P F Nettleton

Moredun Research Institute, Penicuik, Scotland, United Kingdom

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Publications (96)204.05 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: This report describes the full-length genome sequence of the pestivirus strain H2121 which was recently isolated from Pyrenean chamois and typed as Border disease virus (BDV) genotype 4. Comparison with full-length genomic sequences of the approved pestivirus species Bovine viral diarrhea virus-1 (BVDV-1), BVDV-2, BDV, and Classical swine fever virus, the tentative species represented by strain Giraffe-1, as well as the atypical pestivirus strain Th/04_KhonKaen confirmed that the chamois pestivirus strain is most similar to BDV. The viral genome of H2121 is 12,305 nucleotides long and contains one large open reading frame. The latter encodes a polyprotein consisting of 3899 amino acids and is flanked with 376 nucleotides long 5' untranslated region (UTR) and 229 nt long 3' UTR. The genome organization of the chamois virus is reminiscent to that of other pestiviruses. Compared to other BDV strains including BDV-1 strain X818 and BDV-2 strain Reindeer-1, the 5' UTR and ORF of the chamois virus are very similar in length, while the 3' UTR of H2121 is 31-44 nucleotides shorter. In contrast to other BDV strains, the genome of the chamois virus contains a unique four amino acid insertion at the N-terminus of NS2.
    Virus Research 09/2010; 152(1-2):164-8. · 2.75 Impact Factor
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    ABSTRACT: The dramatic decline of the native red squirrel in the UK has been attributed to both direct and disease-mediated competition with the grey squirrel where the competitor acts as a reservoir host of squirrelpox virus (SQPV). SQPV is threatening red squirrel conservation efforts, yet little is known about its epidemiology. We analysed seroprevalence of antibody against SQPV in grey squirrels from northern England and the Scottish Borders in relation to season, weather, sex, and body weight using Generalized Linear Models in conjunction with Structural Equation Modelling. Results indicated a heterogeneous prevalence pattern which is male-biased, increases with weight and varies seasonally. Seroprevalence rose during the autumn and peaked in spring. Weather parameters had an indirect effect on SQPV antibody status. Our findings point towards a direct disease transmission route, which includes environmental contamination. Red squirrel conservation management options should therefore seek to minimize squirrel contact points.
    Epidemiology and Infection 07/2010; 138(7):941-50. · 2.87 Impact Factor
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    ABSTRACT: The squirrel poxvirus (SQPV) is the probable mediator of apparent competition between the introduced invading gray squirrel (Sciurus carolinensis) and the red squirrel (Sciurus vulgaris) in the UK, and modeling studies have shown that this viral disease has had a significant impact on the decline of the red squirrel in the UK. However, given our limited understanding of the epidemiology of the disease, and more generally the effects of invasive species on parasite ecology, there is a need to investigate the transmission dynamics and the relative pathogenicity of the virus between species. We aimed to increase our knowledge of these processes through an empirical study in which we: (i) used pathological signs and transmission electron microscopy (TEM) to diagnose SQPV disease in red squirrels found dead during scanning surveillance between 1993 and 2005; (ii) detected antibody to SQPV using an enzyme-linked immunosorbent assay (ELISA) in the same animals; and (iii) mapped cases of the disease, and the gray squirrel distribution, using a geographical information system. We analyzed the distribution of cases of SQPV disease according to woodland type, a measure of squirrel density. SQPV disease occurred only in areas of England also inhabited by seropositive gray squirrels, and as the geographical range of gray squirrels expanded, SQPV disease occurred in these new gray squirrel habitats, supporting a role for the gray squirrel as a reservoir host of the virus. There was a delay between the establishment of invading gray squirrels and cases of the disease in red squirrels which implies gray squirrels must reach a threshold number or density before the virus is transmitted to red squirrels. The spatial and temporal trend in SQPV disease outbreaks suggested that SQPV disease will have a significant effect on Scottish populations of red squirrels within 25 years. The even spread of cases of disease across months suggested a direct rather than vector-borne transmission route is more likely. Eight juvenile and sub-adult free-living red squirrels apparently survived exposure to SQPV by mounting an immune response, the first evidence of immunity to SQPV in free-living red squirrels, which possibly suggests a changing host-parasite relationship and that the use of a vaccine may be an effective management tool to protect remnant red squirrel populations.
