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ABSTRACTMutations in the SARS-CoV-2 genome may negatively impact a diagnostic test, have no effect, or turn into an opportunity for rapid molecular screening of variants. Using an in-house Emergency Use Authorized RT-qPCR-based COVID-19 diagnostic assay, we combined sequence surveillance of viral variants and computed PCR efficiencies for mismatched templates. We found no significant mismatches for the N, E, and S set of assay primers until the Omicron variant emerged in late November 2021. We found a single mismatch between the Omicron sequence and one of our assay's primers caused a > 4 cycle delay during amplification without impacting overall assay performance.Starting in December 2021, clinical specimens received for COVID-19 diagnostic testing that generated a Cq delay greater than 4 cycles were sequenced and confirmed as Omicron. Clinical samples without a Cq delay were largely confirmed as the Delta variant. The primer-template mismatch was then used as a rapid surrogate marker for Omicron. Primers that correctly identified Omicron were designed and tested, which prepared us for the emergence of future variants with novel mismatches to our diagnostic assay's primers. Our experience demonstrates the importance of monitoring sequences, the need for predicting the impact of mismatches, their value as a surrogate marker, and the relevance of adapting one's molecular diagnostic test for evolving pathogens.
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COVID-19 , Humanos , COVID-19/diagnóstico , Teste para COVID-19 , Saúde Pública , SARS-CoV-2/genéticaRESUMO
Rapid and widespread testing of severe acute respiratory coronavirus 2 (SARS-CoV-2) is essential for an effective public health response aimed at containing and mitigating the coronavirus disease 2019 (COVID-19) pandemic. Successful health policy implementation relies on early identification of infected individuals and extensive contact tracing. However, rural communities, where resources for testing are sparse or simply absent, face distinctive challenges to achieving this success. Accordingly, we report the development of an academic, public land grant University laboratory-based detection assay for the identification of SARS-CoV-2 in samples from various clinical specimens that can be readily deployed in areas where access to testing is limited. The test, which is a quantitative reverse transcription polymerase chain reaction (RT-qPCR)-based procedure, was validated on samples provided by the state laboratory and submitted for FDA Emergency Use Authorization. Our test exhibits comparable sensitivity and exceeds specificity and inclusivity values compared to other molecular assays. Additionally, this test can be re-configured to meet supply chain shortages, modified for scale up demands, and is amenable to several clinical specimens. Test development also involved 3D engineering critical supplies and formulating a stable collection media that allowed samples to be transported for hours over a dispersed rural region without the need for a cold-chain. These two elements that were critical when shortages impacted testing and when personnel needed to reach areas that were geographically isolated from the testing center. Overall, using a robust, easy-to-adapt methodology, we show that an academic laboratory can supplement COVID-19 testing needs and help local health departments assess and manage outbreaks. This additional testing capacity is particularly germane for smaller cities and rural regions that would otherwise be unable to meet the testing demand.
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Teste de Ácido Nucleico para COVID-19/instrumentação , COVID-19/diagnóstico , Kit de Reagentes para Diagnóstico , Serviços de Saúde Rural/organização & administração , COVID-19/epidemiologia , COVID-19/prevenção & controle , COVID-19/virologia , Controle de Doenças Transmissíveis/métodos , Controle de Doenças Transmissíveis/organização & administração , Desenho de Equipamento , Humanos , Limite de Detecção , Nasofaringe/virologia , Pandemias/prevenção & controle , Impressão Tridimensional , RNA Viral/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real , SARS-CoV-2/genética , SARS-CoV-2/isolamento & purificação , Sensibilidade e Especificidade , Manejo de Espécimes/instrumentação , Manejo de Espécimes/métodosRESUMO
The organic-inorganic lead-halide perovskites are composed of organic molecules imbedded in an inorganic framework. The compounds with general formula CH3NH3PbX 3 (MAPbX 3) display large photovoltaic efficiencies for halogens X = Cl, Br, and I in a wide variety of sample geometries and preparation methods. The organic cation and inorganic framework are bound by hydrogen bonds that tether the molecules to the halide anions, and this has been suggested to be important to the optoelectronic properties. We have studied the effects of this bonding using time-of-flight neutron spectroscopy to measure the molecular dynamics in CH3NH3PbCl3 (MAPbCl3). Low-energy/high-resolution neutron backscattering reveals thermally activated molecular dynamics with a characteristic temperature of ~95 K. At this same temperature, higher-energy neutron spectroscopy indicates the presence of an anomalous broadening in energy (reduced lifetime) associated with the molecular vibrations. By contrast, neutron powder diffraction shows that a spatially long-range structural phase transitions occurs at 178 K (cubic â tetragonal) and 173 K (tetragonal â orthorhombic). The large difference between these two temperature scales suggests that the molecular and inorganic lattice dynamics in MAPbCl3 are actually decoupled. With the assumption that underlying physical mechanisms do not change with differing halogens in the organic-inorganic perovskites, we speculate that the energy scale most relevant to the photovoltaic properties of the lead-halogen perovskites is set by the lead-halide bond, not by the hydrogen bond.
