ABSTRACT. The efficacy of Mycoplasma hyopneumoniae oral vaccine was investigated in microsphere dosage form. A co-spray drying process was used to apply an encapsulating material, Eudragit L30 D-55, to microspheres containing Mycoplasma hyopneumoniae antigens. The microspheres were generally effective (>93%) with protein release at pH 7.4, but almost none were released at pH 1.2, for 3 hr in an in vitro dissolution test. An SPF-swine model was used to evaluate the effectiveness of the microspheres as an oral vaccine, and the related immune responses. The serum's systemic IgG against M. hyopneumoniae was evoked by ELISA analysis, after a 2nd immunization of all pigs. The vaccinated groups' mean lesion score was significantly lower after the Mycoplasma hyopneumoniae challenge than that of the nonvaccinated/challenged groups (P<0.05). This study strongly suggests that the oral microspheres vaccine prepared by a co-spray drying method can provide effective protection against M. hyopneumoniae infection in pigs. KEY WORDS: co-spray drying, microsphere, Mycoplasma hyopneumoniae, oral vaccine.J. Vet. Med. Sci. 65(1): 69-74, 2003 Mycoplasma hyopneumoniae is widely recognized as a potent pathogen for mycoplasmal pneumonia in swine [2], colonizing the respiratory epithelia and compromising integrity by inducing an inflammatory response [20]. The pathogen also causes significant economic losses by reducing body weight and prolonging periods between feeds [22]. Although some drugs and antibiotics are effective in vitro against M. hyopneumoniae, reducing the disease's clinical signs [12], they have not yet been proven effective in eliminating the pathogen in vivo. Vaccination is an essential strategy in controlling mycoplasmal pneumonia. M. hyopneumoniae infects the ciliated epithelial cells of the respiratory tract [6], and a mucosal immune response may therefore be important in the prevention and control of M. hyopneumoniae-induced pneumonia [23]. The concept of a common mucosal immune system has been supported by several oral immunization reports [8,13]. Numerous experimental systems have proven that oral immunization can induce antibody secretion into the mucus on these surfaces [8,16,18], and is almost completely restricted to the secretory form of IgA. Oral immunization causes this IgA antibody to be secreted from the GALT to distant mucosal tissue, including the salivary, mammary, respiratory, intestine and genital mucosal tissue, protecting these areas from pathogen invasion [5,14].Microspheres have recently received widespread interest as vehicles for the controlled release of drug or bioactive agents; resisting degradation in gastric acid, they can release drugs in the intestines [1]. Oral antigen delivery by pHdependent microspheres can improve local and systemic immune responses [4,16], serving as a potent mucosal immunogen. It is essential to develop a new, simple and effective vaccine process. Microsphere oral vaccines have been previously developed in our laboratory via a novel cospray-drying process [16]. C...