RESUMO
BACKGROUND: Sheep poxvirus (SPPV), Goat poxvirus (GTPV) and Lumpy skin disease virus (LSDV) are the most serious poxviruses of ruminants. They are double stranded DNA viruses of the genus Capripoxvirus, (subfamily Chordopoxvirinae) within the family Poxviridae. The aim of this study was to develop a Loop-mediated isothermal AMPlification (LAMP) assay for the detection of Capripoxvirus (CaPV) DNA. RESULTS: A single LAMP assay targeting a conserved region of the CaPV P32 gene was selected from 3 pilot LAMP assays and optimised by adding loop primers to accelerate the reaction time. This LAMP assay successfully detected DNA prepared from representative CaPV isolates (SPPV, GTPV and LSDV), and did not cross-react with DNA extracted from other mammalian poxviruses. The analytical sensitivity of the LAMP assay was determined to be at least 163 DNA copies/µl which is equivalent to the performance reported for diagnostic real-time PCR currently used for the detection of CaPV. LAMP reactions were monitored with an intercalating dye using a real-time PCR machine, or by agarose-gel electrophoresis. Furthermore, dual labelled LAMP products (generated using internal LAMP primers that were conjugated with either biotin or fluorescein) could be readily visualised using a lateral-flow device. CONCLUSIONS: This study provides a simple and rapid approach to detect CaPV DNA that may have utility for use in the field, or in non-specialised laboratories where expensive equipment is not available.
Assuntos
Capripoxvirus/genética , DNA Viral/genética , Técnicas de Amplificação de Ácido Nucleico/veterinária , Infecções por Poxviridae/veterinária , Animais , Capripoxvirus/química , DNA Viral/análise , Eletroforese em Gel de Ágar/veterinária , Técnicas de Amplificação de Ácido Nucleico/métodos , Infecções por Poxviridae/diagnóstico , Infecções por Poxviridae/genética , Infecções por Poxviridae/virologia , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
The Caribbean enjoys a long-standing eminence as a popular tourist destination; however, over the years it has also amassed the sobriquet "arbovirus hotspot". As the planet warms and vectors expand their habitats, a cognizant working knowledge of the lesser-known arboviruses and the factors that influence their emergence and resurgence becomes essential. The extant literature on Caribbean arboviruses is spread across decades of published literature and is quite often difficult to access, and, in some cases, is obsolete. Here, we look at the lesser-known arboviruses of the insular Caribbean and examine some of the drivers for their emergence and resurgence. We searched the scientific literature databases PubMed and Google Scholar for peer-reviewed literature as well as scholarly reports. We included articles and reports that describe works resulting in serological evidence of the presence of arboviruses and/or arbovirus isolations in the insular Caribbean. Studies without serological evidence and/or arbovirus isolations as well as those including dengue, chikungunya, Zika, and yellow fever were excluded. Of the 545 articles identified, 122 met the inclusion criteria. A total of 42 arboviruses were identified in the literature. These arboviruses and the drivers that affect their emergence/resurgence are discussed.
Assuntos
Infecções por Arbovirus , Arbovírus , Febre de Chikungunya , Dengue , Febre Amarela , Infecção por Zika virus , Zika virus , Humanos , Região do Caribe , Dengue/epidemiologiaRESUMO
Experimental challenge protocols that consistently reproduce clinical signs of peste des petits ruminants in Alpine goats infected with a tissue culture-passaged peste des petits ruminants virus are described. The protocols can be used to carry out quality-controlled vaccine efficacy and pathogenesis studies under experimental conditions.
Assuntos
Modelos Animais de Doenças , Peste dos Pequenos Ruminantes/patologia , Peste dos Pequenos Ruminantes/virologia , Vírus da Peste dos Pequenos Ruminantes/patogenicidade , Animais , CabrasRESUMO
Sheep and goats sampled in Kuwait during February 2010 were seropositive for bluetongue virus (BTV). BTV isolate KUW2010/02, from 1 of only 2 sheep that also tested positive for BTV by real-time reverse transcription-PCR, caused mild clinical signs in sheep. Nucleotide sequencing identified KUW2010/02 as a novel BTV serotype.
