Tracing the origins of genotype VIIh Newcastle disease in southern Africa.

It is widely accepted that Newcastle disease is endemic in most African countries, but little attention has been afforded to establishing the sources and frequency of the introductions of exotic strains. Newcastle disease outbreaks have a high cost in Africa, particularly on rural livelihoods. Genotype VIIh emerged in South-East Asia and has since caused serious outbreaks in poultry in Malaysia, Indonesia, southern China, Vietnam and Cambodia. Genotype VIIh reached the African continent in 2011, with the first outbreaks reported in Mozambique. Here, we used a combination of phylogenetic evidence, molecular dating and epidemiological reports to trace the origins and spread of subgenotype VIIh Newcastle disease in southern Africa. We determined that the infection spread northwards through Mozambique, and then into the poultry of the north-eastern provinces of Zimbabwe. From Mozambique, it also reached neighbouring Malawi and Zambia. In Zimbabwe, the disease spread southward towards South Africa and Botswana, causing outbreaks in backyard chickens in early-to-mid 2013. In August 2013, the disease entered South Africa's large commercial industry, and the entire country was infected within a year, likely through fomites and the movements of cull chickens. Illegal poultry trading or infected waste from ships and not wild migratory birds was the likely source of the introduction to Mozambique in 2011.

Seroprevalence of Newcastle disease and other infectious diseases in backyard chickens at markets in Eastern Shewa zone, Ethiopia.

This study was conducted to estimate the seroprevalence of Newcastle disease (ND), Pasteurella multocida (PM) infection, Mycoplasma gallisepticum (MG) infection, and infectious bursal disease (IBD) and to assess the level of concurrent seropositivity during the dry and wet seasons of the year 2010. In total, 234 and 216 sera were collected during the dry and wet seasons, respectively, from unvaccinated backyard chickens at 4 live poultry markets in 2 woredas (districts) of Eastern Shewa zone, Ethiopia, and were tested using commercial ELISA kits. The overall seroprevalence of ND, PM, MG, and IBD was 5.9, 66.2, 57.7, and 91.9%, respectively, during the dry season, and 6.0, 63.4, 78.7, and 96.3%, respectively, during the wet season. The seroprevalence of MG was higher (P < 0.001) during the wet season than during the dry season and higher (P = 0.002) in Adami-Tulu-Jido-Kombolcha woreda (74%) than in Ada'a woreda (60%). Area and season had no significant effect on the seroprevalence of ND, IBD, and PM, indicating the widespread presence of those pathogens throughout the year in the study area. Of all the chickens tested, 85.6% had antibodies concurrently to more than one of the pathogens investigated. Birds were concurrently seropositive to more diseases during the wet season (median = 3) than during the dry season (median = 2; P = 0.002). As serology is not able to distinguish between strains, further studies are warranted to better understand the circulating strains, their interactions, and their economic effect on backyard poultry production in Ethiopia.

Uterine adenocarcinoma with transcoelomic metastases in breeder hens (Gallus domesticus).

Hens involved in a Newcastle disease study were euthanased at regular intervals according to a designed protocol. Of these, 7.14 % (n = 42) of the 82-week-old specific pathogen-free breeder hens were found to have well-delineated firm white to yellowish nodules of varying sizes in the abdominal cavity. Histologically, the nodules were identified as an adenocarcinoma originating in the uterus. Transcoelomic spread was evidenced by the presence of similar neoplastic cells embedded in the serosa and outer longitudinal muscle layer of the intestines as well as the liver.

Lessons from Nigeria: the role of roads in the geo-temporal progression of avian influenza (H5N1) virus.

The daily progression of the 2006 (January-June) Nigerian avian influenza (AI H5N1) epidemic was assessed in relation to both spatial variables and the generation interval of the invading virus. Proximity to the highway network appeared to promote epidemic dispersal: from the first AI generation interval onwards > 20% of all cases were located at < 5 km from the nearest major road. Fifty-seven per cent of all cases were located 31 km from three highway intersections. Findings suggest that the spatial features of emerging infections could be key in their control. When the spatial location of a transmission factor is well known, such as that of the highway network, and a substantial percentage of cases (e.g. > 20%) are near that factor, early interventions focusing on transmission factors, such as road blocks that prevent poultry trade, may be more efficacious than interventions applied only to the susceptible population.

