Risk factor analysis and spatiotemporal CART model of cryptosporidiosis in Queensland, Australia

Background: It remains unclear whether it is possible to develop a spatiotemporal epidemic prediction model for cryptosporidiosis disease. This paper examined the impact of social economic and weather factors on cryptosporidiosis and explored the possibility of developing such a model using social economic and weather data in Queensland, Australia. ----- ----- Methods: Data on weather variables, notified cryptosporidiosis cases and social economic factors in Queensland were supplied by the Australian Bureau of Meteorology, Queensland Department of Health, and Australian Bureau of Statistics, respectively. Three-stage spatiotemporal classification and regression tree (CART) models were developed to examine the association between social economic and weather factors and monthly incidence of cryptosporidiosis in Queensland, Australia. The spatiotemporal CART model was used for predicting the outbreak of cryptosporidiosis in Queensland, Australia. ----- ----- Results: The results of the classification tree model (with incidence rates defined as binary presence/absence) showed that there was an 87% chance of an occurrence of cryptosporidiosis in a local government area (LGA) if the socio-economic index for the area (SEIFA) exceeded 1021, while the results of regression tree model (based on non-zero incidence rates) show when SEIFA was between 892 and 945, and temperature exceeded 32°C, the relative risk (RR) of cryptosporidiosis was 3.9 (mean morbidity: 390.6/100,000, standard deviation (SD): 310.5), compared to monthly average incidence of cryptosporidiosis. When SEIFA was less than 892 the RR of cryptosporidiosis was 4.3 (mean morbidity: 426.8/100,000, SD: 319.2). A prediction map for the cryptosporidiosis outbreak was made according to the outputs of spatiotemporal CART models. ----- ----- Conclusions: The results of this study suggest that spatiotemporal CART models based on social economic and weather variables can be used for predicting the outbreak of cryptosporidiosis in Queensland, Australia.

Polybrominated diphenyl ether (PBDE) concentrations decrease with age : analysis of pooled human blood serum in the Australian population

Introduction Polybrominated diphenyl ethers (PBDEs) are considered to be a cost effective and efficient way to reduce the possibility of product ignition and inhibit the spread of fire, thereby limiting harm caused by fires. PBDEs are incorporated into a wide variety of manufactured products and are now considered an ubiquitous contaminant found worldwide in biological and environmental samples1 . In comparison to “traditional” persistent organic pollutants (POPs), the exposure modes of PBDEs in humans are less well defined, although dietary sources, inhalation (air/particulate matter) and dust ingestion have been reported 2-4. Limited investigations of population specific factors such as age or gender and PBDE concentrations report: no conclusive correlation by age in adults; higher concentrations in children ; similar concentrations in maternal and cord blood; and no gender differences. After preliminary findings of higher PBDE concentrations in children than in adults in Australia11 we sought to investigate at what age the PBDE concentrations peaked in an effort to focus exposure studies. This investigation involved the collection of blood samples from young age groups and the development of a simple model to predict PBDE concentrations by age in Australia.

Bayesian network for risk of diarrhea associated with the use of recycled water

Estimating potential health risks associated with recycled (reused) water is highly complex given the multiple factors affecting water quality. We take a conceptual model, which represents the factors and pathways by which recycled water may pose a risk of contracting gastroenteritis, convert the conceptual model to a Bayesian net, and quantify the model using one expert’s opinion. This allows us to make various predictions as to the risks posed under various scenarios. Bayesian nets provide an additional way of modeling the determinants of recycled water quality and elucidating their relative influence on a given disease outcome. The important contribution to Bayesian net methodology is that all model predictions, whether risk or relative risk estimates, are expressed as credible intervals.

Incorporating parameter uncertainty into quantitative microbial risk assessment (QMRA)

Modern statistical models and computational methods can now incorporate uncertainty of the parameters used in Quantitative Microbial Risk Assessments (QMRA). Many QMRAs use Monte Carlo methods, but work from fixed estimates for means, variances and other parameters. We illustrate the ease of estimating all parameters contemporaneously with the risk assessment, incorporating all the parameter uncertainty arising from the experiments from which these parameters are estimated. A Bayesian approach is adopted, using Markov Chain Monte Carlo Gibbs sampling (MCMC) via the freely available software, WinBUGS. The method and its ease of implementation are illustrated by a case study that involves incorporating three disparate datasets into an MCMC framework. The probabilities of infection when the uncertainty associated with parameter estimation is incorporated into a QMRA are shown to be considerably more variable over various dose ranges than the analogous probabilities obtained when constants from the literature are simply ‘plugged’ in as is done in most QMRAs. Neglecting these sources of uncertainty may lead to erroneous decisions for public health and risk management.

