Methicillin-resistant Staphylococcus aureus (MRSA) and anti-MRSA activities of extracts of some medicinal plants: A brief review

The increasing emergence of multidrug-resistant infection causing microorganisms has become a significant burden globally. Despite the efforts of pharmaceuticals in producing relatively new antimicrobial drugs, they have resulted in a high rate of mortality, disability and diseases across the world especially in developing countries. Supporting this claim was the report of the Centre for Disease Control and Prevention (CDC) who estimated that over 2 million illnesses and 23,000 deaths per year are attributable to antibiotic resistant pathogens in the United States. They include Methicillin-resistant Staphylococcus aureus (MRSA), Vancomycin-intermediate Staphylococcus aureus (VISA), Vancomycin-resistant Staphylococcus aureus (VRSA), Vancomycin-resistant enterococci (VRE), Extended spectrum beta-lactamases (ESBLs) producing gram-negative bacilli, Multidrug-resistant Streptococcus pneumoniae (MDRSP), Carbapenem-resistant Enterobacteriaceae (CRE) and Multidrug-resistant Acinetobacter baumannii. For MRSA, resistance is as a result of Methicillin-sensitive S. aureus (MSSA) strains that have acquired Staphylococcal Cassette Chromosome mec (SCCmec) which carries mecA gene. The gene encodes the penicillin-binding protein (PBP2a) which confers resistance to all β-lactam antibiotics. Vancomycin was previously the widely preferred drug for the treatment of MRSA infections. It is no longer the case with the emergence of S. aureus strains with reduced vancomycin sensitivity limiting the conventional treatment options for MRSA infections to very scanty expensive drugs. Presently, many researchers have reported the antibacterial activity of many plant extracts on MRSA. Hence, these medicinal plants might be promising candidates for treatment of MRSA infections. This work is a brief review on Methicillin-resistant Staphylococcus aureus (MRSA) and the anti-MRSA activities of extracts of selected medicinal plants.


Introduction
Multidrug-resistant bacteria (MDRB) are microorganisms that are resistant to one or more antimicrobial agents. They are usually resistant to all but one or two commercially available antimicrobial agents. This definition includes microbes that have acquired resistance to at least one agent in three or more antimicrobial categories.  [1][2][3] .
Infectious diseases caused by MDRB are an important burden globally. They have for centuries been among the leading causes of death, disability, growing challenges to health security and human progress, especially in developing countries [4].
Although, many new antibacterial drugs have been produced, bacteria exhibiting resistance to them have increased and is becoming a global concern as we are fast running out of therapeutic options [5,6]. The challenges of antimicrobial resistance are faced in both the health care and community settings, necessitating a broad approach with multiple partners across the continuum of care. For example, 18-33% of MRSA colonized patients subsequently developed MRSA infections. Community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) strains also constitute an increasing proportion of hospital-onset MRSA infections. The Centre for Disease Control and Prevention (CDC) estimated that over 2 million illnesses and 23,000 deaths per year are attributable to antibiotic resistance in the United States [3].
Vancomycin is widely prescribed for the treatment of infections caused by MRSA; but the emergence of VISA and VRSA has been reported by many authors. Really, teicoplanin, daptomycin, linezolid, etc are expensive drugs which are currently prescribed when faced with MRSA with low sensitivity to vancomycin. However, development of resistance to these drugs has been identified worldwide [7][8][9][10][11].
Usage of plants in fighting against illnesses and diseases has deep roots in man's history. Researchers are interested in plant extracts as medicines because there are several reports regarding the antimicrobial activity of their crude extracts which might be better substitutes for conventional antibiotics. Recent published reports opined that medicinal plants with anti-MRSA activity can be considered for treatment of MRSA infections [8,12]. This present work is a brief review on MRSA, VISA, VRSA and some medicinal plants with anti-MRSA activities.