Molecular characterization and epidemiology of the highly pathogenic avian influenza H5N1 in Nigeria.

Avian influenza caused infection and spread throughout Nigeria in 2006. Carcass samples (lung, liver, spleen, heart, trachea and intestine) from the different regions of Nigeria were processed for virus isolation. Infective allantoic fluids were tested for avian influenza viruses (AIV) and Newcastle disease virus using monospecific antisera. Thirty-five isolates were generated and characterized molecularly using the haemagglutinin gene. The molecular analysis indicated that different sublineages of the highly pathogenic avian influenza (HPAI) H5N1 viruses spread throughout Nigeria. We compared the Nigerian isolates with others from Africa and results indicated close similarities between isolates from West Africa and Sudan. Some of the analysed viruses showed genetic drift, and the implications of these for future epidemiology and ecology of avian influenza in Africa require further evaluation. The spread of primary outbreaks was strongly linked to trade (legal and illegal), live bird markets, inappropriate disposal, and poorly implemented control measures. No strong correlation existed between wild birds and HPAI H5N1 in Nigeria.

Efficacy of a genotype 2 Newcastle disease vaccine (Avinew) against challenge with highly virulent genotypes 5d and 3d.

Since 2002, following its introduction, the lineage 5d Newcastle disease virus (so-called Goose paramyxovirus - GPMV) strain has caused numerous disease outbreaks among commercial and backyard poultry in South Africa, raising questions about the ability of commercially available Newcastle disease vaccines to fully protect poultry against the strain. This study aimed to determine whether there are differences in the level of protection offered by Avinew Newcastle disease vaccine against GPMV virus as compared with a 3d Newcastle disease virus isolated in South Africa in 1993 (Rainbow challenge virus - RCV) strain. Six groups of 10-day-old, specific pathogen-free chickens were vaccinated with doses of 10(3.0), 10(4.5) and 10(6.0) EID50 of Avinew vaccine and challenged at 4 weeks of age intramuscularly at a dose of 10(5.3) EID50/0.2 ml/bird of GPMV and RCV. No statistically significant difference could be found in the protection offered by Avinew vaccine against GPMV as compared to RCV challenge. The protection offered against the ND challenge was found to be dose dependent. At the recommended field dose of 10(6.0) EID50 the vaccine gave 100% protection from mortality against both the challenge viruses, but not against infection and replication of the viruses, as gross lesions were evident even in apparently healthy birds that survived the challenge. The protective dose (PD90) of the Avinew vaccine against GPMV challenge was calculated at 10(4.38) and against that of RCV at 10(4.43).

Characterization of pigeon paramyxoviruses (Newcastle disease virus) isolated in South Africa from 2001 to 2006.

Pigeon paramyxovirus type 1 (PPMV-1), a variant of Newcastle disease virus that primarily affects doves and pigeons has been isolated in South Africa since the mid-1980s. Phylogenetic evidence indicates that pigeon paramyxovirus type 1 viruses were introduced into South Africa on multiple occasions, based on the presence of two separate lineages, 4bi and 4bii, that have been circulating in Europe and the Far East since the early 1990s. During 2006, a PPMV-1 virus was isolated from an African ground hornbill (Bucorvus leadbeateri) which became acutely infected with PPMV-1 and died, probably after scavenging off infected dove carcasses in the region, since a closely-related PPMV-1 strain was also isolated from doves collected nearby. The hornbill isolate had ICPI and MDT values characteristic of PPMV-1 strains. The threat of PPMV-1 to poultry production and biodiversity in southern Africa highlights the importance of monitoring the spread of this strain.

A comparison of the oral application and injection routes using the onderstepoort biological products fowl typhoid vaccine, its safety, efficacy and duration of protection in commercial laying hens.

