Persistent chlamydial infections : role in inflammation and challenges for vaccine development.

The development of chlamydial vaccines continues to be a major challenge for researchers. While acute infections are the main target of vaccine development groups, Chlamydia is well known for its ability to establish chronic or persistent infections in its host. To date, little effort has focussed specifically on the challenges of vaccines to successfully combat the chronic or persistent phase of the disease and yet this will be a necessary aspect of any fully successful chlamydial vaccine. This short review specifically examines the phenomenon of chlamydial persistence and the unique challenges that this brings to vaccine development.

Evidence that human chlamydia pneumoniae was zoonotically acquired

Zoonotic infections are a growing threat to global health. Chlamydia pneumoniae is a major human pathogen that is widespread in human populations, causing acute respiratory disease, and has been associated with chronic disease. C. pneumoniae was first identified solely in human populations; however, its host range now includes other mammals, marsupials, amphibians, and reptiles. Australian koalas (Phascolarctos cinereus) are widely infected with two species of Chlamydia, C. pecorum and C. pneumoniae. Transmission of C. pneumoniae between animals and humans has not been reported; however, two other chlamydial species, C. psittaci and C. abortus, are known zoonotic pathogens. We have sequenced the 1,241,024-bp chromosome and a 7.5-kb cryptic chlamydial plasmid of the koala strain of C. pneumoniae (LPCoLN) using the whole-genome shotgun method. Comparative genomic analysis, including pseudogene and single-nucleotide polymorphism (SNP) distribution, and phylogenetic analysis of conserved genes and SNPs against the human isolates of C. pneumoniae show that the LPCoLN isolate is basal to human isolates. Thus, we propose based on compelling genomic and phylogenetic evidence that humans were originally infected zoonotically by an animal isolate(s) of C. pneumoniae which adapted to humans primarily through the processes of gene decay and plasmid loss, to the point where the animal reservoir is no longer required for transmission.

Morphologic and genomic characterisation of the koala Chlamydia pneumoniae strain

