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.

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.

Comparison of koala LPCoLN and human strains of Chlamydia pneumoniae highlights extended genetic diversity in the species

Background Chlamydia pneumoniae is a widespread pathogen causing upper and lower respiratory tract infections in addition to a range of other diseases in humans and animals. Previous whole genome analyses have focused on four essentially clonal (> 99% identity) C. pneumoniae human genomes (AR39, CWL029, J138 and TW183), providing relatively little insight into strain diversity and evolution of this species. Results We performed individual gene-by-gene comparisons of the recently sequenced C. pneumoniae koala genome and four C. pneumoniae human genomes to identify species-specific genes, and more importantly, to gain an insight into the genetic diversity and evolution of the species. We selected genes dispersed throughout the chromosome, representing genes that were specific to C. pneumoniae, genes with a demonstrated role in chlamydial biology and/or pathogenicity (n = 49), genes encoding nucleotide salvage or amino acid biosynthesis proteins (n = 6), and extrachromosomal elements (9 plasmid and 2 bacteriophage genes). Conclusions We have identified strain-specific differences and targets for detection of C. pneumoniae isolates from both human and animal origin. Such characterisation is necessary for an improved understanding of disease transmission and intervention.

Identification of a cryptic Prokaryotic promoter within the cDNA encoding the 5′ End of Dengue Virus RNA Genome

Infectious cDNA clones of RNA viruses are important research tools, but flavivirus cDNA clones have proven difficult to assemble and propagate in bacteria. This has been attributed to genetic instability and/or host cell toxicity, however the mechanism leading to these difficulties has not been fully elucidated. Here we identify and characterize an efficient cryptic bacterial promoter in the cDNA encoding the dengue virus (DENV) 5′ UTR. Following cryptic transcription in E. coli, protein expression initiated at a conserved in-frame AUG that is downstream from the authentic DENV initiation codon, yielding a DENV polyprotein fragment that was truncated at the N-terminus. A more complete understanding of constitutive viral protein expression in E. coli might help explain the cloning and propagation difficulties generally observed with flavivirus cDNA.

Partial protection against chlamydial reproductive tract infection by a recombinant major outer membrane protein/CpG/cholera toxin intranasal vaccine in the guinea pig Chlamydia cavaie model

Chlamydia trachomatis is a major cause of sexually transmitted diseases worldwide. There currently is no vaccine to protect against chlamydial infection of the female reproductive tract. Vaccine development has predominantly involved using the murine model, however infection of female guinea pigs with Chlamydia caviae more closely resembles chlamydial infection of the human female reproductive tract, and presents a better model to assess potential human chlamydial vaccines. We immunised female guinea pigs intranasally with recombinant major outer membrane protein (r-MOMP) combined with CpG-10109 and cholera toxin adjuvants. Both systemic and mucosal immune responses were elicited in immunised animals. MOMP-specific IgG and IgA were present in the vaginal mucosae, and high levels of MOMP-specific IgG were detected in the serum of immunised animals. Antibodies from the vaginal mucosae were also shown to be capable of neutralising C. caviae in vitro. Following immunisation, animals were challenged intravaginally with a live C. caviae infection of 102 inclusion forming units. We observed a decrease in duration of infection and a significant (p<0.025) reduction in infection load in r-MOMP immunised animals, compared to animals immunised with adjuvant only. Importantly, we also observed a marked reduction in upper reproductive tract (URT) pathology in r-MOMP immunised animals. Intranasal immunisation of female guinea pigs with r-MOMP was able to provide partial protection against C. caviae infection, not only by reducing chlamydial burden but also URT pathology. This data demonstrates the value of using the guinea pig model to evaluate potential chlamydial vaccines for protection against infection and disease pathology caused by C. trachomatis in the female reproductive tract.

A comparison of the effects of a chlamydial vaccine administered during or after a C. muridarum urogenital infection of female mice

There are approximately 92 million new chlamydial infections of the genital tract in humans diagnosed each year, costing health care systems billions of dollars in treatment not only of acute infections, but also of associated inflammatory sequelae, such as pelvic inflammatory disease (PID) and ectopic pregnancy. These numbers are increasing at a steady rate and, due to the asymptomatic nature of infections, the incidence may be underestimated and the costs of treatment therefore higher. Over the previous few decades there has been a large amount of research into the development of an efficacious vaccine against genital tract chlamydial infections. The majority of this research has focused on females, due to the high rate of development of associated diseases, including PID, which can lead to ectopic pregnancy and infertility. In light of the increasing infection rates that have occurred despite the availability of antibiotics, and the asymptomatic nature of chlamydial infections, it is imperative that an efficacious vaccine that protects against infection and associated pathology be developed.