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nodes, spleen tissue and blood samples from slaughteredinfected animals should be tested by bacteriological examina-tion and PCR in order to isolate and/or detect Brucella cells.

The comparison between the ELISA and RBT as screeningmethods showed that the ELISA demonstrated a higher num-ber of false positive cases, while the latter detected a highernumber of false negatives in infected herds from theCampania region. An acceptable method of screening shouldminimise false negative results, especially when used for thedetection of infections with serious consequences both foranimals and humans, such as brucellosis. The number of falsepositives was considered to be more acceptable in this studybecause of subsequent confirmation by CFT analysis.

The ELISA used in this study proved to be more sensitivethan the RBT, and it may have potential as a screening test forthe detection of brucellosis outbreaks in buffaloes, providedthat the CFT is applied to confirm the positive results obtained.Finally, since conventional serological tests have not yet beenstandardised for water buffaloes, this indirect ELISA is the onlystandardised test known so far to diagnose brucellosis in thisanimal species.

ACKNOWLEDGEMENTS

The authors are grateful to Raffaele Riccio for his translationof this article.

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ReferencesALTON, G. G., JONES, L. M., ANGUS, R. D. & VERGER, J. M. (1988)Techniques for the Brucellosis Laboratory. Paris, Institut National de laRecherche Agronomique

BIANCIFIORI, F., GIOVANNIN, A., NANNINI, D. & DI MATTEO, A. (1997)Standardizzazione di una tecnica ELISA per la ricerca degli anticorpi brucel-lari nel latte bovino. Veterinaria Italiana 32,42-46

GARIN-BASTUJI, B. (1993) Brucellosis in cattle, pigs, sheep and goats. Presentepidemiological situation in Europe. Proceedings of the 46th Annual Meetingof the European Association for Animal Production. Prague, Czech Republic,September 4 to 7, 1995. p 142

MAYFIELD, J. E., BANTLE, J. A., EWALT, D. R., MEADOR, V. P. & TATABAI,L. B. (1988) Detection of Brucella cells and cell components. In AnimalBrucellosis. Boca Raton, Chemical Rubber

MORGAN,W. T. B. & MCDIARMID, G. (1960) The excretion of Brucella abor-tus in the milk of experimentally infected cattle. Research in Veterinary Science1,53-56

NIELSEN, K., SMITH, P., GALL, D., PEREZ, B., COSMA, C., MUELLER, P.,TROTTIER, J., COTE, G., BOAG, L. & BOSSE, J. (1996) Development andvalidation of an indirect enzyme immunoassay for detection of antibody to

Brucella abortus in milk. Veterinary Microbiology 52, 165-173UZAL, F. A., CARASCO, A. E., NIELSEN, K., ECHAIDE, S. & CABRERA, R. F.

(1996) An indirect ELISA using a monoclonal anti IgG I enzyme conjugate forthe diagnosis of bovine brucellosis. Veterinary Microbiology 52, 175-180

WEYNANTS, V., TIBOR, A., DENOEL, P. A., SAERGERMAN, C., GODFROID,J., THIANGE, P. & LETESSON, J1 P. (1996) Infection of cattle with Yersiniaenterocolitica 09 a cause of the false positive serological reactions in bovinebrucellosis diagnostic tests. Veterinary Microbiology 48, 101-112

WRIGHT, P. P., NILSSON, E., VAN ROOJ, E. M. A., LELENTA, M. & JEGGO,M. H. (1993) Standardisation and validation of ELISA for the detection ofantibody in infectious diseases diagnosis. Revue Scientifique et Technique -Office International des Epizooties 12,435-450

CorrectionDiagnostic investigation of chronic porcine reproductiveand respiratory syndrome virus in a breeding herd of pigsby M. D. Bierk, S. A. Dee, K. D. Rossow, J. E. Collins, M. I.Guedes, C. Pijoan, T. W. Molitor (VR, 148, June 2, 2001, pp687-690). In Table 3, the number of dead fetuses from sowB should have read 0 and not as stated. The error is regretted.

