Restriction of Intravenous Fluid in ICU Patients with Septic Shock.

BACKGROUND: Intravenous fluids are recommended for the treatment of patients who are in septic shock, but higher fluid volumes have been associated with harm in patients who are in the intensive care unit (ICU). METHODS: In this international, randomized trial, we assigned patients with septic shock in the ICU who had received at least 1 liter of intravenous fluid to receive restricted intravenous fluid or standard intravenous fluid therapy; patients were included if the onset of shock had been within 12 hours before screening. The primary outcome was death from any cause within 90 days after randomization. RESULTS: We enrolled 1554 patients; 770 were assigned to the restrictive-fluid group and 784 to the standard-fluid group. Primary outcome data were available for 1545 patients (99.4%). In the ICU, the restrictive-fluid group received a median of 1798 ml of intravenous fluid (interquartile range, 500 to 4366); the standard-fluid group received a median of 3811 ml (interquartile range, 1861 to 6762). At 90 days, death had occurred in 323 of 764 patients (42.3%) in the restrictive-fluid group, as compared with 329 of 781 patients (42.1%) in the standard-fluid group (adjusted absolute difference, 0.1 percentage points; 95% confidence interval [CI], -4.7 to 4.9; P = 0.96). In the ICU, serious adverse events occurred at least once in 221 of 751 patients (29.4%) in the restrictive-fluid group and in 238 of 772 patients (30.8%) in the standard-fluid group (adjusted absolute difference, -1.7 percentage points; 99% CI, -7.7 to 4.3). At 90 days after randomization, the numbers of days alive without life support and days alive and out of the hospital were similar in the two groups. CONCLUSIONS: Among adult patients with septic shock in the ICU, intravenous fluid restriction did not result in fewer deaths at 90 days than standard intravenous fluid therapy. (Funded by the Novo Nordisk Foundation and others; CLASSIC ClinicalTrials.gov number, NCT03668236.).

Deep learning nuclei detection: A simple approach can deliver state-of-the-art results.

BACKGROUND: Deep convolutional neural networks have become a widespread tool for the detection of nuclei in histopathology images. Many implementations share a basic approach that includes generation of an intermediate map indicating the presence of a nucleus center, which we refer to as PMap. Nevertheless, these implementations often still differ in several parameters, resulting in different detection qualities. METHODS: We identified several essential parameters and configured the basic PMap approach using combinations of them. We thoroughly evaluated and compared various configurations on multiple datasets with respect to detection quality, efficiency and training effort. RESULTS: Post-processing of the PMap was found to have the largest impact on detection quality. Also, two different network architectures were identified that improve either detection quality or runtime performance. The best-performing configuration yields f1-measures of 0.816 on H&E stained images of colorectal adenocarcinomas and 0.819 on Ki-67 stained images of breast tumor tissue. On average, it was fully trained in less than 15,000 iterations and processed 4.15 megapixels per second at prediction time. CONCLUSIONS: The basic PMap approach is greatly affected by certain parameters. Our evaluation provides guidance on their impact and best settings. When configured properly, this simple and efficient approach can yield equal detection quality as more complex and time-consuming state-of-the-art approaches.

A Design Study Investigating Augmented Reality and Photograph Annotation in a Digitalized Grossing Workstation.

