Identification of novel markers for uncomplicated lower genital tract infections and upper genital tract pathology due to Chlamydia trachomatis.

Background: Untreated Chlamydia trachomatis infections in women can result in disease sequelae such as salpingitis and pelvic inflammatory disease (PID), ultimately culminating in tubal occlusion and infertility. Whilst nucleic acid amplification tests can effectively diagnose uncomplicated lower genital tract (LGT) infections, they are not suitable for diagnosing upper genital tract (UGT) pathological sequelae. As a consequence, this study aimed to identify serological markers that can, with a high degree of sensitivity and specificity, discriminate between LGT infections and UGT pathology. Methods: Plasma was collected from 73 women with a history of LGT infection, UGT pathology due to C. trachomatis or no serological evidence of C. trachomatis infection. Western blotting was used to analyse antibody reactivity against extracted chlamydial proteins. Sensitivity and specificity of differential markers were also calculated. Results: Four antigens (CT157, CT423, CT727 and CT396) were identified and found to be capable of discriminating between the infection and disease sequelae state. Sensitivity and specificity calculations showed that our assay for diagnosing LGT infection had a sensitivity and specificity of 75% and 76% respectively, whilst the assay for identifying UGT pathology demonstrated 80% sensitivity and 86% specificity. Conclusions: The use of these assays could potentially facilitate earlier diagnoses in women suffering UGT pathology due to C. trachomatis.

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.

Rapid displacement of dengue virus type 1 by type 4, Pacific region, 2007-2009

Since 2000-2001, dengue virus type 1 has circulated in the Pacific region. However, in 2007, type 4 reemerged and has almost completely displaced the strains of type 1. If only 1 serotype circulates at any time and is replaced approximately every 5 years, DENV-3 may reappear in 2012.

Chlamydia pneumoniae is genetically diverse in animals and appears to have crossed the host barrier to humans on (at least) two occasions

Chlamydia pneumoniae is a common human and animal pathogen associated with a wide range of diseases. Since the first isolation of C. pneumoniae TWAR in 1965, all human isolates have been essentially clonal, providing little evolutionary insight. To address this gap, we investigated the genetic diversity of 30 isolates from diverse geographical locations, from both human and animal origin (amphibian, reptilian, equine and marsupial). Based on the level of variation that we observed at 23 discreet gene loci, it was clearly evident that the animal isolates were more diverse than the isolates of human origin. Furthermore, we show that C. pneumoniae isolates could be grouped into five major genotypes, A-E, with A, B, D and E genotypes linked by geographical location, whereas genotype C was found across multiple continents. Our evidence strongly supports two separate animal-to-human cross species transfer events in the evolutionary history of this pathogen. The C. pneumoniae human genotype identified in the USA, Canada, Taiwan, Iran, Japan, Korea and Australia (non- Indigenous) most likely originated from a single amphibian or reptilian lineage, which appears to have been previously geographically widespread. We identified a separate human lineage present in two Australian Indigenous isolates (independent geographical locations). This lineage is distinct and is present in Australian amphibians as well as a range of Australian marsupials.

Mycobacterium lentiflavum in Drinking Water Supplies, Australia

Mycobacterium lentiflavum, a slow-growing nontuberculous mycobacterium, is a rare cause of human disease. It has been isolated from environmental samples worldwide. To assess the clinical significance of M. lentiflavum isolates reported to the Queensland Tuberculosis Control Centre, Australia, during 2001-2008, we explored the genotypic similarity and geographic relationship between isolates from humans and potable water in the Brisbane metropolitan area. A total of 47 isolates from 36 patients were reported; 4 patients had clinically significant disease. M. lentiflavum was cultured from 13 of 206 drinking water sites. These sites overlapped geographically with home addresses of the patients who had clinically significant disease. Automated repetitive sequence-based PCR genotyping showed a dominant environmental clone closely related to clinical strains. This finding suggests potable water as a possible source of M. lentiflavum infection in humans.

