Leishmaniosis

Leishmanioses are a group of zoonotic diseases transmitted to humans and animals by the bite of phlebotomine sand flies. The diseases are named after Sir William Boog Leishman, Director General Army Medical Services 1923-1926. Leishmanioses are caused by intracellular parasitic protozoans. Worldwide, they are one of the most important vector-borne diseases.

Two types of leishmanioses can broadly be distinguished: 1. Zoonotic leishmanioses, in which the reservoir hosts are wild animals, commensals or domestic animals and 2. Anthroponotic leishmanioses, in which the reservoir is man. Clinical manifestation of the disease depends on the type of pathogen and thus the type of leishmaniosis. It can be a life-threatening systemic infection (visceral form), go along with chronic skin sores (cutaneous form), or show dreaded metastatic complications, causing facial disfigurement (mucocutaneous form).

In the veterinary field of all domestic animals the dog is the main reservoir for Leishmania infantum that causes a visceral human leishmaniosis form. The dog is also the domestic animal most severely affected by the parasite itself. In countries endemic for leishmaniosis, veterinary treatment is often not affordable, whereas in areas as the Mediterranean basin diseased dogs represent a typical veterinary patient.

Regarding the disease in humans, an estimated number of over 1 billion people are living in endemic areas at risk of infection according to the WHO, with 600,000 to 1 million new cases of cutaneous and 50,000 to 90,000 new cases of visceral leishmaniosis annually. Urbanization due to ecological, demographic and environmental changes appears to be one of the major worldwide risk factors for leishmaniosis and largely contributes to the persistence of the burden of the disease, especially in anthroponotic foci.

Pathogens

The genus Leishmania is divided into two subgenera on the basis of the development in sand flies, namely Leishmania and Viannia.

Most of the Leishmania species annotated to date were originally described on the basis of clinical, epidemiological and biological features. The subgenus Leishmania includes besides others medically important species of the Old World species (e.g., L. tropica, L. aethiopica, L. major, L. infantum and L. donovani) and medically important species of the mexicana group restricted to the New World (e.g., L. mexicana, L. amazonensis, L. venezuelensis). The subgenus Viannia is found only in Central and South America. Most important species are L. braziliensis, L. guyanensis, L. panamensis and L. peruviana all of which cause human disease.

The veterinary important species are Leishmania infantum in the Mediterranean area and Leishmania chagasi, meanwhile found identical with L. infantum, in Latin America.

Leishmania infantum, the infectious agent of canine leishmaniosis (CanL), is an obligate heteroxeneous parasite, i.e. it needs two hosts to develop, one vertebrate host and one terminal insect host. In the vertebrate host the parasite is found intracellular as so-called amastigote. In the insect host L. infantum is found extracellular in the gut as promastigote.

The amastigote shows a round to oval body, about 1.5 to 3 x 3 to 6.5 µm in size, depending on the species. It possesses a single nucleus and a rod-shaped kinetoplast. There is no free flagellum, but a rudimentary one is present. The reproduction of this form is by longitudinal binary fission. Amastigotes can be cultured in appropriate cell cultures.

The promastigote varies from 16 to 40 µm length and 1.5 to 3 µm width. Promastigotes are longer than amastigotes, with a central nucleus and anterior kinetoplast and a well-developed flagellum, which is used either for propulsion or for attachment.
 

Promastigotes may be cultured in various media, mostly containing defibrinated blood and inactivated serum.

Epidemiology

Canine leishmaniosis (CanL)

Canine leishmaniosis is endemic in more than 70 countries in the world. It can be found in regions of southern Europe, Africa, Asia, South and Central America and has recently emerged in the USA. Canine leishmaniosis is also an important concern in non-endemic countries where the imported disease may constitute a veterinary and public health problem.

Seroprevalence rates found in studies carried out in the Mediterranean basin range between 10% and 37%, and 70% by using the PCR method for detecting leishmanial DNA. Based on the seroprevalence data it has been estimated that 2.5 million dogs in France, Italy, Portugal and Spain are infected. The number of infected dogs in south America is also estimated in millions with high infection rates in some areas of Brazil and Venezuela.

Canine leishmaniosis (CanL) is caused by the protozoan Leishmania infantum. Dogs are the most important reservoir of the pathogen and are mainly responsible for the persistence of CanL and also human visceral leishmaniosis in the Palaearctic and Neotropical regions. Other hosts have been found naturally infected (foxes, cats, rodents, horses), or serologically positive, but it is unlikely that they can act as effective reservoirs. The dog is such an excellent reservoir for L. infantum for three reasons: 1) a very long pre-patent period of infection, 2) a high concentration of protozoan amastigotes in the skin and 3) a high percentage of relapses together with uncertain parasitological sterilization after treatment.

More than 50% of dogs with proven established infections are apparently healthy on clinical diagnosis (asymptomatic). They are either progressing towards overt disease (pre-patent cases), remaining symptomless for prolonged periods (even for life), or exhibit spontaneous healing; the latter groups are regarded as resistant.

The long pre-patent period of infection produces a high percentage of asymptomatic dogs; however, they are able to infect the sand fly vector. Furthermore, depending on the type of cellular immune response, dogs may not show any clinical signs or develop only cutaneous nodules at the site of inoculation. Nevertheless, the ability to infect sand flies has been observed to be similar in both asymptomatic carriers and animals with different degrees of disease, although it was shown that the rate of infected sand flies increased with the appearance and severity of the clinical signs. About half of the seropositive dogs identified in field studies are asymptomatic. In a conclusion, apart from untreated diseased dogs and free-ranging ones, asymptomatic dogs are one of the most important reservoirs of CanL: they are able to infect sand flies and they might remain asymptomatic for years and therefore may not be recognized by pet owners and veterinarians (if diagnostic tests fail).