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The relaxor PbMg1/3Nb2/3O3 (PMN) has received attention due to its potential applications as a piezoelectric when doped with PbTiO3 (PT). Previous results have found that there are two phases existing in the system, one linked to the near-surface regions of the sample, the other in the bulk. However, the exact origin of these two phases is unclear. In this paper, depth dependant analysis results from negative muon implantation experiments are presented. It is shown that the Pb content is constant throughout all depths probed in the sample, but the Mg and Nb content changes in the near-surface region below 100 µm. At an implantation depth of 60 µm, it is found that there is a 25% increase in Mg content, with a simultaneous 5% decrease in Nb content in order to maintain charge neutrality. These results show that the previously observed skin effects in PMN are due to a change in concentration and unit cell.
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AIM: The children's health state preferences learnt from animation (CHILDSPLA) project developed an interactive application presented on a touch screen device using an animated character to collect information from children about their health. BACKGROUND: The underlying hypothesis was that health information could be directly collected from children as young as 4 years old by the use of animated characters. This paper describes in detail how children were involved in the development of the application, and recounts both the challenges and benefits of that process. A child psychologist and an animation filmmaker worked closely with children to design a character and to animate it to represent different health states. Children were recruited from a local primary school (n = 38) and a paediatric specialist hospital (n = 36). Diverse interactive activities were organized to help children give feedback and guide the design process. The activities for each session were adjusted to the children's needs, based on the experience of previous sessions. RESULTS: The character and the animations were modified according to the feedback provided by the children. CONCLUSIONS: Developing the CHILDSPLA app in collaboration with children was a worthwhile and enriching experience, despite the required iteration and extension of the design process, as it enabled us to adjust the tool to the children's needs.
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Comportamento Cooperativo , Indicadores Básicos de Saúde , Aplicativos Móveis , Relações Profissional-Paciente , Interface Usuário-Computador , Jogos de Vídeo , Adolescente , Criança , Pré-Escolar , Retroalimentação , Feminino , Humanos , Masculino , EscóciaRESUMO
The Renal and Lung Living Donors Evaluation Study assesses outcomes of live lung (lobectomy) donors. This is a retrospective cohort study at University of Southern California (USC) and Washington University (WASHU) Medical Centers (19932006), using medical records to assess morbidity and national databases to ascertain postdonation survival and lung transplantation. Serious complications were defined as those that required significant treatment, were potentially life-threatening or led to prolonged hospitalization. The 369 live lung donors (287 USC, 82 WASHU) were predominantly white, non-Hispanic and male; 72% had a biological relationship to the recipient, and 30% were recipient parents. Serious complications occurred in 18% of donors; 2.2% underwent reoperation and 6.5% had an early rehospitalization. The two centers had significantly different incidences of serious complications (p < 0.001). No deaths occurred and no donors underwent lung transplantation during 4000+ person-years of follow-up (death: minimum 4, maximum 17 years; transplant: minimum 5, maximum 19). Live lung donation remains a potential option for recipients when using deceased donor lungs lacks feasibility. However, the use of two live donors for each recipient and the risk of morbidity associated with live lung donation do not justify this approach when deceased lung donors remain available. Center effects and long-term live donor outcomes require further evaluation.
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Doadores Vivos/estatística & dados numéricos , Pneumopatias/mortalidade , Pneumopatias/cirurgia , Transplante de Pulmão , Adolescente , Adulto , Estudos de Coortes , Bases de Dados Factuais , Feminino , Humanos , Tempo de Internação , Pulmão/cirurgia , Masculino , Pessoa de Meia-Idade , Controle de Qualidade , Projetos de Pesquisa , Estudos Retrospectivos , Resultado do Tratamento , Adulto JovemRESUMO
In the present study we examined the effects of normal aging in the hippocampus and cerebellum, as well as behaviors associated with these substrates. A total of 67 CB6F1 hybrid mice were tested at one of five ages (4, 8, 12, 18 or 25 months) on the context pre-exposure facilitation effect (CPFE) modification of fear conditioning, rotorod, Barnes maze, acoustic startle, Morris water maze (MWM) and 500-ms trace eyeblink classical conditioning (EBCC). Behavioral tasks were chosen to increase the ability to detect age-related changes in learning, as trace EBCC is considered a more difficult paradigm (compared to delay EBCC) and the CPFE has been found to be more sensitive to hippocampus insults than standard contextual fear conditioning. To assess the effects of age on the brain, hippocampus volume was calculated and unbiased stereology was used to estimate the number of Purkinje neurons in the cerebellar cortex. A significant, age-related loss of Purkinje neurons was found-beginning at 12 months of age-and hippocampus volume remained stable over the adult life span. Age-related impairment was found, beginning at 12-18 months in the rotorod, and mice with fewer Purkinje neurons showed greater impairment in this task. CB6F1 mice retained auditory acuity across the life span and mice aged 25 months showed significant age-related impairment in the EBCC task; however, deficits were not associated with the loss of Purkinje neurons. Although the CPFE task is considered more sensitive to hippocampus insult, no age-related impairment was found. Spatial memory retention was impaired in the Barnes maze at 25 months, but no significant deficits were seen in the MWM. These results support the finding of differential aging in the hippocampus and cerebellum.
