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 Table of Contents  
ORIGINAL ARTICLE
Year : 2014  |  Volume : 27  |  Issue : 4  |  Page : 825-832

Characterization of vancomycin-resistant Staphylococcus aureus in the National Liver Institute


1 Microbiology Department, National Liver Institute
2 Microbiology Department, Menoufia University

Date of Submission12-Dec-2013
Date of Acceptance17-Feb-2014
Date of Web Publication22-Jan-2015

Correspondence Address:
Hala A El Refai Khalil
41 Madkhal Cairo Street, Cairo 62825

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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/1110-2098.149802

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  Abstract 

Objective
The aim of the study was to assess vancomycin-resistant Staphylococcus aureus (VRSA) in the National Liver Institute.
Background
VRSA refers to the strains of S. aureus that have become resistant to vancomycin. Three classes of VRSA have emerged: vancomycin intermediate-resistant S. aureus, heterogenous vancomycin-intermediate S. aureus, and high-level VRSA.
Materials and methods
The study was carried out on 705 samples that were collected from patients, personnel, and hospital environments. Group I (the patient group) included samples from 555 patients. From them different microbiological samples were collected (blood, urine, sputum, wound aspiration, nasal swab, throat swab, drains, and endotracheal tubes). Group II (the personnel group) included 45 samples from staff members who were in contact with the patients. Swabs were taken from their noses, throats, and fingertips. Group III (the hospital environment) included samples from 105 swabs.
Samples from all groups were subjected to culture and isolation of the bacteria and identification of S. aureus isolates. These isolates were subjected to an antibiogram using the disc diffusion method, and testing for vancomycin susceptibility by the disc diffusion method, an E-test (E-test strip is Hi Media laboratories Pvt. Limited, Mumbai, India), and Vitek-2 compact (Vitek-2 is bioMιrieux, Inc., Durham, NC). VRSA isolates were subjected to analysis of plasmid DNA profile and restriction fragment length polymorphism (RFLP) PCR of isolated plasmids to detect the vanA gene.
Results
This study showed that, of 145 isolated S. aureus strains, 58.64% were vancomycin-sensitive S. aureus, 20.68% were vancomycin intermediate-resistant S. aureus, and 20.68% were VRSA. Their plasmid profile showed that 10% had no plasmid, 26.7% had one plasmid, 50% had two plasmids, and 13.3% had three plasmids. The molecular weight of most plasmids was 100 kb (36%). RFLP PCR for detection of the vanA gene in the isolated plasmids was positive in 51.9%.
Conclusion
VRSA prevalence in the National Liver Institute is 20.68%. The majority (90%) carried plasmids. Most plasmids had a molecular weight of 100 kb, and the vanA gene, as detected by RFLP PCR, was positive in 51.9%.

Keywords: RFLP PCR, Staphylococcus aureus plasmids, Staphylococcus aureus, vanA gene, vancomycin-resistant Staphylococcus aureus


How to cite this article:
Ghoniem EM, El Hendawy GR, Abdel Moteleb TM, Hassan HA, El Refai Khalil HA. Characterization of vancomycin-resistant Staphylococcus aureus in the National Liver Institute. Menoufia Med J 2014;27:825-32

How to cite this URL:
Ghoniem EM, El Hendawy GR, Abdel Moteleb TM, Hassan HA, El Refai Khalil HA. Characterization of vancomycin-resistant Staphylococcus aureus in the National Liver Institute. Menoufia Med J [serial online] 2014 [cited 2020 Apr 3];27:825-32. Available from: http://www.mmj.eg.net/text.asp?2014/27/4/825/149802


  Introduction Top


The genus Staphylococcus comprises 38 validly described species and subspecies of Gram-positive, catalase-positive cocci [1]. The most commonly associated with human infections are Staphylococcus aureus (most virulent), Staphylococcus epidermidis, Staphylococcus haemolyticus, and Staphylococcus saprophyticus. S. aureus produces coagulase, whereas all other species are referred to as coagulase-negative staphylococci [2].

