Gastrointestinal Coinfection Promotes Chlamydial Pathogenicity in the Genital Tract.

Sexually transmitted Chlamydia, which can cause fibrotic pathology in women's genital tracts, is also frequently detected in the gastrointestinal tract. However, the medical significance of the gastrointestinal Chlamydia remains unclear. A murine Chlamydia readily spreads from the mouse genital tract to the gastrointestinal tract while inducing oviduct fibrotic blockage or hydrosalpinx. We previously proposed a two-hit model in which the mouse gastrointestinal Chlamydia might induce the second hit to promote genital tract pathology, and we are now providing experimental evidence for testing the hypothesis. First, chlamydial mutants that are attenuated in inducing hydrosalpinx in the genital tract also reduce their colonization in the gastrointestinal tract, leading to a better correlation of chlamydial induction of hydrosalpinx with chlamydial colonization in the gastrointestinal tract than in the genital tract. Second, intragastric coinoculation with a wild-type Chlamydia rescued an attenuated Chlamydia mutant to induce hydrosalpinx, while the chlamydial mutant infection in the genital tract alone was unable to induce any significant hydrosalpinx. Finally, the coinoculated gastrointestinal Chlamydia failed to directly spread to the genital tract lumen, suggesting that gastrointestinal Chlamydia may promote genital pathology via an indirect mechanism. Thus, we have demonstrated a significant role of gastrointestinal Chlamydia in promoting pathology in the genital tract possibly via an indirect mechanism. This study provides a novel direction/dimension for further investigating chlamydial pathogenic mechanisms.

Epidermal Growth Factor Receptor and Transforming Growth Factor ß Signaling Pathways Cooperate To Mediate Chlamydia Pathogenesis.

Human genital Chlamydia infection is a major public health concern due to the serious reproductive system complications. Chlamydia binds several receptor tyrosine kinases (RTKs) on host cells, including the epidermal growth factor receptor (EGFR), and activates cellular signaling cascades for host invasion, cytoskeletal remodeling, optimal inclusion development, and induction of pathogenic epithelial-mesenchyme transition (EMT). Chlamydia also upregulates transforming growth factor beta (TGF-ß) expression, whose signaling pathway synergizes with the EGFR cascade, but its role in infectivity, inclusions, and EMT induction is unknown. We hypothesized that the EGFR and TGF-ß signaling pathways cooperate during chlamydial infection for optimal inclusion development and stable EMT induction. The results revealed that Chlamydia upregulated TGF-ß expression as early as 6 h postinfection of epithelial cells and stimulated both the EGFR and TGF-ß signaling pathways. Inhibition of either the EGFR or TGF-ßR1 signaling substantially reduced inclusion development; however, the combined inhibition of both EGFR and TGF-ßR1 signaling reduced inclusions by over 90% and prevented EMT induction. Importantly, EGFR inhibition suppressed TGF-ß expression, and an inhibitory thrombospondin-1 (Tsp1)-based peptide inhibited chlamydia-induced EMT, revealing a major source of active TGF-ß during infection. Finally, TGF-ßR signaling inhibition suppressed the expression of transforming acidic coiled-coil protein-3 (TACC3), which stabilizes EGFR signaling, suggesting reciprocal regulation between TGF-ß and EGFR signaling during chlamydial infection. Thus, RTK-mediated host invasion by chlamydia upregulated TGF-ß expression and signaling, which cooperated with other cellular signaling cascades and cytoskeletal remodeling to support optimal inclusion development and EMT induction. This finding may provide new targets for chlamydial disease biomarkers and prevention.

Chlamydia: what is on the outside does matter.

This review summarises major highlights on the structural biology of the chlamydial envelope. Chlamydiae are obligate intracellular bacteria, characterised by a unique biphasic developmental cycle. Depending on the stage of their lifecycle, they appear in the form of elementary or reticulate bodies. Since these particles have distinctive functions, it is not surprising that their envelope differs in lipid as well as in protein content. Vice versa, by identifying surface proteins, specific characteristics of the particles such as rigidity or immunogenicity may be deduced. Detailed information on the bacterial membranes will increase our understanding on the host-pathogen interactions chlamydiae employ to survive and grow and might lead to new strategies to battle chlamydial infections.

