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Transition of the mRNA sequence downstream from the initiation codon into a single-stranded conformation is strongly promoted by binding of the initiator tRNA

Using an RNA footprinting technique, accessible sites on the mRNA initiation region bound to the ribosome have been determined. Chemical probing experiments have been done both in the presence and absence of the initiator tRNA with dimethyl sulfate, kethoxal and carbodiimide as reagent probes. As an mRNA, a mini-mRNA containing the initiation region of bacteriophage λ gene cro has been used. This region is characterized by a long single-stranded Shine-Dalgarno (SD) sequence followed by two hairpin structures of which the first one comprises in its loop the initiation codon. As compared to a free mRNA, the only nucleotides additionally protected in the binary mRNA-ribosome complex have been those which belong to the S-D sequence and the initiation codon. The protection of other nucleotides has not changed. Addition of the initiator RNA results in the modification of nucleotides in the stems of the downstream hairpin structures of the initiation region. This reflects their transition into a single-stranded conformation promoted by tRNA. A possible implication of these findings for the decoding process is discussed.

Study of Fusion Protein and Attachment Glycoprotein of Nipah Virus Expressed in Recombinant Baculovirus

The envelope attachment glycoprotein (G) and fusion protein (F′) of Nipah virus (NiV) play a key role in viral entry and induction of neutralization antibody. In this study, recombinant baculoviruses, rBac-NF and rBac-NG, were generated to express F and G proteins of NiV. The expressions of recombinant G (rNG) and F (rNF) proteins in rBac-NF and rBac-NG-infected cells were confirmed by Western blot. Both rNG and rNF showed sensitive and specific antigenic reaction to rabbit serum anti-Nipah virus in indirect immunofluorescence detection and indirect ELISA. Immunization with rBac-NF and rBac-NG-infected insect cells elicited G and F protein-specific antibody responses in mice. Furthermore, the G and F protein-specific antibodies could neutralize the infectivity of the VSVΔG*F/G, the NiV F and G envelope glycoproteins of pseudotype recombinant Vesicular Stomatitis Virus expressing green fluorescence protein. The results demonstrated that the F and G proteins expressed by the recombinant baculoviruses could be safe diagnostic antigens for the surveillance and monitoring of NiV and could also be promising subunit vaccines for the prevention of NiV.

Applications of quantitative PCR in the biosafety and genetic stability assessment of biotechnology products

High throughput screening, increased accuracy and the coupling of real-time quantitative PCR (Q-PCR) to robotic set-up systems are beginning to revolutionise biotechnology. Applications of Q-PCR within biotechnology are discussed with particular emphasis on the following areas of biosafety and genetic stability testing: (a) determination of the biodistribution of gene therapy vectors in animals; (b) quantification of the residual DNA in final product therapeutics; (c) detection of viral and bacterial nucleic acid in contaminated cell banks and final products; (d) quantification of the level of virus removal in process validation viral clearance studies; (e) specific detection of retroviral RT activity in vaccines with high sensitivity; and (f) transgene copy number determination for monitoring genetic stability during production. Methods employed for Q-PCR assay validation as required in ICH Topic Q2A Validation of Analytical Methods: Definitions and Terminology (1st June 1995) are also reviewed.

Application of Optical Protein-chip in Detecting Phage M13KO7

Avidin layer was bound on the substrate surface of Silicon wafer modified with aldehyde. The interaction between avidin and biotin was adopted for the immobilization of mouse monoclonal biotin-anti-M13 (antibody GP3)-labeled biotin. The surface was incubated in a solution containing phage M13KO7, which was trapped by the antibody GP3 with the interaction between phage M13KO7 and antibody GP3, resulting in a variation of layer thickness that was detected by imaging ellipsometry. The results showed a saturated layer of antibody GP3 with a thickness about 6.9 nm on the surface of the silicon wafer. The specific interaction between phage M13KO7 and antibody GP3 resulted in a variation of layer thickness. The layer of phage M13KO7 bound with antibody GP3 was 17.5 nm in the concentration of 1.1×10(10) pfu/mL. Each variation of the layer thickness corresponded to a concentration of phage M13KO7 in the range of 0.1×10(10)–2.5×10(10) pfu/mL, with the sensitivity of 10(9) pfu/mL. Compared with other methods, the optical protein-chip, requiring only short measurement time, label free, is a quantitative test, and can be visualized. This study could be significant on the interactions between the antibody and the virus, showing potential in the early diagnosis of virosis.

Chemokines and Their Receptors: Roles in Specific Clinical Conditions and Measurement in the Clinical Laboratory

Considerable progress has been achieved in our knowledge of the function of the chemokine system and in understanding its role in the pathophysiology of human diseases. This complex system, presently including approximately 50 cytokines and 20 receptors, coordinates leukocyte recruitment in a variety of human diseases, ranging from infectious and inflammatory diseases to cancer. A large body of literature has been published describing various assays for the measurement of chemokines in biologic fluids and tissues. We review information available on the role of chemokines in selected human diseases and provide examples of clinical situations in which chemokine determination might be of practical value, and we describe the currently available assays for their measurement.

Detecting geospatial patterns of Plasmodium falciparum parasite migration in Cambodia using optimized estimated effective migration surfaces

BACKGROUND: Understanding the genetic structure of natural populations provides insight into the demographic and adaptive processes that have affected those populations. Such information, particularly when integrated with geospatial data, can have translational applications for a variety of fields, including public health. Estimated effective migration surfaces (EEMS) is an approach that allows visualization of the spatial patterns in genomic data to understand population structure and migration. In this study, we developed a workflow to optimize the resolution of spatial grids used to generate EEMS migration maps and applied this optimized workflow to estimate migration of Plasmodium falciparum in Cambodia and bordering regions of Thailand and Vietnam. METHODS: The optimal density of EEMS grids was determined based on a new workflow created using density clustering to define genomic clusters and the spatial distance between genomic clusters. Topological skeletons were used to capture the spatial distribution for each genomic cluster and to determine the EEMS grid density; i.e., both genomic and spatial clustering were used to guide the optimization of EEMS grids. Model accuracy for migration estimates using the optimized workflow was tested and compared to grid resolutions selected without the optimized workflow. As a test case, the optimized workflow was applied to genomic data generated from P. falciparum sampled in Cambodia and bordering regions, and migration maps were compared to estimates of malaria endemicity, as well as geographic properties of the study area, as a means of validating observed migration patterns. RESULTS: Optimized grids displayed both high model accuracy and reduced computing time compared to grid densities selected in an unguided manner. In addition, EEMS migration maps generated for P. falciparum using the optimized grid corresponded to estimates of malaria endemicity and geographic properties of the study region that might be expected to impact malaria parasite migration, supporting the validity of the observed migration patterns. CONCLUSIONS: Optimized grids reduce spatial uncertainty in the EEMS contours that can result from user-defined parameters, such as the resolution of the spatial grid used in the model. This workflow will be useful to a broad range of EEMS users as it can be applied to analyses involving other organisms of interest and geographic areas.

Serial change of neutrophil extracellular traps in tracheal aspirate of patients with acute respiratory distress syndrome: report of three cases

BACKGROUND: Neutrophil extracellular traps (NETs) are fibrous structures released from activated neutrophils. NET formation has been reported to be associated with acute respiratory distress syndrome (ARDS). However, there are no reports dealing with serial changes of NET formation in tracheal aspirate of ARDS patients. CASE PRESENTATION: We report three cases of ARDS. Case 1 is a 69-year-old man with necrotizing fasciitis of the buttocks, case 2 is a 49-year-old woman with extensive burns (80% of total body surface), and case 3 is a 73-year-old woman with severe bacterial pneumonia. We found abundant expression of citrullinated histone H3 (Cit-H3) and the formation of NETs at the onset of ARDS in all cases. The amounts of Cit-H3 and NETs decreased with the amelioration of respiratory failure in cases 1 and 2. In case 2, the amounts of Cit-H3 and NETs increased with aggravation of infection and respiratory status. In case 3, the abundant expression of Cit-H3 and NETs persisted; the patient did not recover from ARDS and eventually died. Cit-H3 and NETs were found in tracheal aspirates even if the patients had no direct injury to the lung as in cases 1 and 2. CONCLUSIONS: In these three cases, the formation of NETs was observed in tracheal aspirate of patients with ARDS by either direct or indirect insults to the lung. The amount of NET formation changed dynamically over the clinical course of each patient.

Moving beyond the mousetrap: current and emerging humanized mouse and rat models for investigating prevention and cure strategies against HIV infection and associated pathologies

The development of safe and effective combination antiretroviral therapies for human immunodeficiency virus (HIV) infection over the past several decades has significantly reduced HIV-associated morbidity and mortality. Additionally, antiretroviral drugs have provided an effective means of protection against HIV transmission. Despite these advances, significant limitations exist; namely, the inability to eliminate HIV reservoirs, the inability to reverse lymphoid tissues damage, and the lack of an effective vaccine for preventing HIV transmission. Evaluation of the safety and efficacy of therapeutics and vaccines for eliminating HIV reservoirs and preventing HIV transmission requires robust in vivo models. Since HIV is a human-specific pathogen, that targets hematopoietic lineage cells and lymphoid tissues, in vivo animal models for HIV-host interactions require incorporation of human hematopoietic lineage cells and lymphoid tissues. In this review, we will discuss the construction of mouse models with human lymphoid tissues and/or hematopoietic lineage cells, termed, human immune system (HIS)-humanized mice. These HIS-humanized mouse models can support the development of functional human innate and adaptive immune cells, along with primary (thymus) and secondary (spleen) lymphoid tissues. We will discuss applications of HIS-humanized mouse models in evaluating the safety and efficacy of therapeutics against HIV reservoirs and associated immunopathology, and delineate the human immune response elicited by candidate HIV vaccines. In addition to focusing on how these HIS-humanized mouse models have already furthered our understanding of HIV and contributed to HIV therapeutics development, we discuss how emerging HIS-humanized rat models could address the limitations of HIS-mouse models.

HijAkt: The PI3K/Akt Pathway in Virus Replication and Pathogenesis

As obligate parasites of cellular processes, viruses must take over cellular macromolecular machinery. It is also becoming clear that viruses routinely control intracellular signaling pathways through the direct or indirect control of kinases and phosphatases. This control of cellular phosphoproteins is important to promote a variety of viral processes, from control of entry to nuclear function to the stimulation of viral protein synthesis. This review focuses on the takeover of the cellular phosphatidylinositol-3-kinase (PI3K)/Akt signaling pathway by a variety of retroviruses, DNA viruses, and RNA viruses, highlighting the functions ascribed to virus activation of PI3K and Akt activity. This review also describes the role that the PI3K/Akt pathway plays in the host response, noting that it that can trigger anti- as well as proviral functions.

Inflammatory demyelinating diseases of the central nervous system

Inflammatory demyelinating diseases are a heterogeneous group of disorders, which occur against the background of an acute or chronic inflammatory process. The pathologic hallmark of multiple sclerosis (MS) is the presence of focal demyelinated lesions with partial axonal preservation and reactive astrogliosis. Demyelinated plaques are present in the white as well as gray matter, such as the cerebral or cerebellar cortex and brainstem nuclei. Activity of the disease process is reflected by the presence of lesions with ongoing myelin destruction. Axonal and neuronal destruction in the lesions is a major substrate for permanent neurologic deficit in MS patients. The MS pathology is qualitatively similar in different disease stages, such as relapsing remitting MS or secondary or primary progressive MS, but the prevalence of different lesion types differs quantitatively. Acute MS and Balo's type of concentric sclerosis appear to be variants of classic MS. In contrast, neuromyelitis optica (NMO) and spectrum disorders (NMOSD) are inflammatory diseases with primary injury of astrocytes, mediated by aquaporin-4 antibodies. Finally, we discuss the histopathology of other inflammatory demyelinating diseases such as acute disseminated encephalomyelitis and myelin oligodendrocyte glycoprotein antibody-associated demyelination. Knowledge of the heterogenous immunopathology in demyelinating diseases is important, to understand the clinical presentation and disease course and to find the optimal treatment for an individual patient.

Regulatory T Cells in Infection

Infectious agents have intimately co-evolved with the host immune system, acquiring a portfolio of highly sophisticated mechanisms to modulate immunity. Among the common strategies developed by viruses, bacteria, protozoa, helminths, and fungi is the manipulation of the regulatory T cell network in order to favor pathogen survival and transmission. Treg activity also benefits the host in many circumstances by controlling immunopathogenic reactions to infection. Interestingly, some pathogens are able to directly induce the conversion of naive T cells into suppressive Foxp3-expressing Tregs, while others activate pre-existing natural Tregs, in both cases repressing pathogen-specific effector responses. However, Tregs can also act to promote immunity in certain settings, such as in initial stages of infection when effector cells must access the site of infection, and subsequently in ensuring generation of effector memory. Notably, there is little current information on whether infections selectively drive pathogen-specific Tregs, and if so whether these cells are also reactive to self-antigens. Further analysis of specificity, together with a clearer picture of the relative dynamics of Treg subsets over the course of disease, should lead to rational strategies for immune intervention to optimize immunity and eliminate infection.

Modern Approaches in Probiotics Research to Control Foodborne Pathogens

Foodborne illness is a serious public health concern. There are over 200 known microbial, chemical, and physical agents that are known to cause foodborne illness. Efforts are made for improved detection, control and prevention of foodborne pathogen in food, and pathogen associated diseases in the host. Several commonly used approaches to control foodborne pathogens include antibiotics, natural antimicrobials, bacteriophages, bacteriocins, ionizing radiations, and heat. In addition, probiotics offer a potential intervention strategy for the prevention and control of foodborne infections. This review focuses on the use of probiotics and bioengineered probiotics to control foodborne pathogens, their antimicrobial actions, and their delivery strategies. Although probiotics have been demonstrated to be effective in antagonizing foodborne pathogens, challenges exist in the characterization and elucidation of underlying molecular mechanisms of action and in the development of potential delivery strategies that could maintain the viability and functionality of the probiotic in the target organ.

Optimization of 5′ Untranslated Region of Modified mRNA for Use in Cardiac or Hepatic Ischemic Injury

Modified mRNA (modRNA) is a gene-delivery platform for transiently introducing a single gene or several genes of interest to different cell types and tissues. modRNA is considered to be a safe vector for gene transfer, as it negligibly activates the innate immune system and does not compromise the genome integrity. The use of modRNA in basic and translational science is rising, due to the clinical potential of modRNA. We are currently using modRNA to induce cardiac regeneration post-ischemic injury. Major obstacles in using modRNA for cardiac ischemic disease include the need for the direct and single administration of modRNA to the heart and the inefficient translation of modRNA due to its short half-life. Modulation of the 5′ untranslated region (5′ UTR) to enhance translation efficiency in ischemic cardiac disease has great value, as it can reduce the amount of modRNA needed per delivery and will achieve higher and longer protein production post-single delivery. Here, we identified that 5′ UTR, from the fatty acid metabolism gene carboxylesterase 1D (Ces1d), enhanced the translation of firefly luciferase (Luc) modRNA by 2-fold in the heart post-myocardial infarction (MI). Moreover, we identified, in the Ces1d, a specific RNA element (element D) that is responsible for the improvement of modRNA translation and leads to a 2.5-fold translation increment over Luc modRNA carrying artificial 5′ UTR, post-MI. Importantly, we were able to show that 5′ UTR Ces1d also enhances modRNA translation in the liver, but not in the kidney, post-ischemic injury, indicating that Ces1d 5′ UTR and element D may play a wider role in translation of protein under an ischemic condition.

Structural Insight into Paramyxovirus and Pneumovirus Entry Inhibition

Paramyxoviruses and pneumoviruses infect cells through fusion (F) protein-mediated merger of the viral envelope with target membranes. Members of these families include a range of major human and animal pathogens, such as respiratory syncytial virus (RSV), measles virus (MeV), human parainfluenza viruses (HPIVs), and highly pathogenic Nipah virus (NiV). High-resolution F protein structures in both the metastable pre- and the postfusion conformation have been solved for several members of the families and a number of F-targeting entry inhibitors have progressed to advanced development or clinical testing. However, small-molecule RSV entry inhibitors have overall disappointed in clinical trials and viral resistance developed rapidly in experimental settings and patients, raising the question of whether the available structural information may provide a path to counteract viral escape through proactive inhibitor engineering. This article will summarize current mechanistic insight into F-mediated membrane fusion and examine the contribution of structural information to the development of small-molecule F inhibitors. Implications are outlined for future drug target selection and rational drug engineering strategies.