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Envelhecimento/metabolismo , Cerebelo/metabolismo , Condicionamento Palpebral/fisiologia , Hipocampo/metabolismo , Aprendizagem/fisiologia , Longevidade/fisiologia , Animais , Cerebelo/patologia , Hipocampo/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Tamanho do ÓrgãoRESUMO
INTRODUCTION: Expert guidelines indicate that normalised ratios are preferred to clotting times for lupus anticoagulant (LA) assays to mitigate analytical variation. We investigated the effects of deriving normalised ratios from the reference interval (RI) mean or different normal pooled plasmas (NPP). METHODS: Screen, confirm and mixing tests for dilute Russell's viper venom time and APTT were converted to normalised ratios and interpreted for LA. RESULTS: Of 1000 clinical samples, 824 generated identical interpretations using RI mean or NPP-derived ratios and 57 identified LAs in one or both assays via either denominator. Separate RIs were applied for normalised ratios derived from the NPP or RI mean. Applying percentage correction index (PCI) to screen and confirm assays irrespective of screen elevation increased agreement to 92.5%. Two frozen and one lyophilised NPP were then used to derive ratios for 204 samples and 130 generated identical interpretations with all NPPs, 14 had overall interpretation parity and 19 overall agreement via PCI, giving 79.9% overall agreement. The results derived from each NPP were interpreted against RIs derived from RI means to reflect differences resulting from NPPs with clotting times dissimilar to RI means. CONCLUSIONS: Disparities were largely a function of closeness of NPP clotting times to test RI means and not owing to clotting factor level differences and likely related to manufacturing variables. Diagnostic benefit of normalised ratios can be maximised by matching NPP values to RI means. If RI mean is employed, and it likely requires re-establishing with new reagent batches.
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Síndrome Antifosfolipídica/diagnóstico , Testes de Coagulação Sanguínea/estatística & dados numéricos , Testes de Coagulação Sanguínea/normas , Inibidor de Coagulação do Lúpus/sangue , Interpretação Estatística de Dados , Erros de Diagnóstico , HumanosRESUMO
Holsteins (HH), Jerseys (JJ), and their crosses in first (n=157) and second (n=107) lactation were used to determine if reproduction, progesterone (P4), insulin-like growth factor 1 (IGF-1), insulin, nonesterified fatty acids (NEFA), and milk production differed between genetic groups. Thirty-four cows were Holstein-Jersey (HJ) crosses, 46 were Jersey-Holstein (JH) crosses, 48 were purebred Holsteins (HH), and 29 were purebred Jerseys (JJ) in first lactation, whereas the second-lactation animals included 23 HJ, 35 JH, 35 HH, and 14 JJ. Blood samples were collected weekly for the first 10 wk postpartum. Analyses were conducted using the MIXED, chi-square, and GLIMMIX procedures (SAS Institute Inc., Cary, NC). Seasons of calving were cold (November to May) and hot (June to October) and were combined with year to form 8 year-seasons. Days open and number of services were affected by genetic group. The HH were open 169±8 d, which was greater than HJ (143±9 d), JJ (132±10 d), and JH (127±8 d). The HH had 2.4±0.1 services per pregnancy, which was greater than JH (1.9±0.1), but not different from HJ (2.1±0.2) or JJ (2.1±0.2). Concentrations of NEFA were greater in lactation 2 (0.52±0.02 mEq/L) than in lactation 1 (0.45±0.02 mEq/L) and decreased over the 10-wk period. Concentrations of NEFA were greater in the cold season except in yr 3. Insulin in lactation 1 (0.81±0.03 ng/mL) was greater than in lactation 2 (0.72±0.03 ng/mL); insulin decreased to wk 2 then gradually increased. The HJ had the greatest insulin concentrations (0.87±0.04 ng/mL) and the JJ had the lowest (0.66±0.04 ng/mL), and IGF-1 gradually increased over the 10-wk period. Milk production (actual yield in the first 305 d, not adjusted for fat and protein) was affected by genetic group, lactation number, year-season, and wk 1 insulin. The HH produced 10,348±207 kg of milk, which was greater than the HJ (9,129±230 kg), the JH (9,384±190 kg), and the JJ (7,080±240 kg). Milk production in lactation 2 (9,676±163 kg) was greater than that in lactation 1 (8,294±160 kg). The JJ (10.3±4.7%) had the highest frequency of mastitis. The chance of getting mastitis for HH (1.1±0.9%) differed from that for HJ (9.4±4.1%), JH (8.1±3.4%), and JJ (10.3±4.7%). Genetic group affected hormones and metabolites, which may partially explain differences in reproductive measures and milk yield.
