Publications and studies

Find all Scanelis publications, the list of articles resulting from scientific collaborations, and the results of the latest Scanelis studies.

Scanelis Publications and Scientific Collaborations

Below is a list of the most recent scientific publications. When available, feel free to download them.

Publications Scanelis

	Distemper Boucraut-Baralon, C., C. Trumel. 2000. Diagnosis of distemper, the choice of a technique. The New Veterinary Practitioner August-September-October 2000: 51-53 Boucraut-Baralon, C. . 2006. How to diagnose and prevent distemper in dogs and ferrets. The New Canine Veterinary Practitioner, feline Special issue 2006: 83-87
	Mycobacteria Etienne C.-L., F. Granat, C. Trumel, I. Raymond-Letron, M.-N. Lucas, C. Boucraut-Baralon, J.-L. Pingret, L. Magne, M. Delverdier. 2013. A mycobacterial coinfection in a dog suspected on blood smear. 2013 42/4 : 516-521
	Enteritic pathogens Broussou, D., H. Mila, A. Grellet, A. Feugier, C. Mariani, J.-L. Pingret, C. Boucraut-Baralon, S. Chastant-Maillard. 2016. Excretion of canine parvovirus type 2 (CPV-2) during gestation and lactation in bitches and puppies >> Download this poster Grellet, A., R.M. Heilmann, B. Polack, A. Feugier, C. Boucraut-Baralon, D. Grandjean, N.Grüntzner, J.S. Suchodolski, J.M. Steiner, and S. Chastant-Maillard. 2016. Influence of Breed Size, Age, Fecal Quality, and Enteropathogen Shedding on Fecal Calprotectin and Immunoglobulin A Concentrations in Puppies During the Weaning Period. Journal of Veterinary Internal Medicine 2016. >> Download this article Grellet, A., S. Chastant-Maillard, A. Feugier, C. Boucraut-Baralon, D. Grandjean, B. Polack. Risk factors of weaning diarrhea in puppies housed in breeding kennels. Prev Veterinary Medecine 117(1) : 260-5 Download this article
	*Hepatozoon canis* Criado-Fornelio, A., A. Buling , N.A. Cunha-Filho, J.L. Ruas, N.A.R. Farias, C. Rey-Valeiron, J.L. Pingret, M. Etievant, J.C. Barba-Carretero. 2007. Development and evaluation of a quantitative PCR assay for detection of Hepatozoon sp.. Veterinary Parasitology 150 352–356
	Hémoprotozoaires Criado-Fornelio, A., A. Buling, J-L. Pingret, M. Etievant, C. Boucraut-Baralon, A. Alongi, A. Agnone, A. Torina. 2009. Hemoprotozoa of domestic animals in France : Prevalence and molecular characterization. Veterinary Parasitology 159 73–76
	Diagnostic Boucraut-Baralon, C. . 2001. La PCR, un nouvel outil pour le diagnostic des maladies infectieuses chez le chien. Prat Méd Chir Anim Comp 36 : 515-521
	Rétroviroses félines Boucraut-Baralon, C. . 2000. Laboratory diagnosis of feline retroviruses. The New Veterinary Practitioner June-July 2000: 45-47 Boucraut-Baralon, C. . 2006. Vaccination of an FIV-infected cat. The Veterinary Point, 271 12-13
	Eye diseases De Geyer, G., C. Boucraut-Baralon. 2001. Feline Herpesvirus-1 and Eye Disease in Cats. Prat med chir anim comp 36: 461-471
	Coryza Boucraut-Baralon, C. . 2002. Contagious feline coryza. Encycl Méd Chir (Editions Scientifiques et Médicales Elsevier SAS, Paris, all rights reserved), Encyclopédie vétérinaire (Elsevier, Paris), Médecine générale, 1800, 2002, 7p.
	Calicivirus Reynolds, B., C. Boucraut-Baralon. 2006. Diagnostic, traitement et prévention des caliciviroses félines. Le Nouveau Praticien Vétérinaire canine, féline Hors-série 2006 : 61-63 Reynolds, B. H. Poulet, J.-L. Pingret, D. Jas, S.Brunet, C. Lemeter, M.Etievant, C. Boucraut-Baralon. 2009. A nosocomial outbreak of feline calicivirus associated virulent systemic disease in France. Journal Of Feline Medicine and Surgery 11 : 633-644
	Herpèsvirose féline Donzel, E., C. Boucraut-Baralon, S. Mazzucchelli, N. Otero Coves, T. Verite, F. Laguna, J. Piteux, C. Molas, G. Storms, G. Payen, S. Chahory. 2012. Effect of fluorescein and topical anaesthetics on pcr used for the diagnosis of feline herpesvirus related conjunctivitis.
	Parvovirose féline (Typhus / Panleucopénie) Barrault, M-J., D. Rivière, S. Guionie. 2011. Deux cas d’épanchement pleural associés à une infection par le parvovirus félin. Le Point Vétérinaire 319 : 54-58
	Mycobacteria Rivière, D., J.-L. Pingret, M. Etievant, A. Jechoux, D. Lanore, I. Raymond-Letron, C. Corine Boucraut-Baralon. 2011. Disseminated Mycobacterium avium subspecies infection in a cat. Journal Of Feline Medicine and Surgery 13 : 125-128
	NHEO Guerin, J.-L., J. Gelfi, L. Dubois, A. Vuillaume, C. Boucraut-Baralon and J.-L. Pingret. May-2000. A novel polyomavirus (goose hemorrhagic polyomavirus) is the agent of hemorrhagic nephritis enteritis of geese. J Virol 74 : 4523-9. >>Download this article Lacroux, C., O. Andreoletti, B. Payre, J.-L. Pingret, A. Dissais and J.-L. Guerin. Jun-2004. Pathology of spontaneous and experimental infections by Goose haemorrhagic polyomavirus. Avian Pathol 33: 351-8. Pingret, J.-L., C. Boucraut-Baralon and J-L. Guérin. 2008. Goose haemorrhagic polyomavirus infection in ducks. Vet Record 2008 162:164. Derzsy and Parvovirus Muscovy Duck Pingret, J.-L., C. Zadjian, S. Lemière, and C. Boucraut-Baralon. 2005. Detection of palmiped parvoviruses by real-time PCR. Poultry Research Days 6: 423-7. >> Download this poster and this article Fontaine, J., C. Facon, M. Castets, X. Banse, O. Albaric, J.-L. Pingret, J.-Y. Douet and J.-L. Guérin. 2009. Necrotizing splenitis in mule ducklings : case report and etiological investigations. XVIth World Veterinary Poultry Association Congress Marrakesh.
	Chlamydiose, BFDV et APV Lafon, S., J.-L. Pingret et C. Boucraut-Baralon. 2005. Real-Time PCR for diagnosis of three common infectious diseases in caged birds : Chlamydophilosis, Beak and Feather Disease and Avian Polyomavirosis. 8th European AAV Conference.
	Mycobacteria Huynh, M., J.-L. Pingret, A. Nicolier. 2014. Disseminated Mycobacterium genavense Infection in a chinchilla. J Comp Pathol 151(1) : 122-5
	Parvovirus des rongeurs Lafon, S., J.-L. Pingret, N. Fiks, C. Médaille et C. Boucraut-Baralon. 2004.Real-Time PCR diagnostic test of rat Parvovirus infections and genetic strain identification - Comparative study with serological patterns. 9th FELASA symposium >> Download this poster
	Transgéniques Lafon, S., J.-L. Pingret, M. Le Roux, V. Turquois, P.J. Ripoll, C. Boucraut-Baralon.2007. Accurate method for determination of transgene copy number using Real-time PCR in rabbits. 10th FELASA symposium >> Nous contacter pour obtenir ce poster.
	Diagnostic Boucraut-Baralon, C. . 2009. Nouvelle méthode d’investigation, la PCR en dermatologie. Le Nouveau Praticien Vétérinaire canine, féline Juin 2009 : 38-40 Boucraut-Baralon, C. . 2009. PCR: Selection and collection of samples, interpretation of results. PratiqueVet 44: 42-45 Boucraut-Baralon, C. . 2008. PCR: Interest, Limitations and Key Indications. PratiqueVet 43: 670-673 Boucraut-Baralon, C. . 2006. The Contribution of Biological Techniques in the Diagnosis of Infectious Diseases. The New Canine Veterinary Practitioner, feline Special issue 2006: 83-87 Boucraut-Baralon, C. . 2002. PCR or Gene Amplification. Prat med chir anim comp 37: 303-304 Boucraut-Baralon, C. . 2001. Molecular Diagnostics - PCR Techniques and Pathogen Detection. The New Veterinary Practitioner November-December-January 2001: 63-66 Boucraut-Baralon, C. . 2001. La PCR, un nouvel outil pour le diagnostic des maladies infectieuses chez le chien. Prat Méd Chir Anim Comp 36 : 515-521 Boucraut-Baralon, C. . 2001. Diagnostic - Le western blot ou « immunoblotting ». Février-Mai 2001 : 70-72 Trumel, C., C. Boucraut-Baralon. 2001. Analysis and Comments - Interpretation of Bronchoalveolar Lavage Cytology. The New Veterinary Practitioner November-December-January 2001: 51-52

