|Year : 2022 | Volume
| Issue : 3 | Page : 219-229
Effect of immunoglobulin Y formulations on oral microorganisms in human subjects - A systematic review and meta-analysis of clinical trials
Ashwin M Jawdekar, Vatsala Srivastava, Urvashi Tank, Laresh Naresh Mistry
Department of Pediatric and Preventive Dentistry, Bharati Vidyapeeth Deemed to be University Dental College and Hospital, Navi Mumbai, Maharashtra, India
|Date of Submission||06-May-2022|
|Date of Decision||22-Jun-2022|
|Date of Acceptance||24-Jun-2022|
|Date of Web Publication||18-Oct-2022|
Ashwin M Jawdekar
Department of Pediatric and Preventive Dentistry, Bharati Vidyapeeth Deemed to be University Dental College and Hospital, Sector 7, CBD Belapur, Navi Mumbai - 400 614, Maharashtra
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Background: Passive immunization using egg yolk-based antibodies has been tested against oral microorganisms. Our study assessed the effect of immunoglobulin Y (IgY) formulations on Streptococcus mutans, Porphyromonas gingivalis, and Candida albicans in human subjects. Highlights: VS and UT independently searched articles using keyword combinations in four search engines; studies in English were selected. Either parallel-arm or split-mouth randomized controlled trials on healthy human subjects were considered. Ten studies remained in the selection; six studies compared the effect of IgY formulations on S. mutans, three on P. gingivalis, and one on C. albicans. Five studies (422 subjects) compared the effect of IgY formulations on S. mutans. When fixed-effect model (FEM) was applied, the risk ratio (RR) (confidence interval [CI]) was found to be 7.81 (6.00, 10.18). Three studies (167 subjects) compared the effect of IgY formulations on P. gingivalis. When FEM was applied, the RR (CI) was found to be 0.06 (−0.03, 0.15) in relation to reduction in probing depth. When FEM was applied, for percentage reduction in bleeding on probing (BOP), the RR (CI) was 1.99 (1.64, 2.41). Only one study (26 subjects) was available of IgY formulation and C. albicans; hence meta-analysis was not performed.The search was extended using Google Scholar, Semantic Scholar, cross-references and by contacting authors and researchers in the field which further yielded five articles. . Conclusions: IgY formulations were effective in the reduction of S. mutans. They were not effective on P. gingivalis in relation to probing depth but were effective in relation to reduction in BOP. No harms were reported. Evidence is of low quality due to high heterogeneity. The ROB was moderate and publication bias was low.
Keywords: Candida albicans, immunoglobulin Y, Porphyromonas gingivalis, passive immunization, Streptococcus mutans
|How to cite this article:|
Jawdekar AM, Srivastava V, Tank U, Mistry LN. Effect of immunoglobulin Y formulations on oral microorganisms in human subjects - A systematic review and meta-analysis of clinical trials. J Indian Soc Pedod Prev Dent 2022;40:219-29
|How to cite this URL:|
Jawdekar AM, Srivastava V, Tank U, Mistry LN. Effect of immunoglobulin Y formulations on oral microorganisms in human subjects - A systematic review and meta-analysis of clinical trials. J Indian Soc Pedod Prev Dent [serial online] 2022 [cited 2023 Feb 4];40:219-29. Available from: http://www.jisppd.com/text.asp?2022/40/3/219/358832
| Introduction|| |
Why it is important to do this review?
Oral microbiome is diverse, and over 700 species of microorganisms have been identified which constitute the microflora. Streptococcus mutans is a known initiator of dental caries, Porphyromonas gingivalis is a known “invasive and evasive” periodontal pathogen, and Candida albicans is the most common opportunistic fungus of the oral cavity.,,
Although antimicrobials such as antibiotics and topical antiseptics have been available, the strategy to “remove or kill all” microorganisms have been debated against the oral microorganisms. The emergent threat of antimicrobial resistance is also a consideration against their use.