Antiviral Activity of Benzavir-2 against Emerging Flaviviruses

Most flaviviruses are arthropod-borne viruses, transmitted by either ticks or mosquitoes, and cause morbidity and mortality worldwide. They are endemic in many countries and have recently emerged in new regions, such as the Zika virus (ZIKV) in South-and Central America, the West Nile virus (WNV) in North America, and the Yellow fever virus (YFV) in Brazil and many African countries, highlighting the need for preparedness. Currently, there are no antiviral drugs available to treat flavivirus infections. We have previously discovered a broad-spectrum antiviral compound, benzavir-2, with potent antiviral activity against both DNA- and RNA-viruses. Our purpose was to investigate the inhibitory activity of benzavir-2 against flaviviruses. We used a ZIKV ZsGreen-expressing vector, two lineages of wild-type ZIKV, and other medically important flaviviruses. Benzavir-2 inhibited ZIKV derived reporter gene expression with an EC(50) value of 0.8 ± 0.1 µM. Furthermore, ZIKV plaque formation, progeny virus production, and viral RNA expression were strongly inhibited. In addition, 2.5 µM of benzavir-2 reduced infection in vitro in three to five orders of magnitude for five other flaviviruses: WNV, YFV, the tick-borne encephalitis virus, Japanese encephalitis virus, and dengue virus. In conclusion, benzavir-2 was a potent inhibitor of flavivirus infection, which supported the broad-spectrum antiviral activity of benzavir-2.

Topology, Antiviral Functional Residues and Mechanism of IFITM1

Interferon-inducible transmembrane proteins (IFITM1/2/3) have been reported to suppress the entry of a wide range of viruses. However, their antiviral functional residues and specific mechanisms are still unclear. Here, we firstly resolved the topology of IFITM1 on the plasma membrane where N-terminus points into the cytoplasm and C-terminus resides extracellularly. Further, KRRK basic residues of IFITM1 locating at 62–67 of the conserved intracellular loop (CIL) were found to play a key role in the restriction on the Zika virus (ZIKV) and dengue virus (DENV). Similarly, KRRK basic residues of IFITM2/3 also contributed to suppressing ZIKV replication. Finally, IFITM1 was revealed to be capable of restricting the release of ZIKV particles from endosome to cytosol so as to impede the entry of ZIKV into host cells, which was tightly related with the inhibition of IFITM1 on the acidification of organelles. Overall, our study provided topology, antiviral functional residues and the mechanism of interferon-inducible transmembrane proteins.

Berberine Hampers Influenza A Replication through Inhibition of MAPK/ERK Pathway

Background: Berberine (BBR) is an isoquinoline alkaloid which exhibits a variety of biological and therapeutic properties, and has been reported by some to block replication of the influenza virus. However, contradictory results have also been presented, and the mechanistic explanation is lacking. Methods: A panel of cell lines (Madin–Darby canine kidney (MDCK), adenocarcinoma human alveolar basal epithelial cells (A549), lung epithelial type I (LET1)) and primary human airway epithelial cells (HAE) susceptible to influenza virus infection were infected with a seasonal influenza A virus in the presence or absence of BBR. Cytotoxicity towards cell lines was measured using XTT assay. The yield of the virus was analyzed using RT-qPCR. To study the molecular mechanism of BBR, confocal microscopy and Western blot analyses of cellular fractions were applied. Results and conclusions: Our results show cell-type-dependent anti-influenza properties of BBR in vitro which suggests that the compound acts on the cell and not the virus. Importantly, BBR hampers influenza replication in primary human airway epithelium 3D cultures that mimic the natural replication site of the virus. Studies show that the influenza A virus upregulates the mitogen-activated protein kinase/extracellular signal-related kinase (MAPK/ERK) pathway and hijacks this pathway for nucleolar export of the viral ribonucleoprotein. Our results suggest that BBR interferes with this process and hampers influenza A replication.

Harnessing the Potential of Stem Cells for Disease Modeling: Progress and Promises

Ex vivo cell/tissue-based models are an essential step in the workflow of pathophysiology studies, assay development, disease modeling, drug discovery, and development of personalized therapeutic strategies. For these purposes, both scientific and pharmaceutical research have adopted ex vivo stem cell models because of their better predictive power. As matter of a fact, the advancing in isolation and in vitro expansion protocols for culturing autologous human stem cells, and the standardization of methods for generating patient-derived induced pluripotent stem cells has made feasible to generate and investigate human cellular disease models with even greater speed and efficiency. Furthermore, the potential of stem cells on generating more complex systems, such as scaffold-cell models, organoids, or organ-on-a-chip, allowed to overcome the limitations of the two-dimensional culture systems as well as to better mimic tissues structures and functions. Finally, the advent of genome-editing/gene therapy technologies had a great impact on the generation of more proficient stem cell-disease models and on establishing an effective therapeutic treatment. In this review, we discuss important breakthroughs of stem cell-based models highlighting current directions, advantages, and limitations and point out the need to combine experimental biology with computational tools able to describe complex biological systems and deliver results or predictions in the context of personalized medicine.

Pulmonary vasodilators can lead to various complications in pulmonary “arterial” hypertension associated with congenital heart disease

Congenital heart disease-associated pulmonary arterial hypertension (CHD-PAH) is one of the major complications in patients with CHD. A timely closure of the left-to-right shunt will generally result in the normalization of the pulmonary hemodynamics, but a few patients have severe prognosis in their early childhood. We hypothesized that wide-ranging pathological mechanism in PAH could elucidate the clinical state of severe CHD-PAH. Using electronic medical records, we retrospectively analyzed six infants with severe CHD-PAH who had treatment-resistant PH. All patients were born with congenital malformation syndrome. After starting on a pulmonary vasodilator, five of the six patients developed complications including pulmonary edema and interstitial lung disease (ILD), and four patients had alveolar hemorrhage. After steroid therapy, the clinical condition improved in four patients, but two patients died. The autopsy findings in one of the deceased patients indicated the presence of recurrent alveolar hemorrhage, pulmonary venous hypertension, ILD, and PAH. Based on the clinical course of these CHD-PAH in patients and the literature, CHD-PAH can occur with pulmonary vascular obstructive disease (PVOD)/pulmonary capillary hemangiomatosis (PCH), ILD, and/or alveolar hemorrhage. The severity of CHD-PAH may depend on a genetic disorder, respiratory infection, and upper airway stenosis. Additionally, pulmonary vasodilators may be involved in the development of PVOD/PCH and ILD. When patients with CHD-PAH show unexpected deterioration, clinicians should consider complications associated with PVOD/PCH and/or pulmonary disease. In addition, the choice of upfront combination therapy for pediatric patients with CHD-PAH should be selected carefully.

Research on covert communication channel based on modulation of common compressed speech codec

As is well known, multimedia has been widely used in VoIP and mobile communications. Research on how to establish covert communication channel over the above popular public applications has been flourishing in recent years. This paper tries to present a novel and effective method to construct a covert channel over common compressed speech stream by embedding sense information into it. In our method, after analysing the characteristic features of the excitation pulse positions of the ITU-T G.723.1 and G.729A speech codec, we design a novel and effective covert communication channel by finely modulating the codes of excitation pulse positions of the above two codecs in line with the secret information to be hidden. To improve the embedding capacity of the proposed method, we also use all the odd/even characteristics of pulse code positions to conduct information hiding. To test and verify the proposed approach, experiments are conducted on several different scenarios. Experimental results show that our methods and algorithms perform a higher degree of secrecy and sound information embedding efficacy compared with exiting similar methods.

Isolation of influenza A virus, subtype H5N2, and avian paramyxovirus type 1 from a flock of ostriches in Europe

A total of 146 of 506 ostriches (Struthio camelus) introduced into a quarantine in Denmark died within the first 23 days. The majority of deaths were in young birds up to 10 kg body weight. Avian influenza A viruses (AIVs) were isolated from 14 pools of organ tissues representing seven groups each of three or four ostriches, which died over the first 3 weeks. The AIVs were detected in respiratory tissues, kidneys and intestines. All were subtype H5N2. The intravenous pathogenicity index of each isolate for chickens was 0.0 and the four isolates examined each had the amino acid sequence -P-Q-R-E-T-R*G-L-F- at the cleavage site of the haemagglutinin protein, typical of non-pathogenic AIVs. In addition, an avirulent avian paramyxovirus type 1 virus was isolated from one pool of kidney tissues. Bacteriological examination gave no significant results. The most characteristic pathological findings were impaction of the proventriculus and gizzard, enteritis with stasis and multi-focal necrotic hepatitis.

Antigenic and immunogenic characterization of infectious bronchitis virus strains isolated in China between 1986 and 1995

Eight strains of infectious bronchitis virus (IBV) were isolated between 1986 and 1995 from broilers and layers at eight different farms in four provinces in China. The viruses were isolated from flocks which suffered from either respiratory disease or nephritis and the majority had not been vaccinated against IBV. Six strains were shown by monoclonal antibodies to differ from H120, Connecticut and Arkansas 99 strains of IBV and also to differ from each other. Four of these strains were serotyped; one (NRZ) was of the Massachusetts serotype, three (HV, NB-90 and TJ) shared a degree of antigenic similarity and were of a serotype that differed from Massachusetts and Connecticut. NB-90 was similar to both Gray and T strains whereas TJ shared some similarity with the T strain. Four strains, HV, NB-90, YY and TJ induced 33, 47, 60 and 90% mortality, respectively, in 3-week-old specific pathogen-free chickens. Clinical signs and post-mortem findings were identical to those induced by the nephropathogenic T strain. Chickens vaccinated with H120 strain, and then challenged with four highly pathogenic strains HV, NB-90, YY and TJ were not protected as determined by both virus isolation and mortality. The results show that highly pathogenic IBV strains which induce clinical nephritis occur frequently in poultry flocks in China. They also confirm field observations on the lack of protection by currently used IB vaccines of the Massachusetts serotype against challenge with these nephropathogenic strains.

An adaptive parareal algorithm()

In this paper, we consider the problem of accelerating the numerical simulation of time dependent problems by time domain decomposition. The available algorithms enabling such decompositions present severe efficiency limitations and are an obstacle for the solution of large scale and high dimensional problems. Our main contribution is the improvement of the parallel efficiency of the parareal in time method. The parareal method is based on combining predictions made by a numerically inexpensive solver (with coarse physics and/or coarse resolution) with corrections coming from an expensive solver (with high-fidelity physics and high resolution). At convergence, the algorithm provides a solution that has the fine solver’s high-fidelity physics and high resolution. In the classical version, the fine solver has a fixed high accuracy which is the major obstacle to achieve a competitive parallel efficiency. In this paper, we develop an adaptive variant that overcomes this obstacle by dynamically increasing the accuracy of the fine solver across the parareal iterations. We theoretically show that the parallel efficiency becomes very competitive in the ideal case where the cost of the coarse solver is small, thus proving that the only remaining factors impeding full scalability become the cost of the coarse solver and communication time. The developed theory has also the merit of setting a general framework to understand the success of several extensions of parareal based on iteratively improving the quality of the fine solver and re-using information from previous parareal steps. We illustrate the actual performance of the method in stiff ODEs, which are a challenging family of problems since the only mechanism for adaptivity is time and efficiency is affected by the cost of the coarse solver.

Cause and predictors of neonatal mortality among neonates admitted to neonatal intensive care units of public hospitals in eastern Ethiopia: a facility-based prospective follow-up study

BACKGROUND: The first month is the most crucial period for child survival. Neonatal mortality continues to remain high with little improvement over the years in Sub-Saharan Africa, including Ethiopia. This region shows the least progress in reducing neonatal mortality and continues to be a significant public health issue. In this study setting, the causes and predictors of neonatal death in the neonatal intensive care units are not well documented. Hence, this study aimed to determine the causes and predictors of neonatal mortality among infants admitted to neonatal intensive care units in eastern Ethiopia. METHODS: A facility-based in prospective follow-up study was conducted among neonates admitted to neonatal intensive care units of public hospitals of eastern Ethiopia from November 1 to December 30, 2018. Data were collected using a pre-tested structured questionnaire and a follow-up checklist. The main outcomes and causes of death were set by pediatricians and medical residents. EpiData 3.1 and Statistical Package for Social Sciences Version 25 software were used for data entry and analysis, respectively. Multivariable logistic regression was used to identify the predictors of facility-based neonatal mortality. RESULTS: The proportion of facility-based neonatal mortality was 20% (95% CI:16.7–23.8%). The causes of death were complications of preterm birth (28.58%), birth asphyxia (22.45%), neonatal infection (18.36%), meconium aspiration syndrome (9.18%), respiratory distress syndrome (7.14%), and congenital malformation (4.08%). Low birth weight, preterm births, length of stay of the neonatal intensive care unit, low 5 min APGAR score, hyperthermia, and initiation of feeding were predictors of neonatal death among infants admitted to the neonatal intensive care units of public hospitals in eastern Ethiopia. CONCLUSIONS: The proportion of facility-based neonatal deaths was unacceptably high. The main causes of death were preventable and treatable. Hence, improving the timing and quality of antenatal care is essential for early detection, anticipating high-risk newborns, and timely interventions. Furthermore, early initiation of feeding and better referral linkage to tertiary health facilities could lead to a reduction in neonatal death in this setting.

Clinical characteristics and outcomes of methamphetamine-associated versus non-methamphetamine intracerebral hemorrhage

Methamphetamine use has emerged as a risk factor for intracerebral hemorrhage (ICH). We aim to investigate the clinical characteristics and outcomes of methamphetamine-associated ICH (Meth-ICH) versus Non-Meth-ICH. Patients with ICH between January 2011 and December 2017 were studied. Meth-ICH and Non-Meth-ICH were defined by history of abuse and urine drug screen (UDS). The clinical features of the 2 groups were explored. Among the 677 consecutive patients, 61 (9.0%) were identified as Meth-ICH and 350 as Non-Meth ICH. Meth-ICH was more common in Hispanics (14.6%) and Whites (10.1%) as compared to Asians (1.2%). Patients with Meth-ICH were more often younger (51.2 vs. 62.2 years, p < 0.001), male (77.0% vs. 61.4.0%, p < 0.05), and smokers (44.3% vs. 13.4%, p < 0.001). Non-Meth-ICH was more likely to have history of hypertension (72.61% v. 59%, p < 0.05) or antithrombotic use (10.9% vs. 1.6%, p < 0.05). There was no significant difference in clinical severity, hospital length of stay (LOS), rate of functional independence (29.5% vs. 25.7%, p = 0.534), or mortality (18.0% vs. 24.6%, p = 0.267) between the 2 groups. Methamphetamine use was not an independent predictor of poor outcome. Despite difference in demographics, Meth-ICH is similar to Non-Meth ICH in hospital course and outcome.

Suppressed humoral immunity is associated with dengue nonstructural protein NS1-elicited anti-death receptor antibody fractions in mice

Dengue virus (DENV) infections may cause life-threatening dengue hemorrhagic fever (DHF). Suppressed protective immunity was shown in these patients. Although several hypotheses have been formulated, the mechanism of DENV-induced immunosuppression remains unclear. Previously, we found that cross-reactive antibodies against tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor 1 (death receptor 4 [DR4]) were elicited in DHF patients, and that anti-DR4 autoantibody fractions were elicited by nonstructural protein 1 (NS1) immunizations in experimental mice. In this study, we found that anti-DR4 antibodies could suppress B lymphocyte function in vitro and in vivo. Treatment with the anti-DR4 immunoglobulin (Ig) induced caspase-dependent cell death in immortalized B lymphocyte Raji cells in vitro. Anti-DR4 Igs elicited by NS1 and DR4 immunizations markedly suppressed mouse spleen transitional T2 B (IgM(+)IgD(+)), bone marrow pre-pro-B (B220(+)CD43(+)), pre-B (B220(+)CD43(−)), and mature B cell (B220(+)IgD(+)) subsets in mice. Furthermore, functional analysis revealed that the pre-elicitation of anti-NS1 and anti-DR4 Ig titers suppressed subsequently neutralizing antibody production by immunization with DENV envelop protein. Our data suggest that the elicitation of anti-DR4 titers through DENV NS1 immunization plays a suppressive role in humoral immunity in mice.

