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Chlamydiae have a worldwide distribution causing a wide range of diseases in human hosts, livestock, and companion animals as well as in wildlife and exotic species. Moreover, they can persist in their hosts as asymptomatic infections for extended periods of time. The introduction of molecular techniques has revolutionized the Chlamydia field by expanding the host range of known chlamydial species but also by discovering new species and even new families of bacteria in the broader order Chlamydiales. The wide range of hosts, diseases, and tissues affected by chlamydiae complicate the diagnosis such that standard diagnostic approaches for these bacteria are rare. Bacteria of the Chlamydiales order are small and their inclusions are difficult to detect by standard microscopy. With the exception of avian and ovine chlamydiosis, macroscopic and/or histologic changes might not be pathognomic or indicative for a chlamydial infection or even not present at all. Moreover, detection of chlamydial DNA in specimens in the absence of other methods or related pathological lesions questions the significance of such findings. The pathogenic potential of the majority of recently identified Chlamydia-related bacteria remains largely unknown and awaits investigation through experimental or natural infection models including histomorphological characterization of associated lesions. This review aims to summarize the historical background and the most important developments in the field of animal chlamydial research in the past 5 years with a special focus on pathology. It will summarize the current nomenclature, present critical thoughts about diagnostics, and give an update on chlamydial infections in domesticated animals such as livestock, companion animals and birds, as well as free-ranging and captive wild animals such as reptiles, fish, and marsupials.

An intracellular bacterium was isolated from fruit bats (Artibeus intermedius) in Cocoyoc, Mexico. The bacterium caused severe lesions in the lungs and spleens of bats and intracytoplasmic vacuoles in cell cultures. Sequence analyses showed it is related to Waddlia spp. (order Chlamydiales). We propose to call this bacterium Waddlia cocoyoc.

CPAF is a conserved and secreted protease from obligate intracellular bacteria of the order Chlamydiales. Recently, it was demonstrated that most of its host targets are an artifact of inaccurate methods. This review aims to summarize key features of CPAF and propose new approaches for evaluating its role in chlamydial pathogenesis.

An epidemiological survey was performed to detect the presence of Chlamydophila (C.) abortus and other members of the order Chlamydiales in ovine and caprine flocks with a history of abortion in southern Italy. Four pairs of primers were compared to evaluate their ability to detect Chlamydiales using purified DNA preparations and tissue samples from aborted foetuses with suspected chlamydial infections. As expected, amplification of DNA of the reference strain C. abortus using primer pairs U23F/23Sigr, 16SF2/23R, CTU/CTL and CpsiA/CpsiB produced fragments of about 600 bp, 585 bp, 1000 bp and 300 bp, respectively. The detection limits of the four PCR tests performed on serial DNA dilutions of the C. abortus reference strain were of 10 pg, 0.1 pg, 0.1 pg and 1 fg of DNA, respectively. The most sensitive amplification of DNA extracted from the organ tissues was obtained with primer pairs CpsiA/CpsiB, which detected Chlamydophila spp. DNA in all infected tissue samples. Only C. abortus was identified during the survey. The presence of this agent was confirmed in 3 out of 27 ovine and caprine flocks included in the survey suggesting that abortion due to C. abortus is uncommon in southern Italy.

Chlamydial infections of fish are emerging as an important cause of disease in new and established aquaculture industries. To date, epitheliocystis, a skin and gill disease associated with infection by these obligate intracellular pathogens, has been described in over 90 fish species, including hosts from marine and fresh water environments. Aided by advances in molecular detection and typing, recent years have seen an explosion in the description of these epitheliocystis-related chlamydial pathogens of fish, significantly broadening our knowledge of the genetic diversity of the order Chlamydiales. Remarkably, in most cases, it seems that each new piscine host studied has revealed the presence of a phylogenetically unique and novel chlamydial pathogen, providing researchers with a fascinating opportunity to understand the origin, evolution and adaptation of their traditional terrestrial chlamydial relatives. Despite the advances in this area, much still needs to be learnt about the epidemiology of chlamydial infections in fish if these pathogens are to be controlled in farmed environments. The lack of in vitro methods for culturing of chlamydial pathogens of fish is a major hindrance to this field. This review provides an update on our current knowledge of the taxonomy and diversity of chlamydial pathogens of fish, discusses the impact of these infections on the health, and highlights further areas of research required to understand the biology and epidemiology of this important emerging group of fish pathogens of aquaculture species.

Waddlia chondrophila is a strict intracellular microorganism belonging to the order Chlamydiales that has been isolated twice from aborted bovine fetuses, once in USA and once in Germany. This bacterium is now considered as an abortigenic agent in cattle. However, no information is available regarding the presence of this bacterium in Africa. Given the low sensitivity of cell culture to recover such an obligate intracellular bacterium, molecular-based diagnostic approaches are warranted. This report describes the development of a quantitative SYBR Green real-time PCR assay targeting the recA gene of W. chondrophila. Analytical sensitivity was 10 copies of control plasmid DNA per reaction. No cross-amplification was observed when testing pathogens that can cause abortion in cattle. The PCR exhibited a good intra-run and inter-run reproducibility. This real-time PCR was then applied to 150 vaginal swabs taken from Tunisian cows that have aborted. Twelve samples revealed to be Waddlia positive, suggesting a possible role of this bacterium in this setting. This new real-time PCR assay represents a diagnostic tool that may be used to further study the prevalence of Waddlia infection.