Immunological approaches for the management of oral pathogens are not new, while early attempts of developing caries vaccine (active immunization) targeting S. mutans and passive immunization with mucosal antibodies (1990s) had several concerns and limitations. Development of egg yolk-based antibodies has ushered in a new era in the prevention of oral infections.
Specific antimicrobial protection is possible with passive immunological approaches. Immunoglobulin Y (IgY) was first conceptualized in the late 1800s. IgY is functional equivalent of IgG in mammals extracted from egg yolk (hence, the name Ig “Y”) after the hen is injected with a suitable antigen. IgY egg yolk-based antibody against a specific antigen does not activate complement or interact with Fc receptors. Industrial production of IgY formulations is feasible as well as economical. A single egg may contain up to 200 mg of IgY. IgY-based formulations have been used in diagnostic and biomarker discovery as well as in therapeutics. IgY-based formulations have been tested against dental caries (specific antigens of S. mutans such as glucosyltransferase), periodontal disease (antigen: P. gingivalis), oral thrush (antigen: C. albicans), gastric ulcers (antigen: Helicobacter pylori), and rotavirus infections.
Since there have been published several randomized trials over the last few decades reporting the effect of IgY on oral microorganisms, this investigation was planned to address a specific question that has not been previously answered with a systematic review and meta-analysis (MA).
How effective are immunoglobulin Y formulations against oral microorganisms in human subjects?
The study had the following aim:
- To evaluate the effect of IgY formulations on oral microorganisms in human subjects.
The primary objectives were:
- To evaluate the effect of IgY formulations on S. mutans in human subjects
- To evaluate the effect of IgY formulations on P. gingivalis in human subjects
- To evaluate the effect of IgY formulations on C. albicans in human subjects.
The secondary objectives were:
- To assess harms, if any, of IgY-based interventions.
The PICOS was explained as below:
- P - Human subjects
- I - IgY-based topical formulations
- C - Positive/negative control
- O - Change in counts of oral microorganisms or other reported measures
- S - Randomized controlled trials (RCTs) (all types).
| Methods|| |
Criteria for considering studies for this review
Only well-controlled RCTs were identified. Either parallel-arm or split-mouth (only one study: Yokohama et al., 2007) RCTs were considered. A few RCTs had a negative/placebo control. A few RCTs had positive controls such as known antimicrobial formulations. One study used three different formulations of IgY. Following were the considerations for selection.
Types of participants
All studies carried out on healthy human subjects (children as well as adults) were included. Animal and laboratory (in vitro) studies were excluded.
Types of interventions
Interventions included use of different formulations of IgY such as mouthrinse, lozenge, and ointment in the treatment groups. Either placebo or antimicrobials such as chlorhexidine or xylitol-based formulations were used in the control groups.
Types of outcome measures
The independent and dependent variables were topical IgY formulations and changes in microbial counts (either mean + standard deviation) or percentage of subjects showing reduction in microbial counts, respectively. Change in the microbial levels or percentage of subjects showing changed counts of microorganisms was considered as primary outcomes. Change in the probing depth and bleeding on probing (BOP) were reported as outcome measures in the studies in relation to P. gingivalis. No harms were reported in any of the interventions which could be assessed as outcomes.
Search methods for identification of studies
Two researchers, VS and UT, independently searched articles using appropriate keyword combinations (”IgY,” “IgY AND dental caries,” “IgY AND oral,” “IgY AND antibodies,” “IgY AND passive immunization,” “IgY AND S. mutans,” “IgY AND P. gingivalis,” “IgY AND candida,” “IgY AND caries AND human,” “P. gingivalis AND Passive Immunization,” “S. mutans AND Passive Immunization,” “dental AND passive immunization,” etc.) in two search engines; PubMed and Cochrane Library. Studies either reported in English or complete translations available in English were selected. Studies in Mandarin had to be omitted due to unavailability of translated text. The search was conducted till November 2021. The search was further extended using Google Scholar, Semantic Scholar, cross-references and by contacting authors and researchers in the field, which yielded five more articles. Selection of articles was finalized after the agreement was reached between VS and UT. Conflicts were resolved by AJ and LM. The selection of articles is outlined in [Figure 1].