S. aureus, a major cause of potentially life-threatening infections acquired in healthcare and community settings, has developed resistance to most classes of antimicrobial agents. A dramatic increase in the number of healthcare-associated infections due to methicillin-resistant S. aureus (MRSA) in the 1990s and the recent emergence of MRSA in community-associated infections highlight the growth of this species as a pathogen and its ability to adapt under pressure from antimicrobial agents. Glycopeptides such as vancomycin provide effective therapy against most multidrug-resistant strains of S. aureus [3].

However, since 2002, nine MRSA strains that were also resistant to vancomycin (VRSA) have been reported in the USA. This can be explained by an increase in vancomycin use that has led to the emergence of two types of glycopeptide-resistant S. aureus. The first one, vancomycin intermediate-resistant S. aureus (VISA), is associated with a thickened and poorly cross-linked cell wall, resulting in accumulation of acyl-d-alanyl-d-alanine (X-d-Ala-d-Ala) targets in the periphery that sequester glycopeptides. The second type, VRSA, is due to acquisition of the vanA operon from Enterococcus spp., resulting in high-level resistance. Seven types of resistance (VanA, -B, -C, -D, -E, -G, and -L) in enterococci have been described; these correspond to specific operons (vanA, -B, -C, -D, -E, -G, and -L). VanA-type resistance, which was the first to be elucidated and which is the most common, is characterized by high levels of resistance to glycopeptides, vancomycin, and teicoplanin and is chromosomally or plasmid-located [4].

Vancomycin acts by inhibiting proper cell wall synthesis in Gram-positive bacteria. Because of the different mechanisms by which Gram-negative bacteria produce their cell walls and the various factors related to permeating the outer membrane of Gram-negative organisms, vancomycin is not active against Gram-negative bacteria (except some non-gonococcal species of  Neisseria More Details) [5].

In enterococci, glycopeptide resistance is due to the acquisition of transferable operons containing genes that enable the organism to synthesize cell walls from precursors in which lactate replaces the terminal alanine, rendering vancomycin incapable of binding to its d-Ala-d-Ala target [6].

The concentration of vancomycin required to inhibit most strains of S. aureus is typically 2 mg/l or lower. S. aureus isolates for which vancomycin minimum inhibitory concentrations (MICs) are 4-8 mg/l are currently classified as VISA, and isolates for which vancomycin MICs are at least 16 mg/l are classified as VRSA [7].


  Materials and methods Top


This study was conducted from May 2010 to July 2012 in the National Liver Institute, Menoufia University.

Participants

The studied participants were classified into three groups as follows:

Group I (the patient group)

This group included 555 patients (322 male and 233 female patients), their ages ranging from 1 day to 70 years. Different microbiological samples, such as blood, urine, sputum, wound aspiration, nasal swab, throat swab, drains, and endotracheal tubes, were collected from them.

Group II (the personnel group)

This group included 45 individuals from the hospital staff (doctors, nurses, and workers) who were in contact with the patients. Swabs were taken from their noses, throats, and fingertips.

Group III (hospital environment)

This included samples from within the hospital environment, including walls, floor, trolleys, sinks, beds, and suction devices.

Collection of samples

Group I (patient samples)

One clinical sample was collected from each patient as follows:

  1. Sputum samples (45) and endotracheal aspirates (30).
  2. Blood samples (80).
  3. Urine samples (80).
  4. Samples from wounds and other soft tissue infections (100).
  5. Aspiration by syringe from drains and stents from surgical site infections (120).


Group II (medical staff)

Swabs were obtained from the noses and throats (45) of staff members who were in contact with patients [8]. Swabs from their fingertips (45) were also collected [9].

Group III (the environment)

Broth-moistened swabs were taken from walls, floor, trolleys, sinks, beds, and suction devices and were transported quickly to the laboratory to avoid any false results [10].

