Recombinant Shiga toxin B subunit can induce neutralizing immunoglobulin Y antibody.

Previously, we have shown that chickens immunized with Shiga toxin (Stx) produce Stx-neutralizing egg yolk immunoglobulin Y (IgY) antibody. The anti-Stx-1 IgY and anti-Stx-2 IgY exert their neutralizing activity through their antibody activity against the B subunit of the toxin but not the A subunit. In the present study, chickens were immunized with recombinant Stx-1 B subunit (rStx-1B) and recombinant Stx-2 B subunit (rStx-2B). Induced anti-rStx-1B and anti-rStx-2B IgY neutralized the toxicity of Stx-1 and Stx-2 against HeLa 229 cells. The neutralizing activity of anti-rStx-1B IgY on Stx-1 was almost 10 times stronger than that of anti-Stx-1 IgY, and that of anti-rStx-2B IgY was 2.6 times stronger than that of anti-Stx-2 IgY. Anti-rStx-1B and anti-rStx-2B IgY reacted with multimeric and monomeric forms of the B subunits in contrast to anti-Stx-1 and anti-Stx-2 IgY that reacted with only the multimeric form. These results indicated that recombinant B subunits were promising antigens for induction of neutralizing antibodies in chickens.

Evaluation of lozenges containing egg yolk antibody against Porphyromonas gingivalis gingipains as an adjunct to conventional non-surgical therapy in periodontitis patients: A randomized controlled clinical trial.

BACKGROUND: In a previous pilot study, one-time application of anti-Porphyromonas gingivalis gingipain egg yolk immunoglobulin (IgY) into scaling and root planing (SRP)-treated periodontal pockets showed profound improvement of clinical and bacteriological parameters in patients with chronic periodontitis. The present study aims to evaluate the efficacy of daily use of lozenges fortified with the antibody as an adjunct to non-surgical therapy in patients with periodontitis. METHODS: Sixty-four patients with periodontitis were divided randomly into a test and a placebo group. The groups were treated by SRP followed by a daily use of lozenges containing either specific IgY against P. gingivalis gingipains (test) or a sham-immune IgY (placebo). Gingival bleeding index (GBI), probing pocket depth (PD) and quantitation of P. gingivalis in the gingival pockets were assessed at baseline and 8 weeks after the initiation of treatment and compared by using Wilcoxon signed rank test, Mann-Whitney U-test or t test. RESULTS: Both groups showed significant improvement of all parameters at 8 weeks post treatment (P < 0.001). There was a significant difference in reduction of GBI (P < 0.001) and P. gingivalis cell counts (P < 0.05) in the test group compared with the placebo group. The reduction of PD was greater in the test group compared with the placebo group although there was no statistically significant difference between the two groups. CONCLUSIONS: The adjunctive use of lozenges containing IgY antibody against gingipains from P. gingivalis resulted in clinical and microbiological benefits in the treatment for chronic periodontitis. Additional investigations are needed to examine if the IgY brings benefits to case patients who do not receive SRP.

Effects of egg yolk antibody against Porphyromonas gingivalis gingipains in periodontitis patients.

Porphyromonas gingivalis gingipains is suspected to be one of the most important causative agents of periodontitis. We postulated that the inhibition of gingipains may reduce the pathogenic nature of P. gingivalis. Anti-P. gingivalis egg yolk antibody (IgY-GP) was isolated from the yolks of hens immunized with purified gingipains. We applied IgY-GP gel subgingivally in periodontitis patients who harbored P. gingivalis in their subgingival flora. Five pairs of contralateral anterior single-rooted teeth were selected. One tooth in each contralateral pair was randomly treated with IgY-GP and subgingival scaling and root planing, whereas the other tooth was treated with SRP alone. The number of P. gingivalis bacteria was assessed by real-time PCR. Bacterial levels were expressed as the percentage of total bacteria. The IgY-GP group had a significant reduction in probing depth. BOP significantly decreased in the IgY-GP group compared to the control group at week 4. The levels of P. gingivalis significantly increased in the control group at week 4, whereas the reduction in the levels of P. gingivalis was sustained in the IgY-GP group. Within the limitations of the present study, IgY-GP was shown to be an effective immunotherapeutic agent in the treatment of periodontitis.

Chicken Egg Yolk Antibodies (IgY) for Prophylaxis and Treatment of Rotavirus Diarrhea in Human and Animal Neonates: A Concise Review.