Details such as author, year, study design, study settings, number of subjects with age, results of the interventions and control groups with respect to variables of interest such as type of formulation of IgY and change in the levels of S. mutans, P. gingivalis, C. albicans, etc., were extracted from selected articles. In relation to P. gingivalis, changes in the reported variables such as depth of probing in millimeters and BOP were recorded. Only one study was found of IgY and C. albicans; hence, the same was not considered for MA [Table 1], [Table 2], [Table 3].
Risk of bias assessment
Data segregation and analyses (risk of bias [ROB], publication bias, and MAs) were done by all four authors (VS, UT, AJ, and LM) working together. Assessment of ROB in included studies was completed using the Cochrane Tool. The RCTs were evaluated for the risk in the domains: selection bias (randomization and allocation), performance bias (blinding of participants), detection bias (blinding of outcome assessment), attrition bias (missing data due to loss to follow-up), reporting bias (selective reporting), and other biases, if any. The bias is shown color-coded as red (high), yellow and question mark (unclear), and green (low). Majority of the studies exhibited high ROB in the domain of selection, while most studies exhibited little reporting bias [Figure 2]a and [Figure 2]b.
|Figure 2: Assessment of risk of bias using Cochrane tool. (a) In each domain. (b) In each study|
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Assessment of heterogeneity
Due to methodological variations such as differences in age groups of subjects, types of formulations, variations in control groups, and study design (one study having a split-mouth design), the heterogeneity was high. Thus, random-effects models (REMs) were used in the MAs wherever necessary. In addition, additional analyses to exclude a study from each MA were performed.
| Results|| |
Description of studies
The articles were searched up to November 2021. The detailed search is available at https://docs.google.com/document/d/1IxOC0Np3i-ElnBwET-0kFxFljo7jS3c-/edit?usp=sharingandouid=100613536407 046279863andrtpof=true and sd=true.
After excluding studies, a total of 10 studies remained for final selection. Of these 10 studies, 6 studies compared the effect of IgY formulations on S. mutans, 3 studies compared the effect of IgY formulations on P. gingivalis, and 1 study studied the effect on IgY on C. albicans. The selection summary of articles for systematic review and MA is summarized in a flowchart [Figure 1].
Data extraction was done by AJ and VS and LM and UT for S. mutans and P. gingivalis studies, respectively. After data extraction from available studies, studies were segregated and tabulated as shown in [Table 1], [Table 2], [Table 3] (S. mutans, P. gingivalis, and C. albicans, respectively). [Table 1] shows the effect of IgY formulations on S. mutans wherein two studies compared the effect of mouth rinse on S. mutans, and two studies studied the effect of lozenges on S. mutans. In one study (Nguyen et al., 2011), three formulations of IgY corresponding to three antigen types of S. mutans (c, e, and f) were available for comparison. [Table 2] shows the effect of IgY formulations on P. gingivalis wherein one study each studied effects of ointment, lozenge, and mouth rinse on P. gingivalis, respectively. [Table 3] shows the effect of oral moisturizing gel on C. albicans. After data extraction, we could consider MA for five studies on S. mutans and three studies on P. gingivalis.
Characteristics of participants
The participants for the studies included human subjects of different age groups with both adult and child participants. A total of 10 studies were considered for systematic review with 615 participants: 422 for S. mutans, 167 for P. gingivalis, and 26 for C. albicans.
Characteristics of outcome measures
Variable recorded for S. mutans and C. albicans was change in colony forming unit (CFU) ml; whereas that for P. gingivalis were changes in BOP and probing depth in millimeters.
The quantitative assessment with MA was based on the calculation of risk ratio (RR) and confidence interval (CI) from the available data.
The MA was carried out for comparison of outcomes against S. mutans and P. gingivalis using REM and fixed-effect model (FEM).
