SP-D is a pulmonary collectin important in lung immunity. SP-D-deficient mice (Sftpd^-/-) are reported to be susceptible to ovalbumin (OVA)- and fungal allergen-induced pulmonary inflammation, while treatment with exogenous SP-D has therapeutic effects in such disease models. β-glucans are a diverse group of polysaccharides previously suggested to serve as fungal ligands for SP-D. We set out to investigate if SP-D could interact with 1,3-β-glucan and attenuate allergic pulmonary inflammation in the presence of 1,3-β-glucan. Allergic airway disease was induced in Sftpd^-/- and Sftpd^+/+ mice by OVA sensitization and subsequent challenge with OVA, 1,3-β-glucan or OVA/1,3-β-glucan together. Mice in the combined treatment group were further treated with a high dose of recombinant fragment of human SP-D (rfhSP-D). We demonstrated direct interaction between SP-D and 1,3-β-glucan. OVA-induced mucous cell metaplasia was increased in Sftpd^-/- mice, supporting previously reported protective effects of endogenous SP-D in allergy. OVA-induced parenchymal CCL11 levels and eosinophilic infiltration in brochoalveolar lavage (BAL) were unaffected by 1,3-β-glucan but were reversed with rfhSP-D treatment. 1,3-β-glucan-treatment did, however, induce pulmonary neutrophilic infiltration and increased TNF-α levels in BAL, independently of OVA-induced allergy. This infiltration was also reversed by treatment with rfhSP-D. 1,3-β-glucan reduced OVA-induced mucous cell metaplasia, Th2 cytokines and IFN- production. rfhSP-D treatment further reduced mucous metaplasia and Th2 cytokine secretion to background levels. In summary, rfhSP-D treatment resulted in attenuation of both allergic inflammation and 1,3-β-glucan mediated neutrophilic inflammation. Our data suggest that treatment with high dose SP-D protects from mold-induced exacerbations of allergic asthma.