Physical and functional interaction of the archaeal single-stranded DNA-binding protein SSB with RNA polymerase

Archaeal transcription utilizes a complex multisubunit RNA polymerase and the basal transcription factors TBP and TF(II)B, closely resembling its eukaryal counterpart. We have uncovered a tight physical and functional interaction between RNA polymerase and the single-stranded DNA-binding protein SSB in Sulfolobus solfataricus. SSB stimulates transcription from promoters in vitro under TBP-limiting conditions and supports transcription in the absence of TBP. SSB also rescues transcription from repression by reconstituted chromatin. We demonstrate the potential for promoter melting by SSB, suggesting a plausible basis for the stimulation of transcription. This stimulation requires both the single-stranded DNA-binding domain and the acidic C-terminal tail of the SSB. The tail forms a stable interaction with RNA polymerase. These data reveal an unexpected role for single-stranded DNA-binding proteins in transcription in archaea.

Substrate recognition and catalysis by the Holliday junction resolving enzyme Hje

Two archaeal Holliday junction resolving enzymes, Holliday junction cleavage (Hjc) and Holliday junction endonuclease (Hje), have been characterized. Both are members of a nuclease superfamily that includes the type II restriction enzymes, although their DNA cleaving activity is highly specific for four-way junction structure and not nucleic acid sequence. Despite 28% sequence identity, Hje and Hjc cleave junctions with distinct cutting patterns—they cut different strands of a four-way junction, at different distances from the junction centre. We report the high-resolution crystal structure of Hje from Sulfolobus solfataricus. The structure provides a basis to explain the differences in substrate specificity of Hje and Hjc, which result from changes in dimer organization, and suggests a viral origin for the Hje gene. Structural and biochemical data support the modelling of an Hje:DNA junction complex, highlighting a flexible loop that interacts intimately with the junction centre. A highly conserved serine residue on this loop is shown to be essential for the enzyme's activity, suggesting a novel variation of the nuclease active site. The loop may act as a conformational switch, ensuring that the active site is completed only on binding a four-way junction, thus explaining the exquisite specificity of these enzymes.

hSSB1 interacts directly with the MRN complex stimulating its recruitment to DNA double-strand breaks and its endo-nuclease activity

hSSB1 is a recently discovered single-stranded DNA binding protein that is essential for efficient repair of DNA double-strand breaks (DSBs) by the homologous recombination pathway. hSSB1 is required for the efficient recruitment of the MRN complex to sites of DSBs and for the efficient initiation of ATM dependent signalling. Here we explore the interplay between hSSB1 and MRN. We demonstrate that hSSB1 binds directly to NBS1, a component of the MRN complex, in a DNA damage independent manner. Consistent with the direct interaction, we observe that hSSB1 greatly stimulates the endo-nuclease activity of the MRN complex, a process that requires the C-terminal tail of hSSB1. Interestingly, analysis of two point mutations in NBS1, associated with Nijmegen breakage syndrome, revealed weaker binding to hSSB1, suggesting a possible disease mechanism.

hSSB1 rapidly binds at the sites of DNA double-strand breaks and is required for the efficient recruitment of the MRN complex

hSSB1 is a newly discovered single-stranded DNA (ssDNA)-binding protein that is essential for efficient DNA double-strand break signalling through ATM. However, the mechanism by which hSSB1 functions to allow efficient signalling is unknown. Here, we show that hSSB1 is recruited rapidly to sites of double-strand DNA breaks (DSBs) in all interphase cells (G1, S and G2) independently of, CtIP, MDC1 and the MRN complex (Rad50, Mre11, NBS1). However expansion of hSSB1 from the DSB site requires the function of MRN. Strikingly, silencing of hSSB1 prevents foci formation as well as recruitment of MRN to sites of DSBs and leads to a subsequent defect in resection of DSBs as evident by defective RPA and ssDNA generation. Our data suggests that hSSB1 functions upstream of MRN to promote its recruitment at DSBs and is required for efficient resection of DSBs. These findings, together with previous work establish essential roles of hSSB1 in controlling ATM activation and activity, and subsequent DSB resection and homologous recombination (HR).

In situ preparation and protein delivery of silicate/alginate composite microspheres with core-shell structure