CONTEXT: Within digital pathology, digitalization of the grossing procedure has been relatively underexplored in comparison to digitalization of pathology slides. AIMS: Our investigation focuses on the interaction design of an augmented reality gross pathology workstation and refining the interface so that information and visualizations are easily recorded and displayed in a thoughtful view. SETTINGS AND DESIGN: The work in this project occurred in two phases: the first phase focused on implementation of an augmented reality grossing workstation prototype while the second phase focused on the implementation of an incremental prototype in parallel with a deeper design study. SUBJECTS AND METHODS: Our research institute focused on an experimental and "designerly" approach to create a digital gross pathology prototype as opposed to focusing on developing a system for immediate clinical deployment. STATISTICAL ANALYSIS USED: Evaluation has not been limited to user tests and interviews, but rather key insights were uncovered through design methods such as "rapid ethnography" and "conversation with materials". RESULTS: We developed an augmented reality enhanced digital grossing station prototype to assist pathology technicians in capturing data during examination. The prototype uses a magnetically tracked scalpel to annotate planned cuts and dimensions onto photographs taken of the work surface. This article focuses on the use of qualitative design methods to evaluate and refine the prototype. Our aims were to build on the strengths of the prototype's technology, improve the ergonomics of the digital/physical workstation by considering numerous alternative design directions, and to consider the effects of digitalization on personnel and the pathology diagnostics information flow from a wider perspective. A proposed interface design allows the pathology technician to place images in relation to its orientation, annotate directly on the image, and create linked information. CONCLUSIONS: The augmented reality magnetically tracked scalpel reduces tool switching though limitations in today's augmented reality technology fall short of creating an ideal immersive workflow by requiring the use of a monitor. While this technology catches up, we recommend focusing efforts on enabling the easy creation of layered, complex reports, linking, and viewing information across systems. Reflecting upon our results, we argue for digitalization to focus not only on how to record increasing amounts of data but also how these data can be accessed in a more thoughtful way that draws upon the expertise and creativity of pathology professionals using the systems.

Influenza gain-of-function experiments: their role in vaccine virus recommendation and pandemic preparedness.

In recent years, controversy has arisen regarding the risks and benefits of certain types of gain-of-function (GOF) studies involving avian influenza viruses. In this article, we provide specific examples of how different types of data, including information garnered from GOF studies, have helped to shape the influenza vaccine production process-from selection of candidate vaccine viruses (CVVs) to the manufacture and stockpiling of safe, high-yield prepandemic vaccines for the global community. The article is not written to support a specific pro- or anti-GOF stance but rather to inform the scientific community about factors involved in vaccine virus selection and the preparation of prepandemic influenza vaccines and the impact that some GOF information has had on this process.

Development of a latex agglutination test with recombinant variant surface glycoprotein for serodiagnosis of surra.

Serodiagnosis of surra is commonly performed with the CATT/Trypanosoma evansi direct agglutination test. This antibody detection test is based on lyophilised bloodstream form trypanosomes propagated in rats and presenting the predominant variant surface glycoprotein (VSG) RoTat 1.2 on their surface. Recently, the N-terminal fragment of VSG RoTat 1.2 has been expressed as a recombinant protein in the yeast Pichia pastoris and showed diagnostic potential in ELISA. This recombinant antigen has now been incorporated in a latex agglutination test, the rLATEX/T. evansi. In this study, we compared the diagnostic accuracy of rLATEX/T. evansi and CATT/T. evansi with immune trypanolysis (TL) as reference test on a total of 1717 sera from camels, horses, bovines, water buffaloes, dogs and sheep. The rLATEX/T. evansi displayed a slightly better agreement with TL than CATT/T. evansi (kappa [κ] respectively 0.84 and 0.72). The sensitivities of rLATEX/T. evansi (84.2%, 95% CI 80.8-87.1) and CATT/T. evansi (84.0%, 95% CI 80.6-87.0) were similar, but rLATEX/T. evansi was significantly more specific (97.7%, 95% CI 96.7-98.4) than CATT/T. evansi (89.4%; 95% CI 87.6-91.1). We consider the rLATEX/T. evansi an alternative for the CATT/T. evansi, with the advantage that the use of a purified recombinant antigen leads to a more standardised diagnostic test with an improved specificity. Moreover, it eliminates the use of laboratory animals and can be easily scaled-up, e.g. in biofermentors.

Review of influenza A virus in swine worldwide: a call for increased surveillance and research.