Pathologicalinvestigations onanthracosis in cattleK. OZCAN, E. BEYTUTPNEUMOCONIOSIS is a chronic inflammatory reactionbrought about by the inhalation of various types of mineralparticle (Smith and Jones 1961). The condition is more com-mon in humans than in other animals because humans are

often exposed to this kind of pollutant (Schoning and others1996). However, pneumoconiosis has also been reported inurban dogs (Metin and others 1990). Anthracosis, a type ofpneumoconiosis (Seaton 1983), occurs as a result of theinhalation of carbon particles by humans (Seaton 1983,Oshima 1990, Vakharia and others 1990) and animals(Thomson 1984). Anthracosis has been known in miningcommunities since the 18th century (Oshima 1990), and ithas also been detected in the lungs of Egyptian mummies(Walker and others 1987).

The lungs and related lymph nodes are the primary sitesfor anthracosis (Dahme and others 1982, Thomson 1984,Kumar and others 1992). However, in humans, anthracosishas been detected in various other sites such as the liver andspleen (LeFevre and others 1982), and the oesophagus(Vakharia and others 1990). Macroscopically, affected lungsare greyish in colour, and the related lymph nodes are black(Dahme and others 1982, Thomson 1984). Microscopically,in humans (Grobbelaar and Bateman 1991, Murty and Das1993), dogs (Day and others 1996) and rats (Takaki and oth-ers 1989) deposition of carbon particles has been recordedin the lungs, particularly around the respiratory bronchioles.Furthermore, fibrosis in the interalveolar septa of humans(Oshima 1990, Grobbelaar and Bateman 1991) and rats(Kukharenko and others 1988, Takaki and others 1989) withanthracosis has also been documented. Carbon particles weremainly observed in the cytoplasm of sinusoidal macrophagesand free in the parenchyma of the lymph nodes (Dahme andothers 1982, Thomson 1984, Argani and others 1989, Rothand others 1998).

To the authors' knowledge there have been no previousreports of anthracosis in cattle. The aim of this short com-munication is to describe the macroscopic and microscopicfeatures of anthracosis in cattle in the Turkish city of Kars andthe surrounding area.

Samples were obtained from 700 cattle of various breedsand ages, slaughtered in an abattoir during 1998. All theslaughtered animals were from the same region of Turkey andhad been brought from rural areas. However, as the cattlewere bought from the general animal market it was impossi-ble to determine their precise place of origin. The lungs andmediastinal and bronchial lymph nodes of the slaughteredcattle were examined. Subsequently, the lung lobes and asso-

ciated lymph nodes were sectioned and the cut surfaces exam-ined. Lung and lymph node samples with pigmentation were

fixed in 10 per cent formalin and processed routinely. The sec-

tions were cut to 5 [.m thick and stained with haematoxylinand eosin. Some sections were stained with Van Gieson,Gomory's iron reaction and Von Kossa (Luna 1970) stains.For the identification of pigmentation, concentrated sul-phuric acid, hydrogen peroxide, potassium permanganate andMayer's chlorine bleaching methods (Culling and others1985) were applied to the lung and lymph node samples, andexamined under a light microscope.

Pigmentation was found in the lungs, mediastinal andbronchial lymph nodes of 27 (3.85 per cent) of the 700 cattle.Black pigmented areas were observed on the pleural surface

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FIG 1: Lung,disseminated blackpigmentation areas

of all the lung lobes in a disseminated pattern (Fig 1). Similarpigment deposits were seen on the cut surfaces, particularlyin the areas surrounding the bifurcation of the bronchioles.The lungs and lymph nodes with pigment deposits wererespectively greyish and black in colour. On the cut surfacesof the lymph nodes, in most cases, the pigmentation was veryintense and covered the entire surface (Fig 2), in other casesthe pigmentation was present to a lesser degree. In the lungs,black carbon particles were generally seen in the peribron-chiolar and perivascular interstitium. Carbon particles wereparticularly observed in the cytoplasm of alveolarmacrophages and free in the interalveolar septa as well asextending into the subpleural cavities. Van Gieson stainingrevealed a low level fibrotic response. Accumulation of car-bon particles predominated in the lymph nodes rather thanthe lungs. In the lymph nodes, carbon particles were seenespecially in the cytoplasm of sinusoidal macrophages (Fig 3)and free in the parenchyma of the medullary region. Thenucleus of macrophages with phagocytised carbon particleswas not discernible. To differentiate carbon particles fromother pigments, particularly melanin, bleaching methods,including concentrated sulphuric acid, hydrogen peroxide,potassium permanganate and Mayer's chlorine, were appliedto both the lymph nodes and lungs. The black carbon parti-cles were resistant to all the bleaching methods mentionedabove. Further histochemical Von Kossa staining for calcifi-cation and Gomory's iron reaction for iron were negative,confirming anthracotic pigment.

