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Epidemiology of recent outbreaks of infectious laryngotracheitis in poultry in Australia

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HP Blacker,a* NC Kirkpatrick,a A Rubite,b D ORourkea and AH Noormohammadia

Objective Over the past 3 years, numerous outbreaks of infectious laryngotracheitis (ILT) have occurred in poultry in Australia. The objectives of this study were to identify the viral strains involved in the recent outbreaks and to determine possible epidemiological links between these outbreaks. Procedure A combination of polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analyses of several genes of the ILT virus was used to identify genetic dierences in eld/vaccine ILT virus isolates. In a previous study, these procedures had demonstrated ve classes (15) in Australia. Results Analysis of 92 eld ILT viruses demonstrated four new classes: 6, 7, 8 and 9. Class 6 was responsible for four outbreaks in one Victorian broiler company and demonstrated to be distinct from other Australian strains of ILT. Class 7 was the Nobilis ILT vaccine (Intervet Pty Ltd). Class 8 was responsible for the majority of the outbreaks in New South Wales and was phylogenetically close to class 7. On one occasion, classes 7 and 8 were identied in an outbreak on a Victorian farm that had used the Nobilis ILT vaccine. Class 9, also phylogenetically close to classes 7 and 8, was found only in New South Wales. The previously identied class 2 was also found to be responsible for a large number of outbreaks, mainly in Victoria. Conclusion The results demonstrate that, epidemiologically, most outbreaks of ILT in New South Wales are unrelated to those in Victoria and suggest a link between classes 8 and 9 and the Nobilis ILT vaccine (class 7). Keywords epidemiology; herpesvirus; infectious laryngotracheitis; poultry; polymerase chain reaction; restriction fragment length polymorphism Abbreviations CEK, chicken embryo kidney; ICP, infected cell protein; ILT, infectious laryngotracheitis; ILTV, infectious laryngotracheitis virus; PCR, polymerase chain reaction; RFLP, restriction fragment length polymorphism; TK, thymidine kinase
Aust Vet J 2011;89:8994 doi: 10.1111/j.1751-0813.2010.00665.x

like other herpes viruses, can induce a state of latency in carrier birds.24 ILT can be spread between farms in close vicinity by fomites and/or aerosols.5 Darkling beetles (also called litter beetles) have also been found to act as vectors for several poultry pathogens,6 including ILTV.7 It is common practice for layer farms to routinely vaccinate for ILTV, but broiler farms tend to vaccinate only in an outbreak situation, which causes diculties when both farm types are in the same area and vaccination is not being consistently performed. The virus is considered endemic in parts of Australia and outbreaks commonly recur in both Victoria (VIC) and New South Wales (NSW). During 2007 and 2008, particularly, an escalation of ILT outbreaks was recorded and control of these outbreaks was complicated by a shortage of two commercially available Australian vaccines, SA2 and A20 (Fort Dodge Australia Pty Ltd, Baulkham Hills, NSW, Australia), but especially the comparatively less virulent A20. A third vaccine, Nobilis ILT (Intervet Schering-Plough Animal Health, Bendigo, VIC, Australia) was registered for use in Australia in 2006 and also used during some of the outbreaks. The vaccine strain, SA2, originated from an Australian eld isolate and was attenuated through sequential passages in chicken embryos,8 whereas the A20 strain was generated by further passages of the SA2 strain in chicken embryonic cell culture to lessen residual virulence (T Bagust, pers. commun.) and to make it suitable for broiler vaccination. The Nobilis ILT vaccine is the Serva strain of ILTV grown in embryonated eggs.9 Embryo-propagated ILT vaccines have been recently found accountable for eld outbreaks in the USA10,11 so the ability to dierentiate between wild-type and vaccine strains of ILTV is important for epidemiological purposes and is useful for designing vaccination strategies to prevent ILT outbreaks. A range of studies in various countries have focused on the dierentiation of ILTV genotypes by using restriction fragment length polymorphism (RFLP) patterns from endonuclease-digested PCR products1216 and this has since become the most widely-accepted method for genotyping ILTV isolates. Some research has focused on the dierentiation of vaccine and eld isolates using these patterns.12,13 More recently, a combination of RFLP patterns for a number of ILTV genes has been used for classication of ILTV in Australia17 and elsewhere.11 Application of this methodology by Kirkpatrick et al. on historical ILTV isolated from dierent locations in Australia resulted in identication of ve dierent ILTV classes, with the SA2 and A20 vaccine strains classied as class 1, and the remaining four classed as eld strains.17 The main purpose of the current study was to use this methodology to identify the ILTV classes responsible for the recent outbreaks of ILT in Australia, to establish if there are any links