 

Human visceral leishmaniosis (VL)

Although transmission of Leishmania infantum is the predominant cause of human VL in the Mediterranean basin, infection with L. donovani, is responsible for a large part of fatalities in humans. Worldwide, there are approximately 500,000 new cases of visceral leishmaniosis annually.

Human VL is concentrated in Eastern Africa (particularly Sudan and Kenya), and on the Indian subcontinent (Bangladesh, North-east India and Nepal). Infantile VL is found in the Mediterranean basin, extending east through South-west Asia to China, and west to Central and South America. Particularly in North-east Brazil more than 90% of all the cases in Latin America are found. The infantile type has been recognized to expand to infect immunocompromised adults either suffering from HIV infection or any other type of immunosuppression.

Human CL in the Old World is mainly found in arid or semi-arid areas. The anthroponotic type is usually found as epidemics in densely populated cities of central and west Asia. The zoonotic form is characteristic of semidesert areas in Asia and North Africa, associated with colonies of reservoir hosts.

Urbanization due to ecological, demographic and environmental changes appears to be one of the major worldwide risk factors for leishmaniosis and largely contributes to the persistence of the burden of the disease, especially in anthroponotic foci. It affects all four eco-epidemiological entities (cutaneous vs. visceral, anthroponotic vs. zoonotic).

 

Prevalence of canine leishmaniosis

Generally it can be distinguished between stable and unstable endemic conditions of CanL.

Stable CanL is viewed as an endemic situation in which dog-to-dog transmission of Leishmania and therefore the appearance of new cases invariably occurs every season.

In Europe stable transmission is usually from May-June to September-October and associated with optimal ecological conditions: rural and peri-urban environment; hilly territory, commonly with southerly exposure; altitude from 0 to 6-900m above sea level and a lack of extreme conditions (strong winds, absence of vegetation etc.).

Unstable is defined as an endemic condition in which autochthonous cases, usually in low numbers, occur sporadically or periodically, but not every year. It is generally found at the border of a stable endemic distribution. For prevalence studies serology is supposed to remain the method of choice for large-scale screening of dogs. Using this method not all infections are detected so that seroprevalence studies only represent a fraction of all infected animals. Prevalence rates are not uniform in any of the territories investigated. Large variations have been observed in dog populations a few kilometres apart, a situation that is presumed to be caused by strict ecological requirements for Leishmania transmission.

 

Infections in non-endemic countries

Numerous case reports and personal communications about Leishmania infected dogs exist in North America and Northern Europe. In Germany, a total of 20,000 infected dogs is calculated by third party organizations. These are mainly imported animals, but also a lot of dogs travelling with their owners to endemic areas.

Autochthonous cases of CanL in non-endemic countries have been described in Switzerland, Germany, England, Belgium, Netherlands and in the USA. Cases with no travel history or any source of infection and vector identification in the USA have been reported from the following states: Texas, Ohio, Oklahoma and Maryland.

In the USA the emergence of visceral leishmaniosis in a relatively large and widespread population of foxhounds was reported during the last 28 years. Even though sand fly species are present in most of the respective areas, vector transmission has not been demonstrated and is furthermore not supported by the fact that solely foxhounds in distinct kennels are infected. Starting with single foxhounds in 1980, nowadays several thousand dogs in foxhound hunt clubs in some 21 US states and two Canadian provinces have been detected to be infected. In some of the respective areas transmitting vectors are even lacking. Non-vectorial transmission (see below) and a possible unique susceptibility of foxhounds and/or subtle behaviour that maximizes the possibility of exchange of blood and secretions with other foxhounds are suggested. Other potential sources of transmission as congenital transmission and direct transmission by contact or reuse of needles have also been considered. Nevertheless the epidemiology of canine visceral leishmaniosis in the USA remains unclear. Besides the foxhounds, sporadic cases in dogs other than foxhounds have been reported in the USA.

Transmission

Developmental cycle

Leishmania infantum, the infectious agent of canine leishmaniosis (CanL), is an obligate heteroxeneous parasite, i.e. it needs two hosts to develop, one vertebrate host and one terminal insect host. In the vertebrate host the parasite is found intracellular as so-called amastigote. In the insect host L. infantum is found extracellular in the gut as promastigote.

Amastigotes are found in reticuloendothelial cells. They multiply by asexual binary fission forming 'cell nests'. After a rupture of the host cell the progeny is taken up by other local or circulating macrophages. Depending on the parasite species the amastigotes either remain in the superficial tissues, continuing their reproductive cycle or settle in macrophages in the deep organs of the reticuloendothelial system such as lymph nodes, bone marrow, spleen and liver. One single bite might be enough to cause an infection of the vertebrate host.

Amastigotes in macrophages from the skin or peripheral blood circulation are taken up when the female sand fly feeds on an infected host. Inside the insect host a conversion to promastigotes takes place in the midgut. Multiplication and translocation to the oesophagus follow. From there the promastigotes are transmitted to a vertebrate host during feeding of the sand fly.

