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Adam Polkinghorne, Institute of Health and Biomedical Innovation, Queensland University of Technology

Posted Flock & Herd March 2011


Despite the apparent limitations of their obligate intracellular niche, Chlamydia are highly successful pathogens that can infect and cause disease in animals at nearly every level of taxonomy. In humans, chlamydial infections are the leading cause of preventable blindness and sexually transmitted disease and also cause respiratory infections amongst other chronic disease sequelae.

In non-human mammals, chlamydial infections are a significant cause of disease in both domestic animals and the livestock industry. Common chlamydial infections in domestic animals include chlamydial conjunctivitis in cats, guinea pigs and birds. While information on the role of chlamydial infections in Australian livestock is limited, on other continents (particularly Europe), chlamydial infections are a major cause of morbidity in cattle, pigs and small ruminants. Notably, related chlamydial strains are also a significant cause of disease in koalas and produce similar clinical disease manifestations. The zoonotic potential of avian chlamydial strains has been recognised for over 100 years, but more recently, reports of the zoonotic transmission of chlamydial strains in sheep, cattle, cats and guinea pigs emphasises the need for awareness of chlamydial infections for both human and animal health.

A confounding feature in the diagnosis and treatment of chlamydial infections is that, despite the diverse consequences of infection in humans and animals, chlamydial infections are, however, quite often asymptomatic.


Recent years have seen a number of changes to the taxonomic description of the chlamydiae. Prior to 1999, a one-family genus system was used to classify all members of the order Chlamydiales. Under this system, the genus Chlamydia, the sole member of the family Chlamydiaceae, contained four species (C. trachomatis, C. pneumoniae, C. pecorum and C. psittaci).

Prompted by an awareness of the increased diversity in the order Chlamydiales, a new classification system was proposed based on a range of phenotypic, bio-ecological and genotypic data1. This system separated the family Chlamydiaceae into two lineages of nine species, separating the predominately animal pathogens (C. psittaci, C. pecorum, and C. pneumoniae) into a new genus, Chlamydophila. Three new families (Parachlamydiaceae,Waddliaceae and Simkaniaceae) were also proposed to classify a range of novel Chlamydia-like organisms that are phylogenetically related and share an obligate intracellular lifestyle. This taxonomy was never completely accepted by the chlamydial research community but did highlight the diversity within the chlamydial strains that infect and cause disease in animals.

In 2009, a sub-committee of the International Committee on Systematics of Prokaryotes re-examined the taxonomy and nomenclature issues and a decision was made to remerge species in the genusChlamydophila into Chlamydia2. At this time, the family Chlamydiaceae comprises nine species in the genus Chlamydia: (1) C. trachomatis is a causative agent of sexually transmitted and ocular diseases in humans; (2) C. pneumoniae causes atypical pneumonia in humans and is associated with diseases in reptiles, amphibians and marsupials; (3) C. suis, found only pigs; (4) C. muridarum, found in mice (formerly C. psittaci murine biovar); (5) C. felis (formerlyC. psittaci feline subtype), the causative agent of kerato-conjunctivitis in cats; (6) C. caviae, whose natural host is the guinea pig (formerly C. psittaci guinea pig subtype); (7) C. pecorum, the etiological agent of a range of clinical disease manifestations in cattle, small ruminants and marsupials; (8) C. psittaci, comprised of the avian subtype and etiological agent of psittacosis in birds and humans and; (9) C. abortus, the causative agent of ovine enzootic abortion.

Evidence is building that the Chlamydia-like organisms have also been implicated in animal disease. Two classified species, Parachlamydia acanthamoebae and Waddlia chondrophila, members of the families Parachlamydiaceae and Waddliaceae, have been implicated in abortion in small ruminants. Additionally, sequences of uncultured Chlamydia-like organisms, detected in a range of hosts including humans, koalas, cats, amphibians and fish, are suspected to have a causal role in clinical disease.