Effect of female sex hormones on the developmental cycle of Chlamydia abortus compared to a penicillin-induced model of persistent infection.

BACKGROUND: Chlamydia abortus, an obligate intracellular pathogen with an affinity for placenta, causes reproductive failure. In non-pregnant animals, an initial latent infection is established until the next gestation, when the microorganism is reactivated, causing abortion. The precise mechanisms that trigger the awakening of C. abortus are still unknown. Sexual hormones such as estradiol and progesterone have been shown to affect the outcome of infection in other species of the family Chlamydiaceae, while estrogens increase chlamydial infection, progesterone has the opposite effect. To try to establish whether there is a relationship between these events and the latency/ reactivation of C. abortus in the reproductive tract of small ruminants, ovine endometrial (LE) and trophoblastic (AH-1) cells were treated with estradiol or progesterone prior to their infection with C. abortus. The results are compared with those obtained for treatment with penicillin prior to infection, which is a well-established model for studying persistent infection in other chlamydial species. Cells were examined by transmission electron microscopy, and an mRNA expression analysis of 16 genes related to the chlamydial developmental cycle was made. RESULTS: The changes observed in this study by the action of sex hormones seem to depend on the type of cell where the infection develops. In addition, while the changes are morphologically similar to those induced by treatment with penicillin, the patterns of gene expression are different. Gene expression patterns therefore, seem to depend on the persistence induced models of C. abortus used. Hormone treatments induced aberrant forms in infected endometrial cells but did not affect the chlamydial morphology in trophoblast cells. At the genetic level, hormones did not induce significant changes in the expression of the studied genes. CONCLUSIONS: The results suggest that penicillin induces a state of persistence in in vitro cultured C. abortus with characteristic morphological features and gene transcriptional patterns. However, the influence of hormones on the C. abortus developmental cycle is mediated by changes in the host cell environment. Furthermore, a persistent state in C. abortus cannot be characterised by a single profile of gene expression pattern, but may change depending on the model used to induce persistence.

Differential signaling pathways are initiated in macrophages during infection depending on the intracellular fate of Chlamydia spp.

Chlamydia muridarum and Chlamydia caviae have equivalent growth rates in mouse epithelial cells but only C. muridarum replicates inside mouse macrophages, while C. caviae does not. Macrophages infected with C. muridarum or C. caviae were used to address the hypothesis that the early signaling pathways initiated during infection depend on the fate of chlamydiae in the host cell. Transmission electron microscopy of C. muridarum-infected macrophages showed intact chlamydial elementary bodies and reticulate bodies 2 h postinfection in compact vacuoles. Conversely, in macrophages infected with C. caviae, chlamydiae were observed in large phagocytic vacuoles. Furthermore, C. caviae infections failed to develop into inclusions or produce viable bacteria. Expression of proinflammatory cytokines TNFα, IL-1ß and MMP13 was similar in C. caviae- or C. muridarum-infected macrophages at 3 h postinfection, indicating that chlamydial survival is not required for initiation of these responses. IL-1ß secretion, dependent on inflammasome activation, occurred in C. caviae-infected macrophages despite no chlamydial growth. Conversely, IFNß mRNA was observed only in C. muridarum- but not in C. caviae-infected macrophages. These data demonstrate that differential signaling events are initiated during a productive versus nonproductive chlamydial infection in a macrophage.

Chlamydia spp. development is differentially altered by treatment with the LpxC inhibitor LPC-011.