Simkania negevensis is an obligate intracellular bacterium grouped into the order Chlamydiales. This new amoeba-resistant bacterium represents a novel aetiologic agent of bronchiolitis and community-acquired pneumonia in both adults and children. It has been suggested that Simkania could be an ubiquitous microorganism presented in water environments. In the natural history of infections with amoeba-related bacteria encountered in aquatic habitats, the transmissions by environmental aerosols or contaminated water/air systems have been extensively recognized. Therefore, understanding the feasibility of Simkania infection by these or similar routes is relevant. In the present work, we investigated the prevalence of this novel disease-associated microorganism in water samples from different sources by real-time PCR (qPCR). Our results show Simkania detection in 5 of 185 water analyzed samples (2.7%: 2 of 88 cooling towers and 3 of 8 waste water samples). However, no Simkania was detected in a drinking water.

Growing evidence suggests that a novel member of the Chlamydiales order, Waddlia chondrophila, is a potential agent of miscarriage in humans and abortion in ruminants. Due to the lack of genetic tools to manipulate chlamydia, genomic analysis is proving to be the most incisive tool in stimulating investigations into the biology of these obligate intracellular bacteria. 454/Roche and Solexa/Illumina technologies were thus used to sequence and assemble de novo the full genome of the first representative of the Waddliaceae family, W. chondrophila. The bacteria possesses a 2'116'312 bp chromosome and a 15'593 bp low-copy number plasmid that might integrate into the bacterial chromosome. The Waddlia genome displays numerous repeated sequences indicating different genome dynamics from classical chlamydia which almost completely lack repetitive elements. Moreover, W. chondrophila exhibits many virulence factors also present in classical chlamydia, including a functional type III secretion system, but also a large complement of specific factors for resistance to host or environmental stresses. Large families of outer membrane proteins were identified indicating that these highly immunogenic proteins are not Chlamydiaceae specific and might have been present in their last common ancestor. Enhanced metabolic capability for the synthesis of nucleotides, amino acids, lipids and other co-factors suggests that the common ancestor of the modern Chlamydiales may have been less dependent on their eukaryotic host. The fine-detailed analysis of biosynthetic pathways brings us closer to possibly developing a synthetic medium to grow W. chondrophila, a critical step in the development of genetic tools. As a whole, the availability of the W. chondrophila genome opens new possibilities in Chlamydiales research, providing new insights into the evolution of members of the order Chlamydiales and the biology of the Waddliaceae.

Chlamydia comprises a group of obligate intracellular bacterial parasites responsible for a variety of diseases in humans and animals, including several zoonoses. Chlamydia trachomatis causes diseases such as trachoma, urogenital infection and lymphogranuloma venereum with severe morbidity. Chlamydia pneumoniae is a common cause of community-acquired respiratory tract infections. Chlamydia psittaci, causing zoonotic pneumonia in humans, is usually hosted by birds, while Chlamydia abortus, causing abortion and fetal death in mammals, including humans, is mainly hosted by goats and sheep. We used multi-locus sequence typing to asses the population structure of Chlamydia. In total, 132 Chlamydia isolates were analyzed, including 60 C. trachomatis, 18 C. pneumoniae, 16 C. abortus, 34 C. psittaci and one of each of C. pecorum, C. caviae, C. muridarum and C. felis. Cluster analyses utilizing the Neighbour-Joining algorithm with the maximum composite likelihood model of concatenated sequences of 7 housekeeping fragments showed that C. psittaci 84/2334 isolated from a parrot grouped together with the C. abortus isolates from goats and sheep. Cluster analyses of the individual alleles showed that in all instances C. psittaci 84/2334 formed one group with C. abortus. Moving 84/2334 from the C. psittaci group to the C. abortus group resulted in a significant increase in the number of fixed differences and elimination of the number of shared mutations between C. psittaci and C. abortus. C. psittaci M56 from a muskrat branched separately from the main group of C. psittaci isolates. C. psittaci genotypes appeared to be associated with host species. The phylogenetic tree of C. psittaci did not follow that of its host bird species, suggesting host species jumps. In conclusion, we report for the first time an association between C. psittaci genotypes with host species.

Epitheliocystis is an infectious disease in freshwater and marine fish species throughout the world. It affects the gills and skin epithelium and is characterised by the presence of hypertrophied cells, which contain granular inclusions (Nowak and LaPatra 2006). Presently, epitheliocystis has been reported in more than 50 species of freshwater, marine and wild fish species worldwide (Nowak and LaPatra 2006, Karlsen and others 2008, Mitchell and others 2010). Epitheliocystis is caused by a genetically diverse group of intracellular Gram-negative bacteria belonging to the order Chlamydiales (Meijer and others 2006, Nowak and LaPatra 2006, Karlsen and others 2008). The objective of this study was to investigate the epitheliocystis agent in grass carp. In September 2011, grass carp (n=450, average length 10 cm) were sampled from a fish farm in Austria for routine diagnostic examination, and maintained at 25°C in our laboratory. After three weeks, the collected fish demonstrated clinical signs, such as: lethargy, respiratory distress and development of transparent cysts on the gill lamellae. After euthanasia by MS222, tissues specimens (gills, kidney, liver and spleen) were sampled from 20 fish for diagnosis. For bacteriological examination, the samples were investigated using different bacteriological media, and incubated for up to five days at different temperatures under aerobic and anaerobic conditions. For histopathological examination, tissues were fixed in 10 per cent neutral buffered formalin, sectioned, stained with haematoxylin and eosin (H&E) and examined by light microscope. Genomic DNA was extracted from infected …

2 chlamydia-like microorganisms are associated with fatal diseases of the spiderPisaura mirabilis Cl. The general characteristics of these organisms are indicated and place them inside the genusPorochlamydia (order Chlamydiales).

It is proposed that the family Chlamydiaceae, which contains only one recognized genus, Chlamydia, be placed in a new order, Chlamydiales, coordinate with Rickettsiales Buchanan and Buchanan 1938. The principal argument for this change is that chlamydiae multiply by a developmental cycle that is unique among all other forms of bacteria.