Theileriosis of sheep and goats

Previous Authors: 

Current Authors:
J A LAWRENCE - Extraordinary Professor, DPhil, BSc, MRCVS (ret.), DTVM, Department of Paraclinical Veterinary Science, University of Zimbabwe, Harare, Zimbabwe 
C BYARUHANGA - Post-doc Fellow, BVM, MVPM, PhD, Department of Veterinary Tropical Diseases, Para-clinical Building, Faculty of Veterinary Science, University of Pretoria, 100 Old Soutpan Road, Onderstepoort, Pretoria, Gauteng, 0110, South Africa
M OOSTHUIZEN - Parasitology, PhD, Department of Veterinary Tropical Diseases, Faculty of Veterinary Science, Private Bag X04, University of Pretoria, Pretoria, Gauteng, 0110, South Africa
B J MANS - Principal Researcher, BSc, BSc (Hons) Biochemistry, MSc (Biochemistry), PhD (Biochemistry), Agricultural Research Council, Onderstepoort Veterinary Research, 100 Old Soutpan Road, Pretoria, Gauteng, 0110, South Africa

GENERAL INTRODUCTION: THEILERIOSES OF CATTLE

Introduction

Theileria is a genus comprising tick-transmitted parasitic protozoa in the family Theileriidae, order Piroplasmida, subclass Piroplasmia, phylum Apicomplexa. It includes parasites previously allocated to the genera Cytauxzoon, Gonderia and Haematoxenus. The taxonomy of the genus at species level on morphological and biological characteristics in the past has been confused; taxonomy is now based on sequences of the 18S rRNA genes. Three Theileria species infecting small ruminants have been shown to be responsible for heavy mortality. Two of them, T. uilenbergi and T. luwenshuni were first described in China, whereas T. lestoquardi causes malignant theileriosis which is a major problem in Sudan and the Middle East. Other small ruminant Theileria species are considered as non-pathogenic; their importance lies in the confusion they may cause in diagnosis. The name T. recondita was given in the past to a non-pathogenic parasite in Europe transmitted by Haemaphysalis spp. but is regarded as invalid ⁸;the parasite was probably T. luwenshuni.

Table 1: Theileria species affecting sheep and goats

The life cycle of the Theileria species of sheep and goats is similar to that of the bovine Theileria species (See East Coast fever). The parasite is transmitted to its vertebrate host by the bite of an infected tick. Sporozoites injected into the host invade mononuclear leucocytes, either lymphocytes or macrophages, depending on the species, where they develop into schizonts. Some species have the ability to transform the host cell so that it can proliferate and disperse throughout the body, others do not. The schizonts release merozoites into the circulation, where they can be seen as piroplasms in erythrocytes. Ticks ingest the organisms when they feed. The organism undergoes multiplication and a sexual cycle in the tick gut and moves to the salivary gland where it is ready to start the next cycle. The organisms are transmitted from stage to stage of the tick, not transovarially from one tick generation to the next.

Malignant Ovine Theileriosis

Aetiology and life cycle

Theileria lestoquardi (T. hirci) is the best known causal organism of malignant theileriosis of small ruminants and has a life cycle and morphology typical of the genus Theileria. Theileria lestoquardi schizonts infect macrophages and B lymphocytes ¹⁴. The parasite infects both sheep and goats. Antigenically it is closely related to T. annulata of cattle31, and it is occasionally detected by PCR in this species ⁵¹.The parasiteis transmitted from stage to stage through the ticks Hyalomma anatolicum anatolicum, H. impeltatum, H. excavatum, H. scupense (H. detritum), Rhipicephalus sanguineus and R. turanicus, and through vertical transmission ¹⁴.

Schizont-infected cells can be grown readily in suspension culture, in a manner similar to those of T. parva and T. annulata (see East Coast fever and Theileria annulata theileriosis) ²⁶ Cultured schizonts are of value in immunization and as antigen for indirect immunofluorescence testing.³¹

Epidemiology

Malignant theileriosis of sheep and goats caused by T. lestoquardi occurs in the Mediterranean basin, the Middle East, Sudan, western Asia and the Indian subcontinent ¹⁴, and infection has been identified in the absence of clinical disease in Tanzania ⁴⁹ and southern Tunisia ⁴⁶. The disease is widespread in some important sheep breeding areas and causes heavy losses in both indigenous and exotic breeds and their crosses, especially in Iran, Sudan and the Sultanate of Oman; the disease is particularly severe when susceptible sheep are introduced into endemic areas ¹⁴. Morbidity and mortality among non-immunized animals can reach 77% and 46%, respectively. There is disagreement in the literature as to whether lambs are more or less resistant than adults. Breed differences in severity of infection have been demonstrated ¹⁵.  Animals that survive infection are resistant to further challenge, and indigenous sheep and goats usually acquire immunity at an early age 14. However, the development of endemic stability with respect to theileriosis may be hampered by the practice of transhumance in sheep-raising areas, and reported differences in susceptibility of various classes of stock probably reflect differences in the epidemiological status in different situations. The disease usually occurs over a period of two months at the end of spring, when ticks are active.²²