BACKGROUND: Chlamydia species are obligate intracellular bacteria that infect a broad range of mammalian hosts. Members of related genera are pathogens of a variety of vertebrate and invertebrate species. Despite the diversity of Chlamydia, all species contain an outer membrane lipooligosaccharide (LOS) that is comprised of a genus-conserved, and genus-defining, trisaccharide 3-deoxy-D-manno-oct-2-ulosonic acid Kdo region. Recent studies with lipopolysaccharide inhibitors demonstrate that LOS is important for the C. trachomatis developmental cycle during RB- > EB differentiation. Here, we explore the effects of one of these inhibitors, LPC-011, on the developmental cycle of five chlamydial species. RESULTS: Sensitivity to the drug varied in some of the species and was conserved between others. We observed that inhibition of LOS biosynthesis in some chlamydial species induced formation of aberrant reticulate bodies, while in other species, no change was observed to the reticulate body. However, loss of LOS production prevented completion of the chlamydial reproductive cycle in all species tested. In previous studies we found that C. trachomatis and C. caviae infection enhances MHC class I antigen presentation of a model self-peptide. We find that treatment with LPC-011 prevents enhanced host-peptide presentation induced by infection with all chlamydial-species tested. CONCLUSIONS: The data demonstrate that LOS synthesis is necessary for production of infectious progeny and inhibition of LOS synthesis induces aberrancy in certain chlamydial species, which has important implications for the use of LOS synthesis inhibitors as potential antibiotics.

Culture-independent genomic characterisation of Candidatus Chlamydia sanzinia, a novel uncultivated bacterium infecting snakes.

BACKGROUND: Recent molecular studies have revealed considerably more diversity in the phylum Chlamydiae than was previously thought. Evidence is growing that many of these novel chlamydiae may be important pathogens in humans and animals. A significant barrier to characterising these novel chlamydiae is the requirement for culturing. We recently identified a range of novel uncultured chlamydiae in captive snakes in Switzerland, however, nothing is known about their biology. Using a metagenomics approach, the aim of this study was to characterise the genome of a novel chlamydial taxon from the choana of a captive snake. In doing so, we propose a new candidate species in the genus Chlamydia (Candidatus Chlamydia sanzinia) and reveal new information about the biological diversity of this important group of pathogens. RESULTS: We identified two chlamydial genomic contigs: a 1,113,073 bp contig, and a 7,504 bp contig, representing the chromosome and plasmid of Ca. Chlamydia sanzinia strain 2742-308, respectively. The 998 predicted coding regions include an expanded repertoire of outer membrane proteins (Pmps and Omps), some of which exhibited frameshift mutations, as well as several chlamydial virulence factors such as the translocating actin-recruitment phosphoprotein (Tarp) and macrophage inhibition potentiator (Mip). A suite of putative inclusion membrane proteins were also predicted. Notably, no evidence of a traditional chlamydial plasticity zone was identified. Phylogenetically, Ca. Chlamydia sanzinia forms a clade with C. pneumoniae and C. pecorum, distinct from former "Chlamydophila" species. CONCLUSIONS: Genomic characterisation of a novel uncultured chlamydiae from the first reptilian host has expanded our understanding of the diversity and biology of a genus that was thought to be the most well-characterised in this unique phylum. It is anticipated that this method will be suitable for characterisation of other novel chlamydiae.

[A FITZ-HUGH-CURTIS SYNDROME AS A PREMISE OF THE INFLAMMATORY DISEASES OF A HEPATOPANCREATOBILIARY ZONE ORGANS].

A content of a biliary ways microflora and its correlation with clinical form and severity of inflammatory-destructive processes in hepatopancreatobiliary zone were studied up. The investigation objective was the optimization of a treatment--diagnostic tactics for the complications and remote recurrences rate reduction in a complicated biliary calculous disease. There was established, that a standard bacteriological investigation do not give a possibility to estimate some causes of a biliary ducts affection, a chlamidial infection in particular. So on, for the individual antibacterial therapy choosing a more detailed and modern examination of patients is needed.

Phosphoproteomic analysis of the Chlamydia caviae elementary body and reticulate body forms.

