GENERAL INTRODUCTION: POXVIRIDAE

Introduction

Camelpox is a systemic infection of dromedary camels (Camelus dromedarius) characterized by fever and the development of papules, vesicles, pustules and scabs in the skin particularly of the nostrils, lips and eyelids, caused by a species of orthopoxvirus. It has not been reported in bactrian camels (Camelus bactrianus). The camelpox virus is specific to dromedary camels and is found throughout the camel’s distribution in Africa and Asia,^{9, 25, 30, 36} but is absent from the wild dromedary population of Australia. The disease causes major production losses to camel producers.

Both bactrian and dromedary camels are susceptible to infection with a parapoxvirus which causes a localized infection only with lesions similar to those of orf in sheep and goats known as camel ecthyma.

Camelpox is classified within Risk Group 2 for human infection and requires appropriate biosafety and biosecurity handling. Although camelpox is considered as presenting limited public health risk, it is likely that more human cases of camelpox virus will occur due to reduced immunity to orthopoxvirus following the eradication of smallpox.³²

Aetiology

Camelpox belongs to the genus Orthopoxvirus in the family Poxviridae (OPV), subfamily Chordopoxvirinae (see General Introduction: Poxviridae). The viral genome contains 211 putative genes and is 205 719 base pairs long. It contains a unique region of approximately 3 kb, which encodes three ORFs (CMLV185, CMLV186, CMLV187) absent in other OPVs.¹ Phylogenetic analysis indicates that camelpox virus is clearly distinct but most closely related to variola virus (smallpox).¹⁸

There was considerable concern towards the end of the human smallpox eradication programme due to the fact that camelpox virus appeared indistinguishable from smallpox virus, and may cause disease in humans.⁴ However, it was subsequently shown to produce a different cytopathic effect in HeLa and certain other cell lines in which it generated the formation of multinucleate giant cells, unlike smallpox virus.⁵

When grown on the chorioallantoic membrane of embryonated chicken eggs at 37 °C, the lesions it induces are dense white pocks, very similar in appearance to those produced by smallpox virus. However, when cultivated at 34.5 °C the lesions can be distinguished from those of smallpox virus in that pocks with a haemorrhagic centre are produced. Camelpox virus also grows in Vero-, BHK-, lamb testis-, and lamb and calf kidney cells.

Camelpox virus haemagglutinates cockerel erythrocytes.¹⁰

Isolates of camelpox virus were previously identified using restriction endonucleases, and shown to be different from other species of orthopoxvirus.¹⁵ Today, full or partial genome sequencing is used to identify differences between camelpox viruses.¹⁶ More work is required to determine whether differences observed in the genomes of different camelpox virus isolates are responsible for differences in virulence.

Under the electron microscope camelpox virus is morphologically typical of other orthopoxviruses with a characteristic brick shape and distinct from ovoid parapoxviruses.

Epidemiology

The dromedary camel is found in Kenya, Somalia, Ethiopia and the Saharan countries, throughout the Arabian Peninsula, Iraq, Syria, Turkey, Iran, Afghanistan, Pakistan, southern countries of the former USSR, north India, west China and Mongolia. Seroprevalence rates of camelpox virus are high in most regions where the infection is present. In Morocco, seroprevalence rates were between 23-42 per cent in 2003 and 37-56 per cent in 2009 depending on the geographic region and higher in regions with a higher camel population and illegal trans-border animal movement.³³

In endemic situations camelpox is a disease of young camels, affecting them as they lose their colostrum-derived immunity. It possesses many of the epidemiological characteristics of smallpox and sheep- and goatpox, depending for its survival on a constant supply of susceptible camels. It has been thought that infection probably induces life-long sterile immunity, there being no carrier state, but more recently the virus has been identified in atypical skin lesions of camels that suffered from the typical camelpox 12 months previously. Naïve camels that were in contact with camels presenting atypical minute pox-like skin lesions became infected within two weeks, raising the possibility of virus persistence or a carrier in the herd.³⁹ Since atypical pox-like skin lesions could easily be missed by veterinarians and camel handlers, it is possible that these lesions could be important for the maintenance of camelpox virus. More investigative work is required on this issue.

 A reservoir host other than camels is unlikely to exist. It would seem that the virus survives mainly by constant transmission from infected to susceptible animals, and therefore requires a certain minimum size of susceptible population. The size of this population depends on the susceptibility of the host population and, ultimately, on Ro (the basic reproductive number), i.e. the number of susceptible animals infected, on average, by a single diseased animal. Where a population of camels is unable to maintain the virus, the susceptible population will also include adult animals, so that when the virus is re-introduced, all age groups will develop disease. However, the movement of camels over long distances usually ensures the spread of virus. Transmission is by contact between infected and susceptible camels and the virus is probably spread in aerosols, but the involvement of arthropod vectors such as the camel tick, Hyalomma dromedarii, and environmental contamination with scabs are also suspected to play a role in its transmission.³⁶ Disease is reported to occur more frequently and in a more severe form during the rainy season.