Immunoglobulin Y and Streptococcus mutans
Five studies compared the effect of IgY formulations on S. mutans. When FEM was applied, the RR (CI) was found to be 7.81 (6.00, 10.18) indicative of statistically significant difference in percentage reduction of CFU/ml (P = 0.0007) in favor of IgY formulations [Figure 3]a. However, due to high heterogeneity (I2 = 72%), when REM was applied, the RR (CI) was 7.20 (4.18, 12.38) indicative of statistically significant difference (P = 0.0007) in favor of IgY formulations [Figure 3]b.
|Figure 3: Meta-analysis forest plot - IgY formulations versus control (S. mutans)|
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Immunoglobulin Y and Porphyromonas gingivalis
Three studies compared the effect of IgY formulations on P. gingivalis. When FEM was applied, the RR (CI) was found to be 0.06 (−0.03, 0.15) indicative of statistically nonsignificant difference (P = 0.48) in relation to reduction in probing depth [Figure 4]. Heterogeneity was found to be low (I2 = 0). When FEM was applied, for percentage reduction in BOP, the RR (CI) was 1.99 (1.64, 2.41) suggestive of statistically significant difference (P = 0.0002) [Figure 5]a. However, due to high heterogeneity (I2 = 88%), when REM was applied, RR (CI) was 2.34 (1.16, 4.72) indicative of statistically significant difference (P = 0.0002) with the same outcome [Figure 5]b.
|Figure 4: Meta-analysis forest plot - IgY formulations versus control (P. gingivalis - probing depth) - FEM. FEM = Fixed-effect model, IgY = Immunoglobulin Y, P. gingivalis = Porphyromonas gingivalis, CI = Confidence interval, SD = Standard deviation|
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|Figure 5: Meta-analysis forest plot - IgY formulations versus control (P. gingivalis - bleeding on probing)|
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Immunoglobulin Y and Candida albicans
Only one study was available. MA could not be performed.
Additional analysis: Immunoglobulin Y-Streptococcus mutans
Additional analysis was carried out with the exclusion of a study (Hatta et al., 1997) that used 0.12% chlorhexidine as a positive control. Four studies compared the effect of IgY formulations on S. mutans. When FEM was applied, the RR (CI) was found to be 7.89 (5.92, 10.51) indicative of statistically significant difference (P < 0.00001) in percentage reduction of CFU/ml [Figure 6]a. However, due to high heterogeneity (I2 = 76%), when REM was applied, with RR (CI) was found to be 7.23 (3.79, 13.80) indicative of statistically significant difference (P < 0.00001) with the same outcome [Figure 6]b.
|Figure 6: Meta-analysis forest plot - IgY formulations versus control after omitting Hatta et al. 1997 study (S. mutans)|
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Additional analysis - Porphyromonas gingivalis
Additional analysis was carried out with the exclusion of a study (Yokoyama et al., 2007) that used a split-mouth design. Two studies compared the effect of IgY formulations on P. gingivalis (probing depth). When FEM was applied, the RR (CI) was 0.05 (−0.04, 0.14) indicative of statistically nonsignificant difference (P = 0.26) in percentage reduction in probing depth [Figure 7].
|Figure 7: Meta-analysis forest plot - IgY formulations versus control (P. gingivalis - probing depth) after omitting Yokoyama et al., 2007 study - FEM. FEM = Fixed-effect model, IgY = Immunoglobulin Y, P. gingivalis = Porphyromonas gingivalis, CI = Confidence interval|
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When FEM was applied for percentage reduction in BOP, the RR (CI) was 1.75 (1.42, 2.15) suggestive of statistically significant difference (P < 0.00001). However, due to high heterogeneity (I2 = 89%), when REM was applied, RR (CI) was 2.12 (0.81, 5.58) indicative of statistically nonsignificant difference (P = 0.13) with the same outcome [Figure 8]a and [Figure 8]b.
|Figure 8: Meta-analysis forest plot-IgY Formulations versus Control (P. gingivalis - bleeding on probing) after omitting Yokoyama et al., 2007 study|
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No subgroup or sensitivity analysis was carried out.