Chlamydia are Gram-negative, obligate intracellular bacteria responsible for significant diseases in humans and economically important domestic animals. These pathogens undergo a unique biphasic developmental cycle transitioning between the environmentally stable elementary body (EB) and the replicative intracellular reticulate body (RB), a conversion that appears to require extensive regulation of protein synthesis and function. However, Chlamydia possess a limited number of canonical mechanisms of transcriptional regulation. Ser/Thr/Tyr phosphorylation of proteins in bacteria has been increasingly recognized as an important mechanism of post-translational control of protein function. We utilized 2D gel electrophoresis coupled with phosphoprotein staining and MALDI-TOF/TOF analysis to map the phosphoproteome of the EB and RB forms of Chlamydia caviae. Forty-two non-redundant phosphorylated proteins were identified (some proteins were present in multiple locations within the gels). Thirty-four phosphorylated proteins were identified in EBs, including proteins found in central metabolism and protein synthesis, Chlamydia-specific hypothetical proteins and virulence-related proteins. Eleven phosphorylated proteins were identified in RBs, mostly involved in protein synthesis and folding and a single virulence-related protein. Only three phosphoproteins were found in both EB and RB phosphoproteomes. Collectively, 41 of 42 C. caviae phosphoproteins were present across Chlamydia species, consistent with the existence of a conserved chlamydial phosphoproteome. The abundance of stage-specific phosphoproteins suggests that protein phosphorylation may play a role in regulating the function of developmental-stage-specific proteins and/or may function in concert with other factors in directing EB-RB transitions.

Targeting eukaryotic Rab proteins: a smart strategy for chlamydial survival and replication.

Chlamydia, an obligate intracellular bacterium which passes its entire lifecycle within a membrane-bound vacuole called the inclusion, has evolved a variety of unique strategies to establish an advantageous intracellular niche for survival. This review highlights the mechanisms by which Chlamydia subverts vesicular transport in host cells, particularly by hijacking the master controllers of eukaryotic trafficking, the Rab proteins. A subset of Rabs and Rab interacting proteins that control the recycling pathway or the biosynthetic route are selectively recruited to the chlamydial inclusion membrane. By interfering with Rab-controlled transport steps, this intracellular pathogen not only prevents its own degradation in the phagocytic pathway, but also creates a favourable intracellular environment for growth and replication. Chlamydia, a highly adapted and successful intracellular pathogen, has several redundant strategies to re-direct vesicles emerging from biosynthetic compartments that carry host molecules essential for bacterial development. Although current knowledge is limited, the latest findings have shed light on the role of Rab proteins in the course of chlamydial infections and could open novel opportunities for anti-chlamydial therapy.

The broad-spectrum antiviral compound ST-669 restricts chlamydial inclusion development and bacterial growth and localizes to host cell lipid droplets within treated cells.

Novel broad-spectrum antimicrobials are a critical component of a strategy for combating antibiotic-resistant pathogens. In this study, we explored the activity of the broad-spectrum antiviral compound ST-669 for activity against different intracellular bacteria and began a characterization of its mechanism of antimicrobial action. ST-669 inhibits the growth of three different species of chlamydia and the intracellular bacterium Coxiella burnetii in Vero and HeLa cells but not in McCoy (murine) cells. The antichlamydial and anti-C. burnetii activity spectrum was consistent with those observed for tested viruses, suggesting a common mechanism of action. Cycloheximide treatment in the presence of ST-669 abrogated the inhibitory effect, demonstrating that eukaryotic protein synthesis is required for tested activity. Immunofluorescence microscopy demonstrated that different chlamydiae grow atypically in the presence of ST-669, in a manner that suggests the compound affects inclusion formation and organization. Microscopic analysis of cells treated with a fluorescent derivative of ST-669 demonstrated that the compound localized to host cell lipid droplets but not to other organelles or the host cytosol. These results demonstrate that ST-669 affects intracellular growth in a host-cell-dependent manner and interrupts proper development of chlamydial inclusions, possibly through a lipid droplet-dependent process.

The chlamydial transcriptional regulator Euo is a key switch in cell form developmental progression but is not involved in the committed step to the formation of the infectious form.