Pigs and humans have shared influenza A viruses (IAV) since at least 1918, and many interspecies transmission events have been documented since that time. However, despite this interplay, relatively little is known regarding IAV circulating in swine around the world compared with the avian and human knowledge base. This gap in knowledge impedes our understanding of how viruses adapted to swine or man impacts the ecology and evolution of IAV as a whole and the true impact of swine IAV on human health. The pandemic H1N1 that emerged in 2009 underscored the need for greater surveillance and sharing of data on IAV in swine. In this paper, we review the current state of IAV in swine around the world, highlight the collaboration between international organizations and a network of laboratories engaged in human and animal IAV surveillance and research, and emphasize the need to increase information in high-priority regions. The need for global integration and rapid sharing of data and resources to fight IAV in swine and other animal species is apparent, but this effort requires grassroots support from governments, practicing veterinarians and the swine industry and, ultimately, requires significant increases in funding and infrastructure.

Recombinant expression of trypanosome surface glycoproteins in Pichia pastoris for the diagnosis of Trypanosoma evansi infection.

Serodiagnosis of surra, which causes vast economic losses in livestock, is still based on native antigens purified from bloodstream form Trypanosoma (T.) evansi grown in rodents. To avoid the use of laboratory rodents in antigen preparation we expressed fragments of the invariant surface glycoprotein (ISG) 75, cloned from T. brucei gambiense cDNA, and the variant surface glycoprotein (VSG) RoTat 1.2, cloned from T. evansi gDNA, recombinantly in Pichia (P.) pastoris. The M5 strain of this yeast has an engineered N-glycosylation pathway resulting in homogenous Man5GlcNAc2 N-glycosylation which resembles the predominant Man9-5GlcNAc2 oligomannose structures in T. brucei. The secreted recombinant antigens were affinity purified with yields of up to 10mg and 20mg per liter cell culture of rISG 7529-465-E and rRoTat 1.223-385-H respectively. In ELISA, both recombinant proteins discriminated between pre-immune and immune serum samples of 25 goats experimentally infected with T. evansi. The diagnostic potential of rRoTat 1.223-385-H but not of rISG 7529-465-E was confirmed with sera of naturally infected and control dromedary camels. The results suggest that rRoTat 1.223-385-H expressed in P. pastoris requires further evaluation before it could replace native RoTat 1.2 VSG for serodiagnosis of surra, thus eliminating the use of laboratory animals for antigen production.

Carpal tunnel syndrome diagnosed by general practitioners: an observational study.

The aim of this study was to investigate the results of both clinical testing and standardised nerve conduction studies performed on patients with Carpal tunnel syndrome (CTS) complaints, who had been referred to the neurologist by their general practitioners. Analysis of the data of neurological examination and electrodiagnostic tests (EDX) were performed on patients that had been referred by general practitioners. A total of 232 patients with clinically defined CTS, who had been referred by general practitioners, were seen by a neurologist and subsequently underwent electrodiagnostic testing. The diagnosis of CTS made by general practitioners was clinically confirmed by the neurologist in 187 of 232 (81%) patients. In these 187 patients, EDX confirmed CTS clinical diagnosis in 180. In 40 (17%), the neurologists disagreed with the clinical diagnosis of CTS because signs and symptoms were not those of clinical CTS. We showed that general practitioners are very well capable of making a clinical diagnosis of CTS. Therefore, direct referral of patients by general practitioners for nerve conduction studies to have their diagnosis of CTS confirmed is a desirable and time-saving procedure.

A review of trends in the distribution of vector-borne diseases: is international trade contributing to their spread?

It is difficult to determine the part that international trade has played in the expansion of vector-borne diseases, because of the multitude of factors that affect the transformation of habitats and the interfaces between vectors and hosts. The introduction of pathogens through trade in live animals or products of animal origin, as well as the arrival of arthropod vectors, is probably quite frequent but the establishment of an efficient transmission system that develops into a disease outbreak remains the exception. In this paper, based on well-documented examples, the authors review the ecological and epidemiological characteristics of vector-borne diseases that may have been affected in their spread and change of distribution by international trade. In addition, they provide a detailed analysis of the risks associated with specific trade routes and recent expansions of vector populations. Finally, the authors highlight the importance, as well as the challenges, of preventive surveillance and regulation. The need for improved monitoring of vector populations and a readiness to face unpredictable epidemiological events are also emphasised, since this will require rapid reaction, not least in the regulatory context.

