Search

Plant-pathogenic bacteria cause numerous diseases in host plants and can result in serious damage. Timely and accurate diagnostic techniques are, therefore, crucial. While advances in molecular techniques have led to diagnostic systems able to distinguish known plant pathogens at the species or strain level, systems covering larger categories are mostly lacking. In this study, a specific and universal LAMP-based diagnostic system was developed for phytoplasmas, a large group of insect-borne plant-pathogenic bacteria that cause significant agricultural losses worldwide. Targeting the 23S rRNA gene of phytoplasma, the newly designed primer set CaPU23S-4 detected 31 ' Candidatus Phytoplasma' tested within 30 min. This primer set also showed high specificity, without false-positive results for other bacteria (including close relatives of phytoplasmas) or healthy plants. The detection sensitivity was ~10,000 times higher than that of PCR methods for phytoplasma detection. A simple, rapid method of DNA extraction, by boiling phytoplasma-infected tissues, was developed as well. When used together with the universal LAMP assay, it enabled the prompt and accurate detection of phytoplasmas from plants and insects. The results demonstrate the potential of the 23S rRNA gene as a versatile target for the LAMP-based universal detection of bacteria at the genus level and provide a novel avenue for exploring this gene as molecular marker for phytoplasma presence detection.IMPORTANCEPhytoplasmas are associated with economically important diseases in crops worldwide, including lethal yellowing of coconut palm, "flavescence dorée" and "bois noir" of grapevine, X-disease in stone fruits, and white leaf and grassy shoot in sugarcane. Numerous LAMP-based diagnostic assays, mostly targeting the 16S rRNA gene, have been reported for phytoplasmas. However, these assays can only detect a limited number of ' Candidatus Phytoplasma' species, whereas the genus includes at least 50 of these species. In this study, a universal, specific, and rapid diagnostic system was developed that can detect all provisionally classified phytoplasmas within 1 h by combining the LAMP technique targeting the 23S rRNA gene with a simple method for DNA extraction. This diagnostic system will facilitate the on-site detection of phytoplasmas and may aid in the discovery of new phytoplasma-associated diseases and putative insect vectors, irrespective of the availability of infrastructure and experimental resources., Competing Interests: The authors declare no conflict of interest.

Despite accumulating evidence that suggests a unique gut microbiota composition in athletes, a comprehensive understanding of this phenomenon is lacking. Furthermore, seasonal variation in the gut microbiota of athletes, particularly during the off-season, remains underexplored. This study aimed to compare the gut microbiotas between athletic subjects (AS) and non-athletic subjects (NS), and to investigate variations between athletic and off-season periods. The data were derived from an observational study involving Japanese male handball players. The results revealed a distinct gut microbiota composition in AS compared with NS, characterized by significantly higher alpha-diversity and a greater relative abundance of Faecalibacterium and Streptococcus . Moreover, a comparative analysis between athletic and off-season periods in AS demonstrated a significant change in alpha-diversity. Notably, AS exhibited significantly higher alpha-diversity than NS during the athletic season, but no significant difference was observed during the off-season. This study demonstrates the characteristics of the gut microbiota of Japanese handball players and highlights the potential for changes in alpha-diversity during the off-season. These findings contribute to our understanding of the dynamic nature of the gut microbiota of athletes throughout the season.

Visceral fat accumulation is considered to be associated with a higher risk of chronic diseases. We investigated the effects of Bifidobacterium longum subsp. longum ( B. longum ) BB536 and Bifidobacterium breve ( B. breve ) MCC1274 on body composition, including visceral fat, in a randomized, parallel-group, placebo-controlled study. Participants were between 29 and 64 years of age and had a body mass index (BMI) of greater than 23 and less than 30. One hundred participants were randomly assigned to the probiotics group or placebo group. Participants were administered probiotic capsules containing 1 × 10 10 colony-forming units (CFUs) of B. longum BB536 and 5 × 10 9 CFU of B. breve MCC1274 or placebo capsules without bifidobacteria for 16 weeks. In the probiotics group, abdominal visceral fat area, total abdominal fat area, and serum triglyceride levels were significantly decreased compared to those in the placebo group. Additionally, the increase in BMI observed in the placebo group was significantly suppressed in the probiotics group. This study showed that B. longum BB536 and B. breve MCC1274 reduced abdominal visceral fat and total fat levels in healthy normal and overweight adults, suggesting their beneficial effects on body composition.

Probiotics are widely used in food for their health benefits to the host. Inactivated probiotics also reportedly improve the intestinal environment and immune regulation. Our previous studies showed that heat-killed Lacticaseibacillus paracasei MCC1849 (hk-MCC1849) effectively induced IL-12 production in mouse spleen cells and significantly reduced cold symptoms in clinical trial subjects. To further elucidate the mechanism of host immune regulation by hk-MCC1849, human peripheral blood mononuclear cells (PBMCs) were cocultured with hk-MCC1849. The Toll-like receptor 9 ligands CpG-ODN 2216 and hk-MCC1849 and the heat-killed Lacticaseibacillus rhamnosus ATCC53103 were used as positive and negative controls, respectively. The results showed that, compared with the control, hk-MCC1849 significantly increased the expression of the plasmacytoid dendritic cell (pDC) marker CD86 ( p  < .0001) and the pDC marker HLA-DR ( p  < .001) in PBMCs. The expression levels of the IL-12p40 , IFNα , IFNα1 , IFNγ , and ISG15 genes were significantly increased after coculture with hk-MCC1849 ( p  < .05, p  < .05, p  < .05, p  < .05, and p  < .05, respectively, vs. control). Furthermore, to confirm whether hk-MCC1849 directly interacted with pDCs, DCs were enriched with PBMCs following 24 h of coculture with hk-MCC1849. Phagocytosis of fluorescently labeled hk-MCC1849 by pDCs was observed, and there were significant increases in CD86 ( p  < .05) and HLA-DR ( p  < .0001) expression in pDCs. These results suggest that hk-MCC1849 exerts a potential immunomodulatory effect on the host through the activation of peripheral pDCs., Competing Interests: We declare no conflict of interest associated with this manuscript., (© 2024 The Authors. Food Science & Nutrition published by Wiley Periodicals LLC.)