Assessment of reporting bias
The reporting bias was assessed for all studies based on standard error (Log [RR]). The dispersion around the central line was fairly equal with a very little skew on the right side indicative of low publication bias [Figure 9]a and [Figure 9]b.
|Figure 9: Publication bias across studies: (a) IgY formulations Vs S. mutans. (b) IgY formulations Vs P. gingivalis|
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| Discussion|| |
Dental caries and periodontal disease are a result of changed ecology as emerges from the current understanding. Although dental caries is multifactorial, the role of mutans streptococci is still the most important one from preventive perspective. Similarly, P. gingivalis is the most important bacterium for therapeutic interventions in the periodontal disease. Assessment of BOP is practiced commonly to measure gingival inflammation, and the clinical decision-making often depends on a simple clinical technique, i.e., probing depth measurement., Both active and passive immunization attempts have been made to target the specific virulence factors of S. mutans and P. gingivalis, with the recent focus being on passive immunization. In contrast with a nonspecific germ-killing approach of antimicrobials, the use of IgY-passive immunization has attracted attention for diseases of mucosal pathogens, particularly of alimentary track. [25,26]
The paucity of Systematic Review and Meta-Analyses (SRMA) on the effectiveness of IgY formulations prompted this investigation. We attempted to review the studies systematically of IgY-based interventions on oral microorganisms and associated short-term clinical outcomes such as BOP or probing depth. Among the most common causative microorganisms in oral diseases, S. mutans, P. gingivalis, and C. albicans were considered; however, only studies on S. mutans and P. gingivalis could be included in the MA.
The MA results indicate that IgY-based formulations were effective in the reduction of S. mutans compared to control interventions in all studies and the summary statistic was statistically significant. IgY-based formulations were effective in relation to P. gingivalis in terms of reduction in BOP but not in terms of probing depth. None of the studies reported any harms of the intervention.
The studies included had high clinical and statistical heterogeneity that can be attributed to the differences in study designs (types of RCTs - parallel arm, split mouth, placebo/negative/positive control), age groups (adults and children), population characteristics (oral hygiene habits, diets, fluoride exposure, etc.), formulations (mouth washes, lozenges, gels, etc.), statistical parameters, etc., Based on statistical heterogeneity, REM was used in MA for most forest plots.
It is important to note that oral diseases are multifactorial. Changes in the levels of S. mutans, P. gingivalis or probing depth and BOP are not actual measures of disease levels. Therefore, the effectiveness of IgY formulations must be evaluated prospectively using long-term trials measuring changes in the disease status (caries, periodontal disease, or thrush).
Findings from our MA, with limited numbers of studies, have poor representation from different parts of the world and therefore have limited generalizability. However, this being the first of its kind assessment has important implications, particularly for further research. Research is needed on the indications and safety aspects of these agents in children with higher risk of new caries and for the prevention of transmission from the caregiver. More research is needed on IgY formulations against C. albicans.
| Conclusions|| |
In our SRMA, IgY formulations were effective in reduction of S. mutans compared to control intervention: RR (CI) 7.58 (4.11, 13.97). IgY formulations were not as effective on P. gingivalis: RR (CI): 0.06 (−0.03, 0.15) in relation to reduction in probing depth; RR (CI): 1.99 (1.64, 2.41) in relation to reduction in BOP. Only one study on C. albicans reported reduction in the counts; however, no MA could be done. No harms were reported of any IgY formulations. Evidence is of low quality due to high heterogeneity. The ROB was moderate and publication bias was low. Thus, we can conclude that IgY formulations are potentially effective against oral microorganisms in human subjects.
This review being the first and only of its kind lays a foundation for future research. IgY formulations can be considered by clinicians for caries prevention targeting S. mutans. More research is needed to establish the benefits, however.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7], [Figure 8], [Figure 9]
[Table 1], [Table 2], [Table 3]