Apoptosis characterization in mononuclear blood leukocytes of HIV patients during dengue acute disease

Dengue virus (DENV) co-circulation in Brazil represents a challenge for treatment and vaccine development. Despite public health impact, the occurrence of coinfections with other viruses is a common event. Increased T cell activation and altered inflammatory response are found during DENV coinfection with Human Immunodeficiency Virus (HIV) impacting HIV-pathogenesis. Even with Antiretroviral therapy (ART), HIV- treated patients had chronic immune activation and lymphocyte apoptosis. However, apoptotic mechanisms have not been investigated during coinfection with DENV. Our attention was attracted to apoptotic cell markers expressions in PBMCs from DENV and DENV/HIV coinfected patients. We found CD4/CD8 ratio inversion in most coinfected patients. CD4 T and CD8 T-cell subsets from DENV and DENV/HIV groups expressed low levels of anti-apoptotic protein Bcl-2. Furthermore, CD8 CD95 double positive cells frequency expressing low levels of Bcl-2 were significantly higher in these patients. Additionally, the density of Bcl-2 on classical monocytes (CD14(++)CD16(−)) was significantly lower during DENV infection. Upregulation of pro-apoptotic proteins and anti-apoptotic proteins were found in DENV and DENV/HIV, while catalase, an antioxidant protein, was upregulated mainly in DENV/HIV coinfection. These findings provide evidence of apoptosis triggering during DENV/HIV coinfection, which may contribute to knowledge of immunological response during DENV acute infection in HIV-patients treated with ART.

Colloidal Silicon Dioxide in Tablet form (Carbowhite) Efficacy in Patients with Acute Diarrhea: Results of Randomized, Double-Blind, Placebo-Controlled, Multi-Center Study

The acute diarrhea is a wide-spread disease. The prescription of enterosorbents is appropriate as a primary measure for the treatment of the acute diarrhea for effective prevention of the fluid and electrolyte loss, as well as method for symptom relief of the attack of the disease. Aim of the study - the antidiarrheal efficacy and safety study of high-dispersion silicon dioxide enterosorbent in tablet dosage form in patients with acute diarrhea. This was randomized, double-blind, placebo-controlled, 4-center study. Acute diarrhea was defined as three and more episodes of watery stool per day either during 48 hours or less before study entry in the patients having normal stool recently. It has been postulated that symptoms and signs of acute diarrhea have to be caused by direct infection of the gastrointestinal tract and did not associated with moderate-to-severe systemic states. 144 patients with established acute diarrhea were randomized into treatment group (enterosorbent “Carbowhite”, n = 120) or placebo group. Date collection including severity diarrhea, systemic symptoms was performed at baseline and daily during 7 days. Stool examination and serological assay were performed at baseline. The primary end points were declared as time to complete recovery from acute diarrhea. It has been found that the use of the siliceous enterosorbent (“Carbowhite”) allowed to reduce (p < 0.001) the treatment period averagely for 0.9 days (95% confidence interval 0.5–1.2 days) in comparison with placebo. Data of safety monitoring has revealed that both patient groups had negative stool culture, while initiation of antibiotic treatment was run more frequently in placebo group (8.3%) compared to investigational product group (4.1%, P = 0.044). The siliceous enterosorbent “Carbowhite” was well tolerated and reduced the recovery time of the acute episode of the diarrhea in the clinically significant form.

The essential role of fructose-1,6-bisphosphatase 2 enzyme in thermal homeostasis upon cold stress

Skeletal muscle is a major organ for glucose disposal and thermogenesis. While hepatic fructose-1,6-bisphosphatase is well known as a key enzyme for gluconeogenesis, the role of muscle fructose-1,6-bisphosphatase 2 (Fbp2) in glucose disposal and thermogenesis is unknown. Here, using Fbp2 knockout (KO) mice, we assessed the physiological role of Fbp2 in energy and glucose metabolism and thermogenesis. In vivo assessments of energy metabolism, glucose metabolism, and thermogenesis were performed by indirect calorimetry, hyperinsulinemic-euglycemic clamp, and cold challenge studies, respectively. Under both feeding and fasting conditions, Fbp2 KO mice showed similar phenotypes regarding energy and glucose metabolism compared to wild-type (WT) mice. However, Fbp2 KO mice were severely intolerant to cold challenge under fasting conditions. Mechanistically, the cold-induced intramuscular conversion of lactate to glycogen (glyconeogenesis) is completely abolished in the KO muscle, which leads to a lack of glycogen source for thermogenesis in Fbp2 KO mice. The cold-intolerant phenotype of KO mice disappeared after feeding, and the KO mice were equally as cold tolerant as the WT mice and survived during the cold challenge for three weeks. Taken together, these data demonstrate that Fbp2 is essential for muscle thermogenesis by replenishing the intramuscular glycogen pool through glyconeogenesis when the exogenous glucose source is limited. These data imply the physiological importance of Fbp2 in thermal homeostasis and suggest a potential novel therapy targeted to increase glycogen replenishment upon cold stress.

Prognostic role of serum high mobility group box 1 concentration in cardiac surgery

Outcomes of cardiac surgery are influenced by systemic inflammation. High mobility group box 1 (HMGB1), a pivotal inflammatory mediator, plays a potential role as a prognostic biomarker in cardiovascular disease. The aim of this prospective, observational study was to investigate the relationship between serum HMGB1 concentrations and composite of morbidity endpoints in cardiac surgery. Arterial blood samples for HMGB1 measurement were collected from 250 patients after anaesthetic induction (baseline) and 1 h after weaning from cardiopulmonary bypass (post-CPB). The incidence of composite of morbidity endpoints (death, myocardial infarction, stroke, renal failure and prolonged ventilator care) was compared in relation to the tertile distribution of serum HMGB1 concentrations. The incidence of composite of morbidity endpoints was significantly different with respect to the tertile distribution of post-CPB HMGB1 concentrations (p = 0.005) only, and not to the baseline. Multivariable analysis revealed post-CPB HMGB1 concentration (OR, 1.072; p = 0.044), pre-operative creatinine and duration of CPB as independent risk factors of adverse outcome. Accounting for its prominent role in mediating sterile inflammation and its relation to detrimental outcome, HMGB1 measured 1 h after weaning from CPB would serve as a useful biomarker for accurate risk stratification in cardiac surgical patients and may guide tailored anti-inflammatory therapy.

Broad Protection of Pigs against Heterologous PRRSV Strains by a GP5-Mosaic DNA Vaccine Prime/GP5-Mosaic rVaccinia (VACV) Vaccine Boost

Background: Porcine reproductive and respiratory syndrome (PRRS) viruses are a major cause of disease and economic loss in pigs worldwide. High genetic diversity among PRRSV strains is problematic for successful disease control by vaccination. Mosaic DNA and vaccinia (VACV) vaccines were developed in order to improve protection against heterologous PRRSV strains. Methods: Piglets were primed and boosted with GP5-Mosaic DNA vaccine and recombinant GP5-Mosaic VACV (rGP5-Mosaic VACV), respectively. Pigs vaccinated with rGP5-WT (VR2332) DNA and rGP5-WT VACV, or empty vector DNA and empty VACV respectively, served as controls. Virus challenge was given to separate groups of vaccinated pigs with VR2332 or MN184C. Necropsies were performed 14 days after challenge. Results: Vaccination with the GP5-Mosaic-based vaccines resulted in cellular reactivity and higher levels of neutralizing antibodies to both VR2332 and MN184C PRRSV strains. In contrast, vaccination of animals with the GP5-WT vaccines induced responses only to VR2332. Furthermore, vaccination with the GP5-Mosaic based vaccines resulted in protection against challenge with two heterologous virus strains, as demonstrated by the significantly lower viral loads in serum, tissues, porcine alveolar macrophages (PAMs), and bronchoalveolar lavage (BAL) fluids, and less severe lung lesions after challenge with either MN184C or VR2332, which have only 85% identity. In contrast, significant protection by the GP5-WT based vaccines was only achieved against the VR2332 strain. Conclusions: GP5-Mosaic vaccines, using a DNA-prime/VACV boost regimen, conferred protection in pigs against heterologous viruses.

MVA Vectored Vaccines Encoding Rift Valley Fever Virus Glycoproteins Protect Mice against Lethal Challenge in the Absence of Neutralizing Antibody Responses

In vitro neutralizing antibodies have been often correlated with protection against Rift Valley fever virus (RVFV) infection. We have reported previously that a single inoculation of sucrose-purified modified vaccinia Ankara (MVA) encoding RVFV glycoproteins (rMVAGnGc) was sufficient to induce a protective immune response in mice after a lethal RVFV challenge. Protection was related to the presence of glycoprotein specific CD8+ cells, with a low-level detection of in vitro neutralizing antibodies. In this work we extended those observations aimed to explore the role of humoral responses after MVA vaccination and to study the contribution of each glycoprotein antigen to the protective efficacy. Thus, we tested the efficacy and immune responses in BALB/c mice of recombinant MVA viruses expressing either glycoprotein Gn (rMVAGn) or Gc (rMVAGc). In the absence of serum neutralizing antibodies, our data strongly suggest that protection of vaccinated mice upon the RVFV challenge can be achieved by the activation of cellular responses mainly directed against Gc epitopes. The involvement of cellular immunity was stressed by the fact that protection of mice was strain dependent. Furthermore, our data suggest that the rMVA based single dose vaccination elicits suboptimal humoral immune responses against Gn antigen since disease in mice was exacerbated upon virus challenge in the presence of rMVAGnGc or rMVAGn immune serum. Thus, Gc-specific cellular immunity could be an important component in the protection after the challenge observed in BALB/c mice, contributing to the elimination of infected cells reducing morbidity and mortality and counteracting the deleterious effect of a subneutralizing antibody immune response.

Wild Birds in Live Birds Markets: Potential Reservoirs of Enzootic Avian Influenza Viruses and Antimicrobial Resistant Enterobacteriaceae in Northern Egypt

Wild migratory birds are often implicated in the introduction, maintenance, and global dissemination of different pathogens, such as influenza A viruses (IAV) and antimicrobial-resistant (AMR) bacteria. Trapping of migratory birds during their resting periods at the northern coast of Egypt is a common and ancient practice performed mainly for selling in live bird markets (LBM). In the present study, samples were collected from 148 wild birds, representing 14 species, which were being offered for sale in LBM. All birds were tested for the presence of AIV and enterobacteriaceae. Ten samples collected from Northern Shoveler birds (Spatula clypeata) were positive for IAV and PCR sub-typing and pan HA/NA sequencing assays detected H5N8, H9N2, and H6N2 viruses in four, four, and one birds, respectively. Sequencing of the full haemagglutinin (HA) gene revealed a high similarity with currently circulating IAV in Egypt. From all the birds, E. coli was recovered from 37.2% and Salmonella from 20.2%, with 66–96% and 23–43% isolates being resistant to at least one of seven selected critically important antimicrobials (CIA), respectively. The presence of enzootic IAV and the wide prevalence of AMR enterobacteriaceae in wild birds highlight the potential role of LBM in the spread of different pathogens from and to wild birds. Continued surveillance of both AIV and antimicrobial-resistant enterobacteriaceae in wild birds’ habitats is urgently needed.

Exploiting B Cell Receptor Analyses to Inform on HIV-1 Vaccination Strategies

The human antibody repertoire is generated by the recombination of different gene segments as well as by processes of somatic mutation. Together these mechanisms result in a tremendous diversity of antibodies that are able to combat various pathogens including viruses and bacteria, or malignant cells. In this review, we summarize the opportunities and challenges that are associated with the analyses of the B cell receptor repertoire and the antigen-specific B cell response. We will discuss how recent advances have increased our understanding of the antibody response and how repertoire analyses can be exploited to inform on vaccine strategies, particularly against HIV-1.

Humanized Mice for Live-Attenuated Vaccine Research: From Unmet Potential to New Promises

Live-attenuated vaccines (LAV) represent one of the most important medical innovations in human history. In the past three centuries, LAV have saved hundreds of millions of lives, and will continue to do so for many decades to come. Interestingly, the most successful LAVs, such as the smallpox vaccine, the measles vaccine, and the yellow fever vaccine, have been isolated and/or developed in a purely empirical manner without any understanding of the immunological mechanisms they trigger. Today, the mechanisms governing potent LAV immunogenicity and long-term induced protective immunity continue to be elusive, and therefore hamper the rational design of innovative vaccine strategies. A serious roadblock to understanding LAV-induced immunity has been the lack of suitable and cost-effective animal models that can accurately mimic human immune responses. In the last two decades, human-immune system mice (HIS mice), i.e., mice engrafted with components of the human immune system, have been instrumental in investigating the life-cycle and immune responses to multiple human-tropic pathogens. However, their use in LAV research has remained limited. Here, we discuss the strong potential of LAVs as tools to enhance our understanding of human immunity and review the past, current and future contributions of HIS mice to this endeavor.

Synthetic DNA Vaccines Adjuvanted with pIL-33 Drive Liver-Localized T Cells and Provide Protection from Plasmodium Challenge in a Mouse Model

The need for a malaria vaccine is indisputable. A single vaccine for Plasmodium pre-erythrocytic stages targeting the major sporozoite antigen circumsporozoite protein (CSP) has had partial success. Additionally, CD8+ T cells targeting liver-stage (LS) antigens induced by live attenuated sporozoite vaccines were associated with protection in human challenge experiments. To further evaluate protection mediated by LS antigens, we focused on exported pre-erythrocytic proteins (exported protein 1 (EXP1), profilin (PFN), exported protein 2 (EXP2), inhibitor of cysteine proteases (ICP), transmembrane protein 21 (TMP21), and upregulated in infective sporozoites-3 (UIS3)) expressed in all Plasmodium species and designed optimized, synthetic DNA (synDNA) immunogens. SynDNA antigen cocktails were tested with and without the molecular adjuvant plasmid IL-33. Immunized animals developed robust T cell responses including induction of antigen-specific liver-localized CD8+ T cells, which were enhanced by the co-delivery of plasmid IL-33. In total, 100% of mice in adjuvanted groups and 71%–88% in non-adjuvanted groups were protected from blood-stage disease following Plasmodium yoelii sporozoite challenge. This study supports the potential of synDNA LS antigens as vaccine components for malaria parasite infection.

Formulation of Colostrum Supplements, Colostrum Replacers and Acquisition of Passive Immunity in Neonatal Calves

Provision of an adequate mass of IgG from maternal colostrum is essential to health and survival of neonatal calves. Colostrum supplements (CS) have been developed to provide supplemental immunoglobulin when maternal colostrum is of poor quality. However, colostrum replacers (CR) that provide ≥100 g of IgG have not been formulated. Our objective was to determine the absorption of IgG in newborn calves fed CS derived from bovine serum or CR derived from bovine immunoglobulin concentrate. The CS were prepared by collecting, processing, and spray drying bovine serum and blending with other ingredients to provide 45 to 50 g of IgG per dose. The CR were prepared by further processing bovine serum to increase IgG concentration to >50% IgG and blending with other ingredients to provide 100 to 122 g of IgG per dose. Holstein calves (n = 160) were fed 90 to 244 g of IgG from CS or CR in 1 or 2 feedings in two experiments. Blood was collected from each calf by jugular venipuncture at 0 and 24 h of age and plasma IgG was determined by turbidimetric immunoassay. Apparent efficiency of IgG absorption was calculated. Plasma IgG concentrations at 24 h of age were indicative of IgG intake and averaged 5.5 to 14.1 g/L in calves fed CS and CR. Mean apparent efficiency of IgG absorption in calves fed CS was 25 and 28% in experiments 1 and 2, respectively. Mean apparent efficiency of IgG absorption in calves fed CR ranged from 19 to 32% and were affected by method of processing and number of times fed. Treatment of plasma with polyethylene glycol reduced the efficiency of IgG absorption in experiment 1. The addition of animal fat to CR had no effect on IgG absorption. A second feeding of CR increased plasma IgG, but efficiency of absorption was reduced. Mean body weights at 60 d of age were not affected by treatment and ranged from 64.3 to 78.2 kg. Plasma IgG concentration in calves fed ≥122 g of IgG from Ig concentrate approached (9.9 g/L) or exceeded 10 g/L, indicating successful transfer of passive immunity. Provision of IgG to prevent failure of passive transfer is possible with CR containing >20% IgG when fed at 454 g per dose.

