Beneficial Effects of Green Tea Catechin on Veterinary Sciences and Bacterial Infections Mini Review Approaches in Poultry, Dairy & Veterinary Sciences

The anti-bacterial effects of green tea catechins (GTCs) have been extensively studied. (-)-Epigallocatechin-3-O-gallate (EGCG), the most abundant in GTCs, has shown the highest anti-microbial effect among GTCs. This review focuses on anti-bacterial effects and veterinary use of GTCs and EGCG. EGCG shows inhibition of bacterial growth and bio-film formation, and bactericidal effect, resulting from stress response and damage of cell membrane. In veterinary use, feed addition of EGCG and TE brought health improvement to livestock.


Anti-Bacterial Effects
Anti-bacterial effects of tea and GTCs have been demonstrated against Gram-positive and Gram-negative bacteria including Staphylococcus aureus, Vibrio cholerae, Escherichia coli, Shigella spp., Salmonella spp., Bacillus spp., Klebsiella spp. and Pseudomonas aeruginosa [8]. Tea extracts were also bactericidal to staphylococci and Yersinia enterocolitica [9]. EGCG inhibits the growth of S. aureus and Escherichia coli [10]. EGCG at the concentrations between 78 and 625μg/mL inhibited multi-drug resistant Acinetobacter baumannii [11]. Various bacteria isolated from canine oral cavity were sensitive to the polyphenolic compounds mix and EGCG [12]. Minimum inhibitory concentration (MIC) ranges were 0.1-0.8mg/mL for poly-phenolic compounds and 0.0125-0.1mg/mL for EGCG. It is well-known that green tea is effective for caries control. Streptococcus mutans, which is a causative agent of caries, produces insoluble glucan and organic acids in the process of biofilm (plaque) formation. The growth of S. mutans was inhibited by EGCG, ECG, and EGC (the order of their inhibitory effect: EGCG>ECG>EGC), while EC did not show inhibitory function against S. mutans. The MIC and minimum bactericidal concentration (MBC) values of EGCG against S. mutans were 0.125 and 0.1mg/mL, respectively. Tea and tea ingredients showed not only anti-bacterial but also antifungal and antiviral properties.

Anti-Biofilm Effects
Sudano et al. [13] demonstrated that EGCG brought a decrease of slime production and inhibition of biofilm formation by ocular S. aureus and S. epidermidis isolates [13]. Moreover, the colony forming unit (CFU) of S. mutans obtained from biofilm formed in the presence of EGCG were less than that formed in the control culture. These indicate that EGCG had an inhibitory effect on biofilm formation [12]. It has been reported that in addition to binding to lipid layers and peptidoglycan, EGCG interferes with extracellular polymeric material so-called glycocalyx. GTCs significantly inhibit mixed biofilm formation, and reduce the synthesis of auto inducer 2 which is signaling molecule used in quorum sensing [14]. EGCG can eliminate the bio-film matrix by inducing the σE cell envelope stress response and thereby reducing the expression of CsgD [15].

Interaction of Bacterial Cell Components
Initial experiments suggested that negatively charged EGCG exerts its anti-bactericidal activity by binding to the positively charged lipids of the bacterial cell membrane, causing damage to the lipid layer. Subsequently, the interaction of catechins including EGCG with lipid bilayers has been studied in more detail [16,17]. Results from Zhao et al. [10] indicated that EGCG binds directly or indirectly to the peptidogly can of the bacterial cell wall and inhibits the penicillinase activity, protecting penicillin from inactivation [10].

In-vivo model
Steinmann et al. reviewed that the properties of EGCG in various trials against infectious diseases such as methicillinresistant Staphylococcus aureus (MRSA), Stenotrophomonas maltophila, Helicobacter pylori, E. coli and others [5]. We observed that gnotobiotic mice with GTCs had significantly lower Shiga toxin levels than the untreated control group after Enterohemorrhagic E. coli (EHEC) infection [18]. This report also includes the inhibition of bacterial growth in vivo. The untreated controls developed neurological and systemic symptoms, usually culminating in death, whereas none of the mice received green tea extracts exhibited any clinical symptoms or died. The combination of green tea extract and levofloxacin increased survival rates and reduced damage to target organs in orally EHEC infected gnotobiotic mice [19]. Thus, GTCs and EGCG has significant direct and indirect anti-pathogenic effects against food borne bacteria and other bacteria, including multidrug-resistant strains.