Clinical signs

Malignant theileriosis of sheep and goats caused by T. lestoquardi may be acute, subacute or chronic. The most prominent clinical signs are generalized enlargement of the superficial lymph nodes, high fever, listlessness, anorexia, emaciation, intermittent diarrhoea or constipation and loss of condition ¹⁴. Initially, infected animals have an apparently normal appetite, but a few days after the onset of fever they cease eating and later on they become progressively emaciated. Other signs described are mucosal pallor, nasal discharge, hyperaemia of the conjunctivae, oedema of the throat, icterus and transitory haemoglobinuria. Clinical signs in subacute and chronic cases are similar but less marked, and anaemia and emaciation may be pronounced. The severity of the clinical signs is significantly greater in sheep than in goats ²³.

Death follows within four or five days; recovery, if it occurs, is slow.

Pathogenesis and pathology

The pathogenesis and pathology of malignant theileriosis caused by T. lestoquardi are similar to those of Theileria annulata. Both schizonts and piroplasms play a role, the former by inducing macrophage and lymphoid proliferation and dissemination and by disrupting the cell-mediated immune response and stimulating overproduction of pro-inflammatory cytokines, the latter by causing haemolysis, anaemia and icterus.

Salient features at post-mortem examination in acute cases include subcutaneous and intramuscular oedema, especially in the ventral parts of the head, neck and abdomen; enlargement of the spleen and lymph nodes; icterus; enlargement, yellow discolouration and subcapsular haemorrhage of the liver; pulmonary oedema and hydrothorax; red or white nodules of lymphoid hyperplasia in the kidney cortex (so-called infarcts) and perirenal oedema; haemorrhages on serosal surfaces and beneath the epicardium and endocardium; and haemorrhagic lesions and necrosis of the mucosa of the abomasum and large intestine. In chronic cases, anaemia, emaciation and generalized oedema may be prominent.

Diagnosis and differential diagnosis

The diagnosis of malignant theileriosis of sheep and goats is based on the characteristic clinical signs and lesions and may be confirmed by the demonstration of schizonts in lymph node and liver smears; piroplasms may be present in blood smears, but may be infrequent or even absent in the acute disease.¹⁷ Specific diagnosis can be made retrospectively by means of the indirect immunofluorescent antibody test.³¹ An ELISA based on an unknown surface protein (clone 5) identified as a K+-dependent Na+/Ca+ exchanger by conserved domain database analysis has been developed and shows 94.6% sensitivity and 88% specificity compared to the IFAT ⁷.   Assays to directly detect parasite DNA using molecular methods have been developed. Conventional PCR assays based on the 30 kDa merozoite surface protein of T. lestoquardi may be used for detection of parasite in blood or infected ticks ^{29, 30}. A reverse line blot assay that specifically targets the 18S rRNA region of Theileria of small ruminants can detect T. lestoquardi, T. luwenshuni, T. uilenbergi, T. ovis and T. separata 49. A PCR that targets the T. lestoquardi ms1-2 gene was used to detect parasites in stained blood smears ⁵⁰. Simultaneous detection of T. annulata, T. lestoquardi and T. ovis was achieved by nested PCR followed by restriction fragment length polymorphism (RFLP) analysis ^{16, 25, 28}.  A loop mediated isothermal amplification assay (LAMP) specific for T. lestoquardi shows 92.1% specificity and 87.5% sensitivity when compared with conventional PCR assays ⁴⁸.

Control

In endemically unstable environments or when susceptible sheep are introduced to endemic areas tick control or some other disease control measure is essential ¹⁴. Eradication is not a practical proposition due to environmental, managerial and resource constraints and to the lack of a strategy to generate infection-free animals, vector or environment.

Buparvaquone may be used in the treatment of malignant theileriosis in valuable animals.^{17, 22}

Attenuated schizont vaccines for T. lestoquardi prepared in cell culture have proved very successful when tested in Iraq, Iran and Sudan ¹⁴. The parasite is passaged in culture until it causes only mild clinical signs and low parasitaemia in susceptible lambs in a similar way to T. annulata.  

In endemic areas, tick control is either not practised, or used only occasionally to reduce excessive tick burdens as indigenous sheep rarely show disease. Intensive tick control of susceptible sheep is difficult to maintain, and any breakdown is likely to result in an outbreak of disease.

