Innate transcriptional networks activated in bladder in response to uropathogenic Escherichia coli drive diverse biological pathways and rapid synthesis of IL-10 for defense against bacterial urinary tract infection

Early transcriptional activation events that occur in bladder immediately following bacterial urinary tract infection (UTI) are not well defined. In this study, we describe the whole bladder transcriptome of uropathogenic Escherichia coli (UPEC) cystitis in mice using genome-wide expression profiling to define the transcriptome of innate immune activation stemming from UPEC colonization of the bladder. Bladder RNA from female C57BL/6 mice, analyzed using 1.0 ST-Affymetrix microarrays, revealed extensive activation of diverse sets of innate immune response genes, including those that encode multiple IL-family members, receptors, metabolic regulators, MAPK activators, and lymphocyte signaling molecules. These were among 1564 genes differentially regulated at 2 h postinfection, highlighting a rapid and broad innate immune response to bladder colonization. Integrative systems-level analyses using InnateDB (http://www.innatedb.com) bioinformatics and ingenuity pathway analysis identified multiple distinct biological pathways in the bladder transcriptome with extensive involvement of lymphocyte signaling, cell cycle alterations, cytoskeletal, and metabolic changes. A key regulator of IL activity identified in the transcriptome was IL-10, which was analyzed functionally to reveal marked exacerbation of cystitis in IL-10–deficient mice. Studies of clinical UTI revealed significantly elevated urinary IL-10 in patients with UPEC cystitis, indicating a role for IL-10 in the innate response to human UTI. The whole bladder transcriptome presented in this work provides new insight into the diversity of innate factors that determine UTI on a genome-wide scale and will be valuable for further data mining. Identification of protective roles for other elements in the transcriptome will provide critical new insight into the complex cascade of events that underpin UTI.

Phage library screening for the rapid identification and in vivo testing of candidate genes for a DNA vaccine against Mycoplasma mycoides subsp. mycoides small colony biotype

A new strategy for rapidly selecting and testing genetic vaccines has been developed, in which a whole genome library is cloned into a bacteriophage λ ZAP Express vector which contains both prokaryotic (Plac) and eukaryotic (PCMV) promoters upstream of the insertion site. The phage library is plated on Escherichia coli cells, immunoblotted, and probed with hyperimmune and/or convalescent-phase antiserum to rapidly identify vaccine candidates. These are then plaque purified and grown as liquid lysates, and whole bacteriophage particles are then used directly to immunize the host, following which PCMV-driven expression of the candidate vaccine gene occurs. In the example given here, a semirandom genome library of the bovine pathogen Mycoplasma mycoides subsp. mycoides small colony (SC) biotype was cloned into λ ZAP Express, and two strongly immunodominant clones, λ-A8 and λ-B1, were identified and subsequently tested for vaccine potential against M. mycoides subsp. mycoides SC biotype-induced mycoplasmemia. Sequencing and immunoblotting indicated that clone λ-A8 expressed an isopropyl-β-d-thiogalactopyranoside (IPTG)-inducible M. mycoides subsp. mycoides SC biotype protein with a 28-kDa apparent molecular mass, identified as a previously uncharacterized putative lipoprotein (MSC_0397). Clone λ-B1 contained several full-length genes from the M. mycoides subsp. mycoides SC biotype pyruvate dehydrogenase region, and two IPTG-independent polypeptides, of 29 kDa and 57 kDa, were identified on immunoblots. Following vaccination, significant anti-M. mycoides subsp. mycoides SC biotype responses were observed in mice vaccinated with clones λ-A8 and λ-B1. A significant stimulation index was observed following incubation of splenocytes from mice vaccinated with clone λ-A8 with whole live M. mycoides subsp. mycoides SC biotype cells, indicating cellular proliferation. After challenge, mice vaccinated with clone λ-A8 also exhibited a reduced level of mycoplasmemia compared to controls, suggesting that the MSC_0397 lipoprotein has a protective effect in the mouse model when delivered as a bacteriophage DNA vaccine. Bacteriophage-mediated immunoscreening using an appropriate vector system offers a rapid and simple technique for the identification and immediate testing of putative candidate vaccines from a variety of pathogens.

