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There is no doubt that molecular biologic technologies such as PCR testing have dramatically increased the sensitivity of diagnostic tests designed to detect the presence of several infectious pathogens including Streptococcus equi subsp. equi (Strep equi), Salmonella sp., CEMO and Dermatophytosis. Following on from an outbreak of strangles it is important to establish freedom from infection in all affected animals before “a line is drawn” under the outbreak and horses are free to mix and attend shows. Evidence suggests that guttural pouches are the only logical target for post-outbreak sampling with latest research indicating a greater than 50 times chance of detecting the presence of Strep equi in the guttural pouches of carriers compared to their nasopharynx (Boyle and others 2017). Furthermore, recent studies indicate that around 40% of culture-negative strangles submissions may be positive by PCR (Boyle and others 2017; Pusterla and others 2018).
However, the main drawback to reliance on PCR testing is that the technique simply detects the presence or absence of the DNA or RNA of the organism in question, whether or not that material is present in live or dead organisms. Indeed one recent study indicated that less than 20% of PCR positive and culture negative samples contained viable organisms (Pusterla and others 2018). It has been suggested that the lack of mucociliary clearance mechanisms in the guttural pouch could facilitate retention of DNA from dead Strep equi organisms leading to positive PCR results that do not actually indicate any risk of contagion.
A new PCR assay has been developed at Liphook which can distinguish live from dead Strep equi organisms. Use of this PCR will enable much greater confidence in the clinical relevance of positive PCR results. Results from examination of 10 washes, 5 of which had been boiled to kill the organisms, is shown in the graph below. The PCR was able to distinguish the live from dead organisms. Further validation in greater numbers of samples is ongoing.
In order to avoid the possible confounding effect of death of organisms between sampling and testing, it is important that samples are shipped in chilled saline (ACTH chiller packs for example), as previous evidence indicates that viability of Strep equi is excellent in cold and wet conditions (Durham and others 2018).
BOYLE, A. G., STEFANOVSKI, D. & RANKIN, S. C. (2017) Comparison of nasopharyngeal and guttural pouch specimens to determine the optimal sampling site to detect Streptococcus equi subsp equi carriers by DNA amplification. BMC Vet Res 13, 75
DURHAM, A. E., HALL, Y. S., KULP, L. & UNDERWOOD, C. (2018) A study of the environmental survival of Streptococcus equi subspecies equi. Equine Vet J
PUSTERLA, N., LEUTENEGGER, C. M., BARNUM, S. M. & BYRNE, B. A. (2018) Use of quantitative real-time PCR to determine viability of Streptococcus equi subspecies equi in respiratory secretions from horses with strangles. Equine Vet J 50, 697-700
How does the seasonal influence on the pituitary gland influence the diagnosis of PPID at this time of the year? And how does it impact follow-up blood samples in cases that were previously under control? What happens if you try to start pergolide treatment in July or August?
Let us take the stress out of managing your PPID cases – download our latest (free) podcast with Victoria South and Andy Durham who discuss what happens to our basal ACTH reference intervals at this time of the year, and chat through some common case scenarios that are affected by the pituitary hyperactivity observed at this time of the year – Seasonal PPID Podcast
We have upgraded our Endocrinology Analysers and are proud to introduce the Immulite 2000 xpi System.
Our driving ethos at Liphook has always been to stay ahead of the game and offer the latest and best to all of our laboratory users. Thus, we have recently purchased two Siemens Immulite 2000 xpi analysers, the newest and most proficient chemiluminescent immunoassay system available in this format which supersedes the older Immulite 1000 system used by most other labs. These new analysers allow greater precision, accuracy and throughput and, given that we have duplicate machines, this means that we can guarantee uninterrupted output of results even in the unlikely event of a machine malfunction.
It is well recognised that different assays and analysers generally return different results, and even the change from Immulite 1000 to Immulite 2000 xpi is no different in this respect. We have spent the past 6 months validating the new analysers and re-establishing appropriate cutoffs for diagnostic use and so you will see slight differences on forthcoming laboratory reports.
We apologise as I’m sure we’d all prefer to stay with our previous familiar values, but this change not only improves test reliability, but will in time be inevitable as the old Immulite 1000 system is withdrawn. We felt that as reference intervals (for ACTH at least) are about to change with the season in any case, this was the best time to implement this change. Should you require any assistance with interpretation or comparison with previous results then don’t hesitate to contact us to discuss.
Effects on ACTH reporting
As a general guideline the newly generated ACTH values will be slightly lower than previously (by around -10%) meaning lower cutoffs. In general terms this means a cutoff between 23-26 pg/mL for most of the year, rising slowly from July to a peak of 50 pg/mL in late September and returning to baseline by mid-November. Liphook Equine hospital remains the only laboratory internationally with valid ACTH reference ranges available for every week of the year, allowing greater accuracy of interpretation.
Effects on Insulin reporting
As a general guideline the newly generated insulin values will be higher than previously (by around +30%) meaning higher cutoffs. Resting insulin values in normal horses are expected to be no higher than 30-50 mU/L depending on the diet, and the response to 0.45 mL/kg Karo Light Corn Syrup should be no higher than 63 mU/L between 60-90 mins post-dosing.
We appreciate that is can be tricky to keep in touch with the latest publications on endocrinopathic laminitis especially those relevant to the diagnostic tests that we use in practice. We have produced this handy summary of the best evidence-based protocol for the Karo Light Challenge Test. Please click here to view the protocol.
We are all now recognising that most cases of liver disease that we encounter are part of a wider outbreak amongst herd-mates, even if the others appear externally healthy. This suggests a common toxin or infectious aetiology. It is useful to rule out “old favourites” such as ragwort by biopsy. Fluke is rare and almost impossible to rule out as no antemortem tests are reliable.
Work performed at Liphook over the last few years has identified fumonisin B1 contamination of forage as a possible cause of some outbreaks and international work looking at various causes of viral hepatitis in horses has identified a number of candidates including equine Hepacivirus, equine Pegivirus, Theiler’s disease associated virus and equine Parvovirus.
See Cornell University Laboratory website:
and recent publication describing equine Parvovirus: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5782890/pdf/17-1031.pdf
Continued work at Liphook has identified equine Parvovirus in some horses with liver disease and we hope to offer this, and other, new PCR assays shortly.
Liphook Equine Hospital are pleased to announce the date of our first Equine Veterinary CPD of 2018.
The event will take place on Wednesday 21st March at The Petersfield School, Petersfield, and is very kindly sponsored by Dechra.
To book your FREE place, please contact our hospital reception team on: 01428 727200, or via email to firstname.lastname@example.org
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