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An epidemic of high-pathogenicity avian influenza (HPAI) A virus subtype H7N7 occurred in The Netherlands in 2003 that affected 255 flocks and led to the culling of 30 million birds. To evaluate the effectiveness of the control measures, we quantified between-flock transmission characteristics of the virus in 2 affected areas, using the reproduction ratio Rh. The control measures markedly reduced the transmission of HPAI virus: Rh before detection of the outbreak in the first infected flock was 6.5 (95% confidence interval [CI], 3.1-9.9) in one area and 3.1 in another area, and it decreased to 1.2 (95% CI, 0.6-1.9) after detection of the first outbreak in both areas. The observation that Rh remained >1 suggests that the containment of the epidemic was probably due to the reduction in the number of susceptible flocks by complete depopulation of the infected areas rather than to the reduction of the transmission by the other control measures.

SUMMARYFour UK strains of three different serotypes were found to differ by only 2-3% of their S1 amino acids. The S1 sequences were also very similar to those of three Dutch isolates (D207, D274 and D3896), the greatest difference between two of the seven isolates being 4.4%. The few amino acid differences between the seven isolates were located largely between residues 19-122 and 251-347 of the mature S1 subunit.The seven isolates could be differentiated using 16 monoclonal antibodies in an enzymelinked immunosorbent assay. Some virus neutralizing (VN) antibody-inducing epitopes were common to all seven isolates even though the strains had been differentiated into three serotypes by polyclonal sera. The results indicate that the most antigenic of the VN antibody-inducing epitopes are formed by very few amino acids and that these occur in the first and third quarters of the S1 subunit. We suggest that serology-based epizootiological studies of IBV should, therefore, be augmented by the inclusion of nucleic acid sequencing and/or monoclonal antibody analysis.

Monoclonal antibodies (MAbs) directed against structural proteins of infectious bronchitis virus (IBV) were produced to analyse the antigenic structure of this virus. Competitive binding of enzyme-labelled and unlabelled MAbs to IBV peplomer protein was analysed in an antibody binding assay to test the relatedness of the epitopes defined by the MAbs. Based on the competition groups, eight epitope clusters were defined (S-A to S-H); six of these clusters (S1-A to S I-F) were located on the S1 subunit and two (S2-G and S2-H) on the $2 subunit of the peplomer protein.Epitope clusters S 1-A and S1-B overlapped extensively. The biological activities of the MAbs were determined and correlated to the epitope clusters. Monoclonal antibodies directed against epitope clusters S 1-A to S1-E and one MAb directed against cluster S2-G moderately to strongly neutralized IBV at titres higher than 2 log~o, whereas the remaining MAbs, directed against S1 and $2, neutralized at titres lower than 2 log~o. One MAb, directed against cluster S1-D, inhibited the agglutination of chicken erythrocytes.

Neutralizing monoclonal antibodies directed against five antigenic sites on the spike (S) S 1 glycopolypeptide of avian infectious bronchitis virus (IBV) were used to select neutralization-resistant variants of the virus. By comparing the nucleotide sequence of such variants with the sequence of the IBV parent strain, we located five antigenic sites on the amino acid sequence of the S 1 glycopolypeptide. The variants had mutations within three regions corresponding to amino acid residues 24 to 61, 132 to 149 and 291 to 398 of the S1 glycopolypeptide. The location of three overlapping antigenic sites on the IBV spike protein was similar to the location of antigenic sites on the spike protein of other coronaviruses.