Chlamydia pneumoniae is a common human and animal pathogen associated with a wide range of upper and lower respiratory tract infections. In more recent years there has been increasing evidence to suggest a link between C. pneumoniae and chronic diseases in humans, including atherosclerosis, stroke and Alzheimer’s disease. C. pneumoniae human strains show little genetic variation, indicating that the human-derived strain originated from a common ancestor in the recent past. Despite extensive information on the genetics and morphology processes of the human strain, knowledge concerning many other hosts (including marsupials, amphibians, reptiles and equines) remains virtually unexplored. The koala (Phascolarctos cinereus) is a native Australian marsupial under threat due to habitat loss, predation and disease. Koalas are very susceptible to chlamydial infections, most commonly affecting the conjunctiva, urogenital tract and/or respiratory tract. To address this gap in the literature, the present study (i) provides a detailed description of the morphologic and genomic architecture of the C. pneumoniae koala (and human) strain, and shows that the koala strain is microscopically, developmentally and genetically distinct from the C. pneumoniae human strain, and (ii) examines the genetic relationship of geographically diverse C. pneumoniae isolates from human, marsupial, amphibian, reptilian and equine hosts, and identifies two distinct lineages that have arisen from animal-to-human cross species transmissions. Chapter One of this thesis explores the scientific problem and aims of this study, while Chapter Two provides a detailed literature review of the background in this field of work. Chapter Three, the first results chapter, describes the morphology and developmental stages of C. pneumoniae koala isolate LPCoLN, as revealed by fluorescence and transmission electron microscopy. The profile of this isolate, when cultured in HEp-2 human epithelial cells, was quite different to the human AR39 isolate. Koala LPCoLN inclusions were larger; the elementary bodies did not have the characteristic pear-shaped appearance, and the developmental cycle was completed within a shorter period of time (as confirmed by quantitative real-time PCR). These in vitro findings might reflect biological differences between koala LPCoLN and human AR39 in vivo. Chapter Four describes the complete genome sequence of the koala respiratory pathogen, C. pneumoniae LPCoLN. This is the first animal isolate of C. pneumoniae to be fully-sequenced. The genome sequence provides new insights into genomic ‘plasticity’ (organisation), evolution and biology of koala LPCoLN, relative to four complete C. pneumoniae human genomes (AR39, CWL029, J138 and TW183). Koala LPCoLN contains a plasmid that is not shared with any of the human isolates, there is evidence of gene loss in nucleotide salvage pathways, and there are 10 hot spot genomic regions of variation that were previously not identified in the C. pneumoniae human genomes. Sequence (partial-length) from a second, independent, wild koala isolate (EBB) at several gene loci confirmed that the koala LPCoLN isolate was representative of a koala C. pneumoniae strain. The combined sequence data provides evidence that the C. pneumoniae animal (koala LPCoLN) genome is ancestral to the C. pneumoniae human genomes and that human infections may have originated from zoonotic infections. Chapter Five examines key genome components of the five C. pneumoniae genomes in more detail. This analysis reveals genomic features that are shared by and/or contribute to the broad ecological adaptability and evolution of C. pneumoniae. This analysis resulted in the identification of 65 gene sequences for further analysis of intraspecific variation, and revealed some interesting differences, including fragmentation, truncation and gene decay (loss of redundant ancestral traits). This study provides valuable insights into metabolic diversity, adaptation and evolution of C. pneumoniae. Chapter Six utilises a subset of 23 target genes identified from the previous genomic comparisons and makes a significant contribution to our understanding of genetic variability among C. pneumoniae human (11) and animal (6 amphibian, 5 reptilian, 1 equine and 7 marsupial hosts) isolates. It has been shown that the animal isolates are genetically diverse, unlike the human isolates that are virtually clonal. More convincing evidence that C. pneumoniae originated in animals and recently (in the last few hundred thousand years) crossed host species to infect humans is provided in this study. It is proposed that two animal-to-human cross species events have occurred in the context of the results, one evident by the nearly clonal human genotype circulating in the world today, and the other by a more animal-like genotype apparent in Indigenous Australians. Taken together, these data indicate that the C. pneumoniae koala LPCoLN isolate has morphologic and genomic characteristics that are distinct from the human isolates. These differences may affect the survival and activity of the C. pneumoniae koala pathogen in its natural host, in vivo. This study, by utilising the genetic diversity of C. pneumoniae, identified new genetic markers for distinguishing human and animal isolates. However, not all C. pneumoniae isolates were genetically diverse; in fact, several isolates were highly conserved, if not identical in sequence (i.e. Australian marsupials) emphasising that at some stage in the evolution of this pathogen, there has been an adaptation/s to a particular host, providing some stability in the genome. The outcomes of this study by experimental and bioinformatic approaches have significantly enhanced our knowledge of the biology of this pathogen and will advance opportunities for the investigation of novel vaccine targets, antimicrobial therapy, or blocking of pathogenic pathways.

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.

Tubal damage, infertility and tubal ectopic pregnancy : chlamydia trachomatis and other microbial aetiologies

Infertility is a worldwide health problem with one in six couples suffering from this condition and with a major economic burden on the global healthcare industry. Estimates of the current global infertility rate suggest that 15% of couples are infertile (Zegers-Hochschild et al 2009) defined as: (1) failure to conceive after 12 months of unprotected sexual intercourse (i.e. infertility); (2) repeated implantation failure following ART cycles; or (3) recurrent miscarriage without difficulty conceiving (natural conceptions). Tubal factor infertility is among the leading causes of female factor infertility accounting for 7-9.8% of all female factor infertilities. Tubal disease directly causes from 36% to 85% of all cases of female factor infertility in developed and developing nations respectively and is associated with polymicrobial aetiologies. One of the leading global causes of tubal factor infertility is thought to be symptomatic (and asymptomatic in up to 70% cases) infection of the female reproductive tract with the sexually transmitted pathogen, Chlamydia trachomatis. Infection-related damage to the Fallopian tubes caused by Chlamydia accounts for more than 70% of cases of infertility in women from developing nations such as sub-Saharan Africa (Sharma et al 2009). Bacterial vaginosis, a condition associated with increased transmission of sexually transmitted infections including those caused by Neisseria gonorrhoeae and Mycoplasma genitalium is present in two thirds of women with pelvic inflammatory disease (PID). This review will focus on (1) the polymicrobial aetiologies of tubal factor infertility and (2) studies involved in screening for, and treatment and control of, Chlamydial infection to prevent PID and the associated sequelae of Fallopian tube inflammation that may lead to infertility and ectopic pregnancy.

