Newcastle disease

Many species of bird are susceptible to Newcastle disease virus (NDV). In addition to the domestic avian species, natural or experimental infection with NDV has been demonstrated in at least 236 species from 27 of the 50 orders of birds. It is generally agreed that aquatic birds such as ducks and geese are least susceptible to NDV (Bolte et al., 2001), while the most susceptible are chickens, and gregarious birds forming temporary or permanent flocks (Kaleta and Baldauf, 1988; Kouwenhoven, 1993). The breeds used in the poultry industry are equally susceptible, only some indigenous breeds are less susceptible (Kouwenhoven, 1993).

APMV-2 viruses are mainly found in passerines and turkeys as primary hosts, and furthermore in chickens, psittacines and rails. APMV-3 viruses are primary isolated from turkeys (see table).

Table: Avian paramyxoviruses (APMVs), main hosts and disease*

*adapted from Alexander, 1993, Alexander, 1997, Kouwenhoven, 1993.

** these APMVs have also been isolated from captive cage birds (Alexander, 1993)

Newcastle disease (ND) is endemic in many countries of the world. However, some European countries have been free of the disease for years (OIE, 2000).

The first outbreaks of ND are thought to have occurred in 1926 in Java, Indonesia and in Newcastle-upon-Tyne, UK (Alexander, 1997). Different clinical representations of the disease gave rise to different names, such as pseudo-fowl pest, avian distemper or avian pneumo-encephalitis, until serological evaluations revealed that these syndromes were due to the same causative agent (Alexander, 1997). It is speculated that the first panzootic spread of NDV started in the 1920s in Southeast Asia, and took about 30 years to spread around the globe. A second panzootic may have begun in the Middle East in the late 1960s and global spread occurred much more rapidly due to the intensification of the poultry industry worldwide, reaching most countries in about a decade. Imported NDV-carrying psittacine birds captured in the wild are associated with this second panzootic of ND. A third panzootic spread of ND, around the late 1970s, has been related to pigeons and doves (Columba livia) that were kept for leisure. It reached Europe in the 1980s (Alexander, 1997).

International reporting and recording of ND is carried out by the Food and Agricultural Organisation (FAO) of the United Nations. Due to the widespread use of vaccines, the true prevalence of the disease is difficult to assess, but ND is still widespread in many countries in Asia, Africa, and the Americas (Marin et al., 1996). Only countries of Oceania, and some in Europe, are relatively free of the disease. During the 1980s most countries in Europe remained free of ND, but since 1991 many outbreaks have occurred in Western Europe. There have been outbreaks in Belgium, Scandinavia, Denmark, The Netherlands, Luxembourg, Germany, Switzerland, Italy, Spain, Portugal, Malta, the UK and France (Alexander, 1997; Alexander et al., 1998; Lomniczi et al., 1998; OIE, 2005; OIE, 2006). Importantly, outbreaks tend to occur increasingly in backyard flocks, rather than in large commercial flocks. This makes it even more difficult to control and prevent new outbreaks, and studies of backyard birds are scarce (Gutierrez-Ruiz et al., 2000).

For current information on disease incidence, see OIE's WAHID Interface.

Thirty one African countries covering west, east and southern Africa regions reported ND to the AU-IBAR in 2011. Overall, the disease affected a total of 1,031 epidemiological units involving 487,206 cases and 326,706, with a case fatality rate of 67.1% (see table below). The three countries with the highest number of outbreaks are Ghana (216), Benin (152) and Uganda (120). Sierra Leon and Liberia reported ND for the first time in 2010 and continued reporting in 2011 indicating the impacts of capacity building programs provided through SPINAP and VACNADA projects for improving disease surveillance and reporting. Generally, all other countries have consistently reported ND during the past four years, consistent with the known endemicity of the disease on the African continent.

Countries reporting ND to AU-IBAR in 2011

NS: Not specified

There appears to be no temporal trend for ND occurrence on the continent, suggesting the lack of seasonality for the risk factors that determine occurrence and maintenance of the disease.