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Bovinos/fisiologia , Reprodução/fisiologia , Animais , Cruzamento , Bovinos/metabolismo , Ácidos Graxos não Esterificados/sangue , Feminino , Insulina/sangue , Fator de Crescimento Insulin-Like I/análise , Lactação/fisiologia , Gravidez , Progesterona/sangue , Especificidade da EspécieRESUMO
In mid-June 2008, distinct mosaic leaves were observed on a cluster of clover (Trifolium spp.) with light pink and white flowers growing at the edge of a lawn in Palmer, AK. Virus minipurification from leaves of affected clover and protein extractions on a polyacrylamide electrophoresis implicated a ~35-kDa putative coat protein (CP). Subsequent western blots and ELISA with a universal potyvirus antiserum (Agdia Inc., Elkhart, IN) confirmed potyvirus identity. Total RNA extracts (RNeasy Plant Mini Kit, Qiagen Inc., Valencia, CA) from the same plant were used for reverse transcription (RT)-PCR. Three sets of degenerate primers that targeted potyvirus-specific genes, HC-Pro (helper component protease) and CI (cylindrical inclusion protein) and the genomic 3'-terminus that included a partial NIb (nuclear inclusion), CP (coat protein), and UTR (untranslated region), produced the expected PCR segments (~0.7, ~0.7, and ~1.6 kbp, respectively) on 1% agarose gels (1). Direct sequencing of the HC-Pro (GenBank No. GQ181115), CI (GQ181116), and CP (GU126690) segments revealed 98, 97, and 99% nucleotide identities (no gaps), respectively, to Bean yellow mosaic virus (BYMV)-chlorotic spot (CS) strain, GenBank No. AB373203. The next closest BYMV percent identity comparisons decreased to 79% for HC-Pro (GenBank No. DQ641248; BYMV-W), 79% for CI (U47033; BYMV-S) partial genes, and 96% for CP (AB041971; BYMV-P242). Mechanical inoculations of purified virus preparations produced local lesions on Chenopodium amaranticolor Coste & A. Reyn. (2 of 5) and C. quinoa Willd. (6 of 7), and mosaic on Nicotiana benthamiana Domin (5 of 5). BYMV was specifically confirmed on tester plants using a double-antibody sandwich (DAS)-ELISA BYMV (strain 204 and B25) kit (AC Diagnostics, Inc., Fayetteville, AR) as directed. The absence of another potyvirus commonly found in clover, Clover yellow vein virus (ClYVV), was verified in parallel DAS-ELISA ClYVV assays (AC Diagnostics, Inc). The BYMV isolate was maintained in N. benthamiana, and virion or sap extracts inoculated to the following host range (number of infected/total inoculated plants [verified by BYMV ELISA]): Cucumis sativus L. 'Straight Eight' (0/5), Gomphrena globosa L. (1/4), Nicotiana clevelandii A. Gray (4/7), Phaseolus vulgaris L. 'Bountiful' (1/3), Pisum sativum L. (Germplasm Resources Information Network Accession Nos. -PI 508092 (8/12), -W6 17525 (13/13), -W6 17529 (0/13), -W6 17530 (13/14), -W6 17537 (0/12), -W6 17538 (0/12), and -W6 17539 (0/21), Tetragonia tetragoniodes (2/2), Trifolium pretense L. 'Altaswede' (6/10), T. repens L. 'Pilgrim' (0/8), and Vicia faba L. (1/3). All infected plants had symptoms ranging from systemic mosaic (T. pretense, P. sativum) to leaf distortions (N. clevelandii, Tetragonia tetragoniodes). Interestingly, the host range and genomic sequences of the BYMV Alaskan strain resemble the BYMV-CS (chlorotic spot) strain that was originally isolated from a diseased red clover (T. pretense) plant in Japan more than 40 years ago (2). Although BYMV occurs worldwide and has a wide host range in dictoyledonous and monocotyledonous plants (3), to our knowledge, this is the first report of a natural occurrence of BYMV in Alaska. The incidence and distribution of BYMV in clover and other plant species are not known in Alaska. References: (1) C. Ha et al. Arch. Virol. 153:36, 2008. (2) H. Kume et al. Mem. Fac. Agric. Hokkaido Univ. 7:449, 1970. (3) S. J. Wylie et al. Plant Dis. 92:1596, 2008.
