Characterization and comparison of commercial oils used for human embryo culture.

STUDY QUESTION: Are there significant differences between the available commercial oil brands used for human IVF? SUMMARY ANSWER: Important differences have been detected among the tested oil brands in their potential to stabilize culture conditions and, more importantly, in their direct effect on embryo development and viability. WHAT IS KNOWN ALREADY: Mineral oil is a critical component of the human culture system due to its protective and stabilizing roles during in vitro embryo development. Many different oils are available on the market, with differences in their viscosity, density and overall quality. STUDY DESIGN, SIZE, DURATION: Thirteen different commercial oil brands were compared. PARTICIPANTS/MATERIALS, SETTING, METHODS: Each oil was firstly analyzed to assess its viscosity, density, peroxide value and potential oxidation. Secondly, the capacity of each oil to reduce pH, osmolality and temperature fluctuations during embryo culture and manipulation was compared. Lastly, a sensitive mouse embryo assay (MEA) protocol, previously optimized to detect toxicity in oils samples, was used to compare the overall quality of the different brands in terms of embryo developmental rates up to the blastocyst stage. At the end of the MEAs, a triple labeling protocol was applied to analyze Oct4+ cells, apoptotic cells and total cell counts in the blastocysts obtained by fluorescence microscopy. MAIN RESULTS AND THE ROLE OF CHANCE: Significant divergences were detected in the rise of osmolality and the equilibration and stability of pH between different oils, which could be correlated to their physico-chemical characteristics. In particular, oil samples with a higher viscosity tended to offer an additional protection against fluctuations in the culture conditions, however, the differences in temperature stability between oils were minor. Two out of the 13 oil samples, which were commercially available, were identified as embryo-toxic by applying the MEA protocol with increased sensitivity for toxicity detection. Additionally, substantial differences in the total number of cells and the number of cells in the inner cell mass of the obtained blastocysts were also detected between oil groups. LIMITATIONS, REASONS FOR CAUTION: A single lot of oil was used for each brand and, thus, lot-to-lot variations in oil quality could not be determined. However, several bottles from the same oil were included to account for potential intra-lot variability. WIDER IMPLICATIONS OF THE FINDINGS: Commercial oils differ in both their physical characteristics and their performance in maintaining the stability of the culture conditions during in vitro embryo culture. Oil selection is important for embryo culture success. Additionally, the detection of embryo-toxic oils which had already been released to the human IVF market showcases the importance of applying sensitive MEA protocols for a better detection of toxicity in this type of samples. STUDY FUNDING/COMPETING INTEREST(S): This study was privately funded. TRIAL REGISTRATION NUMBER: N/A.

Factors of the human embryo culture system that may affect media evaporation and osmolality.

STUDY QUESTION: Which lab-related factors impact the culture system's capacity to maintain a stable osmolality during human embryo culture? SUMMARY ANSWER: Incubator humidity, the volume of mineral oil, the type of culture media and the design of time-lapse dishes have been identified as important parameters that can cause an impact on media evaporation and consequently osmolality during culture. WHAT IS KNOWN ALREADY: Culture medium is a critical component in human embryo culture. Minimizing its evaporation during culture is an adequate strategy to stabilize osmolality and, as a result, improving culture conditions and clinical outcomes. STUDY DESIGN, SIZE, DURATION: The studied variables included media composition and supplementation; volume of mineral oil; incubator humidification; and the type of dish and incubator used. Additionally, six time-lapse dish models were compared in their ability to prevent evaporation. PARTICIPANTS/MATERIALS, SETTING, METHODS: Dishes were incubated in parallel to analyze osmolality during culture between groups: synthetic oviductal medium enriched with potassium versus human tubal fluid medium; protein versus no protein supplementation; dry versus humid atmosphere; high versus low volume of mineral oil. Additionally, media evaporation was compared between six models of time-lapse dishes with distinct designs, cultured in a joint incubator. Two of them were retested in their corresponding incubator to analyze the dish-incubator fit. Daily osmolality measurements were compared between groups. Linear regression was performed to analyze evaporation rates. MAIN RESULTS AND THE ROLE OF CHANCE: Protein supplementation did not significantly affect evaporation. Contrarily, humidity levels inside the incubators, the volume of mineral oil and the type of culture media, played an important role in osmolality stabilization. The design of time-lapse dishes and their recommended preparation protocol heavily influenced their evaporation rates, which were further altered by each incubator's characteristics. Media with initially high osmolalities had a bigger risk of reaching hypertonic levels during culture. LIMITATIONS, REASONS FOR CAUTION: While numerous, the studied variables are limited and therefore other factors could play a role in osmolality dynamics, as well. Incontrollable atmospheric factors could also result in some variation in the observed results between different centers and laboratories. WIDER IMPLICATIONS OF THE FINDINGS: Published literature has extensively described how hypertonic media may impair embryo development and negatively affect clinical outcomes; therefore, maintaining a stable osmolality during culture should be considered essential. This work is of interest both for embryologists when analyzing their culture system and methodologies, as well as manufacturers in charge of designing IVF consumables. STUDY FUNDING/COMPETING INTEREST(S): This study was privately funded. TRIAL REGISTRATION NUMBER: N/A.