OHS liability, corporate officers and corporate groups

This article examines the new model for corporate officer liability under section 144 of the Occupational Health and Safety Act 2004 (Vic), and explores the extent to which this might effectively extend responsibility for OHS offences to members of corporate groups, such as holding companies. In doing so, the authors canvass the failure of corporate law to impose such obligations on corporate officers in general, and on holding companies as shadow officers. It is argued that provisions such as section 144 of the Victorian Act should be included in all OHS legislation.

Methylproamine protects against ionizing radiation by preventing DNA double-strand breaks

Purpose The majority of cancer patients will receive radiotherapy (RT), therefore, investigations into advances of this modality are important. Conventional RT dose intensities are limited by adverse responses in normal tissues and a primary goal is to ameliorate adverse normal tissue effects. The aim of these experiments is to further our understanding regarding the mechanism of radioprotection by the DNA minor groove binder, methylproamine, in a cellular context at the DNA level. Materials and methods We used immunocytochemical methods to measure the accumulation of phosphorylated H2AX (γH2AX) foci following ionizing radiation (IR) in patient-derived lymphoblastoid cells exposed to methylproamine. Furthermore, we performed pulsed field gel electrophoresis DNA damage and repair assays to directly interrogate the action of methylproamine on DNA in irradiated cells. Results We found that methylproamine-treated cells had fewer γH2AX foci after IR compared to untreated cells. Also, the presence of methylproamine decreased the amount of lower molecular weight DNA entering the gel as shown by the pulsed field gel electrophoresis assay. Conclusions These results suggest that methylproamine acts by preventing the formation of DNA double-strand breaks (dsbs) and support the hypothesis that radioprotection by methylproamine is mediated, at least in part, by decreasing initial DNA damage.

Moderate physical activity level as a protective factor against metabolic syndrome in middle-aged and older women

Aims and objectives To investigate whether physical activity is a protective factor against metabolic syndrome in middle-aged and older women. Background Socio-demographic and lifestyle behaviour factors contribute to metabolic syndrome. To minimise the risk of metabolic syndrome, several global guidelines recommend increasing physical activity level. However, only limited research has investigated the relationship between physical activity levels and metabolic syndrome in middle-aged and older women after adjusting for socio-demographic and lifestyle behaviour factors. Design Cross-sectional design. Methods A convenience sample of 326 middle-aged and older women was recruited. Metabolic syndrome was confirmed according to the National Cholesterol Education Program, Adult Treatment Panel III guidelines, and physical activity levels were measured by the International Physical Activity Questionnaire. Results The sample had a mean age of 60•9 years, and the prevalence of metabolic syndrome was 43•3%. Postmenopausal women and women with low socioeconomic status (low-education background, without personal income and currently unemployed) had a significantly higher risk of developing metabolic syndrome. After adjusting for significant socio-demographic and lifestyle behaviour factors, the women with moderate or high physical activity levels had a significantly lower (OR = 0•10; OR = 0•11, p < 0•001) risk of metabolic syndrome and a lower risk for each specific component of metabolic syndrome, including elevated fasting plasma glucose (OR = 0•29; OR = 0•26, p = 0•009), elevated blood pressure (OR = 0•18; OR = 0•32, p = 0•029), elevated triglycerides (OR = 0•41; OR = 0•15, p = 0•001), reduced high-density lipoprotein (OR = 0•28; OR = 0•27, p = 0•004) and central obesity (OR = 0•31; OR = 0•22, p = 0•027). Conclusions After adjusting for socio-demographic and lifestyle behaviour factors, physical activity level was a significant protective factor against metabolic syndrome in middle-aged and older women. Higher physical activity levels (moderate or high physical activity level) reduced the risk of metabolic syndrome in middle-aged and older women. Relevance to clinical practice Appropriate strategies should be developed to encourage middle-aged and older women across different socio-demographic backgrounds to engage in moderate or high levels of physical activity to reduce the risk of metabolic syndrome.