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Wild larkspur, Delphinium glaucum S. Watson, grows throughout most of Alaska along roadsides and in forests and is planted as an ornamental. Leaves containing distinct vein-clearing and chlorotic mosaic symptoms were first noticed on several D. glaucum plants during 2000 at the Georgeson Botanical Garden in Fairbanks, AK. Although affected plants continued to produce normal flowers, by 2008, the plants developed overall stunting. Initially, virus presence was determined by a general differential centrifugation extraction and concentration protocol followed by examination of the partially purified virus and leaf sap by electron microscopy. Filamentous particles approximately 725 nm long were observed. Virion protein extractions analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a putative coat protein (CP) of ~35 kDa. Potyvirus identity (family Potyviridae) was confirmed with universal potyvirus antiserum in western blots and ELISA assays (Agdia, Inc., Elkhart, IN). Exotic larkspur plants, D. elatum L., growing next to diseased D. glaucum plants, did not exhibit symptoms nor were they positive for potyvirus when tested serologically as described previously. Total RNA was extracted from potyvirus-infected leaves and used in reverse transcriptase-PCR assays that specifically targeted potyviruses (2,4) to generate genomic segments for identification and sequence analysis. Fragments representing portions of the helper component protease gene, HC-Pro (~700 bp), the cylindrical inclusion gene, CI (~700 bp), and the 3'-end (~1.7 kbp) were purified, cloned, sequenced, and deposited in GenBank (Accession Nos. FJ349329, FJ349328, and FJ349327, respectively). The sequenced 3'-end (1,674 nt) revealed a partial nuclear inclusion protein gene, NIb (1 to 630 nt), a CP gene (631 to 1,443 nt), and a 3'-untranslated region (1,447 to 1,674 nt) attached to a poly (A) tail. Blast searches in GenBank for percent identities of the nucleotide and amino acid comparisons resulted in highest similarities in conserved regions among members in the genus Potyvirus. For example, the highest CI, CP, and HP amino acid identities (0 gaps) were 67% with Potato virus A (Accession No. AF543709), 74% with Araujia mosaic virus (Accession No. EF710625), and 65% with Potato virus A (Accession No. AJ131403), respectively. However, none of the identities were sufficient for inclusion with an existing potyvirus species, whereby the CP amino acid sequence identity must be at least 80% (1). Mechanical transmission of purified virus to Chenopodium amaranticolor, C. quinoa, D. elatum, D. glaucum, and Nicotiana benthamiana seedlings was unsuccessful. We conclude that the isolated virus is a new species in the genus Potyvirus and propose the name Delphinium vein-clearing virus (DeVCV). To our knowledge, this is the first report of a virus isolated from D. glaucum and is representative of the growing number of viruses found in native plants (3). The distribution of DeVCV-infected larkspur is not known in managed or natural ecosystems. Identification of new viruses from native plants is important, in that, the host plant may act as a virus reservoir for transmission to other ornamental and crop plants. References: (1) P. H. Berger et al. Family Potyviridae. Page 819 in: Virus Taxonomy-8th Report of the ICTV. C. M. Fauquet et al., eds. Elsevier Academic Press, San Diego, CA, 2005. (2) J. Chen et al. Arch. Virol. 146:757, 2001. (3) I. Cooper and A. C. Jones. Adv. Virus Res. 67:1, 2006. (4) C. Ha et al. Arch. Virol. 153:25, 2008.