The virus is stable in dried scabs for at least 4 months,²⁹ allowing the scabs to contaminate the environment. Transmission can occur by either direct contact between infected and susceptible animals or by indirect contact with a contaminated environment. Infection can occur through inhalation or through skin abrasions. Virus occurs in saliva, nasal and ocular secretions as well as in milk, although neither the quantity of virus nor duration of shedding has been evaluated in experimentally infected camels.

Field investigations have shown that camelpox virus only rarely causes disease in humans, characterized by mild skin rash. When camelpox was experimentally inoculated into the skin of rhesus monkeys (Macaca mulatta) it caused only a localized pustule.⁴ Cases among camel handlers and attendants with papules, vesicles, ulceration and scabs on their fingers and hands were confirmed as resulting from camelpox infection in India.⁷ Three camel herders with clinical signs (erythema, vesicles, and pustules that involved arms, hands, legs, back, and abdomen that resolved within two months without further transmission) were described in Sudan.²³

Pathogenesis, clinical signs and pathology

The natural incubation period following contact exposure is approximately 9-13 days.

In experimental cases of the disease induced by intradermal or subcutaneous inoculation of the virus, the incubation period was five days. At the site of inoculation a papule developed which, over a 9- to 10-day period, progressed to form a vesicle, pustule and scab.

Generalization, characterized by fever and the development of multiple skin lesions at other sites, occurred 9 to 11 days after inoculation. Enlarged lymph nodes and viraemia within the first week of clinical signs are also characteristic.²⁹

The clinical disease can vary from inapparent or small nodular lesions confined to the skin to severe systemic disease. It is likely that genetic differences in the virus and the host as well as immune status of the host play a role in the severity of infection. During an outbreak, affected camels first develop mild fever, followed one to three days later by the appearance of multiple lesions in the skin, particularly of the head, lips, nares, eyelids, neck and limbs. The skin of the mammary glands and external genitalia is also frequently affected. In severe cases, the lips and nasal mucosa become severely swollen and some animals develop severe lesions in the eyes and blindness. Skin lesions can take four to six weeks to heal. Other signs include salivation, lacrimation, mucopurulent nasal and eye discharges, diarrhoea and abortion in some pregnant animals suffering severe systemic disease.

Death may occur during the acute stage of infection, possibly related to the observation that the virus is capable of replication in myocardial cells. Duration of the clinical disease is usually between 10 and 30 days, unless secondary bacterial infections occur. Case mortality rates of 5 to 28 per cent have been reported in adult animals,^{25, 30, 36} and in young animals mortality rates vary from 25 to 100 per cent.²⁶

At necropsy, apart from the skin lesions, multifocal necrotic lesions may be present in the mucosa of the trachea and oesophagus, and nodular lesions up to 50 mm in diameter may occur in the lungs.

Microscopic examination of the skin lesions reveals cytoplasmic swelling, vacuolation and ballooning of keratinocytes of the outer stratum spinosum and perivascular infiltration by mononuclear cells, eosinophils and neutrophils (see General Introduction: Poxviridae).

Immunohistochemistry performed on lung lesions revealed staining of some bronchial epithelial cells, as well as antigen in infiltrating macrophages in the bronchial mucosa and alveolar macrophages.²⁴

Diagnosis and differential diagnosis

A presumptive diagnosis can usually be made on the basis of clinical signs which, when generalized, are likely to be confused with little else. Localized lesions may be mistaken for papilloma or parapox virus infections as well as skin reactions to insect bites.

Several diagnostic methods available for camelpox infection are described in detail in the chapter on camelpox of the OIE 2017, Manual of Diagnostic Tests and Vaccines for Terrestrial Animals,http://www.oie.int/fileadmin/Home/eng/About_us/docs/pdf/Session/2017/A_FR_2017_public.pdf, Jun 15, 2017.²⁹ These diagnostic tests include identification of poxvirus virions using transmission electron microscopy, virus isolation, immunohistochemistry, molecular identification of camelpox virus genetic material and serology to detect orthopoxvirus antibodies. Where possible more than one diagnostic method should be used to achieve a confirmed diagnosis of disease.

Samples of skin lesions are most useful for diagnostic purposes. Blood can also be used but not recommended as viraemia is transient and contains lower amounts of virus than skin lesions.  Nasal or oral swabs are also useful specimens.

Samples of scab or skin biopsy material contain typical orthopox virions that can be identified and differentiated from parapox virions by transmission electron microscopy(TEM). Unfortunately TEM can only identify the virus as an orthopox virus and not specifically as camelpox virus.

