Synthesis and crystal structure of the MgCl2(CH3COOC2H5)2· (CH3COOC2H5) adduct

The reaction of α-MgCl₂ with boiling ethyl acetate affords MgCI₂(CH₃COOC₂H₅)₂· (CH₃COOC₂H₅), which is obtained as crystals suitable for X-ray analysis only from the mother liquor. M=315.5, orthorhombic, space group P2₁22₁ (No. 18), a=25.077(3), b=8.616(1), c=7.345(1) Å, V=1587.0(3) ų, Z=4, D_x=1.32 g cm⁻³,λ A(Mo Kα)=0.71069 Å, μ=4.17 cm⁻¹, F(000)=664, T=298 K, observed reflections: 1667, R=0.059 and R_w=0.069. The structure is composed of polymeric chains of MgCl₂(CH₃COOC₂H₅₎₂ and the ethyl acetate molecules occupy a mutually trans position.     

Sigma Factor SigB Is Crucial to Mediate Staphylococcus aureus Adaptation during Chronic Infections

Staphylococcus aureus is a major human pathogen that causes a range of infections from acute invasive to chronic and difficult-to-treat. Infection strategies associated with persisting S. aureus infections are bacterial host cell invasion and the bacterial ability to dynamically change phenotypes from the aggressive wild-type to small colony variants (SCVs), which are adapted for intracellular long-term persistence. The underlying mechanisms of the bacterial switching and adaptation mechanisms appear to be very dynamic, but are largely unknown. Here, we analyzed the role and the crosstalk of the global S. aureus regulators agr, sarA and SigB by generating single, double and triple mutants, and testing them with proteome analysis and in different in vitro and in vivo infection models. We were able to demonstrate that SigB is the crucial factor for adaptation in chronic infections. During acute infection, the bacteria require the simultaneous action of the agr and sarA loci to defend against invading immune cells by causing inflammation and cytotoxicity and to escape from phagosomes in their host cells that enable them to settle an infection at high bacterial density. To persist intracellularly the bacteria subsequently need to silence agr and sarA. Indeed agr and sarA deletion mutants expressed a much lower number of virulence factors and could persist at high numbers intracellularly. SigB plays a crucial function to promote bacterial intracellular persistence. In fact, ΔsigB-mutants did not generate SCVs and were completely cleared by the host cells within a few days. In this study we identified SigB as an essential factor that enables the bacteria to switch from the highly aggressive phenotype that settles an acute infection to a silent SCV-phenotype that allows for long-term intracellular persistence. Consequently, the SigB-operon represents a possible target to develop preventive and therapeutic strategies against chronic and therapy-refractory infections.     

Novel anti-ErbB3 monoclonal antibodies show therapeutic efficacy in xenografted and spontaneous mouse tumors

The role of the ErbB3 receptor in signal transduction is to augment the signaling repertoire of active heterodimeric ErbB receptor complexes through activating the PI3K/AKT pathway, which in turn promotes survival and proliferation. ErbB3 has recently been proposed to be involved in acquired resistance to tyrosine kinase inhibitors (TKIs), and is therefore a promising new drug cancer target. Since ErbB3 is a kinase defective receptor, it cannot be targeted by small molecule inhibitors, whereas monoclonal antibodies may offer a viable strategy for pharmacological intervention. In this study, we have utilized DNA electroporation (DNA-EP) to generate a set of novel hybridomas directed against human ErbB3, which have been characterized for their biochemical and functional properties and selected for their ability to negatively regulate the ErbB3-mediated signaling pathway. In vitro, the anti-ErbB3 antibodies modulate the growth rate of cancer cells of different origins. In vivo they show antitumoral properties in a xenograft model of human pancreatic tumor and in the ErbB2-driven carcinogenesis genetically engineered mouse model (GEMM) for mammary tumor, the BALB/neuT. Our data confirm that downregulating the ErbB3-mediated signals with the use of anti-ErbB3 monoclonal antibodies is both feasible and relevant for therapeutic purposes and provides new opportunities for novel anti-ErbB3 combinatory strategies for cancer treatment.     