Inhibition of ribonucleotide reductase and growth of human colon carcinoma HT-29 cells and mouse leukemia L1210 cells by N-hydroxy-N′-aminoguanidine derivatives

A series of N-hydroxy-N′-aminoguanidine (HAG) derivatives were studied and compared for their effects on ribonucleotide reductase activity in cell-free extracts; on nucleic acid synthesis and the growth of human colon carcinoma HT-29 cells; and on mouse leukemia L1210 cells in culture. The HAG derivatives [RCHNNHC(NH)NHOH-tosylate] studied could be grouped as; (1) hydroxy-benzylidines; (2) methoxybenzylidines; and (3) nitrobenzylidines substituted at the R position. 2′-Hydroxybenzylidine-HAG, the lead compound, was relatively active in both HT-29 cells and L1210 cells (20 ± 5 and 13 ± 4 μM for 50% inhibition of HT-29 and L1210 cell growth respectively). The monohydroxybenzylidene compounds were generally more active than the dihydroxy- and trihy-droxybenzylidene-HAG derivatives. The methoxybenzylidene-HAGs were as active as the monohy-droxybenzylidene-HAGs. 2′-Hydroxy-4′-methoxybenzylidene-HAG was much more active than 2′,4′-dihydroxybenzylidene-HAG. The mononitrobenzylidene-HAGs were more active than the dinitro-benzylidene-HAG compound. In general, L1210 cells were more sensitive to the effects of the HAG compounds than were HT-29 cells. There was good agreement between the concentration of drug required to inhibit the growth of HT-29 cells and that required to inhibit the growth of L1210 cells. There was also good correlation between the ability of HAG derivatives to inhibit ribonucleotide reductase activity and to inhibit tumor cell growth. Some derivatives, such as 2′,3′,4′- and 3′,4′,5′-trihydroxybenzylidene-HAG inhibited L1210 cell growth by 50% at lower concentrations (7.8 and 11.9 μM respectively) than the concentrations needed for 50% inhibition of HT-29 cell growth (196 and 234 μM respectively) and ribonucleotide reductase activity (122 and 188 μM respectively). The studies of nucleic acid synthesis in L1210 cells using [(3)H]cytidine as a precursor showed that 2′,3′,4′-trihy-droxybenzylidine-HAG inhibited DNA synthesis at a lower concentration (29 μM for 50% inhibition) than was needed for the inhibition of RNA synthesis and formation of [(3)H]deoxycytidine nucleotides in the acid-soluble fraction (320 and 820 μM for 50% inhibition respectively). These results indicate that 2′,3′,4′-trihydroxybenzylidine-HAG inhibits DNA synthesis in L1210 cells through other mechanisms rather than exclusively through the inhibition of ribonucleotide reductase activity.

I, 2. Physiology and pathophysiology of the gut in relation to viral diarrhea

Many advances have been made in the understanding of intestinal electrolyte transport from the molecular to the whole-tissue level. This chapter discusses the molecular mechanisms of intestinal epithelial ion transport processes, as well as the intra- and extracellular factors involved in their regulation, as a framework for the understanding of virus-induced gastroenteritis. Based on the present knowledge of the effects of rotavirus (RV) infection on the physiology of the intestine at different levels of organization, a working model for the pathogenesis of RV diarrhea is presented in the chapter. The understanding of the pathogenic processes of viral diarrheas may serve as the basis for a rational approach in the design of novel therapeutic strategies and the search for new antiviral drugs.

The uses and limitations of a hand-held germicidal ultraviolet wand for surface disinfection

The morbidity and mortality from healthcare associated infections has raised concern that conventional disinfection methods are inadequate and that other adjunct methods such as room fumigation and ultraviolet irradiation may be needed. There is also concern that these alternative methods may pose a risk to workers and patients. Objectives. (1) Determine the efficacy of a germicidal UV-C wand for surface disinfection, (2) evaluate changing relative humidity (RH) and different target distances on bacteria kill rates, and (3) assess potential exposure concerns. Methods. This study investigates whether a hand-held germicidal wand can efficaciously disinfect surfaces treated with either a vegetative or spore forming bacterium and to evaluate the effect of changing environmental conditions such as relative humidity (RH), target position, and target distances on microbial kill rates. Results. Kill rate was best at 40–65% RH at a temperature range of 21–24°C. Both high and low RH interfered with the ability of UV-C to kill the vegetative microbe. In the case of the spore forming bacterium, increased surface drying time was the most significant factor increasing kill rate. Conclusions. This research demonstrates that UV-C was efficacious under optimal conditions, a direct beam exposure, and a short target distance (12.7 cm). However, there are limitations when used in non-optimal conditions. Increased distance and indirect beam angles resulted in lower kill rates. It is also important to minimize unnecessary patient and worker exposure during its use.

Dapsone as treatment adjunct in ARDS

Multiple pharmacological interventions tested over the last decades have failed to reduce ARDS mortality. This short note recounts past data indicating that (i) neutrophils home along an IL-8 gradient, (ii) in ARDS, massive neutrophil accumulation and degranulation in and along bronchoalveolar spaces contributes to damage and hypoxia, (iii) large increases in IL-8 are one of the chemotaxic signals drawing neutrophils to the ARDS lung, and (iv) old data from dermatology and glioblastoma research showed that the old drug against Hansen’s disease, dapsone, inhibits neutrophils’ chemotaxis to IL-8. Therefore dapsone might lower neutrophils’ contributions to ARDS lung pathology. Dapsone can create methemoglobinemia that although rarely problematic it would be particularly undesirable in ARDS. The common antacid drug cimetidine lowers risk of dapsone related methemoglobinemia and should be given concomitantly.

A comparison of facemask and respirator filtration test methods

NIOSH published a Federal Register Notice to explore the possibility of incorporating FDA required filtration tests for surgical masks (SMs) in the 42 CFR Part 84 respirator certification process. There have been no published studies comparing the filtration efficiency test methods used for NIOSH certification of N95 filtering facepiece respirators (N95 FFRs) with those used by the FDA for clearance of SMs. To address this issue, filtration efficiencies of “N95 FFRs” including six N95 FFR models and three surgical N95 FFR models, and three SM models were measured using the NIOSH NaCl aerosol test method, and FDA required particulate filtration efficiency (PFE) and bacterial filtration efficiency (BFE) methods, and viral filtration efficiency (VFE) method. Five samples of each model were tested using each method. Both PFE and BFE tests were done using unneutralized particles as per FDA guidance document. PFE was measured using 0.1 µm size polystyrene latex particles and BFE with ∼3.0 µm size particles containing Staphylococcus aureus bacteria. VFE was obtained using ∼3.0 µm size particles containing phiX 174 as the challenge virus and Escherichia coli as the host. Results showed that the efficiencies measured by the NIOSH NaCl method for “N95 FFRs” were from 98.15–99.68% compared to 99.74–99.99% for PFE, 99.62–99.9% for BFE, and 99.8–99.9% for VFE methods. Efficiencies by the NIOSH NaCl method were significantly (p = <0.05) lower than the other methods. SMs showed lower efficiencies (54.72–88.40%) than “N95 FFRs” measured by the NIOSH NaCl method, while PFE, BFE, and VFE methods produced no significant difference. The above results show that the NIOSH NaCl method is relatively conservative and is able to identify poorly performing filtration devices. The higher efficiencies obtained using PFE, BFE and VFE methods show that adding these supplemental particle penetration methods will not improve respirator certification.

Effect of multiple alcohol-based hand rub applications on the tensile properties of thirteen brands of medical exam nitrile and latex gloves

Current CDC guidance for the disinfection of gloved hands during the doffing of personal protective equipment (PPE) following the care of a patient with Ebola recommends for multiple applications of alcohol-based hand rub (ABHR) on medical exam gloves. To evaluate possible effects of ABHR applications on glove integrity, thirteen brands of nitrile and latex medical exam gloves from five manufacturers and two different ABHRs were included in this study. A pair of gloves were worn by a test operator and the outside surfaces of the gloves were separately treated with an ABHR for 1–6 applications. Tensile strength and ultimate elongation of the gloves without any ABHR treatments (control gloves) and gloves after 1–6 ABHR applications were measured based on the ASTM D412 standard method. In general, tensile strength decreased with each ABHR application. ABHRs had more effect on the tensile strength of the tested nitrile than latex gloves, while ethanol-based ABHR (EBHR) resulted in lesser changes in tensile strength compared to isopropanol-based ABHR (IBHR). The results show that multiple EBHR applications on the latex gloves and some of the nitrile gloves tested should be safe for Ebola PPE doffing based on the CDC guidance. Appropriate hospital staff practice using ABHR treatment and doffing gloves is recommended to become more familiar with changes in glove properties.

Mobile Phones as a Potential Vehicle of Infection in a Hospital Setting

The objective of this article is to investigate the potential role of mobile phones as a reservoir for bacterial colonization and the risk factors for bacterial colonization in a hospital setting. We screened 226 staff members at a regional Australian hospital (146 doctors and 80 medical students) between January 2013 and March 2014. The main outcomes of interest were the types of microorganisms and the amount of contamination of the mobile phones. This study found a high level of bacterial contamination (n = 168/226, 74%) on the mobile phones of staff members in a tertiary hospital, with similar organisms isolated from the staff member's dominant hand and mobile phones. While most of the isolated organisms were normal skin flora, a small percentage were potentially pathogenic (n = 12/226, 5%). Being a junior medical staff was found to be a risk factor for heavy microbial growth (OR 4.00, 95% CI 1.54, 10.37). Only 31% (70/226) of our participants reported cleaning their phones routinely, and only 21% (47/226) reported using alcohol containing wipes on their phones. This study demonstrates that mobile phones are potentially vehicles for pathogenic bacteria in a hospital setting. Only a minority of our participants reported cleaning their phones routinely. Disinfection guidelines utilizing alcohol wipes should be developed and implemented.

Validation of in-house liquid direct agglutination test antigen: the potential diagnostic test in visceral Leishimaniasis endemic areas of Northwest Ethiopia

BACKGROUND: Visceral leishmaniasis in Ethiopia is a re-emerging threat to public health, with increased geographical distribution and number of cases. It is a fatal disease without early diagnosis and treatment; thus, the availability of affordable diagnostic tools is crucial. However, due to delays caused by import regulations, procurement and late delivery of imported test kits, accessibility remains a problem in the control program. Therefore, we aimed to produce and evaluate the performance of an in-house liquid (AQ) direct agglutination test (DAT) antigen. RESULT: The AQ-DAT was produced at the Armauer Hansen Research Institute, using Leishmania donovani strain (MHOM/ET/67/L82). Sera from 272 participants; 110 microscopically confirmed cases of VL, 76 apparently healthy and 86 patients who had infectious disease other than VL were tested with AQ-DAT, and standard kits: Freeze-dried DAT (FD-DAT) and rK39. Taking microscopy as a gold standard; the sensitivity and specificity of the AQ-DAT were 97.3 and 98.8%, respectively. It had high degrees of agreement (k > 0.8), with a significant (P < 0.05) correlation compared to microscopy, FD-DAT, and rK39. CONCLUSION: Although further standardization is required, the in-house AQ-DAT could improve diagnostic accessibility, minimize intermittent stock outs and strengthen the national VL control program.

Clinical outcomes of empirical high-dose meropenem in critically ill patients with sepsis and septic shock: a randomized controlled trial

BACKGROUND: Appropriate antimicrobial dosing is challenging because of changes in pharmacokinetics (PK) parameters and an increase in multidrug-resistant (MDR) organisms in critically ill patients. This study aimed to evaluate the effects of an empirical therapy of high-dose versus standard-dose meropenem in sepsis and septic shock patients. METHODS: We performed a prospective randomized open-label study to compare the changes of modified sequential organ failure assessment (mSOFA) score and other clinical outcomes of the high-dose meropenem (2-g infusion over 3 h every 8 h) versus the standard-dose meropenem (1-g infusion over 3 h every 8 h) in sepsis and septic shock patients. Patients’ characteristics, clinical and microbiological outcomes, 14 and 28-day mortality, vasopressor- and ventilator-free days, intensive care unit (ICU) and hospital-free days, percent of the time of antibiotic concentrations above the minimum inhibitory concentration (%T>MIC), and safety were assessed. RESULTS: Seventy-eight patients were enrolled. Median delta mSOFA was comparable between two groups (– 1 in the high-dose group vs. – 1 in the standard-dose group; P value = 0.75). There was no difference between the two groups regarding clinical and microbiological cure, 14- and 28-day mortality, vasopressor- and ventilator-free days, and ICU- and hospital-free days. In patients admitted from the emergency department (ED) with a mSOFA score ≥ 7, the high-dose group demonstrated significantly better microbiological cure compared with the standard-dose group (75% (9/12 patients) vs. 20% (2/10 patients); P value = 0.03). Likewise, the high-dose group presented higher microbiological cure rate in patients admitted from ED who had either APACHE II score > 20 (83.3% (10/12) vs. 28.6% (2/7); P value = 0.045) or on mechanical ventilator (87.5% (7/8) vs. 23.1% (3/13); P value = 0.008) than the standard-dose group. Adverse events were comparable between the two groups. CONCLUSIONS: Empirical therapy with the high-dose meropenem presented comparable clinical outcomes to the standard-dose meropenem in sepsis and septic shock patients. Besides, subgroup analysis manifested superior microbiological cure rate in sepsis or septic shock patients admitted from ED. TRIAL REGISTRATION: ClinicalTrials.gov, NCT03344627, registered on November 17, 2017

Spinal cord autoregulation using near-infrared spectroscopy under normal, hypovolemic, and post-fluid resuscitation conditions in a swine model: a comparison with cerebral autoregulation

BACKGROUND: Few studies have investigated spinal cord autoregulation using near-infrared spectroscopy (NIRS). Here, we assessed spinal cord autoregulation under normal, hypovolemic, and post-fluid resuscitation conditions compared with cerebral autoregulation. METHODS: Ten pigs (36.1 ± 1.1 kg) were anesthetized with 2.5% isoflurane, before phenylephrine administration at 0.5, 1, 2, and 5 μg kg(−1) min(−1) in a stepwise fashion at 10-min intervals (baseline), followed by similar administration of sodium nitroprusside (SNP). Hypovolemia was induced by a 600-ml bleed (25% estimated total blood volume). Only phenylephrine was readministered (same protocol). Hypovolemia was reversed by infusing 600 ml hydroxyethyl starch, before readministering phenylephrine and SNP. The relationships between mean arterial pressure (MAP) and cerebral, thoracic, and lumbar spinal cord tissue oxygenation indices (TOIs) were evaluated. RESULTS: Thoracic and lumbar spinal cord TOIs were approximately 15% and 10% lower, respectively, than the cerebral TOI at similar MAPs. The average relationship between MAP and each TOI showed an autoregulatory pattern, but negative correlations were observed in the cerebral TOI during phenylephrine infusion. A 600-ml bleed lowered each relationship < 5% and subsequent fluid resuscitation did not change the relationship. Individual oxygenation responses to blood pressure indicated that the spinal cord is more pressure-passive than the cerebrum. Paradoxical responses (an inverse relationship of tissue oxygenation to MAP) were observed particularly in cerebrum during phenylephrine infusion and were rare in the spinal cord. CONCLUSIONS: Spinal cord autoregulation is less robust than cerebral autoregulation and more pressure-dependent. Similar to cerebral oxygenation, spinal cord oxygenation is volume-tolerant but is more sensitive to hypotension.