Assuntos
Vírus Bluetongue/classificação , Vírus Bluetongue/isolamento & purificação , Bluetongue/virologia , Animais , Vírus Bluetongue/genética , Proteínas do Capsídeo/genética , Cabras , Kuweit , Filogenia , RNA Viral/genética , Homologia de Sequência , Sorotipagem , OvinosRESUMO
Wildlife, especially Cape buffalo (Syncerus caffer), are thought to act as a reservoir for many of the important tick-borne pathogens of cattle. In this study, we have determined the prevalence of the most significant tick-borne haemoparasites in wildlife (buffalo, impala, eland and bushbuck) as well as in cattle grazing inside and neighbouring Lake Mburo National Park (LMNP) in Uganda. A high percentage of buffalo were carriers of Theileria parva, Theileria mutans, Theileria velifera, Theileria buffeli and Theileria sp. (buffalo) as well as Anaplasma marginale and Anaplasma centrale. The majority of impala sampled were carriers of A. centrale, and all were carriers of an unidentified Babesia/Theileria species. The eland and bushbuck sampled were all carriers of Theileria taurotragi and Theileria buffeli, and the majority were carriers of T. mutans. The bushbuck sampled were also carriers for Erhlichia bovis. There were some differences in the prevalence of haemoparasites between the calves sampled inside and neighbouring LMNP. In order to address the question of whether there is evidence for interbreeding between buffalo-associated and cattle-associated T. parva populations, multi-locus genotypes (MLGs) of T. parva (based on micro-satellite markers) from buffalo and from calves grazing inside and outside LMNP were compared, and the results revealed that buffalo and cattle gene pools were distinct, showing no evidence for transmission of buffalo-derived T. parva genotypes to the cattle population.
Assuntos
Búfalos/parasitologia , Doenças dos Bovinos/epidemiologia , Bovinos/parasitologia , Variação Genética , Theileria parva/genética , Theileriose/epidemiologia , Animais , Doenças dos Bovinos/parasitologia , Genótipo , Insetos Vetores/parasitologia , Reação em Cadeia da Polimerase , Prevalência , Theileriose/parasitologia , Doenças Transmitidas por Carrapatos/epidemiologia , Doenças Transmitidas por Carrapatos/parasitologia , Carrapatos , Uganda/epidemiologiaRESUMO
The live infection and treatment (ITM) vaccination procedure using the trivalent Muguga cocktail is increasingly being used to control East Coast fever, with potential implications for Theileria parva population genetic structure in the field. Transmission of the Kiambu V T. parva component to unvaccinated cattle has previously been described in Uganda. We monitored the T. parva carrier state in vaccinated and control animals on a farm in West Kenya where an ITM stabilate derived from the Kenyan T. parva Marikebuni stock was evaluated for field efficacy. A nested PCR-based Marikebuni-specific marker identified a carrier state in nine of ten vaccinated animals, detectable for a period of two years. We used 22 variable number tandem repeat (VNTR) markers to determine multilocus genotypes (MLGs) of 19 T. parva schizont-infected lymphocyte isolates derived from cattle and field ticks. Two isolates from unimmunized cattle were identical to the Marikebuni vaccination stock. Two cattle isolates were identical to a Muguga cocktail component Kiambu V. Seven isolates from ticks exhibited MLGs that were identical to the Serengeti/Muguga vaccine stocks. Six cattle and two tick-derived stocks exhibited unique MLGs. The data strongly suggest transmission of immunizing genotypes, from Marikebuni vaccine-induced carrier cattle to unimmunized cattle. It is possible that genotypes similar to those in the Muguga cocktail are present in the field in Western Kenya. An alternative hypothesis is that these parasites may have originated from vaccine trial sites in Eastern Uganda. If correct, this suggests that T. parva stocks used for immunization can potentially be disseminated 125 km beyond the immediate vaccination site. Regardless of their origin, the data provide evidence that genotypes similar to those in the Muguga cocktail are circulating in the field in East Africa, alleviating concerns about dissemination of 'alien' T. parva germplasm through live vaccination.