The rotavirus-induced diarrhea of human and animal neonates is a major public health concern worldwide. Until recently, no effective therapy is available to specifically inactivate the rotavirion particles within the gut. Passive immunotherapy by oral administration of chicken egg yolk antibody (IgY) has emerged of late as a fresh alternative strategy to control infectious diseases of the alimentary tract and has been applied in the treatment of diarrhea due to rotavirus infection. The purpose of this concise review is to evaluate evidence on the properties and performance of anti-rotavirus immunoglobulin Y (IgY) for prevention and treatment of rotavirus diarrhea in human and animal neonates. A survey of relevant anti-rotavirus IgY basic studies and clinical trials among neonatal animals (since 1994-2015) and humans (since 1982-2015) have been reviewed and briefly summarized. Our analysis of a number of rotavirus investigations involving animal and human clinical trials revealed that anti-rotavirus IgY significantly reduced the severity of clinical manifestation of diarrhea among IgY-treated subjects relative to a corresponding control or placebo group. The accumulated information as a whole depicts oral IgY to be a safe and efficacious option for treatment of rotavirus diarrhea in neonates. There is however a clear need for more randomized, placebo controlled and double-blind trials with bigger sample size to further solidify and confirm claims of efficacy and safety in controlling diarrhea caused by rotavirus infection especially among human infants with health issues such as low birth weights or compromised immunity in whom it is most needed.

Passive protection of dogs against clinical disease due to Canine parvovirus-2 by specific antibody from chicken egg yolk.

The protective effect of immunoglobulins derived from chicken egg yolk (IgY) against infection by Canine parvovirus 2 (CPV-2) was evaluated in 10 beagle dogs orally challenged with a strain of the virus. The 2-mo-old dogs were divided into 3 groups and treated with powders containing CPV-2 IgY or normal egg yolk for 7 d after the challenge. The 4 dogs receiving normal egg yolk (control group) demonstrated mild symptoms typical of CPV-2 infection, such as vomiting, diarrhea, and weight loss. No symptoms were observed by 16 d after challenge in the 3 dogs receiving 2 g of IgY powder. Of the 3 dogs receiving 0.5 g of IgY powder, 2 had clinical CPV-2 disease; however, the manifestations were less severe than in the control group. Furthermore, the IgY-treated groups had significantly greater weight gain and shorter duration of virus shedding than the control group. These results indicate that IgY is useful in protecting dogs from CPV-2-induced clinical disease.

High Prevalence of G12 Human Rotaviruses in Children with Gastroenteritis in Myanmar.

Human rotavirus samples from 54 children with acute gastroenteritis in Myanmar in 2011 were subjected to reverse transcription-PCR to determine their G and P types. On G typing, G2 (24/54; 44.4%) was found to be the most prevalent, followed by G12 (17/54; 31.5%) and G1 (1/54; 1.9%). Mixed cases with G2 and G12 were found in 12 of the 54 (22.2%) samples. On P typing, P[4] was found to be the most predominant (29/54; 53.7%), followed by P[8] (17/54; 31.5%) and P[6] (4/54; 7.4%). Mixed cases with P[4] and P[8] were detected in 4 of 54 (7.4%) samples. Thus, occurrence of G2 and unusual G12 in high proportions was characteristic of human rotaviruses in Myanmar in this study setting.

Whole Genomic Analysis of Human G12P[6] and G12P[8] Rotavirus Strains that Have Emerged in Myanmar.

