The increasing microbial resistance to drugs is a great threat that hampers the treatment of many illnesses. The indiscriminate abuse of antimicrobials together with the lack of proper identification of the causative organism and patient demands, have contributed to the rise of resistant strains of microbes to various drugs. Currently more than 70% of bacteria associated with hospital-acquired infections are now resistant to drugs that were once effective against it (Brunton et. l. 2006).
Although new drugs are being created to overcome this problem, most of these new drugs are merely modifications of pre-existing drugs. It is feared that the rate of new drug production against currently resistant microbes might not be fast enough to counter the effects of increasing antimicrobial resistance (Brunton et. al. 2006). Rather than modifying pre-existing drugs, this research hopes to see if mature guava leaves will exhibit antimicrobial activity against Pseudomonas aeruginosa, a gram-negative bacillus, well-known for its increasing resistance to many drugs.
If this research proves successful, it is hoped that future researches will dwell deeper into the chemical properties of the leaves and identify its active antimicrobial component(s). This could potentially lead to a brand new drug which could be used against the increasing resistance of Pseudomonas, and possibly of other organisms. Review of Related Literature Psidium guajava, from the family Myrtaceae, is considered native to Mexico and extends throughout South America, Europe, Africa and Asia It grows in all the tropical and subtropical areas of the world and adapts to different climatic conditions but prefers dry climates.
P. guajava is a small tree with a height of 10m having thin, smooth, patchy, peeling bark. Guava leaves are opposite, short-petiolate, the blade oval with prominent pinnate veins, 5–15 cm long. Flowers are somewhat showy, petals whitish up to 2 cm long and with numerous stamens. Its fruit is yellow, fleshy, globose to ovoid berry about 5 cm in diameter with an edible pink mesocarp containing numerous small hard white seeds. There has been a tremendous interest in this plant as evidenced by the voluminous work (Gutierrez et al. 2008).
Different parts of the plant are used in the indigenous system of medicine for the treatment of various human ailments such as wounds, ulcers, and cholera. Pharmacological investigations indicated that its bark, fruit, and leaves possess antibacterial, hypoglycemic, anti-inflammatory, analgesic, antipyretic, spasmolytic, and CNS depressant activities (Macatol et. al 1998). In the Philippines, the unripe fruit, the leaves, the cortex of the bark and the roots are used for washing ulcers, wounds, astringent, and as treatment for diarrhea (Gutierrez et al 2008).
This study will make use of guava leaf, as previous studies showed that the leaf extracts demonstrated antimicrobial activity. alcoholic extracts of The inhibitory effects of aqueous and P. guajava leaf on the growth of Staphylococcus aureus, Streptococcus mutans, Pseudomonas aeruginosa, Salmonella enteritidis, Bacillus cereus, Proteus spp. , Shigella spp. and Escherichia coli were examined using the in vitro agar well diffusion method. The results revealed that alcoholic extract of guava leaf prevented the growth of 81. % of the test organisms (Chah, et al, 2006).
Flavonoids are hydroxylated phenolic substances that are known to be synthesized by plants in response to microbial infection, which is not surprising that they have been found in vitro to be effective antimicrobial substances against a wide array of microorganisms. Their activity is probably due to their ability to complex with extracellular and soluble proteins and to complex with bacterial cell walls. More lipophilic flavonoids may also disrupt microbial membranes (Cowan 1999).
In mature leaves, the greatest concentrations of flavonoids were found, namely, Myricetin (208. 44 mg kg 1), quercetin (2883. 08 mg kg? 1), luteolin (51. 22 mg kg 1) and kaempferol (97. 25 mg kg? 1) (Gutierrez et al. , 2008). P. aeruginosa is a gram-negative, motile, non-fermentative, pigment- producing, aerobic bacilli. This organism grows at 37-42°C in a wide variety of culture media, forming smooth round colonies with a fluorescent greenish color. non-fluorescent, bluish pigment called pyocyanin. It also produces a Aside from these pigments, it ometimes produces a sweet or grape-like or corn taco- like odor which is helpful in the identification of the organism. (Brooks 2010).
