Development, Validation, and Utility of Species-Specific Diagnostic Markers for Detection of Peronospora belbahrii.

Peronospora belbahrii is an oomycete and the cause of basil downy mildew, one of the most destructive diseases affecting basil production worldwide. Disease management is challenging due to wind-dispersed sporangia and contaminated seed; therefore, identifying P. belbahrii in seed lots before sale or planting or in the field before symptoms develop could allow for timely deployment of disease management strategies. In this study, a draft genome assembly and next-generation sequencing reads for P. belbahrii, as well as publicly available DNA-seq and RNA-seq reads of several other downy mildew pathogens, were incorporated into a bioinformatics pipeline to predict P. belbahrii-specific diagnostic markers. The specificity of each candidate marker was validated against a diverse DNA collection of P. belbahrii, host tissue, and related oomycetes using PCR. Two species-specific markers were identified and used as templates to develop a highly sensitive probe-based real-time quantitative PCR (qPCR) assay that could detect P. belbahrii in leaf tissue and seed samples. Both markers were capable of reliably detecting as low as 500 fg/µl of P. belbahrii genomic DNA and as few as 10 sporangia. The qPCR assay was then validated with seed samples collected from a basil cultivar experiment. In total, 48 seed samples were collected and tested; P. belbahrii was detected in samples of all cultivars at estimated concentrations of 600 fg/µl up to 250 pg/µl and at as few as 10 sporangia up to >1,000 sporangia. The markers and assays are valuable for diagnostics and identifying P. belbahrii-contaminated seed lots to mitigate the effects of future basil downy mildew epidemics.

Genome Sequencing and Transcriptome Analysis of the Hop Downy Mildew Pathogen Pseudoperonospora humuli Reveal Species-Specific Genes for Molecular Detection.

Pseudoperonospora humuli is an obligate oomycete pathogen of hop (Humulus lupulus) that causes downy mildew, an important disease in most production regions in the Northern Hemisphere. The pathogen can cause a systemic infection in hop, overwinter in the root system, and infect propagation material. Substantial yield loss may occur owing to P. humuli infection of strobiles (seed cones), shoots, and cone-bearing branches. Fungicide application and cultural practices are the primary methods to manage hop downy mildew. However, effective, sustainable, and cost-effective management of downy mildew can be improved by developing early detection systems to inform on disease risk and timely fungicide application. However, no species-specific diagnostic assays or genomic resources are available for P. humuli. The genome of the P. humuli OR502AA isolate was partially sequenced using Illumina technology and assembled with ABySS. The assembly had a minimum scaffold length of 500 bp and an N50 (median scaffold length of the assembled genome) of 19.2 kbp. A total number of 18,656 genes were identified using MAKER standard gene predictions. Additionally, transcriptome assemblies were generated using RNA-seq and Trinity for seven additional P. humuli isolates. Bioinformatics analyses of next generation sequencing reads of P. humuli and P. cubensis (a closely related sister species) identified 242 candidate species-specific P. humuli genes that could be used as diagnostic molecular markers. These candidate genes were validated using polymerase chain reaction against a diverse collection of isolates from P. humuli, P. cubensis, and other oomycetes. Overall, four diagnostic markers were found to be uniquely present in P. humuli. These candidate markers identified through comparative genomics can be used for pathogen diagnostics in propagation material, such as rhizomes and vegetative cuttings, or adapted for biosurveillance of airborne sporangia, an important source of inoculum in hop downy mildew epidemics.

Interleukin-1α is the major alarmin of lung epithelial cells released during photodynamic therapy to induce inflammatory mediators in fibroblasts.