    EcoHealth 11/2008; 5(3):305-16. · 2.20 Impact Factor
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    ABSTRACT: Squirrelpox virus (SQPV) causes a fatal disease in free-living red squirrels (Sciurus vulgaris) which has contributed to their decline in the United Kingdom. Given the difficulty of carrying out and funding experimental investigations on free-living wild mammals, data collected from closely monitored natural outbreaks of disease is crucial to our understanding of disease epidemiology. A conservation programme was initiated in the 1990s to bolster the population of red squirrels in the coniferous woodland of Thetford Chase, East Anglia. In 1996, 24 red squirrels were reintroduced to Thetford from Northumberland and Cumbria, while in 1999 a captive breeding and release programme commenced, but in both years the success of the projects was hampered by an outbreak of SQPV disease in which seven and four red squirrels died respectively. Valuable information on the host-pathogen dynamics of SQPV disease was gathered by telemetric and mark-recapture monitoring of the red squirrels. SQPV disease characteristics were comparable to other virulent poxviral infections: the incubation period was <15 days; the course of the disease an average of 10 days and younger animals were significantly more susceptible to disease. SQPV disease places the conservation of the red squirrel in jeopardy in the United Kingdom unless practical disease control methods can be identified.
    Epidemiology and Infection 07/2008; 137(2):257-65. · 2.87 Impact Factor
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    ABSTRACT: Bovine respiratory syncytial virus is an agent involved in calf pneumonia complex, a disease of significant economic importance. Accurate diagnosis of the agents involved on farm premises is important when formulating disease control measures, including vaccination. We have developed a real time reverse transcriptase polymerase chain reaction (rtRT-PCR) and compared it with the diagnostic tests currently available in the United Kingdom: immunohistochemistry (IHC) and immunofluorescence antibody test (IFAT). The rtRT-PCR had a detection limit of 10 gene copies and was 96% efficient. Recent UK isolates and clinical samples were tested; the rtRT-PCR was more sensitive than both conventional tests.
    Veterinary Microbiology 02/2008; 126(1-3):264-70. · 3.13 Impact Factor
  • P. F. Nettleton, K. Willoughby
    Diseases of Sheep, Fourth Edition, 01/2008: pages 119 - 127; , ISBN: 9780470753316
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    The Veterinary record 12/2007; 161(19):660-1. · 1.80 Impact Factor
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    The Veterinary record 05/2007; 160(15):532. · 1.80 Impact Factor
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    ABSTRACT: (S)-1-[3-hydroxy-2-(phosphonomethoxy)propyl]cytosine [corrected] (HPMPC, cidofovir, CDV, Vistide) is an acyclic nucleoside analogue with a potent and selective activity against a broad spectrum of DNA viruses including the poxviruses. In this study we present the results of different treatment regimens in lambs experimentally infected with orf virus with different cidofovir formulations prepared in Beeler basis and Unguentum M. Our results show that choice of excipient, concentration of codofovir [corrected] and treatment regimen were all important to the clinical outcome of the therapy. Whilst one particular regimen appeared to exacerbate the lesion, treatment with 1% (w/v) cidofovir cream, prepared in Beeler basis, for 4 consecutive days did result in milder lesions that resolved in milder lesions that resolved [corrected] more quickly than untreated lesions. Furthermore the scabs of the treated animals contained significantly lower amounts of viable virus meaning there should be less contamination of the environment with virus than would normally occur.