Trypanosoma brucei gambiense: HMI-9 medium containing methylcellulose and human serum supports the continuous axenic in vitro propagation of the bloodstream form.

Trypanosoma brucei (T.b.) gambiense causes the chronic form of human African trypanosomiasis or sleeping sickness. One of the major problems with studying T.b. gambiense is the difficulty to isolate it from its original host and the difficult adaptation to in vivo and in vitro mass propagation. The objective of this study was to evaluate if an established method for axenic culture of pleomorphic bloodstream form T.b. brucei strains, based on methylcellulose containing HMI-9 medium, also facilitated the continuous in vitro propagation of other bloodstream form Trypanozoon strains, in particular of T.b. gambiense. Bloodstream form trypanosomes from one T.b. brucei, two T.b. rhodesiense, one T. evansi and seven T.b. gambiense strains were isolated from mouse blood and each was concurrently cultivated in liquid and methylcellulose-containing HMI-9 based medium, either with or without additional human serum supplementation, for over 10 consecutive sub passages. Although HMI-9 based medium supplemented with 1.1% (w/v) methylcellulose supported the continuous cultivation of all non-gambiense strains better than liquid media could, the in vitro cultivation of all gambiense strains was only achieved in HMI-9 based medium containing 1.1% (w/v) methylcellulose, 15% (v/v) fetal calf serum and 5% (v/v) heat-inactivated human serum.

Efficacy of Cymelarsan and Diminasan against Trypanosoma equiperdum infections in mice and horses.

Trypanocidal sensitivity studies were conducted to assess the efficacy of Diminazene diaceturate (Diminasan) and Bis (aminoethylthio) 4-melaminophenylarsine dihydrochloride (Cymelarsan) against Trypanosoma equiperdum (isolated from two mares with chronic cases of dourine) 713/943 and 834/940 Dodola strains in experimentally infected mice and horses. Diminasan at doses from 3.5 mg/kg to 28 mg/kg and Cymelarsan at doses of 0.25 mg/kg and 0.5 mg/kg body weight failed to cure any of the mice, indicating a clear dose dependent relationship in the mean time of relapse observed in mice. Indeed, mice treated with lower doses relapsed after a shorter time than mice treated with higher doses. However, mice treated with Cymelarsan at doses of 1.0 mg/kg and 2.0 mg/kg body weight were cured and no parasitemia was observed for 60 days. The efficacy of Cymelarsan was also tested in horses. Two groups of horses containing two animals each were infected with T. equiperdum 834/940 Dodola strain and treated with Cymelarsan at a dose rate of 0.25 mg/kg and 0.5 mg/kg, respectively. Cymelarsan at 0.25 mg/kg and 0.5 mg/kg body weight cleared parasitemia within 24 h post treatment and none of the animals were found to show relapse throughout the 320 days of observation. The sensitivity of the particular trypanosome strain to Cymelarsan was also supported by the relative improvement in the mean PCV levels of horses following treatment. A statistically significant difference (p<0.01) in the mean PCV levels of horses treated with Cymelarsan was observed between day 20 at peak parasitemia and days 40 as well as 60 of observation. The mean PCV levels of horses in the control group progressively decreased within the first 60 days of post infection. Two of the horses in the control group developed chronic form of dourine manifested by genital as well as nervous signs with progressive loss of body condition within 320 days post infection. The efficacy of Cymelarsan against the chronic form of dourine was confirmed after treatment of one of the control horses with Cymelarsan at a dose rate of 0.25 mg/kg body weight at day 282 post infection. It was noted that the treated horse improved overall body condition and clinical signs such as incoordination of hind legs, weakness and ventral oedema disappeared within 10 days of treatment. Thus, Cymelarsan was found to be quite effective in curing horses in acute as well as chronic form of dourine. The results obtained from the present study will be important for designing effective control measures against dourine.

Usefulness of additional measurements of the median nerve with ultrasonography.