Enhancement of Biological Activity by Chemical Modification
The functional and structural differences between these catechins are attributed to the number of hydroxyl groups on the B-ring and the presence or absence of a galloyl moiety. Some chemical modifications have been performed to enhance the biological activity of catechins. Galloyl modification is one of the method to enhance catechins [20]. Galloyl moiety is important to the activity of catechin, it is known that catechins without galloyl moiety have low anti-microbial activity [21]. It is expected that inducing galloyl moiety to original catechins could enhance antimicrobial activity. We demonstrated anti-microbial activity of (-)-epicatechin 3,5-O-digallate (EC35G) [22], galloyl modified catechin from EC. This catechin is made from EC by inducing two galloyl moieties to position 3,5. EC35G inhibited the growth of MRSA, but EC did not.

Use of Tea Extract (TE) for Food Products
The safety and quality of food products are important. The use of antimicrobial film is one of tools for contaminated pathogenic 3/4

Appro Poult Dairy & Vet Sci Approaches in Poultry, Dairy & Veterinary Sciences
bacteria. For example, incorporation of TE into chitosan-coated films enhanced their effectiveness against Listeiria monocytogenes [23]. Packaging film using TEs could improve the safety of ham steak during room/refrigerated storage. Edible polymer coting with TE was had a significant effect in reducing the fat oxidation of chicken nuggets [24]. Wagh et al. [25] described that plantderived extracts such as TE can be valuable to the modification of frankfurter formulations for improve oxidative stability [25]. Actually, TE showed strong antioxidant activity [26]. Thus, TE could be useful for the safety from bacterial contamination and quality for inhibition of fat oxidation for meat industry and market.

Application of Catechins for Veterinary Science
Much attention was focused on biological activities of catechins in promotion of animals, because human have long applied catechins as dairy drinking. Several are already being tested for medial application. In veterinary science, TE has been used as supplement of feeding for livestock animals (Table 1) [27][28][29][30][31][32][33]. Hara et al. [27] demonstrated that feeding the diet supplemented with 0.2% of tea polyphenols to pigs for 2 weeks, the odor of the feces was reduced [27]. During tea polyphenols administration, the level of lactobacilli was increased whereas bacteroidaceae and clostridia were decreased. Moreover, fecal phenol, p-cresol, and skatole were reduced.
Supplementing broiler chicken diets with TE showed antimicrobial effects including anti-coccidal [34] and anti-influenza activity [29]. As long -term effects in chicken, tea polyphenols improved lipid metabolism and digestive enzymatic activities, and affected inflammatory status [35]. ECG supplementation significantly down-regulated the expression of fatty acid synthesis and β-oxidation/lipolysis genes [30]. They suggested that EGCG could alleviate fat deposition via the improved metabolism. Tea poly-phenols reduced glucocorticoid-induced growth inhibition and oxidative stress in chicken [36]. Tea poly-phenol was also effective to laying hens in the view point of anti-oxidant status, performance of reproduction and egg quality [37].
Mastitis in daily cows is a disease of economic considerable importance [38]. S. aureus is the most common cases of mastitis in dairy cows. In Belgium, MRSA isolates obtained from bovine mastitis cases was first reported in the 1970 [39]. EGCG shows antibacterial effects to not only S. aureus but also MRSA. This suggests that EGCG is useful for treating mastitis caused by MRSA.

Conclusion
GTCs show anti-bacterial effect to Gram-positive and Gramnegative bacteria. This indicates that EGCG is useful for prevention and treatment of infectious diseases in domestic animals. Together with the veterinary use described above, GTCs will be an excellent medicine that brings various efficacies to livestock.