It is predicted that with increased life expectancy in the developed world, there will be a greater demand for synthetic materials to repair or regenerate lost, injured or diseased bone (Hench & Thompson 2010). There are still few synthetic materials having true bone inductivity, which limits their application for bone regeneration, especially in large-size bone defects. To solve this problem, growth factors, such as bone morphogenetic proteins (BMPs), have been incorporated into synthetic materials in order to stimulate de novo bone formation in the center of large-size bone defects. The greatest obstacle with this approach is that the rapid diffusion of the protein from the carrier material, leading to a precipitous loss of bioactivity; the result is often insufficient local induction or failure of bone regeneration (Wei et al. 2007). It is critical that the protein is loaded in the carrier material in conditions which maintains its bioactivity (van de Manakker et al. 2009). For this reason, the efficient loading and controlled release of a protein from a synthetic material has remained a significant challenge. The use of microspheres as protein/drug carriers has received considerable attention in recent years (Lee et al. 2010; Pareta & Edirisinghe 2006; Wu & Zreiqat 2010). Compared to macroporous block scaffolds, the chief advantage of microspheres is their superior protein-delivery properties and ability to fill bone defects with irregular and complex shapes and sizes. Upon implantation, the microspheres are easily conformed to the irregular implant site, and the interstices between the particles provide space for both tissue and vascular ingrowth, which are important for effective and functional bone regeneration (Hsu et al. 1999). Alginates are natural polysaccharides and their production does not have the implicit risk of contamination with allo or xeno-proteins or viruses (Xie et al. 2010). Because alginate is generally cytocompatible, it has been used extensively in medicine, including cell therapy and tissue engineering applications (Tampieri et al. 2005; Xie et al. 2010; Xu et al. 2007). Calcium cross-linked alginate hydrogel is considered a promising material as a delivery matrix for drugs and proteins, since its gel microspheres form readily in aqueous solutions at room temperature, eliminating the need for harsh organic solvents, thereby maintaining the bioactivity of proteins in the process of loading into the microspheres (Jay & Saltzman 2009; Kikuchi et al. 1999). In addition, calcium cross-linked alginate hydrogel is degradable under physiological conditions (Kibat PG et al. 1990; Park K et al. 1993), which makes alginate stand out as an attractive candidate material for the protein carrier and bone regeneration (Hosoya et al. 2004; Matsuno et al. 2008; Turco et al. 2009). However, the major disadvantages of alginate microspheres is their low loading efficiency and also rapid release of proteins due to the mesh-like networks of the gel (Halder et al. 2005). Previous studies have shown that a core-shell structure in drug/protein carriers can overcome the issues of limited loading efficiencies and rapid release of drug or protein (Chang et al. 2010; Molvinger et al. 2004; Soppimath et al. 2007). We therefore hypothesized that introducing a core-shell structure into the alginate microspheres could solve the shortcomings of the pure alginate. Calcium silicate (CS) has been tested as a biodegradable biomaterial for bone tissue regeneration. CS is capable of inducing bone-like apatite formation in simulated body fluid (SBF) and its apatite-formation rate in SBF is faster than that of Bioglass® and A-W glass-ceramics (De Aza et al. 2000; Siriphannon et al. 2002). Titanium alloys plasma-spray coated with CS have excellent in vivo bioactivity (Xue et al. 2005) and porous CS scaffolds have enhanced in vivo bone formation ability compared to porous β-tricalcium phosphate ceramics (Xu et al. 2008). In light of the many advantages of this material, we decided to prepare CS/alginate composite microspheres by combining a CS shell with an alginate core to improve their protein delivery and mineralization for potential protein delivery and bone repair applications

Early osteogenic response of osteoprogenitor cells on modified titanium implant surfaces leads to improved osteogenesis

Background: Implant surface micro-roughness and hydrophilicity are known to improve the osteogenic differentiation potential of osteoprogenitor cells. This study was aimed to determine whether topographically and chemically modified titanium implant surfaces stimulate an initial osteogenic response in osteoprogenitor cells, which leads to their improved osteogenesis. ----- ----- Methods: Statistical analysis of microarray gene expression profiling data available from studies (at 72 hours) on sand-blasted, large grit acid etched (SLA) titanium surfaces was performed. Subsequently, human osteoprogenitor cells were cultured on SLActive (hydrophilic SLA), SLA and polished titanium surfaces for 24 hours, 3 days and 7 days. The expression of BMP2, BMP6, BMP2K, SP1, ACVR1, FZD6, WNT5A, PDLIM7, ITGB1, ITGA2, OCN, OPN, ALP and RUNX2 were studied using qPCR. ----- ----- Results: Several functional clusters related to osteogenesis were highlighted when genes showing statistically significant differences (from microarray data at 72 hours) in expression on SLA surface (compared with control surface) were analysed using DAVID (online tool). This indicates that differentiation begins very early in response to modified titanium surfaces. At 24 hours, ACVR1 (BMP pathway), FZD6 (Wnt pathway) and SP1 (TGF-β pathway) were significantly up-regulated in cultures on the SLActive surface compared to the other surfaces. WNT5A and ITGB1 also showed higher expression on the modified surfaces. Gene expression patterns on Day 3 and Day 7 did not reveal any significant differences.----- ----- Conclusion: These results suggest that the initial molecular response of osteoprogenitor cells to modified titanium surfaces may be responsible for an improved osteogenic response via the BMP and Wnt signalling pathways.