Scientific collaborations

	Oncology Lanore, D., D. Rivière, C. Delprat. 2010. Suspicion of piroplasmosis in a Bernese Mountain Dog: a diagnosis of hystiocytic sarcoma. The Bottom Line 167 24-28. Lanore, D., D.Rivière. 2010. Assessment of extension of canine cutaneous mastocytoma; Chemotherapy is indicated for stage 2. Veterinary Week 1396 32-33.
	Leishmaniasis Collignon, C., A. Zahra, L. Guénego, R. Gautier, A. Madelenat. 2009. Medical and Surgical Practice of the Pet 44 27-34
	Enteritic pathogens Solano-Gallego, L., L. Kidd, M. Trotta, M. Di Marco, M. Caldin, T. Furlanello, E. Breitschwerdt. 2006. Emerging Infectious Diseases Vol 12 (12): 1985
	Hemobartonella Beaufils, J-P.. 2012. Hemolytic anemia in a dog infected with Mycoplasma haemocanis. Medical and Surgical Practice of the Pet 47 43-47

Scanelis study: prevalence of H1N1 in cats in France

Reference of the article published by Scanelis, presenting the results of the epidemiological study of H1N1 in cats in France:

Vet Rec. 2010 Mar 6; 166(10):307. Epidemiological survey of H1N1 influenza virus in cats in France.
Pingret JL, Rivière D, Lafon S, Etiévant M, Boucraut-Baralon C.

on request to contact@scanelis.com, we will be pleased to send you the PDF file of this article.

However, a case was described in France at the end of 2009 (AFP announcement by the Director General of Health, Didier Houssin, 7/12/09; mention in a ProMed announcement of 11/12/09: INFLUENZA PANDEMIC (H1N1) 2009, ANIMAL (37) - USA (OREGON, CALIFORNIA) FELINE; article in the Veterinary Week n°1385 of December 18, 2009, page 16).

Scanelis Study: Kennel Cough

The Scanelis laboratory conducted a retrospective study on the prevalence of Bordetella bronchiseptica (Bb), canine adenovirus type II or respiratory adenovirus (Cav2) and canine pararainfluenza (CPI) in kennel cough syndrome or infectious tracheobronchitis.

138 dogs with clinical signs suggestive of kennel cough were included in this study. Biological samples of various types (oropharyngeal, nasal and/or conjunctival cells, bronchoalevolar fluid or organs) received between 2001 and 2005 at the Scanelis laboratory were analyzed by real-time PCR for the detection and quantification of the 3 pathogens mentioned above.