Recent outbreaks of highly pathogenic avian influenza (HPAI) viruses in poultry and their threatening zoonotic consequences emphasize the need for effective control measures. Although vaccination of poultry against avian influenza provides a potentially attractive control measure, little is known about the effect of vaccination on epidemiologically relevant parameters, such as transmissibility and the infectious period. We used transmission experiments to study the effect of vaccination on the transmission characteristics of HPAI A͞Chicken͞Netherlands͞03 H7N7 in chickens. In the experiments, a number of infected and uninfected chickens is housed together and the infection chain is monitored by virus isolation and serology. Analysis is based on a stochastic susceptible, latently infected, infectious, recovered (SEIR) epidemic model. We found that vaccination is able to reduce the transmission level to such an extent that a major outbreak is prevented, important variables being the type of vaccine (H7N1 or H7N3) and the moment of challenge after vaccination. Two weeks after vaccination, both vaccines completely block transmission. One week after vaccination, the H7N1 vaccine is better than the H7N3 vaccine at reducing the spread of the H7N7 virus. We discuss the implications of these findings for the use of vaccination programs in poultry and the value of transmission experiments in the process of choosing vaccine.SEIR model ͉ highly pathogenic avian influenza ͉ final size ͉ generalized linear model ͉ reproduction ratio H ighly pathogenic avian influenza (HPAI) is a disease of poultry caused by H5 or H7 AI A strains, with mortality that ranges up to 100%. The number of outbreaks in the last few years has been

Knowledge on the transmission tree of an epidemic can provide valuable insights into disease dynamics. The transmission tree can be reconstructed by analysing either detailed epidemiological data (e.g. contact tracing) or, if sufficient genetic diversity accumulates over the course of the epidemic, genetic data of the pathogen. We present a likelihood-based framework to integrate these two data types, estimating probabilities of infection by taking weighted averages over the set of possible transmission trees. We test the approach by applying it to temporal, geographical and genetic data on the 241 poultry farms infected in an epidemic of avian influenza A (H7N7) in The Netherlands in 2003. We show that the combined approach estimates the transmission tree with higher correctness and resolution than analyses based on genetic or epidemiological data alone. Furthermore, the estimated tree reveals the relative infectiousness of farms of different types and sizes.

Virulence of Newcastle disease virus (NDV) is mainly determined by the amino acid sequence surrounding the fusion (F) protein cleavage site, since host proteases that cleave the F protein of virulent strains are present in more tissues than those that cleave the F protein of non-virulent strains. Nevertheless, comparison of NDV strains that carry exactly the same F protein cleavage site shows that significant differences in virulence still exist. For instance, virulent field strain Herts/33 with the F cleavage site 112 RRQRRF 117 had an intracerebral pathogenicity index of 1?88 compared with 1?28 for strain NDFLtag, which has the same cleavage site. This implies that additional factors contribute to virulence. After generating an infectious clone of Herts/33 (FL-Herts), we were able to map the location of additional virulence factors by exchanging sequences between FL-Herts and NDFLtag. The results showed that, in addition to the F protein cleavage site, the haemagglutinin-neuraminidase (HN) protein also contributed to virulence. The effect of the HN protein on virulence was most prominent after intravenous inoculation. Interestingly, both the stem region and the globular head of the HN protein seem to be involved in determining virulence. INTRODUCTIONNewcastle disease is a highly contagious and fatal viral disease affecting most species of birds. Because chickens are the most susceptible birds, the disease is frequently responsible for devastating losses in poultry (Alexander, 2000(Alexander, , 2001 NDV is an enveloped virus with two membrane proteins: the haemagglutinin-neuraminidase (HN) protein involved in cell attachment and release, and the fusion (F) protein involved in mediating fusion of the viral envelope with cellular membranes. The F protein is synthesized as a precursor, F0, and is only fusogenic after cleavage into disulfide-linked F1 and F2 polypeptides. However, cleavage is not sufficient for the fusion process. The HN protein is also required for fusion to occur by cooperating with the F protein (Deng et al., 1997). The consensus sequence of the F protein cleavage site of velogenic and mesogenic strains is 112 (R/K)RQ(R/K)RF 117. The consensus sequence of the lentogenic F cleavage site is 112 (G/E)(K/R)Q(G/ E)RL 117. The different F protein cleavage sites are the substrates for different types of cellular proteases (Kawahara et al., 1992;Sakaguchi et al., 1991). The F protein of lentogenic viruses can be cleaved only by trypsin-like enzymes, as found in the respiratory and intestinal tracts, whereas the F protein of virulent viruses can be cleaved by a hostThe GenBank/EMBL/DDBJ accession number of the sequence determined in this work is AY741404. 0008-0822 G 2005 SGM Printed in Great Britain