Comparison of intranasal and transcutaneous immunization for induction of protective immunity against chlamydia muridarum respiratory tract infection

Chlamydia pneumoniae causes a range of respiratory infections including bronchitis, pharyngitis and pneumonia. Infection has also been implicated in exacerbation/initiation of asthma and chronic obstructive pulmonary disease (COPD) and may play a role in atherosclerosis and Alzheimer's disease. We have used a mouse model of Chlamydia respiratory infection to determine the effectiveness of intranasal (IN) and transcutaneous immunization (TCI) to prevent Chlamydia lung infection. Female BALB/c mice were immunized with chlamydial major outer membrane protein (MOMP) mixed with cholera toxin and CpG oligodeoxynucleotide adjuvants by either the IN or TCI routes. Serum and bronchoalveolar lavage (BAL) were collected for antibody analysis. Mononuclear cells from lung-draining lymph nodes were stimulated in vitro with MOMP and cytokine mRNA production determined by real time PCR. Animals were challenged with live Chlamydia and weighed daily following challenge. At day 10 (the peak of infection) animals were sacrificed and the numbers of recoverable Chlamydia in lungs determined by real time PCR. MOMP-specific antibody-secreting cells in lung tissues were also determined at day 10 post-infection. Both IN and TCI protected animals against weight loss compared to non-immunized controls with both immunized groups gaining weight by day 10-post challenge while controls had lost 6% of body weight. Both immunization protocols induced MOMP-specific IgG in serum and BAL while only IN immunization induced MOMP-specific IgA in BAL. Both immunization routes resulted in high numbers of MOMP-specific antibody-secreting cells in lung tissues (IN > TCI). Following in vitro re-stimulation of lung-draining lymph node cells with MOMP; IFNγ mRNA increased 20-fold in cells from IN immunized animals (compared to non-immunized controls) while IFNγ levels increased 6- to 7-fold in TCI animals. Ten days post challenge non-immunized animals had >7000 IFU in their lungs, IN immunized animals <50 IFU and TCI immunized animals <1500 IFU. Thus, both intranasal and transcutaneous immunization protected mice against respiratory challenge with Chlamydia. The best protection was obtained following IN immunization and correlated with IFNγ production by mononuclear cells in lung-draining LN and MOMP-specific IgA in BAL.

Chlamydia

Chlamydial infections of humans can cause blindness and infertility as a result of diseases such as keratoconjunctivitis (trachoma), urethritis and cervicitis. However, in greater than half of all chlamydial diseases in males and females there are no signs or symptoms of infection. Chlamydia trachomatis is the causative bacterial organism responsible for the global estimate of 40.6 million people currently suffering with active trachoma and for the five million new cases of sexually transmitted infections each year in the United States of America. Even though antibiotics are available to treat Chlamydia, the incidence of each of these primarily asymptomatic infections continues to increase. In this Chapter we review the current knowledge of C.trachomatis including clinicial diseases and sequelae, the chlamydial developmental cycle in vivo, immunobiology and immune responses to infections, chlamydial genomics and vaccine development.

Development of novel vaccine strategies to prevent genital tract chlamydial infections