Medical negligence, causation and liability for non-disclosure of risk : a post-wallace framework and critique

Through an examination of Wallace v Kam, this article considers and evaluates the law of causation in the specific context of a medical practitioner’s duty to provide information to patients concerning material risks of treatment. To supply a contextual background for the analysis which follows, Part II summarises the basic principles of causation law, while Part III provides an overview of the case and the reasoning adopted in the decisions at first instance and on appeal. With particular emphasis upon the reasoning in the courts of appeal, Part IV then examines the implications of the case in the context of other jurisprudence in this field and, in so doing, provides a framework for a structured consideration of causation issues in future non-disclosure cases under the Australian civil liability legislation. As will become clear, Wallace was fundamentally decided on the basis of policy reasoning centred upon the purpose behind the legal duty violated. Although the plurality in Rogers v Whitaker rejected the utility of expressions such as ‘the patient’s right of self-determination’ in this context, some Australian jurisprudence may be thought to frame the practitioner’s duty to warn in terms of promoting a patient’s autonomy, or right to decide whether to submit to treatment proposed. Accordingly, the impact of Wallace upon the protection of this right, and the interrelation between it and the duty to warn’s purpose, is investigated. The analysis in Part IV also evaluates the courts’ reasoning in Wallace by questioning the extent to which Wallace’s approach to liability and causal connection in non-disclosure of risk cases: depends upon the nature and classification of the risk(s) in question; and can be reconciled with the way in which patients make decisions. Finally, Part V adopts a comparative approach by considering whether the same decision might be reached if Wallace was determined according to English law.

Circulating antibodies against Plasmodium falciparum histidine-rich proteins 2 interfere with antigen detection by rapid diagnostic tests

Background Rapid diagnostic tests (RDTs) for detection of Plasmodium falciparum infection that target P. falciparum histidine-rich protein 2 (PfHRP2), a protein that circulates in the blood of patients infected with this species of malaria, are widely used to guide case management. Understanding determinants of PfHRP2 availability in circulation is therefore essential to understanding the performance of PfHRP2-detecting RDTs. Methods The possibility that pre-formed host anti-PfHRP2 antibodies may block target antigen detection, thereby causing false negative test results was investigated in this study. Results Anti-PfHRP2 antibodies were detected in 19/75 (25%) of plasma samples collected from patients with acute malaria from Cambodia, Nigeria and the Philippines, as well as in 3/28 (10.7%) asymptomatic Solomon Islands residents. Pre-incubation of plasma samples from subjects with high-titre anti-PfHRP2 antibodies with soluble PfHRP2 blocked the detection of the target antigen on two of the three brands of RDTs tested, leading to false negative results. Pre-incubation of the plasma with intact parasitized erythrocytes resulted in a reduction of band intensity at the highest parasite density, and a reduction of lower detection threshold by ten-fold on all three brands of RDTs tested. Conclusions These observations indicate possible reduced sensitivity for diagnosis of P. falciparum malaria using PfHRP2-detecting RDTs among people with high levels of specific antibodies and low density infection, as well as possible interference with tests configured to detect soluble PfHRP2 in saliva or urine samples. Further investigations are required to assess the impact of pre-formed anti-PfHRP2 antibodies on RDT performance in different transmission settings.