Assuntos
Doenças dos Bovinos/parasitologia , Imunização/veterinária , Theileria parva/genética , Theileriose/parasitologia , Doenças Transmitidas por Carrapatos/parasitologia , Carrapatos/parasitologia , Vacinação/veterinária , Animais , Bovinos , Doenças dos Bovinos/prevenção & controle , Doenças dos Bovinos/transmissão , Genótipo , Quênia/epidemiologia , Tipagem de Sequências Multilocus/veterinária , Reação em Cadeia da Polimerase/veterinária , Theileria parva/imunologia , Theileriose/prevenção & controle , Theileriose/transmissão , Doenças Transmitidas por Carrapatos/prevenção & controle , Doenças Transmitidas por Carrapatos/transmissão , Uganda , Vacinas Atenuadas/imunologiaRESUMO
Control of African swine fever (ASF) in countries in Eastern, Central and Southern Africa (ECSA) is particularly complex owing to the presence of all three known epidemiological cycles of maintenance of the virus, namely an ancient sylvatic cycle involving the natural hosts and vectors of the disease as well as domestic cycles with and without involvement of natural vectors. While the situation is well documented in some of the countries, for others very little information is available. In spite of the unfavourable ASF situation, the pig population in the sub-region has grown exponentially in recent decades and is likely to continue to grow in response to rapid urban growth resulting in increasing demand for animal protein by populations that are no longer engaged in livestock production. Better management of ASF will be essential to permit the pig sector to reach its full potential as a supplier of high quality protein and a source of income to improve livelihoods and create wealth. No vaccine is currently available and it is likely that, in the near future, the sub-region will continue to rely on the implementation of preventive measures, based on the epidemiology of the disease, to avoid both the devastating losses that outbreaks can cause and the risk the sub-region poses to other parts of Africa and the world. The current situation in the ECSA sub-region is reviewed and gaps in knowledge are identified in order to support ongoing strategy development for managing ASF in endemic areas.
Assuntos
Febre Suína Africana , Surtos de Doenças/veterinária , África Central/epidemiologia , África Oriental/epidemiologia , África Austral/epidemiologia , Febre Suína Africana/epidemiologia , Febre Suína Africana/prevenção & controle , Febre Suína Africana/transmissão , Animais , SuínosRESUMO
African swine fever virus (ASFV) infection usually results in an acute haemorrhagic disease with a mortality rate approaching 100% in domestic pigs. However, pigs can survive infection with less-virulent isolates of ASFV and may become chronically infected. Surviving animals are resistant to challenge with homologous or, in some cases, closely related isolates of the virus indicating that pigs can develop protective immunity against ASFV. During asymptomatic, non-virulent ASFV infections natural killer cell activity increases in pigs, suggesting this cell type plays a role in ASFV immunity. Furthermore, depletion of CD8(+) lymphocytes from ASFV immune pigs demolishes protective immunity against related virulent viruses. This suggests that ASFV specific antibody alone is not sufficient for protection against ASFV infection and that there is an important role for the CD8(+) lymphocyte subset in ASFV protective immunity. These results were supported by DNA immunization studies, demonstrating a correlation between the protection afforded against lethal challenge and the detection of a large number of vaccine-induced antigen-specific CD8(+) T-cells. Peripheral blood mononuclear cells (PBMCs) from ASF immune pigs protected from clinical disease show higher proportions of ASFV specific CD4(+)CD8(high+) double positive cytotoxic T cells than PBMCs from ASF immune but clinically diseased pig. The frequency of ASFV specific IFNγ producing T cells induced by immunization correlates to the degree of protection from ASFV challenge, and this may prove to be a useful indicator of any potential cross-protection against heterologous ASFV isolates.
Assuntos
Vírus da Febre Suína Africana/imunologia , Febre Suína Africana/imunologia , Imunidade Celular , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , DNA Viral/química , DNA Viral/genética , Interferon gama/metabolismo , Células Matadoras Naturais/imunologia , Dados de Sequência Molecular , Análise de Sequência de DNA , Suínos , Linfócitos T Citotóxicos/imunologia , Proteínas Virais/genética , Proteínas Virais/imunologiaRESUMO
Lumpy skin disease (LSD) is an economically important acute or sub-acute disease of cattle that occurs across Africa and in the Middle East. The aim of this study was to assess whether Rhipicephalus decoloratus ticks were able to transmit lumpy skin disease virus (LSDV) transovarially. Uninfected, laboratory-bred R. decoloratus larvae were placed to feed on experimentally infected "donor" cattle. After completion of the life cycle on donor animals, fully engorged adult female ticks were harvested and allowed to lay eggs. Larvae that hatched from these eggs were then transferred to feed on uninfected "recipient" cattle. The latter became viraemic and showed mild clinical disease with characteristic skin lesions and markedly enlarged precrural and subscapular lymph nodes. This is the first report of transovarial transmission of poxviruses by R. decoloratus ticks, and the importance of this mode of transmission in the spread of LSDV in endemic settings requires further investigation.