nfectious laryngotracheitis (ILT) is a worldwide problem for the intensive poultry industry. In Australia, the disease causes substantial economic losses to both layer and broiler production companies, often with high rates of mortality on farms. It is a highly contagious viral respiratory illness aecting gallinaceous birds and is associated with acute respiratory distress and reduced egg production.1 The virus (ILTV) is a member of the herpesviridiae family and,

*Corresponding author. a School of Veterinary Science, The University of Melbourne, 250 Princes Highway, Werribee, Victoria 3030, Australia; hblacker@unimelb.edu.au b Baiada Poultry, Laverton, Victoria, Australia

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between the outbreaks occurring in dierent states and to investigate if the vaccine strains were linked to these outbreaks. Material and methods
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Viruses The protocol originally used by Kirkpatrick et al.17 to classify ILTV genotypes was applied in the current study (Table 1). The commercial ILTV vaccine strains, SA2, A20 and Serva, together with ILTV eld isolates collected over a 3-year period were compared (Table 2). Submission of eld samples was at the discretion of the eld veterinarian. All eld isolates were isolated from the upper respiratory tract and/or conjunctiva of infected birds during outbreaks of ILT in Australia. All but two submissions examined for this study were made from commercial layer and broiler farms reporting mortalities associated with ILT. Two submissions were from ocks of game birds (Table 2). A vaccine strain associated solely with morbidity, but not with signicant mortality, from vaccine reactions was not included in this study. A total of 92 ILTV isolates were examined, of which 52 were from NSW, 36 were from VIC and 4 were from Queensland (QLD). Extraction of viral DNA DNA was extracted from infected chicken embryo kidney (CEK) cell supernatant, commercial vaccines or directly from swabs of the trachea of an infected bird using a method described previously.17 If insucient amplication was achieved in the PCR procedure, virus isolates were propagated on CEK cells using standard techniques.18