Bacteria in the genus Chlamydia are a significant health burden worldwide. They infect a wide range of vertebrate animals, including humans and domesticated animals. In humans, C. psittaci can cause zoonotic pneumonia, while C. pneumoniae causes a variety of respiratory infections. Infections with C. trachomatis cause ocular or genital infections. All chlamydial species are obligate intracellular bacteria that replicate exclusively inside of eukaryotic host cells. Chlamydial infections are dependent on a complex infection cycle that depends on transitions between specific cell forms. This cycle consists of cell forms specialized for host cell invasion, the elementary body (EB), and a form specialized for intracellular replication, the reticulate body (RB). In addition to the EB and RB, there is a transitionary cell form that mediates the transformation between the RB and the EB, the intermediate body (IB). In this study, we ectopically expressed the regulatory protein Euo and showed that high levels of expression resulted in reversible arrest of the development cycle. The arrested chlamydial cells were trapped phenotypically at an early IB stage of the cycle. These cells had exited the cell cycle but had not shifted gene expression from RB like to IB/EB like. This arrested state was dependent on continued expression of Euo. When ectopic expression was reversed, Euo levels dropped in the arrested cells which led to the repression of native Euo expression and the resumption of the developmental cycle. Our data are consistent with a model where Euo expression levels impact IB maturation to the infectious EB but not the production of the IB form. IMPORTANCE: Bacterial species in the Chlamydiales order infect a variety of vertebrate animals and are a global health concern. They cause various diseases in humans, including genital and respiratory infections. The bacteria are obligate intracellular parasites that rely on a complex infectious cycle involving multiple cell forms. All species share the same life cycle, transitioning through different states to form the infectious elementary body (EB) to spread infections to new hosts. The Euo gene, encoding a DNA-binding protein, is involved in regulating this cycle. This study showed that ectopic expression of Euo halted the cycle at an early stage. This arrest depended on continued Euo expression. When Euo expression was reversed, the developmental cycle resumed. Additionally, this study suggests that high levels of Euo expression affect the formation of the infectious EB but not the production of the cell form committed to EB formation.

Lipid acquisition by intracellular Chlamydiae.

Chlamydia species are obligate intracellular pathogens that are important causes of human genital tract, ocular and respiratory infections. The bacteria replicate within a specialized membrane-bound compartment termed the inclusion and require host-derived lipids for intracellular growth and development. Emerging evidence indicates that Chlamydia has evolved clever strategies to fulfil its lipid needs by interacting with multiple host cell compartments and redirecting trafficking pathways to its intracellular niche. In this review, we highlight recent findings that have significantly expanded our understanding of how Chlamydia exploit lipid trafficking pathways to ensure the survival of this important human pathogen.

Chlamydia bacteriophages.

Phages are called "good viruses" due to their ability to infect and kill pathogenic bacteria. Chlamydia are small, Gram-negative (G-) microbes that can be dangerous to human and animals. In humans, these bacteria are etiological agents of diseases such as psittacosis or respiratory tract diseases, while in animals, the infection may result in enteritis in cattle and chronic bowel diseases, as well as miscarriages in sheep. The first-known representative of chlamydiaphages was Chp1. It was discovered in Chlamydia psittaci isolates. Since then, four more species of chlamydiaphages have been identified [Chp2, Chp3, φCPG1 φCPAR39 (φCpn1) and Chp4]. All of them were shown to infect Chlamydia species. This paper described all known chlamydiaphages. They were characterised in terms of origin, host range, and their molecular structure. The review concerns the characterisation of bacteriophages that infects pathogenic and dangerous bacteria with unusual, intracellular life cycles that are pathogenic. In the era of antibiotic resistance, it is difficult to cure chlamydophilosis. Those bacteriophages can be an alternative to antibiotics, but before this happens, we need to get to know chlamydiaphages better.

Disulfide bonding within components of the Chlamydia type III secretion apparatus correlates with development.