Temperature and the susceptibility of the invertebrate host determine the cycle within the sand fly. A minimum of 10°C is necessary for initiation of the invertebrate cycle and rising temperature is continuously shortening the time needed for completion of the insect phase, with at least 6 days for L. infantum.

 

Non-vectorial transmission

Slappendel and Teske (1999) judge the uptake of amastigotes by an intact gastrointestinal tract as questionable, but presume that it seems quite possible that macrophages may pick up ingested amastigotes via a mucosal lesion or a bite wound. They propose that transmission from dog to dog in the absence of sand flies is not merely a theoretical possibility and may occur more often than recognized in endemic regions, but the possibility of humans being directly infected by dogs is according to them virtually negligible, provided immunological resistance is normal. They advise contact of infected dogs with babies, toddlers, and HIV or otherwise immunodeficient people to be avoided.

  • From mother to offspring:

Transmission of infection from a bitch to a puppy has been described or presumed. In man, diaplacental transmission in a child from Sudan, respectively intrauterine transmission has been reported as well.

  • Via infected blood:

Transmission via blood transfusion has been reported. Furthermore transmission via contaminated needles between drug addicts has been observed.

  • Via other body secretions:

In canine semen and urine as well as in human urine samples leishmanian parasites have been isolated. In human saliva and urine of patients with the Indian type of visceral leishmaniosis (kala azar), amastigotes of Leishmania have been detected.

  • Via other insects:

Mechanical contamination by biting flies has been documented, but there is no evidence for or against a spreading of the disease through this type of transmission. Ticks have been suspected of transmitting leishmaniosis but, though their gastrointestinal tract may harbour infectious promastigotes, their role in transmitting the disease is believed doubtful due to their life cycle.

Pathogenesis

In natural conditions, low numbers of promastigotes (100-1000) are transmitted by sand flies, which are sufficient to induce infection in susceptible dogs. Interestingly, infected sand flies are known to require more blood feedings from different animals. As a consequence, the sand fly sucks less blood from a single vertebrate and therefore attacks additional mammals, spreading the Leishmania parasites to a wider range of hosts.

Most parasites are killed by complement factors of the vertebrate host, but a few survive, using different strategies. The promastigotes adhere to resident or recruited cells of the monocyte / macrophage lineage. Complement-dependent adhesion is followed by internalization through phagocytosis. Inside the phagolysosomes, the promastigotes transform into non-motile amastigotes, accompanied by a rapid drop in pH to ~5.

In a susceptible host, dissemination from the skin to lymph nodes, spleen and bone marrow takes place within the first few hours. In resistant animals, parasites remain localized in the skin or at maximum in the local lymph node.

 

Immunology

Protective immune response requires the presentation of appropriate antigens by antigen-presenting cells (APCs; macrophages, dendritic and Langerhans' cells most important in leishmaniosis), the induction and expansion CD4+ Th1-lymphocytes, and the activation of macrophages for efficient parasite killing. Superoxide (O2-) and nitric oxide (NO) are the two major effector mechanisms for eliminating Leishmania parasites.

The type of immune response (Th1 or Th2) is determined mainly by the cytokine milieu that the T-cells face when they first encounter the antigen, with IL-4 being most important. Early up-regulation of IL-4 is seen in susceptible animals subsequently inducing IL-12 unresponsiveness. Overall, the central decisive element in the development of susceptibility or resistance after Leishmania infection appears whether CD4+ T-cells remain responsive to IL-12 by remaining expression of the IL-12 receptor for more than 48 hours following infection. In susceptible dogs during the course of infection, T-lymphocyte regions become cell-depleted and B-cell regions proliferate.
Potential hazard has further been observed by large amounts of circulating immune complexes (CICs). Autoantibodies against erythrocytes, thrombocytes and nuclear proteins have also been documented.

With macrophages as main cellular type for promastigotes the parasite has chosen a cell type which normally is playing an important role in the induction as well as expression of an immune response. How the parasites are actually managing this paradoxon is not completely clarified.

After different periods of incubation (2-12 months) dogs may either develop clinical signs that are often fatal or they might develop resistance to infection. The latter was found to be associated with a strong parasite-specific cellular immune response and the production of cytokines such as IL-2 and TNF. Dogs may furthermore develop transient, self-limiting infections. They may also spontaneously convert from seropositive infected to negative dogs or do not develop antibodies at all.

Immune responses to Leishmania may result in a polarization of T-cell activity towards a distinct helper T (Th) phenotype. The cytokines produced may activate effector mechanisms that can result in either protective immunity or exacerbation of the disease (Pinelli et al., 1999). Infected macrophages rely mainly on nitric oxide production as an innate mechanism of killing Leishmania. The parasite inhibits this mechanism and is able to multiply in the parasitophorous vacuole. Eventually infected macrophages rupture, and amastigotes are taken up by new phagocytic cells. Macrophages and dendritic cells present Leishmania antigens to T-cells, and either an effective cellular immune response results (a Th1 pattern) or an ineffective humoral response occurs (Th2 pattern). Although several factors influence the development of Th-cell subsets, the most important one may be their early exposure to cytokines (Pinelli et al., 1999).

Diagnosis

 

Clinical diagnosis 

Clinical diagnosis (including non-specific laboratory parameters) is unreliable for the following reasons:

  • More than 50% of dogs with proven established infections are apparently healthy on clinical diagnosis (asymptomatic). They are either progressing towards overt disease (pre-patent cases), remaining symptomless for prolonged periods (even for life), or healing spontaneously, the latter two groups regarded as resistant.
  • When present, clinical signs can be variable and mimic those caused by other diseases.
  • Atypical forms are reported in dogs.

