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Feline calicivirus (FCV) is a common feline pathogen with a potential for antigenic diversity. This study aimed to evaluate and characterize the protective efficacy of the FCV-F9 valency of a tetravalent vaccine, Leucofeligen, against challenge with an unrelated strain. Ten 9-week-old kittens were vaccinated while 10 remained as unvaccinated controls. The vaccinated cats received Leucofeligen twice subcutaneously with a 3-week interval. Four weeks after the second vaccination, all cats were challenged with virulent heterologous FCV and followed up for 21 days, monitoring their general condition, clinical signs, and immunological responses. During the vaccination phase, rectal temperatures and body weights were indistinguishable between the two groups. Only vaccinated cats showed FCV-specific seroconversion (both total and neutralizing antibodies). In the first week after challenge, the vaccinated cats had an 82.6% reduction in median clinical score compared to controls. Leucofeligen was thus shown to provide a significant clinical protection to kittens challenged with heterologous virulent FCV. This protection was similar whether the cats had neutralizing antibody or not, indicating a key role for cellular immunity in the overall protection. This also suggests that previously reported seroneutralisation studies may underestimate the level of cross-protection against field strains obtained with this modified live FCV-F9 vaccine.

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The regulation of cell adhesion in epithelia is a fundamental process governing morphogenesis in embryos and a key step in the progression of invasive cancers. Here, we have analysed the molecular pathways controlling the epithelial organisation of somites. Somites are mesodermal epithelial structures of vertebrate embryos that undergo several changes in cell adhesion during early embryonic life. We show that Wnt6 in the ectoderm overlaying the somites, but not Wnt1 in the neighbouring neural tube, is the most likely candidate molecule responsible for the maintenance of the epithelial structure of the dorsal compartment of the somite: the dermomyotome. We characterised the signalling pathway that mediates Wnt6 activity. Our experiments suggest that the Wnt receptor molecule Frizzled7 probably transduces the Wnt6 signal. Intracellularly, this leads to the activation of the beta-catenin/LEF1-dependent pathway. Finally, we demonstrate that the bHLH transcription factor paraxis, which was previously shown to be a major player in the epithelial organisation of somites, is a target of the beta-catenin signal. We conclude that beta-catenin activity, initiated by Wnt6 and mediated by paraxis, is required for the maintenance of the epithelial structure of somites.

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In ovo electroporation is a well-established method of gene transfer into neural and mesenchymal tissue in chick embryos. Electroporation of somites, however, has been hampered by low efficiency due to technical difficulties. Here, we present a powerful technique to electroporate avian somites and subpopulations of somitic cells at high efficiency in ovo. We demonstrate specific targeting of distinct somitic compartments and their derivatives using single or combinations of plasmid expression vectors. This technique opens new perspectives to investigate the morphologic and genetic basis of somite development.

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In vertebrates, despite the evidence that extrinsic factors induce myogenesis in naive mesoderm, other experiments argue that the initiation of the myogenic program may take place independent of these factors. To resolve this discrepancy, we have re-addressed this issue, using short-term in vivo microsurgery and culture experiments in chick. Our results show that the initial expression of the muscle-specific markers Myf5 and MyoD is regulated in a mesoderm-autonomous fashion. The reception of a Wnt signal is required for MyoD, but not Myf5 expression; however, we show that the source of the Wnt signal is intrinsic to the mesoderm. Gain- and loss-of-function experiments indicate that Wnt5b, which is expressed in the presomitic mesoderm, represents the MyoD-activating cue. Despite Wnt5b expression in the presomitic mesoderm, MyoD is not expressed in this tissue: our experiments demonstrate that this is due to a Bmp inhibitory signal that prevents the premature expression of MyoD before somites form. Our results indicate that myogenesis is a multistep process which is initiated prior to somite formation in a mesoderm-autonomous fashion; as somites form, influences from adjacent tissues are likely to be required for maintenance and patterning of early muscles.

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