Previous clinical studies have shown that heat-killed Lacticaseibacillus paracasei MCC1849 suppresses subjective symptoms among healthy adults. However, the mechanism underlying this beneficial effect remains unclear. This clinical study aimed to investigate the effects of MCC1849 on immune functions in humans. In this randomized, double-blind, placebo-controlled, parallel-group study, 100 healthy adults were randomly divided into MCC1849 or placebo groups. Participants ingested test powder with 5 × 10 10 MCC1849 cells or placebo powder for 4 weeks. Immune functions were evaluated using expression levels of CD86 and HLA-DR on dendritic cells (DCs), neutrophils, and natural killer cells. The expression levels of interferon (IFN)-α, -β, and -γ in peripheral blood mononuclear cells incubated with Cpg2216 in vitro were quantified. Efficacy analysis was performed on participants in the per-protocol set (placebo group; n = 47, MCC1849 group; n = 49). The expression level of CD86 on pDCs and the gene expression levels of IFN-α, -β, and -γ upon TLR9 agonist stimulation were significantly higher in the MCC1849 group at 4 weeks. No side effects were observed. This is the first report to show the positive effects of MCC1849 on human immune cells. These findings reveal one possible mechanism of how MCC1849 suppresses subjective symptoms.

Phytoplasmas infect a wide variety of plants and can cause distinctive symptoms including the conversion of floral organs into leaf-like organs, known as phyllody. Phyllody is induced by an effector protein family called phyllogens, which interact with floral MADS-box transcription factors (MTFs) responsible for determining the identity of floral organs. The MTF/phyllogen complex then interacts with the proteasomal shuttle protein RADIATION SENSITIVE23 (RAD23), which facilitates delivery of the MTF/phyllogen complex to the host proteasome for MTF degradation. Previous studies have indicated that the MTF degradation specificity of phyllogens is determined by their ability to bind to MTFs. However, in the present study, we discovered a novel mechanism determining the degradation specificity through detailed functional analyses of a phyllogen homologue of rice yellow dwarf phytoplasma (PHYL RYD ). PHYL RYD degraded a narrower range of floral MTFs than other phyllody-inducing phyllogens, resulting in compromised phyllody phenotypes in plants. Interestingly, PHYL RYD was able to bind to some floral MTFs that PHYL RYD was unable to efficiently degrade. However, the complex of PHYL RYD and the non-degradable MTF could not interact with RAD23. These results indicate that the MTF degradation specificity of PHYL RYD is correlated with the ability to form the MTF/PHYL RYD /RAD23 ternary complex, rather than the ability to bind to MTF. This study elucidated that phyllogen target specificity is regulated by both the MTF-binding ability and RAD23 recruitment ability of the MTF/phyllogen complex., (© 2023 The Authors. Molecular Plant Pathology published by British Society for Plant Pathology and John Wiley & Sons Ltd.)

The interaction between the gut microbiota and the host can influence the host's immune system. Bifidobacterium , a commensal genus of gut bacteria, seems to have positive effects on host health. Our previous clinical research showed that B. longum subsp. longum BB536 enhanced innate and adaptive immune responses in elderly individuals with a lower grade of immunity, but the immunomodulatory mechanism is still unclear. In this study, dendritic cell (DC) surface markers in peripheral blood mononuclear cells isolated from healthy individuals were evaluated through coculture with heat-killed BB536. DC markers, innate immune activity and cytokine levels in plasma were also evaluated by a randomized, double-blind, placebo-controlled, parallel-group study (UMIN000045564) with 4 weeks of continuous live BB536 intake. BB536 significantly increased the expression of CD86 and HLA-DR on plasmacytoid DCs (pDCs) in vitro. Compared to placebo ( n = 48), a significant increase in the expression of CD86 on peripheral pDCs was detected at week 4 of live BB536 intake ( n = 49; 1 × 10 10 CFU/day). Furthermore, coculture with hk-BB536 significantly increased the IFNγ expression level and demonstrated trends of increased IFNα1 and IFNβ expression. These findings suggest that consumption of BB536 has potential immunomodulatory effects on healthy individuals through the activation of peripheral pDCs.

We previously reported that the intake of heat-killed Lacticaseibacillus paracasei MCC1849 suppressed the onset of cold-like symptoms in healthy young women who were susceptible to colds. This study aimed to investigate the effects of MCC1849 on subjective symptoms of physical condition in healthy adults of a wide age range. In this randomized, double-blind, placebo-controlled, parallel-group study, 200 healthy adults were randomly divided into the MCC1849 group or placebo group. The participants received test powder with 50 billion MCC1849 cells or placebo powder without MCC1849 for 24 weeks. Subjective symptoms were assessed by diary scores. Analysis was performed on 183 participants (MCC1849 group; n = 91, placebo group; n = 92) in the per-protocol set. The number of days of stuffy nose and cold-like symptoms was significantly reduced in the MCC1849 group compared with the placebo group. In addition, the duration of stuffy nose, sore throat and cold-like symptoms was significantly lower in the MCC1849 group. No side effects were observed. Therefore, oral intake of MCC1849 suppressed subjective symptoms in healthy adults of a wide age range. These data suggest that MCC1849 may help maintain physical condition.

