New Zealand Veterinary Journal
ISSN: 0048-0169 (Print) 1176-0710 (Online) Journal homepage: http://www.tandfonline.com/loi/tnzv20
Avian reoviruses in New Zealand A.F. Green M.Sc. , J.K. Clarke B.Sc. Ph.D. & J.E. Lohr Dr. Med. Vet. To cite this article: A.F. Green M.Sc. , J.K. Clarke B.Sc. Ph.D. & J.E. Lohr Dr. Med. Vet. (1976) Avian reoviruses in New Zealand, New Zealand Veterinary Journal, 24:8, 181-183, DOI: 10.1080/00480169.1976.34312 To link to this article: http://dx.doi.org/10.1080/00480169.1976.34312
Published online: 23 Feb 2011.
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Date: 07 November 2015, At: 08:41
1976
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AVIAN REOVIRUSES IN NEW ZEALAND A. F.
GREEN,
J. K.
CLARKE AND
INTRODUCTION
and 1)ropagatiO'n O'f viruses O'f domestic hens in New Zealand has been carried out mainly by using embryO'nated eggs. Although this technique of viral pro pagation is the methO'd O'f choice for a range of agents, there are advantages in using cell-culture techniques for sO'me viruses that infect domestic he.ns. The present study was undertaken to find if reoviruses (respiratory enteric orPhan) could be recO'vered from domestic hens, using chick-kidney-cell cultures. DETECTION
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MATERIALS AND METHODS
Fertile eggs were incubated and hatch ed in isolation. Soon after hatching, the kidneys were removed, trypsinized and! sedimented by low speed centrifugation. Monolayers were grown in rolled tubes using Hank's lactalbumin medium con taining 10% calf serum. When monolay ers were formed the growth medium was replaced by Earle's lactalbumin medium containing 2% foetal calf serum. Speci mens were taken from the alimentary and resDiratory tracts of diseased and healthy birds, and processed for virus iso lation as described by McFerran et al. ( 1972). TO' identify agents causing a cytopathic effect, Iysates, prepared by freezing and thawing cell cultures in a few drops of distille.d water, were applied to carbon coated grids, negatively stained with 3% sodium phosphotungstate at pH 6.5, and examined in an electron microscope. Un inoculated cultures in no case develope.d a CPE nor were any virus particles detect ed in such cultures by negative contrast ele.ctron microscopy. *A F. Green. M.Sc .• J. K. Clarke. B.Sc .• Ph.D .• De partment of Microbiology and Genetics. Massey University. Palmerston North. J. E. Lohr. Dr Med. Vet .• Department of Veteri nary Pathology and Public Health. Massey Univer sity. Palmerston North.
J. E.
LoHR*
Chloroform sensitivity was tested by the methO'd O'f Feldman and Wang (1961). The effect O'f cations on thermal stability was tested as described by Wallis et al. (1962), and the effect O'f 1O-4M 5-iodo 2-deoxyuridine (IOU) was tested as de scribed by McFerran et al. (1972). Anti se.rum fO'r neutralization tests was pre pared in rabbits by multiple intramuscu lar inoculation of viral preparation, puri fied -b three passages at limiting dilution in cel culture.
I
RESULTS
Ouring the course of this study five agents subsequently identifie.d as reovirus es were recovered from 120 specimens. All five agents caused a cytopathic effect (CPE) in cell cultures characterized by the formation of syncytia (Fig. 1). Follow iing primary isolation of the agents all .gave a readily recogI}izable CPE 2 to 3 daysafte.r passage of cell-culture super natant and inoculated cell cultures be came totally degenerate about 2 days after a CPE was first observed. The con centration of infectious virus in the super natant usually had a titre of about 104 TCIOso per ml. The five isolates were ob ,served by negative contrast electron microscopy, and in e,ach case viral cap sids were readily detected (Fig. 2). The average capsid diameter measured 75 nm, -and in pene,trated particles, an inner shell was visible, and -had an average diameter of 46 nm. The five isolates were morpho logically indistinguishable from one an Qther and frO'm type 3 mammalian reo virus. Antiserum to one of the five agents neutralized the other four isolates, sO' physico-chemical tests were confined to the original isolate. The infectivity of this agent was not diminished by chloroform; its replication was not affected by IOU at a concentration that strongly inhibited a strain of avian adenovirus (McFe,rran et al., 1972). Following heating at serc in cell-culture medium for one hour, the viral titre diminished 5- to 6-fold. A
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FIG. 1: Chick kidney cell culture inoculated with an
FIG. 2: Avian reovirulS stained with sodium phos· photungstate. All particles are penetrated by stain and show an inner shell surrounded by an outer layer 0/ capsomeres. Some broken particles are present.
similar small loss o.f infectivity was ob served in the presence of 2.0 M Na +, 1.0M Ca++ and 1.0 M Mg++.
mon in O'ther countries (see Menendez et al., 1975), and reovirus is alsO' a com mO'n contaminant of avian cell cultures (Mustaffa-Babjee; and Spradbmw, 1971, 1973). The causative agent of viral arth ritis (O'r tenosynovitis) has been classifi ed as a reovirus by Olson and Weiss (1972) and Glass et al. (1973). A reovirus was also isolated from ruptured gastro cnemius tendons of chickens in England (Jones et al., 1975). The Fahey-Crawley virus, isolated from chickens with acute or chmnic respiratory disease (Fahey'and Crawley, 1954) is a reovirus (Peteik et al., 1967) serolo¢callv related to the causa tive agent of viral arthritis (Olson and Weiss, 1972). An Australian reovirus was reported to' be invol~fld in an