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In the present study, mutants defective in <jats:italic>\u03b2<\/jats:italic>\u20101,6\u2010glucan synthesis, including <jats:italic>kre1\u2206<\/jats:italic>, <jats:italic>kre6\u2206<\/jats:italic>, <jats:italic>kre9\u2206<\/jats:italic> and <jats:italic>big1\u2206<\/jats:italic>, were sporulated to analyse the effect of <jats:italic>\u03b2<\/jats:italic>\u20101,6\u2010glucan defects on the spore wall. Except for <jats:italic>kre6\u2206<\/jats:italic>, these mutant spores were sensitive to treatment with ether, suggesting that the mutations perturb the integrity of the spore wall. Morphologically, the mutant spores were indistinguishable from wild\u2010type spores. They lacked significant sporulation defects partly because the chitosan layer, which covers the glucan layer, compensated for the damage. The proof for this model was obtained from the effect of the additional deletion of <jats:italic>CHS3<\/jats:italic> that resulted in the absence of the chitosan layer. Among the double mutants, the most severe spore wall deficiency was observed in <jats:italic>big1\u2206<\/jats:italic> spores. The majority of the <jats:italic>big1\u2206chs3\u2206<\/jats:italic> mutants failed to form visible spores at a higher temperature. Given that the <jats:italic>big1\u2206<\/jats:italic> mutation caused a failure to attach a GPI\u2010anchored reporter, Cwp2\u2010GFP, to the spore wall, <jats:italic>\u03b2<\/jats:italic>\u20101,6\u2010glucan is involved in tethering of GPI\u2010anchored proteins in the spore wall as well as in the vegetative cell wall. Thus, <jats:italic>\u03b2<\/jats:italic>\u20101,6\u2010glucan is required for proper organization of the spore wall. 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