In modern times humans and animals have been exposedto increasing environmental pollution caused by variouschemical and mineral contaminants. Air pollution is of par-ticular importance as it may cause various health problems(Kumar and others 1992). In general, it is difficult to deter-mine the constituents of atmospheric pollutants and specialtechniques are required (Kukharenko and others 1988, Takakiand others 1989). In the present study, the use of bleaching

FIG 3: Macrophageswith phagocytisedcarbon particles in themediastinal lymphnodes. Haematoxylinand eosin. x 200

FIG 2: Mediastinal lymph node, heavy pigmentation on thecut surface

methods, including Mayer's chlorine technique (Culling andothers 1985) and others, revealed bleach-resistant pigmentparticles, suggesting carbon accumulation in the tissues.Furthermore, the results were supported by macroscopic andmicroscopic findings yielded by the various staining methodsused.

There are no previous reports of anthracosis in cattle.However, anthracosis has been reported in humans (Seaton1983, Vakharia and others 1990, Grobbelaar and Bateman1991, Murty and Das 1993, Roth and others 1998), dogs (Dayand others 1996) and in birds, reptiles and various othermammals kept in captivity (Choudary and others 1986). Inaddition, anthracosis has been induced experimentally in rats(Kukharenko and others 1988, Takaki and others 1989).Macroscopically, parenchymal deposits of particles in thelungs and lymph nodes appear as grey and black, respectively(Dahme and others 1982, Thomson 1984). In the presentstudy, the lungs appeared grey because of intense particledeposition in the pleural and cut surfaces, particularly in theareas surrounding the bifurcation of the bronchioles.Pigmentation was recorded in all the lymph nodes, and inmost cases the lymph nodes were heavily infiltrated by par-ticles and appeared black in colour.

Inhaled particles are normally entrapped and subse-quently transported out of the upper airways by ciliary action.Some particles may reach the conducting airways and bephagocytosed by alveolar macrophages and removed via thelymphatic channels. In the present study, the peribronchiolarand perivascular deposition of carbon particles can beexplained by the impairment of alveolar clearance which mayoccur in heavily polluted environments. Particles may then bedeposited in the alveoli and around bronchioles (Thomson1984). Perivascular and peribronchiolar accumulation of car-bon particles has been previously reported (Dahme and oth-ers 1982, Takaki and others 1989, Grobbelaar and Bateman1991, Murty and Das 1993, Day and others 1996). Subpleuralpigment deposition was also observed in the present studyand similar findings have been reported in rats affected bypoor ventilation of subpleural cavities which were experi-mentally exposed to exhaust fumes (Takaki and others 1989).

Fibrosis of the lungs has also been reported in humanswith anthracosis (Oshima 1990, Grobbelaar and Bateman1991). In a study on rats, it was reported that polycarboxylicacid by-products of the oxidative-hydrolytic breakdown ofcoal damaged the microcirculatory bed of the lungs and thestructures of the aerohaematic barrier in the organism(Kukharenko and others 1988). In another study on rats withexperimentally induced anthracosis, fibrosis, adenoma, ade-nocarcinoma and squamous cell carcinoma were observed(Takaki and others 1989). Takaki and others (1989) attributedthese pulmonary changes to carbon particle deposition in the

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pulmonary tissue. In the present study, slight fibrosis wasobserved in the alveolar septa. This lower level fibrosis may beattributed to the young age of the cattle at slaughter. The pres-ence of carbon particles free in the tissue and especially in thecytoplasm of sinusoidal macrophages of the lymph nodes hasalso been reported by various authors (Dahme and others1982, Thomson 1984, Argani and others 1989).