Scrapings from the trachea and/or conjunctiva of aected birds were diluted in cell culture medium and inoculated onto CEK cells. Once infected, CEK cells were frozen at -80C, thawed, centrifuged at 500g for 5 min, the supernatant was then removed and used for DNA extraction. The extracted DNA samples were used immediately or stored at -20C before the genes of interest were amplied by PCR. PCR Primers for the thymidine kinase (TK), infected cell protein 4 (ICP4) and ICP18.5 genes were used in PCR as described previously17 with some modications. The ICP4 and ICP18.5 PCRs were carried out by subjecting the reactions to 94C for 1 min followed by 35 cycles of 94C for 15 s, 64C for 45 s and 68C for 5.5 min, and a nal extension of 68C for 10 min. All PCRs were performed using Platinum Taq DNA polymerase high delity (Invitrogen, Carlsbad, CA, USA). A control tube containing distilled H2O, instead of extracted DNA, was included as the negative control in all PCR series. The PCR products were separated by electrophoresis in 1% agarose gels, stained with GelRed (Biotium, Hayward, CA, USA), and exposed to ultraviolet light for visualisation using the Kodak Gel Logic 1500 Imaging System with Kodak Molecular Imaging Software (Version 4.0.5, 2005). RFLP and staining In this procedure, 10 mL of PCR products were digested separately with the restriction endonucleases at 37C for 2 h as described previously.17 The restriction endonuclease enzyme MspI was used for the TK amplicon and HaeIII for the ICP4 and ICP18.5 amplicons. After digestion, the resultant DNA fragments were separated in a 15% polyacrylamide gel and visualised by staining with GelRed (Biotium) and digital imaging using the Kodak Gel Logic 1500 Imaging System with Kodak Molecular Imaging Software (Version 4.0.5, 2005). Cluster analysis The ILTV RFLP pattern combinations were subjected to a cluster analysis using the Free-Tree software.19,20 The data input was established by the presence or absence of a particular restriction site and designated either 1 or 0 at each position. Similarity coecients were calculated using the method described previously.21 An unrooted dendogram was constructed using the unweighted pair group method, and testing for robustness was obtained by bootstrapping using a count of 500 repetitions. Results New restriction pattern for ICP4 gene Examination of the ILTV vaccine strain, Serva, revealed a new RFLP pattern (D) for the ICP4 gene when digested with the restriction enzyme HaeIII (Figure 1). Using RFLP patterns generated from the TK, ICP4 and ICP18.5 genes, ILTV isolates occurred in seven dierent classes (Tables 1, 2), three of which have been reported previously.17 The four new classes (6, 7, 8 and 9) were named in chronological order of their detection (Table 1). ILTV class 7 was the Serva strain and classes 6, 8 and 9 were eld isolates. Epidemiological relationship of ILT outbreaks Samples from a total of 92 outbreaks in NSW, VIC and QLD were referred to our laboratory for detection and strain identication
2011 The Authors Australian Veterinary Journal 2011 Australian Veterinary Association

Table 1. Classication of ILTV eld and vaccine strains

Class

RFLP pattern

Vaccine strain equivalent A B C C A C C A20/SA2 (Fort Dodge) Serva (Intervet Nobilis ILT)

1a 2a 3a 4a 5ab 6c 7c 8c 9c Restriction enzyme PCR product

A B B B A B B A A MspI TK

A B A C A B D D D HaeIII ICP4

C A HaeIII ICP18.5

Classication is based on the combination of restriction fragment length polymorphoism (RFLP) patterns, using two dierent restriction enzymes, of the polymerase chain reaction (PCR) products from three separate genes. ICP, infected cell protein; ILTV, infectious laryngotracheitis virus; TK, thymidine kinase. a Classes identied by Kirkpatrick et al.17 b Can only be distinguished from class 1 when subjected to additional RFLP, using the Fok1 enzyme.17 C Identied during the current study.

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Table 2. Field isolates typed in the current 3-year studya

Year 2007

ILTV class 2 2 6 7 2 2 2 2 8 9 7+8 7+8 8+9 7+8+9 2 2 3 3 8 9 8+9 7+8+9

Region of Australia Mornington Peninsula, VIC North-east Melbourne, VIC North Central VIC Sydney basin, NSW Mornington Peninsula, VIC North-east Melbourne, VIC Geelong outskirts, VIC Central NSW Sydney basin, NSW Sydney basin, NSW Geelong outskirts, VIC Sydney basin, NSW Sydney basin, NSW Sydney basin, NSW Mornington Peninsula, VIC Gold Coast Hinterland, QLD Wide Bay-Burnett region, QLD Central NSW Sydney basin, NSW Sydney basin, NSW Sydney basin, NSW Sydney basin, NSW

Flock type Broilers Layers Broilers Layers Broilers/layers Broilers Broilers Unknown Broilers/layers Broilers Layers Layers Layers Unknown Broilers Broilers Game birds Game birds Broilers Broilers Broilers Unknown

No. of isolates 8 1 4 1 14 2 2 2 29 2 1 1 1 1 4 3 1 1 8 4 1 1 92

2008

2009

Total
a

Submissions made to the typing laboratory were at the discretion of eld veterinarians, so not all outbreaks have been identied.