Temporal dynamics of Middle East respiratory syndrome coronavirus in the Arabian Peninsula, 2012-2017.

Middle East respiratory syndrome coronavirus (MERS-CoV) remains a notable disease and poses a significant threat to global public health. The Arabian Peninsula is considered a major global epicentre for the disease and the virus has crossed regional and continental boundaries since 2012. In this study, we focused on exploring the temporal dynamics of MERS-CoV in human populations in the Arabian Peninsula between 2012 and 2017, using publicly available data on case counts and combining two analytical methods. Disease progression was assessed by quantifying the time-dependent reproductive number (TD-Rs), while case series temporal pattern was modelled using the AutoRegressive Integrated Moving Average (ARIMA). We accounted for geographical variability between three major affected regions in Saudi Arabia including Eastern Province, Riyadh and Makkah. In Saudi Arabia, the epidemic size was large with TD-Rs >1, indicating significant spread until 2017. In both Makkah and Riyadh regions, the epidemic progression reached its peak in April 2014 (TD-Rs > 7), during the highest incidence period of MERS-CoV cases. In Eastern Province, one unique super-spreading event (TD-R > 10) was identified in May 2013, which comprised of the most notable cases of human-to-human transmission. Best-fitting ARIMA model inferred statistically significant biannual seasonality in Riyadh region, a region characterised by heavy seasonal camel-related activities. However, no statistical evidence of seasonality was identified in Eastern Province and Makkah. Instead, both areas were marked by an endemic pattern of cases with sporadic outbreaks. Our study suggested new insights into the epidemiology of the virus, including inferences about epidemic progression and evidence for seasonality. Despite the inherent limitations of the available data, our conclusions provide further guidance to currently implement risk-based surveillance in high-risk populations and, subsequently, improve related interventions strategies against the epidemic at country and regional levels.

Retaining or excising the supraspinatus tendon in complex proximal humeral fractures treated with reverse prosthesis: a biomechanical analysis in two different designs.

PURPOSE: We aimed to biomechanically evaluate the effect of the supraspinatus tendon on tuberosity stability using two different reverse shoulder arthroplasty (RSA) models for complex proximal humeral fractures (PHFs). METHODS: Four-part proximal humeral fractures were simulated in 20 cadaveric shoulders. Two different RSA designs were implemented: a glenosphere-medialized model and a glenosphere-lateralized model. Tuberosities were reconstructed, and displacement of bony fragments was measured (mm) by placing three sensors: in the humeral diaphysis (D), in the greater tuberosity (GT), and in the lesser tuberosity (LT). Axial forces were induced and measured in Newton (N). The test was performed twice in each specimen, with and without the supraspinatus tendon. The regression line (RL) was measured in mm/N. RESULTS: In the medialized model, the GT-D displacement was greater in the supraspinatus preserving model than that in the tendon excision model (p < 0.001), as well as for the LT-D displacement (p < 0.001). In the lateralized model, GT-D displacement and GT-LT distance were greater in the preserving model than that in the excision model (p < 0.001, p = 0.04). CONCLUSION: The supraspinatus tendon resection leads to a more biomechanically stable tuberosity construct when performing RSA for PHFs, while the rest of the rotator cuff tendons (infraspinatus and teres minor) are retained in the greater tuberosity. LEVEL OF EVIDENCE: Basic science study. Cadaveric study.

