in the U.S., Canada, Great Britain, Australia, and New Zealand. The survey follows 12 years after an earlier initial survey reported by Dempsey et al. (2005). Approximately 1221 eligible participants were invited by e-mail to participate, and 405 surveys were included in the final analysis. The survey queried use of basic instruments relevant to ergonomic practice as well as more specific analytical tools such as observational techniques for assessing postural demands of work and instrumentation for direct measurement of such demands. Some ergonomic assessment methods appear to have increased in their overall use by U.S. ergonomists compared to 2005 data. This was observed for: RULA, REBA, Psychophysical Upper Extremity Data, Strain Index, and ACGIH TLV for Hand Activity Level. There is minimal evidence of increased overall use of direct measurement approaches in the U.S. There appear to be geographic differences between countries/continents in terms of use of various methods. The use of mobile device/smart phone "apps" by ergonomists was queried and these technologies presently appear to be in early adoption phase with 24-28% of practitioners reporting use of an app in their ergonomics practice.
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Ergonomía/métodos , Adulto , Ergonomía/instrumentación , Femenino , Humanos , Masculino , Enfermedades Musculoesqueléticas/etiología , Enfermedades Musculoesqueléticas/prevención & control , Medición de Riesgo , Teléfono Inteligente/tendencias , Encuestas y CuestionariosRESUMEN
Fish immunization has been carried out for over 50 years and is generally accepted as an effective method for preventing a wide range of bacterial and viral diseases. Vaccination efforts contribute to environmental, social, and economic sustainability in global aquaculture. Most licensed fish vaccines have traditionally been inactivated microorganisms that were formulated with adjuvants and delivered through immersion or injection routes. Live vaccines are more efficacious, as they mimic natural pathogen infection and generate a strong antibody response, thus having a greater potential to be administered via oral or immersion routes. Modern vaccine technology has targeted specific pathogen components, and vaccines developed using such approaches may include subunit, or recombinant, DNA/RNA particle vaccines. These advanced technologies have been developed globally and appear to induce greater levels of immunity than traditional fish vaccines. Advanced technologies have shown great promise for the future of aquaculture vaccines and will provide health benefits and enhanced economic potential for producers. This review describes the use of conventional aquaculture vaccines and provides an overview of current molecular approaches and strategies that are promising for new aquaculture vaccine development.
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BACKGROUND: Autosomal-dominant optic atrophy (ADOA) is one of the most common types of inherited optic atrophy. We identify OPA1 pathogenic variants and assess the clinical features of a cohort of Chinese ADOA patients Materials and Methods: Detailed clinical evaluations were performed and genomic DNA was extracted from peripheral blood for all the participants. Sanger sequencing was used to analyze all exons and exon/intron junctions of OPA1 for eight pedigrees. Target exome capture plus next-generation sequencing (NGS) were applied for one atypical family with photophobia. Reverse transcription polymerase chain reaction was carried out to further characterize the mRNA change of selected splicing alteration. RESULTS: All 17 patients had impaired vision and optic-disk pallor; however, the clinical severity varied markedly. Two patients complicated with hearing loss. Six novel and two reported pathogenic variants in OPA1 (GenBank Accession No. NM_130837.2) were identified including four nonsynonymous variants (c.2400T > G, c.1468T > C, c.1567A > G and c.1466T > C), two splicing variants (c.2984-1_2986delGAGA and c.2983 + 5G > A), one small deletion (c.2960_2968delGCGTTCAAC), and one small insertion (c.3009_3010insA). RNA analysis revealed the splicing variant c.2984-1_2986delGAGA caused small deletion of mRNA (r.2983_2988del). CONCLUSIONS: ADOA patients presented variable clinical manifestations. Novel OPA1 pathogenic variants are the main genetic defect for Chinese ADOA cases. NGS may be a useful molecular testing tool for atypical ADOA.
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Pueblo Asiatico/genética , GTP Fosfohidrolasas/genética , Mutación , Atrofia Óptica Autosómica Dominante/genética , Adolescente , Adulto , Niño , Preescolar , Análisis Mutacional de ADN , Femenino , Humanos , Lactante , Masculino , Persona de Mediana Edad , Atrofia Óptica Autosómica Dominante/patología , Linaje , Pronóstico , Adulto JovenRESUMEN
INTRODUCTION: There is limited research to guide physicians and patients in deciding whether it is safe to drive while wearing various forms of upper extremity immobilization. The purpose of this study is to evaluate the effect of below-elbow removable splints and fiberglass casts on automobile driving performance. METHODS: 20 healthy subjects completed 10 runs through a closed, cone-marked driving course while wearing a randomized sequence of four different types of immobilization on each extremity (short arm thumb spica fiberglass cast, short arm fiberglass cast, short arm thumb spica splint, and short arm wrist splint). The first and last driving runs were without immobilization and served as controls. Performance was measured based on evaluation by a certified driving instructor (pass/fail scoring), cones hit, run time, and subject-perceived driving difficulty (1-10 analogue scoring). RESULTS: The greatest number of instructor-scored failures occurred while immobilized in right arm spica casts (n=6; p=0.02) and left arm spica casts (n=5; p=0.049). The right arm spica cast had the highest subject-perceived difficulty (5.2±1.9; p<0.001). All forms of immobilization had significantly increased perceived difficulty compared to control, except for the left short arm splint (2.5±1.6; p>0.05). There was no significant difference in number of cones hit or driving time between control runs and runs with any type of immobilization. CONCLUSIONS: Drivers should use caution when wearing any of the forms of upper extremity immobilization tested in this study. All forms of immobilization, with exception of the left short arm splint significantly increased perceived driving difficulty. However, only the fiberglass spica casts (both left and right arm), significantly increased drive run failures due to loss of vehicle control. We recommend against driving when wearing a below-elbow fiberglass spica cast on either extremity.