It also has special structures, enzymes, and toxins which makes it highly virulent when introduced into areas devoid of normal defenses. The structures help the organism attach to host cells, avoid the action of phagocytes, and form biofilms, while the enzymes breakdown tissues causing necrosis. On the other hand, toxins produced by P. aeruginosa inhibit protein synthesis by interfering with adenosine diphosphateribosylation of elongation factor-2 (Braunwald 2008).
It is a bacterium responsible for severe nosocomial infections, life-threatening infections in immunocompromised persons, and chronic infections in cystic fibrosis patients. The bacterium’s virulence depends on a large number of cell-associated and extracellular factors. Cell-to-cell signalling systems control the expression and allow a coordinated, cell-density–dependent production of many extracellular virulence factors (Van Delden, 1998). This study will utilize P. aeruginosa as the test organism because of reported cases where the organism is highly resistant to commonly used antibiotics. Todar, 2011)
The reason why it has a low antibiotic susceptibility is because of its efflux pumps with chromosomally encoded antibiotic resistance genes whose net effect is to effectively reduce the amount of drug that enter the bacterium. Since the drug does not reach high enough concentrations within the bacteria it is unable to kill it. Another tactic employed by the bacterium is the low permeability of the cell wall which further reduces the amount of drug that enters the cell. Most of the commonly-used antibiotics, particularly the penicillins and first generation cephalosporins, are not effective against Pseudomonas (Zinsser, 1992).
This study will use Amikacin as the positive control. Amikacin is a semisynthetic aminoglycoside antibiotic produced by acylation of Kanamycin A. It works by binding to the bacterial 30S ribosomal subunit interfering with the correct transcription of the mRNA resulting in the inhibition of bacterial protein biosynthesis. It is very effective against Pseudomonas species. It is recommended as a single dose or divided doses into two or three equal portions of 15 mg/kg/day but the dosage and interval must be altered for patients with renal failure.
Amikacin may be given intramuscularly or intravenously (Brunton, 2006). Amikacin resists degradation by most aminoglycoside inactivating enzymes known to affect gentamicin, tobramycin, and kanamycin (Bauer et al, 2004). Disc Susceptibility Test method, which requires the measurement of the diameters of the zone of inhibition, give the most precise estimates of antibiotic susceptibility. One such procedure has been recommended for use with discs to test susceptibility to Amikacin. Interpretation involves correlation of the diameters obtained in the disc test with MIC values for Amikacin.
When the causative organism is tested by the KirbyBauer method of disc susceptibility, a 30-mcg Amikacin disc should give a zone of 17 mm or greater to indicate susceptibility. Zone sizes of 14 mm or less indicate resistance. “Susceptible” indicates that the infecting organism is likely to respond to therapy while a report of “resistant” indicates that the infecting organism is not likely to respond to therapy. Since guava has been used against many types of ailments including those brought about by bacteria, it would be beneficial to test if its extracts are effective against P. aeruginosa.
Plants have been a source of many drugs since some of its compounds are known to be antimicrobial such as flavonoids. It is hoped that guava may have such compounds or possibly other unidentified compound which can aid against Pseudomonas. P. aeruginosa which utilizes many tactics such as efflux pump and a membrane that greatly limits entry of drugs is making it highly resistant to many drugs. If left untreated, it can be lethal. Significance of the Study This research will be conducted hoping that it will benefit the following:
- The general public, that they may be able to expand their knowledge of the eneficial effects of natural products.
- The Department of Health, Department of Science and Technology and other concerned agencies, that they do further studies on guava and determine the exact active component(s) to help in developing a new drug.
- Future researchers that they may use this study as basis for similar studies to be conducted that would involve determining the medicinal benefits of guava leaves.