 

For final diagnosis three types of tests are available:

  • parasitological methods aiming at the detection of parasites
  • serological methods with detection of anti-leishmanial antibodies in the blood
  • molecularbiological techniques like the polymerase-chain reaction (PCR), detecting the parasite DNA in host tissue after multiplication.

 

No consensus on the relative diagnostic efficacies of the different techniques exists, particularly with regard to specificity and sensitivity. Parameters may vary according to materials used (e.g. culture medium, antigen, oligonucleotides) and methods employed (serological technique, PCR protocol). Multi-diagnostic approaches should be followed as no single technique identifies all infected animals from cross-sectional samples.

Comparing the performance of diverse techniques in a longitudinal study, Quinnell et al. (2001) recorded highest sensitivity of PCR shortly after the infection (88%), declining to around 50% in the following months. Conversely, the sensitivity of serology was low (41%) at the beginning of the infection, but high (93-100%) thereafter. PCR thus is better at detecting the early stages of established infections and transient and self-limiting infections, whereas a reliable serological technique is better at detecting advanced stages of infection, both in patent and asymptomatic dogs (although some positive cases may convert to seronegative during the course of infection).

 

Parasitological diagnosis

Parasitological methods possess 100% specificity but are not very sensitive.

  • Smears

Fine needle biopsies are taken from bone marrow or lymph nodes and ideally stained with May-Grünwald-Giemsa stain. Examination is under a x100 oil immersion lens. Parasites (here amastigotes) are small, oval bodies (1.5 to 3 x 3 to 6.5 µm) with dark nucleus and small kinetoplast in vertical position. No correlation between the clinic and the number of Leishmania found in cytological preparations could be found (Denerolle, 1996). Sometimes the parasites can be found in impression smear or fine needle aspirates from skin nodules. The sensibility of the cytology is too low for proper diagnosis: Bone marrow smears are positive in 50 to 70% of infected animals, lymph node smears in about 30%. The competence of the examiner and the time taken for examination are decisive for the success of the method. In bone marrow smears Leishmania are nearly exclusively found in macrophages while in cytological preparations from lymph nodes they are mainly extracellular. Infected macrophages often burst while being smeared.

Microscopic image of a Giemsa stained smear showing Leishmania amastigotes with the cytoplasma of cells.
Leishmania amastigotes within the cytoplasm in a smear stained with Giemsa
  • Culture

Culturing of Leishmania should be performed at 26-28°C in suitable media, e.g. Novy-MacNeal-Nicolle (NMN)-medium. Positive cultures will have promastigotes visible within two weeks.

 

Serological diagnosis 

Seroprevalence, regarded as an intermediate measure between disease and the prevalence of infection, is influenced by the fact that on the one hand serology can reveal a high proportion of asymptomatic carriers which represent half of all seropositive animals; and on the other hand the limitation due to the long periods of serological latency. Antibodies are often detectable only several months after infection. Furthermore, a proportion of seropositive infected dogs spontaneously converts to negative, or does not develop specific antibodies at all (Acedo-Sànchez et al., 1998). Thus a maximum estimated sensitivity of the most efficient technique (IFAT) hardly reaches 80% and only during a few months after exposure to Leishmania in the previous season.

Several techniques can be used for the detection of antileishmanial antibodies. Commercially available are IFAT, Dot-ELISA and DAT (direct agglutination test). They possess high specificity and sensitivity (80-100%) but should not be used solely as diagnostic means. Further techniques are competitive ELISA, K39-ELISA, Western blot (WB) and Latex test. Ideally at least a repetition should be performed in six to eight weeks due to false positive and negative results (Noli, 1999). No correlation has been found between the degree of the clinical appearance and the serological titer. Serology is not useful for the control of a success in treatment, because antibodies may remain even after clinical recovery.

  • IFAT

Universally recognized as the gold standard is the indirect immunofluorescent antibody test (IFAT), being the most sensitive and specific test. IFAT is considered impractical for the examination of large numbers of sera, and there is no consensus on dilutions at which serum considered to be positive. Reported threshold titers range from 1/20 to 1/160.

 

Molecularbiological diagnosis

With the molecularbiological technique of PCR, a highly sensitive and specific method has been developed for the diagnosis of leishmaniosis. Parasite kinetoplast-DNA in liver, spleen, lymph node or bone marrow biopsies can be detected after multiplication. Fresh as well as formaline fixed paraffin embedded samples can be examined.

  • PCR

PCR is generally thought to approach a 'gold standard' status but has not been successful for mass screening e.g. in American cutaneous leishmaniosis epidemiological studies (with 31% positive by PCR versus 81% by ELISA) (Reithinger et al, 2003).
PCRs using different primers have been developed for the detection of leishmanial DNA in dogs. Alvar et al. (2002) described a nested PCR, targeting the sSSUrRNA gene (LnPCR) with very high specificity and sensitivity, which is ideal for diagnosis, monitoring the success of treatment and for predicting relapses in man (slightly less sensitive in dogs).

 

Xenodiagnosis

It is a technique in which the natural arthropod vector is used for the detection and isolation of the pathogen. This technique cannot be used as a routine measure but has been employed to solve epidemiological questions about the role of the clinical status and drug treatment of CanL.