bp

MW

viruses were detected in specimens from a range of commercial ocks, including broilers, layers or game birds, comprising in total 52 submitted from NSW, 36 from VIC and 4 from QLD. The majority (37/52) of the submissions from NSW were found to contain only class 8 ILTV and 4/52 were found to contain only class 9 or class 2 ILTVs (Figure 2). A number of submissions contained a mixture of two, or occasionally three, classes of ILTV, with class 8 isolated in all cases of infections (Table 2). In contrast, 31 of the 36 submissions from VIC were found to contain only class 2 ILTV (Figure 2), with 4 submissions from an isolated poultry company in central VIC found to contain ILTV class 6. There was a further single submission from a commercial layer ock that had recently been vaccinated with the Serva strain (class 7) and it was found to contain a mixture of classes 7 and 8 ILTV. Of four submissions from QLD, three were found to contain class 2 and one contained class 3 ILTV (Figure 2). Cluster analysis of the RFLP patterns The cluster analysis of the RFLP patterns for all identied Australian ILTV classes clustered classes 1, 3 and 5 in one group, classes 79 in a second group, and classes 2, 4 and 6 in a third group (Figure 3). Classes 1 and 5 are only distinguishable by a further RFLP digest using the

750 500 300

150

50

Figure 1. Representative gel showing the fragmentation patterns A, B, C and D generated by restriction enzyme digest of the ICP4 amplicon by HaeIII, separated in 15% polyacrylamide gel electrophoresis.

between 2007 and 2009 (Table 2). These excluded cases of no or low incidence of mortality related to a vaccination reaction (as conrmed by detection of class 1 or 7 genotype only). From outbreaks associated with high mortality, ILTV classes 2, 3, 6, 7, 8 and 9 were detected. These
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Class 3

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Class 1

Class 5

Class 7

QLD
3 2

Brisbane

Class 8

NSW

Class 9
2 3 7, 8, 9

Sydney
Class 2

VIC
6 2 2, 7, 8 2

Melbourne

Figure 2. Geographical distribution of outbreaks of infectious laryngotracheitis (ILT) in the eastern states of Australia, with prevalent ILT virus genotypes represented for each region.

Class 4

Fok1 enzyme,17 which was not performed in this study, so for the purpose of this analysis they have been presented without horizontal branches. Discussion In this study, a combination of the RFLP patterns of a number of ILTV genes was used to investigate the source of, and a possible link between, recent outbreaks of ILT occurring predominantly in two states of Australia, VIC and NSW. The application of this technique resulted in the discovery of four new Australian ILTV classes (69) in addition to those reported previously (classes 15).17 Examination of the ILTV isolates collected during a 3-year period from NSW and VIC revealed that outbreaks in those two states were largely epidemiologically unrelated. The outbreaks in VIC were mainly caused by class 2 ILTV, a class that has been in circulation in that state since at least 1999.17 This class was distinct from ILTV classes 79 by cluster analysis, and also distinct from the vaccine strains SA2 and A20 that are predominantly used for the control of ILT in VIC. Although cluster analysis may be considered a crude means of establishing the relationships between ILTV strains, it is still a useful technique for clustering of the strains generated from RFLP. The results obtained by cluster analysis should be conrmed by analy-

0.1

Class 6

Figure 3. Dendogram constructed using cluster analysis of RFLP patterns for the ILTV genes, TK, ICP4 and ICP18.5, for the nine classes of ILTV. Bar, 0.1, represents the phylogenetic distance (percentage of divergence divided by 100). Similarity coecients were calculated based on Nei-Li and DICE and a bootstrap resampling method was used for 500 repetitions. ICP, infected cell protein; ILTV, infectious laryngotracheitis virus; RFLP, restriction fragment length polymorphism; TK, thymidine kinase.