MRSA
Staphylococcus aureus is a Gram-positive coccoid bacterium. The cells are arranged in irregular grape-like appearance and they are usually found as normal flora in humans and animals. It is ubiquitous in the human population and 30-40% of adults are asymptomatic carriers. It is also a major pathogen of human and can cause a range of infections from mild skin infections and food poisoning, to life threatening infections [13][14][15][16][17].
Resistance to methicillin by S. aureus was initially observed in 1961 shortly after the antibacterial agent was introduced clinically and since then, there has been a global epidemic of Methicillin-resistant Staphylococcus aureus (MRSA) in both healthcare and community settings [18][19][20]. MRSA isolates from the UK and Denmark in the early 1960s constituted the very first epidemic MRSA clone soon after methicillin was introduced and it has since emerged as an important pathogen in human medicine [21][22][23]. Although, methicillin is no longer prescribed for patients and has been replaced by isoxazolyl penicillins, particularly flucloxacillin in the UK, the acronym MRSA has stayed [24]. It is characterized by antibiotic resistance to penicillins, cephalosporins, carbapenems and has tendency of developing resistance to quinolones, aminoglycosides, and macrolides [10,25,26].
The origination of MRSA was as a result of Staphylococcal Cassette Chromosome mec (SCCmec) genes acquired by methicillin-susceptible S. aureus (MSSA). The SCCmec harbours the mecA gene which encodes the penicillin-binding protein (PBP2a) that confers resistance to all βlactam antibiotics [10,[27][28][29]. SCCmec also contains the cassette chromosome recombinases (ccr) gene complex. The ccr genes (composed of ccrC or a pair of ccrA and ccrB) encode recombinases mediating integration and excision of SCCmec into or from the chromosome. The ccr genes and surrounding genes form the ccr gene complex. In addition to ccr and mec gene complexes, SCCmec contains a few other genes and various other mobile genetic elements such as: insertion sequences, transposons and plasmids [30,31].
Eleven different types of SCCmec (I-XI) and five allotypes of the ccr gene complexes (ccrAB1, ccrAB2, ccrAB3, ccrAB4 and ccrC) have been reported. Generally, SCCmec types I, II, III, VI and VIII are called hospital-acquired MRSA or (HA-MRSA). Types IV, V and VII as communityacquired (CA-MRSA) while types IX, X and XI as livestock-associated MRSA (LA-MRSA) [31,32]. Expression of methicillin resistance in S. aureus is commonly under regulatory control by mecI or blaI gene. The mecI and blaI repressors are controlled by the mecRI and blaRI transducers [20].
MRSA remains a major public health concern worldwide and a therapeutic challenge as the antibacterial drugs effective for treatment are scanty and costly. The changing epidemiology of MRSA infections, varying resistance to commonly used antibiotics and involvement in hospital and community infections are influencing the use and clinical outcomes of currently available antiinfective agents [33].

Resistance of vancomycin by S. aureus
Vancomycin is an antibacterial agent that inhibits cell wall production by binding with the Dalanyl-D-alanine C terminus of the bacterial cell wall precursors, and subsequently preventing crosslinking by transpeptidation. Vancomycin acts extracellularly and inhibits late-stage peptidoglycan biosynthesis which results in the intracellular accumulation of UDP-linked MurNAc-pentapeptide precursors. The vancomycin complex involves a number of hydrogen bonds between the peptide component of vancomycin and the D-Ala-D-Ala residue. Any process that interferes with vancomycin binding to D-Ala-D-Ala residues in the cell wall will decrease the potency of the drug [13,36].
Vancomycin was widely utilized for the treatment of MRSA infections and has led to the emergence of vancomycin-intermediate and vancomycin-resistant S. aureus (VISA and VRSA) [37]. This also triggered off alarms in the medical community as S. aureus causes life-threatening infections in hospitalized and non-hospitalized patients [38].  [34][35][36].
VISA isolates emerged as a result of mutations (not their acquisition of foreign genetic elements) in MRSA isolates during treatment of patients with vancomycin. The comparison of vancomycinsusceptible and -resistant isolates to the VISA isolates showed that the mutations often occurred in the walkR, vraSR, rpoB (ribosomal) genes and the yvqF/vraSR system. Usually, the relevant mutated genes seemed to be directly or indirectly involved with the biosynthesis/metabolism of the staphylococcal cell wall [41].
Often, there were treatment failures when VISA infections were treated with vancomycin [41]. It was observed that under vancomycin selective pressure usually during treatment, the VISA strains with a vancomycin MIC of 8 µg/ml have emerged and led to therapy failure. However, the nature of this resistance phenotype (VISA) was unstable especially when vancomycin selective pressure is removed as some strains reverted back to vancomycin-susceptible strains with MIC at 2 µg/ml [36].