Clinical Signs

Natural infections with Leishmania infantum in dogs may remain asymptomatic for long periods before clinical signs develop. But once the disease becomes patent, progression is usually rapid and death occurs within a few weeks to months. Clinically affected dogs usually exhibit one or more of nine main clinical features: 1. skin lesions, 2. loss of weight or poor appetite, 3. local or generalized lymphadenopathy, 4. ocular lesions, 5. epistaxis, 6. lameness, 7. anemia, 8. renal failure or 9. diarrhoea. No predilection of the infection for any age, breed or sex is observed, but Ferrer (1999) suggested from epidemiological data that mongrel dogs and some Spanish autochthonous breeds are more resistant. Small breeds seem to be affected less often, presumably due to their mainly indoor living. Despite high percentages of dogs that had contact in endemic areas (60 to 80%), many show no clinical signs. The incubation period also varies independent of age, sex or breed and generally takes some weeks or months up to several years. Atypical forms of canine leishmaniosis (CanL) with haemostatic problems, disorders of the cardiovascular, respiratory and musculo-skeletal systems, chronic colitis, and renal failure (without any other signs) have also been described.

 

Clinic in 'resistant' dogs 

Depending on the type of cellular immune response some dogs do not show any clinical signs or develop only cutaneous nodules at the place of inoculation, sometimes termed 'inoculation chancres'. They are usually located on the nose or ears, typical sites of sand fly bites, nodules of 1 to 3 cm, alopecic, ulcerated and crusted, non-pruritic and only mildly painful. The general appearance is that of a healthy dog, and most chancres disappear spontaneously in 1 to 6 months (Ferrer, 1999).

 

Clinic in 'susceptible' dogs 

The incubation period may vary between one month and seven years. During this time the parasites disseminate with predilection sites in the following organs: bone marrow, lymph nodes, spleen and liver. The damage caused by the parasite is dependent on two factors:

  • direct effect onto the tissue, causing non-purulent, infectious lesions in skin, liver, gut, kidneys, eyes and bones
  • indirect damage due to the deposit of immune complexes in joints and basal membranes of the kidneys, vessels and eyes, causing vasculitis, glomerulonephritis, polyarthritis and uveitis.

 

With exclusion of the CNS, numerous organs can be affected thus causing different clinic. Main symptoms are weakness, reduced physical activity, skin lesions/alterations and weight loss.

The first stage, displaying with an enlargement of the popliteal lymph nodes and deterioration in the texture of the dog's coat, co-exists with seroconversion and the presence of parasites in the reticuloendothelial system. The severity of the skin lesions seems to be depending on the state of immunity of the infected animal. Ill patients generally look older due to a marked muscle atrophy, especially in the head region. Immune complex deposits in the kidneys, leading to glomerulonephritis, are seen in 32 % of all clinical cases, often associated with a severe proteinuria. Nephropathies and associated symptoms represent the main cause of death in dogs.

 

Clinical manifestations and laboratory abnormalities found in canine leishmaniosis due to Leishmania infantum (Solano-Gallego et al., 2011)

Clinical manifestations

Laboratory abnormalities

General
  • Generalized lymphadenomegaly
  • Loss of body weight
  • Decreased or increased appetite
  • Lethargy
  • Mucous membranes pallor
  • Splenomegaly
  • Polyuria and polydypsia
  • Fever
  • Vomiting
  • Diarrhoea (including chronic colitis)
Serum proteins and electrophoretogram
  • Hyperglobulinaemia Polyclonal beta and/or gammaglobulinaemia
  • Hypoalbuminaemia
  • Decreased albumin/globulin ratio

 

 

Cutaneous
  • Non-pruritic exfoliative dermatitis with or without alopecia
  • Erosive-ulcerative dermatitis
  • Nodular dermatitis
  • Papular dermatitis
  • Pustular dermatitis
  • Onychogryphosis
Complete blood count (CBC)/Haemostasis
  • Mild to moderate non-regenerative anaemia
  • Leukocytosis or leukopenia
  • Thrombocytopathy
  • Thrombocytopenia
  • Impaired secondary haemostasis and fibrinolysis
Ocular
  • Blepharitis (exfoliative, ulcerative, or nodular) and conjunctivitis (nodular)
  • Keratoconjunctivitis, either common or sicca
  • Anterior uveitis/Endophtalmitis
Biochemical profile/urinalysis
  • Mild to severe proteinuria
  • Renal azotaemia
  • Elevated liver enzyme activities
Other
  • Mucocutaneous and mucosal ulcerative or nodular lesions (oral, genital and nasal)
  • Epistaxis
  • Lameness (erosive or non-erosive polyarthritis, osteomyelitis, polymyositis)
  • Atrophic masticatory myositis
  • Vascular disorders (systemic vasculitis, arterial thromboembolism)

 

 

Cutaneous leishmaniosis in dogs

Skin lesions are the most common manifestation of CanL in dogs admitted for treatment due to the disease (Ciaramella et al., 1997; Koutinas et al., 1999). They may be seen along with other clinical signs and/or clinicopathological abnormalities, be the only reported abnormality or may be absent. Several dermatological entities have been described (Ferrer et al., 1988; Koutinas et al., 1992): (1) non-pruritic exfoliative dermatitis with or without alopecia which can be generalized or localized over the face, ears and limbs, (2) ulcerative dermatitis over bony prominences, mucocutaneous junctions, paws, ear pinnae, (3) focal or multifocal nodular dermatitis, (4) mucocutaneous proliferative dermatitis and (5) papular dermatitis (Ordeix et al., 2005; Bottero et al., 2006).