BACKGROUND: Photodynamic therapy (PDT) causes tissue damage that initiates a local inflammatory response. Post-PDT reactions are considered to assist in mobilising the immune system thereby affecting tumour recurrence. The initiating process of the PDT-dependent tissue reaction remains to be determined. METHODS: Primary cultures of human lung cells were established. The photoreaction mediated by pyropheophorbide-a, at specific subcellular sites and levels resulting in the release of alarmins by epithelial cells (Eps), was defined by immunoblot analyses and expression profiling. The activity of Ep-derived factors to stimulate expression of proinflammatory mediators, including IL-6, and to enhance neutrophil binding by fibroblasts (Fbs) was determined by functional bioassays. RESULTS: Epithelial cells release IL-1ß as the primary Fb-stimulatory activity under basal conditions. Intracellular IL-1α, externalised following photoreaction, accounts for most of the PDT-mediated Fb activation. Expression of IL-1 is subject to increase or loss during oncogenic transformation resulting in altered alarmin functions mobilisable by PDT. Photoreaction by a cell surface-bound photosensitiser (PS) is 10-fold more effective than PSs localised to mitochondria or lysosomes. High-dose intracellular, but not cell surface, photoreaction inactivates IL-1 and reduces Fb stimulation. CONCLUSION: These in vitro data suggest that the subcellular site and intensity of photoreaction influence the magnitude of the stromal cell response to the local damage and, in part, support the relationship of PDT dose and level of post-PDT inflammatory response observed in vivo.

Two new methods of improving optical image quality.

Two simple methods are presented by which the coherence between a laser light source and an illuminated object can be destroyed in optical systems where this coherence is undesirable. Both methods employ rotating phase changing disks; the first introduces a random time varying phase relationship across the area illuminated in the object plane, whereas the second produces a rapid, regular phase variation across this area. Results are claimed to be comparable to those obtained using normal incoherent light.

The participation of women in education in the third world.

PIP: In this descriptive and exploratory discussion of the participation of women in education in the 3rd world, focus is only on participation in schools. The index used is years of schooling for each sex. It provides measures of utilization. The question is how far do girls go in school, compared with boys, and what do they study. Attention is directed to the following: participation versus access; literacy and primary schooling across the generations; enrollment rates and wastage around the world (overview of enrollment rates, wastage and promotion and retardation, early marriage and schooling, ambiguities of coeducation, women's schooling in Muslim and in Latin American countries); intracountry variations in schooling of girls (spatial diffusions of schooling, sex and social selection for schooling, and the assessment of progress). The availability of educational options does not ensure their utilization, and in the less developed countries (LDCs) this distinction between provision and utilization is basic for policy. Whether schooling of a daughter is considered valuable will be influenced by perceptions of the effects of schooling on jobs, on acquisition of a "better" husband, on quality of domestic life, on the daughter's personality development, and on the well-being of her children. How girls perform in school compared with boys is affected by the same factors determining initial access. The situation regarding differences in literacy and primary schooling between men and women is presented in tables to illustrate 4 distinctive patterns of change. Sex differentials in schooling among children 6-11 are negligible in European countries and in Latin America, although the rates in Latin America are lower. In these regions only small differentials occur for ages 12-17, and sex contrasts continue to be moderate at ages 18-23. In the 3rd world the situation is different. In Asian countries (excluding Japan), the rates for 6-11 year olds are 71 and 50%, respectively, and for Africa 59 and 43%; at ages 12-17, in Asia 83 and 22%, and in Africa 39 and 24%. Everywhere outside the most developed regions intercountry variations in enrollment rates are very large for both sexes, as are disparities in those rates. A country can rank quite differently at each level of school for the proportion of girls, and countries vary greatly in female shares of pupils. In countries where girls are married at ages 15-19, their enrollments at those ages are lower, but there is no simple trade-off between marriage and schooling. It appears that there are common causes for both early marriage and low school attendance. In all aspects of girls' schooling, the availability of women teachers is salient as both an instrument and a product. The proportion of girls who are in school can vary more among provinces than among countries taken as a whole. The factors affecting educational opportunities are numerous and cross cutting. Possibly the firmest generalization regarding social selectivity in the education of girls is that socioeconomic status of parents has more influence on the schooling of girls than of boys. This influence frequently is greater in rural localities or among disadvantaged ethnic groups than it is in favored segments of the population. Probit and logit models are becoming widely used in many different kinds of studies of educational participation.^ieng

A clinic within a general hospital for the assessment of urgent psychiatric problems.

Of 100 consecutive referrals to a clinic for urgent psychiatric assessment only a third (34 patients) were considered to warrant such urgent attention. In 29 of the 34 cases the patient's behaviour--either intent to harm himself or unpredictable behaviour that alarmed the referring agent--was the most important consideration for urgent assessment.

Thyroid hormone-induced precocious growth and maturation of the embryonic chicken liver.