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Conducción de Automóvil , Inmovilización , Extremidad Superior/fisiopatología , Adolescente , Adulto , Anciano , Moldes Quirúrgicos/efectos adversos , Femenino , Lateralidad Funcional , Conocimientos, Actitudes y Práctica en Salud , Humanos , Masculino , Persona de Mediana Edad , Tiempo de Reacción , Seguridad , Férulas (Fijadores)/efectos adversos , Análisis y Desempeño de Tareas , Estados Unidos , Adulto JovenRESUMEN
Leber congenital amaurosis (LCA) is a severe disease that leads to complete blindness in children, typically before the first year of life. Due to the clinical and genetic heterogeneity among LCA and other retinal diseases, providing patients with a molecular diagnosis is essential to assigning an accurate clinical diagnosis. Using our gene panel that targets 300 genes that are known to cause retinal disease, including 24 genes reported to cause LCA, we sequenced 43 unrelated probands with Brazilian ancestry. We identified 42 unique variants and were able to assign a molecular diagnosis to 30/43 (70%) Brazilian patients. Among these, 30 patients were initially diagnosed with LCA or a form of early-onset retinal dystrophy, 17 patients harbored mutations in LCA-associated genes, while 13 patients had mutations in genes that were reported to cause other diseases involving the retina.
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Here we report a method for the preparation of anomerically pure ß-S-glycopyranosides (1,2-trans-glycosides) from the corresponding peracetate donors. S-glycosylation was performed in CHCl3 at reflux in the presence of a catalytic amount of InBr3. Deacylation of the intermediate peracetates were achieved under Zemplén conditions. Five pyranose examples, monosaccharides D-glucose and D-galactose and disaccharides cellobiose, maltose, and lactose, were used as donors, and five thiols including an α/ω dithiol and Fmoc-L-cysteine were used as acceptors. Melting points, high res MS, [α]D and NMR data ((1)H and (13)C, including COSY, HSQC and HMBC) are reported for compounds not previously described.
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Cisteína/química , Ácidos de Lewis/química , Tensoactivos/química , Tioglicósidos/química , Acilación , Catálisis , Indio/químicaRESUMEN
Naturally occurring glycopeptides and glycoproteins play important roles in biological processes. Glycosylation is one of the most common post-translational modifications in vivo. Glycopeptides are involved in cell signaling and sorting, providing cell surface markers for recognition. From the drug design and synthesis perspective, modification of a peptide through glycosylation results in increased bioavailability and bioactivity of glycopeptides in living systems with negligible toxicity of degradation products. Glycopeptide synthesis can be accomplished through incorporation of a glycosylated amino acid in solid phase peptide synthesis (SPPS) to form the desired peptide, or via incorporation of sugar-amino acid moieties. Additionally, research indicates that glycosylation increases penetration of the blood-brain barrier (BBB) by peptides, which may lead to novel therapeutics for neurological disorders. Recent applications of glycopeptides have focused on the in vivo central nervous system (CNS) effects after peripheral administration of centrally active peptides modified with various carbohydrates.
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STUDY DESIGN: This study utilized a quasi-experimental design in which subjects served as their own controls. OBJECTIVE: To determine whether heart rate, pain threshold velocity, and pain perception varied in patients running on a soft-belt treadmill versus a standard hard-belt treadmill. BACKGROUND: According to promotional literature, the relatively new Orbiter soft-belt tread produces a greater increase in heart rate at a given velocity as well as a higher velocity tolerance while walking or running. The manufacturer also asserts that decreased forces transmitted through the lower extremity should decrease pain levels while exercising on the soft-belt treadmill. METHODS AND MEASURES: Twenty-seven subjects walked or ran on each of 2 treadmills at incrementally increasing velocities until they experienced either the onset of pain or an increase in pain from baseline levels. Locomotion continued for 2 minutes after that, during which time heart rate and pain level on a visual analog scale (VAS) were recorded. RESULTS: Two univariate paired t tests and a Wilcoxon's signed rank test revealed a greater heart rate and pain threshold velocity when using the soft-belt treadmill with no statistical difference in the pain reported between the 2 treadmills. CONCLUSION: Our study revealed a 10% higher heart rate and a 14.5% higher pain threshold velocity with the soft-belt treadmill compared to a hard-belt treadmill. These differences are considered clinically meaningful.