 

Relative prevalences (%) of different skin lesions (in relation to all affected animals) (Noli, 1999)

Clinical signs

Relative prevalence (%)

Dry, exfoliative dermatitis

56-90.9

Ulcers

32.8-40

Periocular alopecia

18

Diffuse alopecia

14

Onychogryposis

24-54.5

Paronchia

13.6

Sterile, pustular dermatitis

1.6-13.6

Nasal depigmentation

4.5

Nasal/digital hyperkeratosis

4.5

Non-ulcerative nodules

4.5-16.8

But besides these skin lesions occurring along with a L. infantum/L. chagasi infection, dogs can also be infected with pathogenic agents of cutaneous leishmaniosis (in man). Generally, human cutaneous leishmaniosis (CL)/oriental sore may be anthroponotic, caused by L. tropica (in Asia) or L. peruviana, or zoonotic, caused by L. tropica (host: hyrax) (in Africa), L. major (host: gerbil), L. aethiopica, L. mexicana (respectively L. mexicana complex: L. mexicana, L. amazonensis, L. venezuelensis), L. amazonensis, L. panamensis, L. guyanensis or L. braziliensis (respectively L. braziliensis complex: L. braziliensis, L. panamensis, L. guyanensis). In CL, amastigotes multiply in lesions at the inoculation site, typically on the arms, legs, face or ears. Lesions are nodular or ulcerative, single or multiple. The typical lesion is a chronic 2-5 cm ulcer with indurated margins. Self-healing times are ≤5 months (L. major), ≤8 months (L. mexicana) and in ~1 year (L. tropica and L. braziliensis). Old World CL clinical features in man differ between and within regions. A 'classical' lesion starts as a nodule at the site of inoculation. A crust develops centrally, which may fall off exposing an ulcer which heals gradually, leaving a depressed scar with altered pigment. Satellite nodules at the edge of the lesion are common (WHO, 1990). New World CL has a wide variety of clinical manifestations (WHO, 1990). In endemic areas some forms are titled specifically.

L. braziliensis, L. peruviana and L. panamensis are frequently found in dogs as non-fatal cutaneous infections (Miles et al., 1999). L. braziliensis here is the main causative agent of cutaneous leishmaniosis in dogs in South America (Reithinger and Davies, 1999). Most of the dogs infected by L. braziliensis live in rural areas and they may present single cutaneous or mucosal lesions (Madeira et al., 2005). Dogs have been suspected to play a role in the domestic transmission cycle of L. braziliensis and L. peruviana in some areas of South America, but there is only circumstantial evidence supporting this hypothesis (Reithinger and Davies, 1999). In fact, the role of dogs in the maintenance of these parasites is probably minor (Dantas-Torres, 2007). A summary of CL pathogens detected in dogs in South America is listed below.

 

Leishmania species causing cutaneous leishmaniosis (CL) infecting dogs in South America (modified after Dantas-Torres, 2009)

Species

Disease form

Suspected/proven vectorsa

Geographical distribution

Leishmania braziliensis

Cutaneous

Lu. whitmani, among other

Argentina, Bolivia, Brazil, Colombia, Peru, Venezuela

Leishmania mexicana

Cutaneous

Lu. ayachuchensis

Ecuador

Leishmania panamensis

Cutaneous

Lu. trapido

Colombia, Ecuador

Leishmania peruviana

Cutaneous

Lu. peruensis, Lu. verrucarum

Peru

Leishmania pifanoi

Cutaneous

Unknown

Ecuador

Lutzomyia spp. that have been suspected to be involved in the transmission of Leishmania spp. to dogs in South America. Further information on the phlebotomine sand flies have been implicated as vectors of Leishmania spp. in this region can be found elsewhere (Lainson and Shaw, 2005; Young and Duncan, 1994).


Concomitant diseases

Probably due to a weakened cellular immunity infected dogs often suffer under accompanying diseases such as demodicosis and dermatophytosis. Concomitant infestation with other ectoparasites, especially ticks, lead to pathogen transmission (e.g. ehrlichiosis). Further concomitant diseases can be hepatozoonosis, and cryptococcosis. Generally, co-infection can alter clinical signs, duration of disease and the chances for successful treatment.

Treatment & Prevention

 

Treatment

The disease can be classified into four stages for determining the appropriate therapy:

  • Stage I: mild disease
  • Stage II: moderate disease
  • Stage III: severe disease
  • Stage IV: very severe disease

The detailed staging criteria can be found on the LeishVet homepage: http://www.leishvet.org/fact-sheet/clinical-staging/. According to this clinical staging, different treatment regiments are recommended.

 

Treatment in dogs 

In dogs anti-leishmanial treatment with current drugs often achieves temporary clinical improvement, but no elimination of the parasite.

The different drugs used for treatment of CanL are:

  • Allopurinol
  • Meglumine antimoniate
  • Miltefosine

Anti-leishmanial therapy can be practiced as monotherapy or in combination: http://www.leishvet.org/fact-sheet/therapy/

Furthermore genetic composition of the individual dog, determining the nature of immune response, as well as the susceptibility to drugs and the natural virulence of the infecting parasite may play an important role in therapy response and dog recovery. Parameters that should be considered before starting anti-leishmanial treatment are hemogram, renal and hepatic functions, electrophoretic protein pattern, antileishmania antibody titers, and bone marrow and lymph node parasite load.

