Risk factors for avian influenza and Newcastle disease in smallholder farming systems, Madagascar highlands.

Newcastle disease (ND) and avian influenza (AI) are issues of interest to avian producers in Madagascar. Newcastle disease virus (NDV) is the major constraint for village aviculture, and avian influenza viruses type A (AIAV) are known to circulate in bird flocks. This study aims at classifying smallholder poultry farms, according to the combination of risk factors potentially associated with NDV and AIAV transmission and to assess the level of infection for each farm class. Two study sites, Lake Alaotra and Grand Antananarivo, were chosen with respect to their differences in terms of agro-ecological features and poultry productions. A typology survey involving 526 farms was performed to identify possible risk factors for (i) within-village, and (ii) between-village virus transmission. A cross-sectional serological study was also carried out in 270 farms to assess sero-prevalences of NDV and AIAV for each farm class and the link between them and risk factor patterns. For within-village transmission, four classes of farms were identified in Grand Antananarivo and five in Lake Alaotra. For between-village virus transmission, four classes of farms were identified for each site. In both sites, NDV sero-prevalence was higher than for AIAV. There was no evidence of the presence of H5 or H7 subtypes of AIAV. Sero-prevalences were significantly higher in Lake Alaotra than in Grand Antananarivo for both viruses (OR=2.4, p=0.02 for NDV, and OR=9.6, p<0.0001 for AIAV). For within-village NDV transmission in Grand Antananarivo, backyard chicken farms (OR=3.6, p<0.001), and chicken farms with biosecurity awareness (OR=3.4, p<0.01) had greater odds of having antibodies against NDV than the others. For between-village virus transmission, farms with multiple external contacts, and farms using many small markets had greater odds of having antibodies against NDV than the others (OR=5.4, p<0.01). For AIAV, there were no differences in sero-prevalences among farm classes. In Lake Alaotra, the observed high density of palmipeds and widespread rice paddies were associated with high sero-prevalences for both viruses, and a homogeneous risk of virus transmission between the different farm classes. In Grand Antananarivo, farm visits by collectors or animal health workers, and farm contacts with several markets were identified as potential risk factors for NDV transmission. Further studies are needed to identify the circulating virus genotypes, model their transmission risk, and provide adapted control measures.

Identification of a natural multi-recombinant of Newcastle disease virus.

Newcastle disease (ND), caused by ND virus (NDV), is one of the most serious illnesses of birds, particularly chickens, and has been one of the major causes of economic losses in the poultry industry. Live vaccines are widely used to prevent chicken from NDV all over the world. Given the implications that recombination has for RNA virus evolution, it is clearly important to determine the extent to which recombination plays a role in NDV evolution. In this study, we performed the phylogenetic and recombination analysis on complete NDV genomes. A natural multi-recombinant cockatoo/Indonesia/14698/90 (AY562985) was identified. Its two minor parental-like strains might be from the NDV vaccine lineage and anhinga/U.S.(Fl)/44083/93 lineage, respectively. Our study suggests that recombination plays a role in NDV evolution. Especially, the study also suggests that live vaccines have capacity to play roles in shaping NDV evolution by homologous recombination with circulating virus.

The hemagglutinin-neuraminidase protein of Newcastle disease virus determines tropism and virulence.

The hemagglutinin-neuraminidase (HN) protein of Newcastle disease virus (NDV) plays a crucial role in the process of infection. However, the exact contribution of the HN gene to NDV pathogenesis is not known. In this study, the role of the HN gene in NDV virulence was examined. By use of reverse genetics procedures, the HN genes of a virulent recombinant NDV strain, rBeaudette C (rBC), and an avirulent recombinant NDV strain, rLaSota, were exchanged. The hemadsorption and neuraminidase activities of the chimeric viruses showed significant differences from those of their parental strains, but heterotypic F and HN pairs were equally effective in fusion promotion. The tissue tropism of the viruses was shown to be dependent on the origin of the HN protein. The chimeric virus with the HN protein derived from the virulent virus exhibited a tissue predilection similar to that of the virulent virus, and vice versa. The chimeric viruses with reciprocal HN proteins either gained or lost virulence, as determined by a standard intracerebral pathogenicity index test of chickens and by the mean death time in chicken embryos (a measure devised to classify these viruses), indicating that virulence is a function of the amino acid differences in the HN protein. These results are consistent with the hypothesis that the virulence of NDV is multigenic and that the cleavability of F protein alone does not determine the virulence of a strain.