Characterization of influenza outbreaks in Lebanon during the 2013/14 and 2014/15 seasons.

Despite the significant burden of influenza outbreaks, active disease monitoring has been largely absent in the Middle East, including Lebanon. In this study we characterized influenza virus in 440 nasopharyngeal swabs collected from patients with acute respiratory infections during two influenza seasons in Lebanon. Influenza A(H3N2) was dominant in the 2013/14 season while the A(H1N1)pdm09 and B/Yamagata strains were most prevalent in the 2014/15 season. All tested isolates were susceptible to 4 neuraminidase inhibitors (oseltamivir, zanamivir, peramivir and laninamivir). Genetic analysis of the haemagglutinin gene revealed multiple introductions of influenza viruses into Lebanon from different geographic sources during each season. Additionally, large data gaps were identified in the Middle East region, as indicated by the lack of current influenza sequences in the database from many countries in the region.

Characterization of influenza outbreaks in Lebanon during the 2013/14 and 2014/15 seasons

Despite the significant burden of influenza outbreaks, active disease monitoring has been largely absent in the Middle East, including Lebanon. In this study we characterized influenza virus in 440 nasopharyngeal swabs collected from patients with acute respiratory infections during two influenza seasons in Lebanon. Influenza A[H3N2] was dominant in the 2013/14 season while the A[H1N1]pdm09 and B/Yamagata strains were most prevalent in the 2014/15 season. All tested isolates were susceptible to 4 neuraminidase inhibitors [oseltamivir, zanamivir, peramivir and laninamivir]. Genetic analysis of the haemagglutinin gene revealed multiple introductions of influenza viruses into Lebanon from different geographic sources during each season. Additionally, large data gaps were identified in the Middle East region, as indicated by the lack of current influenza sequences in the database from many countries in the region

Malgré la lourde charge que représentent les flambées de grippe, la surveillance active de la maladie était jusqu'à présent inexistante au Moyen-Orient, et notamment au Liban. Dans la présente étude, le virus de la grippe a été caractérisé dans 440 sécrétions rhinopharyngées prélevées par écouvillonnage chez des patients ayant souffert d'infections respiratoires aiguës pendant deux saisons grippales au Liban. Le virus de la grippe A[H3N2] était prédominant pendant la saison 2013/2014, tandis que celui de la grippe A[H1N1]pdm09 et les souches de grippe B/Yamagata étaient les plus courants pendant la saison 2014/2015. Tous les isolats testés étaient sensibles à quatre inhibiteurs de la neuraminidase [l'oseltamivir, le zanamivir, le peramivir, et le laninamivir]. L'analyse génétique du gène de l'hémagglutinine a révélé de multiples introductions des virus de la grippe au Liban, depuis différentes sources géographiques au cours de chaque saison. De plus, d'importantes lacunes dans les données ont été constatées dans la région du Moyen-Orient, comme le montre l'absence des séquences génétiques actuelles de la grippe dans les bases de données de nombreux pays de la region

Clinical Sentinel Surveillance of Equine West Nile Fever, Spain.

West Nile fever (WNF) is a viral zoonotic infection caused by a mosquito-borne flavivirus of the Flaviviridae family. According to a comparative study, the passive surveillance of horses by equine veterinarians appeared to be the most cost-effective system in the European context of WNF. Clinical data issued from a passive epidemiosurveillance network from September 2010 to December 2011 on horses in Spain were statistically compared and used to develop a predictive diagnostic decision tree, both with the aim to improve the early clinical detection of WNF in horses. Although clinical signs were variable in horses affected by WNF, four clinical signs and the month of occurrence were identified as useful indicators to distinguish between WNF-related and WNF-unrelated cases. The signs that pointed out a presumptive diagnosis of WNF in horses were cranial nerves deficits, limb paralysis, photophobia and nasal discharge. Clinical examination of horses with neurological signs that are not vaccinated against WNV could provide important clues for the early clinical detection of WNF and therefore serve as an alert for possible human viral infections. The study of the clinical pattern of WNF in horses is of importance to enhance awareness and better understanding and to optimize surveillance designs for clinical detection of WNF in horses in advance of epidemic activity affecting humans.