Assuntos
Doença Nodular Cutânea/transmissão , Vírus da Doença Nodular Cutânea/fisiologia , Rhipicephalus/classificação , Rhipicephalus/virologia , Animais , Bovinos , Feminino , Larva/virologia , Doença Nodular Cutânea/virologiaRESUMO
Capripoxviruses, which are endemic in much of Africa and Asia, are the aetiological agents of economically devastating poxviral diseases in cattle, sheep and goats. The aim of this study was to validate a high-throughput real-time PCR assay for routine diagnostic use in a capripoxvirus reference laboratory. The performance of two previously published real-time PCR methods were compared using commercially available reagents including the amplification kits recommended in the original publication. Furthermore, both manual and robotic extraction methods used to prepare template nucleic acid were evaluated using samples collected from experimentally infected animals. The optimised assay had an analytical sensitivity of at least 63 target DNA copies per reaction, displayed a greater diagnostic sensitivity compared to conventional gel-based PCR, detected capripoxviruses isolated from outbreaks around the world and did not amplify DNA from related viruses in the genera Orthopoxvirus or Parapoxvirus. The high-throughput robotic DNA extraction procedure did not adversely affect the sensitivity of the assay compared to manual preparation of PCR templates. This laboratory-based assay provides a rapid and robust method to detect capripoxviruses following suspicion of disease in endemic or disease-free countries.
Assuntos
Capripoxvirus/isolamento & purificação , DNA Viral/isolamento & purificação , Ensaios de Triagem em Larga Escala/métodos , Infecções por Poxviridae/veterinária , Reação em Cadeia da Polimerase em Tempo Real/métodos , Medicina Veterinária/métodos , Virologia/métodos , Animais , Automação/métodos , Capripoxvirus/genética , Bovinos , Doenças dos Bovinos/diagnóstico , Doenças dos Bovinos/virologia , DNA Viral/genética , Doenças das Cabras/diagnóstico , Doenças das Cabras/virologia , Cabras , Infecções por Poxviridae/diagnóstico , Sensibilidade e Especificidade , Ovinos , Doenças dos Ovinos/diagnóstico , Doenças dos Ovinos/virologiaRESUMO
Epizootic hemorrhagic disease virus (EHDV), an arthropod-borne orbivirus (family Reoviridae), is an emerging pathogen of wild and domestic ruminants that is closely related to bluetongue virus (BTV). The present study examines the outcome of an experimental EHDV-7 infection of Holstein cattle and East Frisian sheep. Apart from naïve animals that had not been exposed to BTV, it included animals that had been experimentally infected with either BTV-6 or BTV-8 two months earlier. In addition, EHDV-infected cattle were subsequently challenged with BTV-8. Samples were tested with commercially available ELISA and real-time RT-PCR kits and a custom NS3-specific real-time RT-PCR assay. Virus isolation was attempted in Vero, C6/36 and KC cells (from Culicoides variipennis), embryonated chicken eggs and type I interferon receptor-deficient IFNAR(-/-) mice. EHDV-7 productively infected Holstein cattle, but caused no clinical signs. The inoculation of East Frisian sheep, on the other hand, apparently did not lead to a productive infection. The commercial diagnostic kits performed adequately. KC cells proved to be the most sensitive means of virus isolation, but viremia was shorter than 2 weeks in most animals. No interference between EHDV and BTV infection was observed; therefore the pre-existing immunity to some BTV serotypes in Europe is not expected to protect against a possible introduction of EHDV, in spite of the close relation between the viruses.