 

Monitoring of canine therapy

The recommended monitoring during and after treatment of CanL was published by LeihVet group: http://www.leishvet.org/fact-sheet/monitoring/

 

Prognosis

Parasitological cure is often an exception in the course of treatment and relapses are frequently seen. Additionally dogs with a reduced renal function at the start of treatment possess the worst prognosis in general. A high proportion of dogs remain parasitologically positive after therapy and clinical cure, and therefore remain infectious to sand flies.

 

Treatment in humans

In humans, different drugs, dosages and treatment regimens are used. The WHO published a treatment regime for humans: https://www.who.int/leishmaniasis/research/978924129496_pp67_71.pdf?ua=1

 

Prevention

Prevention of canine leishmaniosis should include the application of long-acting topical insecticides to dogs with repellent activity against sand flies. The different recommendations can be found on the LeishVet homepage: http://www.leishvet.org/fact-sheet/prevention/

 

Vaccines

The LeishVet group recommends the preventative treatment with additional vaccination: http://www.leishvet.org/fact-sheet/vaccines/

Situation in Cats

For feline leishmaniosis (FeL) only limited data are available. This disease caused by Leishmania infantum is a worldwide problem. While subclinical feline infections are common in areas endemic for canine leishmaniosis, clinical illness due to L. infantum in cats is rare (Pennisi et al, 2015).

The prevalence rates of feline infection ranges from 0% to more than 60%. Due to the fact that cats are able to infect sand flies, they may act as a secondary reservoir, with dogs being the primary natural reservoir.

Cats can show clinical signs like lymph node enlargement and skin lesions such as ulcerative, exfoliative, crusting or nodular dermatitis, ocular lesions, feline chronic gingivostomatitis syndrome, mucocutaneous ulcerative or nodular lesions, hypergammaglobulinaemia and mild normocytic normochromic anaemia. In case the cat has a retroviral coinfection or an immunosuppressive therapy, clinical illness is more frequently compared to healthy cats.

Diagnosis of FeL is based on serology, PCR, cytology, histology, immunohistochemistry (IHC) or culture.

The most common treatment used in cats is allopurinol. Meglumine antimoniate has been administered in very few reported cases. Both drugs are administered alone and most cats recover clinically after therapy. Follow-up of treated cats with routine laboratory tests, serology and PCR is essential for prevention of clinical relapses (Pennisi et al, 2015).

Comparisons of populations of cats and dogs exposed to sand flies and L. infantum under the same conditions indicated that although a high rate of exposure was detected in cats as manifested by a significantly greater degree of seropositivity, dogs had significantly higher blood parasite loads, and were likely to be more infectious to sand flies than cats (Baneth et al., 2020).

References

Transmission

Slappendel RJ, Teske E: A review of canine leishmaniasis presenting outside endemic areas. In: Killick-Kendrick R (ed.): Canine leishmaniasis: an update. Proc. Int. Can. Leishm. Forum, Barcelona, Spain, 1999, Intervet Int., Boxmeer, The Netherlands, 1999, 54-9

 

Pathogenesis

Pinelli E, van Kaaij SY, Slappendel R, et al.: Detection of canine cytokine gene expression by reverse transcription-polymerase chain reaction. Vet Immunol Immunopathol. 1999, 69, 121-6

 

Diagnosis

Acedo-Sànchez C, Morillas-Màrquez F, Sanchìz-Marìn MC, et al.: Changes in antibody titres against Leishmania infantum in naturally infected dogs in southern Spain. Vet Parasitol. 1998, 75, 1-8

Alvar J, Cruz I, Morales MA, et al.: Molecular biology tools in leishmaniasis diagnosis and epidemiology. In: Killick-Kendrick R (ed.): Canine leishmaniasis: moving towards a solution. Proc. 2nd Int. Can. Leishm. Forum, Sevilla, Spain, 2002, Intervet Int., Boxmeer, The Netherlands, 2002, 25-30

Denerolle P: Leishmaniose canine: difficulté du diagnostic et du traitement. Prat Méd Chir Anim Comp. 1996, 31, 137-45

Noli C: Leishmaniose des Hundes. Waltham Focus 1999, 9, 16-24

Quinnell RJ, Courtenay O, Davidson S, et al.: Detection of Leishmania infantum by PCR, serology and cellular immune response in a cohort study of Brazilian dogs. Parasitol. 2001, 122, 253-61 

Reithinger R, Espinoza JC, Courtenay O: Evaluation of PCR as a diagnostic mass-screening tool to detect Leishmania (Viannia) spp. in domestic dogs (Canis familiaris). J Clin Microbiol. 2003, 41, 1486-93

 

Clinical Signs

Bottero E, Poggi M, Viglione M: Lesioni papulari indotte da Leishmania spp. in cani giovani. Veterinaria 2006, 1, 33-6

Ciaramella P, Oliva G, Luna RD, et al.: A retrospective clinical study of canine leishmaniasis in 150 dogs naturally infected by Leishmania infantum. Vet Rec. 1997, 141, 539-43

Dantas-Torres F: The role of dogs as reservoirs of Leishmania parasites, with emphasis on Leishmania (Leishmaniainfantum and Leishmania (Vianniabraziliensis. Vet Parasitol. 2007, 149, 139-46

Dantas-Torres F: Canine leishmaniosis in South America. Parasit Vectors. 2009, 2 (Suppl 1), S1

