Immune and allergenic effects of the microalga Coccomyxa sp. strain KJ in healthy humans: A pilot study.

BACKGROUND: The Coccomyxa sp. strain KJ (Coccomyxa KJ), a microalga found in Japan, has a potential function in controlling viral infections. Recently, its dry powder has been marketed as a health food product. OBJECTIVES: This pilot study investigated the effects of Coccomyxa KJ powder tablet intake on allergic reactions and immune functions in healthy participants. MATERIAL AND METHODS: Nine healthy volunteers (4 males and 5 females) who expressed interest in foods containing Coccomyxa KJ, and were willing to undergo blood tests, were recruited. Each individual was asked to take 2 Coccomyxa KJ powder tablets (0.3 g) before breakfast once a day for 4 weeks. The salivary immunoglobulin A (IgA) level and blood parameters (white blood cell (WBC) count, eosinophil and lymphocyte counts and percentages, natural killer (NK) cell activity, interleukin (IL)-6 level, and T helper (Th)1/Th2 cell ratio) were evaluated at baseline and weeks 2 and 4. RESULTS: The 4-week intake of Coccomyxa KJ did not affect salivary IgA levels, WBC count, eosinophil and lymphocyte counts and percentages, or the Th1/Th2 ratio. There were significant differences in the NK cell activity after 4 weeks, with an average increase of 11.78 (95% confidence interval (95% CI): 6.80-16.76). None of the patients experienced adverse reactions during or after the study. CONCLUSIONS: Long-term Coccomyxa KJ intake improved NK cell activity without causing adverse effects on the indicators of local immunity, systemic inflammation and immune response balance. This study suggests that Coccomyxa KJ powder tablets can induce beneficial immune modifications without causing any adverse effects.

Electrical biosensing system utilizing ion-producing enzymes conjugated with aptamers for the sensing of severe acute respiratory syndrome coronavirus 2.

Viral outbreaks, which include the ongoing coronavirus disease 2019 (COVID-19) pandemic provoked by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), are a major global crisis that enormously threaten human health and social activities worldwide. Consequently, the rapid and repeated treatment and isolation of these viruses to control their spread are crucial to address the COVID-19 pandemic and future epidemics of novel emerging viruses. The application of cost-efficient, rapid, and easy-to-operate detection devices with miniaturized footprints as a substitute for the conventional optic-based polymerase chain reaction (PCR) and immunoassay tests is critical. In this context, semiconductor-based electrical biosensors are attractive sensing platforms for signal readout. Therefore, this study aimed to examine the electrical sensing of patient-derived SARS-CoV-2 samples by harnessing the activity of DNA aptamers directed against spike proteins on viral surfaces. We obtained rapid and sensitive virus detection beyond the Debye length limitation by exploiting aptamers coupled with alkaline phosphatases, which catalytically generate free hydrogen ions which can readily be measured on pH meters or ion-sensitive field-effect transistors. Furthermore, we demonstrated the detection of the viruses of approximately 100 copies/µL in 10 min, surpassing the capability of typical immunochromatographic assays. Therefore, our newly developed technology has great potential for point-of-care testing not only for SARS-CoV-2, but also for other types of pathogens and biomolecules.

Phenol Derivatives Obtained from Grape Seed Extract Show Virucidal Activity against Murine Norovirus.

Human noroviruses are the most common pathogens known to cause acute gastroenteritis, a condition that can lead to severe illness among immunocompromised individuals such as organ transplant recipients and the elderly. To date, no safe and effective vaccines or therapeutic agents have been approved for treating norovirus infections. Therefore, we aimed to demonstrate the virucidal activity of grape seed extract (GSE), which contains >83% proanthocyanidins, against murine norovirus (MNV), a surrogate for human norovirus. GSE showed virucidal activity against MNV in a dose- and time-dependent manner. Atomic force microscopic analysis showed viral particle aggregates after treatment of MNV with GSE. MNV treated with 50 µg/mL of GSE for 10 min resulted in the absence of pathogenicity in an animal model of infection, indicating that GSE has irreversible virucidal activity against MNV particles. Thus, GSE may aid in the development of treatments for norovirus infections.

Dietary Exposure of the Japanese General Population to Elements: Total Diet Study 2013-2018.