The pathogens of interest were considered to be involved in the clinical signs observed when viral or bacterial loads were significant.
Au contraire les faibles charges ont été comptabilisées dans la catégorie « Trace vaccinale » ou « Portage asymptomatique » selon l’historique vaccinal de l’animal (date et type de vaccin utilisé).
These 2 categories have been grouped together in the table below:

Prevalence of Bordetella bronchiseptica (Bb), canine adenovirus type II or respiratory adenovirus (Cav2) and canine pararainfluenza (CPI) in kennel cough syndrome

Pathogen(s)	Positive Animals	Prevalence
Bordetella bronchiseptica	42	30,4 %
Canine adenovirus type II	4	2,9 %
Canine parainfluenza	2	1,4 %
Bordetella bronchiseptica AND Canine Adenovirus Type II	3	2,2 %
Bordetella bronchiseptica AND Canine Parainfluenza	5	3,6 %
Bordetella bronchiseptica, Canine Adenovirus Type II AND Canine Parainfluenza	1	0,7 %
Trace vaccination or asymptomatic carriage of one or more of these agents	31	22,5 %
None of these 3 agents	50	36,2 %
TOTAL	142

In more than 40% of cases, at least one of the 3 pathogens of interest may be considered to be involved in the observed clinical signs.
In 5.8% of cases, 2 agents explain the symptoms and in one dog Bordetella bronchiseptica, canine adenovirus type II and canine parainfluenza were detected in significant amounts.

Individual prevalence of Bordetella bronchiseptica (Bb), canine adenovirus type II or respiratory adenovirus (Cav2) and canine pararainfluenza (CPI) in kennel cough syndrome

Pathogen(s)	Positive animals (significant load)	Prevalence
Bordetella bronchiseptica	51	37,0 %
Canine adenovirus type II	8	5,8 %
Canine parainfluenza	8	5,8 %

Of these 3 agents, Bordetella bronchiseptica is most frequently implicated in the clinical signs of kennel cough. Both viruses (respiratory adenovirus and canine parainfluenza) have a much lower but identical prevalence.

Prevalence of Bordetella bronchiseptica according to the age of animals suspected of kennel cough

	Dogs under 6 months of age	Dogs over 6 months of age
Negative	48 (45,7 %)	25 (86,2 %)
Significant Bb Load	47 (44,8 %)	1 (3,4 %)
Trace vaccination or asymptomatic carriage	10 (9,5 %)	3 (10,3 %)

In nearly 1 in 2 dogs suspected of kennel cough under 6 months of age, Bordetella bronchiseptica is implicated in the observed clinical signs. On the contrary, the bacterium is rarely implicated in kennel cough syndrome in dogs over 6 months of age (1/29).

Other infectious agents are described as possibly involved in the Kennel Cough complex (canine herpesvirus, Coronavirus, distemper virus, etc.). The prevalence of these viruses will be the subject of a future study.

Scanelis Study: Leishmanian Meningitis

Clinical case: Leishmanian meningitis, about 3 cases
S. CATHELAND (1) , D. RIVIERE (2), C. TRUMEL (3)
(1) Cabassu Veterinary Clinic, 12 avenue du Prado, 13010 Marseille, Tel: 04 91 37 16 30, catheland@wanadoo.fr
(2) SCANELIS Laboratory, Allée Charles Cros, 31770 Colomiers, Tel: 05 34 50 50 90, delphine.riviere@scanelis.com
(3) Central Laboratory of Clinics, National Veterinary School, 23 chemin des Capelles, 31300 Toulouse, c.trumel@envt.fr

Three dogs, Dora, a 1.5-year-old Brittany spaniel, Missi and Vanka, two Border Collies aged 5 and 9 respectively, are referred to the Cavassis clinic for neurology. Dora is presented for recurrent pain syndrome, Missi for hyperthermia and pain, Vanka for tetraparesis.

On the day of the consultation, Dora had no clinical abnormalities. Missi has pain in the thoracolumbar region and Vanka is tetraparetic.

CT scans were performed on the 3. Discrete abdominal adenomegaly is visualized for Missi and Dora. Meningitis is suspected for Vanka due to strengthening of the meningeal contours. Cerebrospinal fluid (CSF) punctures are performed. Cytological analyses reveal pleiocytosis corresponding to an inflammatory infiltrate of mixed mononuclear cells (macrophages and lymphoid cells) or exclusively macrophage for Missi. PCR analyses of distemper, toxoplasmosis, neosporosis, ehrlichiosis and leishmaniasis are requested. Tests are only positive for leishmaniasis for all 3 CSFs. On the other hand, serological blood tests are negative for Dora and Missi (not performed at Vanka).

A combination of prednisolone (1mg/kg/day/5 to 10 days then CJA), meglumine antimoniate (Glucantime®, 100mg/kg/28d) and allopurinol (Zyloric®, 10mg/kg/bid) is initiated. An improvement in general condition is visible quickly in all 3 patients. A PCR analysis on CSF was performed again for one of the 3 dogs, 8 days after the start of treatment but with discontinuation of corticosteroids for 4 days. The result is negative. At the same time, the cytology of this CSF shows only a very slight granulomatous inflammation with a clear decrease in cellularity. (Appendix 1).

The first case of Leishmanian meningitis in dogs was published in 1996, since then the cases described are few and mainly concern Brazil with Leishmania chagasi. Less than 5% of Leishmanian dogs are found to have neurological disorders. L. chagasi and infantum have been found in the brains of dogs with visceral leishmaniasis. Amastigotes appear to infiltrate mainly via the communicating junctions of the choroid plexuses and the leptomeninge, but also via the fenestrated capillaries of the circumventricular organs that do not possess a blood-brain barrier. However, immunohistochemical analyses do not always show the presence of leishmania in the brain of dogs with neurological signs. Leishmania fragments or usually unsecreted intracellular proteins, which have been demonstrated in recent literature, may be sufficient to elicit a CD3-dominant LT-dominant immune response in CSF. The question remains about the very low leishmania loads detected by PCR: are the variations in the quantities of leishmania detected by PCR related to both the quantity and the quality of the inflammatory cells infiltrating the CSF (lymphocytes versus macrophages)?