The impact of visual feedback on the motor control of the upper-limb.

Stroke is a leading cause of adult disability with upper-limb hemiparesis being one of the most frequent consequences. Given that stroke only affects the paretic arm's control structure (the set of synergies and activation vectors needed to perform a movement), we propose that the control structure of the non-affected arm can serve as a physiological reference to rehabilitate the paretic arm. However, it is unclear how rehabilitation can effectively tune the control structure of a patient. The use of Visual Feedback (VF) is recommended to boost stroke rehabilitation, as it is able to positively modify neural mechanisms and improve motor performance. Thus, in this study we investigate whether VF can effectively modify the control structure of the upper-limb. We asked six neurologically intact subjects to perform a complete upper-limb rehabilitation routine comprised of 12 movements in absence and presence of VF. Our results indicate that VF significantly increases interlimb similarity both in terms of synergies and activation coefficients. However, the magnitude of improvement depended upon each subject. In general, VF brings the control structure of the nondominant side closer to the control structure of dominant side, suggesting that VF modifies the control structure towards more optimized motor patterns. This is especially interesting because stroke mainly affects the activation coefficients of patients and because it has been shown that the control of the affected side resembles that of the nondominant side. In conclusion, VF may enhance motor performance by effectively tuning the control-structure. Notably, this finding offers new insights to design improved stroke rehabilitation.

Synergy analysis as a tool to design and assess an effective stroke rehabilitation.

The poor rehabilitation success rate, including the cases of ineffective and detrimental adaptations, make stroke a leading cause of disability. Thus, it is essential to recognize the mechanisms driving healthy motor recovery to improve such rate. Stroke alters the Synergy Architecture (SA), the modular muscle control system. So SA analysis may constitute a powerful tool to design and assess rehabilitation procedures. However, current impairment scales do not consider the patient's neuromuscular state. To gain insights into this hypothesis, we recorded multiple myoelectric signals from upper-limb muscles, in healthy subjects, while executing a set of common rehabilitation exercises. We found that SA reveals optimized motor control strategies and the positive effects of the use of visual feedback (VF) on motor control. Furthermore we demonstrate that the right and left arm's SA share the basic structure within the same subject, so we propose using the unaffected limb's SA as a reference motion pattern to be reached through rehabilitation.

Deregulation of tissue homeostasis in endometria from patients with polycystic ovarian syndrome with and without endometrial hyperplasia.

OBJECTIVE: To study the proteins involved in endometrial homeostasis in PCOS women. METHODS: Protein expression of Ki67, Bcl-2, Bax, Pro-Caspase-3 and Caspase-3 by immunohistochemistry and/or Western blot, and DNA fragmentation using in situ 3'-end labeling of apoptotic cells, was measured in 9 samples of normal endometrium (NE), 12 PCOS endometria without treatment (PCOSE), 7 endometria from PCOS women with endometrial hyperplasia (HPCOSE) and 9 endometria from patients with endometrial hyperplasia (HE). RESULTS: Cell proliferation was higher in epithelium from PCOSE (P<0.05), HPCOSE and HE vs NE. A higher Bcl-2/Bax relative ratio in PCOSE and HPCOSE was observed, in absence of active Caspase-3 and scarce DNA fragmentation in the four groups of endometria studied. CONCLUSION: As the apoptosis was scarce in all of the groups studied, endometrial homeostasis deregulation in PCOS could be a result of increased proliferation. Therefore, the onset of endometrial hyperplasia in PCOS endometrium could be linked to inadequate cell proliferation, and concomitantly to inadequate cell survival.