Recombinant lipidated Zika virus envelope protein domain III elicits durable neutralizing antibody responses against Zika virus in mice

BACKGROUND: The emergence of Zika virus (ZV) in tropical and subtropical areas of the world has created an urgent need for vaccines against ZV. However, approved vaccines that prevent ZV infection are not available. To develop an effective vaccine against ZV infection, a lipidated form of ZV envelope protein domain III that possesses an intrinsic adjuvant property was rationally designed. Our goal was to examine the immunogenicity of recombinant lipidated ZV envelope protein domain III (rLZE3) and evaluate its potential as a vaccine candidate against ZV. METHODS: Recombinant ZV envelope protein domain III (rZE3) and rLZE3 were prepared with an Escherichia coli-based system. Dendritic cell surface marker expression and cytokine production upon stimulation were analyzed to evaluate the function of rLZE3. Neutralizing antibody capacities were evaluated using focus reduction neutralization tests after immunization. To investigate the protective immunity in immunized mice, serum samples collected from immunized mice were adoptively transferred into AG129 mice, and then viremia levels and survival times were examined after ZV challenge. RESULTS: rLZE3 alone but not rZE3 alone efficiently activated dendritic cells in vitro and was taken up by dendritic cells in vivo. Immunization of C57BL/6 mice with rLZE3 alone (without exogenous adjuvant) could induce ZV-specific neutralizing antibody responses. Furthermore, serum samples obtained from rLZE3-immunized mice provided protection as indicated by a reduction in viremia levels and prolongation of survival times after ZV challenge. CONCLUSION: These results indicate that rLZE3 is an excellent vaccine candidate and has great potential that should be evaluated in further preclinical studies.

Forecasting the 2017/2018 seasonal influenza epidemic in England using multiple dynamic transmission models: a case study

BACKGROUND: Since the 2009 A/H1N1 pandemic, Public Health England have developed a suite of real-time statistical models utilising enhanced pandemic surveillance data to nowcast and forecast a future pandemic. Their ability to track seasonal influenza and predict heightened winter healthcare burden in the light of high activity in Australia in 2017 was untested. METHODS: Four transmission models were used in forecasting the 2017/2018 seasonal influenza epidemic in England: a stratified primary care model using daily, region-specific, counts and virological swab positivity of influenza-like illness consultations in general practice (GP); a strain-specific (SS) model using weekly, national GP ILI and virological data; an intensive care model (ICU) using reports of ICU influenza admissions; and a synthesis model that included all data sources. For the first 12 weeks of 2018, each model was applied to the latest data to provide estimates of epidemic parameters and short-term influenza forecasts. The added value of pre-season population susceptibility data was explored. RESULTS: The combined results provided valuable nowcasts of the state of the epidemic. Short-term predictions of burden on primary and secondary health services were initially highly variable before reaching consensus beyond the observed peaks in activity between weeks 3–4 of 2018. Estimates for R(0) were consistent over time for three of the four models until week 12 of 2018, and there was consistency in the estimation of R(0) across the SPC and SS models, and in the ICU attack rates estimated by the ICU and the synthesis model. Estimation and predictions varied according to the assumed levels of pre-season immunity. CONCLUSIONS: This exercise successfully applied a range of pandemic models to seasonal influenza. Forecasting early in the season remains challenging but represents a crucially important activity to inform planning. Improved knowledge of pre-existing levels of immunity would be valuable.

Infektionsschutzrecht nach Inkrafttreten des Masernschutzgesetzes

On 1 March 2020, the amendments to the German Protection Against Infection Act that were introduced by the act to protect against measles and strengthen vaccination prevention (Measles Protection Act) entered into force. The reason for the changes is that the number of individuals with measles has significantly increased in recent years. To protect public health, the Measles Protection Act has implemented regulations requiring that persons in certain institutions must either have adequate protection against measles or have immunity to measles. In this article the current legal situation with regard to health care facilities is presented.

CpG Dinucleotides Inhibit HIV-1 Replication through Zinc Finger Antiviral Protein (ZAP)-Dependent and -Independent Mechanisms

CpG dinucleotides are suppressed in the genomes of many vertebrate RNA viruses, including HIV-1. The cellular antiviral protein ZAP (zinc finger antiviral protein) binds CpGs and inhibits HIV-1 replication when CpGs are introduced into the viral genome. However, it is not known if ZAP-mediated restriction is the only mechanism driving CpG suppression. To determine how CpG dinucleotides affect HIV-1 replication, we increased their abundance in multiple regions of the viral genome and analyzed the effect on RNA expression, protein abundance, and infectious-virus production. We found that the antiviral effect of CpGs was not correlated with their abundance. Interestingly, CpGs inserted into some regions of the genome sensitize the virus to ZAP antiviral activity more efficiently than insertions into other regions, and this sensitivity can be modulated by interferon treatment or ZAP overexpression. Furthermore, the sensitivity of the virus to endogenous ZAP was correlated with its sensitivity to the ZAP cofactor KHNYN. Finally, we show that CpGs in some contexts can also inhibit HIV-1 replication by ZAP-independent mechanisms, and one of these is the activation of a cryptic splice site at the expense of a canonical splice site. Overall, we show that the location and sequence context of the CpG in the viral genome determines its antiviral activity. IMPORTANCE Some RNA virus genomes are suppressed in the nucleotide combination of a cytosine followed by a guanosine (CpG), indicating that they are detrimental to the virus. The antiviral protein ZAP binds viral RNA containing CpGs and prevents the virus from multiplying. However, it remains unknown how the number and position of CpGs in viral genomes affect restriction by ZAP and whether CpGs have other antiviral mechanisms. Importantly, manipulating the CpG content in viral genomes could help create new vaccines. HIV-1 shows marked CpG suppression, and by introducing CpGs into its genome, we show that ZAP efficiently targets a specific region of the viral genome, that the number of CpGs does not predict the magnitude of antiviral activity, and that CpGs can inhibit HIV-1 gene expression through a ZAP-independent mechanism. Overall, the position of CpGs in the HIV-1 genome determines the magnitude and mechanism through which they inhibit the virus.

Nanoplasmid Vectors Co-expressing Innate Immune Agonists Enhance DNA Vaccines for Venezuelan Equine Encephalitis Virus and Ebola Virus

DNA vaccines expressing codon-optimized Venezuelan equine encephalitis virus (VEEV) and Ebola virus (EBOV) glycoprotein genes provide protective immunity to mice and nonhuman primates when delivered by intramuscular (IM) electroporation (EP). To achieve equivalent protective efficacy in the absence of EP, we evaluated VEEV and EBOV DNA vaccines constructed using minimalized Nanoplasmid expression vectors that are smaller than conventional plasmids used for DNA vaccination. These vectors may also be designed to co-express type I interferon inducing innate immune agonist genes that have an adjuvant effect. Nanoplasmid vaccinated mice had increased antibody responses as compared to those receiving our conventional pWRG7077-based vaccines when delivered by IM injection, and these responses were further enhanced by the inclusion of the innate immune agonist genes. The Nanoplasmid VEEV DNA vaccines also significantly increased protection against aerosol VEEV challenge as compared to the pWRG7077 VEEV DNA vaccine. Although all mice receiving the pWRG7077 and Nanoplasmid EBOV DNA vaccines at the dose tested survived EBOV challenge, only mice receiving the Nanoplasmid EBOV DNA vaccine that co-expresses the innate immune agonist genes failed to lose weight after challenge. Our results suggest that Nanoplasmid vectors can improve the immunogenicity and protective efficacy of alphavirus and filovirus DNA vaccines.

Anaerobic antibiotic usage for pneumonia in the medical intensive care unit

BACKGROUND AND OBJECTIVE: Pneumonia is a common admitting diagnosis in the intensive care unit (ICU). When aspiration is suspected, antibiotics to cover anaerobes are frequently used, but in the absence of clear risk factors, current guidelines have questioned their role. It is unknown how frequently these guidelines are followed. METHODS: We conducted a single‐centre observational study on practice patterns of anaerobic antibiotic use in consecutive patients admitted to the ICU with aspiration pneumonia (Asp), community‐acquired pneumonia (CAP) and healthcare‐associated pneumonia (HCAP). RESULTS: A total of 192 patients were studied (Asp: 20, HCAP: 107, CAP: 65). Overall, 59 patients received anaerobic antibiotics (Asp: 90%, HCAP: 28%, CAP 17%) but a significant proportion of these patients did not meet criteria to receive them. Inappropriate anaerobic antibiotic use was 12/20 for Asp, 27/107 for HCAP and 9/65 for CAP. Mortality probability model III at zero hours (MPM0) score and a diagnosis of Asp were predictors of receiving inappropriate anaerobic antibiotics. Receiving inappropriate anaerobic antibiotics was associated with a longer ICU length of stay (LOS; 7 days (interquartile range (IQR): 7–21) vs 4 days (IQR:2–9), P = 0.017). CONCLUSION: For patients in the ICU admitted with pneumonia, there is a high occurrence of inappropriately prescribed anaerobic antibiotics, the use of which was associated with a longer ICU LOS.

Enteritis in sheep, goats and pigs due to Yersinia pseudotuberculosis infection

SUMMARY The features of naturally occurring Yersinia pseudotuberculosis serotype III infections in 16 sheep, one goat and 3 pigs, and Y. pseudotuberculosis serotype I infections in 3 goats, are described. Affected animals usually had diarrhoea and were in poor condition or emaciated. A number were moribund or dead when submitted for necropsy. Thickening of the caecal and colonic mucosa was the only gross lesion attributable to Y. pseudotuberculosis infection, with liver or other visceral abscesses not being seen. Characteristic microabscesses were demonstrated in the intestinal mucosa of 10 sheep, one goat and one pig infected with Y. pseudotuberculosis serotype III and one goat infected with Y. pseudotuberculosis serotype I. Sheep, goats and pigs dosed orally with Y. pseudotuberculosis serotype III, the serotype isolated most commonly from these species, developed intestinal infection. In sheep and pigs, infection was accompanied by diarrhoea. Haematological changes and specific antibodies were elicited in all 3 species in response to infection. Microabscesses were seen in the intestinal mucosa of all experimentally exposed animals. The occurrence of field cases and the results of experimental exposure confirm that Y. pseudotuberculosis serotype III is an enteropathogen of sheep, goats and pigs. The association of Y. pseudotuberculosis serotype I with lesions in a goat, indicates that this bacterium may also be a pathogen of this species. It is concluded that Y. pseudotuberculosis serotype III is an enteric pathogen of a wide range of ungulate species including cattle, buffalo, deer, antelopes, sheep, goats and pigs. Serotypes I and II, while having a more restricted host range, are probably also pathogens of ungulates and, in particular, deer, antelopes and goats.

ISOLATION OF ROTAVIRUS FROM FOALS WITH DIARRHOEA

A rotavirus, morphologically similar to other known rotaviruses, was demonstrated in the faeces of 5 foals with diarrhoea on two properties. Four of these 5 samples produced specific intracytoplasmic fluorescence in cell culture when reacted with calf rotavirus antiserum conjugate. Sixteen affected foals from both properties were depressed, did not suckle and became recumbent. Most had a watery diarrhoea which lasted for 3 days and resulted in some dehydration and loss of body condition. Sick foals were separated from their mothers following the onset of diarrhoea and given fluid therapy and antibiotics, but despite these measures 4 of 12 affected foals on one property died. Acute and convalescent sera were collected from 6 foals, and in each case a rise in the titres of complement fixing and neutralising antibodies was demonstrated. Faecal filtrates containing foal rotaviruses were fed to gnotobiotic piglets and the effects compared with those of other rotaviruses. Viral isolates from both properties produced an asymptomatic infection in piglets. This contrasts with the other rotaviruses isolated in Australia that have been shown to cause diarrhoea in gnotobiotic piglets.

Intelligent Adversary Risk Analysis: A Bioterrorism Risk Management Model

The tragic events of 9/11 and the concerns about the potential for a terrorist or hostile state attack with weapons of mass destruction have led to an increased emphasis on risk analysis for homeland security. Uncertain hazards (natural and engineering) have been successfully analyzed using probabilistic risk analysis (PRA). Unlike uncertain hazards, terrorists and hostile states are intelligent adversaries who can observe our vulnerabilities and dynamically adapt their plans and actions to achieve their objectives. This article compares uncertain hazard risk analysis with intelligent adversary risk analysis, describes the intelligent adversary risk analysis challenges, and presents a probabilistic defender–attacker–defender model to evaluate the baseline risk and the potential risk reduction provided by defender investments. The model includes defender decisions prior to an attack; attacker decisions during the attack; defender actions after an attack; and the uncertainties of attack implementation, detection, and consequences. The risk management model is demonstrated with an illustrative bioterrorism problem with notional data.

Alterations in T (1) of normal and reperfused infarcted myocardium after Gd‐BOPTA versus GD‐DTPA on inversion recovery EPI

This study tested whether Gd‐BOPTA/Dimeg or Gd‐DTPA exerts greater relaxation enhancement for blood and reperfused infarcted myocardium. Relaxivity of Gd‐BOPTA is increased by weak binding to serum albumin. Thirty‐six rats were subjected to reperfused infarction before contrast (doses = 0.05, 0.1, and 0.2 mmol/kg). ΔR1 was repeatedly measured over 30 min. Gd‐BOPTA caused greater ΔR1 for blood and myocardium than did Gd‐DTPA clearance of both agents from normal and infarcted myocardium was similar to blood clearance; plots of ΔR1myocardium/ΔR1blood showed equilibrium phase contrast distribution. Fractional contrast agent distribution volumes were approximately 0.24 for both agents in normal myocardium, 0.98 and 1.6 for Gd‐DTPA and Gd‐BOPTA, respectively, in reperfused infarction. The high value for Gd‐BOPTA was ascribed to greater relaxivity in infarction versus blood. It was concluded that Gd‐BOPTA/Dimeg causes a greater ΔR1 than Gd‐DTPA in regions which contain serum albumin.

Species groups can be transferred across different scales

Aim To test whether species groups (i.e. assemblages of species co‐occurring in nature) that are statistically derived at one scale (broad, medium, or fine scale) can be transferred to another scale, and to identify the driving forces that determine species groups at the various scales. Location Northern Bohemia (Czech Republic, central Europe) in the Ještědský hřbet mountain range and its neighbourhood. Methods Three data sets were sampled: a floristic data set at the broad scale, another floristic data set at the intermediate scale, and a vegetation data set at the habitat scale. First, in each data set, species groups were produced by the COCKTAIL algorithm, which ensures maximized joint occurrence in the data set using a fidelity coefficient. Corresponding species groups were produced in the individual data sets by employing the same species for starting the algorithm. Second, the species groups formed in one data set, i.e. at a particular scale, were applied crosswise to the other data sets, i.e. to the other scales. Correspondence of a species group formed at a particular scale with a species group at another scale was determined. Third, to highlight the driving factors for the distribution of the plant species groups at each scale, canonical correspondence analysis was carried out. Results Twelve species groups were used to analyse the transferability of the groups across the three scales, but only six of them were found to be common to all scales. Correspondence of species groups derived from the finest scale with those derived at the broadest scale was, on average, higher than in the opposite direction. Forest (tree layer) cover, altitude and bedrock type explained most of the variability in canonical correspondence analysis across all scales. Main conclusions Transferability of species groups distinguished at a fine scale to broader scales is better than it is in the opposite direction. Therefore, a possible application of the results is to use species groups to predict the potential occurrence of missing species in broad‐scale floristic surveys from fine‐scale vegetation‐plot data.

Development of a lateral flow device for in‐field detection and evaluation of PCR‐based diagnostic methods for Xanthomonas campestris pv. musacearum, the causal agent of banana xanthomonas wilt

Xanthomonas campestris pv. musacearum (Xcm) is the causal agent of banana xanthomonas wilt, a major threat to banana production in eastern and central Africa. The pathogen is present in very high levels within infected plants and can be transmitted by a broad range of mechanisms; therefore early specific detection is vital for effective disease management. In this study, a polyclonal antibody (pAb) was developed and deployed in a lateral flow device (LFD) format to allow rapid in‐field detection of Xcm. Published Xcm PCR assays were also independently assessed: only two assays gave specific amplification of Xcm, whilst others cross‐reacted with non‐target Xanthomonas species. Pure cultures of Xcm were used to immunize a rabbit, the IgG antibodies purified from the serum and the resulting polyclonal antibodies tested using ELISA and LFD. Testing against a wide range of bacterial species showed the pAb detected all strains of Xcm, representing isolates from seven countries and the known genetic diversity of Xcm. The pAb also detected the closely related Xanthomonas axonopodis pv. vasculorum (Xav), primarily a sugarcane pathogen. Detection was successful in both naturally and experimentally infected banana plants, and the LFD limit of detection was 10(5) cells mL(−1). Whilst the pAb is not fully specific for Xcm, Xav has never been found in banana. Therefore the LFD can be used as a first‐line screening tool to detect Xcm in the field. Testing by LFD requires no equipment, can be performed by non‐scientists and is cost‐effective. Therefore this LFD provides a vital tool to aid in the management and control of Xcm.

