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Lung Tuberculosis in a Horse Caused by Mycobacterium Avium Subsp. Avium of Serotype 2: a Case Report

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Lung Tuberculosis in a Horse Caused by Mycobacterium Avium Subsp. Avium of Serotype 2: a Case Report Veterinarni Medicina, 53, 2008 (2): 111–116 Case Report Lung tuberculosis in a horse caused by Mycobacterium avium subsp. avium of serotype 2: a case report I. Pavlik1, P. Jahn2, M. Moravkova1, L. Matlova1, F. Treml2, A. Cizek2, E. Nesnalova2, L. Dvorska-Bartosova1, R. Halouzka2 1Veterinary Research Institute, Brno, Czech Republic 2University of Veterinary and Pharmaceutical Sciences, Brno, Czech Republic ABSTRACT: Interstitial pneumonia (2/3 of the lungs were affected) and diffusely enlarged bronchial and medi- astinal lymph nodes were diagnosed by gross examination of a dead 16-year-old mare. Based on histopathological examination and the detection of acid-fast rods after staining by the Ziehl-Neelsen technique, tuberculosis was suspected. Mycobacterium avium subsp. avium of serotype 2 and IS901+/IS1245+ genotype was isolated from the pulmonary lymphnode after five-week incubation at 37°C. Due to the fact that horses have a naturally high resistance to mycobacterial infections, the high age of the mare most likely contributed to the development of the disease. Keywords: avian mycobacteriosis; avian tuberculosis; Mycobacterium avium complex; zoonosis Mycobacterial infections are very rare in hors- belongs (according to the Commission Decision es. In the first part of 20th century mycobacte- No. 2004/320/EC of March 31, 2004) to those rial infections in horses were mainly caused by European countries officially free of bovine tuber- Mycobacterium bovis. The situation has changed culosis. The causative agent of human tuberculosis, since the second part of 20th century, and the M. tuberculosis, has been found only occasionally most common cause of mycobacterial infections in domestic and wild animals in Central European in horses are now members of the M. avium com- countries during these years (Pavlik, 2006); nev- plex (MAC). Generally, mycobacterial infections in ertheless, potential infection with this pathogen horses arise via ingestion, though primary respira- cannot be precluded (Pavlik et al., 2004). tory infection does occur. The prevailing clinical On the other hand, the description of MAC infec- signs are chronic weight loss, diarrhoea, pyrexia, tion in different animal species is more common. chronic pneumonia, septic arthritis and blindness M. avium subsp. hominissuis (M. a. hominissuis) of in both eyes; abortions have also been documented, IS901–/IS1245+ genotype and serotypes 4–6, 8–11 as analysed in a published review by Pavlik et al. and 21 causes infection mainly in pigs (Matlova et (2004). al., 2004; Pate et al., 2004; Pavlik et al., 2005). It is In the Czech Republic and in other Central commonly isolated from the environment and from European countries, bovine tuberculosis is one of immunosupressed patients (Dvorska et al., 2002; the eradicated infectious diseases of domestic ani- Matlova et al., 2003; Reed et al., 2006). M. avium mals (Pavlik, 2006), wild animals (Trcka et al., 2006) subsp. avium (M. a. avium) of IS901+/IS1245+ and humans (Pavlik, 2006). The Czech Republic genotype and serotypes 1–3 is the agent of avian Supported by the Ministry of Agriculture of the Czech Republic (Grants No. NPV 1B53009 and No. MZE 0002716201) and by EC, Brussels (Grant VENoMYC, No. SSPE-CT-2004-501903). 111 Case Report Veterinarni Medicina, 53, 2008 (2): 111–116 tuberculosis of birds; but it is also occasionally Post mortem findings isolated from other animals such as pigs or cattle (Pavlik et al., 2000; Pate et al., 2004; Moravkova et Necropsy and histopathological examination. al., 2007; Shitaye et al., 2007). At necropsy, a serofibrinous bloodstained effusion The diagnosis of lung tuberculosis in a horse in the thoracic cavity and in the collapsed lung were used for teaching at the University of Veterinary revealed. Coarse confluent nodular lesions of larda- and Pharmaceutical Sciences in Brno was serious; ceous appearance affecting 2/3 of the caudodorsal therefore, the purpose of the present study was to part of diaphragmatic lobes of lung grossly resem- describe the clinical signs, gross findings and the bled a sarcoma. Pulmonary pleura above the area specific identification of the mycobacterial isolate of lung parenchyma consolidation showed diffused in a 16-year-old mare. granular thickenings. The pattern of pale homoge- neous tissue caused enlargement and effacement of bronchial lymph nodes with a loss of corticomed- Case history ullary distinction. No other marked gross lesions were seen except heart hypertrophy and visceral Clinical examination. Inappetence and a fe- organ congestion. brile state not exceeding 39.5°C was detected in a During the necropsy, samples of the