Virus isolation can be attempted using a biopsy specimen that is homogenized and an inoculum placed on susceptible cells. Camelpox virus can be grown in a variety of cell lines including Vero, BS-C-1 (transformed green monkey kidney), MA-104 and MS monkey kidney, baby hamster kidney (BHK), HELA, and Dubai camel skin (Dubca) as well as primary cells such as lamb testis, lamb kidney, camel embryonic kidney, calf kidney, and chicken embryo fibroblasts. The cytopathic effect (CPE) consists of foci or rounded cells, ballooning and giant-cell formation.  Syncytia characterized by large multinucleated cells develop in cultures of lamb testis cells, HeLa cells, BS-C-1 cells and transformed human amnion cells.

Camelpox virus also grows well in cells of the chorioallantoic membrane (CAM) of embryonated chicken eggs (see above). Between 4 and 5 days after inoculation the eggs are opened and characteristic greyish white pox-like lesions observed on the CAM. These pox lesions contain high levels of virus (10⁵ TCID₅₀ per ml in VERO cells).¹⁴

Since camelpox is an Orthopoxvirus all anti-orthopoxvirus sera can be used to stain camelpox virus antigen in infected cells by immunofluorescence.^{14, 37} Immunocytochemistry, using either polyclonal anti-orthopoxvirus sera or an anti-vaccinia monoclonal antibody, can also be used.³⁷

Conventional PCR methods are available to amplify camelpox virus genome. For example, a method based on amplifying the C18L gene and a duplex PCR based on the C18L and DNA polymerase genes was developed in order to identify and differentiate camelpox from other orthopoxviruses, capripoxviruses and parapoxviruses.³ A PCR based on the haemagglutinin protein gene has been developed with primers to allow detection of orthopoxivurus DNA followed by a secondary PCR using specific primers for the same gene in camelpoxvirus generating a different sized DNA fragment.³¹ Similarly, amplification of the A-type inclusion protein gene amplifies orthopoxvirus species and allows the differentiation into vaccinia, mousepox, monkeypox and camelpox based on the size of the amplified fragment.^{27, 28} A multiplex PCR has been developed to diagnose camelpox virus, camel parapox virus  and camelus dromedary papilloma virus that can co-infect camels.²²

LAMP assay targeting the C18L gene sequence of CMLV has been developed.³⁴ A real- time PCR based on the C18L gene has been developed for quantification of camelpoxvirus.³⁵ A real- time PCR assay that differentiates the specific poxvirus using melting curves was developed to support the differential diagnosis of pox like diseases in ruminants.¹⁷

Serological tests are time-consuming, making them impractical for primary diagnosis. However, serology is useful for secondary confirmatory testing and to determine the prevalence of infection in areas where vaccination is not applied. Serology can be performed using a virus neutralization assay on VERO cells.  An ELISA that utilizes purified camelpox virus as the antigen has been described.²

Control

There is no specific treatment for an animal already affected but supportive treatment is essential for valuable animals. This should include antibiotic cover, topical treatment of the lesions, protection from flies and other ectoparasites, assistance with feeding when necessary and, possibly, administration of appropriate quantities of intravenous fluids. Affected animals should be isolated in insect-proof accommodation if that is available. There are no approved antiviral drugs available that can be used for treatment of camelpox virus.

Before specific camelpox virus vaccines were available, effective protection was provided by immunizing uninfected animals with vaccinia, rendered effective by the close antigenic relationship between orthopoxviruses.²¹ It is even possible to protect camels against camelpox using smallpox virus,⁶ although even if that virus were available, this would obviously not be recommended. The efficacy of generating protective immunity against camelpox has also be demonstrated using a modified vaccinia virus (Ankara [MVA]) vaccine expressing the MERS-CoV spike protein.¹⁹ Specific vaccines have more recently been developed using attenuated strains of camelpox virus.^{20, 37} It has been shown that a live attenuated camelpox virus vaccine can provide protection against the disease for at least 6 years.³⁸ Unfortunately the mechanism of attenuation of the camelpox virus is not currently understood. Restriction enzyme analysis of the vaccine demonstrated that there were no major differences between the vaccine and the wild type viruses.³⁷ Analysis using full genome sequencing may allow for the mechanism of attenuation to be determined; however, it is likely that multiple point mutations will be identified similar to attenuated capripoxvirus vaccines. An inactivated camelpox virus vaccine has been developed using a camelpox virus isolated from Morocco in 1984.¹³ The inactivated vaccine provides good protection against camelpox after two vaccinations at a 3 to 6 month interval followed by an annual revaccination.¹²

The limited host range of camelpox and the availability of effective vaccines offers the possibility of camelpoxvirus eradication.¹¹ However, the large population of camels and the long distance movement of camels are hurdles to achieving that end.⁸

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