Chlamydia trachomatis is the most prevalent bacterial sexually transmitted infection in the developed world and the leading cause of preventable blindness worldwide. As reported by the World Health Organization in 2001, there are approximately 92 million new infections detected annually, costing health systems billions of dollars to treat not only the acute infection, but also to treat infection-associated sequelae. The majority of genital infections are asymptomatic, with 50-70% going undetected. Genital tract infections can be easily treated with antibiotics when detected. Lack of treatment can lead to the development of pelvic inflammatory disease, ectopic pregnancies and tubal factor infertility in women and epididymitis and prostatitis in men. With infection rates on the continual rise and the large number of infections going undetected, there is a need to develop an efficacious vaccine which prevents not only infection, but also the development of infection-associated pathology. Before a vaccine can be developed and administered, the pathogenesis of chlamydial infections needs to be fully understood. This includes the kinetics of ascending infection and the effects of inoculating dose on ascension and development of pathology. The first aim in this study was to examine these factors in a murine model. Female BALB/c mice were infected intravaginally with varying doses of C. muridarum, the mouse variant of human C. trachomatis, and the ascension of infection along the reproductive tract and the time-course of infection-associated pathology development, including inflammatory cell infiltration, pyosalpinx and hydrosalpinx, were determined. It was found that while the inoculating dose did affect the rate and degree of infection, it did not affect any of the pathological parameters examined. This highlighted that the sexual transmission dose may have minimal effect on the development of reproductive sequelae. The results of the first section enabled further studies presented here to use an optimal inoculating dose that would ascend the reproductive tract and cause pathology development, so that vaccine efficacy could be determined. There has been a large amount of research into the development of an efficacious vaccine against genital tract chlamydial infections, with little success. However, there have been no studies examining the effects of the timing of vaccination, including the effects of vaccination during an active genital infection, or after clearance of a previous infection. These are important factors that need to be examined, as it is not yet known whether immunization will enhance not only the individual's immune response, but also pathology development. It is also unknown whether any enhancement of the immune responses will cause the Chlamydia to enter a dormant, persistent state, and possibly further enhance any pathology development. The second section of this study aimed to determine if vaccination during an active genital tract infection, or after clearance of a primary infection, enhanced the murine immune responses and whether any enhanced or reduced pathology occurred. Naïve, actively infected, or previously infected animals were immunized intranasally or transcutaneously with the adjuvants cholera toxin and CpG-ODN in combination with either the major outer membrane protein (MOMP) of C. muridarum, or MOMP and ribonucleotide reductase small chain protein (NrdB) of C. muridarum. It was found that the systemic immune responses in actively or previously infected mice were altered in comparison to animals immunized naïve with the same combinations, however mucosal antibodies were not enhanced. It was also found that there was no difference in pathology development between any of the groups. This suggests that immunization of individuals who may have an asymptomatic infection, or may have been previously exposed to a genital infection, may not benefit from vaccination in terms of enhanced immune responses against re-exposure. The final section of this study aimed to determine if the vaccination regimes mentioned above caused in vivo persistence of C. muridarum in the upper reproductive tracts of mice. As there has been no characterization of C. muridarum persistence in vitro, either ultrastructurally or via transcriptome analysis, this was the first aim of this section. Once it had been shown that C. muridarum could be induced into a persistent state, the gene transcriptional profiles of the selected persistent marker genes were used to determine if persistent infections were indeed present in the upper reproductive tracts of the mice. We found that intranasal immunization during an active infection induced persistent infections in the oviducts, but not the uterine horns, and that intranasal immunization after clearance of infection, caused persistent infections in both the uterine horns and the oviducts of the mice. This is a significant finding, not only because it is the first time that C. muridarum persistence has been characterized in vitro, but also due to the fact that there is minimal characterization of in vivo persistence of any chlamydial species. It is possible that the induction of persistent infections in the reproductive tract might enhance the development of pathology and thereby enhance the risk of infertility, factors that need to be prevented by vaccination, not enhanced. Overall, this study has shown that the inoculating dose does not affect pathology development in the female reproductive tract of infected mice, but does alter the degree and rate of ascending infection. It has also been shown that intranasal immunization during an active genital infection, or after clearance of one, induces persistent infections in the uterine horns and oviducts of mice. This suggests that potential vaccine candidates will need to have these factors closely examined before progressing to clinical trials. This is significant, because if the same situation occurs in humans, a vaccine administered to an asymptomatic, or previously exposed individual may not afford any extra protection and may in fact enhance the risk of development of infection-associated sequelae. This suggests that a vaccine may serve the community better if administered before the commencement of sexual activity.

Novel molecular markers of Chlamydia pecorum genetic diversity in the koala (Phascolarctos cinereus)