The B‐cell system of human mucosae and exocrine glands

Summary: The mucosae and exocrine glands harbour the largest activated B‐cell system of the body, amounting to some 80–90% of all immunoglobulins (Ig)‐producing cells. The major product of these immunocytes is polymeric (p)IgA (mainly dimers) with associated J chain. Both pIgA and pentameric IgM contain a binding site for the polymeric Ig receptor (pIgR), or secretory component (SC), which is a requirement for their active external transport through secretory epithelia. The pIgR/SC binding site depends on covalent incorporation of the J chain into the quaternary structure of the polymers when they are produced by the local immunocytes. This important differentiation characteristic appears to be sufficient functional justification for the J chain to be expressed also by most B cells terminating at secretory effector sites with IgD or IgG production; they probably represent a ‘spin‐off’ from sequential downstream C(H) switching on its way to pIgA expression, thus apparently reflecting a maturational stage of effector B‐cell clones compatible with homing to these sites. Observations in IgA‐deficient individuals suggest that the magnitude of this homing is fairly well maintained even when the differentiation pathway to IgA is blocked. Certain microenvironmental elements such as specific cytokines and dendritic cells appear to be required for induction of IgA synthesis, but it remains virtually unknown why this isotype normally is such a dominating product of local immunocytes and why they have such a high level of J chain expression. Also, despite the recent identification of some important requirements in terms of adhesion molecules (e.g. integrin α4β7 and MAdCAM‐1) that explain the “gut‐seeking” properties of enterically induced B cells, the origin of regionalized homing of B cells to secretory effector sites outside the gut remains elusive. Moreover, little is known about immune regulation underlying the striking disparity of both the class (IgD, IgM) and subclass (IgA1, IgA2, IgGI, IgG2) production patterns shown by local iinmttnocytes in various regions of the body, although the topical microbiota and other environmental stimuli might be important. Rational design of local vaccines will depend on better knowledge of both inductive and migratory properties of human mucosal B cells.

A contour‐based topographic model for hydrological and ecological applications

A digital model for discretizing three‐dimensional terrain into small irregularly shaped polygons or elements based on contour lines and their orthogonals is described. From this subdivision the model estimates a number of topographic attributes for each element including the total upslope contributing area, element area, slope, and aspect. This form of discretization of a catchment produces natural units for problems involving water flow as either a surface or subsurface flow phenomenon. The model therefore has wide potential application for representing the three‐dimensionality of natural terrain and water flow processes in the fields of hydrology, sedimentology, and geomorphology. Three example applications are presented and discussed. They are the prediction of zones of surface saturation, the prediction of the distribution of potential daily solar radiation, and the prediction of zones of erosion and deposition in a catchment.

Recent developments in CE and CEC of peptides

The article brings a comprehensive survey of recent developments and applications of high‐performance capillary electromigration methods, zone electrophoresis, ITP, IEF, affinity electrophoresis, EKC, and electrochromatography, to analysis, preparation, and physicochemical characterization of peptides. New approaches to the theoretical description and experimental verification of electromigration behavior of peptides and to methodology of their separations, such as sample preparation, adsorption suppression, and detection, are presented. Novel developments in individual CE and CEC modes are shown and several types of their applications to peptide analysis are presented: conventional qualitative and quantitative analysis, purity control, determination in biomatrices, monitoring of chemical and enzymatical reactions and physical changes, amino acid and sequence analysis, and peptide mapping of proteins. Some examples of micropreparative peptide separations are given and capabilities of CE and CEC techniques to provide important physicochemical characteristics of peptides are demonstrated.

Loss-of-function tolerance of enhancers in the human genome

Previous studies have surveyed the potential impact of loss-of-function (LoF) variants and identified LoF-tolerant protein-coding genes. However, the tolerance of human genomes to losing enhancers has not yet been evaluated. Here we present the catalog of LoF-tolerant enhancers using structural variants from whole-genome sequences. Using a conservative approach, we estimate that individual human genomes possess at least 28 LoF-tolerant enhancers on average. We assessed the properties of LoF-tolerant enhancers in a unified regulatory network constructed by integrating tissue-specific enhancers and gene-gene interactions. We find that LoF-tolerant enhancers tend to be more tissue-specific and regulate fewer and more dispensable genes relative to other enhancers. They are enriched in immune-related cells while enhancers with low LoF-tolerance are enriched in kidney and brain/neuronal stem cells. We developed a supervised learning approach to predict the LoF-tolerance of all enhancers, which achieved an area under the receiver operating characteristics curve (AUROC) of 98%. We predict 3,519 more enhancers would be likely tolerant to LoF and 129 enhancers that would have low LoF-tolerance. Our predictions are supported by a known set of disease enhancers and novel deletions from PacBio sequencing. The LoF-tolerance scores provided here will serve as an important reference for disease studies.

Far‐Red Fluorescent Lipid‐Polymer Probes for an Efficient Labeling of Enveloped Viruses

Far‐red emitting fluorescent lipid probes are desirable to label enveloped viruses, for their efficient tracking by optical microscopy inside autofluorescent cells. Most used probes are rapidly released from membranes, leading to fluorescence signal decay and loss of contrast. Here, water‐soluble lipid‐polymer probes are synthesized harboring hydrophilic or hydrophobic far‐red emitting dyes, and exhibiting enhanced brightness. They efficiently label Hepatitis C Virus pseudotyped particles (HCVpp), more stably and reproducibly than commercial probes, and a strong fluorescence signal is observed with a high contrast. Labeling with such probes do not alter virion morphology, integrity, nor infectivity. Finally, it is shown by fluorescence microscopy that these probes enable efficient tracking of labeled HCVpp inside hepatocarcinoma cells used as model hepatocytes, in spite of their autofluorescence up to 700 nm. These novel fluorescent lipid‐polymer probes should therefore enable a better characterization of early stages of infection of autofluorescent cells by enveloped viruses.

Attribute‐based classification of European hydrophytes and its relationship to habitat utilization

1.. Here we classify selected European hydrophytes into ‘attribute groups’ based on the possession of homogenous sets of characteristics, and explore the correspondence between these attribute groups, or individual attributes, and habitat use. 2.. Non‐hierarchical clustering was used to assign 120 species to twenty groups based on a matrix of categorical scores for literature‐ and field‐derived information covering seventeen intrinsic morphological and life‐history traits. Subdivision of some of these traits produced a total of 58 attributes (i.e. modalities). The robustness of this classification was confirmed by a high rate of reclassification (92%) under multiple discriminant analysis (MDA). The phylogenetic contribution was explored using ordination methods with taxonomy at family level acting as a covariable. 3.. Our approach differed from earlier classifications based on growth or life form because we regarded growth form plasticity as a property of the species and its range of growing conditions, rather than of each individual population, and we considered additional (e.g. regenerative) traits. However, some conventional life form groups were preserved (i.e. utricularids, isoetids, hydrocharids and lemnids). 4.. Some parallels existed with established theory on terrestrial plant growth strategies, but we used strictly intrinsic attributes relevant specifically to hydrophytes and our groups could not be decomposed into three or four primary strategies. Only finer levels of partitioning appear to be of fundamental and applied ecological relevance in hydrophytes. 5.. A principal components analysis ordination based on 26 attributes related to physical habitat utilization separated species and their attribute groups along axes relating to: (a) flow, substratum grade and organic matter content, scour frequency, and sedimentation; and (b) depth, water level stability and biotic disturbance. A MDA applied to species ordination scores indicated only a modest overall correspondence between attribute groups and habitat use (54% correct reclassification). Poor reclassification was the result of intergroup overlap (indicating alternative sets of attributes for a given habitat) or high intragroup variance in habitat utilization (indicating commonality of attributes between different habitats). These results are interpreted in terms of trade‐offs between resistance and resilience traits, ‘functional plasticity’ in traits, phylogenetic dependence in some groups and methodological constraints. The predictive potential of hydrophyte groups and their limitations are discussed. 6.. Redundancy analysis revealed a highly significant correlation between traits and habitat use (P < 0.01). Our attribute matrix explained 72% of variation in physical habitat use with eight attributes (i.e. turions, anchored emergent leaves, high or low body flexibility, high root:shoot biomass ratio, free‐floating surface or free‐floating submerged growth form, and annual life history) explaining half of this variation. 7.. Most attributes were mapped in accordance with habitat template predictions, although tests were confounded by the underlying correlation between spatial and temporal heterogeneity. The main features were: (a) a trade‐off between resistance‐type traits (related to stream lining, flexibility and anchorage) in more spatially heterogenous riverine and littoral zone habitats, and resilience type traits (i.e. turions, very small body size and free‐floating growth forms) in spatially simple, rarely disturbed habitats, such as backwaters and canals; and (b) a shift from high investment competitive traits with a low reproductive output in deep stable habitats to classically ruderal and desiccation resistance traits in shallow fluctuating habitats.

Seasonal and geographical distribution of cave‐dwelling bats in Romania: implications for conservation

Caves offer bats refuges for hibernation, breeding and other social events. Their quality is important for species distribution. The role of cave microclimate as well as other environmental factors influencing the distribution of cave‐dwelling species, is poorly known. We tested the significance of cave variables (length, temperature, elevation, occurrence of water) and geographical location for the presence of bats during hibernation and the breeding season in five regions in Romania. To detect species' environmental relationships, we used canonical correspondence analyses for winter bat aggregations and principal components analysis for maternity colonies. We analysed the factors influencing the distribution of bats by using two sets of explanatory variables reflecting cave characteristics and geographical locations. Winter aggregation was divided into three groups: (1) bat species that prefer high temperatures (Rhinolophus euryale, Myotis cappacinii) and hibernate at a low altitude; (2) species ranging from mid‐ to high elevation and low temperature (Myotis myotis/oxygnathus group); (3) species that hibernate in large, cold cave systems with a constant flow of the water (Pipistrellus pipistrellus, Nyctalus noctula, Barbastella barbastellus). Maternity colonies were divided into those that select either high (rhinolophids) or low temperatures (My. myotis/oxygnathus and Miniopterus schreibersii). The most important factors influencing the distribution of bats are the temperature in caves and their geographical location. This information was combined with IUCN's Red List data as well as with the number of individuals occurring in caves with the aim of identifying the key sites for conservation. The majority of these sites, which also constitute the refuges for vulnerable species, are located in west and south‐western Romania. Seven caves provide shelter throughout the year for 122 000 individuals of 14 species.

A Stable Calcium Alumanyl

A seven‐membered N,N′‐heterocyclic potassium alumanyl nucleophile is introduced and utilised in the metathetical synthesis of Mg−Al and Ca−Al bonded derivatives. Both species have been characterised by experimental and theoretical means, allowing a rationalisation of the greater reactivity of the heavier group 2 species implied by an initial assay of their reactivity.

Observations on different methods of aphid trapping

Cylindrical and horizontal sticky traps painted in a range of spectral colours were used to determine the flight and landing behaviour of aphids. Data are also presented on aphid catches in suction traps at two heights and in light traps. Apart from colour sensitivity (yellow versus white) there was apparently a separate response to colour which in some species varied with season. Within yellow‐sensitive species there was also a differential response to colour. With the experimental methods used, it was not possible to define mathematically the active and passive landing components on cylindrical traps. Though the active landing component was large it varied between aphid species. Most species caught on horizontal traps at ground level had been flying above 1 m. In some species the response of males to colour and their landing behaviour differed from that of viviparae and oviparae. Four years data from suction traps suggest that aphid species can be divided into three categories on the basis of the height at which they normally fly. One group, mostly tree‐feeders, always show the greatest density at a high level (12.2 m) throughout the season. The second group always have the highest density at a low level (1 m) whilst a third group of species change at a specific date each autumn from a maximum density at 1–12.2 m. Attraction to light (moth trap) appeared to be linked with the grouping of species by height of flight. The interpretation of catch data is discussed in the light of these observations.

Dormancy of alpine and subalpine perennial forbs

Depth of dormancy of alpine and subalpine perennial forbs in autumn was investigated, which was judged by the number of days required for growth initiation at 24 °C. The depth of dormancy differed depending on Raunkiaer's life‐form and shoot habits. Chamaephytes with perennial shoot‐axes showed shallower dormancy than hemicryptophytes with annual shoot‐axes, and geophytes with annual shoot‐axes showed the deepest dormancy. The results strongly suggest that the dormancy is more endogenously controlled in forbs less hardy to freezing stress. Potential growth ability of alpine herbaceous chamaephytes in autumn is an adaptive advantage, since they utilize the short vegetative period as long as possible. All of the species with annual shoot‐axes had winter buds covered with scales. In plants with perennial shoot‐axes, percentage of winter buds covered with scales increased with increasing depth of dormancy. The results indicate that the shoot apices are well protected by bud scales in forbs with a long endogeneous‐dormant period.

MS‐2 and T4 phage removal in an anaerobic membrane bioreactor (AnMBR): effect of gas sparging rate

BACKGROUND: The enhanced removal of viruses in wastewater treatments plant is important due to concerns about public health. Bacteriophages (or phages) are often used to model the behavior of pathogenic human viruses as they are similar in size, structure and behavior. This study investigated the removal of phages MS‐2 (25 nm) and T4 (200 nm) in an anaerobic membrane bioreactor (AnMBR) with a membrane pore size of 0.4 µm. RESULTS: The membrane reactor without biomass was assessed and its log removal was 0.7 ± 0.4 log for the MS‐2 phage, and 2.3 ± 0.2 log for the T4. When anaerobic biomass was added to the reactor the log removal for both phages increased, and this was thought to be due to a complex relationship with the biofilm on the membrane. CONCLUSIONS: Overall MS‐2 rejections ranged from 1.75 up to 5.5 log, with the highest rejections observed at the highest sparging rates after extensive fouling had occurred. For T4, removal in the AnMBR ranged from 5 log up to complete removal (>log 7). © 2014 Society of Chemical Industry

Pathogenesis of Mouse Hepatitis Virus Infection: The Role of Nasal Epithelial Cells as a Primary Target of Low‐Virulence Virus, MHV‐S

The pathogenesis of mouse hepatitis virus (MHV‐S) infection in suckling and weanling mice was comparatively studied after intranasal inoculation. In sucklings, infectious virus as well as specific antigen was first detected in the nasal mucosa at 12 hr, then in the nerve cells of the olfactory bulbs. At this stage viral particles were demonstrated both in the supporting cells and olfactory cells of the nasal mucosa. In the posterior part of the brain and spinal cord, virus was detected on days 3 to 4 postinoculation when viral growth was clearly demonstrable in the liver, spleen and intestines. In weanlings too, infection was first established in the nasal mucosa, shedding infectious virus in the nasal washing until day 6 postinoculation, and later infection spread to the brain and spinal cord. In weanling mice, however, neither infectious virus nor viral antigen was detected in the liver or other visceral organs, while serum neutralizing antibody became detectable on day 5 postinoculation, increasing in titer thereafter. Histopathologically degenerative and necrotic changes were observed in the nasal mucosa and central nervous system of both age groups of animals coincidentally with the presence of viral specific antigen, while inflammatory response was much less prominent in sucklings. In the liver, spleen and intestines, however, some lesions were observed only in sucklings.

Infectious disease and the conservation of free‐ranging large carnivores

Large carnivores are of vital importance to the stability and integrity of most ecosystems, but recent declines in free‐ranging populations have highlighted the potentially devastating effect of infectious diseases on their conservation. We reviewed the literature on infectious diseases of 34 large (maximum body mass of adults >20 kg) terrestrial carnivore species, 18 of which are considered to be threatened in the wild, and examined reports of antibody prevalence (seroprevalence) and cases of infection, mortality and population decline. Of 52 diseases examined, 44% were viral, 31% bacterial and the remainder were protozoal or fungal. Many infections were endemic in carnivores and/or infected multiple taxonomic families, with the majority probably occurring via inhalation or ingestion. Most disease studies consisted of serological surveys for disease antibodies, and antibody detection tended to be widespread implying that exposure to micro‐organisms was common. Seroprevalence was higher in tropical than temperate areas, and marginally higher for infections known to occur in multiple carnivore groups. Confirmation of active infection via micro‐organism recovery was less common for ursids than other taxonomic groups. Published descriptions of disease‐induced population decline or extinction were rare, and most outbreaks were allegedly the result of direct transmission of rabies or canine distemper virus (CDV) from abundant carnivore species to less‐common large carnivores. We conclude that the threat of disease epidemics in large carnivores may be serious if otherwise lethal infections are endemic in reservoir hosts and transmitted horizontally among taxa. To prevent or mitigate future population declines, research efforts should be aimed at identifying both the diseases of potential importance to large carnivores and the ecological conditions associated with their spread and severity.