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Peonies (Paeonia sp.) are highly valued for their large showy flowers in home gardens and commercially in the cut flower industry. In 2007, scattered peony (Paeonia lactiflora 'Sarah Bernhardt') plants cultivated on small plots at the University of Alaska Experimental Station in Fairbanks displayed distinct leaf ringspot patterns. Symptoms were more severe during the cooler months of the growing season (June and September), with symptom remission in the intervening warmer months. Leaf samples from six symptomatic plants were collected in July and from 20 symptomatic plants in September and assayed for viruses. Leaf samples (1 g) were assayed with a general protocol for plant virus extraction and partial purification with differential centrifugation followed by protein detection on stained sodium dodecyl sulfate-polyacrylamide gel electrophoresis (1). No distinct proteins indicative of viral coat protein(s) were detected. Tomato spotted wilt virus (TSWV) and Tobacco rattle virus (TRV), known pathogens of peony, were then specifically targeted. Total RNA was extracted from each sample with an RNeasy Plant Mini kit (Qiagen Inc., Valencia, CA) and used as the template for reverse transcription (RT)-PCR with random primers. TSWV was not detected by RT-PCR with tospovirus group-specific primers (Agdia, Inc., Elkhart, IN). A nested set of primers designed from the TRV 16-kDa protein gene on RNA1 (4) amplified an ~600-bp fragment from one of the symptomatic plants. This DNA was directly sequenced (GenBank Accession No. FJ357572) and BLAST searches in GenBank revealed as much as 95% nucleotide (nt) identity with TRV accessions J04347 and X03685. Additional primer pairs specific for TRV (2) amplified overlapping fragments with expected sizes of ~818, ~515, and ~290 bp from the 29- and 16-kDa protein genes on the 3'-end of RNA1 that were directly sequenced. Assembly of these sequences in Sequencher 4.8 (Gene Codes Corp., Ann Arbor, MI) resulted in a 1,422-nt sequence (Accession No. FJ357571) and Clustal X analysis (3) showed 93 to 94% nt identity to TRV isolates, -ORY (AF034622), -PpK20 (AF314165), -Pp085 (AJ586803), and -SYM (D00155). Mechanical inoculation of partially purified virions from the confirmed TRV-infected peony plant to Nicotiana benthamiana gave no symptoms to occasional ringspots, faintly curled leaves, and chlorotic blotches on N. tabacum 'Samsun', and local lesions on Chenopodium amaranticolor. TRV infection of these hosts was confirmed by RT-PCR. With electron microscopy, rod-shaped particles similar to TRV with a distinct central canal characteristic of TRV were seen occasionally only from inoculated N. benthamiana. On the basis of the biological and molecular data, we have determined the virus in the peony to be an isolate of TRV, tentatively named TRV-Peony. TRV was confirmed in only one other peony based on a sequenced 290-nt PCR fragment with 95% identity with the sequence from the other TRV-infected peony. Lack of TRV detection in the other symptomatic peonies was possibly due to low viral concentrations and interfering plant substances. Documentation of TRV in peonies is especially important to help avoid distribution of virus-infected vegetative propagation material. To our knowledge, this is the first report of TRV in this host in Alaska, but also of this virus in Alaska. References: (1) L. C. Lane. Methods Enzymol. 118:687, 1986. (2) D. J. Robinson. J. Phytopathol. 152:286, 2004. (3) J. D. Thompson et al. Nucleic Acids Res. 24:4882, 1997. (4) F. Van Der Wilk et al. Eur. J. Plant Pathol.100:109, 1994.
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Experimental unilateral ureteral obstruction (UUO) is widely used to study renal fibrosis; however, renal injury can only be scored semiobjectively by histology. We sought to improve the UUO model by reimplanting the obstructed ureter followed by removal of the contralateral kidney, thus allowing longitudinal measurements of renal function. Mice underwent UUO for different lengths of time before ureteral reimplantation and contralateral nephrectomy. Measurement of blood urea nitrogen (BUN) allows objective evaluation of residual renal function. Seven weeks after reimplantation and contralateral nephrectomy, mean BUN levels were increased with longer duration of UUO. Interstitial expansion, fibrosis, and T-cell and macrophage infiltration were similar in kidneys harvested after 10 days of UUO or following 10 weeks of ureter reimplantation, suggesting that the inflammatory process persisted despite relief of obstruction. Urinary protein excretion after reimplantation was significantly increased compared to control animals. Our study shows that functional assessment of the formerly obstructed kidney can be made after reimplantation and may provide a useful model to test therapeutic strategies for reversing renal fibrosis and preserving or restoring renal function.
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Modelos Animais de Doenças , Rim/fisiopatologia , Ureter/cirurgia , Obstrução Ureteral/cirurgia , Animais , Feminino , Rim/patologia , Camundongos , Camundongos Endogâmicos C57BL , NefrectomiaRESUMO
OBJECTIVES: To assess whether the route by which neonatal congenital heart disease (CHD) is first recognised influences outcome after surgery. METHODS: Surgical neonates admitted to a tertiary cardiac unit between March 1999 and February 2002 were retrospectively reviewed with analysis of risk factors for outcome. Three routes to initial recognition of CHD were compared: antenatal diagnosis, detection on the postnatal ward, and presentation after discharge to home. Outcome measures were mortality and duration of perioperative ventilation. RESULTS: 286 neonates had cardiac surgery with a median duration of ventilation of 101 h and in-hospital mortality of 12%. Recognition of CHD was antenatal in 20%, on the postnatal ward in 55% and after discharge to home in 25%. Multiple regression analyses, including the cardiac diagnosis, associated problems and other risk factors, indicated that severe cardiovascular compromise on admission to the cardiac unit was significantly related to mortality and prolonged ventilation. Considered in isolation, the route to recognition of heart disease did not influence mortality or ventilation time. Route to initial recognition did, however, influence the patient's condition on admission to the cardiac unit. Cardiovascular compromise and end organ dysfunction were least likely when recognition was antenatal and most common when presentation followed discharge to home. CONCLUSION: The setting in which neonatal CHD is first recognised has an impact on preoperative condition, which in turn influences postoperative progress and survival after surgery. Optimal screening procedures and access to specialist care will improve outcome for neonates undergoing cardiac surgery.