Only the analysis of the CSF makes it possible to highlight the presence of leishmania for these three cases. Indeed, cytology points to an inflammatory process and quantitative PCR makes it possible to identify the pathogen. All the results of the additional examinations associated with the very good response to treatment make it possible to direct the diagnosis towards Leishmanian meningitis.

However, studies on a larger number of cases may help to understand the pathogenesis of these sequestered forms. In addition, it would be interesting to conduct comparative studies on dogs that do not live in endemic areas in order to avoid asymptomatic carrying bias.

Appendix 1

	Dora Brittany Spaniel	Border collie Missi	Border collie Vanka
LCS Cytology	Cellularity: 350/μl - Lymphocyte macrophages 72% - GNN 28%	Cellularity: 45/μl - Macrophages 98% - GNN 2%	Cellularity: 3600 /μl - Macrophages 51% - Lymphocytes 47% - GNN 2%
PCR before treatment	LCS: Very Low Load (VLC)	LCS: CTF	LCS: CTF - MO: CTF
PCR after start of treatment		Blood: CTF - CSF: negative - Peripheral NL: negative - MO: negative	LCS to be punctured in the coming days
Response to treatment	Very good	Very good	Very good

Scanelis Study: FIP or Typhus?

AFVAC-FECAVA 2009 and ECVIM 2010 (Delphine Rivière, Scanelis)

Parvovirus and coronavirus are two viruses responsible respectively for feline panleukopenia (commonly known as typhus) and feline infectious peritonitis (FIP) in cats. The purpose of this presentation is to: to show that feline parvovirus should be part of the differential diagnosis of macroscopically citrine yellow abdominal effusions in young cats in the same way as FIP.

Material and method

Quantitative PCR parvovirus analysis is performed on macroscopically citrine yellow abdominal effusions and collected from patients suspected of FIP. Coronavirus tests carried out by the same technique and requested as a first-line treatment by veterinarians were negative or very weakly positive.

Results

Epidemiology: 8-month-old male Burmese sacred, 3-month-old European female, 6-month-old European female, 2-month-old European male.
General symptoms: dejection, digestive disorders (vomiting, diarrhoea sometimes haemorrhagic), hypo- or hyperthermia, nervous disorders (ataxia).
PCR diagnosis: real-time PCR analyses show high loads of parvovirus in the order of 10⁶ to 10⁸ copies of viruses per mL of effusion. At the same time, the amounts of coronavirus are very low (1 in 4 cats) or below the detection limit (111 copies of target sequence per analysis).
PCR analyses also performed on 2 of the cats on rectal swabs confirmed parvovirus, with high parvovirus loads (around 10⁸ to 10¹⁰ copies of viruses by sampling).

Discussion

The macroscopic citrine yellow appearance with or without fibrin of abdominal effusion, associated with a favourable epidemiological context (breeding kitten, certain breeds including the Sacrés de Burma, etc.) is suggestive of a wet form of FIP in the kitten. However, parvovirus (cat typhus) can cause the same clinical signs.
These cases also illustrate the value of real-time PCR compared to conventional PCR. Indeed, a qualitatively positive coronavirus result on abdominal effusion, even in a context of strong clinical suspicion, is not pathognomonic of FIP. The detection of a small amount of virus by the quantitative PCR method makes it possible to cast doubt on the hypothesis of FIP and therefore leads to the search for another cause.

Conclusion

Feline panleukopenia is far from uncommon and should be suspected in animals with unknown or incomplete vaccination status. Its diagnosis is all the more important as the risk of contamination, treatments and prognosis are different for these two diseases.

Scanelis study: in the footsteps of FIP

AFVAC 2008 Conference, Strasbourg (Corine Boucraut-Baralon, Scanelis)

Infection with feline coronaviruses is now widespread in feline communities. So widespread in fact that many breeders wonder about the interest of its screening compared to the real risk - quite low, of seeing cases of Feline Infectious Peritonitis (FIP), insofar as no screening method has a predictive value on the fate of the infection and the cost of this screening is not negligible.
However, this number of FIP cases, even though it represents a small percentage of cats infected with coronaviruses, is constantly increasing and it is still very difficult for a breeder to manage the diagnosis of the disease in one of his kittens, either at home or more often in the weeks or months following the sale.
Cases of FIP will appear in animals that have been infected with a common enteritic coronavirus, often within weeks of birth (mass infection is likely to be an important risk factor). This infection is a necessary but not sufficient factor: stress is also a major factor, as is individual susceptibility (a strongly suspected genetic predisposition). The most frequently put forward scientific hypothesis is that so-called "enteritic" coronaviruses (still found under the name FECV in the Anglo-Saxon literature) with little or no pathogenic disease can undergo genetic modifications that will result in the appearance of highly pathogenic viruses (FIPV), capable of replicating at a high level in monocytes and macrophages and causing the appearance of very characteristic lesions in certain organs.

Pathogenesis of infection

Classically, two biotypes of the virus are distinguished: enteritic coronaviruses, which mainly infect the epithelial cells of the digestive tract, which are minimally pathogenic, or not pathogenic, and pathogenic coronaviruses, which cause systemic infection and replicate mainly in blood monocytes and macrophages.
This dichotomy is quite theoretical, since coronaviruses classified as enteritic are also responsible for systemic infections, even if their replication in monocytes and macrophages is limited.
After a primary infection, a peak of viral shedding is observed in the faeces one week after an infection of SPF (Specific pathogen free) cats via the fecal-oral route (orally administered enteritic strain).
These animals remain asymptomatic or have transient diarrhea.
The level of carriage is higher in kittens than in adult cats and remains high for 2 to 10 months. Then three profiles are observed according to the animals:

persistent carriage (at least 9 months): the virus is excreted in large quantities and almost continuously
intermittent carrying: the level of excretion is lower and there is an alternation between phases of excretion and phases of non-excretion
Transient carriage: animals completely stop shedding the virus after 5 to 19 months, the level of shedding is lower than in the previous two categories.