ICS 2017 Abstracts

https://www.ics.org/2017/videos

Losing uniqueness – shifts in carabid species composition during dry grassland and heathland succession

Dry sand ecosystems, such as dry grasslands and heathlands, have suffered habitat loss and degradation due to land‐use changes and are today among the most endangered habitats in Central Europe. To evaluate the impact of degradation processes on habitat quality, we investigated how succession from sparse vegetated sand ecosystems to grass‐invaded and tree‐dominated ecosystems and the environmental parameters associated with it influences carabid assemblages. We also determined to what extent typical xerophilic species assemblages still exist. Pitfall trapping at 28 study sites in northwestern Germany yielded 111 carabid species that were grouped using Kendall's W coefficient of concordance. Ordination revealed that the differences between the four species groups resulted from vegetation cover and soil humidity, indicating that carabid distribution clearly reflects degradation processes. Our results suggest that areas in which succession proceeds were unsuitable for assemblages typical of dry grasslands and heathlands. In all, 35 species are lost due to succession from dry grassland and heathland to grass‐invaded and tree‐dominated sites. We discuss implications for habitat management and restoration, since dry sand ecosystems comprise a very high number of specialized and endangered species.

The Use of Narrative Evidence and Explicit Likelihood by Decisionmakers Varying in Numeracy

Decisionmakers are often presented with explicit likelihood assessments (e.g., there is a 10% chance that an attack will occur over the next three months) and supporting narrative evidence in forecasting and risk communication domains. Decisionmakers are thought to rely on both numerical and narrative information to the extent that they perceive the information to be diagnostic, accurate, and trustworthy. In two studies, we explored how lay decisionmakers varying in numeracy evaluated and used likelihood assessments and narrative evidence in forecasts. Overall, the less numerate reported higher risk and likelihood perceptions. In simple probabilistic forecasts without narrative evidence, decisionmakers at all levels of numeracy were able to use the stated likelihood information, although risk perceptions of the less numerate were more affected by likelihood format. When a forecast includes narrative evidence, decisionmakers were better able to use stated likelihood in a percentage as compared to frequency or verbal formats. The more numerate used stated likelihood more in their evaluations whereas the less numerate focused more on the narrative evidence. These results have important implications for risk analysts and forecasters who need to report the results of their analyses to decisionmakers. Decisionmakers varying in numerical ability may evaluate forecasts in different ways depending on the types of information they find easiest to evaluate.

Dormancy and germination characteristics of herbaceous species in the seasonally dry tropics of northern Australia

This study investigated changes in dormancy and germination over 8 months for 23 common species (annual and perennial grasses, legumes and other dicotyledons) from herbaceous communities in northern Australia. Seeds were exposed to three storage treatments: relatively constant laboratory conditions, an oven with fluctuating temperatures similar to those found on the soil surface (25/60°C), or exposed on the soil surface at Townsville. There were wide ranges of initial levels of dormancy (9–100%), rates of change of dormancy and response to the different storage conditions showing that species with several types of dormancy characteristics are able to coexist in these communities. The general trend in dormancy levels was a decline with time with the rate of decline greatest for seeds exposed on the soil surface and least for those stored in the laboratory. The species were divided into groups based on dormancy levels in seeds on the soil surface during the late dry and mid wet seasons. The dormancy characteristics of the groups were related to the ecology of the species in the groups. There was an approximately linear increase in germination rate (i.e. a decrease in the number of days to 50% of final germination) over time for all storage treatments; rates for seeds on the soil surface increased more rapidly than those of seeds in laboratory and oven samples.

Sap flow variation in selected riparian woodland species in the Okavango Delta, Botswana

In the tropical Okavango Delta, transpiration by trees is an important process partly responsible for maintaining the basin as a freshwater environment. Quantification of evapotranspiration from terrestrial landforms of the delta, fringed by riparian woodlands, is one of the main contributors to uncertainty in current hydrological modelling. We investigated sap flow of common trees in the distal, mid‐ and upper delta in July–August 2012, November–December 2012 and February–April 2013 using the compensation heat pulse velocity method. In the distal delta, four Diospyros mespiliformis individuals of different sizes were studied. Four trees of different species were studied in the mid‐ and upper delta. Sap flow density (SFD; flow per unit cross‐sectional area) was used as a common unit to facilitate comparison. Sap flow varied with tree size, species, season and location. It was positively correlated with tree size (r (2) = 0.67). Sap flow variation between seasons and across locations in all the species studied indicated two distinct groups. Group 1 transpired the least during the hottest season, November–December, and Group 2 the most. In Group 1, the highest average SFD was 1.17 l cm(−2 )day(−1) during July–August; in Group 2, it was 1.07 l cm(−2) day(−1) during November–December. Changes in the hydrology of the delta would negatively affect the riparian woodland.

Allozyme variation and genetic relationships among species in the C arex willdenowii complex (Cyperaceae)

A taxonomic study by Naczi, Reznicek, and Ford (American Journal of Botany, 85, 434–447, 1998) has determined that three species (Carex willdenowii, C. basiantha, and C. superata) can be recognized within the C. willdenowii complex. To determine the amount of genetic divergence within and between these species, allozyme analyses were conducted on 14 populations distributed from Pennsylvania to eastern Texas. Seventeen loci were surveyed, 13 of which were polymorphic, with all populations being polymorphic at one or more loci. Interspecific genetic identities ranged from 0.560 (C. willdenowii and C. basiantha) to 0.807 (C. basiantha and C. superata). Alleles for the isozymes Aat‐1, Dia‐1, Idh‐2, Mdh‐2, Per‐1, Pgm‐1, and Pgm‐2 served to distinguish C. willdenowii from C. basiantha and C. superata. Carex basiantha and C. superata were recognized by alleles of Mdh‐2, Pgm‐1, and Tpi‐2. The genetic identities of populations within species were high and exceeded 0.957. A caespitose growth habit and perigynia in close proximity to the staminate flowers suggest adaptations for selfing and therefore low levels of heterozygosity. Paradoxically, the values for expected heterozygosities (H (exp)) were always lower than those obtained by direct count (H (obs)): F values were highly negative, indicating heterozygous excess. Disassortative mating and selection are discussed as possible mechanisms for maintaining heterozygous excess within populations.

Phylogenetic relationships in the Gesnerioideae (Gesneriaceae) based on nrDNA ITS and cpDNA trnL‐F and trnE‐T spacer region sequences

The Gesnerioideae includes most of the New World members of the Gesneriaceae family and is currently considered to include five tribes: Beslerieae, Episcieae, Gesnerieae, Gloxinieae, and Napeantheae. This study presents maximum parsimony and maximum likelihood phylogenetic analyses of nuclear ribosomal DNA internal transcribed spacer regions (ITS), and the chloroplast DNA trnL intron, trnL‐trnF intergenic spacer region, and trnE‐trnT intergenic spacer region sequences. The ITS and cpDNA data sets strongly support the monophyly of a Beslerieae/Napeantheae clade; an Episcieae clade; a Gesnerieae clade; a Gloxinieae clade minus Sinningia, Sinningia relatives, and Gloxinia sarmentiana; and a Sinningia/Paliavana/Vanhouttea clade. This is the first study to provide strong statistical support for these tribes/clades. These analyses suggest that Sinningia and relatives should be considered as a separate tribe. Additionally, generic relationships are explored, including the apparent polyphyly of Gloxinia. Chromosome number changes are minimized on the proposed phylogeny, with the exception of the n = 11 taxa of the Gloxinieae. Scaly rhizomes appear to have been derived once in the Gloxinieae sensu stricto. The number of derivations of the inferior ovary is unclear: either there was one derivation with a reversal to a superior ovary in the Episcieae, or there were multiple independent derivations of the inferior ovary.

Maternal Antibodies Protect Immunoglobulin Deficient Neonatal Mice From Mouse Hepatitis Virus (MHV)‐Associated Wasting Syndrome

PROBLEM: Neonatal mice nursed by dams lacking immunoglobulins (Igs) may often suffer from lethal runting if raised under conventional conditions. The present study was performed in order to clarify a) the cause of the wasting syndrome and b) the protective role of antigen‐specific milk antibodies. METHOD: Ig‐deficient mouse embryos in a conventional environment were embryo‐transferred to specified pathogen free (SPF) dams. Neonatal growth, mortality, and health status of mice from both environments was recorded. Suspected presence of mouse hepatitis virus (MHV) was tested by RT‐PCR. Protective effects on neonatal mortality of milk containing different titers of anti‐MHV antibodies were investigated in cross‐fostering experiments. RESULTS: The SPF colony of Ig‐deficient mice exhibited no breeding problems, whereas Ig‐deficient neonates in the conventional environment suffered from lethal wasting syndrome. Serological screening of the mice kept in the two environments revealed that mice in the conventional room had high titers of antibodies against mouse hepatitis virus. Presence of MHV in runting neonates was confirmed by pathological examinations and RT‐nested‐PCR using MHV genome specific primers. Milk containing high titers of anti‐MHV antibodies, when provided for 8 days or more, completely prevented Ig‐deficient neonates from developing wasting syndrome in the conventional environment. CONCLUSION: These findings show that the neonatal wasting syndrome is associated with the presence of MHV and that neonates are efficiently protected by MHV‐specific antibodies in the milk.

Genomic investigation of piglet resilience following porcine epidemic diarrhea outbreaks

Porcine epidemic diarrhea virus (PEDV) belongs to the Coronaviridae family and causes malabsorptive watery diarrhea, vomiting, dehydration and imbalanced blood electrolytes in pigs. Since the 1970s, PED outbreaks have become a source of problems in pig producing countries all over the world, causing large economic losses for pig producers. Although the infection in adults is not fatal, in naïve suckling piglets mortality is close to 100%. In this study, we investigated genome‐wide differences between dead and recovered suckling piglets from commercial farms after PED outbreaks. Samples from 262 animals (156 dead and 106 recovered) belonging to several commercial lines were collected from five different farms in three different countries (USA, Canada and Germany) and genotyped with the porcine 80K SNP chip. Mean F (st) value was calculated in 1‐Mb non‐overlapping windows between dead and recovered individuals, and the results were normalized to find differences within the comparison. Seven windows with high divergence between dead and recovered were detected—five on chromosome 2, one on chromosome 4 and one on chromosome 15—in total encompassing 152 genes. Several of these genes are either under‐ or overexpressed in many virus infections, including Coronaviridae (such as SARS‐CoV). A total of 32 genes are included in one or more Gene Ontology terms that can be related to PED development, such as Golgi apparatus, as well as mechanisms generally linked to resilience or diarrhea development (cell proliferation, ion transport, ATPase activity). Taken together this information provides a first genomic picture of PEDV resilience in suckling piglets.

Pathologic characterization of short‐chain acyl‐CoA dehydrogenase deficiency in BALB/cByJ mice

BALB/cByJ mice have a deficiency of short‐chain acyl‐CoA dehydrogenase (SCAD) and are a useful model for studying the inborn errors of fatty acid metabolism which affect humans. Patients with some of these disorders present with hypoglycemia, hyperamonemia, and microvesicular fatty change of hepatocytes. In the present study we examined pathogen‐free, SCAD deficient BALB/cByJ mice and control BALB/cBy mice for biochemical and tissue changes following fasting or salicylate challenge. We observed mitochondrial swelling and microvesicular fatty changes in hepatocytes in mutant mice, especially severe following a fast. However, fasting did not alter their blood ammonia and there was no apparent clinical disease. Similarly, salicylates did not produce disease in the BALB/cByJ mice. We did detect in mice an alternative pathway for salicylate metabolism, by‐passing glycine conjugation which is the principal metabolic pathway in humans. © 1993 Wiley‐Liss, Inc.

Morphological diversity and evolution of Centrolepidaceae (Poales), a species‐poor clade with diverse body plans and developmental patterns

• Premise of the study: The small primarily Australian commelinid monocot family Centrolepidaceae displays remarkably high structural diversity that has been hitherto relatively poorly explored. Data on Centrolepidaceae are important for comparison with other Poales, including grasses and sedges. • Methods: We examined vegetative and reproductive morphology in a global survey of Centrolepidaceae based on light and scanning electron microscopy of 18 species, representing all three genera. We used these data to perform a cladistic analysis to assess character evolution. • Key results: Each of the three genera is monophyletic; Centrolepis is sister to Aphelia. Some Centrolepidaceae show a change from spiral to distichous phyllotaxy on inflorescence transition. In Aphelia and most species of Centrolepis, several morphologically distinct leaf types develop along the primary shoot axis and flowers are confined to dorsiventral lateral spikelets. Centrolepis racemosa displays secondary unification of programs of leaf development, absence of the leaf hyperphyll and loss of shoot dimorphism. Presence or absence of a leaf ligule and features of inflorescence and flower morphology are useful as phylogenetic characters in Centrolepidaceae. • Conclusions: Ontogenetic changes in phyllotaxy differ fundamentally between some Centrolepidaceae and many grasses. Inferred evolutionary transformations of phyllotaxy in Centrolepidaceae inflorescences also differ from those in grasses. In contrast with grasses, some Centrolepidaceae possess ligulate leaves where the ligule represents the boundary between the bifacial hypophyll and unifacial hyperphyll. All the highly unusual features of the morphological‐misfit species Centrolepis racemosa could result from the same saltational event. Centrolepidaceae offer good perspectives for studies of evolutionary developmental biology.

Oral transmission of transmissible gastroenteritis virus by muscle and lymph node from slaughtered pigs

SUMMARY A study was conducted in the USA to determine whether transmissible gastroenteritis (TGE) virus could be transmitted from carcases of slaughtered pigs. Transmissible gastroenteritis virus was transmitted to 6‐day‐old piglets by dosing with homogenates of muscle and lymph node collected from 500 clinically normal pigs at the time of slaughter. All piglets in 2 separately housed litters showed clinical signs of TGE with 5 piglets dying within 10 d of oral dosing with homogenates. Transmissible gastroenteritis virus was isolated from 2 of these piglets and all piglets developed TGE antibody. Transmissible gastroenteritis virus was not isolated in tissue culture from muscle and lymph node homogenates, but was isolated from 4 (0.8%) of 500 tonsil samples collected from the same pigs. A survey of 250 serum samples provided an estimate of the prevalence of slaughtered pigs with TGE antibody of 34.8% in the sample population. The results indicate that carcases of some pigs from TGE endemic areas contain viable TGE virus, and that there would be a substantial risk of introducing TGE virus into Australia by the importation of uncooked pig meat from these areas.

Monitoring of dairy herds for Brucella abortus infection when prevalence is low

A total of 2,698 dairy herds were surveyed in 1981–1982 in New South Wales and north eastern Victoria in a review of the methods used to monitor them for the presence of Brucella abortus., The methods used to monitor dairy herds were testing of all breeding cows over 1 year of age using the rose bengal test (RBT) and complement fixation test (CFT), the bulk milk ring test (BMRT), and testing of blood samples collected at abattoirs using the RBT and CFT. The surveyed herds had at least one whole herd test, and BMRT was done at regular intervals in the period of the survey. Of the 99 (3.7%) herds that reacted to the BMRT, 91 (3.4%) herds had false positive reactions and 8 (0.3%) herds were declared infected on follow‐up herd testing. False‐positive reactions were obtained in 22 herds on more than one occasion. Common causes of false positive reactions to the BMRT were thought to be previous vaccination with Strain 19 and sampling in very early or late lactation. Of the 98 (3.63%) herds that reacted to the whole herd serological tests, 80 (2.96%) herds had false‐positive reactions and 18 (0.67%) herds were declared infected. Strain 19 vaccination was thought to be an important cause of false‐positive reactions. Fifty‐three (2.0%) herds showed suspicious reactions on abattoir monitoring but none was declared infected on follow‐up testing. Of the 18 herds with infected or equivocal status, the BMRT identified 8. In a further 6 herds, the infected cattle were not in the milking herd. Four other herds had milkers with high CFT titres which could not be confirmed as infected on culture. In no herds were culture positive RBT or CFT reactors from the milking herd detected without the BMRT being positive. The proportion of false‐positive reactions to the BMRT was high but the BMRT proved very useful in identifying dairy herds infected with B. abortus, when the prevalence of brucellosis was very low. Aust Vet J, 64: 97–100

The TO strains of Theiler's viruses cause “slow virus—like” infections in mice

Intracerebral inoculation of mice with tissue culture–adapted TO strains of Theiler's mouse encephalomyelitis viruses results in a clinical disease consisting of spastic paralysis due to demyelination after a lengthy incubation period. Thus, in effect, these ordinary picornaviruses are capable of causing a slow infection in their natural host, the mouse. In addition, through the use of tissue culture–adapted virus stocks, virus content in mouse tissues now can be accurately quantified by standard plaque assay.