Background Chlamydia pecorum is an obligate intracellular bacterium and the causative agent of reproductive and ocular disease in several animal hosts including koalas, sheep, cattle and goats. C. pecorum strains detected in koalas are genetically diverse, raising interesting questions about the origin and transmission of this species within koala hosts. While the ompA gene remains the most widely-used target in C. pecorum typing studies, it is generally recognised that surface protein encoding genes are not suited for phylogenetic analysis and it is becoming increasingly apparent that the ompA gene locus is not congruent with the phylogeny of the C. pecorum genome. Using the recently sequenced C. pecorum genome sequence (E58), we analysed 10 genes, including ompA, to evaluate the use of ompA as a molecular marker in the study of koala C. pecorum genetic diversity. Results Three genes (incA, ORF663, tarP) were found to contain sufficient nucleotide diversity and discriminatory power for detailed analysis and were used, with ompA, to genotype 24 C. pecorum PCR-positive koala samples from four populations. The most robust representation of the phylogeny of these samples was achieved through concatenation of all four gene sequences, enabling the recreation of a "true" phylogenetic signal. OmpA and incA were of limited value as fine-detailed genetic markers as they were unable to confer accurate phylogenetic distinctions between samples. On the other hand, the tarP and ORF663 genes were identified as useful "neutral" and "contingency" markers respectively, to represent the broad evolutionary history and intra-species genetic diversity of koala C. pecorum. Furthermore, the concatenation of ompA, incA and ORF663 sequences highlighted the monophyletic nature of koala C. pecorum infections by demonstrating a single evolutionary trajectory for koala hosts that is distinct from that seen in non-koala hosts. Conclusions While the continued use of ompA as a fine-detailed molecular marker for epidemiological analysis appears justified, the tarP and ORF663 genes also appear to be valuable markers of phylogenetic or biogeographic divisions at the C. pecorum intra-species level. This research has significant implications for future typing studies to understand the phylogeny, genetic diversity, and epidemiology of C. pecorum infections in the koala and other animal species.

Vaccination of healthy and diseased koalas (Phascolarctos cinereus) with a Chlamydia pecorum multi-subunit vaccine : evaluation of immunity and pathology

Chlamydial infections represent a major threat to the long-term survival of the koala and a successful vaccine would provide a valuable management tool. Vaccination however has the potential to enhance inflammatory disease in animals exposed to a natural infection prior to vaccination, a finding in early human and primate trials of whole cell vaccines to prevent trachoma. In the present study, we vaccinated both healthy koalas as well as clinically diseased koalas with a multi-subunit vaccine consisting of Chlamydia pecorum MOMP and NrdB mixed with immune stimulating complex as adjuvant. Following vaccination, there was no increase in inflammatory pathological changes in animals previously infected with Chlamydia. Strong antibody (including neutralizing antibodies) and lymphocyte proliferation responses were recorded in all vaccinated koalas, both healthy and clinically diseased. Vaccine induced antibodies specific for both vaccine antigens were observed not only in plasma but also in ocular secretions. Our data shows that an experimental chlamydial vaccine is safe to use in previously infected koalas, in that it does not worsen infection-associated lesions. Furthermore, the prototype vaccine is effective, as demonstrated by strong levels of neutralizing antibody and lymphocyte proliferation responses in both healthy and clinically diseased koalas. Collectively, this work illustrates the feasibility of developing a safe and effective Chlamydia vaccine as a tool for management of disease in wild koalas.

Evaluation of the cost-effectiveness of strategies claiming to reduce the risk of surgical site infections following primary total hip arthroplasty