Heterogeneous VISA (hVISA)
In 1997, the first case of hVISA was reported in Japan. The cultures of hVISA strains contain both low-frequency subpopulations of bacteria with increased vancomycin MIC value and high frequency of bacteria with low vancomycin MIC values (close to those of susceptible strains) [41]. The MIC for hVISA strains was defined by the presence of subpopulations of VISA at a rate of one organism per 105 to 106 organisms [42,43]. The hVISA strains were detected using vancomycin population analysis profile (PAP) which was proposed as the most accurate method for hVISA detection; however, it is relatively time-consuming and requires the use of a spiral plater. The hVISA strain has generally required formal population analysis using the serial passage of screened isolates of S. aureus on selective agar containing increasing concentrations of vancomycin for its detection [13]. Results are generally not ready until at least 3 to 5 days [36].
VISA and hVISA strains have thickened cell wall with reduced glycopeptide cross-linking as a result of the complex reorganization of cell wall metabolism. It has been proposed that the thickened cell wall may trap and sequester vancomycin and consequently, interferes with its mode of action [13]. This could be due to alteration in peptidoglycan production leading to increased residues of D alanyl-Dalanine, which bind vancomycin molecules and prevent them from reaching the target sites [18][19][20].

Vancomycin-resistant S. aureus (VRSA)
In 2002, the first hospital strain of Vancomycin-resistant S. aureus (VRSA) was reported in the United States [44]. The acquisition of vanA gene from vancomycin-resistant enterococci resulted in the emergence of vancomycin-resistant strains of S. aureus (VRSA) with vancomycin MIC value greater than 16 µg/ml [36,41,45].

Prevalence of MRSA and S. aureus with reduced sensitivity to vancomycin
MRSA has spread worldwide, and its prevalence has increased in both health-care and community environments. The proportion of MRSA varied among countries such as for instance: 0.4% in Sweden [24]; 25% in western part to 50% in southern India [10]; 33%-43% in Nigeria [46]; 37-56% in Greece, Portugal and Romania in 2014 [47]. High prevalence of MRSA with rates greater than 50% has also been reported in hospitals worldwide including in Asia, Malta, North and South America [29,48]. Variation in the prevalence rates of MRSA was due to different epidemiological factors such as geographical and health system capability in running infection control program [49].
Akanbi and Mbe [50] reported a prevalence range of 0% to 6% VRSA in southern parts of Nigeria among clinical isolates and also 57.7% in Zaria, northern Nigeria. Goud, et al., [51] reported a vancomycin resistance in 1.4% of S. aureus isolates in southern India. Other countries such as: Australia, Korea, Hong Kong, Scotland, Israel, Thailand, South Africa, etc have also reported S. aureus with vancomycin sensitivity reduction with prevalence ranges from 0-74% [20,36,52].

Therapeutic measures
Currently, there are seven common antibiotics used against MRSA, which are: vancomycin, daptomycin, linezolid, Sulfamethoxazole and trimethoprim (TMP-SMZ), quinupristin-dalfopristin, clindamycin and tigecycline. These antibiotics are gradually losing their efficiency as MRSA strains are developing resistance against them [8,20,53]. Presently, the therapeutic alternatives available for treatment of infections caused by MRSA and S. aureus with reduced vancomycin susceptibility are limited. Therefore, there is a global urgency for the development of novel drugs that will be effective in the treatment of S. aureus exhibiting multidrug resistance so as to combat the scourge caused by the microorganism in the globe [52].