sis of the entire genomic sequence of the ILTV strains, but this was outside the scope of the current study. A number of outbreaks involving a single broiler company in central VIC in 2007 were caused by ILTV class 6, a class not reported elsewhere in Australia. The company did not report further outbreaks. The geographical isolation of this particular operation, as well as eective biosecurity measures, are likely to have contributed to conning further outbreaks (personal communication with the company veterinarian). However, biosecurity measures may not be as eectively implemented in farms operating on a smaller scale, such as game bird ocks. The presence of a class 3 ILTV isolate in game bird ocks in both NSW and QLD suggests movement of the class 3 virus in association with game-bird transport. Records from our laboratory indicate that this genotype was in circulation in VIC and South Australia
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in 2004,17 which implies that class 3 ILTV has migrated to backyard and other small-scale poultry operations, which are not usually vaccinated and therefore may act as a reservoir of infection for Australian commercial poultry, and vice versa. In NSW, ILTV class 8 was responsible for the majority of outbreaks, a class that by cluster analysis was closely related to the Serva vaccine strain (class 7; Figure 3) commonly used in NSW for the control of ILT outbreaks. ILTV class 9, also closely related to the Serva vaccine strain, was isolated from a number of ILT outbreaks in NSW. It is notable that in only one case (a poultry (layer) farm in VIC), a mixed infection of ILTV classes 7 + 8 was detected in association with severe clinical signs and mortality. The history with this submission indicated administration of the Serva vaccine strain before samples were obtained from the outbreak. This was the only VIC case to reveal a class 7 or 8 ILTV in the period of this study. Several cases from NSW, however, revealed mixed infections of classes 7 + 8, 8 + 9 or 7 + 8 + 9 (Table 2). The results presented here suggest that outbreaks caused by ILTV classes 8 and 9 may have been related to the use of the Serva vaccine strain. It seems unlikely that these ILTV classes are true eld isolates, which by chance show molecular similarity to class 7 ILTV, particularly because these genotypes were not detected in Australia before the introduction of the Serva vaccine strain.17 Using viral plaquepurication techniques, a previous study in the USA demonstrated the presence of quasispecies within ILT vaccines of chicken embryo origin.22 Therefore, it is possible that ILTV classes 8 and 9 exist as subpopulations within the Nobilis ILT vaccine preparation, which are then subjected to selective pressure in the host species, allowing them to dominate in vivo. Reversion to virulence of the ILT vaccine strains after passage in vivo has been previously documented,23 which is an alternative explanation of the source of classes 8 and 9 ILTV in Australia. Studies in Northern Ireland,13 Taiwan12 and the USA22 have all speculated that since the introduction of live-attenuated ILTV vaccines, vaccine strain viruses have displaced the wild-type viruses and been responsible for many of the ILT outbreaks occurring in those regions. It therefore seems prudent that the importation of a live ILT vaccine, which has the potential to produce wild-type-like outbreaks of a foreign ILTV strain, should be allowed only after vigorous testing of safety and reversion to virulence. In this study, genotyping of the ILTV isolates by PCRRFLP lead to an understanding of the behaviour and spread of ILTV in Australia. The technique was adopted from a previous study that screened a number of ILTV genes, genomic regions and restriction enzymes to select the most useful combination for ILTV typing.17 The ILTV genomic region, ORFB-TK, which has been described previously,17 was not examined in this study, because assessment of the earlier isolates did not appear to increase the discriminatory power of the RFLP system. In addition, RFLP only detects nucleotide sequence variations within the restriction sites used and there may be other variations in the genome that are undetected by this technique. However, the relatively stable nature of herpesvirus genomes means that nucleotide changes usually confer separate virus strains. Complete nucleotide sequences of the genes used in this study, or of other ILTV genes such as gM/UL9 and gG/UL47,11 may be useful to further conrm the relatedness of the classes designated in this study. It would be desirable to examine ILTV
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classes found in Australia against those found elsewhere; however, because dierent ILTV genes are used for strain identication in different laboratories, such an examination would require a coordinated international eort. Alternatively, complete genomic sequence analyses of the ILTV isolates characterised in this study may enable a full understanding of the relationship of the viral isolates. Information derived from full genome sequencing will also be useful for developing more rapid classication techniques, such as real time PCR with high-resolution melt curve analysis or single-strand conformation polymorphism analyses.