This study was undertaken to establish whether the Onderstepoort Biological Products Fowl Typhoid (OBPft) vaccine registered as an injectable vaccine was effective and safe when administered orally to commercial layers. Its efficacy and duration of protection were compared with application by intramuscular injection. Commercial brown layer hens were used as they were found to be highly susceptible to Salmonella gallinarum infections. In the vaccine safety trial birds were euthanased at timed intervals spanning 4 weeks postvaccination. Necropsies were performed and samples were taken and tested. No clinical signs or mortalities could be attributed to the OBPft vaccine nor could active shedding of the vaccine strain be detected. Slight pathological changes were noted with both routes of vaccination; however, these changes were transient, returning to normal within the observation period. The injected groups showed a better serological response with the rapid serum plate agglutination (RSPA) test than the orally vaccinated groups. In the duration of protection trial, birds were challenged at 3-8-week intervals post-vaccination. All unvaccinated birds died. Protection 8 and 16 weeks after vaccination was above 60 %,by 24 weeks after challenge, the vaccine protection was below 30 %. It was found that there was no significant difference (P < 0.05) in the protection offered by either the oral or injected route of vaccination with the OBPft vaccine.

The financial cost implications of the highly pathogenic notifiable avian influenza H5N1 in Nigeria.

Nigeria and several other nations have recently been affected by outbreaks of the Asian H5N1 strain of highly pathogenic notifiable avian influenza (HPNAI) virus, which affects the poultry sector most heavily. This study analysed previous methods of assessing losses due to avian influenza, and used a revised economic model to calculate costs associated with the current avian influenza outbreaks. The evaluation used epidemiological data, production figures and other input parameters to determine the final costs. An infection involving 10% of the commercial bird population will cost Nigeria about $245 million and a worse scenario may lead to a loss of around $700 million. The results urge governments to invest more in measures aimed at the effective prevention of HPNAI and to consider the huge economic losses associated with the disease. Finally, an inter-disciplinary approach to managing and controlling HPNAI outbreaks is encouraged.

The value of plant extracts with antioxidant activity in attenuating coccidiosis in broiler chickens.

Coccidiosis remains one of the most important diseases in the poultry industry and results in the annual loss of millions of US dollars by the poultry industry. In South Africa and other developing countries where a large percentage of the population is unemployed, cheap food production is necessary. If the control of the coccidian parasite could be made more economical, these savings could be passed on to the consumer. In Europe, where the economics are different, people are becoming more aware of the potential dangers of using antimicrobials in producing animal protein. A solution to both these problems could be the use of plant products that function by mechanisms other than those of chemotherapeutics, with the additional advantage of a natural origin. Antioxidant compounds could hold promise for the control of Eimeria infections due to the association of coccidial infection with lipid peroxidation of the intestinal mucosa. Four plant extracts with antioxidant activity were screened for their anticoccidial activity in vivo with toltrazuril as the positive control. Combretum woodii (160 mg/kg) proved to be extremely toxic to the birds, while treatment with Tulbaghia violacea (35 g/kg), Vitis vinifera (75 mg/kg) and Artemisia afra (150 mg/kg) resulted in feed conversion ratios similar to toltrazuril, and higher than the untreated control. T. violacea also significantly decreased the oocyst production in the birds. From this study we conclude that antioxidant-rich plant extracts have potential benefits in treating coccidial infections. The promising results obtained with T. violacea justify further studies on the potential value of the plant as a therapeutic or prophylactic anticoccidial agent.

Phylogenetic analysis of low-pathogenicity avian influenza H6N2 viruses from chicken outbreaks (2001-2005) suggest that they are reassortants of historic ostrich low-pathogenicity avian influenza H9N2 and H6N8 viruses.

Low-pathogenicity (LPAI) and high-pathogenicity (HPAI) avian influenza viruses are periodically isolated from South African ostriches, but during 2002 the first recorded outbreak of LPAI (H6N2) in South African chickens occurred on commercial farms in the Camperdown area of KwaZulu/Natal (KZN) Province. Sequence analysis of all eight genes were performed and phylogenetic analysis was done based on the hemagglutinin and neuraminidasc sequences. Results from phylogenetic analyses indicated that the H6N2 chicken viruses most likely arose from a reassortment between two South African LPAI ostrich isolates: an H9N2 virus isolated in 1995 and an H6N8 virus isolated in 1998. Two cocirculating sublineages of H6N2 viruses were detected, both sharing a recent common ancestor. One of these sublineages was restricted to the KZN province. The neuraminidase gene contained a 22-amino acid deletion in the NA-stalk region, which is associated with adaptation to growth in chickens, whereas the other group, although lacking the NA-stalk deletion, spread to commercial farms in other provinces. The persistence of particular H6N2 types in some regions for at least 2 yr supports reports from Asia and southern California suggesting that H6N2 viruses can form stable lineages in chickens. It is probable that the ostrich H6N8 and H9N2 progenitors of the chicken H6N2 viruses were introduced to ostriches by wild birds. Ostriches, in which AI infections are often subclinical, may serve as mixing vessels for LPAI strains that occasionally spill over into other poultry.