Prospects of medicinal plants as therapeutic option for MRSA
Natural products including medicinal plants have contributed immensely to human health, wellbeing and development of novel drugs. They are useful natural blueprints for the development of new drugs (especially in western countries) or/and phytomedicines purified to be used for the treatment of disease (commonly in developing countries and Europe) [54]. Medicinal plants can be valuable therapeutic resources. In numerous developing countries, including Nigeria, 80% of patients use home-made phytomedicines to treat infectious diseases. Despite the availability of modern medicine in some communities, the use of medicinal plants has remained high due to their efficacy, popularity and low cost. They also represent sources of potentially important new pharmaceutical substances since all the plants parts are utilized in traditional treatment and can therefore, act as lead compounds ( Table 1).
The applications of phytomedicines for human well being and as blueprints for developing novel useful drugs have drastically increased worldwide in recent years [77].
The emergence of multidrug-resistant infectious agents associated with over-and inappropriate use of antibiotics has necessitated the World Health Organization (WHO) to acknowledge and pronounce the urgent need to develop novel antimicrobials and/or new approaches to tackle the menace caused by them in the globe; these have subsequently led to the resuscitation of the interest in medicinal plants [78]. The most common bacteria that have been used in susceptibility tests with numerous medicinal plants include: Staphylococcus aureus, methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococcus (VRE), Pseudomonas aeruginosa Helicobacter pylori, etc [54]. Presently, numerous studies have reported the antibacterial activity of many plant extracts against MRSA. In this study, only fifty-one (51) plants with anti-MRSA activities from thirtyfive (35) families were mentioned ( Table 1). The minimum inhibitory concentrations (MIC) values of the plants on the tested MRSA strains were between 1.25 µg/ml to 6.30 mg/ml. Twenty-nine of the plants had MIC values < 1.0 mg/ml while the remaining twenty-two MIC values were > 1.0 mg/ml but < 8.0 mg/ml. Extracts exhibiting activities with MIC values below 8 mg/ml are widely accepted to possess some antimicrobial activity while those with values below 1 mg/ml are considered noteworthy [77,79]. However, most of the plants in this review were not tested on S. aureus strains with reduced vancomycin susceptibility.
The solvents used for the medicinal plants extraction in this review were ethanol and methanol (Table 1). This is probably because alcoholic extracts have higher antimicrobial activity than aqueous extracts. It has been reported that ethanolic extracts have higher antimicrobial activity than aqueous extracts because of the presence of higher amounts of polyphenols. They are more efficient in cell walls and seeds degradation causing polyphenols to be released from cells. Also, the enzyme polyphenol oxidase, degrades polyphenols in water extracts but is inactive in methanol and ethanol. Moreover, water is a better medium for the growth of microorganisms than ethanol [80].
Although, methanol is more polar than ethanol but it is not frequently used for plant extraction due to its cytotoxic nature that may give incorrect results [81].
Extracts of medicinal plants are rich in phytochemicals. Phytochemicals or secondary metabolites are natural protective agents biosynthesized by plants against external stress and pathogenic attack. They are crucial for plant defences and survival. They have been divided into several categories: phenolics, alkaloids, steroids, terpenes, saponins, etc. They exhibit other bioactivities such as antimutagenic, anticarcinogenic, antioxidant, antimicrobial, and antiinflammatory properties and are therefore responsible for the medicinal potential of plants (Table 2). Hence, from this review, anti-MRSA plants have antibacterial effect on MRSA strains and other medicinal/therapeutic uses as depicted in Table 2.  The therapeutic properties of these medicinal plants obtained from their phytochemicals could be employed for drug development [118]. The antibacterial (anti-MRSA) activity of these plants is attributed to their phytochemical contents. For instance, flavonoids complex with bacterial cell wall, extracellular and soluble protein while tannins inactivate microbial adhesions, enzymes and cell envelop proteins [55,[67][68][69]119].
Although, these anti-MRSA plants are likely promising candidates for drug development for MRSA infections, it has been reported that most plants contain potentially toxic, mutagenic, and/or carcinogenic substances. Therefore, it is highly recommended that medicinal plants undergo a critical sequential antimicrobial, pharmacological, and toxicology screening to ascertain their safety and selection as good candidates for novel drug development [77,79,120].

Conclusion
S. aureus is a common microorganism that is widely spread in the human population with many being asymptomatic carriers. It can also cause life-threatening infections and its strains have evolved into MRSA and strains with reduced vancomycin susceptibility (VISA, hVISA and VRSA). These strains cause infections and diseases that are either difficult to treat or resistant to the empiric antibiotics usually prescribed for treatment. The globe is running short of drugs/antibiotics available for therapy as a result of infections associated with this organism.
Many research studies have reported that some medicinal plants in different countries have anti-MRSA activities due to their phytochemical contents. These plants can be employed as alternative candidates for drug development to halt or/and control the infections of multi-drug resistant S. aureus. However, there is a need for further studies to adequately determine the safety and clinical efficacy of anti-MRSA plants to man.