The effects of thyroid hormones thyroxine (T4) or triiodothyronine (T3) on the ontogeny of chicken embryonic liver were studied. Two micrograms of T4 administered to chicken embryos, prior to day 11 of incubation, was found to be least toxic and effective in increasing liver weights, total protein and DNA and RNA, over those of controls. A non-toxic dose of T4 (0.1 microgram) had no effect on embryonic chicken liver. Injection of 125I-labelled T4 or T3 into chick embryos showed that T4 was taken up in greater amounts by the liver than was T3. Uptake of both hormones by the liver increased dramatically around day 9 of incubation. Induction of hypothyroidism by methimazole (a goitrogen) suppressed the natural increase in liver weight.

Utility of blood cultures in pediatric patients found to have pneumonia in the emergency department.

STUDY OBJECTIVE: To determine the prevalence of bacteremia in pediatric patients with radiographic evidence of pneumonia in whom blood cultures were obtained. METHODS: We carried out a retrospective review of the radiology log of a tertiary care children's hospital to identify patients with radiographic evidence of pneumonia seen between August 1991 and July 1992. These patients were cross-referenced with the hospital laboratory information system, yielding results of any CBC or blood cultures. RESULTS: We found 939 patients with chest radiography findings consistent with pneumonia. Blood culturing was performed in 409 (44%). Eleven of these cultures (2.7%) grew pathogenic bacteria. Review of the medical records revealed no changes in clinical management made on the basis of the results of the blood cultures. CONCLUSION: Blood cultures are uncommonly positive in outpatients diagnosed with pneumonia.

Babywalker-related injuries continue despite warning labels and public education.

OBJECTIVE: To describe the epidemiology of babywalker-related injuries to children treated in a pediatric emergency department despite current prevention efforts, and to investigate the beliefs of parents regarding babywalker use. DESIGN: A descriptive study of a consecutive series of patients. SETTING: The emergency department of a large, academic children's hospital. PARTICIPANTS: Children treated for babywalker-related injuries during the 3-year period of March 1993 through February 1996. RESULTS: There were 271 children treated for babywalker-related injuries. The mean age was 9.2 months, and 62% of patients were boys. Ninety-six percent of children were injured when they fell down stairs in their babywalker. The number of stairs that a child fell down was significantly associated with skull fracture and admission to the hospital, and a fall down more than 10 stairs had a relative risk (RR) of skull fracture = 3.28 (95% confidence interval, 1.35 < RR < 7.98). There were 159 children with contusions/abrasions (58.6%), 35 concussions/head injuries (12. 9%), 33 lacerations (12.2%), 26 skull fractures (9.6%), 9 epistaxis (3.3%), 4 nonskull fractures (1.5%), 4 avulsed teeth (1.5%), and 1 burn (0.4%). Three of the skull fractures were depressed, and three also had accompanying intracranial hemorrhage. Ten patients (3.7%) were admitted to the hospital, and all had skull fractures resulting from falls down stairs. Supervision was present in 78% of cases, including supervision by an adult in 69% of cases. Forty-five percent of families kept the walker, and 32% used the walker again for the study patient or another child after the injury episode. Fifty-nine percent of parents acknowledged that they were aware of the potential dangers of babywalkers before the injury event. Fifty-six percent of parents favored a national ban on the sale of walkers, and 20% were opposed. CONCLUSION: Despite the currently used prevention strategies, including adult supervision, warning labels, care giver education programs, and stairway gates, serious injuries associated with babywalkers continue to occur to young children. The US Consumer Product Safety Commission should promulgate a rule, similar to the voluntary standard adopted in Canada, regarding design requirements for babywalkers that will prevent their passage through household doorways at the head of stairs. The manufacture and sale of mobile babywalkers that do not meet this new standard should be banned in the US. A recall or trade-in campaign should be conducted nationally to decrease the number of existing babywalkers.

Age disorientation in schizophrenia: a constant prevalence of 25 per cent in a chronic mental hospital population?

The prevalence of age disorientation was estimated in the population of patients with a diagnosis of schizophrenia in a large mental hospital. Of these 357 patients 25 per cent demonstrated age disorientation, defined as a five-year discrepancy between true and subjective age, and 11 per cent of the population believed themselves to be within five years of the age they were at admission, although they were a mean 28.9 years older. Age-disorientated patients differed from the age-orientated in being significantly older. However, when age-matched, they were younger at first admission and had a longer duration of stay than patients with a diagnosis of schizophrenia without age-disorientation. Age-disorientation may thus be a feature of a type of schizophrenic illness of early onset and poor prognosis.

