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The Veterinary Journal

The Veterinary Journal 175 (2008) 1415 www.elsevier.com/locate/tvjl

Guest Editorial

Canine leishmaniasis: A diagnostic and clinical challenge

Canine leishmaniasis is a severe zoonotic disease that affects millions of dogs in Europe, Asia, Africa and America (Moreno and Alvar, 2002). The dog is considered the reservoir host for Leishmania infantum in the Old World and for its New World synonym Leishmania chagasi. Dogs are infected by Leishmania promastigotes deposited in the skin during the bites of infected female sandy vectors. The promastigotes invade host cells and replicate as intracellular amastigotes. The disease incubation period prior to the appearance of clinical signs may last months to years, during which the parasite disseminates in the body (Oliva et al., 2006). Some dogs develop clinical disease, whereas others remain asymptomatic carriers that are infectious to sandies and may thus transmit disease to other dogs or humans. The clinical signs of canine leishmaniasis are variable. While most symptomatic dogs exhibit dermal abnormalities, some may present other signs in conjunction with or in the absence of skin lesions. These include signs of kidney disease, expistaxis, ocular or musculoskeletal abnormalities. It is presumed that the majority of dogs in highly endemic areas where the transmission of Leishmania is extensive are exposed to infection. Population studies in Leishmania-endemic areas have shown that a proportion of the canine population develops a symptomatic disease, another fraction has persistent asymptomatic infection, while yet another fraction is resistant to the infection or intermittently resolves it without developing clinical signs. Leishmaniasis is a good example of a disease in which infection does not equal clinical illness. This makes canine leishmaniasis a diagnostic challenge for the veterinary practitioner, clinical pathologist and public health ocial in endemic countries as well as non-endemic regions where imported infection is a concern. The publication of a review on biotechnological advances in the diagnosis of canine visceral leishmaniasis in this issue of The Veterinary Journal (Gomes et al., 2008) is a timely opportunity to discuss the complex issue of diagnostic options in canine leishmaniasis. The laboratory assays for the diagnosis of canine leishmanaisis include aspirations and biopsies for cytological and hisopathological detection of the parasites presence in tissues, serologi1090-0233/$ - see front matter 2006 Elsevier Ltd. All rights reserved. doi:10.1016/j.tvjl.2006.11.011

cal tests for detection of serum anti-leishmanial antibodies or antibodies to specic antigens (Scalone et al., 2002), culture of the parasite from tissue samples, or the polymerase chain reaction (PCR) demonstrating presence of Leishmania DNA in tissue or blood samples (Gomes et al., 2008). Each of these diagnostic techniques has advantages and shortcomings, and the diagnostic methods should be selected in light of these considerations. Cytological or histological identication of Leishmania amastigotes, free or contained within macrophages, in stained specimens from lymph nodes, spleen, skin, bone marrow or other organs can provide a potentially quick diagnosis of infection. However, these are neither sensitive nor specic assays as dogs with overt clinical disease may have few or no demonstrable tissue parasites and because other objects viewed by light microscopy can be erroneously considered amastigotes. Detection of serum antileishmanial antibodies is indicative of exposure to infection. Low, possibly transient antibody levels are detected in dogs that have been exposed, but have not developed clinical disease. Dogs with persistent infections usually develop gradually increasing antibody titers over time. High antibody levels are usually associated with symptomatic disease or persistent asymptomatic infection in carrier dogs. Serological cross reactivity with dierent pathogens is possible with some serological tests, especially those based on whole parasite antigens. Cross reactivity with Trypanosome species should be suspected in regions of South and Central America endemic to Trypanosoma cruzi and in other areas where trypanosomes infect dogs. The demonstration of leishmanial DNA in fresh or formalin-xed tissues or blood of infected animals by PCR is a sensitive method (Roura et al., 1999; SolanoGallego et al., 2001; Lachaud et al., 2002). Non-invasive conjunctival PCR has been shown to be helpful in the direct diagnosis of canine infection (Strauss-Ayali et al., 2004). Several PCR protocols are used in dierent laboratories with a number of target gene sequences. The small-subunit (SSU) rRNA gene, the internal transcribed spacer (ITS) of the ribosomal operon and the high copy number sequences of kinetoplast DNA (kDNA) are frequently used as PCR targets (Lachaud et al., 2002). Real

Guest Editorial / The Veterinary Journal 175 (2008) 1415

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time PCR is an additional sensitive diagnostic tool, which is able to quantify the number of parasites in a sample and to detect a minute number of parasite DNA molecules in the tissue (Francino et al., 2006). Studies evaluating PCR from dierent tissues of infected dogs have shown variable and sometimes conicting results. These have often depended on the sampling technique, storage method and PCR protocol employed. Some organs from the same dog may be positive whereas other tissues will be negative (Solano-Gallego et al., 2001). Blood is easy to sample and submit for testing, but is not the best source for leishmanial DNA with some protocols, because this pathogen is not as abundant in blood as in the skin and lymphatics (Strauss-Ayali et al., 2004). Epidemiological studies have shown that dogs can be seronegative for Leishmania while being PCR positive (Oliva et al., 2006), or PCR negative while seropositive (Solano-Gallego et al., 2001). These conicting results are usually obtained from asymptomatic dogs that have been exposed to Leishmania, but at the time of sampling it is not known whether they would develop a persistent infection. Therefore, it is recommended to continue monitoring dogs with no clinical signs of disease and inconclusive infection status, with repeat testing in 3 months time. Symptomatic dogs or those progressing toward symptomatic infection will usually have both positive serology and PCR. A single negative PCR result in a suspected dog with compatible clinical signs is not enough to rule out infection. When the index of suspicion is high, evaluation of additional tissues by PCR is recommended. How should dogs be monitored during and after medical treatment of leishmaniasis? Dogs with symptomatic disease treated with standard drug protocols often resolve their clinical disease, but remain seropositive and parasitologically positive. Clinical illness may recur due to the incomplete elimination of parasites by therapy. None of the currently used anti-leishmanial drugs have been proven to reliably induce parasitological cure in dogs, and repeated treatments are often warranted in order to maintain a clinical remission (Baneth and Shaw, 2002; Noli and Auxilia, 2005). In addition, treated dogs living in endemic areas are constantly exposed to the parasite and may become re-infected. Studies evaluating infectiousness to sandies have indicated that dogs after treatment can be infectious to sandies and may thus serve as a source of infection to other dogs and humans (Alvar et al., 1994). In conclusion, the diagnosis of canine leishmanaisis remains a challenge despite the development of new molecular diagnostic tools and advanced serological tests. More studies are needed to correlate between evidence of natural exposure to the parasite and the prospect of developing a persistent infection. Simple and standard assays should be used for monitoring dogs in endemic areas or dogs visiting these areas to verify if infection has occurred and a

clinical disease may develop. Finally, additional research is warranted to determine how to monitor dogs under treatment and what test results indicate a stable, diseasefree remission or elimination of infection. Gad Baneth Itamar Aroch School of Veterinary Medicine, Hebrew University, P.O. Box 12, Rehovot 76100, Israel E-mail address: baneth@agri.huji.ac.il (G. Baneth)

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