Role of fusion protein cleavage site in the virulence of Newcastle disease virus.

Newcastle disease virus (NDV) causes a highly contagious and economically important disease in poultry. Viral determinants of NDV virulence are not completely understood. The amino acid sequence at the protease cleavage site of the fusion (F) protein has been postulated as a major determinant of NDV virulence. In this study, we have examined the role of F protein cleavage site sequence in NDV virulence using reverse genetics technology. The sequence G-R-Q-G-R present at the cleavage site of the F protein of avirulent strain LaSota was mutated to R-R-Q-K-R, which is present in the F cleavage site of neurovirulent strain Beaudette C (BC). The resultant mutated LaSota V.F. virus did not require exogenous protease for infectivity in cell culture, indicating that the F protein was cleaved by intracellular proteases. The virulence of the mutant and parental viruses was evaluated in vivo by intracerebral pathogenicity index (ICPI) and intravenous pathogenicity index (IVPI) tests in chickens. Our results showed that the modification of the F protein cleavage site resulted in a dramatic increase in virulence from an ICPI value of 0.00 for LaSota to a value of 1.12 for LaSota V.F. However, the ICPI value of LaSota V.F. was lower than that of BC, which had a value of 1.58. Interestingly, the IVPI tests showed values of 0.00 for both LaSota and LaSota V.F. viruses, compared to the IVPI value of 1.45 of BC. In vitro characteristics of the viruses were also studied. Our results demonstrate that the efficiency of cleavage of the F protein plays an important role if the NDV is delivered directly into the brains of chicks, but there could be other viral factors that probably affect peripheral replication, viremia, or entry into the central nervous system.

The interaction between vitamin A status and Newcastle disease virus infection in chickens.

Newcastle disease virus (NDV) infection in chickens differing in vitamin A status has been selected as a model to examine the interrelationship between marginal vitamin A deficiency and the severity of consequences of measles infection in humans. Day-old chickens with limited vitamin A reserves, the progeny of marginally vitamin A-deficient hens, were fed purified diets containing either marginal (120 retinol equivalents/kg diet, ad libitum) or adequate (1200 retinol equivalents/kg diet, ad libitum or pair-fed) levels of vitamin A for a period of 10 wk. At 4 wk of age, half of the chickens in each group were infected intraocularly with the lentogenic, i.e., mildly pathogenic, La Sota strain of NDV. Within 1 wk of infection, plasma retinol levels in the infected, marginally vitamin A-deficient chickens showed a significant and persistent decrease compared to their noninfected counterparts fed the same diet. Moreover, infection with NDV resulted in increased rates of morbidity in the marginally vitamin A-deficient chickens compared with nondeficient chickens. The results of this study indicate that pre-existing marginal vitamin A status increases the severity of disease following NDV infection, and that infection with NDV reduces marginal plasma vitamin A levels to levels which can be regarded as deficient.

Experimental infection and serologic survey for selected paramyxoviruses in red-winged blackbirds (Agelaius phoeniceus).

Red-winged blackbirds (Agelaius phoeniceus) were experimentally exposed to three avian paramyxoviruses: turkey parainfluenza virus, Yucaipa virus, and two strains of Newcastle disease virus. Aerosol exposure resulted in infection but exposure in food or drinking water rarely or never did. Tracheal swabs contained virus for up to eight days post exposure, cloacal swabs were negative. Transmission to contact birds occurred infrequently. Antibody response was of low titer and short duration. No hemagglutination inhibition activity against these viruses was found in 387 sera collected from red-winged blackbirds and tricolored blackbirds (Agelaius tricolor) trapped in six states.