Clinical diagnosis can be made upon respiratory or nervous signs. Birds may suddenly cough, gasp or may droop their wings. The animals may twist their head or neck, or show ataxia or paralysis. Diagnosis may also be made when egg production has decreased or completely stopped, and eggs are misshapen, rough-shelled, thin-shelled and contain watery albumen. Also when birds show greenish diarrhoea, and tissues around the neck are oedematous and swollen, NV may be suspected. However, a presumptive diagnosis of ND must always be confirmed by virus isolation and identification.

Laboratory diagnosis depends on the detection of the agent, because the widespread use of vaccines hampers the interpretation of serological results (OIE, 2008). Although direct detection of NDV antigen by immunohistological techniques may unequivocally reveal the presence of NDV, these methods, including immunofluorescence, impression smears or immunoperoxidase techniques, do not allow further characterization of the virus and are less sensitive than virus isolation. Therefore, virus isolation (VI) is preferred. Inoculation of specific-pathogen-free (SPF) or NDV-antibody free embryonated chicken eggs, incubated 9-11 days before use, is the most sensitive method for isolation of NDV (Kouwenhoven, 1993; Alexander, 1997). Samples to be used for VI are preferentially cloacal swabs or faeces of dead birds, intestinal contents, tracheal material or bone marrow. Whole carcasses should preferably be submitted, without freezing and thawing (Kouwenhoven, 1993; Alexander, 1997). About 0.2 ml supernatant of a clarified 20% w/v suspension of pooled organs containing antibiotics (10 minutes 1000 x g) is injected in the allantoic cavity of at least five embryonated eggs. The embryos frequently die within 3-6 days after inoculation and are subsequently tested for haemagglutinating activity. HI-positive isolates should be further typed with Avian paramyxovirus (APMV)-monospecific sera. In particular APMV-1 should be distinguished from APMV-3 and -7 (OIE, 2000). Blind passaging is seldom required (Alexander, 1997; Kouwenhoven, 1993; OIE, 2000).

The polymerase chain reaction (PCR) is frequently used to detect, and quantifiy, NDV (Fuller et al., 2009; Jang et al., 2011) in conjunction with virus isolation and biological characterization for index cases.

Virulence is traditionally determined using in vivo pathogenicity tests to distinguish between high, moderate and low virulence isolates (OIE, 2008). The pathogenicity of any newly isolated virus can be assessed by determining the mean death-time in eggs, the intracerebral pathogenicity index in 1-day old chickens or by the intravenous pathogenicity index in 6-week old chickens. Increasingly the polymerase chain reaction (PCR), in which the fusion protein (F0) gene is amplified followed by sequencing of that part of the F0 gene that encodes the cleavage site (between the F2 and F1 polypeptides) has been used to differentiate virulent from non-virulent strains (Kant et al., 1997; Nanthakumar et al., 2000; OIE, 2000; Aldous et al., 2001; Al-Garib et al., 2003). At the position where the F0 polypeptide is cleaved, to produce F2 and F1 polypeptides, there are a number of basic amino acid residues (arginine, R, and/or lysine, K) at the C-terminus of the F2 polypeptide. Most NDVs that are pathogenic for chickens have the sequence 112R/K-R-Q-K/R-R116 at the C-terminus of the F2 protein i.e. at least one pair of basic amino acids, whereas viruses of low virulence have the sequence 112G/E-K/R-Q-G/E-R116 i.e. the basic amino acids only occur singly. Furthermore, the virulent and non-virulent strains have a phenylalanine or leucine residue at position 117 (start of the F1 polypeptide), respectively.