Background Total hip arthroplasty (THA) is a commonly performed procedure and numbers are increasing with ageing populations. One of the most serious complications in THA are surgical site infections (SSIs), caused by pathogens entering the wound during the procedure. SSIs are associated with a substantial burden for health services, increased mortality and reduced functional outcomes in patients. Numerous approaches to preventing these infections exist but there is no gold standard in practice and the cost-effectiveness of alternate strategies is largely unknown. Objectives The aim of this project was to evaluate the cost-effectiveness of strategies claiming to reduce deep surgical site infections following total hip arthroplasty in Australia. The objectives were: 1. Identification of competing strategies or combinations of strategies that are clinically relevant to the control of SSI related to hip arthroplasty 2. Evidence synthesis and pooling of results to assess the volume and quality of evidence claiming to reduce the risk of SSI following total hip arthroplasty 3. Construction of an economic decision model incorporating cost and health outcomes for each of the identified strategies 4. Quantification of the effect of uncertainty in the model 5. Assessment of the value of perfect information among model parameters to inform future data collection Methods The literature relating to SSI in THA was reviewed, in particular to establish definitions of these concepts, understand mechanisms of aetiology and microbiology, risk factors, diagnosis and consequences as well as to give an overview of existing infection prevention measures. Published economic evaluations on this topic were also reviewed and limitations for Australian decision-makers identified. A Markov state-transition model was developed for the Australian context and subsequently validated by clinicians. The model was designed to capture key events related to deep SSI occurring within the first 12 months following primary THA. Relevant infection prevention measures were selected by reviewing clinical guideline recommendations combined with expert elicitation. Strategies selected for evaluation were the routine use of pre-operative antibiotic prophylaxis (AP) versus no use of antibiotic prophylaxis (No AP) or in combination with antibiotic-impregnated cement (AP & ABC) or laminar air operating rooms (AP & LOR). The best available evidence for clinical effect size and utility parameters was harvested from the medical literature using reproducible methods. Queensland hospital data were extracted to inform patients’ transitions between model health states and related costs captured in assigned treatment codes. Costs related to infection prevention were derived from reliable hospital records and expert opinion. Uncertainty of model input parameters was explored in probabilistic sensitivity analyses and scenario analyses and the value of perfect information was estimated. Results The cost-effectiveness analysis was performed from a health services perspective using a hypothetical cohort of 30,000 THA patients aged 65 years. The baseline rate of deep SSI was 0.96% within one year of a primary THA. The routine use of antibiotic prophylaxis (AP) was highly cost-effective and resulted in cost savings of over $1.6m whilst generating an extra 163 QALYs (without consideration of uncertainty). Deterministic and probabilistic analysis (considering uncertainty) identified antibiotic prophylaxis combined with antibiotic-impregnated cement (AP & ABC) to be the most cost-effective strategy. Using AP & ABC generated the highest net monetary benefit (NMB) and an incremental $3.1m NMB compared to only using antibiotic prophylaxis. There was a very low error probability that this strategy might not have the largest NMB (<5%). Not using antibiotic prophylaxis (No AP) or using both antibiotic prophylaxis combined with laminar air operating rooms (AP & LOR) resulted in worse health outcomes and higher costs. Sensitivity analyses showed that the model was sensitive to the initial cohort starting age and the additional costs of ABC but the best strategy did not change, even for extreme values. The cost-effectiveness improved for a higher proportion of cemented primary THAs and higher baseline rates of deep SSI. The value of perfect information indicated that no additional research is required to support the model conclusions. Conclusions Preventing deep SSI with antibiotic prophylaxis and antibiotic-impregnated cement has shown to improve health outcomes among hospitalised patients, save lives and enhance resource allocation. By implementing a more beneficial infection control strategy, scarce health care resources can be used more efficiently to the benefit of all members of society. The results of this project provide Australian policy makers with key information about how to efficiently manage risks of infection in THA.

Post-discharge surveillance : can patients reliably diagnose surgical wound infections?

Post-discharge surgical wound infection surveillance is an important part of many infection control programs. It is frequently undertaken by patient self-assessment, prompted either by a telephone or postal questionnaire. To assess the reliability of this method, 290 patients were followed for six weeks postoperatively. Their wounds were photographed and also covertly assessed for signs of infection by two experienced infection control nurses (ICNs). Patients also responded to a postal questionnaire seeking evidence of infection at both week four and week six post-surgery. Correlation between the patient's assessment of their wound and the ICNs diagnosis was poor (r=0.37) with a low positive predictive value (28.7%), although negative predictive value was high (98.2%). Assessment of photos for signs of infection by two experienced clinicians also correlated poorly with the ICNs diagnosis of infection (r=0.54). The patient's recall of prescription of an antibiotic by their general practitioner (GP) for wound infection during the postoperative period correlated best with the ICNs diagnosis (r=0.76). This latter measure, particularly when confirmed by the GP in those patients reporting an infection, appears to provide the most valid and resource efficient marker of post-discharge surgical wound infection.

Comparison of antigen detection and quantitative PCR in the detection of chlamydial infection in koalas (Phascolarctos cinereus)

The gold standard method for detecting chlamydial infection in domestic and wild animals is PCR, but the technique is not suited to testing animals in the field when a rapid diagnosis is frequently required. The objective of this study was to compare the results of a commercially available enzyme immunoassay test for Chlamydia against a quantitative Chlamydia pecorum-specific PCR performed on swabs collected from the conjunctival sac, nasal cavity and urogenital sinuses of naturally infected koalas (Phascolarctos cinereus). The level of agreement for positive results between the two assays was low (43.2%). The immunoassay detection cut-off was determined as approximately 400 C. pecorum copies, indicating that the test was sufficiently sensitive to be used for the rapid diagnosis of active chlamydial infections.