21.15109/ARP/3IIFGYNagy, Eszter ZsófiaEszter ZsófiaNagy57950528900HUN-REN Veterinary Medical Research InstituteSzeredi, LeventeLeventeSzeredi0000-0001-5128-7302University of Veterinary Medicine, Department of Microbiology and Infectious DiseasesFöldi, DorottyaDorottyaFöldi57216506247HUN-REN Veterinary Medical Research InstituteBelecz, NikolettNikolettBelecz57239500800HUN-REN Veterinary Medical Research InstituteKovács, Áron BotondÁron BotondKovács0000-0001-6013-819XHUN-REN Veterinary Medical Research InstituteSulyok, Kinga MáriaKinga MáriaSulyok0009-0004-0352-3306HUN-REN Veterinary Medical Research InstituteGrózner, DénesDénesGrózner0000-0002-2154-7537HUN-REN Veterinary Medical Research InstituteWehmann, EnikőEnikőWehmann0000-0002-1229-1146HUN-REN Veterinary Medical Research InstituteBányai, KrisztiánKrisztiánBányai0000-0002-6270-1772HUN-REN Veterinary Medical Research InstituteMarton, SzilviaSzilviaMarton15045147500HUN-REN Veterinary Medical Research InstituteTenk, MiklósMiklósTenk0000-0003-0399-8147University of Veterinary Medicine, Department of Microbiology and Infectious DiseasesKreizinger, ZsuzsaZsuzsaKreizinger0000-0002-6365-0719HUN-REN Veterinary Medical Research InstituteGyuranecz, MiklósMiklósGyuranecz0000-0002-5698-5713HUN-REN Veterinary Medical Research InstituteARP2026Agricultural Sciences1-Methyl-3-nitro-1-nitrosoguanidineLive attenuated vaccineMycoplasma hyorhinisNTGPorcineTemperature-sensitive strainKreizinger, ZsuzsaZsuzsaKreizingerHUN-REN Veterinary Medical Research Institute2026-03-252026-03-2510.1016/j.vaccine.2026.1282781244692114577770337619440298320577542087290285474641889text/csvtext/csvtext/csvtext/csvtext/csvtext/csvtext/csvtext/csvtext/csvtext/csvtext/csvtext/csvtext/csvtext/csvtext/csvapplication/vnd.openxmlformats-officedocument.spreadsheetml.sheet1.1CC BY-NC 4.0<b>Background</b> <i>Mycoplasma</i> (<i>M</i>.) <i>hyorhinis</i> causes substantial economic losses in swine. Currently, prevention and treatment rely on minimizing risk factors and administering antibiotics, as no vaccines are commercially available in Europe. However, antibiotics often cannot fully eliminate the bacteria. The development of an effective vaccine could lead to a potentially long-term control method. <b>Materials and methods</b> A temperature-sensitive <i>M. hyorhinis</i> 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 <i>M. hyorhinis</i> 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. <b>Results</b> 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 <i>M. hyorhinis</i>-specific IgG antibody levels post-challenge than the positive control group. However, the vaccine candidate did not mitigate the impact of <i>M. hyorhinis</i> 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. <b>Discussion</b> Overall, the attenuated <i>M. hyorhinis</i> strain, combined with the adjuvant, provided protection against <i>M. hyorhinis</i> infection. These results form a basis for the development of a novel vaccine candidate that offers effective prevention.Momentum (Lendület) program of the Hungarian Academy of SciencesLP2022–6/2022Recovery and Resilience Facility (RRF), financed under the National Recovery FundRRF-2.3.1-21-2022-00001