Some countries have conducted a total diet study (TDS) focused on the estimation of specific trace elements. Although some results of a Japanese TDS examining trace elements were published, there have been no reports of a nationwide TDS across Japan over a multi-year period to estimate the level of exposure to multiple elements. In the present study, a TDS using a market basket approach was performed to estimate the dietary exposure levels of the general population of Japan to 15 elements, including aluminum (Al), total arsenic (tAs), boron (B), barium (Ba), cadmium (Cd), cobalt (Co), chromium (Cr), total mercury (THg), molybdenum (Mo), nickel (Ni), lead (Pb), antimony (Sb), selenium (Se), tin (Sn), and uranium (U). Samples prepared in eight regions across Japan over a 6-year period were analyzed using validated methods. The robust mean exposure estimates for Al, tAs, B, Ba, Cd, Co, Cr, THg, Mo, Ni, Pb, Sb, Se, Sn, and U were 48, 4.2, 29, 8.6, 0.35, 0.17, 0.49, 0.14, 4.2, 2.8, 0.15, 0.022, 1.8, 0.10, and 0.021 µg/kg body weight/day, respectively. Although the variability in exposure estimates varied greatly from element to element, the relative standard deviations calculated from the robust means and robust standard deviations were ≤ 50% for all elements except Sn. Compared against the health-based guidance values, none of the robust and precise estimates obtained for the target elements would be associated with urgent health risk concern. In addition, the estimated exposure levels were generally in agreement with previously reported estimates, indicating that health risks associated with exposure to these elements have not changed markedly nationwide in Japan in recent years.

Effect of Oral Administration of Lactiplantibacillus plantarum SNK12 on Temporary Stress in Adults: A Randomized, Placebo-Controlled, Double-Blind, Parallel-Group Study.

Mouse studies have reported anti-stress effects of Lactiplantibacillus plantarum SNK12 (SNK). Specifically, oral SNK administration increased mRNA levels of hippocampal neurotrophic factor and gamma-aminobutyric acid receptor in mice with sub-chronic mild stress-induced social defeat; moreover, it improved depressive behavior. We aimed to evaluate the efficacy of SNK ingestion against stress in healthy adults. We used the Uchida-Kraepelin test for the stress load, with a low-dose (50 mg/day), high-dose (150 mg/day), and placebo groups (dextrin). The primary outcome was the psychological evaluation as measured by the Profile of Mood States 2nd Edition (POMS2) using total mood disturbance (TMD) scores. The secondary outcomes were the score of each POMS2 item, salivary cortisol as a stress marker, and autonomic balance with the low frequency (LF)/ high frequency (HF) ratio. Compared with the placebo group, the SNK ingestion group showed significantly lower TMD scores. Additionally, compared with the placebo group, the high-dose group showed significantly lower scores for Tension-Anxiety and Confusion-Bewilderment, while the low-dose group showed significantly lower Anger-Hostility scores, salivary cortisol levels, and LF/HF scores. Our findings suggest that SNK ingestion could relieve stress (negative feelings, anxiety, tension, embarrassment, confusion, anger, and hostility) resulting from the temporary load caused by work and study.

Virucidal and Immunostimulating Activities of Monogalactosyl Diacylglyceride from Coccomyxa sp. KJ, a Green Microalga, against Murine Norovirus and Feline Calicivirus.

Human noroviruses are the most common pathogens causing acute gastroenteritis and may lead to more severe illnesses among immunosuppressed people, including elderly and organ transplant recipients. To date, there are no safe and effective vaccines or antiviral agents for norovirus infections. In the present study, we aimed to demonstrate the antiviral activity of monogalactosyl diacylglyceride (MGDG) isolated from a microalga, Coccomyxa sp. KJ, against murine norovirus (MNV) and feline calicivirus (FCV), the surrogates for human norovirus. MGDG showed virucidal activities against these viruses in a dose- and time-dependent manner-MGDG at 100 µg/mL reduced the infectivity of MNV and FCV to approximately 10% after 60 min incubation. In the animal experiments of MNV infection, intraoral administration of MGDG (1 mg/day) exerted a therapeutic effect by suppressing viral shedding in the feces and produced high neutralizing antibody titers in sera and feces. When MGDG was orally administered to immunocompromised mice treated with 5-fluorouracil, the compound exhibited earlier stopping of viral shedding and higher neutralizing antibody titers of sera than those in the control mice administered with distilled water. Thus, MGDG may offer a new therapeutic and prophylactic alternative against norovirus infections.