Acknowledgments Funding for this project was provided by the Australian Poultry Cooperative Research Centre (CRC). The authors thank Dr Peter Scott for his scientic input, Peter Cowling, Rebecca Agnew and Kylie Hewson for their technical support, the Victorian and New South Wales Departments of Primary Industries for supplying some of the ILTV isolates, and the eld veterinarians for submission of clinical cases that were suspected to involve ILT viruses.

References
1. Guy JS, Bagust TJ. Laryngotracheitis. 11th edn. Iowa State Press, Ames, IA, 2003. 2. Creelan JL, Calvert VM, Graham DA, McCullough SJ. Rapid detection and characterization from eld cases of infectious laryngotracheitis virus by real-time polymerase chain reaction and restriction fragment length polymorphism. Avian Pathol 2006;35:173179. 3. Hughes CS, Jones RC, Gaskell RM, Jordan FT, Bradbury JM. Demonstration in live chickens of the carrier state in infectious laryngotracheitis. Res Vet Sci 1987;42:407410. 4. Hughes CS, Williams RA, Gaskell RM et al. Latency and reactivation of infectious laryngotracheitis vaccine virus. Arch Virol 1991;121:213218. 5. Johnson YJ, Gedamu N, Colby MM et al. Wind-borne transmission of infectious laryngotracheitis between commercial poultry operations. Int J Poultry Sci 2005;4:263267. 6. Goodwin MA, Waltman WD. Transmission of Eimeria, viruses, and bacteria to chicks: darkling beetles (Alphitobius diaperinus) as vectors of pathogens. J Appl Poultry Res 1996:5;5155. 7. Ou SC, Giambrone JJ, Macklin K. Detection of infectious laryngotracheitis virus from the darkling beetle and its larval stage (lesser mealworm) by real-time PCR. In: Proceeding of the 97th Annual Meeting of the Poultry Science Association, Niagara Falls, Ontario, Canada, 2008. 8. Purcell DA, Surman PG. Aerosol administration of the SA-2 vaccine strain of infectious laryngotracheitis virus [Letter]. Aust Vet J 1974;50:419420. 9. Australian Pesticides and Veterinary Medicines Authority. Notice of registration of agricultural and veterinary chemical products. 3. Veterinary products based on existing active constituents [Gazette]. APVMA, ACT, 2006:13. 10. Davison S, Dufour-Zavala L, Garcia M et al. Vaccinal laryngotracheitis: overview in the United States. In: Proceedings of the 109th Annual Meeting of the United States Animal Health Association, Hershey, PA, USA, 2005:580. 11. Oldoni I, Garcia M. Characterization of infectious laryngotracheitis virus isolates from the US by polymerase chain reaction and restriction fragment length polymorphism of multiple genome regions. Avian Pathol 2007;36:167176. 12. Chang PC, Lee YL, Shien JH, Shieh HK. Rapid dierentiation of vaccine strains and eld isolates of infectious laryngotracheitis virus by restriction fragment length polymorphism of PCR products. J Virol Methods 1997;66:179186. 13. Graham DA, McLaren IE, Calvert V, Torrens D, Meehan BM. RFLP analysis of recent Northern Ireland isolates of infectious laryngotracheitis virus: comparison with vaccine virus and eld isolates from England, Scotland and the Republic of Ireland. Avian Pathol 2000;29:5762. 14. Han MG, Kim SJ. Analysis of Korean strains of infectious laryngotracheitis virus by nucleotide sequences and restriction fragment length polymorphism. Vet Microbiol 2001;83:321331.