    Antiviral Research 04/2007; 73(3):169-74. · 3.93 Impact Factor
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    ABSTRACT: The identification of Louping ill virus (LIV) in clinical specimens has been routinely achieved by virus isolation using susceptible pig kidney cells and subsequent serological analysis. While this method is sensitive and detects infectious virus, it is relatively labour intensive and time-consuming. In view of the veterinary and potential medical importance of LIV, a rapid and precise detection method for routine use that employs the TaqMan reverse transcription polymerase chain reaction (RT-PCR) has been developed to detect LIV RNA extracted from field samples. The TaqMan assay was evaluated against virus isolation using 22 cell culture grown LIV isolates, which had previously been partially characterised by sequencing, and material from 63 suspect field cases. Histopathological and/or serological reports were available for 39 of the suspect cases, providing additional diagnostic information to evaluate the results obtained from the TaqMan RT-PCR assay. The TaqMan assay was as sensitive as the cell culture infectious virus assay currently used and had the advantage that it was able to detect LIV in clinical specimens from which infectious virus could not be isolated possibly due to the presence of high levels of LIV antibody.
    Journal of Virological Methods 11/2006; 137(1):21-8. · 1.90 Impact Factor
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    ABSTRACT: The genome of a virulent squirrelpox virus (SQPV) isolate was characterized in order to determine its relationship with other poxviruses. Restriction enzyme analysis suggested a genome length of approximately 158 kb, whilst sequence analysis of the two ends of the genome indicated a G + C composition of approximately 66 %. Two contiguous stretches of 23 and 37 kb at the left-hand and right-hand ends of the genome, respectively, were sequenced allowing the identification of at least 59 genes contained therein. The partial sequence of a further 15 genes was determined by spot sequencing of restriction fragments located across the genome. Phylogenetic analysis of 15 genes conserved in all the recognized genera of the subfamily Chordopoxvirinae confirmed that the SQPV does not group within the family Parapoxvirinae, but instead partitions on its own in a separate clade of the poxviruses. Analysis of serum from British woodland rodents failed to find any evidence of SQPV infection in wood mice or bank voles, but for the first time serum samples from grey squirrels in the USA were found to contain antibody against SQPV.
    Journal of General Virology 09/2006; 87(Pt 8):2115-25. · 3.13 Impact Factor
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    ABSTRACT: Malignant catarrhal fever (MCF) is a sporadic but fatal lymphoproliferative viral disease of cattle, deer and other ruminants. The causative agents are highly-cell-associated herpesviruses of the subfamily gammaherpesvirinae. In this study, an ELISA (WC11-ELISA) was developed to detect antibody to malignant catarrhal fever virus (MCFV) in cattle serum and compared to the commercially produced competitive-inhibition ELISA (CI-ELISA). Crude lysate antigen from alcelaphine herpesvirus-1 strain WC11 was bound to 96-well microplates and used to capture antibodies to MCFV. Dilutions of test sera were added to wells containing bound MCF antigen and control wells containing uninfected cell lysates. A horseradish peroxidase-labelled rabbit-anti-bovine IgG conjugate detected antibodies to MCF, and the results were expressed as absorbance readings at 450 nm. Samples were selected blind from cattle sera which had been sent to the laboratory for diagnostic testing for MCFV antibodies and were tested in both the WC11-ELISA and the CI-ELISA. Good agreement between the WC11-ELISA and CI-ELISA test (k=0.86, n=95) results was found.
    Veterinary Microbiology 09/2006; 116(1-3):21-8. · 3.13 Impact Factor
  • S Vilcek, P F Nettleton
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    ABSTRACT: Pestiviruses are not strictly host-species specific and can infect not only domestic but also wild animals. The most important pestivirus, CSFV, infects domestic pigs and wild boars, which may cause a major problem for successful CSFV eradication programmes. Mainly BVDV specific antibodies have been reported in captive and free-living animals. Virus has been isolated from some of these animal species, but since BVDV can contaminate cell cultures and foetal calf serum, early reports of BVDV isolation have to be considered with caution. Genetic typing of early pestivirus isolates from wild species revealed that the majority were BVDV-1. Of the pestiviruses identified so far three species (CSFV, BVDV-1, giraffe pestivirus) and three genotypes (BDV-2, BDV-4, pronghorn) appear to circulate in wildlife animal populations. The potential for pestiviruses to spread between farm animals and free-living animals is discussed as are epidemiological and technical problems, and the future direction of research.