Evaluation of the application of a thermostable Newcastle disease vaccine by community volunteers in the North West Province of South Africa.

Participatory research on vaccination of village poultry against Newcastle disease (ND) was carried out in the village of Disaneng, in the North West Province of South Africa. Three application methods for ND Inkukhu vaccine were shown to induce sufficient levels of immunity in back-yard poultry when correctly administered. These are eye-droplet administration to individual fowls, in-feed and in-water administration to small flocks. After a community meeting and group discussion to select methods of vaccination, only 2 of the 3 methods were chosen; the individual administration of droplets into the eyes was considered to be too impractical because back-yard fowls are difficult to catch. Visual and practical training material was prepared and presented to volunteer vaccinators (n = 23). Vaccinators were then required to register all the poultry owners in their ward who wished to have poultry vaccinated. Once an indication of the number of chickens to be vaccinated had been made available, ND Nobilis Inkukhu vaccine was supplied to vaccinators free of charge. Community vaccinators were responsible for the organisation of the vaccination campaign, including storage and preparation of vaccine for application. All 9 wards in the village were initially involved with a total of 482 households, owning 6141 chickens, participating. This represented slightly in excess of 60 % of the fowls in the area. Involvement in a 2nd round of vaccinations, 1 month later, was far poorer with only 211 households owning a total of 1636 chickens participating. Serum samples were collected from vaccinated fowls using systematic random sampling and tested for circulating antibodies. The levels of protection varied, with no significant difference found between in-feed and in-water vaccine administration. Volunteer vaccinators were found to be unreliable, easily demotivated, did not keep good records and left the project when offered permanent employment. Contacting them to make arrangements for delivering vaccine was difficult and time consuming. Structured interviews indicated that deaths in poultry and the attitude of the owners probably contributed to the demotivation of the volunteers used as community vaccinators. It was concluded that volunteers are not the ideal choice for vaccination of village poultry against Newcastle disease.

Phylogenetic analyses of genes from South African LPAI viruses isolated in 2004 from wild aquatic birds suggests introduction by Eurasian migrants.

In 2004, South Africa experienced its first recorded outbreak of a highly pathogenic notifiable avian influenza (HPNAI) viral strain of the H5N2 subtype in ostriches in the Eastern Cape province. The traditional ostrich-farming areas in the Western Cape province report almost yearly outbreaks of low pathogenicity avian influenza (LPAI) in ostriches, which is attributed to introduction by wild birds and certain climatic patterns. During the winter of 2004, LPAI H3N8, H4N8, H5N2 and H5N1 avian influenza viruses were isolated from wild aquatic birds. All eight genes of the H3N8, H4N8 and H5N1 viruses were analysed. The results show that the H5N1 virus does not belong to the HPAI Z/Z+N genotype currently circulating in Asia, but that the most recent common ancestors are Russian H5N2 and H5N3 viruses. The N1 gene lacks the stalk deletion associated with virulence. Internal genes probably originate from a pool containing Chinese, Middle Eastern and Italian viruses. The South African H3N8 and H4N8 viruses appear to have derived their genes from an ecosystem where Asian H5N1, H6N9 and H9N2, Russian H4, and Danish H3N8 viruses have been circulating since 1997. All three viruses share recent nucleoprotein common ancestors with the German and Dutch HPNAI H7N7 viruses from 2003. The diverse pool of genes from which local viruses are derived suggests that reassortment occurred at the Siberian breeding grounds where migratory paths cross, or within the South African ecosystem. This data highlights the importance of surveillance in aquatic migratory birds, particularly members of the Charadriidae, for their potential roles in the introduction of avian diseases to South African poultry and especially ostriches in the case of avian influenza.

Floor pen study to evaluate the serological response of broiler breeders after vaccination with ts-11 strain Mycoplasma gallisepticum vaccine.