Highly discriminatory single-nucleotide polymorphism interrogation of Escherichia coli by use of allele-specific real-time PCR and eBURST analysis

In total, 782 Escherichia coli strains originating from various host sources have been analyzed in this study by using a highly discriminatory single-nucleotide polymorphism (SNP) approach. A set of eight SNPs, with a discrimination value (Simpson's index of diversity [D]) of 0.96, was determined using the Minimum SNPs software, based on sequences of housekeeping genes from the E. coli multilocus sequence typing (MLST) database. Allele-specific real-time PCR was used to screen 114 E. coli isolates from various fecal sources in Southeast Queensland (SEQ). The combined analysis of both the MLST database and SEQ E. coli isolates using eight high-D SNPs resolved the isolates into 74 SNP profiles. The data obtained suggest that SNP typing is a promising approach for the discrimination of host-specific groups and allows for the identification of human-specific E. coli in environmental samples. However, a more diverse E. coli collection is required to determine animal- and environment-specific E. coli SNP profiles due to the abundance of human E. coli strains (56%) in the MLST database.

Chlamydia muridarum major outer membrane protein-specific antibodies inhibit in vitro infection but enhance pathology in vivo

PROBLEM Chlamydia trachomatis is a significant worldwide health problem, and the often-asymptomatic disease can result in infertility. To develop a successful vaccine, a complete understanding of the immune response to chlamydial infection and development of genital tract pathology is required. METHOD OF STUDY We utilized the murine genital model of chlamydial infection. Mice were immunized with chlamydial major outer membrane protein, and vaginal lavage was assessed for the presence of neutralizing antibodies. These samples were then pre-incubated with Chlamydia muridarum and administered to the vaginal vaults of immune-competent female BALB/c mice to determine the effect on infection. RESULTS The administration of C. muridarum in conjunction with neutralizing antibodies reduced the numbers of mice infected, but a surprising finding was that this accelerated the development of severe oviduct pathology. CONCLUSION Antibodies play an under-recognized role in chlamydial infection and pathology development, which possibly involves interaction with Th1 immunity.

Anticipated behaviours of emergency prehospital medical care providers during an influenza pandemic

Introduction: Emergency prehospital medical care providers are frontline health workers during emergencies. However, little is known about their attitudes, perceptions, and likely behaviors during emergency conditions. Understanding these attitudes and behaviors is crucial to mitigating the psychological and operational effects of biohazard events such as pandemic influenza, and will support the business continuity of essential prehospital services. ----- ----- Problem: This study was designed to investigate the association between knowledge and attitudes regarding avian influenza on likely behavioral responses of Australian emergency prehospital medical care providers in pandemic conditions. ----- ----- Methods: Using a reply-paid postal questionnaire, the knowledge and attitudes of a national, stratified, random sample of the Australian emergency prehospital medical care workforce in relation to pandemic influenza were investigated. In addition to knowledge and attitudes, there were five measures of anticipated behavior during pandemic conditions: (1) preparedness to wear personal protective equipment (PPE); (2) preparedness to change role; (3) willingness to work; and likely refusal to work with colleagues who were exposed to (4) known and (5) suspected influenza. Multiple logistic regression models were constructed to determine the independent predictors of each of the anticipated behaviors, while controlling for other relevant variables. ----- ----- Results: Almost half (43%) of the 725 emergency prehospital medical care personnel who responded to the survey indicated that they would be unwilling to work during pandemic conditions; one-quarter indicated that they would not be prepared to work in PPE; and one-third would refuse to work with a colleague exposed to a known case of pandemic human influenza. Willingness to work during a pandemic (OR = 1.41; 95% CI = 1.0–1.9), and willingness to change roles (OR = 1.44; 95% CI = 1.04–2.0) significantly increased with adequate knowledge about infectious agents generally. Generally, refusal to work with exposed (OR = 0.48; 95% CI = 0.3–0.7) or potentially exposed (OR = 0.43; 95% CI = 0.3–0.6) colleagues significantly decreased with adequate knowledge about infectious agents. Confidence in the employer’s capacity to respond appropriately to a pandemic significantly increased employee willingness to work (OR = 2.83; 95% CI = 1.9–4.1); willingness to change roles during a pandemic (OR = 1.52; 95% CI = 1.1–2.1); preparedness to wear PPE (OR = 1.68; 95% CI = 1.1–2.5); and significantly decreased the likelihood of refusing to work with colleagues exposed to (suspected) influenza (OR = 0.59; 95% CI = 0.4–0.9). ----- ----- Conclusions:These findings indicate that education and training alone will not adequately prepare the emergency prehospital medical workforce for a pandemic. It is crucial to address the concerns of ambulance personnel and the perceived concerns of their relationship with partners in order to maintain an effective prehospital emergency medical care service during pandemic conditions.