When mood states are impaired, daily life is severely disrupted. To maintain a specific mood state, both positive and negative moods must be controlled; however, methods to maintain a positive mood have not been fully established. Previous studies have suggested that heat-killed L. helveticus MCC1848 has the potential to improve positive moods. This study aimed to test the efficacy of heat-killed L. helveticus MCC1848 in maintaining and improving a positive mood with PANAS, a questionnaire specifically designed to assess positive and negative mood, as the primary endpoint. Healthy Japanese nursing students ( n = 46) were randomized to receive heat-killed L. helveticus MCC1848 (5 billion/day) or placebo powder for four weeks. Mood state was assessed before and two and four weeks after the intervention began; ingestion of heat-killed L. helveticus MCC1848 significantly improved PANAS 'Positive Affect' compared to the placebo. These results indicate that heat-killed L. helveticus MCC1848 is effective in enhancing positive mood.

Phytoplasmas are obligate intracellular plant pathogenic bacteria that can induce phyllody, which is a type of abnormal floral organ development. Phytoplasmas possess phyllogens, which are effector proteins that cause phyllody in plants. Phylogenetic comparisons of phyllogen and 16S rRNA genes have suggested that phyllogen genes undergo horizontal transfer between phytoplasma species and strains. However, the mechanisms and evolutionary implications of this horizontal gene transfer are unclear. Here, we analyzed synteny in phyllogen flanking genomic regions from 17 phytoplasma strains that were related to six ' Candidatus ' species, including three strains newly sequenced in this study. Many of the phyllogens were flanked by multicopy genes within potential mobile units (PMUs), which are putative transposable elements found in phytoplasmas. The multicopy genes exhibited two distinct patterns of synteny that correlated with the linked phyllogens . The low level of sequence identities and partial truncations found among these phyllogen flanking genes indicate that the PMU sequences are deteriorating, whereas the highly conserved sequences and functions (e.g., inducing phyllody) of the phyllogens suggest that the latter are important for phytoplasma fitness. Furthermore, although their phyllogens were similar, PMUs in strains related to ' Ca . P. asteris' were often located in different regions of the genome. These findings strongly indicate that PMUs drive the horizontal transfer of phyllogens among phytoplasma species and strains. These insights improve our understanding of how symptom-determinant genes have been shared among phytoplasmas., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Tokuda, Iwabuchi, Kitazawa, Nijo, Suzuki, Maejima, Oshima, Namba and Yamaji.)

To understand protein function deeply, it is important to identify how it interacts physically with its target. Phyllogen is a phyllody-inducing effector that interacts with the K domain of plant MADS-box transcription factors (MTFs), which is followed by proteasome-mediated degradation of the MTF. Although several amino acid residues of phyllogen have been identified as being responsible for the interaction, the exact interface of the interaction has not been elucidated. In this study, we comprehensively explored interface residues based on random mutagenesis using error-prone PCR. Two novel residues, at which mutations enhanced the affinity of phyllogen to MTF, were identified. These residues, and all other known interaction-involved residues, are clustered together at the surface of the protein structure of phyllogen, indicating that they constitute the interface of the interaction. Moreover, in silico structural prediction of the protein complex using ColabFold suggested that phyllogen interacts with the K domain of MTF via the putative interface. Our study facilitates an understanding of the interaction mechanisms between phyllogen and MTF., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Kitazawa, Iwabuchi, Maejima, Matsumoto, Suzuki, Matsuyama, Koinuma, Oshima, Namba and Yamaji.)

Bifidobacteria are important intestinal bacteria that provide a variety of health benefits in infants. We investigated the efficacy and safety of Bifidobacterium longum subsp. infantis ( B. infantis ) M-63 in healthy infants in a double-blind, randomized, placebo-controlled trial. Healthy term infants were given B. infantis M-63 (n = 56; 1 × 10 9 CFU/day) or placebo (n = 54) from postnatal age ≤ 7 days to 3 months. Fecal samples were collected, and fecal microbiota, stool pH, short-chain fatty acids, and immune substances were analyzed. Supplementation with B. infantis M-63 significantly increased the relative abundance of Bifidobacterium compared with the placebo group, with a positive correlation with the frequency of breastfeeding. Supplementation with B. infantis M-63 led to decreased stool pH and increased levels of acetic acid and IgA in the stool at 1 month of age compared with the placebo group. There was a decreased frequency of defecation and watery stools in the probiotic group. No adverse events related to test foods were observed. These results indicate that early supplementation with B. infantis M-63 is well tolerated and contributes to the development of Bifidobacterium -predominant gut microbiota during a critical developmental phase in term infants.

We report the complete genome sequence of a Japanese isolate of Tea plant necrotic ring blotch virus (TPNRBV-J). The predicted TPNRBV-J genes have the same organization as those of a Chinese isolate, and the 5' termini of the segments have conserved nucleotide sequences.

Clover yellow mosaic virus (ClYMV) infecting white clover was isolated in Japan, and the complete genome sequence was determined.

Plant pathogenic bacteria have developed effectors to manipulate host cell functions to facilitate infection. A certain number of effectors use the conserved ubiquitin-proteasome system in eukaryotic to proteolyze targets. The proteasome utilization mechanism is mainly mediated by ubiquitin interaction with target proteins destined for degradation. Phyllogens are a family of protein effectors produced by pathogenic phytoplasmas that transform flowers into leaves in diverse plants. Here, we present a noncanonical mechanism for phyllogen action that involves the proteasome and is ubiquitin-independent. Phyllogens induce proteasomal degradation of floral MADS-box transcription factors (MTFs) in the presence of RADIATION-SENSITIVE23 (RAD23) shuttle proteins, which recruit ubiquitinated proteins to the proteasome. Intracellular localization analysis revealed that phyllogen induced colocalization of MTF with RAD23. The MTF/phyllogen/RAD23 ternary protein complex was detected not only in planta but also in vitro in the absence of ubiquitin, showing that phyllogen directly mediates interaction between MTF and RAD23. A Lys-less nonubiquitinated phyllogen mutant induced degradation of MTF or a Lys-less mutant of MTF. Furthermore, the method of sequential formation of the MTF/phyllogen/RAD23 protein complex was elucidated, first by MTF/phyllogen interaction and then RAD23 recruitment. Phyllogen recognized both the evolutionarily conserved tetramerization region of MTF and the ubiquitin-associated domain of RAD23. Our findings indicate that phyllogen functionally mimics ubiquitin as a mediator between MTF and RAD23., (© American Society of Plant Biologists 2022. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

The complete genome sequence of a pokeweed (Phytolacca americana L.) isolate of mirabilis crinkle mosaic virus (MiCMV) in Japan was determined.