lungs, bron- 16-year-old warm-blooded mare from the property chial lymph nodes, liver, spleen and kidneys were of the Faculty of Veterinary Medicine, Brno (Czech collected for bacteriological and histopathological Republic). examinations. Samples were fixed in 10% buffered Clinical examination of the mare did not show formalin and routinely processed by the paraffin any pathology except for an increased breath technique. The thickness of histological sections sound over the entire lung fields. Endoscopic ex- was 5 µm. After deparaffinisation, these were amination of the airways revealed secretion in the stained with haematoxylin and eosin. Samples from trachea. No lesions were found by examination lung tissue and lung lymph nodes were stained by of the upper respiratory tract. Haematological the Ziehl-Neelsen (Z-N) technique for the acid-fast examination results were within reference range. rods (AFR) detection (Vacek, 1972). Alveolar macrophages and a small amount of neu- Histopathological changes of lesioned lung tis- trophils were detected by cytological examination sue were characterised by the obliteration of both of the tracheal secretion, collected by transtra- alveolar and interalveolar spaces by macrophages, cheal aspiration. Bacteria were not detected in epithelioid cells, lymphocytes and a few giant poly- the sample, either by cytological examination or nuclear cells. In the central zone of the granuloma- bacteriological culture. tous inflammation, the proliferative fibrous tissues Therapy and further examinations. The mare with lymphocytes and macrophages, infiltration was administered Procaine penicillin G (Biotika, was observed. Caseation was occasional and of a Slovenska Lupca, Slovak Republic) in the dose of small extension. No calcification was observed. 10 000 IU/kg b.w. for three days. Due to the fact The Z-N staining method showed only sporadic that the clinical state did not improve during that single AFR. time and the auscultatory finding in the chest per- Basic bacteriological examination. The tissue sisted, a latero-lateral radiogram of the chest was samples from lung lymph nodes and lung tissue performed, which showed a nodular interstitial were bacteriologically examined for the presence and alveolar pattern (Figure 1). During the follow- of pathogenic aerobic and anaerobic bacteria. ing four days, gradual deterioration of the clini- Aseptically collected samples were cultured on cal state occurred. The inappetence and febrile Columbia agar (Oxoid, CM331) enriched with 5% state persisted despite the fact that the mare was sheep blood in sodium citrate and MacConkey agar administered Sulfadoxin-Trimethoprim (Borgal, No. 3 (Oxoid, CM115). Seeded agars were incubat- Hoechst, Germany) in the dose of 15 mg/kg b.w. ed at 37°C. The basic bacteriological examination twice a day. Bioptic sample of lung tissue was col- of pulmonary tissue and pulmonary lymphnodes lected. Due to the injury of a large blood vessel, was negative. pulmonary bleeding occurred, which could not Mycobacteriological examination. Two sam- be stopped. The mare died about 15 min after the ples (tuberculous lesioned lung tissue and lung biopsy. lymph node) were also examined by mycobacte- 112 Veterinarni Medicina, 53, 2008 (2): 111–116 Case Report rial culture examination. About 1 g of the sample amined isolate was of serotype 2. Virulence of one was homogenised, treated with 1 mol/l HCl and primo isolate was studied after intramuscular ad- 2 mol/l NaOH and 40 µl was pipetted on solid egg ministration to a chicken by a method previously media according to Stonebrink, two solid Herrold described (Pavlik et al., 2000). The primo isolate was egg yolk media without Mycobactin J and antibiot- virulent (causing miliary tuberculosis of the liver and ics. Decontaminated samples were also inoculated spleen) in the biological experiment on a chicken. onto two liquid serum media according to Sula. The growth of mycobacteria was monitored at 37°C after the first week and then every two weeks for DISCUSSION a period of three months (Matlova et al., 2003). One mycobacterial isolate originating only from the Clinical signs of pulmonary tuberculosis in the lung tissue with tuberculous lesions was detected mare were non-specific. The only data in history by the Z-N staining technique after six weeks of giving evidence of disease were the febrile state and incubation. inappetence. Clinical examination revealed only an PCR identification. The AFR positive isolate was increased breath sounds over the lung fields and in- examined by the PCR method for the detection of creased amount mucus in the trachea. The transtra- 1 030 bp fragments of the 16S rRNA gene specific cheal aspiration sample contained a small amount for the genus Mycobacterium (Wilton and Cousins, of alveolar macrophages and minute amounts of 1992). For identification of the M. tuberculosis neutrophils, which
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