When these cats are re-infected, if they were shedding the virus at that time, there is no change in the level of shedding and if they were no longer shedding, everything happens as it was during the first infection.
In the cat population studied, no effect of gestation, lactation or corticosteroid treatment on the level of excretion or excretion was observed in no longer excreting cats.
Source: Pedersen NC et al, 2008, Pathogenesis of the feline enteritic coronavirus excretion, Journal of feline Medicine and Surgery, 10, 529-541.
One of the most difficult questions about the pathogenesis of this disease concerns the mechanisms that explain why a virus with no or low pathogenic disease can cause a fatal disease in some animals.
A change in the tropism of the virus has been implicated, probably linked to the appearance of mutations in the genome of the virus. However, some of these mutations, characterized a few years ago, have not been found to be specific to pathogenic strains of the virus and have been demonstrated in viruses hosted by fully asymptomatic animals. More than a radical change in tropism (particular sensitivity of monocytes and macrophages), it is rather a difference in the level of replication of the virus in macrophage cells that is observed. Indeed, the quantities of viruses found in the tissues of sick animals are much higher than those found in healthy animals.
At present, there is therefore no formal proof of the existence of these mutations, although this explanation is the most likely.
Moreover, in parallel with this modification of cellular tropism, the animal's immune response seems to be inadequate since the infected cells are not destroyed by the immune system. Thus, it has been shown that macrophage cells infected with the virus internalize viral antigens in the presence of specific antibodies, and are not recognized as infected by the immune system. This phenomenon seems to be specific to this cell type and could explain at least in part the phenomenon of virus escape from the IS, the long incubation period and the persistence of the infection.
The risk of the disease appearing in an infected cat is quite low and increases with the size of the population in which the animal evolves (between 5 and 15% of infected animals will develop the disease). Other risk factors are stress (also related to the size of the workforce but also surgery, change of ownership, breeding, exposure, etc.) which is a major trigger.
On the other hand, there is no predictive test to detect among infected animals those that will develop the disease. The risk seems to be highest in the 6 months following the primary infection (out of 100 cases of wet FIP, 70% concern animals one year old or less than one year old, source Scanelis).
In addition, the disease itself is not contagious or very minimally contagious.

Screening and Diagnosis: A Complex Problem

Screening (collective or individual) makes it possible to know whether or not a workforce is contaminated by the coronavirus. If the screening reveals an absence of infection (cases of low numbers in particular), it is important for the breeder to maintain this status and therefore to avoid any contact with animals carrying coronavirus but also to alert buyers to the risk incurred by a negative animal that would be put in contact with animals shedding coronavirus. Many cases of FIP have been described in these animals which, after a primary infection with a common enteritic coronavirus and under stress (change of environment for example), will often develop the disease quickly, which may lead to the belief that it is ultimately better to live with coronavirus on its farm than without it.
If the population is contaminated (i.e. in most cases), it is possible to limit the circulation of viruses by grouping animals according to their status. Screening of breeding stock in particular, although expensive, makes it possible to know the individual status of each animal and in particular

isolate highly shedding animals and chronic shedders (and especially avoid contact with very young animals)
select animals that are more resistant to infection (animals that are seronegative or that shed little or no virus even though they live in contact with highly contaminated animals).
This screening appears to be necessary at least in populations where several cases of FIP have been reported (importance of the definitive diagnosis of these FIP cases). Infection pressure is generally high in these communities. In addition, some lineages appear to be genetically predisposed.
In general, screening is also important to manage the introduction of a new animal into the workforce or outdoor matings, which are two risk factors for the introduction of the virus.

Means of screening
Two main types of means are available. They are complementary.
Indirect Methods
Serology can detect the presence of antibodies against coronaviruses of any kind. An infected animal becomes positive within 15 days to 1 month after infection most of the time. Shedding of the virus precedes seroconversion. However, a few cases of lack of long-term seroconversion have been described in infected animals carrying viruses.
In France, many tests are available to veterinarians for this screening:
* Rapid immuno-migration tests (qualitative tests). It should be noted that the name PIF test has been dropped. The only test on the market at the moment (and as of December 2008) is the ®Speed F coronavirus from Biovetotest.
* Immunofluorescence (quantitative tests): different laboratories offer this test but the methods vary depending on the laboratory
* ELISA (quantitative or qualitative tests depending on the lab): here again, the methods differ depending on the laboratory.
It is very difficult, if not impossible, to compare results from different laboratories, especially quantitative results, even if they were obtained with the same technique. Even the positivity thresholds can vary (a titer considered low positive in one lab may be considered negative in another). The results obtained in published studies are also difficult to transpose for the same reasons.
The immunomigration test is slightly less sensitive than laboratory tests and the ELISA method is the most sensitive.
These methods have the advantage of being inexpensive and are useful for assessing infection in a community. However, they do not make it possible to know precisely the excretory status of the animal and in particular to detect chronic shedders, whose serological titers are not always different from some transient carrier cats. Animals that have cleared the virus can remain HIV-positive for several weeks or even months, just as recently infected animals can be HIV-negative. During experimental or natural contamination, it has been shown that some animals do not show seroconversion even though they excrete the virus. Overall, however, there is a correlation between seropositivity and viral shedding. Although there appears to be a correlation between antibody titer and persistence of infection, it is not possible to accurately determine the shedding status of an animal based on an isolated serological result. Some animals with high titers may see their antibody titer drop sharply after isolation, while others, chronically infected, still have high titers after separation. It is therefore difficult to correlate the amount of virus excreted and serological titer, especially in view of the multitude of tests carried out in France. Hence the interest of methods for direct detection of the virus.