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Cardiopatias Congênitas/diagnóstico , Análise de Variância , Cardiopatias Congênitas/mortalidade , Cardiopatias Congênitas/cirurgia , Humanos , Recém-Nascido , Cuidados Pré-Operatórios , Prognóstico , Fatores de RiscoRESUMO
OBJECTIVE: To identify predictors of outcome in ex-premature infants supported with extracorporeal membrane oxygenation (ECMO) for acute hypoxic respiratory failure. METHODS: Retrospective review of ex-premature infants with acquired acute hypoxic respiratory failure requiring ECMO support in the United Kingdom from 1992 to 2001. Review of follow up questionnaires completed by general practitioners and local paediatricians. RESULTS: Sixty four ex-premature infants (5-10 each year) received ECMO support, despite increased use of advanced conventional treatments over the decade. The most common infective agent was respiratory syncytial virus (85% of cases). Median birth gestation was 29 weeks and median corrected age at the time of ECMO support was 42 weeks. Median ECMO support duration was relatively long, at 229 hours. Survival to hospital discharge and to 6 months was 80%, remaining similar throughout the period of review. At follow up, 60% had long term neurodisability and 79% had chronic pulmonary problems. Of pre-ECMO factors, baseline oxygen dependence, younger age, and inpatient status were associated with non-survival (p < or = 0.05). Of ECMO related factors, patient complications were independently associated with adverse neurodevelopmental outcome and death (p < 0.01). CONCLUSIONS: Survival rates for ex-premature infants after ECMO support are favourable, but patients suffer a high burden of morbidity during intensive care and over the long term. At the time of ECMO referral, baseline oxygen dependence is the most important predictor of death, but no combination of the factors considered was associated with a mortality that would preclude ECMO support.
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Oxigenação por Membrana Extracorpórea , Recém-Nascido Prematuro , Insuficiência Respiratória/terapia , Doença Aguda , Humanos , Lactente , Recém-Nascido , Prognóstico , Infecções por Vírus Respiratório Sincicial/terapia , Estudos Retrospectivos , Fatores de Risco , Estações do Ano , Análise de Sobrevida , Resultado do TratamentoRESUMO
OBJECTIVE: To delineate predictors of hospital survival in a large series of children with biventricular physiology supported with extracorporeal membrane oxygenation (ECMO) after open heart surgery. RESULTS: 81 children were placed on ECMO after open heart surgery. 58% (47 of 81) were transferred directly from cardiopulmonary bypass to ECMO. Hospital survival was 49% (40 of 81) but there were seven late deaths among these survivors (18%). Factors that improved the odds of survival were initiation of ECMO in theatre (64% survival (30 of 47)) rather than the cardiac intensive care unit (29% survival (10 of 34)) and initiation of ECMO for reactive pulmonary hypertension. Important adverse factors for hospital survival were serious mechanical ECMO circuit problems, renal support, residual lesions, and duration of ECMO. CONCLUSIONS: Hospital survival of children with biventricular physiology who require cardiac ECMO is similar to that found in series that include univentricular hearts, suggesting that successful cardiac ECMO is critically dependent on the identification of hearts with reversible ventricular dysfunction. In our experience of postoperative cardiac ECMO, the higher survival of patients cannulated in the operating room than in the cardiac intensive care unit is due to early effective support preventing prolonged hypoperfusion and the avoidance of a catastrophic cardiac arrest.
Assuntos
Oxigenação por Membrana Extracorpórea/métodos , Cardiopatias Congênitas/cirurgia , Ponte Cardiopulmonar/métodos , Criança , Pré-Escolar , Oxigenação por Membrana Extracorpórea/efeitos adversos , Mortalidade Hospitalar , Humanos , Lactente , Recém-Nascido , Nefropatias/etiologia , Nefropatias/terapia , Hemorragia Pós-Operatória/etiologia , Hemorragia Pós-Operatória/terapia , Análise de Regressão , Paralisia Respiratória/etiologia , Paralisia Respiratória/terapia , Estudos Retrospectivos , Sepse/etiologia , Análise de Sobrevida , Resultado do TratamentoRESUMO
Follicular dendritic cells (FDCs) of the lymphoreticular system play a role in the peripheral replication of prion proteins in some transmissible spongiform encephalopathies (TSEs), including experimental murine scrapie models. Disease-specific PrP (PrPd) accumulation occurs in association with the plasmalemma and extracellular space around FDC dendrites, but no specific immunological response has yet been reported in animals affected by TSEs. In the present study, morphology (light microscopical and ultrastructural) of secondary lymphoid follicles of the spleen were examined in mice infected with the ME7 strain of scrapie and in uninfected control mice, with or without immunological stimulation with sheep red blood cells (SRBCs), at 70 days post-inoculation or at the terminal stage of disease (268 days). Scrapie infection was associated with hypertrophy of FDC dendrites, increased retention of electron-dense material at the FDC plasma membrane, and increased maturation and numbers of B lymphocytes within secondary follicles. FDC hypertrophy was particularly conspicuous in immune-stimulated ME7-infected mice. The electron-dense material was associated with PrP Napoli accumulation, as determined by immunogold labelling. We hypothesize that immune system changes are associated with increased immune complex trapping by hypertrophic FDCs expressing PrP Napoli molecules at the plasmalemma of dendrites, and that this process is exaggerated by immune system stimulation. Contrary to previous dogma, these results show that a pathological response within the immune system follows scrapie infection.