The defining feature of Chlamydia is their unique, obligate intracellular developmental cycle, which involves interconversion between two distinct morphological states and environments. The basic developmental cycle commences with the attachment of the extracellular and infectious elementary body (EB) cell to a specific receptor on the host cell surface. This EB is adapted for prolonged extracellular survival, courtesy of its rigid cell envelope and small size and is metabolically dormant. Following attachment to a specific cell receptor, EBs are taken up by endocytosis inside a membrane-bound vesicle (inclusion), which is resistant to phagolysosomal fusion. In this inclusion, EBs convert into a metabolically active reticulate body (RB). RB growth is thought to be similar to that of extracellular bacteria as cells replicate by binary fission within the inclusion. This process will occur until, at a currently unknown stage, the RBs undergo secondary differentiation into EBs. The EB progeny are then released from the host cell and are available for subsequent infection of neighbouring cells.

Under stress conditions, triggered by a range of factors including the presence of antibiotics, the effects of cytokines such as gamma interferon and the depletion of glucose and essential amino acids, chlamydial RBs may enter an alternative developmental stage which involves the formation of large, pleomorphic cellular forms (aberrant bodies; ABs). ABs will persist inside host cells, coining the term 'persistence', until the developmental trigger is removed or the nutrients are replaced. Importantly in terms of treatment, in vitro studies have indicated that persistent chlamydial infections are refractile to standard antibiotic treatment. While the detection and discrimination between multiple repeat infections and a single chronic infection remains difficult, persistent infections have nevertheless been associated with a range of chronic infections in humans and animals including polyarthritis, infertility and cardiovascular disease.


Aided by a small genome size (ie. approx. 1,000,000 base pairs, 1000 genes), complete nucleotide sequences are available for nearly all members of the Chlamydiaceae, including multiple strains of human C. trachomatis and both human and animal strains of C. pneumoniae. This work has revolutionised our understanding of chlamydial growth and development and has assisted in identifying determinants of Chlamydia's pathogenesis and tissue and host trophisms.

Observations from the initial genome sequences revealed that Chlamydia has more metabolic capabilities than expected. Chief amongst these was the discovery that Chlamydia has a nearly complete peptidoglycan synthesis pathway. This discovery confirmed previous work showing that chlamydial development is highly sensitive to beta-lactam antibiotic treatment (although this is not the preferred antibiotic prescription), despite repeated failures to detect peptidoglycan precursors in chlamydial cells. Other analyses have also revealed that Chlamydia, unexpectedly, can also produce its own ATP which contradicts previous thoughts that Chlamydia was entirely dependent on the host cell's energy reserves.

A major finding in recent years has been the realisation that the ability to evade the host interferon gamma (IFN-γ) immune pathway defines the pathogenicity and tissue trophism of some chlamydial species3. A major consequence of IFN-γ activation is the deletion of host cell tryptophan pools by activation of a tryptophan decyclising enzyme, indoleamine 2,3-digoxygenase (IDO). When this occurs during a chlamydial infection, it has the effect of either completing blocking chlamydial growth or induces the development of persistence. Genital strains of human C. trachomatis are predicted to avoid this pathway by synthesising tryptophan from exogenous indole, produced by lactobacilli and other organisms of the vaginal flora. Ocular strains lack this biosynthetic capability. C. muridarum strains also lack tryptophan synthase but the IFN-γ-mediated pathway is thought to be absent in the mouse genital tract epithelium.

Genome sequencing has also revealed that Chlamydia has a functional Type III secretion system to deliver effector proteins to the host cell cytoplasm4. These effector proteins are thought to be involved in a variety of roles including activation of host cell pathways to form the inclusion and avoid phagolysosomal fusion and inhibition of IDO activation amongst others.


A wide range of test methods have been used to diagnose chlamydial infections in humans and animals.