Direct Methods
In order to determine the individual status of an animal, it is possible to search directly by PCR (RT-PCR) for the genome of coronaviruses in faeces. This method is very sensitive and therefore detects very small amounts of viruses. Qualitative information is of limited interest on a one-time analysis insofar as many animals are positive, some can excrete very large quantities of virus (up to 1016 viral particles in a rectal swab) and others very little. To confirm that a cat that was previously detected positive is no longer shedding virus, it is necessary to obtain several negative results over a few weeks. To assess the status of chronic shedder, it is therefore necessary to repeat the analyses over several months, which is difficult to achieve in practice given the cost. Quantitative methods have been developed (real-time RT-PCR), allowing to determine the level of excretion of the virus in the faeces. Several studies have shown a correlation between the viral load excreted and the frequency of shedding, with chronic shedding animals also being those that excrete the largest amounts of virus continuously. Epidemiologically, these animals are therefore particularly dangerous. In a herd, chronic shedders often make up a very small percentage of the animals, so it is quite simple once they have been detected to isolate them and exclude them from breeding. To determine the chronic shedding status of an adult cat, it is recommended to test the animal once and if the viral load is high, to test it again 1 to 3 months later. If the burden has not changed, it is likely that the animal is a chronic shedder. These analyses are carried out by real-time PCR.
On the other hand, performing quantitative PCR analyses in very young animals (in which the primary infection is recent) will not necessarily provide very relevant information because in these animals, viral loads during primary infection are higher than those found in adults and they can evolve rapidly. A very high viral load at 2 or 3 months of age does not predict the onset of the disease, or even the future status of chronic carrier.
The method of detecting antigens on biopsies or organs is reserved for the diagnosis of the disease and is still little used in France.

Scouting practice and breeding management

Husbandry and preventive measures are obviously very important. Screening is only one tool that makes it possible to make decisions and implement the most appropriate measures for the epidemiological situation of a farm, but also to verify the effectiveness of these measures.
For example, the practice of early weaning gives quite contradictory results depending on the studies. It appears that early weaning is most effective in small numbers (less than 6 cats) and when infectious pressure is low. Thus, early weaning measures that may be very effective in one kennel will not necessarily be effective in another (according to some studies, in a highly infected environment, kittens can become contaminated at 2 weeks, especially if they are born to chronic surrogate mothers). It is therefore interesting to check the effectiveness of early weaning by carrying out serological tests (at 3 months the kittens must be seronegative) and possibly RT-PCR tests.
Knowing the status of your farm with regard to the coronavirus (regular serologies, quantitative RT-PCRs) can therefore be useful for prioritizing. In highly contaminated populations, and especially if cases of FIP have been diagnosed, it can be very effective to separate or even remove the few chronically shedding animals (generally 10-15% of the population) from the farm in order to limit the infectious pressure. If the level of contamination is low overall, it is necessary to monitor new introductions in particular.
Tests are of great interest for the introduction of a new animal into a workforce. It should be isolated for a minimum of 1 month and two serological tests should be carried out at the beginning and end of quarantine to ensure the negative status of the animal. If the breeding is negative, it is important to check for viral shedding by RT-PCR. If a positive animal is to be introduced into an already contaminated population, the level of shedding of that animal must be assessed in order to assess the risk of introduction. This is difficult in practice for animals introduced at a very young age.
Testing for viruses in the blood by RT-PCR is of no interest in screening. Indeed, many animals carrying coronavirus will be negative in the blood, especially adults. In addition, a positive result is not a predictor of the onset of the disease.

Benefits of serological and virological tests for the diagnosis of FIP

As these screening tests are not specific to FIPV, it is not recommended to use them as a first-line treatment. However, when the clinical, haematological and biochemical elements are strongly in favour of FIP or when histopathological examination does not allow to conclude with certainty, RT-PCR can provide relevant additional information (in particular search for the virus in certain organs such as liver or kidney or in effusion fluids).
Serology is very uninformative because of the risk of false positives linked to infection with benign enteritic coronaviruses (in the case of cats living in groups in particular) and the risk of false negatives (in wet forms in particular). Only very high serological titres in private cats can be considered to have diagnostic value. The value of RT-PCR is quite controversial because this test is considered too sensitive and the cause of many false positives. The studies carried out are difficult to compare because they use different methods, and give rather contradictory results.
It appears that once again, quantitative RT-PCR is more interesting than conventional RT-PCR since it allows for a more detailed interpretation of the result.
In wet forms, the presence of large quantities of virus in the effusion fluid has a good diagnostic value, while a low viral load can be found in certain inflammatory pathologies such as pancreatitis, certain cholangiohepatitis, or certain forms of panleukopenia.
In "localized" dry forms such as nerve or ocular forms, the virus will be preferentially tested in the CSF or aqueous humor.
In the other dry forms, the search for the virus in the whole blood collected at the time of hyperthermia peaks, especially if it is combined with a quantitative search in the faeces, is very informative. Even though small amounts of virus can be found in the blood of some animals carrying coronavirus, particularly at the time of primary infection, viremia is more important during FIP. In addition, in the first case, the fecal loads are extremely high, which is not the case with PIF.
In a recent study carried out at Utrecht University, the sensitivity of RT-PCR on blood cells was assessed at 93% (reference method: anatomo-pathological examination). In a study of 35 cases confirmed by the anatomo-pathological examination performed at Scanelis, the sensitivity was 94%. Several studies on large populations of animals that are asymptomatic or with FIP have shown that the diagnostic specificity of the test is greater than or equal to 94% depending on the test used.
Kidney or liver biopsies can also be tested for the virus, as a significant viral load is found in these organs in animals developing FIP. On these organs, antigen testing is considered a reference method because it is very specific (but not very sensitive).
PCR can therefore be a very interesting tool to confirm a diagnosis of FIP, but its use does not exclude a rigorous diagnostic approach, which can often make it possible to exclude FIP without the need for this additional examination. It is also important to use perfectly validated RT-PCR tests to ensure that the specificity and sensitivity are optimal on the samples analyzed, as these parameters vary greatly with the test and the volume of samples analyzed.
Screening for coronavirus infection is a difficult topic. Which tool, when and how often are recurring questions that are not easy to answer unequivocally. The epidemiological context (size of the farm, cases of FIP, possibility of separating animals) but also the evaluation of the cost-benefit ratio, which is very difficult to assess, are all factors that come into play in the prevention strategy of FIP.
This prevention involves first and foremost the search for and isolation of chronic carriers of the virus, who are few in number, who play a major role in the transmission of infection to young people and therefore constitute an important risk factor for the appearance of FIP cases. The quantitative RT-PCR method makes it possible to search for these chronic carriers with less cumbersome protocols than conventional RT-PCR.
But analyses are not everything, they must be accompanied by measures to limit the non-animal source of virus, which is essentially made up of bedding, which must be plentiful, cleaned and disinfected very regularly.