The colonization and persistence of probiotics introduced into the adult human gut appears to be limited. It is uncertain, however, whether probiotics can successfully colonize the intestinal tracts of full-term and premature infants. In this study, we investigated the colonization and the effect of oral supplementation with Bifidobacterium breve M-16V on the gut microbiota of low birth weight (LBW) infants. A total of 22 LBW infants (12 infants in the M-16V group and 10 infants in the control group) were enrolled. B. breve M-16V was administrated to LBW infants in the M-16V group from birth until hospital discharge. Fecal samples were collected from each subject at weeks (3.7-9.3 weeks in the M-16V group and 2.1-6.1 weeks in the control group) after discharge. qPCR analysis showed that the administrated strain was detected in 83.3% of fecal samples in the M-16V group (at log 10 8.33 ± 0.99 cell numbers per gram of wet feces), suggesting that this strain colonized most of the infants beyond several weeks post-administration. Fecal microbiota analysis by 16S rRNA gene sequencing showed that the abundance of Actinobacteria was significantly higher ( P < 0.01), whereas that of Proteobacteria was significantly lower ( P < 0.001) in the M-16V group as compared with the control group. Notably, the levels of the administrated strain and indigenous Bifidobacterium bacteria were both significantly higher in the M-16V group than in the control group. Our findings suggest that oral administration of B. breve M-16V led to engraftment for at least several weeks post-administration and we observed a potential overall improvement in microbiota formation in the LBW infants' guts., Competing Interests: AH, TO, NI, and J-ZX were employees of Morinaga Milk Industry Co., Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Horigome, Hisata, Odamaki, Iwabuchi, Xiao and Shimizu.)

Flower malformation represented by phyllody is a common symptom of phytoplasma infection induced by a novel family of phytoplasma effectors called phyllogens. Despite the accumulation of functional and structural phyllogen information, the molecular mechanisms of phyllody have not yet been integrated with their evolutionary aspects due to the limited data on their homologs across diverse phytoplasma lineages. Here, we developed a novel universal PCR-based approach to identify 25 phytoplasma phyllogens related to nine "Candidatus Phytoplasma" species, including four species whose phyllogens have not yet been identified. Phylogenetic analyses showed that the phyllogen family consists of four groups (phyl-A, -B, -C, and -D) and that the evolutionary relationships of phyllogens were significantly distinct from those of phytoplasmas, suggesting that phyllogens were transferred horizontally among phytoplasma strains and species. Although phyllogens belonging to the phyl-A, -C, and -D groups induced phyllody, the phyl-B group lacked the ability to induce phyllody. Comparative functional analyses of phyllogens revealed that a single amino acid polymorphism in phyl-B group phyllogens prevented interactions between phyllogens and A- and E-class MADS domain transcription factors (MTFs), resulting in the inability to degrade several MTFs and induce phyllody. Our finding of natural variation in the function of phytoplasma effectors provides new insights into molecular mechanisms underlying the aetiology of phytoplasma diseases., (© 2020 The Authors. Molecular Plant Pathology published by British Society for Plant Pathology and John Wiley & Sons Ltd.)

Bifidobacterium breve M-16V is a probiotic bacterial strain with efficacy in infants achieved by suppressing T-helper type (Th) 2 immune responses and modulating the systemic Th1/Th2 balance. Exposure to air pollution during pregnancy increases asthma susceptibility in offspring. The aim of this study was to investigate the effects of the maternal intake of B. breve M-16V on susceptibility to asthma accelerated by prenatal exposure to air pollution. The intake of B. breve M-16V in residual oil fly ash (ROFA)-exposed pregnant mice resulted in fewer eosinophils in the bronchoalveolar lavage fluid of neonatal mice and reduced allergic lung inflammation. The expressions of Th2 cytokines including IL-5 and IL-13 were decreased in neonatal mice from ROFA-exposed mothers fed B. breve M-16V. The analysis of fecal microbiota from neonatal mice revealed that the intake of B. breve M-16V by mothers changed the composition of fecal microbiota in neonatal mice, which resulted in a decreased population of Firmicutes. Moreover, several bacterial strains of fecal microbiota from neonatal mice had a strong correlation with Th2 cytokines and histological score. These results suggest that the maternal intake of M-16V might have beneficial effects in neonates by preventing and/or alleviating allergic reactions accelerated by prenatal exposure to air pollution., Competing Interests: All authors have read the manuscript and have approved this submission. The authors have the journal's policy and have the following conflicts: A. Hiraku, K. Kato and N. Iwabuchi are employees of Morinaga Milk Industry Co., Ltd. The Morinaga Milk Industry Co., Ltd. is the commercial company that sells dairy products like milk, yoghurt, infant formula and so on. The Morinaga Milk Industry Co., Ltd. has filed patent applications on aspects of this work. A. Hiraku, N. Iwabuchi, and Dr. Ito are named as inventors on patent application number WO2019/163257 (patent name: COMPOSITION FOR PREVENTING OR RELIEVING RESPIRATORY DISEASE). These do not alter our adherence to all the PLOS ONE policies on sharing data and materials.

