The intranasal administration of Naegleria fowleri lysates plus cholera toxin (CT) increases protection against N. fowleri meningoencephalitis in mice, suggesting that humoral immune response mediated by antibodies is crucial to induce protection against the infection. In the present study, we applied a protein analysis to detect and identify immunogenic antigens from N. fowleri, which might be responsible for such protection. A Western blot assay of N. fowleri polypeptides was performed using the serum and nasal washes from mice immunized with N. fowleri lysates, either alone or with CT after one, two, three, or four weekly immunizations and challenged with trophozoites of N. fowleri. Immunized mice with N. fowleri plus CT, after four doses, had the highest survival rate (100%). Nasal or sera IgA and IgG antibody response was progressively stronger as the number of immunizations was increased, and that response was mainly directed to 250, 100, 70, 50, 37, and 19 kDa polypeptide bands, especially in the third and fourth immunization. Peptides present in these immunogenic bands were matched by nano-LC–ESI-MSMS with different proteins, which could serve as candidates for a vaccine against N. fowleri infection.

Many pathogenicity factors are involved in the development of primary amoebic meningoencephalitis (PAM) caused by N fowleri. However, most of them are not exclusive for N fowleri and they have not even been described in other nonpathogenic Naegleria species. Therefore, the objective of this work was to identify differential proteins and protein pattern recognition between Naegleria fowleri and Naegleria lovaniensis using antibodies anti‐N fowleri as strategy to find vaccine candidates against meningoencephalitis. Electrophoresis and Western blots conventional and 2‐DE were performed for the identification of antigenic proteins, and these were analysed by the mass spectrometry technique. The results obtained in 2‐DE gels and Western blot showed very notable differences in spot intensity between these two species, specifically those with relative molecular weight of 100, 75, 50 and 19 kDa. Some spots corresponding to these molecular weights were identified as actin fragment, myosin II, heat shock protein, membrane protein Mp2CL5 among others, with differences in theoretical post‐translational modifications. In this work, we found differences in antigenic proteins between both species, proteins that could be used for a further development of vaccines against N fowleri infection.

Barley malting quality depends on seed characteristics achieved during grain development and germination. One important parameter is protein accumulation in the mature seed, which may vary between cultivars. Here we conducted a protein pattern analysis in the range of pI 4–7 of mature grains from five Mexican barley cultivars, commonly used for malt and beer production. Reproducibly distinct protein spots, separated by 2D SDS PAGE, were identified by mass spectrometry and considered as potential markers for cultivars with distinct seed protein accumulation. The expression patterns of glutamate decarboxylase (GAD) and protein disulfide isomerase (PDI1-1) were followed at transcript level during grain development for three independent growth cycles to establish whether differences between cultivars were reproducible. Quantitative determination of PDI1-1 protein levels by ELISA confirmed a reproducibly, distinctive accumulation and post-translational modifications between cultivars, which were independent of plant growth regimes. According to its impact on differential storage protein accumulation, we propose the PDI1-1 protein as potential biomarker for Mexican malting barley cultivars.

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