PCR can be used to detect the virus in clinical specimens as well as following the growth of virus in embryos in the laboratory, the advantage being the extremely rapid demonstration of the presence of virus and even its virulence if primers covering the part of the genome coding for the F0 cleavage site are used (Creelan et al., 2002). However, some studies have shown lack of sensitivity of RT-PCR in detecting virus in some organs and particularly in faeces (Creelan et al., 2002). Multiplex PCR tests have been developed to allow simultaneous detection and differentiation of several avian viruses, such as NDV, avian pneumovirus and avian influenza (Malik et al., 2004). Multiplex PCR techniques have also been used experimentally to differentiate between velogenic, mesogenic and lentogenic strains from chickens (Shan et al., 2003). Real-time RT-PCR techniques have been developed in which detection is faster than conventional RT-PCR, partly because of automation and in part because the PCR product is detected during the actual PCR reaction; a separate detection stage is not required (Jang et al., 2011). Fuller et al. (2009) have developed a real time PCR which simultaneously detects and pathotypes strains of NDV. As with virulence determination, it is important that molecular techniques alone are not used to record a negative result in investigations of suspected ND (OIE, 2004f). Indeed, isolation of virus and assessment of pathogenicity in vivo is an international obligatory requirement at the start of each outbreak.

Table: Methods to determine the pathogenicity of NDV isolates*

* adapted from Anonymous, 2000, ** there seems to be overall the requirement of at least one pair of basic amino acids at residues 116 and 115 plus a phenylalanine at residue 117, and a basic amino acid at (R) at 113 if the virus is to show virulence in chickens (OIE, 2000).

As an example, the Ulster NDV isolate is an avirulent NDV representative with MDT, IVPI and ICPI values of 0. The well known lentogenic LaSota NDV strain has MDT, IVPI and ICPI values of 103, 0 and 0.15, respectively. An example of an extremely virulent strain is the Herts ‘33 NDV strain. However, there is no consequent correlation between the MDT, IVPI and ICPI, and interpretation may be difficult. Notably, the MDT is imprecise in particular for strains of low virulence. The IVPI is particularly useful for classifying moderately and highly virulent NDV isolates (Kouwenhoven, 1993).

Serological tests may be useful for diagnosis provided the vaccination status of the animals is known. Virus neutralisation, HI and ELISA tests are available, and new ELISAs have been described for use with future marker and subunit vaccines (Mackay et al., 1999). As with PCR, rapid field and multiplex versions of serological tests are being developed, for example an immunocomb-based dot-enzyme-linked immunosorbent test for detection of ND, infectious bursal disease and infectious bronchitis (Manoharan et al., 2004). At present, the HI test is most widely used. SPF chicken red blood cells are routinely used, with some variations in test procedures between laboratories. HI tests can also be used to assess the immune status of a flock (OIE, 2000).

Good management and hygiene remain the basis for prevention of ND, but in areas with an intensive poultry industry control of ND without vaccination is uncommon. Only in geographically isolated areas, with a very low risk of introduction of NDV and a relatively small economic impact of an outbreak, may vaccination be reserved for emergency or ring vaccinations (Kouwenhoven, 1993).

Vaccines available against NDV consist of live NDV strains of low virulence or inactivated strains, and recombinant vectored vaccines (OIE, 2008).

Among live avirulent strains Clone 30, Hitchner-B1, La Sota, Queensland V4, Poulvac NDW and F (Asplin) are used extensively worldwide as primary vaccines. Occasionally, new lentogenic field isolates are evaluated as vaccine candidates (Rehmani and Spradbrow, 1996; Murakawa et al., 2000). Live vaccines should contain = 10^6.5-10⁷, 50% egg infectious dose (EID50). Mesogenic strains used in vaccines include Roakin, Mukteswar and Komarov, and are used for secondary vaccinations (Agoha et al., 1992; Kouwenhoven, 1993; Alexander, 1997; Roy et al., 1999; OIE, 2000). The challenge for every vaccine manufacturer is the timing of vaccination, and the balance between attenuation and immunogenicity. For instance, in young birds vaccination with LaSota and Clone strains will usually lead to strong immunity and can overcome certain levels of maternal antibodies, but may also result in adverse effects, as opposed to the Hitchner-B1 strain. For inactivated vaccines, a high virus yield is important to produce a potent vaccine and the Ulster 2C strain has proven very suitable for this purpose (OIE, 2000). The potency of inactivated vaccines is greatly dependent on the antigen dose (Maas et al., 1999; Maas et al., 2000), and it is proposed that inactivated vaccines should preferably contain 50 x 50% protective doses (PD50) per 0.5 ml dose (Kouwenhoven, 1993). A combination of live and inactivated ND vaccine, administered simultaneously, is shown to provide better protection against virulent NDV and has been successfully used in control programmes in areas of intense poultry production (Senne et al., 2004).