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15. Han MG, Kim SJ. Comparison of virulence and restriction endonuclease cleavage patterns of infectious laryngotracheitis viruses isolated in Korea. Avian Pathol 2001;30:337344. 16. Han MG, Kim SJ. Ecacy of live virus vaccines against infectious laryngotracheitis assessed by polymerase chain reaction-restriction fragment length polymorphism. Avian Dis 2003;47:261271. 17. Kirkpatrick NC, Mahmoudian A, ORourke D, Noormohammadi AH. Dierentiation of infectious laryngotracheitis virus isolates by restriction fragment length polymorphic analysis of polymerase chain reaction products amplied from multiple genes. Avian Dis 2006;50:2834. 18. Tripathy DN, Hanson LE. Laryngotracheitis. American Association of Avian Pathologists, College Station, TX, 1980. 19. Pavlicek A, Hrda S, Flegr J. Free-Tree: freeware program for construction of phylogenetic trees on the basis of distance data and bootstrap/jackknife analysis of the tree robustness. Application in the RAPD analysis of genus Frenkelia. Folia Biol (Praha) 1999;45:9799. 20. Free-Tree. http://www.natur.cuni.cz/~egr/programs/freetree. Accessed December 2009. 21. Nei M, Li WH. Mathematical model for studying genetic variation in terms of restriction endonucleases. Proc Natl Acad Sci USA 1979;76:52695273. 22. Garcia M, Riblet SM. Characterization of infectious laryngotracheitis virus isolates: demonstration of viral subpopulations within vaccine preparations. Avian Dis 2001;45:558566. 23. Guy JS, Barnes HJ, Smith L. Increased virulence of modied-live infectious laryngotracheitis vaccine virus following bird-to-bird passage. Avian Dis 1991; 35:348355. (Accepted for publication 9 August 2010)

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BOOK REVIEW

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Notes on small animal dermatology. J Joyce. Wiley-Blackwell, 2010. 365 pages. Price A$79.95. ISBN 978 1 4051 3497 2.

his book by British veterinarian Judith Joyce, who has almost three decades of small animal dermatology experience, is part of the Notes on series, of which Ive previously reviewed the excellent Notes on Canine Internal Medicine and Notes on Feline Internal Medicine. All of these soft-cover books are well produced, compact, well organised and easy to navigate.

next section is titled the Aetiological Approachand is broken down into chapters on disease types, including endocrine disease, neoplastic skin disease and hypersensitivity dermatitis. The following section titled Anatomically Localised Skin Disease has chapters on the dierent areas of the body, such as ear disease, periocular skin disease and the foot. The nal section, Treatment of Skin Diseases, is divided into chapters on treatment of presenting signs and primary skin disease, topical treatments and the use and abuse of corticosteroids. Chapters are easy to navigate, with frequent use of bold type for emphasis, dot point summaries to quickly show relevant information, plus many easy to read tables and ow-charts. There is also useful information on drug dose rates. The book contains many excellent colour photographs throughout, which feature examples of lesions and diagnoses. With dermatology cases being a common part of small animal practice, Notes on Small Animal Dermatology is a concise and wellpresented book, which should prove invaluable to the practicing small animal veterinarian as a quick reference guide in the clinic. This book could also assist veterinary students, through presentation of dierent approaches to the diagnosis of skin diseases. P Tucak Dr Phil Tucak works as both a veterinary surgeon in companion animal practice and as a television producer and journalist doi: 10.1111/j.1751-0813.2010.00671.x

Notes on Small Animal Dermatology begins by covering the investigation and diagnosis of dermatology cases, including details on the types of skin examinations and testing that would be required by the small animal practitioner. The book is then further divided into four sections, oering dierent approaches to diagnosis, each having a dierent appeal, depending on the work method of the practitioner. The rst section is titled Problem-Oriented Approach, which allows practitioners to nd information quickly, according to the skin problem presented, and includes the pruritic patient, the alopecic patient and management of raised and ulcerative skin lesions. The

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