Chlamydia spp. exhibit a unique biphasic developmental cycle whereby infectious elementary bodies (EBs) invade host epithelial cells and differentiate into noninfectious, metabolically active reticulate bodies (RBs). EBs posses a unique outer envelope where rigidity is achieved by disulfide bonding among cysteine-rich envelope-associated proteins. Conversely, these disulfide bonds become reduced in RBs to accommodate vegetative growth, thereby linking the redox status of cysteine-rich envelope proteins with progression of the developmental cycle. We investigated the potential role of disulfide bonding within the chlamydial type III secretion system (T3SS), since activity of this system is also closely linked to development. We focused on structural components of the T3S apparatus that contain an unusually high number of cysteine residues compared to orthologs in other secretion systems. Nonreducing SDS-PAGE revealed that EB-localized apparatus proteins such as CdsF, CdsD, and CdsC form higher-order complexes mediated by disulfide bonding. The most dramatic alterations were detected for the needle protein CdsF. Significantly, disulfide bonding patterns shifted during differentiation of developmental forms and were completely reduced in RBs. Furthermore, at later time points during infection following RB to EB conversion, we found that CdsF is reoxidized into higher-order complexes. Overall, we conclude that the redox status of specific T3SS apparatus proteins is intimately linked to the developmental cycle and constitutes a newly appreciated aspect of functionally significant alterations within proteins of the chlamydial envelope.

Raman microspectroscopy reveals long-term extracellular activity of Chlamydiae.

The phylum Chlamydiae consists exclusively of obligate intracellular bacteria. Some of them are formidable pathogens of humans, while others occur as symbionts of amoebae. These genetically intractable bacteria possess a developmental cycle consisting of replicative reticulate bodies and infectious elementary bodies, which are believed to be physiologically inactive. Confocal Raman microspectroscopy was applied to differentiate between reticulate bodies and elementary bodies of Protochlamydia amoebophila and to demonstrate in situ the labelling of this amoeba symbiont after addition of isotope-labelled phenylalanine. Unexpectedly, uptake of this amino acid was also observed for both developmental stages for up to 3 weeks, if incubated extracellularly with labelled phenylalanine, and P. amoebophila remained infective during this period. Furthermore, P. amoebophila energizes its membrane and performs protein synthesis outside of its host. Importantly, amino acid uptake and protein synthesis after extended extracellular incubation could also be demonstrated for the human pathogen Chlamydia trachomatis, which synthesizes stress-related proteins under these conditions as shown by 2-D gel electrophoresis and MALDI-TOF/TOF mass spectrometry. These findings change our perception of chlamydial biology and reveal that host-free analyses possess a previously not recognized potential for direct experimental access to these elusive microorganisms.

[Relationship of TLR2 and TLR4 gene polymorphism with a penchant for individual urogenital infections].

We investigated the population prevalence of polymorphisms of Arg753Gln TLR2gene, and Asp299Gly, Thr399Ile TLR4 genes among people living in the Poltava region, as well as data communication polymorphisms with the presence of diseases caused by urogenital infections. The group of population control was a random sample of residents of the Poltava region (n = 299). The group of patients with urogenital diseases included 156 people. Genotyping of these groups TLR2 Arg753Gln polymorphism and the gene TLR4 Asp299Gly, Thr399Ile was performed using PCR and subsequent restriction analysis. Statistically significant association of allele A ofgene TLR2 (p = 0.0018) and allele G of gene TLR4 (p = 0.085) with the presence of urogenital diseases was confirmed.

Inclusion membrane proteins of Protochlamydia amoebophila UWE25 reveal a conserved mechanism for host cell interaction among the Chlamydiae.

Chlamydiae are a group of obligate intracellular bacteria comprising several important human pathogens. Inside the eukaryotic cell, chlamydiae remain within a host-derived vesicular compartment, termed the inclusion. They modify the inclusion membrane through insertion of unique proteins, which are involved in interaction with and manipulation of the host cell. Among chlamydiae, inclusion membrane proteins have been exclusively found in members of the family Chlamydiaceae, which predominantly infect mammalian and avian hosts. Here, the presence of inclusion membrane proteins in Protochlamydia amoebophila UWE25, a chlamydial endosymbiont of free-living amoebae, is reported. A genome-wide screening for secondary structure motifs resulted in the identification of 23 putative inclusion membrane proteins for this organism. Immunofluorescence analysis demonstrated that five of these proteins were expressed, and four of them could be localized to a halo surrounding the intracellular bacteria. Colocalization studies showed an almost complete overlap of the signals obtained for the four putative inclusion membrane proteins, and immuno-transmission electron microscopy unambiguously demonstrated their location in the inclusion membrane. The presence of inclusion membrane proteins (designated IncA, IncQ, IncR, and IncS) in P. amoebophila shows that this strategy for host cell interaction is conserved among the chlamydiae and is used by chlamydial symbionts and pathogens alike.