Assuntos
Centro Germinativo/patologia , Proteínas PrPSc/metabolismo , Scrapie/imunologia , Scrapie/patologia , Baço/patologia , Animais , Complexo Antígeno-Anticorpo/imunologia , Células Dendríticas Foliculares/imunologia , Células Dendríticas Foliculares/patologia , Células Dendríticas Foliculares/ultraestrutura , Centro Germinativo/ultraestrutura , Imuno-Histoquímica , Camundongos , Microscopia Eletrônica , Modelos Imunológicos , Baço/imunologia , Baço/ultraestruturaRESUMO
OBJECTIVE: In this report of a near-fatal case of grape aspiration successfully treated with extracorporeal membrane oxygenation (ECMO), we highlight the danger of feeding seedless grapes to young children and demonstrate that ECMO can provide cardiopulmonary support for cases of acquired large-airway disruption and can facilitate therapeutic intervention. DESIGN: Case report. SETTING: A tertiary pediatric intensive care unit and ECMO center. PATIENT: A healthy 14-month-old boy aspirated a seedless grape while playing at home and suffered a cardiopulmonary arrest of 15 mins in duration. He responded to advanced life support with return of cardiac output but developed intractable cardiopulmonary failure secondary to aspirated grape particles and postobstructive pulmonary edema. INTERVENTIONS: The patient was emergently transferred to the regional ECMO center and placed on venoarterial ECMO. Bronchoscopies were performed in the stable environment provided by ECMO, aspirated particles were removed from the large airways, and lung recovery was facilitated. MEASUREMENTS AND MAIN RESULTS: End-organ perfusion was restored via ECMO during a period of severe intractable cardiopulmonary failure. Pulmonary recovery occurred during a 6-day ECMO run and was facilitated by therapeutic bronchoscopy. The patient was reviewed 1 yr later and has made a full neurodevelopmental recovery, despite a 15-min out-of-hospital cardiac arrest. CONCLUSIONS: Aspiration of a seedless grape is a life-threatening event in a small child. This danger is not fully appreciated by parents in the UK. ECMO may be life saving in cases of acquired large-airway disruption resulting in severe cardiopulmonary failure, including foreign body aspiration, as long as end-organ perfusion is maintained.
Assuntos
Oxigenação por Membrana Extracorpórea , Pneumonia Aspirativa/etiologia , Baixo Débito Cardíaco/etiologia , Pré-Escolar , Humanos , Lactente , Masculino , Pneumonia Aspirativa/terapia , Resultado do TratamentoRESUMO
Activation of T cells by Ag or stimulation of monocytes with inflammatory cytokines induces CD44 to bind to hyaluronan (HA), an adhesion event implicated in leukocyte-leukocyte, leukocyte-endothelial cell, and leukocyte-stromal cell interactions. We have previously shown that TNF-alpha induces CD44 sulfation in a leukemic cell line, which correlated with the induction of HA binding and CD44-mediated adhesion. In this study, we establish that TNF-alpha and IFN-gamma induce HA binding and the sulfation of CD44 in CD14(+) PBMC, whereas no induced HA binding or CD44 sulfation was observed in CD14(-) PBMC stimulated with TNF-alpha. Treatment of cells with NaClO(3), an inhibitor of sulfation, prevented HA binding in a significant percentage of CD14(+) PBMC induced by TNF-alpha, LPS, IL-1beta, or IFN-gamma. Furthermore, stimulation with TNF-alpha or IFN-gamma in the presence of NaClO(3) reduced the ability of isolated CD44H to bind HA, demonstrating a direct effect of CD44H sulfation on HA binding. In contrast, the transient induction of HA binding in T cells by PHA was not affected by NaClO(3), suggesting that activated T cells do not use sulfation as a mechanism to regulate HA binding. Overall, these results demonstrate that inducible sulfation of CD44H is one mechanism used by CD14(+) peripheral blood monocytes to induce HA binding in response to inflammatory agents such as TNF-alpha and IFN-gamma.