In conclusion, the present study found anthracosis withmacroscopic and microscopic changes in lung and lymphnode tissues in cattle in Kars city and the surrounding area.In this region heavy industry and coal mining are not presentand may therefore be discounted as possible causes of anthra-cosis. A more likely potential cause is the fact that animalbarns are simple constructions and are generally built adja-cent to houses. Both houses and barns are of simple stone andmud constructions which are quickly eroded. Stoves and clayovens for heating, cooking and making bread are generallybuilt in the part of the house adjacent to the barn. Smoke maytherefore permeate the walls of the houses, entering the barnsand potentially resulting in anthracosis in any animal exposedto it. Further field studies and research are required to inves-tigate this hypothesis.

ReferencesARGANI, P., GHOSSEIN, R. & ROSAI, J. (1989) Anthracotic and anthracosili-

cotic spindle cell pseudotumors of mediastinal lymph nodes: report of fivecases of reactive lesion that simulates malignancy. Human Pathology 29,851-855

CHOUDARY, C., RAO, M. R. K. M. & ALI, S. (1986) Anthracosis in zoo ani-mals and birds. Indian Veterinary Journal 63, 869-870

CULLING, C. F. A., ALLISON, R. T. & BARR, W. T. (1985) Cellular PathologyTechnique. 4th edn. London, Butterworths

DAHME, E., DROMMER, W., KOHLER, H. MESSOW, C., SCHRODER, B. &TRAUTWEON, G. (1982) Lehrbuch Der Allgemeinen Pathologie furTierarzte und Studierende der Tiermedizin. 9th Vollig. Stuttgart,Neubearbeitete Auflage Ferdinand Enke Verlag. p 183

DAY, M. J., PEARSON, G. R., LUCKE, V. M., LANE, S. J. & SPARKS, R. S. J.(1996) Lesions associated with mineral deposition in the lymph node andlung of the dog. Veterinary Pathology 33, 29-42

GROBBELAAR, J. P. & BATEMAN, E. D. (1991) Hut lung: a domesticallyacquired pneumoconiosis of mixed aetiology in rural women. Thorax 46,334-340

KUKHARENKO, T. A., DINKELIS, S. S., YERSHOVA, N. N. & SHKUTIN, A.E. (1988) The role of soluble acids from oxidative-hydrolytic breakdown ofcoal in the development of the fibrous process in anthracosis. Journal ofHygiene Epidemiology, Microbiology and Immunology 32,249-256

KUMAR, V., COTRAN, R. S. & ROBBINS, S. L. (1992) Basic Pathology. 5th edn.Ed V. Kumar. Philadelphia, W. B. Saunders Company. p 220

LEFEVRE, M. E., GREEN, F. H. Y., JOE, D. D. & LAGUEUR, W. (1982)Frequency ofblack pigment in livers and spleens of coal workers: correlationwith pulmonary pathology and occupational information. Human Pathology13,1121-1126

LUNA, C. L. (1970) Manual Histologic Staining Methods of the Armed ForcesInstitute of Pathology. 3rd edn. New York, McGraw Hill Book Company

METIN, N., OZER, H. & KARADAS, E. (1990) Koperklerde pneumoconisis.Firat Uiniversitesi Saglzk Bilimleri Dergisi 4, 111-115

MURTY, D. A. & DAS, D. K. (1993) Pulmonary tuberculosis with anthracosis:an unusual diagnosis by fine needle aspiration cytology. Acta Cytologica 37,639-640

OHSHIMA, S. (1990) Studies on pulmonary anthracosis with special referenceto the mineral constitution of intrapulmonary particulate pollutants in thehuman lung. Acta Pathologica Japonica 40,41-49

ROTH, M. J., GUO-QING, W., LEWIN, K. J., NING, L., DAWSEY, S. M., WES-LEY, M. N., GIFFIN, C., YONG-QIANG, X., MAHER, M. M. & TAYLOR, P.R. (1998) Histopathologic changes seen in esophagectomy specimens fromthe high-risk region of linxian, China: potential clues to etiologic exposure?Human Pathology 29, 1294-1298