Virucidal effect of monogalactosyl diacylglyceride from a green microalga, Coccomyxa sp. KJ, against clinical isolates of SARS-CoV-2 as assessed by a plaque assay.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of coronavirus disease 2019 (COVID-19) and is capable of human-to-human transmission and rapid global spread. Thus, the establishment of high-quality viral detection and quantification methods, and the development of anti-SARS-CoV-2 agents are critical. METHODS: Here, we present the rapid detection of infectious SARS-CoV-2 particles using a plaque assay with 0.5% agarose-ME (Medium Electroosmosis) as an overlay medium. RESULTS: The plaques were capable of detecting the virus within 36-40 h post-infection. In addition, we showed that a monogalactosyl diacylglyceride isolated from a microalga (Coccomyxa sp. KJ) could inactivate the clinical isolates of SARS-CoV-2 in a time- and concentration-dependent manner. CONCLUSIONS: These results would allow rapid quantification of the infectious virus titers and help develop more potent virucidal agents against SARS-CoV-2.

Physical properties of lactic acid bacteria influence the level of protection against influenza infection in mice.

We evaluated whether the water dispersibility of lactic acid bacteria (Enterococcus faecalis KH2) affects their efficacy. When cultured lactic acid bacteria are washed, heat-killed, and powdered, adhesion occurs between results in aggregation (non-treated lactic acid bacteria, n-LAB). However, dispersed lactic acid bacteria (d-LAB) with a lower number of aggregates can be prepared by treating them with a high-pressure homogenizer and adding an excipient during powdering. Mice were administered n-LAB or d-LAB Peyer's patches in the small intestine were observed. Following n-LAB administration, a high amount of aggregated bacteria drifting in the intestinal mucosa was observed; meanwhile, d-LAB reached the Peyer's patches and was absorbed into them. Evaluation in a mouse influenza virus infection model showed that d-LAB was more effective than n-LAB in the influenza yield of bronchoalveolar lavage fluids on day 3 post-infection and neutralizing antibody titers of sera and influenza virus-specific immunoglobulin A in the feces on day 14 post-infection. Therefore, the physical properties of lactic acid bacteria affect their efficacy; controlling their water dispersibility can improve their effectiveness.

Anti-Influenza virus effects of Enterococcus faecalis KH2 and Lactobacillus plantarum SNK12 RNA.

Bacterial RNA has recently emerged as an immune-stimulating factor during viral infection. The immune response in an organism is directly related to the progression of virus infections. Lactic acid bacteria in particular have anticancer, bioprotective, and antiallergic effects by modulating immunity. Here, we aimed to demonstrate the effect of bacterial RNA on in vitro production of IL-12, a proinflammatory cytokine, and on in vivo activity against influenza A virus (IFV) infection. Oral administration of heat-killed Enterococcus faecalis KH2 (KH2) or Lactobacillus plantarum SNK12 (SNK) in IFV-infected mice suppressed viral replication and stimulated production of virus-specific antibodies. However, ribonuclease-treated KH2 or SNK abrogated the effect, reducing IL-12 production in vitro and anti-IFV effects in vivo. Taken together, KH2 or SNK showed antiviral effects in vivo when administered orally, and the RNAs of KH2 and SNK play a part in these effects, despite the phylogenetic differences between the bacteria.

Anti-Influenza A Virus Activity of Rhamnan Sulfate from Green Algae Monostroma nitidum in Mice with Normal and Compromised Immunity.

Influenza viruses cause a significant public health burden each year despite the availability of anti-influenza drugs and vaccines. Therefore, new anti-influenza virus agents are needed. Rhamnan sulfate (RS) is a sulfated polysaccharide derived from the green alga Monostroma nitidum. Here, we aimed to demonstrate the antiviral activity of RS, especially against influenza A virus (IFV) infection, in vitro and in vivo. RS showed inhibitory effects on viral proliferation of enveloped viruses in vitro. Evaluation of the anti-IFV activity of RS in vitro showed that it inhibited both virus adsorption and entry steps. The oral administration of RS in IFV-infected immunocompetent and immunocompromised mice suppressed viral proliferation in both mouse types. The oral administration of RS also had stimulatory effects on neutralizing antibody production. Fluorescent analysis showed that RS colocalized with M cells in Peyer's patches, suggesting that RS bound to the M cells and may be incorporated into the Peyer's patches, which are essential to intestinal immunity. In summary, RS inhibits influenza virus infection and promotes antibody production, suggesting that RS is a potential candidate for the treatment of influenza virus infections.