Background and Purpose- Multilineage-differentiating stress-enduring cells are endogenous nontumorigenic reparative pluripotent-like stem cells found in bone marrow, peripheral blood, and connective tissues. Topically administered human multilineage-differentiating stress-enduring cells into rat/mouse stroke models differentiated into neural cells and promoted clinically relevant functional recovery. However, critical questions on the appropriate timing and dose, and safety of the less invasive intravenous administration of clinical-grade multilineage-differentiating stress-enduring cell-based product CL2020 remain unanswered. Methods- Using an immunodeficient mouse lacunar model, CL2020 was administered via the cervical vein in different doses (high dose=5×10 4 cells/body; medium dose=1×10 4 cells/body; low dose=5×10 3 cells/body) at subacute phase (≈9 days after onset) and chronic phase (≈30 days). Cylinder test, depletion of human cells by diphtheria toxin administration, immunohistochemistry, and human specific-genome detection were performed. Results- Tumorigenesis and adverse effects were not detected for up to 22 weeks. The high-dose group displayed significant functional recovery compared with the vehicle group in cylinder test in subacute-phase-treated and chronic-phase-treated animals after 6 weeks and 8 weeks post-injection, respectively. In the high-dose group of subacute-phase-treated animals, robust and stable recovery in cylinder test persisted up to 22 weeks compared with the vehicle group. In both groups, intraperitoneal injection of diphtheria toxin abrogated the functional recovery. Anti-human mitochondria revealed CL2020 distributed mainly in the peri-infarct area at 1, 10, and 22 weeks and expressed NeuN (neuronal nuclei)- and MAP-2 (microtubule-associated protein-2)-immunoreactivity. Conclusions- Intravenously administered CL2020 was safe, migrated to the peri-infarct area, and afforded functional recovery in experimental stroke.

Peptidoglycan recognition proteins (PGLYRPs) are a family of pattern recognition receptors (PRRs) that are able to induce innate immune responses through their binding to peptidoglycan (PGN), lipopolysaccharide, or lipoteichoic acid, or by interacting with other PRR-ligands. Recently, progress has been made in understanding the immunobiology of PGLYRPs in human and mice, however, their functions in livestock animals have been less explored. In this study, we characterized the expression patterns of PGLYRPs in porcine intestinal epithelial (PIE) cells and antigen-presenting cells (APCs) and their modulation by the interactions of host cells with PRR-ligands and non-viable immunomodulatory probiotics referred to as paraimmunobiotics. We demonstrated that PGLYRP-1, -2, -3, and -4 are expressed in PIE cells and APCs from Peyer's patches, being PGLYPR-3 and -4 levels higher than PGLYRP-1 and -2. We also showed that PGLYRPs expression in APCs and PIE cells can be modulated by different PRR agonists. By using knockdown PIE cells for TLR2, TLR4, NOD1, and NOD2, or the four PGLYRPs, we demonstrated that PGLYRPs expressions would be required for activation and functioning of TLR2, TLR4, NOD1, and NOD2 in porcine epitheliocytes, but PGLYRPs activation would be independent of those PRR expressions. Importantly, we reported for the first time that PGLYRPs expression can be differentially modulated by paraimmunobiotic bifidobacteria in a strain-dependent manner. These results provide evidence for the use of paraimmunobiotic bifidobacteria as an alternative for the improvement of resistance to intestinal infections or as therapeutic tools for the reduction of the severity of inflammatory damage in diseases in which a role of PGLYRPs-microbe interaction has been demonstrated.

Probiotics intervention has been proposed as a feasible preventative approach against adverse health-related complications in infants. Nevertheless, the umbrella concept of probiotics has led to a massive application of probiotics in a range of products for promoting infant health, for which the strain-specificity, safety and efficacy findings associated with a specific probiotics strain are not clearly defined. Bifidobacterium breve M-16V is a commonly used probiotic strain in infants. M-16V has been demonstrated to offer potential in protecting infants from developing the devastating necrotising enterocolitis (NEC) and allergic diseases. This review comprehends the potential beneficial effects of M-16V on infant health particularly in the prevention and treatment of premature birth complications and immune-mediated disorders in infants. Mechanistic studies supporting the use of M-16V implicated that M-16V is capable of promoting early gut microbial colonisation and may be involved in the regulation of immune balance and inflammatory response to protect high-risk infants from NEC and allergies. Summarised information on M-16V has provided conceptual proof of the use of M-16V as a potential probiotics candidate aimed at promoting infant health, particularly in the vulnerable preterm population., Competing Interests: The authors, C.B.W., N.I., and J.-Z.X. are employees of Morinaga Milk Industry Co., Ltd., which has several probiotic products marketed worldwide. This does not alter our adherence to Nutrients policies on sharing data and materials.

The complete genome sequence of the first Japanese isolate of carrot torradovirus 1 (CaTV1-J), which infects Angelica keiskei , was determined. This is the first report of a CaTV1 isolate obtained from A. keiskei ., (Copyright © 2019 Tokuda et al.)

Phytoplasmas are plant-pathogenic bacteria that infect many important crops and cause serious economic losses worldwide. However, owing to an inability to culture phytoplasmas, screening of antimicrobials on media is difficult. The only antimicrobials being used to control phytoplasmas are tetracycline-class antibiotics. In this study, we developed an accurate and efficient screening method to evaluate the effects of antimicrobials using an in vitro plant-phytoplasma co-culture system. We tested 40 antimicrobials, in addition to tetracycline, and four of these (doxycycline, chloramphenicol, thiamphenicol and rifampicin) decreased the accumulation of 'Candidatus (Ca.) Phytoplasma asteris'. The phytoplasma was eliminated from infected plants by the application of both tetracycline and rifampicin. We also compared nucleotide sequences of rRNAs and amino acid sequences of proteins targeted by antimicrobials between phytoplasmas and other bacteria. Since antimicrobial target sequences were conserved among various phytoplasma species, the antimicrobials that decreased accumulation of 'Ca. P. asteris' may also have been effective against other phytoplasma species. These approaches will provide new strategies for phytoplasma disease management.