Comparative Inactivation of Murine Norovirus, Human Adenovirus, and Human JC Polyomavirus by Chlorine in Seawater

Viruses excreted by humans affect the commercial and recreational use of coastal water. Shellfish produced in contaminated waters have been linked to many episodes and outbreaks of viral gastroenteritis, as well as other food-borne diseases worldwide. The risk can be reduced by appropriate treatment following harvesting and by depuration. The kinetics of inactivation of murine norovirus 1 and human adenovirus 2 in natural and artificial seawater by free available chlorine was studied by quantifying genomic copies (GC) using quantitative PCR and infectious viral particles (PFU). Human JC polyomavirus Mad4 kinetics were evaluated by quantitative PCR. DNase or RNase were used to eliminate free genomes and assess potential viral infectivity when molecular detection was performed. At 30 min of assay, human adenovirus 2 showed 2.6- and 2.7-log(10) GC reductions and a 2.3- and 2.4-log(10) PFU reductions in natural and artificial seawater, respectively, and infectious viral particles were still observed at the end of the assay. When DNase was used prior to the nucleic acid extraction the kinetic of inactivation obtained by quantitative PCR was statistically equivalent to the one observed by infectivity assays. For murine norovirus 1, 2.5, and 3.5-log(10) GC reductions were observed in natural and artificial seawater, respectively, while no viruses remained infectious after 30 min of contact with chlorine. Regarding JC polyomavirus Mad4, 1.5- and 1.1-log(10) GC reductions were observed after 30 min of contact time. No infectivity assays were conducted for this virus. The results obtained provide data that might be applicable to seawater used in shellfish depuration.     

Accuracy of Serological Testing for the Diagnosis of Prevalent Neurocysticercosis in Outpatients with Epilepsy,Eastern Cape Province,South Africa

Background

Few studies have estimated prevalence of neurocysticercosis (NCC) among persons with epilepsy in sub-Saharan Africa. While the limitations of serological testing in identification of NCC are well known, the characteristics of persons who are misdiagnosed based on serology have not been explored. The first objective of this pilot study was to estimate the prevalence of NCC in epilepsy outpatients from an area of South Africa endemic for cysticercosis. The second objective was to estimate the accuracy of serological testing in detecting NCC in these outpatients and characterize sources of disagreement between serology and neuroimaging.

Methodology/Principal Findings

All out-patients aged 5 or older attending the epilepsy clinic of St. Elizabeth''s Hospital in Lusikisiki, Eastern Cape Province, between July 2004 and April 2005 were invited to participate. Epidemiological data were collected by local study staff using a standardized questionnaire. Blood samples were tested by ELISA for antibody and antigen for Taenia solium. Four randomly chosen, consenting participants were transported each week to Mthatha for brain CT scan. The proportion of persons with epilepsy attending St. Elizabeth clinic with CT-confirmed NCC was 37% (95% CI: 27%–48%). Using CT as the gold standard, the sensitivity and specificity of antibody testing for identifying NCC were 54.5% (36.4%–71.9%) and 69.2% (52.4%–83.0%), respectively. Sensitivity improved to 78.6% (49.2%–95.3%) for those with active lesions. Sensitivity and specificity of antigen testing were considerably poorer. Compared to false negatives, true positives more often had active lesions. False positives were more likely to keep pigs and to have seizure onset within the past year than were true negatives.

Conclusions/Significance

The prevalence of NCC in South African outpatients with epilepsy is similar to that observed in other countries where cysticercosis is prevalent. Errors in classification of NCC using serology alone may reflect the natural history of NCC.     

Binding studies on aluminum(III)-albumin interaction

The present paper reports a quantitative investigation on the binding of aluminum to human serum albumin. Equilibrium dialysis and a general thermodynamic approach have been used to determine the binding parameters. Two aluminum binding sites have been identified on the albumin molecule, involving sites with single and double occupancy modes. The binding properties of these two distinct sites appear to undergo reciprocal influences, suggesting a possible interaction between the corresponding protein moieties. Both coordination modes proceed from weak interactions, as shown by the binding energies calculated.     

