Parkinson’s disease (PD) is the most prevalent movement disorder known and predominantly affects the elderly. It is a progressive neurodegenerative disease wherein α-synuclein, a neuronal protein, aggregates to form toxic structures in nerve cells. The cause of Parkinson’s disease (PD) remains unknown. Intestinal dysfunction and changes in the gut microbiota, common symptoms of PD, are evidently linked to the pathogenesis of PD. Although a multitude of studies have investigated microbial etiologies of PD, the microbial role in disease progression remains unclear. Here, we show that Gram-negative sulfate-reducing bacteria of the genus Desulfovibrio may play a potential role in the development of PD. Conventional and quantitative real-time PCR analysis of feces from twenty PD patients and twenty healthy controls revealed that all PD patients harbored Desulfovibrio bacteria in their gut microbiota and these bacteria were present at higher levels in PD patients than in healthy controls. Additionally, the concentration of Desulfovibrio species correlated with the severity of PD. Desulfovibrio bacteria produce hydrogen sulfide and lipopolysaccharide, and several strains synthesize magnetite, all of which likely induce the oligomerization and aggregation of α-synuclein protein. The substances originating from Desulfovibrio bacteria likely take part in pathogenesis of PD. These findings may open new avenues for the treatment of PD and the identification of people at risk for developing PD.

Nisin-producing Lactococcus lactis protects its own cell membrane against the bacteriocin with the ABC transporter NisFEG, and the immunity lipoprotein NisI. In this study, in order to localize a site for specific nisin interaction in NisI, a C-terminal deletion series of NisI was constructed, and the C-terminally truncated NisI proteins were expressed in L. lactis. The shortest deletion (5 aa) decreased the nisin immunity capacity considerably in the nisin-negative strain MG1614, resulting in approximately 78 % loss of immunity function compared with native NisI. A deletion of 21 aa decreased the immunity level even more, but longer deletions, up to 74 aa, provided the same level of nisin immunity as the 21 aa deletion, i.e. approximately 14 % of the immunity provided by native NisI. Similar to native NisI, all the C-terminally truncated NisI proteins provided higher immunity to nisin in the NisFEG-expressing strain NZ9840 than in MG1614, i.e. approximately 40-50 % of the immunity capacity of native NisI. Then, it was determined whether the NisI Cterminal 21 aa fragment could protect cells against nisin. To target the 21 aa fragment to its natural location, 21 C-terminal amino acids from the subtilin-specific immunity lipoprotein SpaI were replaced by 21 C-terminal amino acids from NisI. The expression of the SpaI9-9NisI fusion in L. lactis strains significantly increased their nisin immunity. This is the first time the immunity function of a lantibiotic immunity protein has been transferred to another protein. However, unlike native NisI, and the C-terminally truncated NisI fragments, the increase in nisin immunity conferred by the SpaI9-9NisI fusion was the same in both the NisFEG strain NZ9840 and MG1614. In conclusion, the SpaI9-9NisI fusion could not enhance nisin immunity by interacting with NisFEG, whereas the Cterminally truncated NisI fragments and native NisI were able to enhance nisin immunity, probably by co-operation with NisFEG. The results made it evident that the C terminus of NisI is involved in specific interaction with nisin, and that it confers specificity for the NisI immunity lipoprotein.

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Lactococcus lactis cells producing the antibacterial peptide nisin protect their own cytoplasmic membrane by specific immunity proteins, NisI and NisF/E/G. We show here that approximately half of the produced NisI escaped the lipid modification (LF-NisI=lipid-free NisI) and was secreted to the medium, and that LF-NisI had no affinity to cells of L. lactis. The molar ratio of NisI and nisin was determined to be approximately 1:10 on the cell surface and 1:50 in the culture supernatant. Purified LF-NisI was shown to enhance the activity of nisin against several tested indicator strains. The enhancement of nisin activity by LF-NisI was not observed with cells containing the NisFEG transport system.

A nisin-resistant Lactococcus lactis strain TML01 was isolated from crude milk. A gene with 99% homology to the nisin-resistance gene, nsr, was identified. The food-grade secretion plasmid, pLEB690 (3746 bp), was constructed based on this novel nsr gene enabling primary selection with up to 5 μg nisin/ml. The functionality of pLEB690 as a secretion vector was shown by expressing and secreting the pediocin AcH gene papA in L. lactis. pLEB690 is therefore, a functional food-grade secretion vector potentially useful for the food industry.

Nisin-producing Lactococcus lactis cells protect their own cytoplasmic membrane by specific immunity proteins, NisF/E/G and NisI, a transporter complex and a lipoprotein, respectively. A portion of NisI is secreted to the medium in a lipid-free form (LF-NisI). Here, kinetics of the interaction between nisin and LF-NisI was examined by surface plasmon resonance analysis. The affinity constant K D for the interaction was calculated to be in the micromolar range. Contribution of the secreted LF-NisI to nisin immunity was studied by replacing the lipoprotein specific nisI signal sequence with a secretion signal of non-lipoprotein origin. Secretion of LF-NisI in NisF/E/G-expressing L. lactis strain NZ9840 increased significantly its nisin tolerance suggesting that the lipid-free form of NisI could have a supportive role in nisin immunity.

Lactic acid bacteria originating in the intestine have recently undergone intensive study for their potential probiotic properties. Here partial 16S rRNA gene sequencing of 8 Lactobacillus strains proved them to be Lactobacillus crispatus. Fatty acid analysis confirmed strains being closely related. These strains and type strain ATCC33820 were characterized for genetic engineering potential, thus determining aerobic growth, erythromycin sensitivity, and glycine tolerance. Out of 5 plasmids, a 2.9-kb plasmid (pLEB579) was successfully introduced into 4 chicken-originated wild-type L. crispatus strains. Transformation frequency was approximately 30 transformants per microgram of DNA, the first reported electrotransformation into chicken-originated L. crispatus. In spite of its low frequency, transformation enables bioengineering of these strains to improve the probiotic function in feed adsorption, chicken health, and food safety.

Nisin‐producing Lactococcus lactis cells protect their own cytoplasmic membrane by specific immunity proteins, NisF/E/G and NisI, a transporter complex and a lipoprotein, respectively. A portion of NisI is secreted to the medium in a lipid‐free form (LF‐NisI). Here, kinetics of the interaction between nisin and LF‐NisI was examined by surface plasmon resonance analysis. The affinity constant KD for the interaction was calculated to be in the micromolar range. Contribution of the secreted LF‐NisI to nisin immunity was studied by replacing the lipoprotein specific nisI signal sequence with a secretion signal of non‐lipoprotein origin. Secretion of LF‐NisI in NisF/E/G‐expressing L. lactis strain NZ9840 increased significantly its nisin tolerance suggesting that the lipid‐free form of NisI could have a supportive role in nisin immunity.