Attitudes to living and working in pandemic conditions among emergency prehospital medical care personnel

Introduction: Little is known about the risk perceptions and attitudes of healthcare personnel, especially of emergency prehospital medical care personnel, regarding the possibility of an outbreak or epidemic event. Problem: This study was designed to investigate pre-event knowledge and attitudes of a national sample of the emergency prehospital medical care providers in relation to a potential human influenza pandemic, and to determine predictors of these attitudes. Methods: Surveys were distributed to a random, cross-sectional sample of 20% of the Australian emergency prehospital medical care workforce (n = 2,929), stratified by the nine services operating in Australia, as well as by gender and location. The surveys included: (1) demographic information; (2) knowledge of influenza; and (3) attitudes and perceptions related to working during influenza pandemic conditions. Multiple logistic regression models were constructed to identify predictors of pandemic-related risk perceptions. Results: Among the 725 Australian emergency prehospital medical care personnel who responded, 89% were very anxious about working during pandemic conditions, and 85% perceived a high personal risk associated with working in such conditions. In general, respondents demonstrated poor knowledge in relation to avian influenza, influenza generally, and infection transmission methods. Less than 5% of respondents perceived that they had adequate education/training about avian influenza. Logistic regression analyses indicate that, in managing the attitudes and risk perceptions of emergency prehospital medical care staff, particular attention should be directed toward the paid, male workforce (as opposed to volunteers), and on personnel whose relationship partners do not work in the health industry. Conclusions: These results highlight the potentially crucial role of education and training in pandemic preparedness. Organizations that provide emergency prehospital medical care must address this apparent lack of knowledge regarding infection transmission, and procedures for protection and decontamination. Careful management of the perceptions of emergency prehospital medical care personnel during a pandemic is likely to be critical in achieving an effective response to a widespread outbreak of infectious disease.

Can the contra-lateral limb be used as a control with respect to analyses of bone remodelling?

Bone loss may result from remodelling initiated by implant stress protection. Quantifying remodelling requires bone density distributions which can be obtained from computed tomography scans. Pre-operative scans of large animals however are rarely possible. This study aimed to determine if the contra-lateral bone is a suitable control for the purpose of quantifying bone remodelling. CT scans of 8 pairs of ovine tibia were used to determine the likeness of left and right bones. The deviation between the outer surfaces of the bone pairs was used to quantify geometric similarity. The density differences were determined by dividing the bones into discrete volumes along the shaft of the tibia. Density differences were also determined for fractured and contra-lateral bone pairs to determine the magnitude of implant related remodelling. Left and right ovine tibiae were found to have a high degree of similarity with differences of less than 1.0 mm in the outer surface deviation and density difference of less than 5% in over 90% of the shaft region. The density differences (10–40%) as a result of implant related bone remodelling were greater than left-right differences. Therefore, for the purpose of quantifying bone remodelling in sheep, the contra-lateral tibia may be considered an alternative to a pre-operative control.

Diversity of community acquired MRSA carrying the PVL gene in Queensland and New South Wales, Australia

Emergence and dissemination of community acquired methicillin resistant Staphylococcus aureus (CA-MRSA) strains are being reported with increasing frequency in Australia and worldwide. These strains of CA-MRSA are genetically diverse and distinct in Australia. Genotyping of CA-MRSA using eight highly-discriminatory single nucleotide polymorphisms (SNPs) is a rapid and robust method for monitoring the dissemination of these strains in the community. In this study, a SNP genotyping method was used to investigate the molecular epidemiology of 249 community acquired non-multiresistant MRSA (nm-MRSA) isolates over a 12-month period from routine diagnostic specimens. A real-time PCR for the presence of Panton-Valentine leukocidin (PVL) was also performed on these isolates. The CA-MRSA isolates were sourced from a large private laboratory in Brisbane, Australia that serves a wide geographic region encompassing Queensland and Northern New South Wales. This study identified 16 different STs and 98% of the CA-MRSA isolates were positive for the PVL gene. The most common ST was ST93 with 41% of isolates testing positive for this clone.

A mathematical model of chlamydial infection incorporating movement of chlamydial particles

We present a spatiotemporal mathematical model of chlamydial infection, host immune response and spatial movement of infectious particles. The re- sulting partial differential equations model both the dynamics of the infection and changes in infection profile observed spatially along the length of the host genital tract. This model advances previous chlamydia modelling by incorporating spatial change, which we also demonstrate to be essential when the timescale for movement of infectious particles is equal to, or shorter than, the developmental cycle timescale. Numerical solutions and model analysis are carried out, and we present a hypothesis regarding the potential for treatment and prevention of infection by increasing chlamydial particle motility.