All samples were subjected to different microbiological methods including the following:

(1) Culture and isolation of the bacteria [11]:

  1. Samples were inoculated onto nutrient agar, blood agar, MacConkey's agar, and mannitol salt agar plates and incubated at 37°C for 24 h.
  2. Blood cultures were incubated at 37°C for 7 days and subcultured on the same media on the 3rd, 5th, 7th, 10th, and14th day or when any sign of microbial growth appeared in blood cultures.


(2) Identification of S. aureus [11]:

Culture characteristics were analyzed on different media with microscopic examination, evaluation of biochemical characteristics, and using a Vitek-2 compact system.

Testing for vancomycin susceptibility was carried out with the following methods:

  1. Disc diffusion method:

    The disc diffusion method was carried out according to the protocol of the Clinical and Laboratory Standards Institute [12].
  2. MIC determination [13]: using E-test strips.
  3. The Vitek-2 compact system.


VRSA strains were subject to the following:

Antibiogram

This was performed using the disc diffusion method according to the protocol of the Clinical and Laboratory Standards Institute [7,12], using the following antibiotic sensitivity discs supplied by Oxoid (Wade Rd, Basingstoke, Hampshire RG24 8PW, United Kingdom):

Ampicillin 10 mcg/disc, ampicillin-sulbactam 10/10 mcg/disc, amoxicillin 10 mcg/disc, amoxicillin-clavulanic acid 30 mcg/disc, erythromycin 15 mcg/disc, azithromycin 15 mcg/disc, levofloxacin 5 mcg/disc, ciprofloxacin 5 mcg/disc, tetracycline10 mcg/disc, tigecycline 15 mcg/disc, clindamycin 2 mcg/disc, rifampicin 30 mcg/disc, septrin 25 mcg/disc, linezolid 30 mcg/disc, vancomycin 30 mcg/disc, oxacillin 1 μg/disc, gentamicin 10 mcg/disc, and amikacin 30 mcg/disc.

Analysis of plasmid DNA [14]

Preparation of plasmids

Plasmids were prepared by a modified Brinboim and Dolly's method [14]:

  1. A single bacterial colony was used to inoculate a 5 ml nutrient broth, which was incubated overnight in a shaking incubator at 37°C.
  2. A volume of 1.5 ml of each isolate taken in an Eppendorff tube was centrifuged for 5 min using a microfuge.
  3. The supernatant was discarded and the bacterial pellet was resuspended by vortexing in 100 μl of an ice-cold solution of reagent I containing 5 mg lysozyme. After 5 min at room temperature, the cells were lysed by adding 200 μl III to each tube. The contents were mixed thoroughly but gently.
  4. After storing the tubes for 5 min on ice, the bacterial debris was separated by centrifugation of a freshly prepared solution of reagent II. The contents of the tube were mixed by inverting it several times until it became clear and viscous, indicating complete lysis.
  5. After placing the tubes on ice for 5 min, 150 μl of an ice-cold solution of reagent was added for 5 min and the tubes were placed in a microfuge. The supernatant was transferred to fresh sterile tubes and two volumes of ice-cold 95% ethanol was added and the contents were mixed by inverting the tubes 2-3 times. The tubes were kept on ice for 20 min and then centrifuged for 5 min. The supernatant was discarded and the tubes were left in an inverted position on a paper towel to drain the liquid and dry the pellet.
  6. The pellet was resuspended with 1 ml of 70% ethanol and centrifuged for 5 min. The DNA pellet was dissolved in 20 μl of TE buffer (pH 8), and 10 μl of the loading buffer was added.


Agarose gel electrophoresis [14]

Agarose gel electrophoresis was performed using 0.8% agarose, which was melted well with Tris borate buffer in a microwave and mixed well before pouring into the electrophoresis tray to a depth of 7 mm.

Samples of plasmid preparations of the tested MRSA isolates and the standard molecular weight plasmid strain were loaded into wells in the gel made by a comb inserted during casting of the gel. Electrophoresis was performed on a horizontal apparatus and it was run at 60 V. The gel was stained with a solution of ethidium bromide (0.5 μg/ml) in Tris borate buffer for 30 min at room temperature in a dark place. Plasmid DNA was visualized using a UV transilluminator and photographs were taken with a digital camera with 667 instant films.

