A Bovine Model of Respiratory Chlamydia psittaci Infection: Challenge Dose Titration

This study aimed to establish and evaluate a bovine respiratory model of experimentally induced acute C. psittaci infection. Calves are natural hosts and pathogenesis may resemble the situation in humans. Intrabronchial inoculation of C. psittaci strain DC15 was performed in calves aged 2–3 months via bronchoscope at four different challenge doses from 106 to 109 inclusion-forming units (ifu) per animal. Control groups received either UV-inactivated C. psittaci or cell culture medium. While 106 ifu/calf resulted in a mild respiratory infection only, the doses of 107 and 108 induced fever, tachypnea, dry cough, and tachycardia that became apparent 2–3 days post inoculation (dpi) and lasted for about one week. In calves exposed to 109 ifu C. psittaci, the respiratory disease was accompanied by severe systemic illness (apathy, tremor, markedly reduced appetite). At the time point of most pronounced clinical signs (3 dpi) the extent of lung lesions was below 10% of pulmonary tissue in calves inoculated with 106 and 107 ifu, about 15% in calves inoculated with 108 and more than 30% in calves inoculated with 109 ifu C. psittaci. Beside clinical signs and pathologic lesions, the bacterial load of lung tissue and markers of pulmonary inflammation (i.e., cell counts, concentration of proteins and eicosanoids in broncho-alveolar lavage fluid) were positively associated with ifu of viable C. psittaci. While any effect of endotoxin has been ruled out, all effects could be attributed to infection by the replicating bacteria. In conclusion, the calf represents a suitable model of respiratory chlamydial infection. Dose titration revealed that both clinically latent and clinically manifest infection can be reproduced experimentally by either 106 or 108 ifu/calf of C. psittaci DC15 while doses above 108 ifu C. psittaci cannot be recommended for further studies for ethical reasons. This defined model of different clinical expressions of chlamydial infection allows studying host-pathogen interactions

Infection, Disease, and Transmission Dynamics in Calves after Experimental and Natural Challenge with a Bovine <i>Chlamydia psittaci</i> Isolate

<div><p><i>Chlamydia</i> (<i>C.</i>) <i>psittaci</i> is the causative agent of psittacosis, a zoonotic disease in birds and man. In addition, <i>C. psittaci</i> has been repeatedly found in domestic animals and is, at least in calves, also able to induce respiratory disease. Knowledge about transmission routes in cattle herds is still deficient, and nothing is known about differences in host response after either experimental or natural exposure to <i>C. psittaci</i>. Therefore, our recently developed respiratory infection model was exploited to evaluate (i) the presence of the pathogen in blood, excretions and air, (ii) the possibility of transmission and (iii) clinical symptoms, acute phase and immune response until 5 weeks after exposure. In this prospective study, intrabronchial inoculation of 10⁸ inclusion-forming units of <i>C. psittaci</i> (n = 21 calves) led to reproducible acute respiratory illness (of approximately one week), accompanied by a systemic inflammatory reaction with an innate immune response dominated by neutrophils. Excretion and/or exhalation of the pathogen was sufficient to transmit the infection to naïve sentinel calves (n = 3) co-housed with the infected animals. Sentinel calves developed mild to subclinical infections only. Notably, excretion of the pathogen, predominantly via feces, occurred more frequently in animals naturally exposed to <i>C. psittaci</i> (i.e. sentinels) as compared to experimentally-inoculated calves. The humoral immune response was generally weak, and did not emerge regularly following experimental infection; however, it was largely absent after naturally acquired infection.</p></div

Rectal temperature and respiratory rate measured 48–72 hours post inoculation (i.e. peak of clinical signs).

<p>In calves experimentally inoculated with different doses of viable <i>C. psittaci,</i> both rectal temperature (panel A) and respiratory rate (panel B) were significantly increased while no significant changes were seen in control calves. Data are given as Box-and Whisker Plots based on 2 or 3 measurements per calf in controls or infected animals, respectively. Different letters indicate significant differences between groups at given <i>P</i>-level (multiple range test).</p

Markers of pulmonary inflammation assessed in broncho-alveolar lavage fluid (BALF).

<p>Both concentration of total protein (A) and 12-HETE (B) were maximal in calves inoculated with 10⁹ ifu of <i>C. psittaci</i>. Data for control groups are from 2 and 3 dpi combined (Box and Whisker Plots). Data for calves challenged with different doses of viable <i>C. psittaci</i> are given on an individual basis for time points when calves were sacrificed (filled circles: 3 dpi; open circles: 7 dpi and 14 dpi). Kruskal-Wallis test revealed significant differences between groups at given <i>P</i>-level.</p

Assessment of regression lines of the clinical scores given in Figure 1.

<p>Assessment of regression lines of the clinical scores given in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0030125#pone-0030125-g001" target="_blank">Figure 1</a>.</p

Distribution and extent of pulmonary lesions at day 3 pi in a calf inoculated with 10⁹ ifu of <i>C. psittaci</i>.

<p>Dorsal view of the lung and heart (H). Pneumonic lesions present as dark red discolorations (>) in the apical lobes, middle lobe and basal lobes. Note distension of the basal lobes due to severe bronchopneumonia in the inferior segments (hatched lines). Bar = 5 cm.</p