Environmental Factors Influencing Human Viral Pathogens and Their Potential Indicator Organisms in the Blue Mussel, Mytilus edulis: the First Scandinavian Report

This study was carried out in order to investigate human enteric virus contaminants in mussels from three sites on the west coast of Sweden, representing a gradient of anthropogenic influence. Mussels were sampled monthly during the period from February 2000 to July 2001 and analyzed for adeno-, entero-, Norwalk-like, and hepatitis A viruses as well as the potential viral indicator organisms somatic coliphages, F-specific RNA bacteriophages, bacteriophages infecting Bacteroides fragilis, and Escherichia coli. The influence of environmental factors such as water temperature, salinity, and land runoff on the occurrence of these microbes was also included in this study. Enteric viruses were found in 50 to 60% of the mussel samples, and there were no pronounced differences between the samples from the three sites. E. coli counts exceeded the limit for category A for shellfish sanitary safety in 40% of the samples from the sites situated in fjords. However, at the site in the outer archipelago, this limit was exceeded only once, in March 2001, when extremely high levels of atypical indole-negative strains of E. coli were registered at all three sites. The environmental factors influenced the occurrence of viruses and phages differently, and therefore, it was hard to find a coexistence between them. This study shows that, for risk assessment, separate modeling should be done for every specific area, with special emphasis on environmental factors such as temperature and land runoff. The present standard for human fecal contamination, E. coli, seems to be an acceptable indicator of only local sanitary contamination; it is not a reliable indicator of viral contaminants in mussels. To protect consumers and get verification of “clean” mussels, it seems necessary to analyze for viruses as well. The use of a molecular index of the human contamination of Swedish shellfish underscores the need for reference laboratories with high-technology facilities.     

Association between bryophytes and seedlings in Nothofagus sub-Antarctic forests

Bryophytes, which include mosses, liverworts and hornworts, indeed play a significant role in the natural dynamics of native forests. They contribute to fundamental ecological processes such as tree regeneration. In this context, the objective was to analyse the association of bryophytes with Nothofagus seedlings, as well as whether the substrates where bryophytes grew influenced bryophyte-tree seedling association, in sub-Antarctic forests at two contrasting landscapes (coastal and mountain) of Tierra del Fuego (Argentina), to better understand their impacts on the natural regeneration processes. We hypothesized that bryophytes act as a substrate for seed germination and initial regeneration growth in pure and mixed forests of N. pumilio and N. betuloides, resulting in widespread tree seedlings–bryophyte associations in all landscapes. In each forest and landscape location, 60 transects were established (10 m in length) to evaluate bryophyte cover (by point intercept method) and substrate type (bare soil, decaying wood and litter cover) where they were growing. An adaptation of the relative interaction index (RII) based on tree seedling cover associated with bryophyte species (%) and tree seedling cover in the absence of bryophytes (%) was calculated and analysed to assess competition or facilitation between bryophytes and tree seedlings. Nothofagus pumilio seedlings were less abundant in bryophytes compared to other substrates, suggesting an inhibitory effect on the germination and/or survival of N. pumilio. In contrast, the seedlings of N. betuloides, in both pure and mixed forests, exhibited higher abundance on bryophyte substrates, particularly in mosses at mountain landscape (RII = 0.83 ± 0.08 in litter and 0.62 ± 0.11 in decaying wood). These findings suggest that bryophytes play a facilitating role for N. betuloides seedlings during germination and initial phases of growth, mainly in the mountain. Therefore, their conservation promotes N. betuloides forest continuity, both in pure and mixed structures.     

Effects of vasopressin and cyclic AMP on water transport at arachnoid villi of cats.

The effects of vasopressin and cyclic AMP on water transport at arachnoid villi into the superior sagittal sinus were examined using the isolated meninges preparations of cats. The meninges preparation, the superior sagittal sinus of which was opened at the midline of the outer surface, was held between two polyethylene tubes. The tubes were fixed vertically in the way that the opened surface of the sinus was directed downward and arachnoid surface upward. Water transport was determined by measuring the tritiated water dripping through the membrane preparation. Vasopressin from less than 50 to 500 muU/ml accelerated the water transport and this effect was dose-dependent. Cyclic AMP from 0.5 to 10 mM was proved to manifest the same effect as vasopressin. This effect of cyclic AMP appeared rapidly in comparison with that of vasopressin, suggesting that the effect of vasopressin may be manifested through cyclic AMP. From these findings a physiological role of vasopressin in cerebrospinal fluid was discussed regarding the regulation of intracranial pressure.