In vitro and in vivo anti-herpes simplex virus activity of monogalactosyl diacylglyceride from Coccomyxa sp. KJ (IPOD FERM BP-22254), a green microalga.

A monogalactosyl diacylglyceride (MGDG) was isolated as an antiviral component from Coccomyxa sp. KJ (IPOD FERM BP-22254) via bioassay-guided fractionation. α-Linolenic acid (C18:3) and 7,10,13-hexadecatrienoic acid (C16:3) accounted for approximately 72% and 23%, respectively, of the MGDG total fatty acids of the MGDG. The MGDG showed virucidal activity against herpes simplex virus type 2 (HSV-2), a pathogen that causes genital herpes. Physical changes in HSV-2 shape were observed after treatment with MGDG, including a decrease in particle size, and possible damage to the viral envelope, as assessed using electron microscopy. In accordance with the morphological findings, virus particles lost their ability to bind to host cells. HSV-2 treated with high concentrations of MGDG resulted in no pathogenicity in an animal model, indicating that MGDG exhibits irreversible virucidal activity against HSV-2 particles. In the animal model of HSV-2-induced genital herpes, intravaginally administered MGDG exerted a prophylactic effect by suppressing viral yields in the genital cavity and formation of herpetic lesions, resulting in a higher survival rate in treated mice than control mice administered solvent. Thus, MGDG offers a novel prophylactic option against HSV infections.

[Surveillance of Cadmium Concentration in Chocolate and Cocoa Powder Products Distributed in Japan].

Chocolate and cocoa are manufactured from cacao beans produced by the cacao tree (Theobroma cacao). These products may contain cadmium (Cd), which originates from contaminated soil. Here, we surveyed the Cd concentrations in dark chocolate, milk chocolate, white chocolate and cocoa powder products purchased at retail stores in Japan, using inductively coupled plasma mass spectrometry. The Cd concentrations in these chocolate and cocoa powder products ranged from 0.00021 to 2.3 mg/kg and from 0.015 to 1.8 mg/kg, respectively. A weak positive correlation was found between the Cd concentration and the content of cocoa solids stated on the product labels. A comparison between these results and the maximum levels (MLs) set by the European Union revealed that the Cd concentrations in chocolate and cocoa powder products on the Japanese market exceeded the MLs for eight of the 180 chocolate products and 26 of the 140 cocoa powder products.

Correlation between plasma brain natriuretic peptide levels and left atrial appendage flow velocity in patients with non-valvular atrial fibrillation and normal left ventricular systolic function.

BACKGROUND: The left atrial appendage (LAA) flow velocity is an important factor for thrombus formation in patients with non-valvular atrial fibrillation (NV-AF). Recently, the relation of plasma brain natriuretic peptide (BNP) levels and thromboembolism has been reported in patients with NV-AF. The aim of this study was to determine whether the plasma BNP is predictive of lower LAA flow velocity in patients with NV-AF and normal left ventricular (LV) systolic function. METHODS AND RESULTS: A total of 184 patients with NV-AF (132 men; 65 ± 12 years, LV ejection fraction; 65 ± 10%) underwent transthoracic echocardiography, transesophageal echocardiography (TEE), and measurement of plasma BNP. The LAA flow velocity was obtained by pulsed Doppler TEE. Multivariate logistic regression analysis demonstrated that plasma BNP levels, left atrial volume index (LAVI), LV mass index (LVMI), and the CHADS2 score were independent predictors of lower LAA flow velocity (< 20 cm/s). Plasma BNP levels (r = - 0.58, p < 0.001) were correlated with LAA flow velocity. The area under the curve (AUC) for BNP (AUC 0.803) was larger than that for the CHADS2 score (AUC 0.712), LAVI (AUC 0.664) and LVMI (AUC 0.608) with an optimal BNP cut-off value of 164 pg/ml (sensitivity 75.7%, specificity 71.1%). CONCLUSIONS: This study showed that a higher plasma BNP was associated with a lower LAA flow velocity in patients with NV-AF and normal LV systolic function. The plasma BNP may complement the role of the CHADS2 score in predicting lower LAA flow velocity.