Assuntos
Doenças dos Bovinos/virologia , Doenças Transmissíveis Emergentes/veterinária , Vírus da Doença Hemorrágica Epizoótica/isolamento & purificação , Infecções por Reoviridae/veterinária , Doenças dos Ovinos/virologia , Animais , Vírus Bluetongue , Bovinos , Doenças dos Bovinos/diagnóstico , Linhagem Celular , Ensaio de Imunoadsorção Enzimática , Europa (Continente) , Vírus da Doença Hemorrágica Epizoótica/classificação , Vírus da Doença Hemorrágica Epizoótica/genética , Vírus da Doença Hemorrágica Epizoótica/crescimento & desenvolvimento , Infecções por Reoviridae/diagnóstico , Infecções por Reoviridae/virologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Ovinos , Doenças dos Ovinos/diagnóstico , Carneiro DomésticoRESUMO
A loop-mediated isothermal amplification (LAMP) assay was developed for the detection of African swine fever virus (ASFV). This assay targets the topoisomerase II gene of ASFV and its specificity was confirmed by restriction enzyme digestion of the reaction products. The analytical sensitivity of this ASFV LAMP assay was at least 330 genome copies, and the test was able to detect representative isolates of ASFV (n=38) without cross-reacting with classical swine fever virus. The performance of the LAMP assay was compared with other laboratory tests used for ASF diagnosis. Using blood and tissue samples collected from pigs experimentally infected with ASFV (Malawi isolate), there was good concordance between the LAMP assay and real-time PCR. In addition to detecting the reaction products using either agarose gels or real-time PCR machines, it was possible to visualise dual-labelled biotin and fluorescein ASFV LAMP amplicons using novel lateral flow devices. This assay and detection format represents the first step towards developing a practical, simple-to-use and inexpensive molecular assay format for ASF diagnosis in the field which is especially relevant to Africa where the disease is endemic in many countries.
Assuntos
Vírus da Febre Suína Africana/isolamento & purificação , Febre Suína Africana/diagnóstico , DNA Viral/isolamento & purificação , Técnicas de Amplificação de Ácido Nucleico/métodos , Vírus da Febre Suína Africana/genética , Animais , Vírus da Febre Suína Clássica/genética , Reações Cruzadas , DNA Topoisomerases Tipo II/genética , DNA Viral/genética , Sensibilidade e Especificidade , Suínos , Temperatura , Proteínas Virais/genéticaRESUMO
In mid September 2008, clinical signs of bluetongue (particularly coronitis) were observed in cows on three different farms in eastern Netherlands (Luttenberg, Heeten, and Barchem), two of which had been vaccinated with an inactivated BTV-8 vaccine (during May-June 2008). Bluetongue virus (BTV) infection was also detected on a fourth farm (Oldenzaal) in the same area while testing for export. BTV RNA was subsequently identified by real time RT-PCR targeting genome-segment (Seg-) 10, in blood samples from each farm. The virus was isolated from the Heeten sample (IAH "dsRNA virus reference collection" [dsRNA-VRC] isolate number NET2008/05) and typed as BTV-6 by RT-PCR targeting Seg-2. Sequencing confirmed the virus type, showing an identical Seg-2 sequence to that of the South African BTV-6 live-vaccine-strain. Although most of the other genome segments also showed very high levels of identity to the BTV-6 vaccine (99.7 to 100%), Seg-10 showed greatest identity (98.4%) to the BTV-2 vaccine (RSAvvv2/02), indicating that NET2008/05 had acquired a different Seg-10 by reassortment. Although Seg-7 from NET2008/05 was also most closely related to the BTV-6 vaccine (99.7/100% nt/aa identity), the Seg-7 sequence derived from the blood sample of the same animal (NET2008/06) was identical to that of the Netherlands BTV-8 (NET2006/04 and NET2007/01). This indicates that the blood contained two different Seg-7 sequences, one of which (from the BTV-6 vaccine) was selected during virus isolation in cell-culture. The predominance of the BTV-8 Seg-7 in the blood sample suggests that the virus was in the process of reassorting with the northern field strain of BTV-8. Two genome segments of the virus showed significant differences from the BTV-6 vaccine, indicating that they had been acquired by reassortment event with BTV-8, and another unknown parental-strain. However, the route by which BTV-6 and BTV-8 entered northern Europe was not established.