Antigen-specific immunoglobulin (Ig) A plays a major role in host defense against infections in gut mucosal tissue. Follicular helper T (Tfh) cells are located in germinal centers and promote IgA production via interactions with germinal center B cells. Several studies have demonstrated that some lactic acid bacteria (LAB) strains activate the host's acquired immune system, inducing IgA secretion in the intestine. However, the precise molecular mechanisms underlying the effects of LAB on IgA production and Tfh cells are not fully resolved. Lactobacillus paracasei MCC1849 is a probiotic strain isolated from the intestine of a healthy adult. In this study, we investigated the effects of orally administered heat-killed MCC1849 on IgA production in the intestine and on Tfh cell induction in vivo. We found that orally administered MCC1849 induced antigen-specific IgA production in the small intestine, serum and lungs. We also observed that MCC1849 increased the proportion of IgA+ B cells and Tfh cells in Peyer's patches (PPs). In addition, MCC1849 increased the gene expression of IL-12p40, IL-10, IL-21, STAT4 and Bcl-6 associated with Tfh cell differentiation. These results suggest that orally administered MCC1849 enhances antigen-specific IgA production and likely affects Tfh cell differentiation in PPs., Competing Interests: All the authors have read the manuscript and have approved this submission. The authors have the journal's policy and have the following conflicts: S. Arai, N. Iwabuchi, S. Takahashi, J-Z Xiao and F. Abe are employees of Morinaga Milk Industry Co., Ltd. The Morinaga Milk Industry Co., Ltd. is the commercial company that sells dairy products like milk, yoghurt, infant formula and so on. The Morinaga Milk Industry Co., Ltd. has filed patent applications on aspects of this work. S. Arai, N. Iwabuchi, and Dr. Hachimura are named as inventors on patent application number P2016-113378A (patent name: Increasing agent for follicular helper T cells). N. Iwabuchi, and J-Z Xiao are named as inventors with the Morinaga Milk Industry Co., Ltd. on patent application number WO2015/004949 A1 (patent name: Novel lactobacillus and novel lactobacillus-containing medicine, food, beverage and feed). These do not alter our adherence to all the PLOS ONE policies on sharing data and materials.

Accumulating evidence suggests a relationship between the gut microbiota and the development of obesity, indicating the potential of probiotics as a therapeutic approach. Bifidobacterium breve B-3 has been shown to exert anti-obesity effects in high-fat diet-induced obese mice. In the present study, the anti-obesity effects of the consumption of B. breve B-3 by healthy pre-obese (25 ≤ BMI < 30) adults were investigated in a randomized, double-blind, placebo-controlled trial (trial registration: UMIN-CTR No. 000023919; preregistered on September 2, 2016). Eighty participants were randomized to receive placebo or B. breve B-3 capsules (2 × 10 10 CFU/day) daily for 12 weeks. The visceral fat area significantly increased at weeks 4 and 8 in the placebo group only; no significant change was observed in the B-3 group. Body fat mass and percent body fat were significantly lower in the B-3 group than in the placebo group at weeks 8 and 12 (p<0.05, ANCOVA adjusted with baseline values). Although no significant differences were observed in blood parameters between the groups, the intake of B. breve B-3 slightly decreased triglyceride levels and improved HDL cholesterol from the baseline. No serious adverse effects were noted in either group. These results suggest that the probiotic strain B. breve B-3 has potential as a functional food ingredient to reduce body fat in healthy pre-obese individuals.

ABCE-class MADS domain transcription factors (MTFs) are key regulators of floral organ development in angiosperms. Aberrant expression of these genes can result in abnormal floral traits such as phyllody. Phyllogen is a virulence factor conserved in phytoplasmas, plant pathogenic bacteria of the class Mollicutes. It triggers phyllody in Arabidopsis thaliana by inducing degradation of A- and E-class MTFs. However, it is still unknown whether phyllogen can induce phyllody in plants other than A. thaliana, although phytoplasma-associated phyllody symptoms are observed in a broad range of angiosperms. In this study, phyllogen was shown to cause phyllody phenotypes in several eudicot species belonging to three different families. Moreover, phyllogen can interact with MTFs of not only angiosperm species including eudicots and monocots but also gymnosperms and a fern, and induce their degradation. These results suggest that phyllogen induces phyllody in angiosperms and inhibits MTF function in diverse plant species., (© The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

One of the important antiviral genetic strategies used in crop breeding is recessive resistance. Two eukaryotic translation initiation factor 4E family genes, eIF4E and eIFiso4E, are the most common recessive resistance genes whose absence inhibits infection by plant viruses in Potyviridae, Carmovirus, and Cucumovirus. Here, we show that another eIF4E family gene, nCBP, acts as a novel recessive resistance gene in Arabidopsis thaliana toward plant viruses in Alpha- and Betaflexiviridae. We found that infection by Plantago asiatica mosaic virus (PlAMV), a potexvirus, was delayed in ncbp mutants of A. thaliana. Virus replication efficiency did not differ between an ncbp mutant and a wild type plant in single cells, but viral cell-to-cell movement was significantly delayed in the ncbp mutant. Furthermore, the accumulation of triple-gene-block protein 2 (TGB2) and TGB3, the movement proteins of potexviruses, decreased in the ncbp mutant. Inoculation experiments with several viruses showed that the accumulation of viruses encoding TGBs in their genomes decreased in the ncbp mutant. These results indicate that nCBP is a novel member of the eIF4E family recessive resistance genes whose loss impairs viral cell-to-cell movement by inhibiting the efficient accumulation of TGB2 and TGB3.