Some recent bibliographical references to find out more

+ info on the diagnosis of feline corona virus by real-time RT-PCR

Scanelis Study: Diagnosis of Aleutian Disease in Ferrets by Real-Time PCR

AFVAC-FECAVA 2009 Short Paper (S. LAFON, Scanelis)

Introduction

Aleutian disease is caused by a parvovirus, the Aleutian Disease Virus (ADV).
In ferrets, ADV infection causes the formation of immune complexes that can cause multiple organ failures expressed by chronic debilitating disease and/or neurological signs. The signs are therefore non-specific to the disease.
According to the literature, the infection can also be asymptomatic.
To meet the growing demand for diagnosis of ADV infection in ferrets, the Scanelis laboratory has developed a real-time PCR test for the direct detection of the pathogen.

Materials and Methods

A study was carried out on samples taken from 179 ferrets received at the Scanelis laboratory (blood, CSF, organs and rectal swabs).
The following were tested by ADV real-time PCR:
89 sick ferrets, suspected of various infectious diseases (distemper, toxoplasmosis, neosporosis, mycobacteriosis), for which the corresponding tests were negative,
28 ferrets with distemper (diagnosis by real-time PCR, Scanelis), including 6 with neurological disorders
54 ferrets without clinical memorials,
13 healthy ferrets.

Results

Sixteen of the 89 ferrets with clinical signs and negative in the tests initially ordered were positive by ADV PCR: 8 had neurological symptoms and 5 had general signs (3 without information).
The following were negative on the ADV real-time PCR test:
32 ferrets with neurological disorders (and negative in the tests initially requested),
the 28 ferrets with distemper,
the 54 ferrets of unknown clinical status,
the 13 asymptomatic ferrets.

Discussion

Eight of the 46 ferrets with neurological signs were positive on the ADV PCR test, which allows the prevalence and role of this pathogen in these pathologies to be estimated.
Aleutian disease should therefore be part of the differential diagnosis of neurologically predominantly neurological conditions in ferrets.

In 28 ferrets with distemper, no co-infection with ADV was detected.

No ADV infection was detected in healthy animals or animals of unknown clinical status. Thus, the data obtained during this study did not confirm that ferrets can be infected with ADV without expressing symptoms suggestive of Aleutian disease.
The detection limit of this test has been assessed according to the recommendations of the European Pharmacopoeia. The sensitivity of the test is very good and is therefore not called into question by the absence of virus detection in asymptomatic ferrets, in which, if asymptomatic carriage exists, the presence of very low viral loads can be assumed.

However, these healthy ferrets could be a source of infection in the community. It is therefore necessary to continue these investigations on a greater number of asymptomatic animals.
If healthy carriage is eventually found, the comparison of viral loads detected in animals with and without clinical signs should highlight the importance of using a quantitative technique, such as real-time PCR, for interpretation of the result and development of the diagnosis.

Scanelis Study: Feline Respiratory and Eye Diseases

In order to study the Prevalence of infection of infectious agents classically implicated in oculorespiratory disorders in cats, Scanelis conducted a study during the winter of 2009-2010.
Systematic analyses were carried out on oropharyngeal or ocular samples from 98 cats : feline herpesvirus, feline calicivirus, Chlamydophila felis, Bordetella bronchiseptica and influenza H1N1 virus were tested by real-time PCR in cats with acute eye and/or respiratory disorders.

The results of this study were presented at the GEMI annual congress held in Avignon in April 2010:

Prevalence of Feline Herpesvirus Infection, Feline Calicivirus, Chlamydophila felis, Bordetella bronchiseptica and the H1N1 influenza virus in cats with acute eye and/or respiratory disorders during the winter period 2009-2010 in France

C. Boucraut-Baralon, D. Rivière, S. Lafon, M. Etiévant and J.L. Pingret

In order to study the prevalence of infection of infectious agents classically involved in oculo-respiratory disorders in cats, systematic analyses were carried out on oropharyngeal or ocular samples from 98 cats during the period from 10 November 2009 to 15 January 2010.
The choice of this period was justified by the fact that it corresponded in France to the epidemic peak of H1N1 influenza in humans, which made it possible to study in parallel the frequency of infection in the feline species whose sensitivity to the H1N1 influenza virus has been proven (cases described in the USA and Europe).

Materials and Methods

98 samples taken from cats with acute respiratory and/or ocular signs (less than 15 days of evolution) were analyzed by PCR or real-time RT-PCR for FHV (feline herpesvirus), FCV (feline calicivirus), Chlamydophila felis, Bordetella bronchiseptica and the H1N1 flu virus.

Results

65% of cats with acute ocular and/or respiratory signs are carriers of at least one infectious agent among those sought. 30% of cats are carriers of FHV, 30% are carriers of calicivirus, 20% are carriers of Chlamydophila and 7% of Bordetella.
Co-infections with at least two of the infectious agents sought are found in 30% of cats positive for at least one infectious agent, with the most frequent co-infections being herpes virus calicivirus co-infections (8% of cats positive for at least one infectious agent). Herpesvirus/calicivirus/co-infections Bordetella are found in 3% of cats.
The prevalence of infections varies significantly depending on the nature of the clinical signs: Chlamydophila is not found in animals without ocular signs, but is detected in 25% of cats with acute ocular signs without respiratory signs. Infection with Bordetella bronchiseptica is always associated with the presence of respiratory signs and FHV-FCV co-infections are consistently associated with the presence of both respiratory and ocular signs. Isolated FCV infection is 3 times more common in animals with only respiratory signs compared to animals with only ocular signs.
Finally, none of the cats tested were carriers of the H1N1 virus, which confirms that the prevalence of infection in cats was probably very low during the peak epidemic in humans.