Pediatric head injuries: can clinical factors reliably predict an abnormality on computed tomography?

STUDY OBJECTIVE: To assess clinical features that might reliably predict the need for computed tomography (CT) imaging in pediatric head trauma. DESIGN/SETTING/TYPE OF PARTICIPANT: Prospective cohort of 324 head CT scans performed on 322 consecutive trauma patients at an urban children's hospital. RESULTS: Sixty-two percent of patients were male. The mean age was 7.1 years (10 days to 20.6 years); half were less than 5 years of age. The two most frequent mechanisms of injury were falls (32%) and motor vehicle accidents (25%). Abnormalities were detected in 74 scans. Intracranial injuries were apparent in 39 patients (12%); 16 had a concomitant fracture. An isolated cranial abnormality was observed on 35 scans (11%). Loss of consciousness, amnesia for the event, a Glasgow Coma Scale (GCS) of less than 15, and the presence of a neurologic deficit were more common in children with intracranial injury (P < .05). Vomiting, seizures, and headache were not discriminating clinical features. No single characteristic consistently identified the children with an intracranial injury. Of the 195 children who were neurologically intact (GCS, 15) at the time of presentation, 11 (5%) had evidence of intracranial pathology on CT scan. CONCLUSION: This study demonstrates a poor correlation between the clinical symptoms of significant traumatic brain injury and findings on CT.

Pediatric cervical-spine immobilization: achieving neutral position?

This study was designed to evaluate prospectively the ability of current spine-immobilization devices to achieve radiographic-neutral positioning of the cervical spine in pediatric trauma patients. All trauma patients who required spinal immobilization and a lateral cervical spine radiograph were included in the study. A lateral cervical spine radiograph was obtained while the child was immobilized. The Cobb angle (C2-C6) was measured using a handheld goniometer. The method of immobilization, age at injury, and Cobb angle were compared. One hundred and eighteen patients with an average age of 7.9 years were enrolled. The majority were males (71%). The most frequent mechanisms of injury included motor vehicle accidents (35%) and falls (32%). The average Glascow Coma Scale score was 14. Although 31% of the children complained of neck pain, 92% were without neurologic deficits. The Cobb angles ranged from 27 degree kyphosis to 27 degree lordosis, and only 12 of the patients presented in a neutral position (0 degrees). Greater than 5 degrees of kyphosis or lordosis was observed in 60% of the children. Thirty-seven percent of the patients had 10 degrees or greater angulation. The most frequent methods of immobilization included a collar, backboard, and towels (40%), and a collar, backboard, and blocks (20%), but these techniques provided < 5 degrees kyphosis or lordosis in only 45% and 26% of the children respectively. No single method or combination of methods of immobilization consistently placed the children in the neutral position.(ABSTRACT TRUNCATED AT 250 WORDS)

Accuracy of visual determination of neutral position of the immobilized pediatric cervical spine.

BACKGROUND: The definition of neutral position for the immobilized pediatric cervical spine is not well standardized. In this study, we attempted to determine whether 1) physicians and/or paramedics could accurately assess visually if the cervical spine was in a neutral position, 2) the visual assessments of the observers were in agreement, and 3) a radiographic Cobb angle would correlate with the visual determination. METHODS: Children presenting to a pediatric emergency department (ED) in full spinal immobilization were randomly selected (convenience sample) for this prospective study. The emergency physician and transporting paramedic independently determined positioning of the cervical spine. A radiologist, blinded to clinical information, determined Cobb angles from radiographs of the immobilized cervical spines. RESULTS: Of the 59 children studied, the evaluation of cervical spine position by the physician and paramedic correlated in 88% of the cases. For the 22 children with non-neutral Cobb angles (definition of neutral: between 5 degrees flexion and 5 degrees extension), observers agreed in 100% of the cases. However, in 21 of these cases (95%) the position was observed as neutral. CONCLUSIONS: Although visual determinations of neutral position of the cervical spine by two observers may correlate, radiographic studies demonstrate that neutral position was not achieved in 37% of the cases.