One of the plant host resistance machineries to viruses is attributed to recessive alleles of genes encoding critical host factors for virus infection. This type of resistance, also referred to as recessive resistance, is useful for revealing plant-virus interactions and for breeding antivirus resistance in crop plants. Therefore, it is important to identify a novel host factor responsible for robust recessive resistance to plant viruses. Here, we identified a mutant from an ethylmethane sulfonate (EMS)-mutagenized Arabidopsis population which confers resistance to plantago asiatica mosaic virus (PlAMV, genus Potexvirus). Based on map-based cloning and single nucleotide polymorphism analysis, we identified a premature termination codon in a functionally unknown gene containing a GYF domain, which binds to proline-rich sequences in eukaryotes. Complementation analyses and robust resistance to PlAMV in a T-DNA mutant demonstrated that this gene, named Essential for poteXvirus Accumulation 1 (EXA1), is indispensable for PlAMV infection. EXA1 contains a GYF domain and a conserved motif for interaction with eukaryotic translation initiation factor 4E (eIF4E), and is highly conserved among monocot and dicot species. Analysis using qRT-PCR and immunoblotting revealed that EXA1 was expressed in all tissues, and was not transcriptionally responsive to PlAMV infection in Arabidopsis plants. Moreover, accumulation of PlAMV and a PlAMV-derived replicon was drastically diminished in the initially infected cells by the EXA1 deficiency. Accumulation of two other potexviruses also decreased in exa1-1 mutant plants. Our results provided a functional annotation to GYF domain-containing proteins by revealing the function of the highly conserved EXA1 gene in plant-virus interactions., (© 2016 The Authors. The Plant Journal published by Society for Experimental Biology and John Wiley & Sons Ltd.)

We recently reported that a close relationship exists between alkane carbon-chain length, cell growth, and translocation frequency in Rhodococcus. In the present study, we examined the regulation of the spatial arrangement of cells in aqueous-alkane two phase cultures. An analysis of the effects of minerals on cell localization revealed that changes in the concentration of MgSO4 in two phase cultures containing n-dodecane (C12) altered cell localization from translocation to adhesion and vice versa. Our results indicate that the spatial arrangement of cells in two phase culture systems is controlled through the regulation of MgSO4 concentrations.

The 5'-terminal genomic sequence of Cherry virus A (CVA) has long been unknown. We determined the first complete genome sequence of an apricot isolate of CVA (7,434 nucleotides [nt]). The 5'-untranslated region was 107 nt in length, which was 53 nt longer than those of known CVA sequences., (Copyright © 2016 Koinuma et al.)

Hydrangea ringspot virus (HdRSV) is a plant RNA virus, naturally infectingHydrangea macrophylla Here, we report the first genomic sequences of two HdRSV isolates from hydrangea plants in Japan. The overall nucleotide sequences of these Japanese isolates were 96.0 to 96.3% identical to those of known European isolates., (Copyright © 2016 Yusa et al.)

In this work, we aimed to characterize the antiviral response of an originally established porcine intestinal epithelial cell line (PIE cells) by evaluating the molecular innate immune response to rotavirus (RVs). In addition, we aimed to select immunomodulatory bacteria with antiviral capabilities. PIE cells were inoculated with RVs isolated from different host species and the infective titers and the molecular innate immune response were evaluated. In addition, the protection against RVs infection and the modulation of immune response by different lactic acid bacteria (LAB) strains was studied. The RVs strains OSU (porcine) and UK (bovine) effectively infected PIE cells. Our results also showed that RVs infection in PIE cells triggered TLR3-, RIG-I- and MDA-5-mediated immune responses with activation of IRF3 and NF-κB, induction of IFN-β and up-regulation of the interferon stimulated genes MxA and RNase L. Among the LAB strains tested, Bifidobacterium infantis MCC12 and B. breve MCC1274 significantly reduced RVs titers in infected PIE cells. The beneficial effects of both bifidobacteria were associated with reduction of A20 expression, and improvements of IRF-3 activation, IFN-β production, and MxA and RNase L expressions. These results indicate the value of PIE cells for studying RVs molecular innate immune response in pigs and for the selection of beneficial bacteria with antiviral capabilities.

Alternanthera mosaic virus (AltMV) infecting Achyranthes bidentata was first detected in Asia, and the complete genome sequence (6,604 nucleotides) was determined. Sequence identity analysis and phylogenetic analysis confirmed that this isolate is the most phylogenetically distant AltMV isolate worldwide., (Copyright © 2016 Iwabuchi et al.)

Schizophrenia is a chronic psychiatric illness. Disruption of the dopaminergic system has been suggested to be the pathogenic cause of this disease. The effect of Bifidobacterium longum BB536 (BB536) on schizophrenic behavior was investigated in an animal model. Daily administration of BB536 (10(9) CFU/mouse, p.o. for 2 weeks) was found to reduce rearing behavior augmented by the dopamine receptor agonist apomorphine and to decrease the resting level of plasma corticosterone and the ratio of kynurenine to tryptophan. These results suggest the potential of BB536 for supplemental treatment of the symptoms of schizophrenia.

Background: Probiotic administration may be a useful method for preventing allergies in infants; however, there have been controversial results about the efficacy. We investigated the effects of bifidobacterial supplementation on the risk of developing allergic diseases in the Japanese population., Methods: In an open trial, we gave Bifidobacterium breve M-16V and Bifidobacterium longum BB536 prenatally to 130 mothers beginning 1 month prior to delivery and postnatally to their infants for 6 months. Another 36 mother-infant pairs served as controls and did not receive the bifidobacterial supplementation. Development of allergic symptoms in the infants was assessed at 4, 10 and 18 months of age. Fecal samples were collected from the mothers and infants., Results: The risk of developing eczema/atopic dermatitis (AD) during the first 18 months of life was significantly reduced in infants in the probiotic group (OR: 0.231 [95% CI: 0.084-0.628] and 0.304 [0.105-0.892] at 10 and 18 months of age, respectively). Pyrosequencing analyses indicated an altered composition of the fecal microbiota at 4 months for infants who developed eczema/AD at 4 and 10 months of age. The proportion of Proteobacteria was significantly lower (P = 0.007) in mothers at the time of delivery who received the supplementation when compared with the control group and was positively correlated (r = 0.283, P = 0.024) with that of infants at 4 months of age. No adverse effects were related to the use of probiotics., Conclusions: These data suggest that the prenatal and postnatal supplementation of bifidobacteria is effective in primary preventing allergic diseases. Some limited changes in the composition of fecal microbiota by the bifidobacterial supplementation were observed.

