Defining immune correlates during latent and active chlamydial infection in sheep.

Ovine enzootic abortion (OEA) caused by the obligate intracellular bacterial pathogen Chlamydia abortus (C. abortus), is an endemic disease in most sheep-rearing countries worldwide. Following infection, C. abortus establishes a complex host-pathogen interaction with a latent phase in non-pregnant sheep followed by an active disease phase in the placenta during pregnancy leading to OEA. Improved knowledge of the host-pathogen interactions at these different phases of disease will accelerate the development of new diagnostic tests and vaccines to control OEA. Current evidence indicates that cellular immunity is essential for controlling C. abortus infection. We have previously described a model of mucosal (intranasal) infection of non-pregnant sheep with C. abortus that replicates the latent and active phases of OEA. We have investigated antigen-specific recall responses of peripheral blood mononuclear cells (PBMC) in sheep infected with C. abortus via the intranasal route to determine how these change during the latent and active phases of disease. By analysing cytokines associated with the major CD4+ve Thelper (Th) cell subsets (Interferon-gamma (IFN-γ)/Th1; Interleukin (IL)-4/Th2; IL-17A/Th17; IL-10/Tregulatory), we show that there is selective activation of PBMC producing IFN-γ and/or IL-10 during the latent phase following infection. These cytokines are also elevated during the active disease phase and while they are produced by sheep that are protected from OEA, they are also produced by sheep that abort, highlighting the difficulties in finding specific cellular immunological correlates of protection for complex intracellular pathogens.

FERLAVIRUS-RELATED DEATHS IN A COLLECTION OF VIPERID SNAKES.

Between June and October 2013, 26 snakes of six viperid species kept in two adjoining rooms died ( n = 16) or were euthanized on medical (1) or welfare grounds (9). Two were from the main zoo collection, but the other 24 had been imported and quarantined for a minimum of 6 mo. Four of those that died and the single snake euthanized on medical grounds showed minor signs of respiratory disease prior to death, and five were weak, lethargic, and/or poor feeders. Frequent postmortem findings among all snakes were poor body condition (18) and respiratory disease (13). Seventeen cases were examined histologically, and pneumonia, sometimes with air sacculitis and/or tracheitis, was present in 15 individuals. Lung samples from 24 snakes were ferlavirus polymerase chain reaction (PCR) positive, and one of the two snakes for which only liver was available was also positive. The negative liver sample was from a snake that died of sepsis following anesthesia for surgical removal of a spindle cell sarcoma. Correlation with antemortem PCR testing of glottal and cloacal swabs in five cases was poor (sensitivity = 40%). Immunohistochemistry (IHC) for ferlaviruses on the tissues of 13 PCR-positive cases showed positive labeling in 7 only. Tissues samples from 22 ferlavirus PCR-positive snakes were examined for Chlamydia species by PCR, and 9 were positive, although DNA sequencing only confirmed two of three tested as Chlamydia pneumoniae. Immunohistochemistry for Chlamydia pneumoniae of seven cases (two Chlamydiales PCR positive, one of which was sequenced as C. pneumoniae, plus five negative) confirmed the Chlamydia PCR results. These two Chlamydiales PCR and IHC positive snakes were ferlavirus PCR positive, but IHC negative suggesting that, even though a ferlavirus was the predominant cause of the outbreak, in a few cases death may have been due to chlamydiosis with ferlavirus present, but not acting as the primary pathogen.

Processing of Chlamydia abortus polymorphic membrane protein 18D during the chlamydial developmental cycle.

BACKGROUND: Chlamydia possess a unique family of autotransporter proteins known as the Polymorphic membrane proteins (Pmps). While the total number of pmp genes varies between Chlamydia species, all encode a single pmpD gene. In both Chlamydia trachomatis (C. trachomatis) and C. pneumoniae, the PmpD protein is proteolytically cleaved on the cell surface. The current study was carried out to determine the cleavage patterns of the PmpD protein in the animal pathogen C. abortus (termed Pmp18D). METHODOLOGY/PRINCIPAL FINDINGS: Using antibodies directed against different regions of Pmp18D, proteomic techniques revealed that the mature protein was cleaved on the cell surface, resulting in a100 kDa N-terminal product and a 60 kDa carboxy-terminal protein. The N-terminal protein was further processed into 84, 76 and 73 kDa products. Clustering analysis resolved PmpD proteins into three distinct clades with C. abortus Pmp18D, being most similar to those originating from C. psittaci, C. felis and C. caviae. CONCLUSIONS/SIGNIFICANCE: This study indicates that C. abortus Pmp18D is proteolytically processed at the cell surface similar to the proteins of C. trachomatis and C. pneumoniae. However, patterns of cleavage are species-specific, with low sequence conservation of PmpD across the genus. The absence of conserved domains indicates that the function of the PmpD molecule in chlamydia remains to be elucidated.