Construcción de un cuestionario de valoración de la salud social de los usuarios en el ámbito sociosanitario / Creating a social health evaluation questionnaire for users in the socio-sanitary area

Introducción: puede definirse salud social como la situación óptima en cada una de las áreas que configuran el entorno social de la persona. Este trabajo presenta los resultados preliminares de la creación del Qüestionari de Valoració de Salut Social (QVSS), que pretende valorar el estado de salud social de los usuarios del ámbito sociosanitario. Material y métodos: partiendo del análisis de contenido de instrumentos conocidos, el equipo de trabajo elaboró los ámbitos que consideraba necesarios para valorar la salud social, determinó las posibles situaciones en que podía encontrarse un individuo dentro de cada ámbito y formuló las preguntas necesarias para poder describirlas. Posteriormente, en una reunión con un grupo de expertos, se consensuaron los ámbitos y preguntas definitivas. Resultados: los ámbitos, que se corresponden a subescalas independientes del QVSS, fueron: soporte informal, soporte formal, situación económica, vivienda e integración familiar y social. El sexo (p < 0,001), los estudios (p = 0,017) y la clase social (p = 0,029) se asocian con la subescala económica. También el estado civil (p = 0,002), que además se asocia con el soporte informal (p = 0,030). El tiempo medio de respuesta fue de 13,5 min. Conclusiones: el QVSS mostró facilidad en su administración. Parece que no existen grandes diferencias en función de quién lo responda, excepto en la subescala de soporte informal. Se hace necesario validarlo y ahondar en el estudio sobre la relación entre el QVSS respondido por el paciente o por el cuidador

Introduction: social health can be defined as the best possible situation for each of the areas that make up a person's social environment. This article presents the preliminary results of the creation of a Social Health Evaluation Questionnaire (SHEV), aimed at assessing the social health status of users of hospitals for the elderly and disabled. Material and methods: the contents of well-known instruments were analysed and the working team selected the areas considered necessary to be able to evaluate social health status. The team then determined the possible situations in which individuals could find themselves in each of these areas and created questions to help them describe these situations. Subsequently, in a meeting with a group of experts, an overall agreement was reached on the scope and content of the final questions to be included in the questionnaire. Results: the distinct areas, which correspond to independent subscales in the SHEV, were: informal support, formal support, economic situation, housing, and social and family integration. Sex (p < 0.001), educational status (p = 0.017) and social class (p = 0.029) were associated with economic situation. Marital status was associated with economic situation (p = 0.002) and informal support (p = 0.030). The mean answering time was 13.5 minutes. Conclusions: the SHEV proved to be an easy tool to administer. No significant differences based upon responders were found, except in the subscale of informal support. Further validation and a more thorough investigation of the relationship between the SHEV as answered by patients or by their carers is required

Infección por virus de inmunodeficiencia humana en la embarazada. Importancia del conocimiento de la infección en el embarazo y factores de riesgo en la transmisión perinatal / Human immunodeficiency virus infection in pregnant women. The importance of recognizing the infection during pregnancy and risk factors for perinatal transmission

BACKGROUND: Using adequate infection control measures, the rate of vertical transmission of human immunodeficiency virus (HIV) during pregnancy, has been reduced to 3 per cent in Chile. AIM: To determine vertical transmission rate and risk factors associated to perinatal infection in pregnant women with known (KI) and unknown HIV infection (UI). PATIENTS AND METHODS: HIV infected pregnant women whose deliveries were attended at the San Borja Arriaran Hospital were included. Antiretroviral therapy (ART) has been used since 1995 (Zidovudine 13 patients, biOtherapy 4 and triple therapy 14 patients). Newborns have received ART since 1995. Premature labor without evident cause, premature rupture of membranes, and rupture of membranes over 4 h before delivery were evaluated. Delivery was by elective cesarean section since 1993. Breast feeding was avoided. Pregnant women with UI (suspected disease after delivery due to child or mother pathology) did not received ART. Delivery and breast feeding were managed with common obstetrical-neonatal criteria. RESULTS: Fifty three HIV infected pregnant women were studied (43 with KI and 10 with UI). Four children (36.4 per cent) from the KI group and seven (63.6 per cent from the UI group became infected. The global rate of vertical transmission among KI group was significantly lower than UI group: 9.5 per cent (4/42) vs 70.0 per cent(7/10) p < 0.001. Using ART, this rate was further reduced to 6.5 per cent (2/31) and with bitherapy or triple therapy to 0 per cent (0/18). Breast feeding, vaginal delivery, premature delivery with no clinical cause, premature rupture of membranes, rupture of membranes longer than 4 h and lack of ART, were significantly more common in the UI group, compared with KI group. CONCLUSIONS: Vertical transmission in pregnant women with KI is significantly lower compared with UI. Risk factors increasing HIV perinatal infection are: breast feeding, lack of ART, vaginal delivery, premature rupture of membranes, rupture of membranes > 4 h and premature labor without a clinical cause.