Molecular weight estimation of plasmids [14]

The molecular size of unknown plasmids was determined on the basis of their mobility through agarose gel, which was compared with the mobility of known molecular size plasmids present in the standard (140, 10, 4.8, 2.7, 1.6, 1.4, and 1.3 MDa). A standard curve was drawn by plotting the distances traveled by standard plasmids in agarose gel against the logarithm of their known molecular weights. The molecular weight of unknown plasmids was determined using the standard curve.

Restriction fragment length polymorphism polymerase chain reaction for detection of the vanA gene from isolated plasmid [3]

The PCR mixture (100 μl per reaction mixture) consisted of 10 mmol/l Tris-HCl, 50 mmol/l KCl, 2.5 mmol/l MgCl 2 , deoxyribonucleotide as triphosphates (final concentration, 0.2 mmol/l each), 100 pmol of each primer, 100 ng of template DNA, and 1.5 U of Taq DNA polymerase in reactions with the vanA forward and vanA reverse primer sets.

Amplification was performed and consisted of the following steps: predenaturation at 95°C for 5 min, and then 35 cycles for 30 s at 95°C; for 20 s at 25°C; and for 30 s at 72°C. A final extension step at 72°C for 7 min.

DNA digestion with restriction enzyme [15]

Ten microliters of the PCR product was inoculated overnight with 10 U of HindIII according to the manufacturer's recommendations. In a 1.5 ml tube, 50 μl of the following mixture was prepared: PCR product, 10 μl; HindIII, 10 U; NEBuffer2; BSA; and dH 2 O. The compounds were gently mixed by pipetting. The mixture was incubated for 16 h at 37°C. Both the PCR products and the restriction digest fragments were detected by electrophoresis using a 2% agarose gel in the presence of ethidium bromide and then photographed under UV illumination.

Statistical analyses

Using Microsoft Excel 2007 and SPSS (v17.0; SPSS Inc., Chicago, Illinois, USA) for Microsoft Windows 7 the clinical and laboratory data were statistically analyzed to obtain [16]:

Descriptive statistics: These included mean (X -),SD, and range (minimum and maximum).

(1) Analytical statistics: Qualitative variables were expressed as number and % and analyzed by means of the c2 -test.

A P-value less than 0.01 was considered statistically significant.

This work was approved by the Ethics Committee of our University.


  Results Top


S. aureus was the most prevalent isolated organism, being identified in 26.12% of patients, whereas enterococci was the least prevalent isolated organism, representing 2.16% of the patients. The incidence of S. aureus infection was significantly higher in elderly patients and nonsignificantly higher in male patients.

According to methicillin susceptibility, 28.27% of S. aureus isolates were methicillin-sensitive S. aureus (MSSA), whereas 71.72% were MRSA. According to vancomycin susceptibility 58.62% were vancomycin-sensitive S. aureus, 20.68% were VISA, and 20.68% were VRSA.

According to the results of the antibiogram, isolated VRSA showed fair susceptibility to rifampicin (50%), whereas susceptibility to erythromycin, azithromycin, ciprofloxacin, gentamicin, levofloxacin, tetracycline, septrin, tigecycline, amikacin, and clindamycin was diminished, with a percentage of 13.3, 20, 20, 30, 33.3, 33.3, 36.6, 40, 43.3, and 43.3%, respectively. For ampicillin, ampicillin-sulbactam, amoxicillin, and amoxiclav, all isolates were resistant. On the other hand, all VRSA isolates were sensitive to linezolid.

Three (10%) VRSA isolates were without plasmids, whereas 27 (90%) isolates contained plasmids: 26.7% had one plasmid, 50% had two plasmids, and 13.3% had three plasmids. Molecular weights ranged from 4 to 120 kb; the most common weight was 100 kb (36%) and the least was 60 kb (2%).