G12 rotaviruses are emerging rotavirus strains causing severe diarrhea in infants and young children worldwide. However, the whole genomes of only a few G12 strains have been fully sequenced and analyzed. In this study, we sequenced and characterized the complete genomes of six G12 strains (RVA/Human-tc/MMR/A14/2011/G12P[8], RVA/Human-tc/MMR/A23/2011/G12P[6], RVA/Human-tc/MMR/A25/2011/G12P[8], RVA/Human-tc/MMR/P02/2011/G12P[8], RVA/Human-tc/MMR/P39/2011/G12P[8], and RVA/Human-tc/MMR/P43/2011/G12P[8]) detected in six stool samples from children with acute gastroenteritis in Myanmar. On whole genomic analysis, all six Myanmarese G12 strains were found to have a Wa-like genetic backbone: G12-P[8]-I1-R1-C1-M1-A1-N1-T1-E1-H1 for strains A14, A25, P02, P39, and P43, and G12-P[6]-I1-R1-C1-M1-A1-N1-T1-E1-H1 for strain A23. Phylogenetic analysis showed that most genes of the six strains examined in this study were genetically related to globally circulating human G1, G3, G9, and G12 strains. Of note is that the NSP4 gene of strain A23 exhibited the closest relationship with the cognate genes of human-like bovine strains as well as human strains, suggesting the occurrence of reassortment between human and bovine strains. Furthermore, strains A14, A25, P02, P39, and P43 were very closely related to one another in all the 11 gene segments, indicating derivation of the five strains from a common origin. On the other hand, strain A23 consistently formed distinct clusters as to all the 11 gene segments, indicating a distinct origin of strain A23 from that of strains A14, A25, P02, P39, and P43. To our knowledge, this is the first report on whole genome-based characterization of G12 strains that have emerged in Myanmar. Our observations will provide important insights into the evolutionary dynamics of spreading G12 rotaviruses in Asia.

Oral passive IgY-based immunotherapeutics: a novel solution for prevention and treatment of alimentary tract diseases.

This commentary summarizes the laboratory investigations and clinical trials published recently involving per-oral application of IgY supplemented food for specific orogastrointestinal disease prevention and control purposes. The prolonged use and misuse of conventional antibacterial drugs has spawned antibiotic resistant microbes prompting scientists to search for other germ-killing options. In particular, the use of IgY as a novel mode of immunotherapy using oral chicken immunoglobulin (IgY) to confer passive immunity has gained much interest as an inexpensive non-antibiotic alternative for the prophylaxis and treatment of a wide variety of infectious diseases. The stability of IgY in the orogastrointestinal tract and its safety profile has been well-documented. IgY has been used in the treatment or prevention of dental caries, periodontitis and gingivitis, gastritis and gastric ulcer, oral thrush and infant rotavirus diarrhea. The recent clinical trials on IgY with encouraging results has catapulted into the market novel nutraceutical or health supplements for therapeutic or prophylactic intervention based on the consumption of mono-specific or mixed IgY formulations. With recent trends in consumer preference for natural materials to alleviate health concerns, the increasing healthcare costs and the recent advances in drug delivery systems, IgY is likely to shift from its mainly functional food status toward pharmaceuticalization in the foreseeable future.

Anti-obesity activity of hen egg anti-lipase immunoglobulin yolk, a novel pancreatic lipase inhibitor.

BACKGROUND: There is completely no report about both hen egg anti-lipase immunoglobulin yolk (IgY) and its anti-obesity action. Thus, we tried to isolate and characterize a novel anti-lipase immunoglobulin from hen egg yolk. Moreover, we investigated whether hen egg yolk anti-lipase IgY inhibits pancreatic lipase activity in vitro, and examined its ability to prevent obesity in a murine high fat diet-induced obesity model. METHODS: We determined the inhibitory action of Anti-lipase IgY on lipase activity in vitro. We also focused our evaluation on the anti-obesity properties of Anti-lipase IgY in a murine high fat diet-induced obesity model. RESULTS: Anti-lipase IgY blocked porcine lipase activity with an IC50 of 0.49 µM. Supplementing the high fat diet with only 0.2% (w/w) of Anti-lipase IgY for 35 days significantly decreased the weights of intraperitoneal adipose tissues, epididymal, mesenteric, retroperitoneal and perirenal adipose tissues, and the amounts of hepatic total lipid, triglyceride, and cholesterol. This was accompanied by a significant increase in the fecal excretion of triglyceride in the absence of diarrhea. Furthermore, Anti-lipase IgY treatment restored body weight gain to levels similar to mice fed with Control IgY. CONCLUSIONS: This study provides the first report of the development of anti-lipase IgY and the direct evidence that inhibition of pancreatic lipase using Anti-lipase IgY is an effective anti-obesity treatment due to the associated increase in fecal excretion of triglyceride.

Randomized placebo-controlled clinical trial of immunoglobulin Y as adjunct to standard supportive therapy for rotavirus-associated diarrhea among pediatric patients.