Discussion

This study confirms the importance of viral infections in the etiopathogenesis of acute eye and respiratory disorders in cats. Chlamydophila felis is not involved in the etiology of respiratory disorders, but this bacterium is the most frequently isolated agent in acute eye disorders. Finally Bordetella bronchiseptica However, which is rarely sought in cats, it is detected in 7% of animals (11% of positive animals) in association or not with viral agents. However, it is associated with serious disorders and significant mortality in kittens and should therefore be more frequently sought.

To read about it

Vet Rec. 2005 May 21; 156(21):669-73. Factors associated with upper respiratory tract disease caused by feline herpesvirus, feline calicivirus, Chlamydophila felis and Bordetella bronchiseptica in cats : experience from 218 European catteries.
Helps CR, Lait P, Damhuis A, Björnehammar U, Bolta D, Brovida C, Chabanne L, Egberink H, Ferrand G, Fontbonne A, Pennisi MG, Gruffydd-Jones T, Gunn-Moore D, Hartmann K, Lutz H, Malandain E, Möstl K, Stengel C, Harbour DA, Graat EA. University of Bristol, Bristol BS40 5DU.

A full history of the management practices and the prevalence of upper respiratory tract disease (URTD) at 218 rescue shelters, breeding establishments and private households with five or more cats was recorded. Oropharyngeal and conjunctival swabs and blood samples were taken from 1748 cats. The prevalences of feline herpesvirus (FHV), feline calicivirus (FCV), Chlamydophila felis and Bordetella bronchiseptica were determined by PCR on swab samples. An ELISA was applied to determine the prevalence of antibodies to B. bronchiseptica. The rates of detection by PCR of each pathogen in the cats in catteries with and without ongoing URTD were, respectively, FHV 16 per cent and 8 per cent ; FCV 47 per cent and 29 per cent ; C. felis 10 per cent and 3 per cent ; and B. bronchiseptica 5 per cent and 1.3 per cent ; the seroprevalences of B. bronchiseptica were 61 per cent and 41 per cent, respectively. There was evidence that FHV, FCV and B. bronchiseptica played a role in URTD. The risk factors associated with the disease were less than excellent hygiene, contact with dogs with URTD, and larger numbers of cats in the cattery or household.

The ABCD website (European Advisory Board on Cat Diseases)

Download the sheet “L’infection par Bordetella bronchiseptica”
See the group's recommendations (English version available) at “Bordetella bronchiseptica infection in cats”

Scanelis study: feline uveitis of viral origin

Feline uveitis: prevalence of infection by coronaviruses, retroviruses, feline Herpesvirus and Toxoplasma gondii in aqueous humor samples assessed from 91 cases.
C. BOUCRAUT-BARALON, D. RIVIERE and S. LAFON - Scanelis

Feline uveitis is common and may be the first or only manifestation of systemic disease. Trauma, with or without bacterial infection, viral or parasitic infections, and neoplasms are among the classic etiologies of feline uveitis. 91 aqueous humor samples have been systematically tested for feline coronaviruses, FeLV, FIV, feline herpes virus and toxoplasma by real-time PCR techniques.

Materials and Methods

91 aqueous humor samples received in the laboratory in 2009 and taken from cats presenting isolated uveitis or associated with extra-ocular clinical signs were analyzed by real-time RT-PCR for coronavirus, FeLV and FIV and by PCR For Toxoplasma gondii and feline herpes virus. For positive samples, quantification of infectious agents was carried out.

Results

No samples were positive for Toxoplasma, three samples were positive for feline herpes virus, one of which was also positive for coronavirus. The very low viral loads detected do not allow us to conclude with certainty about a possible involvement of FHV in the development of the clinical signs observed.
On the other hand, the coronavirus and FeLV were found at a higher frequency. The coronavirus is the most frequently detected infectious agent (a third of samples) with most often high viral loads suggesting an involvement of viral replication in the pathogenesis of uveitis. The average age of cats with coronavirus uveitis is 16 months compared to 47 months for negative cats and 39 months for all animals included in the study. Two thirds of cats affected by coronavirus uveitis are less than 1 year old.
For coronavirus-positive samples, general signs were associated with uveitis in the majority of cases, hyperthermia being the most common. Nervous, digestive or respiratory signs as well as effusive forms are also observed. When extraocular signs were described, other available samples were analyzed in parallel (blood, effusion, CSF) and also proved positive.

Discussion

If uveitis is a sign classically described in feline infectious peritonitis, no data is currently available in the literature concerning the benefit of searching for the virus directly in the aqueous humor. It appears that among the infectious or parasitic diseases classically involved in the pathogenesis of feline uveitis, FIP is the most common, particularly in young cats, and that it can be associated with other infections (FeLV, FIV, FHV). Although coronavirus uveitis is most often associated with general signs and viremia, there are however certain cases of isolated coronavirus uveitis without waiting for the general condition, in the absence of viremia and for which the Implementation of appropriate glucocorticoid treatment allows for long-term remissions.

Conclusion

Feline Coronavirus and FeLV infections are the most common causes of viral uveitis in cats. The detection of FHV in the aqueous humor is rare and the role of this agent remains to be clarified. Finally, the prevalence of Toxoplasma gondii evaluated from the search for the parasite by PCR directly in the aqueous humor is very low. In our experience, the few confirmed cases (2 cases out of more than 300 aqueous humors analyzed over the last ten years) were associated with parasitemia in a context of generalized toxoplasmosis.

Communication presented at the AFVAC 2010 conference

Scanelis, because PCR is a matter for specialists

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