Lectin microarray analysis of isolated polysaccharides from Sasa veitchii.

We report lectin microarray profile of the polysaccharide fraction derived from Sasa veitchii leaf that exhibits anti-influenza activity. This fraction showed higher reactivities with lectins known as binders to oligo-mannose, fucose, or galactose. Our findings along with previously reported monosaccharide components suggest that the polysaccharide can be cross-reactive with cell surface receptors involved in immune system, thereby exerting anti-influenza activity.

[Surveillance of Chromium(VI) Concentrations in Mineral Water Products].

We developed and evaluated a method of quantifying chromium(VI) in mineral water (MW). After a performance evaluation, we performed a surveillance of chromium(VI) concentrations in 150 kinds of MW products purchased from markets in 2016. Recovery rates, as examined by using 150 spiked samples prepared from the purchased MW products, ranged from 93 to 107%. These results strongly suggest that the validated method is suitable for quantifying chromium(VI) in MW. Chromium(VI) was detected in 65 kinds of MW products; the detection rate was 43%. The minimum concentration of chromium(VI) was 0.0001 mg/L and the maximum was 0.0019 mg/L. The median value of chromium(VI) concentration was 0.0003 mg/L. In most of the products in which chromium(VI) was detected, the concentration ranged from 0.0001 to 0.0002 mg/L. No products were found with concentrations higher than the value (0.05 mg/L) set by the Food Sanitation Law as the specification for MW.

In vitro and in vivo antiherpetic effects of (1R,2R)-1-(5'-methylful-3'-yl)propane-1,2,3-triol.

In this study, we demonstrated the in vitro and in vivo antiherpetic activities of a stable furan derivative, (1R,2R)-1-(5'-methylful-3'-yl)propane-1,2,3-triol (MFPT), which had originally been isolated from Streptomyces sp. strain FV60. In the present study, we synthesized MFPT from (5-methylfuran-3-yl)methanol in 6 steps for use in the experiments. MFPT showed potent in vitro antiviral activities against two acyclovir (ACV)-sensitive (KOS and HF) strains and an ACV-resistant (A4-3) strain of herpes simplex virus type 1 (HSV-1) and an ACV-sensitive HSV type 2 (HSV-2) UW 268 strain, their selectivity indices ranging from 310 to 530. By intravaginal application of MFPT to mice, the virus yields decreased dose-dependently against the three strains of HSV-1 and HSV-2. When MFPT was applied at a dose of 1.0 mg/day, the lesion scores, as clinical signs manifested by viral infection, were extensively suppressed in HSV-1-infected mice, whereas the lesion scores in HSV-2-infected mice were not markedly decreased. Interestingly, MFPT exerted an inhibitory effect against ACV-resistant HSV-1 in mice to a similar degree as in ACV-sensitive HSV-1-infected mice. Therefore, the compound might have potential for developing a topical antiviral agent that could be also applied to the infections caused by ACV-resistant viruses.

Membrane topology of Golgi-localized probable S-adenosylmethionine-dependent methyltransferase in tobacco (Nicotiana tabacum) BY-2 cells.

S-adenosylmethionine (SAM)-dependent methyltransferases (MTases) transfer methyl groups to substrates. In this study, a novel putative tobacco SAM-MTase termed Golgi-localized methyl transferase 1 (GLMT1) has been characterized. GLMT1 is comprised of 611 amino acids with short N-terminal region, putative transmembrane region, and C-terminal SAM-MTase domain. Expression of monomeric red fluorescence protein (mRFP)-tagged protein in tobacco BY-2 cell indicated that GLMT1 is a Golgi-localized protein. Analysis of the membrane topology by protease digestion suggested that both C-terminal catalytic region and N-terminal region seem to be located to the cytosolic side of the Golgi apparatus. Therefore, GLMT1 might have a different function than the previously studied SAM-MTases in plants.

Characterization of a novel mutation in NS1 protein of influenza A virus induced by a chemical substance for the attenuation of pathogenicity.