Oriental Ovine Theileriosis

Aetiology and life cycle

Two species of Theileria, T. luwenshuni, originally known as China ¹, and T. uilenbergi, known as China ², are responsible for fatal theileriosis of sheep and goats in western China, which has been recognised for over 100 years ⁶¹.The disease was attributed initially to T. ovis and later to T. lestoquardi. The species are not closely related to each other antigenically; T. luwenshuni is closely related to T. buffeli/orientalis. Strains of both species in Western Europe and Western Asia are generally benign. Both species are transmitted by Haemophysalis qinghaiensis and H. longicornis ^{35, 36, 61}, while H. punctata is suspected to be the vector in Western Europe ⁴⁵. The parasite has been detected by PCR in Dermacentor niveus ³³, though transmission by this species has not been demonstrated.

Since their identification in sheep and goats, T. luwenshuni has been shown to be widespread in Mongolian gazelle (Procapra gutturosa) ³³ and roe deer, (Capreolus pygargus), sika deer (Cervus nippon) and red deer (Cervus elaphus) ³⁴, and serow (Capricornis crispus) ⁴⁹, while T. uilenbergi has been detected in Reeves' muntjac (Muntiacus reevesi) ⁵⁷ red deer and sika deer ³². Presumably wild deer and gazelle are the original hosts of these species, and infection has spread from them to domestic small ruminants. Theileria luwenshuni has also been detected in hedgehogs in China ¹² and sheepdogs in Iran ²⁰.

Both species have a life cycle and morphology typical of the genus Theileria ⁶¹. Macroschizonts and microschizonts can be found in lymph nodes, spleen, liver, kidney, lung and peripheral blood, but are more often extracellular than intracellular. Three days after the appearance of the schizonts in the lymph nodes, piroplasms of a variety of shapes can be detected in erythrocytes of experimentally infected animals.

Neither species has been established in cell culture.

Epidemiology

The distributions in China of T. uilenbergi and T. luwenshuni overlap ⁶⁰ but T. luwenshuni appears to be much more widespread, as it is often the only species of the two detected in surveys ^{11, 19, 37, 58, 63}. As both species are transmitted by the same ticks, other unidentified epidemiological factors must control their distribution. Depending on the area, the morbidity rate in sheep and goats may reach 65%, while mortality may reach 75% ⁶¹. Up to 47% of the sheep in an endemic area die because of the infection. Highest mortality occurs in lambs and imported exotic animals. Peak infection occurs in spring ⁴¹.

Theileria luwenshuni has also been identified in wildlife in Korea ⁴² and Japan ⁴⁹ and, in the absence of clinical illness, in sheep in Spain, where it was originally named Theileria sp. OT1 ⁴³. Great Britain ⁴⁵; and, together with T. uilenbergi, in Italy ² and Turkey, Iraq and Iran ⁹.  High mortality in sheep in an outbreak in England attributed to T. luwenshuni was probably exacerbated by very heavy tick infestation ⁴⁵. The introduction of the Chinese Theileria into Western Europe may have been in deer imported from China over a century previously ⁴⁵.

Clinical signs, pathogenesis and pathology

Pathogenesis and pathology have not been described from China. In one British sheep examined the lips and tongue were swollen and oedematous, and there was also oedema of the conjunctivae and subcutaneous oedema around the eyes and elsewhere over the face and throat ⁴⁵. There were petechial haemorrhages over the oral mucosa. The lungs were oedematous and mottled and excessive froth was present in the trachea and bronchi. The spleen was enlarged, the kidneys pale and the blood watery.

Diagnosis and differential diagnosis

The diagnosis of oriental theileriosis of sheep and goats is based on the characteristic clinical signs and lesions and may be confirmed by the demonstration of schizonts in lymph node and liver smears; piroplasms may be present in blood smears, but may be infrequent or even absent in the acute disease.¹⁷ Specific diagnosis can be made retrospectively by means of the indirect immunofluorescent antibody test.31 An ELISA based on the recombinant immunodominant protein from T. uilenbergi (rTuIP) was developed for use in sheep ³⁹. Diagnostic sensitivity and specificity was determined at 98.4%. The assay is capable of detecting early infection and was tested up to two months after infection. The ELISA also detect antibodies from T. luwenshuni and may as such be used in epidemiological studies if testing for oriential ovine theileriosis in general. This assay was optimized for rapid screening (15 minutes) in a field setting using a colloidal gold-based immunochromatographic strip and showed 93% sensitivity and 100% specificity compared to the rTuIP ELISA (Lu et al., 2015). An indirect ELISA based on a T. luwenshuni surface protein (rTlSP) does not give cross-reactions with T. uilenbergi ²⁴.  