High resolution sonography is a relatively new diagnostic technique in diagnosing carpal tunnel syndrome (CTS). Normal values in different studies, however, vary and this makes their practical use difficult. The aim of this study was to establish normal values for the median nerve cross-sectional area (CSA) and to investigate the value of measuring additional parameters. Ninety-eight wrists of 29 women and 25 men without signs or symptoms of CTS were included. Width and circumference of the wrist were measured. The CSA of the median nerve at the level of the pisiform bone was measured using ultrasonography. We found a significant correlation between the CSA of the median nerve at the wrist and wrist circumference. Measuring wrist circumference will establish the upper level of normal more accurately compared to predictions solely based upon gender. This has important implications in diagnosing CTS with ultrasonography.

Seroepidemiological survey of trypanozoon infection in horses in the suspected dourine-infected Bale highlands of the Oromia region, Ethiopia.

This paper presents the results of a seroepidemiological survey of trypanozoon infection in horses carried out between September 2007 and June 2008. The survey was conducted to determine the seroprevalence of anti-trypanozoon antibodies in 880 serum samples collected randomly from selected horse-breeding districts of the Bale highlands of Ethiopia. The seroprevalence of trypanozoon infection was found to be 173 (19.66%) and 140 (15.91%) for the CATT/T. evansi and LATEX/T. evansi tests, respectively. The high seroprevalence of trypanozoon infection strongly indicates that the infection is endemic. Neither test can differentiate between anti-trypanozoon antibodies caused by infection with T. equiperdum (the causative agent of dourine) and those of T. evansi (the causative agent of surra). The findings of the present study suggest that field-applicable screening serological tests such as the CATT/T. evansi and LATEX/T. evansi could be useful for epidemiological studies and the control of trypanozoon infection.

Current practice in the use of nerve conduction studies in carpal tunnel syndrome by surgeons in the netherlands.

The current practice in and the opinion about the treatment of carpal tunnel syndrome by surgeons in the Netherlands was evaluated in respect of the extent to which electrodiagnostic studies are used or needed to confirm the diagnosis. Questionnaires were sent to all Dutch surgeons who operate on patients with carpal tunnel syndrome. The response rate was 47% (324 out of 686). The majority of neurosurgeons and orthopaedic surgeons seldom operate without electrodiagnostic confirmation in line with the Dutch consensus guideline on this subject. In contrast, plastic surgeons operate more often on patients with clinically defined carpal tunnel syndrome even with normal electrodiagnostic studies. Knowledge of these strikingly different diagnostic and therapeutic strategies and opinions may influence diagnostic and referral behaviour of clinical neurologists and others.

Validation of a PCR-oligochromatography test for detection of Trypanozoon parasites in a multicenter collaborative trial.

A PCR-oligochromatography test for diagnosis of human and animal trypanosomiasis was evaluated through a multicenter ring trial with six laboratories testing a set of 21 blinded samples, resulting in qualitative data (positive or negative). Results showed an intralaboratory repeatability (accordance) of 88.7% (credible interval [CI], 84.4 to 92.5%) and an interlaboratory repeatability (concordance) of 88.1% (CI, 84.3 to 92.3%).

The invariant surface glycoprotein ISG75 gene family consists of two main groups in the Trypanozoon subgenus.

In Trypanosoma brucei brucei, an invariant surface glycoprotein of molecular weight 75 kDa (ISG75) is uniformly distributed over the surface of a trypanosome and is specific for bloodstream-form parasites. For the other taxa of the Trypanozoon subgenus no data about this surface molecule are available. Therefore, we investigated the ISG75 in the genomes of several pathogenic Trypanozoon by Southern blot, PCR and RT-PCR and sequence analysis. This study reveals that (i) all members of the Trypanozoon subgenus, i.e. T. b. brucei, T. b. gambiense, T. b. rhodesiense, T. evansi and T. equiperdum, harbour ISG75 as multiple gene copies with at least 4-16 copies per genome; (ii) ISG75 gDNA and cDNA sequences are distributed in 2 groups that share at least 75% and 77% identity respectively; (iii) sequences from both groups are transcribed in all species and subspecies of the Trypanozoon subgenus; (iv) the main differences between group I and group II are located in the variable region at the amino-terminus of the putative proteins; (v) however, all the sequences in both groups have some well-conserved features, such as the cysteine residues, an amino-terminal cleavable signal peptide, a single alpha-helix transmembrane domain and a cytoplasmic domain at the carboxy-terminus.

