Supplementary data – Mycoplasma hyorhinis vaccine strain – Nagy, 2026

The development of an effective vaccine could lead to a potentially long-term control method. Materials and methods A temperature-sensitive M. hyorhinis strain was developed using 1-methyl-3-nitro-1-nitrosoguanidine (NTG) mutagenesis. The immunogenicity and efficacy of this vaccine candidate clone were evaluated in combination with an adjuvant.

Three-week-old piglets were immunized with the candidate vaccine strain, and the vaccination site was monitored daily. At six weeks of age, the pigs were challenged intravenously on two subsequent days. Daily clinical examinations were conducted, with blood and nasal swabs collected weekly throughout the study for M. hyorhinis enzyme-linked immunosorbent assay (ELISA), real-time PCR analysis, and isolation.

Three weeks post-challenge, the animals were euthanized for gross and histopathological examinations. Body temperature was recorded daily, and body weight was measured upon arrival, and then at six and nine weeks of age. Results Vaccination significantly reduced clinical signs (p = 0.03), as well as gross pathological (p = 0.01) and histopathological (p = 0.005) lesions compared with the positive control group.

The vaccinated group exhibited an earlier and higher increase in M. hyorhinis-specific IgG antibody levels post-challenge than the positive control group. However, the vaccine candidate did not mitigate the impact of M. hyorhinis infection on the weight gain. After the challenge (days 21–42), both the vaccinated (p = 0.001) and the positive control (p = 0.003) groups exhibited reduced weight gain compared with the negative control group.

Discussion Overall, the attenuated M. hyorhinis strain, combined with the adjuvant, provided protection against M. hyorhinis infection. These results form a basis for the development of a novel vaccine candidate that offers effective prevention.