(a) Traditional methods

Traditional methods for diagnosing Chlamydia involve microscopic investigation of stained smears and examination for specific pathology. These methods are used irregularly now for diagnosis because it is almost impossible to classify the chlamydial species involved and require specific expertise. Isolation of the chlamydial agent in embryonated hen eggs or in eukaryotic cells is a highly successful method for detection and speciation of chlamydial infections but is laborious and costly and has largely been replaced by specific polymerase chain reaction (PCR) assays.

(b) Antigen and antibody detection methods

Serology, particularly in the form of the complementation test (CFT), is still a convenient and commonly used technique for detecting present and past chlamydial infections in humans, cats, birds, cattle and small ruminants. Although the sensitivity is significantly less than 100%, the ease of collection of blood samples and the availability of technology to perform these tests makes these ideal for screening large numbers of samples (particularly suited to screen livestock).

Antigen detection, usually in the form of either plate-based or solid-phase ELISAs (eg. ClearviewTMtest), also has several advantages over traditional methods, including the ability to detect chlamydial antigen in fresh or fixed tissues and/or swab samples collected from various anatomical sites. Antigens primarily detected in this method include the group-specific lipopolysaccharide which is conserved within the genus (making it ideal for veterinary studies) and the chlamydial major outer membrane protein (MOMP), which is a dominant immunogen but can be highly variable between species.

(c) PCR

In recent times, PCR has replaced traditional methods as the preferred method for detecting chlamydial infections in humans and animals. This method is reliable, rapid and highly sensitive and can be used on non-viable specimens. Additional advantages include opportunities for subsequent speciation and fine detailed molecular typing of isolates.

PCR assays targeting a variety of chlamydial genes are available and report a range of specificity and sensitivities. The most common amplification targets are genes encoding chlamydial 16S rRNA, 23S rRNA and ompA (encoding MOMP).

PCRs that target the 16S rRNA-23S rRNA region are generally able to amplify all members of the order Chlamydiales but make subsequent speciation difficult due to conservation between species1. ompA has also been widely used but is highly variable; this makes selection of conserved PCR primers difficult but the variability can be utilised for fine-detailed epidemiological analysis of chlamydial outbreaks in animal populations.

New PCR based technologies are also emerging involving quantitative broad-range PCR to amplify DNA from selected chlamydial species, followed either by high resolution temperature-based analysis of the PCR product5 or hybridisation to specific probes on a DNA chip for speciation6. These methods will further establish PCR-based methods as the gold-standard for detection of chlamydial infections in animals.


  1. Everett KD, Bush RM and Andersen, AA (1999) Emended description of the order Chlamydiales, proposal of Parachlamydiaceae fam. nov. and Simkaniaceae fam. nov., each containing one monotypic genus, revised taxonomy of the family Chlamydiaceae, including a new genus and five new species, and standards for the identification of organisms. Int. J. Syst. Bacteriol. 49: 415-440
  2. Stephens, RS, Myers G, Eppinger, and Bavoil, PM (2009) Divergence without difference: phylogenetics and taxonomy of Chlamydia resolved. FEMS Immunol. Med. Microbiol. 55: 115-119
  3. McClarty, G, Caldwell, HD, and Nelson DE (2007) Chlamydial interferon gamma immune evasion influences infection tropism. Curr. Opin. Microbiol. 10: 47-51
  4. Peters, J, Wilson, DP, Myers, G, Timms, P, and Bavoil, PM. (2007) Type III secretion - la Chlamydia. 15: 241-251
  5. Robertson, T, Bibby, S, O'Rourke, D, Belfiore, T, Lambie, H, and Noomohammadi, AH. (2009). Characterisation of Chlamydiaceae species using PCR and high resolution melt curve analysis of the 16S rRNA gene. J. Appl. Microbiol. 170: 2017-2028
  6. Borel, N, Kempf, E, Hotzel, H, Schubert, E, Torgerson, P, Slickers, P, Ehricht, R, Tasara, T, Pospischil, A, and Sachse, K. (2008). Direct identification of chlamydiae from clinical samples using a DNA microarray assay: a validation study. Mol. Cell Probes. 22: 55-64


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