    Veterinary Microbiology 09/2006; 116(1-3):1-12. · 3.13 Impact Factor
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    ABSTRACT: Squirrel poxvirus (SQPV) is a well-documented example of pathogen-mediated competition between an invasive species, the grey squirrel (Sciurus carolinensis), and a native species, the Eurasian red squirrel (Sciurus vulgaris). SQPV causes disease with high mortality in red squirrels but appears non-pathogenic in grey squirrels. Not all populations of introduced grey squirrels carry the virus, notably those in Scotland and Italy, and the rate of red squirrel replacement by grey squirrels is some twenty times faster in those areas where grey squirrels carry the virus. Here we develop strategies to manage the SQPV disease threat to red squirrels by reference to the largest, designated red squirrel refuge site in England, Kielder Forest (50 000 ha). Using modelling techniques, we identify four main corridors within the buffer zone by which grey squirrels will reach Kielder, initially within two years and in large numbers within 10 years. Assuming that greys will not settle within Kielder because of the unfavourable nature of the spruce habitat, we predict that SQPV disease will burn out at the edges of the forest, although many red squirrels will die. This burn-out is unlikely to be the scenario in other refuge areas where the habitat is more favourable to greys. We conclude that the conservation of red squirrels will depend on minimising contact between red and grey squirrel populations, and we advocate monitoring grey squirrels in corridors within buffer zones around refuge areas, and removing them when detected.
    Biological Conservation 08/2006; 131(2):287-295. · 3.79 Impact Factor
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    ABSTRACT: Red squirrels are declining in the United Kingdom. Competition from, and squirrel poxvirus (SQPV) disease carried by, grey squirrels are assumed to be determining the decline. We analyse the incidence of disease and changes in distribution of the two species in Cumbria, from 1993 to 2003 and compare these to the predictions of an individual-based (IB) spatially explicit disease model simulating the dynamics of both squirrel species and SQPV in the landscape. Grey squirrels increased whilst red squirrels declined over 10 years. The incidence of disease in red squirrels was related to the time since grey squirrels arrived in the landscape. Analysis of rates of decline in red squirrel populations in other areas showed that declines are 17-25 times higher in regions where SQPV is present in grey squirrel populations than in those where it is not. The IB model predicted spatial overlap of 3-4 years between the species that was also observed in the field. The model predictions matched the observed data best when contact rates and rates of infection between the two species were low. The model predicted that a grey squirrel population control of >60% effective kill was needed to stop the decline in red squirrel populations in Cumbria.
    Epidemiology and Infection 06/2006; 134(3):521-33. · 2.87 Impact Factor
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    ABSTRACT: A real time one-step RT-PCR was designed to detect and type border disease virus (BDV), bovine viral diarrhea virus (BVDV) type 1 and BVDV type 2 in ovine samples. The real time RT-PCR was shown to behave in a linear manner and had limits of detection of 100-1000 copies of viral RNA as judged by in vitro transcribed RNA. The real time RT-PCR was validated on 50 clinical samples from UK flocks and was more sensitive than a virus isolation and a classical nested RT-PCR (nRT-PCR). The results of real time RT-PCR virus typing agreed completely with sequencing. The majority of ovine isolates were BDV; a small proportion were BVDV type 1. BVDV type 2 was not detected in any sample. This test appears reliable and can be used for the typing of ovine pestiviruses in the UK.
    Journal of Virological Methods 04/2006; 132(1-2):187-94. · 1.90 Impact Factor
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    ABSTRACT: The population of red squirrels (Sciurus vulgaris) in the British Isles is in decline and is being supplanted by the grey squirrel (Sciurus carolinensis). It has been suggested that parapoxvirus-associated disease has caused significant mortality in red squirrels and that grey squirrels are the source of the virus. A direct enzyme-linked immunosorbent assay (ELISA) was developed for the measurement of antibody to squirrel parapoxvirus. We tested 140 sera from red squirrels and 223 from grey squirrels from different populations in the UK. A high percentage (61%) of apparently healthy grey squirrels, were found to have been exposed to the parapoxvirus. Only 2.86% (4/140) of red squirrels had antibody and three of these animals had parapoxvirus-associated disease. We postulate that the grey squirrel may act as a reservoir host for the virus.