A conventional PCR based on species specific primers that targets the 18S rRNA gene has been developed that distinguish T. luwenshuni and T. uilenbergi ⁵⁹.  LAMP PCR assays was developed for T. luwenshuni based on UTRlu8 and for T. uilenbergi for UTRu6 ³⁸. Compared to RLB both assays showed a sensitivity and specificity of 66% and 97%, respectively. A multiplex PCR based on the 5.8S rRNA for T. ovis and T. luwenshuni and the 18S rRNA gene of T. uilenbergi that is 100% specific, detect parasites up to a parasitemia of 0.001%.  Given the relatively low sensitivity, this assay could be used as a herd test, to determine whether the agents of oriental ovine theileriosis exist in a region, similar to the study of ³⁷ that showed the existence of T. luwenshuni in central China, but not T. ovis or T. uilenbergi. A multiplex DNA bead-based suspension array based on Luminex xMAP technology was developed in Spain to detect T. luwenshuni, T. sp. OT3, T. ovis, Babesia motasi and B. ovis and confirmed that these parasites are endemic to this region ⁴⁷.  

Control

Treatment of affected sheep with Primaquine has proved effective ⁴¹, and it is likely that buparvaquone would also be effective. Tick control is the only method of prophylaxis available; treatment with organophosphate acaricides at 15-20 day intervals reduced infection rates dramatically ⁴¹. No vaccines have been developed.

Benign Ovine Theileriosis

Aetiology and life cycle

Theileria separata, originally described as Haematoxenus separatus, ⁹ is phylogenetically most closely related to Theileria sp. (sable), a parasite responsible for mortality in sable (Hippotragus niger) and roan (Hippotragus equinus) antelope in South Africa.  It has been hypothesized that T. separata was probably passed from sable antelope to domestic sheep ⁴⁹. Theileria separata has also been detected in sable antelope ⁸, common tsessebe (Damaliscus lunatas) ¹⁰, and blue wildebeest (Connochaetes taurinus), black wildebeest (Connochaetes gnou), springbok (Antidorcas marsupialis) and red hartebeest (Alcelaphus buselaphus caama) ⁸. It is transmitted by Rhipicephalus evertsi evertsi and R. e. mimeticus.⁵⁶ The piroplasms are readily transmitted by blood inoculation.⁵³ Infectivity for goats is very low. It is common in eastern, central and southern Africa; reports of its occurrence in Europe and Asia are probably the result of misidentification.
Theileria separata can be distinguished morphologically from the other species of Theileria in small ruminants by the association with the piroplasms of a clearly defined veil, often situated outside the erythrocyte membrane but lying close to it, opposite a corresponding gap in the erythrocyte cytoplasm (see Non-pathogenic Theileria species in Cattle: Figure 35.1). There may also be a small granule, possibly a reduced bar structure of parasitic origin, within the erythrocyte cytoplasm. Veils can be seen associated with a proportion of piroplasms in Giemsa-stained smears but may not be obvious. They are more easily demonstrated in wet preparations by phase-contrast microscopy.⁶²

The T. ovis complex infects both sheep and goats, and its vectors have not been identified with certainty. Rhipicephalus bursa is known to transmit one non-pathogenic ovine Theileria, and Amblyomma spp. are suspected to be the vectors of another,⁵⁵ while Haemaphysalis punctata has been shown to transmit a third.⁵⁶ Theileria ovis has been detected in sheep dogs in Iran ²⁰.

Epidemiology

Little is known of the epidemiology of the benign species of Theileria.  Molecular detection using reverse line blot analysis confirmed the presence of T. ovis in small ruminants in Northern Spain ^{18, 43}. Pakistan ⁶, Sudan ¹⁶ and Turkey ^{4, 5, 6, 44}. Theileria sp. MK was first identified in Turkey ⁴, using reverse line blot and confirmed using a specific PCR ³. Beyond molecular detection, not much is known of this Theileria, except that its geographic distribution is similar to T. ovis. Theileria sp. OT3 was first identified in Spain in sheep using reverse line blot ^{43, 47}.  It shows high prevalance in red deer, roe deer and chamois in northern Spain ¹⁸. It has also been found in sheep in China ⁵², Croatia ¹³, Italy ²¹ and Turkey ^{4, 5}.

Clinical signs

The benign Theileria spp. cause only a mild febrile reaction, moderate swelling of the superficial lymph nodes and possibly mild anaemia.

Pathogenesis and pathology

Enlargement of lymph nodes and mild anaemia are the only lesions to be expected in benign theileriosis.

Diagnosis and differential diagnosis

Benign theileriosis may be suspected when piroplasms alone are found in sheep or goats in the absence of severe disease, but it is necessary to exclude the presence of pathogenic species in a latent or subclinical form by testing for antibodies or parasitic molecules. The characteristic features of the piroplasms of T. separata permit differentiation from other Theileria spp. in sheep. Reverse line blot and gene specific PCR assays have been used to detect and distinguish T. ovis, T. sp. MK and T. sp. OT3 as described under Epidemiology.

Control

There are no indications for the control of the benign species of Theileria.

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