To determine the Mycoplasma gallisepticum (MG) rapid serum plate agglutination (RSPA) test response of broiler breeders after ts-11 strain vaccination, 55 Cobb pullets derived from a nonvaccinated, MG-negative, commercial, broiler breeder grandparent flock were monitored from 8 to 20 wk of age (over a 12-wk trial period). To evaluate the effect of lateral spread of the ts-11 vaccine strain on RSPA test results from commingled and adjacently penned birds, treatment groups included (A) birds vaccinated with ts-11strain MG at 8 wk of age, (B) commingled nonvaccinates in the same pen as the vaccinated birds, (C) nonvaccinates in a second pen separated from the first pen by a distance of 2 m, and (D) birds vaccinated with ts-11 strain MG at 8 wk of age and kept in a separate room. Rapid serum plate agglutination tests were performed once a week for 6 wk and then every 2 wk for 6 more wk, postvaccination. A polymerase chain reaction (PCR) assay specific fbr ts-11 strain MG was used to confirm vaccination, and a second PCR specific for non-ts-11 strain MG was used to confirm the absence of field infection. Seroconversion was first detected by the RSPA test 2 wk postvaccination and attained maximum positive rates of 58% at 12 wk postvaccination in treatment A and 60% at 8 wk postvaccination in treatment D. Seroconversion rates in nonvaccinated, commingled pullets was 10% at 5 wk and 30% at 12 wk after the vaccination of pen mates. The ts-11-specific PCR detected the vaccine strain in 80%-100% of the vaccinated birds 2 wk after vaccination. One of 15 nonvaccinated birds penned 2 m from vaccinated birds yielded ts-11 by PCR assay 12 wk after vaccination, which indicates that the spread of ts-11 over short distances may be possible in situations in which there is a common caretaker. PCR on tracheal swabs taken 12 wk postvaccination detected ts-l1 in 50% and 60% of the vaccinated birds in treatments A and D, respectively; in 30% of the commingled nonvaccinates; and in 6.6% of the separately penned nonvaccinates. In contrast, choanal swabs collected from vaccinated birds at 12 wk were 21% and 40% PCR positive for ts-11 strain MG, while those from nonvaccinates were negative. All samples were PCR negative for field strain MG. The pattern of seroconversion as measured by RSPA test in small groups of broiler breeders was different from that previously reported for leghorns. Lateral spread of the ts-11 strain to commingled pen mates occurred rapidly, causing RSPA seroconversion patterns that mimicked those of the vaccinated pen mates.

Different approaches to the vaccination of free ranging village chickens against Newcastle disease in Qwa-Qwa, South Africa.

The aim of this study was to develop a strategy to control Newcastle disease (ND) in free ranging village chickens using the Nobilis ND Inkukhu vaccine (Intervet South Africa). The study was conducted at Thibella village in Qwa-Qwa, South Africa from April 2001 to October 2002. Three different routes of vaccination (administration via eye-drop, drinking water and feed) were investigated. The haemagglutination inhibition (HI) test was conducted monthly in order to measure the antibody response of village chickens after immunization against Newcastle disease. Using a South African isolate of velogenic ND virus, challenge trials were conducted to determine the efficacy of the vaccine. A questionnaire was provided to evaluate perceptions of farmers on vaccinations. The eye-drop vaccination route produced the highest HI titres ranging between 2.7 and 4.4, followed by the drinking water vaccination route with titres ranging between 2.3 and 4.0. The lowest titres were from the feed vaccination route which ranged between 1.6 and 3.0. Following the challenge, the entire control group died on the third and fourth day after infection. However, 70% of the chickens immunized by using either the eye-drop or drinking water route survived the challenge. Only 20% of the chickens from the group immunized through the feed route survived. Evidently both the eye-drop and drinking water routes were efficient in preventing disease. Necropsies showed that vaccinated chickens had mild lesions whilst control chickens had severe lesions compatible with Newcastle disease. The efficacy of the vaccine using either of the routes can be enhanced by administration of booster vaccinations at 3-month intervals during the first year of a vaccination campaign and then at 6-month intervals from the second year onwards. The majority of the owners indicated that they would prefer to vaccinate their flocks using the drinking water route.

A phylogenetic study of South African Newcastle disease virus strains isolated between 1990 and 2002 suggests epidemiological origins in the Far East.