Rodent blood-stage Plasmodium survive in dendritic cells that infect naive mice.

Plasmodium spp. parasites cause malaria in 300 to 500 million individuals each year. Disease occurs during the blood-stage of the parasite’s life cycle, where the parasite is thought to replicate exclusively within erythrocytes. Infected individuals can also suffer relapses after several years, from Plasmodium vivax and Plasmodium ovale surviving in hepatocytes. Plasmodium falciparum and Plasmodium malariae can also persist after the original bout of infection has apparently cleared in the blood, suggesting that host cells other than erythrocytes (but not hepatocytes) may harbor these blood-stage parasites, thereby assisting their escape from host immunity. Using blood stage transgenic Plasmodium berghei-expressing GFP (PbGFP) to track parasites in host cells, we found that the parasite had a tropism for CD317+ dendritic cells. Other studies using confocal microscopy, in vitro cultures, and cell transfer studies showed that blood-stage parasites could infect, survive, and replicate within CD317+ dendritic cells, and that small numbers of these cells released parasites infectious for erythrocytes in vivo. These data have identified a unique survival strategy for blood-stage Plasmodium, which has significant implications for understanding the escape of Plasmodium spp. from immune-surveillance and for vaccine development.

Transcutaneous immunization with novel lipid-based adjuvants induces protection against gastric Helicobacter pylori infection

The development of vaccines to combat pathogens that infect across mucosal surfaces has been a major goal of vaccine research. Successful mucosal vaccination requires the co-administration of adjuvants that can overcome the state of immune tolerance normally associated with mucosal application of proteins. In the case of oral immunization, delivery systems are also required to protect vaccine antigens against destruction by gastric pH and digestive enzymes. Furthermore, adjuvants used for mucosal delivery must be free of neurotoxic effects like those induced by the commonly used experimental mucosal adjuvant cholera toxin. Maintenance of the "cold chain" is also essential for the effectiveness of any vaccine and adjuvants/delivery systems that enhance the stability of a vaccine would offer a significant advantage. Needle-free methods of vaccination that induce protective immunity at multiple mucosal surfaces are also desirable for rapid vaccination of large populations. In the present study we show that transcutaneous immunization (TCI) using Lipid C, a novel lipid-based matrix originally developed for oral immunization, containing soluble Helicobacter sonicate significantly reduces the gastric bacterial burden in mice following gastric challenge with live Helicobacter pylori. Protection is associated with the production of splenic gamma interferon and gastric IgA and was achieved without the co-administration of potent and potentially toxic adjuvants, although protection was further enhanced by inclusion of CpG-ODN and cholera toxin in the lipid delivery system.

The spermostatic and microbicidal actions of quinones and malemides : towards a dual-purpose contraceptive agent

There is an urgent need to develop safe, effective, dual-purpose contraceptive agents that combine the prevention of pregnancy with protection against sexually transmitted diseases. Here we report the identification of a group of compounds that on contact with human spermatozoa induce a state of “spermostasis,” characterized by the extremely rapid inhibition of sperm movement without compromising cell viability. These spermostatic agents were more active and significantly less toxic than the reagent in current clinical use, nonoxynol 9, giving therapeutic indices (ratio of spermostatic to cytotoxic activity) that were orders of magnitude greater than this traditional spermicide. Although certain compounds could trigger reactive oxygen species generation by spermatozoa, this activity was not correlated with spermostasis. Rather, the latter was associated with alkylation of two major sperm tail proteins that were identified as A Kinase-Anchoring Proteins (AKAP3 and AKAP4) by mass spectrometry. As a consequence of disrupted AKAP function, the abilities of cAMP to drive protein kinase A-dependent activities in the sperm tail, such as the activation of SRC and the consequent stimulation of tyrosine phosphorylation, were suppressed. Furthermore, analysis of microbicidal activity using Chlamydia muridarum revealed powerful inhibitory effects at the same low micromolar doses that suppressed sperm movement. In this case, the microbicidal action was associated with alkylation of Major Outer Membrane Protein (MOMP), a major chlamydial membrane protein. Taken together, these results have identified for the first time a novel set of cellular targets and chemical principles capable of providing simultaneous defense against both fertility and the spread of sexually transmitted disease.