We previously reported that R. erythropolis PR4 translocated from the aqueous to the alkane phase, and then grew in two phase cultures to which long-chain alkanes had been added. This was considered to be beneficial for bioremediation. In the present study, we investigated the proteins involved in the translocation of R. erythropolis PR4. The results of our proteogenomic analysis suggested that GroEL2 was upregulated more in cells that translocated inside of the pristane (C19) phase than in those located at the aqueous-alkane interface attached to the n-dodecane (C12) surface. PR4 (pK4-EL2-1) and PR4 (pK4-ΔEL2-1) strains were constructed to confirm the effects of the upregulation of GroEL2 in translocated cells. The expression of GroEL2 in PR4 (pK4-EL2-1) was 15.5-fold higher than that in PR4 (pK4-ΔEL2-1) in two phase cultures containing C12. The growth and cell surface lipophilicity of PR4 were enhanced by the introduction of pK4-EL2-1. These results suggested that the plasmid overexpression of groEL2 in PR4 (pK4-EL2-1) led to changes in cell localization, enhanced growth, and increased cell surface lipophilicity. Thus, we concluded that the overexpression of GroEL2 may play an important role in increasing the organic solvent tolerance of R. erythropolis PR4 in aqueous-alkane two phase cultures.

We recently reported that the overexpression of GroEL2 played an important role in increasing the alkane tolerance of Rhodococcus erythropolis PR4. In the present study, we examined the effects of the introduction of groEL2 on the alkane tolerance of other Rhodococcus strains. The introduction of groEL2 into Rhodococcus strains led to increased alkane tolerance. The translocation of R. rhodochrous ATCC12674 cells to and survival in the n-octane (C8) phase in two phase culture were significantly enhanced by the introduction of groEL2 derived from strain PR4, suggesting that engineering cells to overexpress GroEL2 represents an effective strategy for enhancing organic solvent tolerance in Rhodococcus.

Bifidobacterium breve MCC-117 is able to significantly reduce the expression of inflammatory cytokines in porcine intestinal epithelial (PIE) cells and to improve IL-10 levels in CD4(+)CD25(high) Foxp3(+) lymphocytes in response to heat-stable enterotoxigenic Escherichia coli (ETEC) pathogen-associated molecular patterns (PAMPs), while the immunoregulatory effect of B. adolescentis ATCC15705 was significantly lower than that observed for the MCC-117 strain. Considering the different capacities of the two bifidobacterium strains to activate toll-like receptor (TLR)-2 and their differential immunoregulatory activities in PIE and immune cells, we hypothesized that comparative studies with both strains could provide important information regarding the molecular mechanism(s) involved in the anti-inflammatory activity of bifidobacteria. In this work, we demonstrated that the anti-inflammatory effect of B. breve MCC-117 was achieved by a complex interaction of multiple negative regulators of TLRs as well as inhibition of multiple signaling pathways. We showed that B. breve MCC-117 reduced heat-stable ETEC PAMP-induced NF-κB, p38 MAPK and PI3 K activation and expression of pro-inflammatory cytokines in PIE cells. In addition, we demonstrated that B. breve MCC-117 may activate TLR2 synergistically and cooperatively with one or more other pattern recognition receptors (PRRs), and that interactions may result in a coordinated sum of signals that induce the upregulation of A20, Bcl-3, Tollip and SIGIRR. Upregulation of these negative regulators could have an important physiological impact on maintaining or reestablishing homeostatic TLR signals in PIE cells. Therefore, in the present study, we gained insight into the molecular mechanisms involved in the immunoregulatory effect of B. breve MCC-117.

Background: We previously showed that evaluation of anti-inflammatory activities of lactic acid bacteria in porcine intestinal epithelial (PIE) cells is useful for selecting potentially immunobiotic strains., Objective: The aims of the present study were: i) to select potentially immunomodulatory bifidobacteria that beneficially modulate the Toll-like receptor (TLR)-4-triggered inflammatory response in PIE cells and; ii) to gain insight into the molecular mechanisms involved in the anti-inflammatory effect of immunobiotics by evaluating the role of TLR2 and TLR negative regulators in the modulation of proinflammatory cytokine production and activation of mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB) pathways in PIE cells., Results: Bifidobacteria longum BB536 and B. breve M-16V strains significantly downregulated levels of interleukin (IL)-8, monocyte chemotactic protein (MCP)-1 and IL-6 in PIE cells challenged with heat-killed enterotoxigenic Escherichia coli. Moreover, BB536 and M-16V strains attenuated the proinflammatory response by modulating the NF-κB and MAPK pathways. In addition, our findings provide evidence for a key role for the ubiquitin-editing enzyme A20 in the anti-inflammatory effect of immunobiotic bifidobacteria in PIE cells., Conclusions: We show new data regarding the mechanism involved in the anti-inflammatory effect of immunobiotics. Several strains with immunoregulatory capabilities used a common mechanism to induce tolerance in PIE cells. Immunoregulatory strains interacted with TLR2, upregulated the expression of A20 in PIE cells, and beneficially modulated the subsequent TLR4 activation by reducing the activation of MAPK and NF-κB pathways and the production of proinflammatory cytokines. We also show that the combination of TLR2 activation and A20 induction can be used as biomarkers to screen and select potential immunoregulatory bifidobacteria strains.