Genetic comparisons were made of the fusion protein sequences of 155 Newcastle disease virus isolates collected in South Africa between 1990 and 2002. Their evolutionary relationships and origins are described. All of the lentogenic field isolates were shown to be derived from commercial vaccines. No true South African lentogenic wild type strain was identified. Furthermore, it was shown that almost all mesogenic isolates had avirulent F(0) cleavage site sequences. Three major epizootics occurred in South Africa during the period of this study. The first outbreak (1990/1991) was caused by viruses endemic to South Africa since the 1960's (genotype VIII) but were occasionally also isolated in 2000. Genotype VIIb viruses, implicated in the severe outbreaks during 1993/1994, persisted until 1999. Genotype VIId viruses, responsible for the most recent outbreak in 1999/2000, had their origins in the Far East like those of the two previous outbreaks.

The use of a bacterin vaccine in broiler breeders for the control of Ornithobacterium rhinotracheale in commercial broilers.

Ornithobacterium rhinotracheale (ORT) is a recently identified bacterial pathogen of poultry, linked to the respiratory disease complex of broilers and the economic losses associated with that disease complex. Present control measures applied for the disease include the continuous use of in-feed antibiotics. A recently developed bacterin vaccine that is applied to broiler-breeder hens to pass on protective immunity to their broiler progeny was tested under large-scale commercial conditions in South Africa. An indirect ELISA test for antibodies to ORT, optimised for use in South Africa, was used to determine antibody levels in breeders and broilers. ELISA test results showed that the vaccine stimulated the development of high antibody titre levels in broiler breeders. The efficacy of the vaccine in protecting the progeny of these birds from ORT challenge could not be determined during the trial, although the progeny of vaccinated hens appeared to perform slightly better under commercial conditions than the progeny of unvaccinated hens.

Diseases of free-ranging chickens in the Qwa-Qwa District of the northeastern Free State province of South Africa.

A total of 177 free-ranging chickens from 19 Qwa-Qwa villages were bled from wing veins over a period of 6 months (June-November 2000). Serological tests indicated that 5% of chickens tested had been exposed to Newcastle disease, 43% to infectious bronchitis and 63% to Mycoplasma gallisepticum infection. McMaster and Visser sieve techniques were used to determine helminth and coccidia from pooled fresh faecal samples. Helminths isolated in 37% of the villages investigated were Heterakis, Ascaridia and Capillaria species. Eimeria species were also isolated in 32% of the villages investigated. The red fowl mite (Dermanyssus gallinae) was isolated from some of the birds and their nests. Data from a questionnaire survey indicated that all farmers interviewed had never received any technical support and that their chickens had never been vaccinated against any avian diseases. Only 10.5% of the owners interviewed had scientific knowledge on poultry diseases. There is an urgent need for the government to support free-ranging poultry farmers by providing subsidised vaccinations and technical support in order to develop and stimulate economic development in impoverished rural areas of South Africa.

The use of dried bakery products in a free-choice feeding method for small-scale broiler production.

Waste material from bakeries is an unconventional energy feed source which is available in sufficient quantities for use in small-scale broiler production in South Africa. Small-scale broiler producers do not have access to the computer programs required to balance home-mixed rations. This investigation confirms that the use of dried bakery products (DBP) in a 2-stage, free-choice method can be used to compensate for this lack. A total of 570 day-old male broiler chickens was assigned to 3 feeding treatments: the control group was fed a 2-stage feeding programme using standard commercial starter and grower rations. The 2nd group received a commercial starter ration up to Day 7 and was thereafter given a choice of a commercial starter ration with normal salt content (0.35%) and DBP. The 3rd group was fed a commercial starter ration up to Day 7, then offered a choice of commercial starter ration with a lower salt content (0.1%) and DBP. The low salt alternative was used to test whether the higher salt percentage in DBP influenced the choice of feed by the birds. It was found that the control group consumed significantly more feed (P < 0.05) and was significantly heavier (P < 0.05) than the experimental groups. However, there was no significant difference between the 2 experimental groups, which indicated that salt content did not play a role in the choice of ration. Feed consumption by both experimental groups was about one-third less than the control group, but the profit margin, as calculated using gross margin analysis, was approximately 15 % higher. It was therefore concluded that dried bakery products can be profitably incorporated as an energy feed source, using the free-choice feeding method, in small-scale broiler enterprises.