Assuntos
Vírus Bluetongue/isolamento & purificação , Bluetongue/diagnóstico , Genoma Viral/genética , Animais , Bovinos , Países Baixos , Filogenia , Análise de Sequência de RNA , VacinasRESUMO
Real-time, reverse transcription polymerase chain reaction (rRT-PCR) has become one of the most widely used methods in the field of molecular diagnostics and research. The potential of this format to provide sensitive, specific and swift detection and quantification of viral RNAs has made it an indispensable tool for state-of-the-art diagnostics of important human and animal viral pathogens. Integration of these assays into automated liquid handling platforms for nucleic acid extraction increases the rate and standardisation of sample throughput and decreases the potential for cross-contamination. The reliability of these assays can be further enhanced by using internal controls to validate test results. Based on these advantageous characteristics, numerous robust rRT-PCRs systems have been developed and validated for important epizootic diseases of livestock. Here, we review the rRT-PCR assays that have been developed for the detection of five RNA viruses that cause diseases that are notifiable to the World Organisation for Animal Health (OIE), namely: foot-and-mouth disease, classical swine fever, bluetongue disease, avian influenza and Newcastle disease. The performance of these tests for viral diagnostics and disease control and prospects for improved strategies in the future are discussed.
Assuntos
Bluetongue/diagnóstico , Peste Suína Clássica/diagnóstico , Febre Aftosa/diagnóstico , Influenza Aviária/diagnóstico , Doença de Newcastle/diagnóstico , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária , Animais , Notificação de Doenças , Agências Internacionais , Aves Domésticas/virologia , Ovinos/virologia , Suínos/virologiaRESUMO
During 2006 the first outbreak of bluetongue ever recorded in northern Europe started in Belgium and the Netherlands, spreading to Luxemburg, Germany and north-east France. The virus overwintered (2006-2007) reappearing during May-June 2007 with greatly increased severity in affected areas, spreading further into Germany and France, reaching Denmark, Switzerland, the Czech Republic and the UK. Infected animals were also imported into Poland, Italy, Spain and the UK. An initial isolate from the Netherlands (NET2006/04) was identified as BTV-8 by RT-PCR assays targeting genome segment 2. The full genome of NET2006/04 was sequenced and compared to selected European isolates, South African vaccine strains and other BTV-8 strains, indicating that it originated in sub-Saharan Africa. Although NET2006/04 showed high levels of nucleotide identity with other 'western' BTV strains, it represents a new introduction and was not derived from the BTV-8 vaccine, although its route of entry into Europe has not been established.
Assuntos
Vírus Bluetongue/classificação , Vírus Bluetongue/genética , Genoma Viral , RNA Viral/genética , Animais , Sequência de Bases , Bluetongue/virologia , Vírus Bluetongue/imunologia , Vírus Bluetongue/isolamento & purificação , Proteínas do Capsídeo/genética , Europa (Continente)/epidemiologia , Dados de Sequência Molecular , Países Baixos/epidemiologia , RNA de Cadeia Dupla/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência , SorotipagemRESUMO
The apicomplexan parasite, Theileria annulata, dedifferentiates and induces continuous division of infected bovine myeloid cells. Re-expression of differentiation markers and a loss of proliferation occur upon treatment with buparvaquone, implying that parasite factors actively maintain the altered status of the infected cell. The factors that induce this unique transformation event have not been identified. However, parasite polypeptides (TashAT family) that are located in the infected leucocyte nucleus have been postulated to function as modulators of host cell phenotype. In this study differential RNA display and proteomic analysis were used to identify altered mRNA and polypeptide expression profiles in a bovine macrophage cell line (BoMac) transfected with TashAT2. One of the genes identified by differential display was found to encode an ubiquitin-like protease (bUBP43) belonging to the UBP43 family. The bUBP43 gene and the gene encoding its ubiquitin-like substrate, bISG15, were expressed at a low level in T. annulata-infected cells. However, infected cells were refractory to induction of elevated bISG15 expression by lipopolysaccharide or type 1 interferons while TashAT2-transfected cells showed no induction when treated with camptothecin. Modulation of the ISGylation system may be of relevance to the establishment of the transformed infected host cell, as ISGylation is associated with resistance to intracellular infection by pathogens, stimulation of the immune response and terminal differentiation of leukaemic cells.