Characteristics of a local virulent strain of Newcastle disease virus.

A local virulent strain, VLT, of Newcastle disease virus formed 3- to 4-mm plaques on monolayers of primary chicken embryo cultures on the 4th day after inoculation. It agglutinated chicken and human 0 erythrocytes. Its hemagglutinin was stable at 56 C when compared with those of Komarov (K) and F vaccinal strains of the same virus. The viral titer of infected allantoic fluid dropped from 10(8.1) plaque-forming units to 10(1.0) plaque-forming units/ml within 2 hours when incubated at 56 C. The strain was ether-sensitive; it adsorbed readily on monolayers of chicken embryo cells and did not diffuse through agar. Its intracerebral pathogenicity index, chicken dose LD50, and embryo mean death time (hours) were 1.8, 9.0, and 48, respectively. Two virulent strains isolated in 1974 and 1975 were found to be identical to the VLT strain in terms of certain physical and biological properties. On the basis of plaque morphologic characteristics, hemagglutination spectrum, and hemagglutinin inactivation at 56 C, it was possible to identify readily the field isolate when it was compared with vaccinal strains (K and F) commonly used in Lebanon.

Mycoplasma-mediated hyporeactivity to various interferon inducers.

Three strains of Mycoplasma arthritidis were shown to induce marked hyporeactivity in mice to interferon induction by both Newcastle disease virus and poly(I:C). In contrast, the interferon response of mice to tilorone was only partially suppressed by pretreatment of the animals with mycoplasms. Hyporeactivity to Newcastle disease virus was maximal 1 and 3 days after mycoplasms treatment, but the interferon response was maximal 1 day after injection of the mycoplasmas and was no longer apparent by 5 days. No relationship was found between the ability of the mycoplasms themselves to induce interferon and the degree of hyporeactivity produced. These results suggest that mycoplasmas may alter virus-host relationships in vivo.

Acute newcastle viral infection of the upper respiratory tract of the chicken. II. The effect of diets deficient in vitamin A on the pathogenesis of the infection.

Keratotic and squamous changes characteristic of vitamin A deficiency were minimal even in chicks which were malnourished and growth stunted and had no vitamin A in their diet. However, when these chicks were infected with Newcastle disease virus (NDV), keratotic changes appeared, most markedly in areas regenerating after infection. In chicks raised on full nutrient diets lacking only vitamin A, keratotic changes appeared in several areas of nasal mucosa but were absent from the mucosa of the inner (under) surface of the maxillary turbinate. Following NDV infection, such changes did appear in the inner lining epithelia. It is suggested that depletion of vitamin A causes regenerating epithelial cells to keratinize. Other effects of combined lack of vitamin A plus NDV infection were exhaustion of lymphoid cells from cranial bone marrow and exhaustion of lymphoid cell systems locally from the nose and paranasal glands.

Acute Newcastle viral infection of the upper respiratory tract of the chicken. I. A model for the study of environmental factors on upper respiratory tract infection.

Acute Newcastle disease virus infection following intranasal inoculation of chicks with a mesogenic strain of the virus produced a localized infection of the middle turbinate which was histologically demonstrable 18 hours after inoculation. There was destruction of mucous cells of individual acini in the under surface of the middle turbinate, and the infection rapidly spread to ciliated and goblet cells and to neighboring acini. By day 2 there was simultaneous remodeling of the mucosa, continued destruction and inflammatory infiltration and frequent loss of cartilage basophilia. By day 3 polymorphonuclear cells almost disappeared, epithelial mitoses commenced, and lymphocyte infiltration intensified; the plasma cells normally present along the lateral nasal gland ducts were often destroyed, very occasionally the glands themselves were destroyed. By days 5 and 6 inflammation greatly decreased, and by day 8 the mucociliated epithelium was essentially normal. The infection is sequentially comparable to acute mild rhinitis of man.