Hepatitis C: es hora de actuar / Hepatitis C: it´s time to act

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Occult cancer in patients with acute pulmonary embolism. A prospective study.

We prospectively studied 78 consecutive patients with acute pulmonary embolism (PE) to determine the most appropriate workup study for searching for hidden cancer. After a careful physical examination, the following tests were performed: erythrocyte sedimentation rate (ESR), complete blood cell counts, biochemistry, carcinoembryonic antigen levels, chest radiograph, upper gastrointestinal endoscopy, and abdominal ultrasound. If a malignant lesion was suspected, further appropriate tests were performed. After hospital discharge, periodic follow-up was performed on all patients in our outpatient clinic. A malignant lesion was detected in 9 of 78 patients: in 7 of them, cancer was diagnosed during the hospital admission because of acute PE. All but one of these 7 patients were asymptomatic, except for PE symptoms. In three of them some abnormalities on physical examination led to the diagnosis of cancer; in the remaining three patients the diagnosis was suspected from abnormal results of blood tests. Cancer was detected several months after hospital discharge in two additional patients: an esophageal cancer was diagnosed 5 months later in one of the 23 patients who refused endoscopy; and a colonic carcinoma was detected 21 months after hospital discharge in a patient in whom colonoscopy was not performed at the time of hospital admission. When considered overall, cancer was more commonly found in patients with "idiopathic" PE as compared with patients with known risk factors for PE development (6 of 21 patients vs 3 of 51 patients; p < 0.05). On the other hand, one patient died because of massive recurrent PE after a biopsy sample was obtained because of a prostatic node. Gross hematuria had developed shortly after biopsy, and any attempt to increase heparin doses was followed by recurrent hematuria. According to our experience, any decision about procedures that potentially involve bleeding should be carefully individualized in patients with acute PE.

[Clinical and microbiological study of acute pelvic inflammatory disease]. / Estudio clínico y microbiológico de la enfermedad inflamatoria pélvica aguda.