Impact of azithromycin resistance mutations on the virulence and fitness of Chlamydia caviae in guinea pigs.

Azithromycin (AZM) is a major drug used in the treatment and prophylaxis of infections caused by Chlamydia, yet no significant clinical resistance has been reported for these obligate intracellular bacteria. Nevertheless, spontaneous AZM resistance (Azm(r)) arose in vitro at frequencies ranging from 3 x 10(-8) to 8 x 10(-10) for clonal isolates of Chlamydia caviae, which is a natural pathogen of guinea pigs. Sequencing of the unique 23S rRNA gene copy in 44 independent Azm(r) isolates identified single mutations at position A(2058) or A(2059) (Escherichia coli numbering system). While SP(6)AZ(1) (A(2058)C) and SP(6)AZ(2) (A(2059)C) Azm(r) mutants showed growth defects in cell culture and were less pathogenic in the guinea pig ocular infection model than in the parent SP(6), the three isogenic C. caviae isolates grew equally well in the animal. On the other hand, coinoculation of the C. caviae parent strain with one of the Azm(r) strains was detrimental for the mutant strain. This apparent lack of association between pathology and bacterial load in vivo showed that virulence of the two Azm(r) mutants of C. caviae was attenuated. While chlamydial growth in vitro reflects the ability of the bacteria to multiply in permissive cells, survival in the host is a balance between cellular multiplication and clearance by the host immune system. The obligate intracellular nature of Chlamydia may therefore limit emergence of resistance in vivo due to the strength of the immune response induced by the wild-type antibiotic-sensitive bacteria at the time of antibiotic treatment.

Mixed infections with Chlamydia and porcine epidemic diarrhea virus - a new in vitro model of chlamydial persistence.

BACKGROUND: Chlamydiae induce persistent infections, which have been associated with a wide range of chronic diseases in humans and animals. Mixed infections with Chlamydia and porcine epidemic diarrhea virus (PEDV) may result in generation of persistent chlamydial infections. To test this hypothesis, an in vitro model of dual infection with cell culture-adapted PEDV and Chlamydia abortus or Chlamydia pecorum in Vero cells was established. RESULTS: Infected cultures were investigated by immunofluorescence (IF), transmission electron microscopy (TEM) and re-infection experiments. By IF, Chlamydia-infected cells showed normal inclusions after 39 hpi. Dual infections with Chlamydia abortus revealed a heterogenous mix of inclusion types including small inclusions consisting of aberrant bodies (ABs), medium-sized inclusions consisting of ABs and reticulate bodies and normal inclusions. Only aberrant inclusions were observable in dual infection experiments with Chlamydia pecorum and PEDV. TEM examinations of mixed infections with Chlamydia abortus and Chlamydia pecorum revealed aberrant chlamydial inclusions containing reticulate-like, pleomorphic ABs, which were up to 2 microm in diameter. No re-differentiation into elementary bodies (EBs) was detected. In re-infection experiments, co-infected cells produced fewer EBs than monoinfected cells. CONCLUSIONS: In the present study we confirm that PEDV co-infection alters the developmental cycle of member species of the family Chlamydiaceae, in a similar manner to other well-described persistence induction methods. Interestingly, this effect appears to be partially species-specific as Chlamydia pecorum appears more sensitive to PEDV co-infection than Chlamydia abortus, as evidenced by TEM and IF observations of a homogenous population of aberrant inclusions in PEDV - Chlamydia pecorum co-infections.