Antibodies against human herpesvirus 8 in black South African patients with cancer

Background Infection with human herpesvirus 8 (HHV-8) has been consistently linked to Kaposi's sarcoma, but its mode of transmission, association with other cancers, and interaction with the human immunodeficiency virus type 1 (HIV-1) are largely unknown. Methods Between January 1992 and December 1997, we interviewed 3591 black patients with cancer in Johannesburg and Soweto, South Africa. Blood was tested for antibodies against HIV-1 and HHV-8 in 3344 of the patients. Antibodies against HHV-8 were detected with an indirect immunofluorescence assay. The intensity of the fluorescent signal correlated well with the titers of antibodies (P<0.001). The relations among the presence of anti–HHV-8 antibodies, sociodemographic and behavioral factors, type of cancer, and the presence or absence of coexistent HIV-1 infection were examined with the use of unconditional logistic-regression models. Results Among the 3293 subjects with cancers other than Kaposi's sarcoma, the standardized seroprevalence of antibodies against HHV-8 was 32 percent, which did not differ significantly from the standardized seroprevalence among black blood donors. Among these 3293 patients, the prevalence of antibodies against HHV-8 increased with increasing age (P<0.001) and an increasing number of sexual partners (P=0.05) and decreased with increasing years of education (P=0.007); it was not strongly associated with HIV-1 infection. Anti–HHV-8 antibodies were more frequent among black than white blood donors (P<0.001). Among the 51 patients with Kaposi's sarcoma, the standardized seroprevalence of antibodies against HHV-8 was 83 percent, significantly higher than the prevalence among those without Kaposi's sarcoma (P<0.001). For 16 other specific types of cancer, including multiple myeloma (108 cases) and prostate cancer (202 cases), the variation in the standardized seroprevalence of antibodies against HHV-8 was not remarkable. At a given intensity of fluorescence of anti–HHV-8 antibodies, Kaposi's sarcoma was more frequent among HIV-1–positive patients than among those who were HIV-1–negative (P<0.001). Conclusions Among black patients with cancer in South Africa, the seroprevalence of anti–HHV-8 antibodies is high and is specifically associated with Kaposi's sarcoma, particularly at high titers.

Examination of the properties of IMRT and VMAT beams and evaluation against pre-treatment quality assurance results

This study aimed to provide a detailed evaluation and comparison of a range of modulated beam evaluation metrics, in terms of their correlation with QA testing results and their variation between treatment sites, for a large number of treatments. Ten metrics including the modulation index (MI), fluence map complexity (FMC), modulation complexity score (MCS), mean aperture displacement (MAD) and small aperture score (SAS) were evaluated for 546 beams from 122 IMRT and VMAT treatment plans targeting the anus, rectum, endometrium, brain, head and neck and prostate. The calculated sets of metrics were evaluated in terms of their relationships to each other and their correlation with the results of electronic portal imaging based quality assurance (QA) evaluations of the treatment beams. Evaluation of the MI, MAD and SAS suggested that beams used in treatments of the anus, rectum, head and neck were more complex than the prostate and brain treatment beams. Seven of the ten beam complexity metrics were found to be strongly correlated with the results from QA testing of the IMRT beams (p < 0.00008). For example, Values of SAS (with MLC apertures narrower than 10 mm defined as “small”) less than 0.2 also identified QA passing IMRT beams with 100% specificity. However, few of the metrics are correlated with the results from QA testing of the VMAT beams, whether they were evaluated as whole 360◦ arcs or as 60◦ sub-arcs. Select evaluation of beam complexity metrics (at least MI, MCS and SAS) is therefore recommended, as an intermediate step in the IMRT QA chain. Such evaluation may also be useful as a means of periodically reviewing VMAT planning or optimiser performance.

Literacy, place, and pedagogies of possibility: Working against residualization effects and deficit discourses in poor school communities