Storage of red blood cells with improved maintenance of 2,3-bisphosphoglycerate.

BACKGROUND: During storage, red blood cells (RBCs) rapidly lose 2,3-bisphosphoglycerate (2,3-DPG) leading to an increase in the affinity for O(2) and a temporary impairment of O(2) transport. Recent clinical evaluations indicate that the quality of transfused RBCs may be more important for patient survival than previously recognized. STUDY DESIGN AND METHODS: Glucose-free additive solutions (ASs) were prepared with sodium citrate, sodium gluconate, adenine, mannitol, and phosphates at high pH, a solution that can be heat-sterilized. CP2D was used as an anticoagulant. Additional CP2D was added to the AS to supply glucose. RBCs were stored at 4 degrees C and assayed periodically for intracellular pH (pHi), extracellular pH, glucose, lactate, phosphate, ATP, 2,3-DPG, hemolysis, and morphology. RESULTS: Storage in 175 mL of the chloride-free, hypotonic medium at a hematocrit (Hct) level of 59 to 60 percent resulted in an elevated pHi and the maintenance of 2,3-DPG at or above the initial value for 2 weeks without loss of ATP. The addition of 400 mL of storage solution followed by centrifugation and removal of 300 mL of excess solution to a Hct level of 60 to 66 percent further reduced the chloride concentration, resulting in the maintenance of 2,3-DPG for 4 weeks. Hemolysis was at 0.1 percent at 6 weeks. CONCLUSION: Improvements in the maintenance of 2,3-DPG were achieved with 175 mL of a chloride-free storage solution with familiar additives at nontoxic concentrations to increase pHi. Adding, instead, 400 mL of storage solution followed by the removal of 300 mL reduced the chloride concentration, increasing the pHi and extending the maintenance of 2,3-DPG to 4 weeks.

Molecular dipstick test for diagnosis of sleeping sickness.

Human African trypanosomiasis (HAT) or sleeping sickness is a neglected disease that affects poor rural populations across sub-Saharan Africa. Confirmation of diagnosis is based on detection of parasites in either blood or lymph by microscopy. Here we present the development and the first-phase evaluation of a simple and rapid test (HAT-PCR-OC [human African trypanosomiasis-PCR-oligochromatography]) for detection of amplified Trypanosoma brucei DNA. PCR products are visualized on a dipstick through hybridization with a gold-conjugated probe (oligochromatography). Visualization is straightforward and takes only 5 min. Controls both for the PCR and for DNA migration are incorporated into the assay. The lower detection limit of the test is 5 fg of pure T. brucei DNA. One parasite in 180 microl of blood is still detectable. Sensitivity and specificity for T. brucei were calculated at 100% when tested on blood samples from 26 confirmed sleeping sickness patients, 18 negative controls (nonendemic region), and 50 negative control blood samples from an endemic region. HAT-PCR-OC is a promising new tool for diagnosis of sleeping sickness in laboratory settings, and the diagnostic format described here may have wider application for other infectious diseases.

Red blood cells intended for transfusion: quality criteria revisited.

Great variation exists with respect to viability and function of fresh and stored red blood cells (RBCs) as well as of the contents of RBC hemoglobin (Hb) in individual units. Improved technology is available for the preparation as well as the storage of RBCs. The authors raise the question whether it may be time to revise current standards for RBC units. The establishment of a standard unit of blood based on Hb content should be a high-priority goal. It is recommended that a standard RBC unit should contain 50 g of Hb. Major organizations concerned with the collection and distribution of blood components should agree on the criteria for a standard unit of RBCs based on Hb content and for the collection of double units. Manufacturers of blood collection equipment should provide suitable technology for collecting a standard unit with defined contents of RBC Hb. Efforts should be directed at the design of storage solutions acceptable for transfusion that maximize the maintenance of both RBC viability and function during storage. The ideal storage protocol would require sterile, high-pH solutions containing both glucose and electrolytes.