Megaloblastic Anemia and Immune Abnormalities in a Patient with Methionine Synthase Deficiency

ABSTRACT. We report a case of methionine synthase deficiency associated with cellular immune deficiency discovered in a 14‐year‐old boy. Principal findings were: developmental delay, recurrent upper and lower respiratory tract infections, megaloblastic anemia, discovered at 3 months of age, unresponsive to cyanocobalamin and poorly responsive to folinic acid. Biochemical studies showed: an abnormal deoxyuridine suppression test despite normal serum folate, cobalamin and transcobalamin levels; a normal intracellular uptake of these two coenzymes; and an absolute requirement of methionine for fibroblast growth, suggestive of defective methionine synthesis. An absence of methionine synthase activity in the patient's bone marrow and a profound depression of this activity in lymphocytes and liver were found. Hypergammaglobulinemia with variable lymphopenia, depressed lymphocyte transformation after lectin or recall‐antigen stimulation, defective delayed‐type hypersensitivity and decreased natural killer activity were noted as well. The patient died at the age of 14.

Clinical response decision tree for the mountain gorilla (Gorilla beringeii) as a model for great apes

Disease is one of the main threats to the remaining great ape populations of the world. The decision to intervene in the health of the great apes for population sustainability is controversial. Humans' increasing negative influence on great ape health has mandated the reevaluation of current management policies. The Mountain Gorilla Veterinary Project (MGVP) has been making health intervention decisions since 1986. The decision to intervene has often been made subjectively due to poorly defined criteria that are often influenced by emotion. This paper provides a consistent framework for evidence‐based health intervention decision‐making. The decision tree is a five‐tier process consisting of routine sentinel health observation, intensive follow‐up veterinary health observation, outbreak assessment, risk assessment, and risk management. Although this paper focuses on the mountain gorillas, it serves as a basis for evidence‐based decision‐making in other species. Am. J. Primatol. 68:909–927, 2006.© 2006 Wiley‐Liss, Inc.

The Tn Antigen—Structural Simplicity and Biological Complexity

Glycoproteins in animal cells contain a variety of glycan structures that are added co‐ and/or posttranslationally to proteins. Of over 20 different types of sugar–amino acid linkages known, the two major types are N‐glycans (Asn‐linked) and O‐glycans (Ser/Thr‐linked). An abnormal mucin‐type O‐glycan whose expression is associated with cancer and several human disorders is the Tn antigen. It has a relatively simple structure composed of N‐acetyl‐d‐galactosamine with a glycosidic α linkage to serine/threonine residues in glycoproteins (GalNAcα1‐O‐Ser/Thr), and was one of the first glycoconjugates to be chemically synthesized. The Tn antigen is normally modified by a specific galactosyltransferase (T‐synthase) in the Golgi apparatus of cells. Expression of active T‐synthase is uniquely dependent on the molecular chaperone Cosmc, which is encoded by a gene on the X chromosome. Expression of the Tn antigen can arise as a consequence of mutations in the genes for T‐synthase or Cosmc, or genes affecting other steps of O‐glycosylation pathways. Because of the association of the Tn antigen with disease, there is much interest in the development of Tn‐based vaccines and other therapeutic approaches based on Tn expression.

Enteritis in cattle due to Yersinia pseudotuberculosis infection

SUMMARY: A selective medium was used to isolate Yersinia sp from the intestinal tract of 222 scouring cattle in Gippsland during 1985 and 1986. Intestinal infection with Y. pseudotuberculosis, particularly of serotype III, was found to be especially prevalent in weaned calves, yearlings and young adult cattle. Clinically affected cattle had a profuse liquid diarrhoea and many were systemically ill. Haematological changes suggestive of infection were present in 38 of 49 of these cattle. At least 35 cattle died and characteristic microabscesses were demonstrated in the intestinal mucosa of 20 of 26 examined histologically. Y. pseudotuberculosis was sensitive to tetracyclines in vitro and this drug produced a rapid bacteriological cure. Yersiniosis occurred during the winter, spring and early summer. Challenge of adult cattle with Y. pseudotuberculosis serotype III did not result in intestinal colonisation or clinical disease. Intestinal infection was, however, established in 4 weaned calves and haematological changes and antibody production were demonstrated in them. Intestinal microabscesses were seen in three calves killed on days 8, 14 and 18 after challenge. The fourth calf eliminated infection by day 18 and no lesions were demonstrated when it was killed on day 72. There is a very high prevalence of antibodies reacting with Y. pseudotuberculosis serotype III in adult cattle. It is concluded that cattle are a common host for this bacterium, infection being frequent, with clinical and fatal disease occurring occasionally. The factors leading to clinical disease are unknown.

Aspects of the diagnosis, pathogenesis and epidemiology of canine parvovirus

Between 18 July 1980 and 2 January 1981, 188 samples (145 faeces and 43 intestinal contents) were submitted from dogs with suspected canine parvovirus (CPV) enteritis. CPV was demonstrated in 56 (30%) of these samples; the weekly rate of positive CPV identification was remarkably constant at ∼30% even though clinical and often post‐mortem findings strongly supported a diagnosis of CPV enteritis. The simplest, most sensitive and most rapid method for detection of virus was haemagglutination (HA) which was twice as sensitive as isolation of virus and 8 times as sensitive as electron microscopy (EM). Forty nine of 56 (88%) samples positive for CPV were from dogs < 1 year old and 44 (79%) CPV‐positive samples were from pups < 6 months old; only one sample from a pup < 2 months old (pup was 7 weeks old) was positive. An additional 68 samples (53 faeces and 15 intestinal contents) were submitted from Beagle dogs that were part of a colony of ∼1200 dogs. Epidemiological data pinpoints the entry of CPV into the colony in November 1978 at which time most dogs including pups < 6 months of age developed antibody to CPV without developing clinical disease. From these data an overview of some aspects of the pathogenesis and epidemiology of CPV is constructed.

The Abstracts of the 4th UQ Symposium on Organisational Psychology: Emmanuel College, University of Queensland

These abstracts form the collection of papers that were presented at the 4th UQ Symposium on Organisational Psychology held at Emmanuel College, University of Queensland, Brisbane on Saturday 24th July, 2004. The UQ Symposium on Organisational Psychology is an annual even organised by the Centre for Organisational Psychology at the University of Queensland. The aim of the symposium is for academic psychologists to present their latest research to fellow academics and practitioners. Papers were accepted for either paper presentation or poster presentation following a peer‐review process. The 100 delegates who attended consisted of practitioners and academics. The invited speaker was Dr Sally Carless (Monash University, Melbourne) who presented her latest research on selection procedures. The 5th Symposium will take place on Saturday 4th June, 2005. For more information about the UQ Symposium on Organisational Psychology series please contact Robin Martin (r.martin@psy.uq.edu.au).

Epidemiology of encephalitis in children: A 20‐Year survey

Four hundred five children from the Helsinki area who were 1 month to 16 years old were treated for acute encephalitis at the Children's Hospital, University of Helsinki, from January 1968 through December 1987. Encephalitis occurred most commonly in children 1 to 1.9 years of age, among whom the incidence was 16.7 per 100,000 child‐years. The incidence remained quite high until the age of 10 years, and then gradually declined to 1.0 per 100,000 child‐years at the age of 15 years. Since 1983, when mumps, measles, and rubella vaccination eradicated the encephalitides associated with these microbes, the major associated agents have been varicella‐zoster, Mycoplasma pneumoniae, and respiratory and enteroviruses. In infants younger than 1 year of age, the major agents were enteroviruses, herpes simplex virus, and the group of “others,” whereas in older children, respiratory viruses and Mycoplasma Pneumoniae as well as varicella‐zoster virus, dominated. In children aged 1 to 11 months, the causal agent could not be identified in one‐half of all cases, whereas in children who were at least 10 years old, the etiology remained unknown in only one‐fourth of cases. Male dominance was most evident in the 4‐ to 9‐year age group. The difference in etiology between males and females was significant (p = 0.02); mumps and varicella were more common in boys, and adenovirus and Mycoplasma pneumoniae were more common in girls. The overall male‐to‐female ratio was 1.4:1. Characteristic seasonal variation occurred in encephalitides associated with mumps, measles, and entero‐ and respiratory viruses. In the whole series, some accumulation appeared in February and March. Less than one‐half of this number appeared in July and August.

Synovial pathologic changes in spontaneous canine rheumatoid‐like arthritis

The synovial fluid and membrane were studied in 10 dogs meeting the American Rheumatism Association criteria for classic human rheumatoid arthritis (RA). Light microscopic pathologic features were consistent with those found in the human disease. Neutrophilic infiltration of synovium was somewhat more prominent than in chronic human RA, and activated lymphocytes in fluid or membrane were less frequent. The proliferative and plasma cell reaction seemed identical. Electron microscopy (EM) suggested microvascular injury with findings which included electron dense deposits in the vessel walls of 2 dogs. Seven dogs had meshworks of 20‐25 mm tubules in tubuloreticular structures (TRS) similar to those seen in human systemic lupus erythematosus and only occasionally in human RA. There were also crystalline arrays of tubules, a configuration previously reported in tumors and virus infections and possibly suggestive of a cellular reaction to virus infection. To date no initiating agent has been identified, but this spontaneous canine disease which is very similar to human RA can provide a valuable model in which to examine pathogenesis of chronic arthritis.

The virology of demyelinating diseases

Infectious agents have been postulated as causes of multiple sclerosis for over a century. The possible role of a virus or viruses is supported by data that (1) a childhood exposure is involved and “viral” infections may precipitate exacerbations of disease, (2) experimental infections in animals and natural infections in humans can cause diseases with long incubation periods, remitting and relapsing courses, and demyelination, and (3) patients with multiple sclerosis have abnormal immune responses to viruses. The pathogenesis of three human demyelinating diseases of known viral etiology is discussed. In progressive multifocal leukoencephalopathy, a papovavirus selectively infects oligodendrocytes and causes focal areas of demyelination. In postmeasles encephalomyelitis, the virus is lymphotrophic and disrupts immune regulation that can result in an autoimmune perivenular demyelinating illness without evidence of infection of the central nervous system. In human immunodeficiency virus‐encephalopathy and myelopathy virus is present in macrophages and microglia and the myelin abnormalities apparently are caused by soluble factors such as viral proteins, cytokines, or neurotoxins. These findings may have implications on how, when, and where to seek viruses in multiple sclerosis.

Dendritic DNA Building Blocks for Amplified Detection Assays and Biomaterials

DNA branches out: Recent advances in the assembly of dendritic DNA structures enable applications in biosensing of pathogens and the generation of novel pads of DNA hydrogel biomaterials (see scheme, left). These pads are immersed in a cell extract containing RNA polymerase (red), ribosomes (yellow), and other components for in vitro protein biosynthesis, where they can be used as templates for cell‐free protein production.[Image: see text]

Immunohistochemical localization of components of the immune barrier in the olfactory mucosae of salamanders and rats

Immunohistochemical techniques were used to investigate the cellular distribution of components of the secretory immune system, including secretory immunoglobulin, secretory piece, and J chain, as well as other immunoglobulins and nonspecific defense factors in the olfactory mucosae of salamanders and rats. In the salamander, secretory immunoglobulin M, and J chain were localized in duct and acinar cells of Bowman's glands, in B lymphocytes, and in sustentacular cells in immature regions of the olfactory mucosa. Lactoferrin and lysozyme were also present in Bowman's glands, in sustentacular cells in immature regions of the olfactory mucosa, and in blood cells in the lamina propria. Olfactory nerve section resulted in the presence of increased numbers of secretory immunoglobulin‐immunoreactive B lymphocytes and in an altered distribution of IgM, secretory piece, and lactoferrin. In the rat, secretory immunoglobulin A and J chain were localized in duct and acinar cells of Bowman's glands and in B lymphocytes in the lamina propria. Secretory piece could be demonstrated in Bowman's glands only in rats that had a prior viral infection. Other defense factors, localized in the lamina propria, included IgG in the connective tissue stroma and in B lymphocytes, IgD‐immunore‐active B lymphocytes, and IgE‐immunoreactive cells that were identified as mucosal mast cells. Lactoferrin and lysozyme were present in serous acinar cells of Bowman's glands and in blood cells. These results demonstrate that the olfactory mucosa is protected from pathogenic invasion by the secretory immune system as well as other immunoglobulins, lactoferrin, and lysozyme.

An estimate of the economic effects of cattle tick (Boophilus microplus) infestation on Queensland dairy farms

Objective To establish the cost to the Queensland dairy industry of cattle tick infestation and its control, excluding the costs incurred from control measures directed specifically at tick fever and morbidity and mortality arising from tick fever. Study design Economic models are described that have been based on empirical data relating to liveweight and milk yield loss, and on a survey of control practices and tick infestation. The first two models were designed to estimate costs of control and losses resulting from tick infestation on a single dairy farm. The third model developed estimates of the cost of tick infestation for each of four regions within the tick‐infested area of Queensland. Results The overall cost to the Queensland dairy industry of the cattle tick (excluding the costs associated specifically with tick fever) and based on 1998 management practices, was $4,096,000 per annum. About 49% of this cost was related to the costs of control and 51% to losses in production. Conclusion Cattle tick infestation represents a significant impost on dairy producers in Queensland, and although the actual cost will change as deregulation results in economic changes in the industry, infestations of ticks will continue to be expensive to control.

Evidence for multiple sclerosis as an infectious disease

Evidence for a viral cause of multiple sclerosis (MS) is indirect since no infectious agent has been reproducibly isolated from MS tissues nor has viral genome or antigen been consistently identified. The occurrence of spontaneous human and animal models of demyelination, serologic studies, and epidemiologic data provide pursuasive circumstantial evidence for an infectious trigger in this disease. Potential mechanisms for viral induced demyelination include persistent infection of host tissues or immune mediated organ damage either in the presence or absence of the infectious agent. Any proposed viral candidate should cause demyelination in animals or man and the pattern of infection should be consistent with the unique geographic features of MS epidemiology. In addition, serologic studies should support an infection by the agent and/or viral genome should be detected in MS tissues. At this time no virus can be unequivocally linked to MS but cumulative evidence is more supportive of canine distemper virus than other viruses.

Experimental colibacillosis in gnotobiotic piglets exposed to 3 enterotoxigenic serotypes

SUMMARY Three strains of enterotoxigenic Escherichia coli (ETEC) (064:KSNT, K88ac; 020:KSNT, K88ac and 08:K85ab, K99) originally cultured from outbreaks of diarrhoea in piglets a few hours old, were administered orally to gnotobiotic piglets. There was a marked age‐related difference in the clinical response to infection between the 3 strains although they all produced heat‐stable toxin. All 3 strains produced severe clinical signs of depression, anorexia, vomiting, diarrhoea, followed by dehydration and death in one‐day‐old piglets. In piglets infected at 3 days of age the two K88+ ETEC caused diarrhoea and death but the K99+ ETEC induced moderate diarrhoea only. In piglets infected at 7 days of age, the 064 strain produced severe diarrhoea and death, the 020 strain caused mild diarrhoea in 3 of 6 piglets with one death while the 08 strain caused no illness. Pathological changes in the intestinal tract associated with these infections were minimal, or absent. Immunofluorescent staining with homologous hyperimmune sera demonstrated adherence of the 3 ETEC strains to the brush border of small intestinal epithelial cells. Fluorescing organisms were observed in all infected piglets irrespective of the severity of clinical signs but the degree and extent of colonisation varied with the age of the piglets and the infecting strain. This may explain the difference in clinical response between the 3 strains.