This study aims to evaluate the effect of hyperimmune immunoglobulin Y (IgY) against human rotavirus (HRV) among pediatric patients receiving standard supportive treatment for rotavirus-associated diarrhea mostly with an enteric non-cholera co-pathogen in a hospital setting. Two natural HRV reassortant clinical strains ATCC VR 2273 and ATCC VR 2274 were used as mixed immunizing antigens in poultry hens to generate anti-HRV IgY (Rotamix IgY). The Rotamix IgY was used in laboratory and clinical studies against control or placebo IgY. The control or placebo IgY was prepared using tissue culture medium from mock-infected MA104 cell line as antigen for poultry immunization. In vitro, Rotamix IgY exhibited multi-serotypic cross neutralization activities along with synergistic effects against major global serotypes G1, G2, G3, G4 and other human or animal rotavirus strains when compared with mono-specific IgY. Suckling mice (ICR strain) pre-treated orally once with Rotamix IgY and then challenged with rotavirus 3h later showed a significant dose-dependent reduction in frequency (p<0.05) and duration (p<0.05) of diarrhea compared to placebo IgY-treated mice. Out of 114 children aged between 3 and 14 months and with diarrhea upon admission in a Myanmar hospital, 54 dehydrated and rotavirus-positive children were randomized into Rotamix IgY group and placebo IgY group. Of these, only 52 children had complete data with n=26 children per study group. Ninety-two percent of patients in each of these groups were positive for co-infecting enteric non-cholera pathogen and all patients received standard supportive therapy for diarrhea. The patients were monitored for volume and duration of oral rehydration fluid (ORF) and intravenous fluid (IVF) intake, daily stool frequency and overall duration of diarrhea, and frequency and duration of rotavirus shedding. Compared to placebo IgY group, the Rotamix IgY group had statistically significant reduction in mean ORF intake (p=0.004), mean duration of intravenous fluid administration (p=0.03), mean duration of diarrhea from day of admission (p<0.01) and mean duration of rotavirus clearance from stool from day of admission (p=0.05). Overall, our novel approach using oral Rotamix IgY for rotavirus-infected children mostly with non-cholera enteric pathogen co-infection appears to be a promising, safe and effective adjunct to management of acute diarrhea in pediatric patients.

Specific egg yolk immunoglobulin as a new preventive approach for Shiga-toxin-mediated diseases.

Shiga toxins (Stxs) are involved in the development of severe systemic complications associated with enterohemorrhagic Escherichia coli (EHEC) infection. Various neutralizing agents against Stxs are under investigation for management of EHEC infection. In this study, we immunized chickens with formalin-inactivated Stx-1 or Stx-2, and obtained immunoglobulin Y (IgY) from the egg yolk. Anti-Stx-1 IgY and anti-Stx-2 IgY recognized the corresponding Stx A subunit and polymeric but not monomeric B subunit. Anti-Stx-1 IgY and anti-Stx-2 IgY suppressed the cytotoxicity of Stx-1 and Stx-2 to HeLa 229 cells, without cross-suppressive activity. The suppressive activity of these IgY was abrogated by pre-incubation with the corresponding recombinant B subunit, which suggests that the antibodies directed to the polymeric B subunits were predominantly involved in the suppression. In vivo, the intraperitoneal or intravenous administration of these IgY rescued mice from death caused by intraperitoneal injection of the corresponding toxin at a lethal dose. Moreover, oral administration of anti-Stx-2 IgY reduced the mortality of mice infected intestinally with EHEC O157:H7. Our results therefore suggest that anti-Stx IgY antibodies may be considered as preventive agents for Stx-mediated diseases in EHEC infection.

In vitro and in vivo effectiveness of egg yolk antibody against Candida albicans (anti-CA IgY).

We prepared anti-Candida albicans antibody in chicken egg yolk (anti-CA IgY) and investigated its in vitro and in vivo effectiveness. Anti-CA IgY significantly reduced the adherence capacity of C. albicans to FaDu cells (human pharynx carcinoma cells) in a dose-dependent manner. The protective efficacy of anti-CA IgY was investigated in experimentally induced oral candidiasis in immunosuppressed mice. Oral administration of anti-CA IgY significantly reduced the number of C. albicans and the scores of the tongue lesions. Moreover, anti-CA IgY reduced the colonization of C. albicans in mice organs. These results indicate that anti-CA IgY has a protective effect against the oral candidiasis of experimentally infected mice and reduces the dissemination of C. albicans. Putting together, these results indicate that anti-CA IgY is effective against C. albicans. This effect might be due to the blocking of the binding of C. albicans to the host cells. Therefore, anti-CA IgY might be considered as a prophylactic immunotherapy or possibly an adjunctive antifungal therapy.