    Animal Conservation 02/2006; 3(3):229 - 233. · 2.69 Impact Factor
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    ABSTRACT: Subspeciation of Campylobacter fetus subsp. fetus (CFF) and Campylobacter fetus subsp. venerealis (CFV) is important for international animal import regulations. Phenotyping can be unreliable, and genotyping by techniques like pulsed field gel electrophoresis is difficult in routine diagnostic laboratories. A PCR subspeciation technique has been reported [Aust Vet J (1997) 75, 827]; we aimed to develop this PCR and investigate its use on UK C. fetus isolates. We augmented the PCR with further primers, and tested 76 isolates of C. fetus and 16 isolates of other Campylobacter spp. PCR failed to correlate well with phenotyping, especially for CFV. We characterized the amplicon of the CFV-specific primers (reported as plasmid derived, but unavailable on the public databases); and predicted a parA gene sequence, anticipated to be plasmid-associated. However, although plasmid isolations from selected CFV isolates demonstrated the presence of several plasmids, there was no correlation between plasmid profile and PCR result. Further, the parA sequence was not detected by PCR in any of the plasmid bands. This PCR is not suitable for subspeciation of C. fetus in the UK. The results suggest that this is a reflection of the presence of an unusual clone of CFV currently present in cattle in this country. PCR cannot substitute for phenotyping of C. fetus isolates in the UK. The reasons for failure of PCR genotyping may reflect local strains and/or plasmid profiles. Further study is required to better elucidate molecular sub-speciation of C. fetus.
    Journal of Applied Microbiology 02/2005; 99(4):758-66. · 2.20 Impact Factor
  • The Veterinary record 02/2005; 156(2):59-60. · 1.80 Impact Factor
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    ABSTRACT: During investigations into recent population decreases in Pyrenean chamois (Rupicapra pyrenaica pyrenaica) 21 animals found dead or dying were necropsied. Immunohistochemistry revealed the presence of a pestivirus in organs from two of the 21 chamois. From one of these animals a pestivirus was isolated from the spleen, skin and serum. The virus had better growth in ovine than in bovine cells and was neutralized most effectively by an anti-border disease virus (BDV) reference antiserum. Using panpestivirus and genotype-specific primers selected from 5'-untranslated region (UTR) of the pestivirus genome, BDV RNA was demonstrated by RT-PCR. Comparison of the chamois sequences from 5'-UTR, entire N(pro) and E2 gene coding regions with those of other pestivirus genotypes revealed that this virus did not fall into any of the pestivirus genotypes identified so far. Results of phylogenetic analysis suggested that the chamois pestivirus was closely related to BDV and it was typed as BDV-4 genotype.
    Journal of General Virology 01/2005; 85(Pt 12):3653-7. · 3.13 Impact Factor

Publication Stats

2k Citations
204.05 Total Impact Points

Institutions

  • 1983–2010
    • Moredun Research Institute
      • • Virus Surveillance Unit
      • • Department of Parasitology
      Penicuik, Scotland, United Kingdom
  • 2007
    • University of Bologna
      Bolonia, Emilia-Romagna, Italy
  • 1995–2006
    • University of Veterinary Medicine in Kosice - Univerzita veterinarneho lekarstva v Kosiciach
      • Department of Parasitology and Infectious Diseases
      Kassa, Košický, Slovakia
  • 2005
    • University of Zaragoza
      • Faculty of Veterinary
      Zaragoza, Aragon, Spain
  • 2002
    • University of Otago
      • Department of Microbiology and Immunology
      Dunedin, Otago, New Zealand
  • 1990
    • The University of Edinburgh
      • Royal (Dick) School of Veterinary Studies
      Edinburgh, Scotland, United Kingdom
  • 1988
    • Sokoine University of Agriculture (SUA)
      • Department of Veterinary Medicine and Public Health
      Morogoro, Morogoro Region, Tanzania