Australia has a long history of policy attention to the education of poor and working-class youth (Connell, 1994), yet currently on standardized educational outcomes measures the gaps are widening in ways that relate to social background, including race, location and class. An economic analysis of school choice in Australia reveals that a high proportion of government school students now come from lower Socio-Economic Status (SES) backgrounds (Ryan & Watson, 2004), indicating a trend towards a gradual residualisation of the poor in government schools, with increased private school enrolments as a confirmed national trend. The spatial distribution of poverty and the effects on school populations are not unique to Australia (Lupton, 2003; Lipman, 2011; Ryan, 2010). Raffo and colleagues (2010) recently provided a synthesis of socially critical approaches towards schooling and poverty arguing that what is needed are shifts in the balances of power to reposition those within the educational system as having some say in the ways schooling is organized. ‘Disadvantaged’ primary schools are not a marginal concern for education systems, but now account for a large and growing number of schools that serve an ever increasing population being made redundant, in part-time precarious work, under-employed or unemployed (Thomson 2002; Smyth, Down et al 2010). In Australia, the notion of the ‘disadvantaged’ school now refers to those, mostly public schools, being residualised by a politics of parental choice that drives neoliberalising policy logic (Bonner & Caro 2007; Hattam & Comber, forthcoming 2014; Thomson & Reid, 2003)...

Professional indemnity insurance, performance and pre-emptive negligence: The impact of ‘non-claims’

As part of Australian licensing requirements professional valuers are required to maintain a level of professional indemnity insurance. A core feature of any insurance cover is that the insured has an obligation to notify their insurer of both actual and potential claims. An actual claim clearly will impact upon future policies and premiums paid. Notification of a potential claim, whether or not the notification crystallises into an actual claim, also can have an impact upon the insured’s claims history and premiums. The Global Financial Crisis continues to impact upon business practices and land transactions both directly and indirectly. The Australian valuation profession is not exempt from this impact. One example of this ongoing impact is reflected in a worrying practice engaged in by some financial institutions in respect of their loan portfolios. That is, even though the mortgagor is not in default, some institutions are pre-emptively issuing notices of demand regarding potential losses. Further, in some instances such demands are based only on mass appraisal valuations without specific consideration being given to the individual lot in question. The author examines the impact of this practice for the valuation profession and seeks to provide guidance for the appropriate handling of such demands.

Duty and control in intermediary copyright liability: An Australian perspective

In the internet age, copyright owners are increasingly looking to online intermediaries to take steps to prevent copyright infringement. Sometimes these intermediaries are closely tied to the acts of infringement; sometimes – as in the case of ISPs – they are not. In 2012, the Australian High Court decided the Roadshow Films v iiNet case, in which it held that an Australian ISP was not liable under copyright’s authorization doctrine, which asks whether the intermediary has sanctioned, approved or countenanced the infringement. The Australian Copyright Act 1968 directs a court to consider, in these situations, whether the intermediary had the power to prevent the infringement and whether it took any reasonable steps to prevent or avoid the infringement. It is generally not difficult for a court to find the power to prevent infringement – power to prevent can include an unrefined technical ability to disconnect users from the copyright source, such as an ISP terminating users’ internet accounts. In the iiNet case, the High Court eschewed this broad approach in favor of focusing on a notion of control that was influenced by principles of tort law. In tort, when a plaintiff asserts that a defendant should be liable for failing to act to prevent harm caused to the plaintiff by a third party, there is a heavy burden on the plaintiff to show that the defendant had a duty to act. The duty must be clear and specific, and will often hinge on the degree of control that the defendant was able to exercise over the third party. Control in these circumstances relates directly to control over the third party’s actions in inflicting the harm. Thus, in iiNet’s case, the control would need to be directed to the third party’s infringing use of BitTorrent; control over a person’s ability to access the internet is too imprecise. Further, when considering omissions to act, tort law differentiates between the ability to control and the ability to hinder. The ability to control may establish a duty to act, and the court will then look to small measures taken to prevent the harm to determine whether these satisfy the duty. But the ability to hinder will not suffice to establish liability in the absence of control. This article argues that an inquiry grounded in control as defined in tort law would provide a more principled framework for assessing the liability of passive intermediaries in copyright. In particular, it would set a higher, more stable benchmark for determining the copyright liability of passive intermediaries, based on the degree of actual, direct control that the intermediary can exercise over the infringing actions of its users. This approach would provide greater clarity and consistency than has existed to date in this area of copyright law in Australia.