It is generally accepted that live attenuated influenza vaccine (LAIV) has the potential for use as a vaccination against flu. In this study, we demonstrated the nature of an influenza A virus (IAV) mutant induced by treating the IAV with a stable furan derivative, (1R,2R)-1-(5'-methylfur-3'-yl)propane-1,2,3-triol (MFPT), which had been isolated from Streptomyces sp. strain FV60 with the objective of it being an LAIV candidate. The resulting MFPT-resistant (MFPTr) IAVs possessed attenuated pathogenicity in vitro and in vivo when compared with that of the parent virus (H1N1 subtype, NWS strain). Sequencing analysis revealed that a novel mutation, C490U in ns gene (P164S in NS1), was detected in all MFPTr virus clones tested. Therefore, NS1 might be a main target of MFPT, and it was suggested that the P164S mutation contributed to the attenuated pathogenicity of the mutants. Although the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway is one of the targets of NS1, the MFPTr virus suppressed the phosphorylation of Akt when compared with the wild-type (WT) virus. It was suggested that this might lead to the subsequent inhibition of the cleavage of PARP-1 and caspase-3, which is important for the progression of apoptosis. At the same time, nucleoprotein (NP) was found to be retained in the nuclei in MFPTr virus-infected cells while nuclear export of NP was detected in WT virus-infected cells. In addition, the expression levels of interferon-ß transcripts were significantly decreased in MFPTr virus-infected cells. From these results it can be shown that the mutation, NS1P164S, might be one of the key residues to control NS1 function concerning the induction of apoptosis. In conclusion, MFPT induced favorable mutation in the ns gene for the attenuation of IAV, and therefore might provide the novel methodology for preparing LAIVs.

Characterization and biological effects of two polysaccharides isolated from Acanthopanax sciadophylloides.

Two polysaccharides abbreviated ANP and AAP were isolated from the young buds of Acanthopanax sciadophylloides. ANP consisted of L-arabinose, D-mannose, D-glucose and D-galactose in a ratio of ca 1.0:2.6:2.5:1.4 and its weight average molecular weight (Mw) was 1.07×10(4). AAP consisted of L-arabinose, D-galactose and 4-O-methyl-D-glucuronic acid in a ratio of ca 5:10:1, and its Mw was estimated to be 8.40×10(4). ANP was suggested to be an acetylated heteropolysaccharide, whereas AAP was speculated to be a type II arabinogalactan on the basis of structural analysis data. Both polysaccharides were found to stimulate NO production and induce the expression of cytokine mRNAs including IL-1ß, IL-6, IL-10 and TNF-α on RAW264.7 cells. They also induced NF-κB activation in RAW-Blue cells. NO production and NF-κB activation by both polysaccharides were decreased by pretreatment with neutralizing anti-TLR-4 and anti-CD14 antibodies but not with anti-TLR-2, anti-SR-A, anti-CD11c, and anti-Dectin-1 antibodies. Therefore, these immunostimulating effects of ANP and AAP were suggested to be promoted by the interaction through the membrane receptors, TLR-4 and CD14. In addition to immunomodulating effects, ANP showed anti-HSV-2 effects in vitro.

Mekabu fucoidan: structural complexity and defensive effects against avian influenza A viruses.

Fucoidan from the sporophyll (Mekabu) of brown seaweed Undaria pinnatifida (wakame) is interesting due to its various biological activities. Mekabu fucoidan (Mw ∼ 9 kDa) of this study (MF) was previously isolated and characterized by chemical and separation methods including GPC and methylation analysis (Lee, Hayashi, Hashimoto, Nakano, & Hayashi, 2004). It was found that this fucoidan composed of partially sulphated (DS ∼ 0.72) fucose and galactose at approximately equal amounts. Methylation analyses revealed complex structure of MF. However, it has been still unclear about the linkages between units and substitution patterns. To solve these structural tasks, spectroscopic methods (FTIR, FT Raman and NMR) were used in the analysis of native MF and its deesterified derivatives. According to obtained results, this polysaccharide was defined as O-acetylated sulphated fucogalactan. The defensive effects of MF were evaluated on mice infected with avian influenza A viruses (H5N3 and H7N2 subtypes); its efficacy was determined in reducing viral replication and increasing antibody production. Oral administration of MF resulted in suppressing virus yields. In addition, the production of neutralizing antibodies and mucosal IgA in the animals inoculated with the avian influenza A viruses was significantly increased. These results suggested that MF could be used for the prevention of viral infection.