SCHONING, P., ABRAHAM, J. L. & BURNETT, B. R. (1996) Silicate and metaldust in lungs of greyhounds. American Journal of Veterinary Research 57,1006-1009

SEATON, A. (1983) Coal and the lung. Thorax 38,241-243SMITH, H. A. & JONES, T. C. (1961) Veterinary Pathology. 2nd edn.Philadelphia, Lea Febiger. p 763

TAKAKI, Y., KITAMURA, S., KUWABARA, N. & FUKUDA, Y. (1989) Long-

term inhalation studies of exhaust from the diesel engine in F-344 rats: thequantitative relationship between pulmonary hyperplasia and anthracosis.Experimental Pathology 37, 56-61

THOMSON, R. G. (1984) General Veterinary Pathology. 2nd edn. Philadelphia,W. B. Saunders Company. p 85

VAKHARIA, B. M., PIETRUK, T. & CALZADA, R. (1990) Anthracosis of esoph-agus. Gastrointestinal Endoscopy 36, 615-617

WALKER, R., PARSCHE, F., BIERBRIER, M. & McKERROW, J. H. (1987)Tissue identification and histologic study of six lung specimens fromEgyptian mummies. American Journal ofPhysical Anthropology 72,43-48

Ruptured dissectingaortic aneurysms intwo red lechwe(Kobus leche)B. A. WOLFE, F. G. DALLDORF, J. F. WRIGHT,M. A. STAMPER, M. R. LOOMIS

ANEURYSMS are uncommon in large animals, but have beendocumented as the cause of sudden death in horses (Brown1985) and sporadically in other exotic hoofstock species(Gupta and Rehbinder 1981, Keep 1973). Trauma, sepsis, par-asite migration, degenerative vascular disease, atherosclerosisand changes as a result of ageing are thought to be involvedin large animal aneurysms (McGuirk and others 1990). Thisshort communication describes ruptured dissecting aorticaneurysms in two red lechwe (Kobus leche).

Two male, half-sibling red lechwe, maintained at the NorthCarolina Zoological Park for 12 years, died suddenly agedbetween 14 and 15 years. The first animal was being immo-bilised for a routine hoof trim and had shown no health prob-lems. Cardiac arrest occurred at the onset of anaesthesia. Thesecond male was found dead without premonitory signs. Bothanimals were housed on pasture containing mainly tall fescue(Festuca arundinacea), and fed a pelleted herbivore diet(Mazuri; Purina Milla).

The necropsy results for both animals were similar. Thefirst animal (animal 1) had a dissecting aortic aneurysm 27x 11 x 9 cm in size, beginning 4 cm distal to the heart. Thefalse lumen did not appear to communicate with the aortaproper and was filled with an organised thrombus. The ven-tral portion of the cranial lobe of the right lung was adheredto the aneurysm (Fig la). The thyroid glands were enlargedand contained multiple cyst-like structures 1 to 3 mm indiameter. Histological examination of this animal's tissuesrevealed fibrosis and mineralisation of the aorta near theaneurysm, arterial sclerosis, atherosclerotic changes and adhe-sion of the aorta to the atelectatic right lung, indicating thatthis aneurysm had been present for an extended period oftime. The atelectasis, fibrosis and adhesion of the lung weremost likely to be secondary to compression by the aneurysm.Colloid cysts of the thyroid were also seen.

Necropsy findings on the second animal (animal 2)included a dissecting aortic aneurysm 5 x 5 x 26 cm in size,beginning 15 cm distal to the heart and extending to the aor-tic bifurcation. The accessory lumen of this aneurysm com-municated with the aorta proper via seven re-entry channels.Histologically, the aorta had multifocal medial fibrosis withosseous metaplasia, and local severe loss of smooth muscleand elastic fibres, with total loss ofboth in the aneurysm (Fig

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B. A. Wolfe, DVM, PhD,F. G. Dalldorf, MD,J. F. Wright, VMD, PhD,M. A. Stamper, DVM,M. R. Loomis, DVM, MS,Division of VeterinaryServices, North CarolinaZoological Park, 4401 ZooParkway, Asheboro, NC27203, USA

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