Immunosuppression, notably after infectious bursal disease infection or vaccination, may reduce the response to ND vaccination (Montgomery et al., 1997). However, some IBD and ND vaccines may provide good immunity after simultaneous vaccination (Kouwenhoven, 1993). The effect of ND on E. coli infections is unclear. It has been reported that NDV vaccination stimulates innate immunity, suppressing the multiplication of E. coli in chickens for a period of 2-8 days post vaccination (Huang and Matsumoto, 2000). However, there are also studies suggesting that ND vaccination may enhance an E. coli infection, if both occur simultaneously (Nakamura et al., 1992).

Live vaccines may be administered by eye drop or intranasal instillation, spray, or drinking water. The administration route has a significant effect on the induced immune response (Mutalib and Boyle, 1994). Eye drop and intranasal installation is laborious, but is more likely to lead to a uniformly high degree of long lasting protection. Furthermore, maternal antibodies do not interfere at the mucosal surfaces of the nose, the Harderian and paranasal glands. Vaccination by aerosol, using sprays, can be administered in coarse or fine droplets. Coarse droplets are bigger and birds must be hit directly. This spray method is usually performed using hand-sprayers, knapsack sprayers or spray cabinets in hatcheries. The ‘Atomist’ atomizer is frequently used for fine-droplet spraying (Kouwenhoven, 1993). One should consider that, for successful immunization using these administration routes, sufficient virus replication is required in the upper respiratory tract, which is a characteristic of many lentogenic isolates. Vaccination via the drinking water gives heterogeneous results due to the variations in water intake by the birds. Essentially, intestinal replication is required for this type of administration. After aerosol vaccination with live lentogenic vaccines clinical signs are inevitable. Coughing, wet tracheas and incidental plugging of the bifurcation of the trachea can occur. Adverse reactions are usually less significant after vaccination via eyedrop or drinking water. Also, lighter breeds such as white layers are more prone to adverse reactions than heavier breeds. Research using both live and inactivated vaccines showed that the ocular route of administration was superior to the drinking water route, which was in turn superior to the spray technique. However, the ocular route may not be economically viable for small flocks (Degefa et al., 2004).

Inactivated vaccines are usually based on oil emulsions and administered by intramuscular or, more seldom, subcutaneous injection (Deguchi et al., 1998). The vaccine in a dose of approximately 0.5 ml per chicken is deposited in the breast or leg muscles. When injecting in the breast, care should be taken not to inject internal organs such as the heart, liver or lungs as this will cause rapid death of the bird. Inactivated vaccines are mainly used as secondary vaccinations at the end of the rearing period (Kouwenhoven, 1993).

The immune status of a flock can be determined using the level of HI antibodies, because there is a strong correlation between protection and HI antibody level in birds older than 6 weeks. This is the basis for the widespread use of blood testing to determine the proper vaccination schedule. Young birds with maternal antibodies, vaccinated at an early age, will respond with low or no rise in HI titre, and are protected less well and for a shorter period than older birds. Re-vaccinations can be timed using HI titres, because re-vaccination of flocks with high HI titres will not result in a titre rise, and the titre may even drop due to antibody consumption.

Because maternal antibodies protect well during the first week of life, vaccination of parent stock with inactivated vaccines is widely practised (van Eck, 1990). Subsequently, young chickens are vaccinated with a lentogenic strain the first week of life, which should provide protection during the next few weeks. Subsequent re-vaccinations should be timed based on the infection pressure and HI titre of the flock (Kouwenhoven, 1993).

Recombinant vectored vaccines against ND that are commercially available include herpesvirus of turkeys (HVT, a Marek’s disease vaccine) and fowl pox virus, both of which express a protective surface antigen of NDV. The HVT+ND vaccine can be given in ovo as well as to chicks.