Upper genital tract infection was investigated in 46 women admitted to hospital with clinic diagnosis of acute pelvic inflammatory disease (PID) and 62 control women accepted to hospital for laparoscopy Fallopian tubes sterilization. Diagnosis was ratified by laparoscopy in mild and moderate salpingitis; culdocentesis and ultrasonography were performed in severe salpingitis and endometrial sample was made in endometritis. Microbiological specimens were taken from the cervix and abdomen. Antecedents and complete clinical studies were obtained. Patients were treated with antibiotic association sodic G penicillin, chloramphenicol and gentamicin. Risk factors to development PID were: single female (p < 0.05), multiple sexual partner (p < 0.01), previous PID (p < 0.05), infertility (p < 0.05), mean year of IUD use in severe salpingitis (p = 0.05) and mean years of age from women with sexually transmitted bacterias (STB) vs endogenous bacterias (EB) (p < 0.05). In the control group no abdomen bacterias were isolated. In patients with PID, C. trachomatis was detected by serology in 28.3%. N. gonorrhoeae was isolated from the cervix in 23.9% and from the abdomen 17.4%. Besides it was isolated from the abdomen: M. hominis 17.3% and E. coli 15.2%. STB were isolated in 54.3% and EB in 47.8% of the patients. Bacterial association was present on the 37%. Cervix isolation of G. vaginalis and Mycoplasma were not correlated with development of PID. Cervix microbiological samples were useful to know abdomen microbic etiology. They coincide with those in the 90.9%. EB were more frequently isolated from severe salpingitis (p = 0.05) and STB from mild and moderate salpingitis (p = 0.05). Antibiotic association cured all the mild and moderate salpingitis with independence of bacterial etiology. Failure occurred in 2 diffuse peritonitis and 13/14 tubo-ovarian abscesses. Surgery used in severe salpingitis and diffuse peritonitis, principally consisted in anexectomy, peritoneal toilet and drainages. No hysterectomies were performed. Colpotomy drainage was used as a laparotomy complement or as unique drainage. Severe complications of surgery occurred in 10.5%. Failure in antibiotic treatment, surgery and complications were present with preference in PID with EB. After PID 26.5% of women had both Fallopian tubes damaged; in 39.7% tube damage was not evaluated and in 34.2% one tube rested in health. Damage did not depend of bacterial etiology. Conclusion on the necessity of adequate prevention of this disease and it should need education related to the roll of STB and standards about the IUD use.

Characterization of recombinant bet vI, the major pollen allergen of Betula verrucosa (white birch), produced by fed-batch fermentation.

The gene encoding the major allergen, Bet v I, from Betula verrucosa (white birch) pollen was cloned by application of the polymerase chain reaction with double-stranded cDNA as template and specific primers based on the published nucleotide sequence of the gene. The gene was inserted into plasmid pKK223-3 and expressed in Escherichia coli K-12 strain JM105 grown to high cell density in a fermenter using a fed-batch procedure. Ample material was provided for a thorough characterization of the epitope structure of recombinant Bet v I contained in an unpurified soluble lysate. The antibody-binding characteristics of recombinant Bet v I was compared with that of the natural allergen using polyclonal rabbit antibodies raised against Bet v I in crossed immunoelectrophoresis, tandem crossed immunoelectrophoresis, and Western blotting. IgE from a pool of 16 allergic patients' serum was applied in crossed radioimmunoelectrophoresis, Western blotting, and a quantitative luminescence inhibition immunoassay. Well-defined epitopes were assayed by the application of a panel of six murine monoclonal antibodies in a quantitative radio inhibition immunoassay. In all assays the activity of recombinant Bet v I was comparable to that of natural Bet v I, and it is concluded that the epitope structure of recombinant Bet v I closely resembles that of natural Bet v I. This result has important implications for the future use of recombinant tree pollen allergens as a model system for the study of allergenic B and T cell epitopes aiming at improvements in reagents used for the management of allergic disease.

Platelet count and venous thromboembolism. A useful test for suspected pulmonary embolism.

We have previously reported that patients with deep vein thrombosis (DVT) and scintigraphic evidence of pulmonary embolism (PE) had a fall in platelet count, as compared with their levels before thrombosis had developed. Otherwise, no changes were found in DVT patients without embolism. We recently conducted a prospective study with a larger series of patients and studied platelet count behavior in 189 consecutive patients with acute venous thromboembolism (VTE) in whom a baseline blood cell count was available (obtained before thromboembolism developed). We found no significant differences in baseline platelet counts between groups. However, at the time of VTE diagnosis the analysis of variance demonstrated that mean platelet count was significantly higher in patients without embolism as compared with PE patients (p less than 0.001). On the other hand, no differences were found between patients with silent PE and those with clinically obvious PE. When patients with postoperative VTE and those with nonpostoperative VTE were analyzed separately, mean platelet count increased only in postoperative DVT patients without embolism (p less than 0.001). In the absence of a previous intervention, DVT did not produce any change in platelet count, while PE significantly reduced platelet number (p less than 0.008). In DVT patients without respiratory symptoms of embolism, we suggest that a lung scan should be performed when platelet count is lower than baseline value. For patients with a higher count, the probability of finding PE is very low, and scintigraphy is not cost-effective.