Ferrer L, Rabanal R, Fondevila D, et al.: Skin lesions in canine leishmaniasis. J Small Anim Pract. 1988, 29, 381-8

Ferrer LM: Clinical aspects of canine leishmaniasis. In: Killick-Kendrick R (ed.): Canine leishmaniasis: An update. Proc. Int. Can. Leishm. Forum, Barcelona, Spain, 1999, Intervet Int., Boxmeer, The Netherlands, 1999, 6-10

Koutinas AF, Scott DW, Kontos V, et al.: Skin lesions in canine leishmaniasis (Kala-Azar): a clinical and histopathological study on 22 spontaneous cases in Greece. Vet Dermatol. 1992, 3, 121-30

Koutinas AF, Polizopoulou ZS, Saridomichelakis MN, et al.: Clinical considerations on canine visceral leishmaniasis in Greece: a retrospective study of 158 cases (1989-1996). J Am Anim Hosp Assoc. 1999, 35, 376-83

Lainson R, Shaw JJ: New World leishmaniasis. In: Cox FEG, Kreier JP, Wakelin D (eds.): Topley & Wilson’s Microbiology and Microbial Infections, Parasitology. 2005, Arnold, London, pp. 313-49

Madeira MF, Schubach AO, Schubach TM, et al.: Is Leishmania (Vianniabraziliensis preferentially restricted to the cutaneous lesions of naturally infected dogs? Parasitol Res. 2005, 97, 73-6

Noli C: Leishmaniose des Hundes. Waltham Focus 1999, 9, 16-24

Ordeix L, Solano-Gallego L, Fondevila D, et al.: Papular dermatitis due to Leishmania spp. infection in dogs with parasite-specific cellular immune responses. Vet Dermatol. 2005, 16, 187-91

Reithinger R, Davies CR: Is the domestic dog (Canis familiaris) a reservoir host of American cutaneous leishmaniasis? A critical review of the current evidence. Am J Trop Med Hyg. 1999, 61, 530-41

Solano-Gallego L, Miró G, Koutinas A, et al.: LeishVet guidelines for the practical management of canine leishmaniosis. Parasit Vectors. 2011, 4, 86

Young DG, Duncan MA: Guide to the identification and geographic distribution of Lutzomyia sand flies in Mexico, the West Indies, Central and South America (Diptera: Psychodidae). Mem Amer Entomol Inst. 1994, 54, 1-881

 

Situation in Cats

Baneth G, Nachum-Biala Y, Zuberi A, et al.: Leishmania infection in cats and dogs housed together in an animal shelter reveals a higher parasite load in infected dogs despite a greater seroprevalence among cats. Parasit Vectors. 2020, 13, 115

Pennisi MG, Cardoso L, Baneth G, et al.: LeishVet update and recommendations on feline leishmaniosis Parasit Vectors. 2015, 8, 302

Further Reading

Baneth G, Koutinas AF, Solano-Gallego L, et al.: Canine leishmaniosis – new concepts and insights on an expanding zoonosis: part one. Trends Parasitol. 2008, 24, 324-30

Baneth G, Nachum-Biala Y, Zuberi A, et al.: Leishmania infection in cats and dogs housed together in an animal shelter reveals a higher parasite load in infected dogs despite a greater seroprevalence among cats. Parasit Vectors. 2020, 13, 115

Bañuls AL, Hide M, Tibayrenc M: Molecular epidemiology and evolutionary genetics of Leishmania parasites. Int J Parasitol. 1999, 29, 1137-47

Ferrer L: The pathology of canine leishmaniasis. In: Killick-Kendrick R (ed.): Canine leishmaniasis: moving towards a solution. Proc. 2nd Int. Can. Leishm. Forum, Sevilla, Spain, Intervet Int., Boxmeer, The Netherlands, 2002, 21-4

Killick-Kendrick R: The life-cycles of Leishmania in the sand fly and transmission of leishmaniasis by bite. In: Killick-Kendrick R (ed.): Canine leishmaniasis: moving towards a solution. Proc. 2nd Int. Can. Leishm. Forum, Sevilla, Spain, Intervet Int., Boxmeer, The Netherlands, 2002, 57-68

Miró G, Cardoso L, Pennisi MG, et al: Canine leishmaniosis – new concepts and insights on an expanding zoonosis: part two. Trends Parasitol. 2008, 371-77

Pennisi MG, Cardoso L, Baneth G, et al.: LeishVet update and recommendations on feline leishmaniosis Parasit Vectors. 2015, 8, 302

Slappendel, R.J., and E. Teske in: Killick-Kendrick, R. (ed.): Canine leishmaniasis: an update. Proc. Int. Can. Leishm. Forum, Barcelona, Spain, 1999, Intervet Int., Boxmeer, The Netherlands, 1999, 54-59

Slappendel RJ, Teske E: A review of canine leishmaniasis presenting outside endemic areas. In: Killick-Kendrick R (ed.): Canine leishmaniasis: an update. Proc. Int. Can. Leishm. Forum, Barcelona, Spain, 1999, Intervet Int., Boxmeer, The Netherlands, 1999, 54-9

Solano-Gallego L, Miró G, Koutinas A, et al.: LeishVet guidelines for the practical management of canine leishmaniosis. Parasit Vectors. 2011, 4, 86

WHO: Control of the leishmaniases. Report of a WHO Expert Committee, Tech. Rep. Ser. No. 793, WHO, Geneva, 1990

www.leishvet.org/

www.who.int/leishmaniasis/en/

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