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INTRODUCTION

Allicin, main component of garlic, is known for its antibacterial activity including Streptococcus species.

Streptococcus agalactiae, or group B streptococcus (GBS), is one of the leading causes of neonatal infections.

GBS is a member of normal human oral, gastrointestinal and vaginal flora. 20-30% of pregnant women are colonized with GBS and are at increased risk of transmitting the bacteria to 50% of neonates during delivery.

In UK, GBS reported cases increased from 643 in 1990 to 1248 in 2005 (Figure 1).

In order to decrease the adverse effects of antibioprophylaxis, other alternatives such as topical antiseptics are studied.

Allicin formulated in gel for topical use may account as an option for prophylaxis against perinatal GBS infection.

Figure 1: Incidence of GBS in UK between 1990 and 2005

13% of isolates were resistant to erythromycin, 95% resistant to tetracycline, and 47% resistant to erythromycin when grown on 5% blood ISA (Figure 2).

No significant difference in determination of IZDs of streptomycin, erythromycin, methicillin, and penicillin by using 5% blood ISA, ISA, and THA.

M11 Antibiotic disc-diffusion testing

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Figure 2: Plate A showed a mucoid resistant strain to all antibiotics except chloramphenicol; Plate B showed inhibition zones around all antibiotics except for streptomycin.

MICs and MBCs of allicin The average MIC of allicin was 70 μg/mL ranging between 35 and 95 μg/mL. MIC was determined by agar and microbroth dilution methods.

87.5% of isolates had MBC of 156.12 μg/mL, but 100% of strains were killed at 312.25 μg/mL.

Figure 3: Determination of allicin’s MIC by agar dilution method at allicin concentrations 30, 50, 80, and 100 μg/mL (Right to Left). The figure showed bacterial growth of 3 mucoid GBS isolates at 80 μg/mL and none at 100 μg/mL.

Antibacterial activity of allicin in gel formulations

Allicin was formulated in 4 different gels. Gels B and C showed the largest IZDs at allicin 500 μg/mL, while gel B was the best at allicin concentration 1000 μg/mL.Figure 4: Plate A showed gel B-allicin (1000 μg/mL) vs gel B base,

while Plate B showed antibacterial activity of aqueous extract of allicin (500 μg/mL) vs allicin-gel B at the same concentration.

MALDI-ToF-MS Analysis

Maldi analysis showed that variations existed between isolates having different resistance patterns, mucoid and nonmucoid morphology.0

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Figure 5 : MALDI-ToF-MS results showing different

peaks between mucoid and nonmucoid isolates

AIM

This research project aimed to study the: Bacteriostatic or bactericidal activity of a stable aqueous extract of allicin against GBS isolates via determination of MIC, MBC, and killing kinetics assays.

Time- or concentration-dependent activity of allicin. Activity of a novel allicin-gel formulation against clinical GBS isolates to be suitable for topical prophylaxis against perinatal GBS infections.

Spectra of cell envelope components of the GBS isolates by MALDI-ToF-MS to determine if the clinical isolates were different.

Andrews, J. M. (2007). Journal of Antimicrobial Chemotherapy. doi:10.1093/jac/dkm110.Ankri, S. and Mirelman, D. (1999). Microbes and Infection; 2:125-129. Centers for Disease Control and Prevention. MMWR 2002; 51:1–18.Gibbs, R.S. et al. (2004) Obstet Gynecol; 104:1062-1076.Heath, P.T. and Schuchat, A. (2007) Best Practice & Research Clinical Obstetrics and Gynaecology; 21(3): 411- 424.Witkin, S., et al. (2007) Best Practice and Research Clinical Obstetrics and Gynaecology; 21(3): 347- 354.

REFERENCES

MIC of allicin was higher than other streptococci species; it ranged between 35 and 95 μg/mL.

Killing kinetics showed that 13/16 isolates were killed at 140 μg/mL, 1/16 at 280 μg/mL, and 1/16 at 1120 μg/mL at < 24 h; 1 strain was not killed.

Allicin showed time-dependent bactericidal activity.

87.5% of isolates had MBC of 156.12 μg/mL and 100% were killed at 312.25 μg/mL.

For the first time, a novel gel formulation of allicin was active against GBS isolates at 500 μg/mL, thus it can be used topically for prophylaxis against perinatal GBS infections.

Further research projects including invivo studies of allicin-gel formulations and molecular studies for GBS isolates can be done.

CONCLUSION

Figure 6: Killing Kinetics of allicin against 4GBS isolates.

16 GBS isolates selected on the basis of different antibiotic resistance patterns and allicin’s MIC and MBC values were studied in killing kinetics.

Combined results of killing kinetics showed that allicin had bactericidal activity against GBS isolates.

Results for individual strains showed that: 13 GBS isolates were killed at 140 μg/mL at 24 h (Fig.7A, B).

1 strain was killed at 280 (Fig.7C) and another at 1120 μg/mL.

1 strain was not killed (Fig.7D).

Killing kinetics of allicin against GBS

METHODS and RESULTS

ALLICIN, a Promising Natural Agent in the Prophylaxis of Perinatal Streptococcus agalactiae Infections

Hussein El Haj Ahmad BPharm, AMRSC

School of Health and Biosciences, University of East London

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Bacterial Strains

76 clinical GBS isolates were studied.

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