Restriction fragment length polymorphism (RFLP) PCR for detection of the vanA gene from isolated plasmids was nonsignificantly negative in 48.1%, whereas it was positive in 51.9% [Figure 1],[Figure 2],[Figure 3],[Figure 4] and [Figure 5] and [Table 1],[Table 2],m[Table 3],[Table 4].
Figure 1: Prevalence of different organisms isolated from the patient group. CONS, coagulase-negative staphylococci.

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Figure 2: Methicillin and vancomycin susceptibility pattern of isolated Staphylococcus aureus. MRSA, methicillin-resistant S. aureus; MSSA, methicillin-sensitive S. aureus; VISA, vancomycin intermediate-resistant S. aureus; VRSA, vancomycin-resistant S. aureus; VSSA, vancomycin-sensitive S. aureus.

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Figure 3: E-test for vancomycin-resistant Staphylococcus aureus isolate.

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Figure 4: Gel electrophoresis of plasmid extracted from four vancomycin-resistant Staphylococcus aureus (VRSA) isolates. L0: ladder 1.3, 1.4, 1.6, 2.7, 4.8, 10, 140, and 210 kb; L1: VRSA isolate with single plasmid 120 kb; L2: VRSA isolate with two plasmids 4 and 100 kb; L3: VRSA isolate with two plasmids 4 and 20 kb; L4: VRSA isolate with single plasmid 100 kb; L5: plasmidless VRSA isolate; L6: plasmidless VRSA isolate.

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Figure 5: Electrophoretic pattern of the vanA gene on the basis of restriction fragment length polymorphism. L0: 1 kb molecular marker (10, 8, 6, 5, 4, 3, 2.5, 2, 1.5, 1 kb); L1, L2, L3, L5, L6: plasmids positive for vanA gene showing different molecular weight patterns; L4: negative for vanA gene.

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Table 1: Antibiotic susceptibility pattern of 30 vancomycin-resistant Staphylococcus aureus isolates

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Table 2: Frequency of plasmid quantity in the vancomycin-resistant Staphylococcus aureus isolates

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Table 3: Molecular weight (in kb) of isolated plasmids from the vancomycin-resistant Staphylococcus aureus isolates

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Table 4: Restriction fragment length polymorphism polymerase chain reaction for detection of the vanA gene in the isolated plasmids

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  Discussion Top


This study showed that the most frequent healthcare-associated isolate was S. aureus (26.12%), followed by Klebsiella spp. (24.50%),  Escherichia More Details coli (23.42%), coagulase-negative staphylococci (12.43%), Streptococcus spp. (4.32%), Pseudomonas spp. (4.32%), Proteus spp. (2.70%), and Enterococcus spp. (2.16%).

These results were comparable to the survey carried out in British hospitals by Wilson et al. [17], which revealed that over 26% of isolates causing healthcare-associated infections were S. aureus. A nearly similar result was found by Fattom et al. [18], who reported that 20-25% of all healthcare-associated infections were caused by S. aureus, as well as by Zaghlol [14] who found that S. aureus accounted for 26.7% of all healthcare-associated isolates. However, a higher rate (70%) of healthcare-associated S. aureus isolates were found in a study carried out by Arakere et al. [19]. This may be because the incidence of S. aureus bacteremia and its complications has increased sharply in recent years because of the increased frequency of invasive procedures, increased numbers of immunocompromised patients, and increased resistance of S. aureus strains to available antibiotics.

Regarding the age of the studied patients, the highest percentage (42.7%) of S. aureus infections was detected in patients above 55 years possibly because of the decrease in initial inflammatory response, slow healing in old age, and weakened immunity in this age group.

Our results coincide with that of Warren [20], who stated that S. aureus infection was significantly correlated with older age. Also, Bradley [21] found the same results but they attributed it to the underlying disease and functional debility, rather than the effect of age itself. However, Munckhof et al. [22] found no significant relationship.

Sex distribution in this study showed that S. aureus infection was higher in male patients (56.55%), although there was no statistical significance. These findings were similar to those of Primo et al. [23] who found that 57% of patients were male.

In the present study, 28.27% of the isolated S. aureus strains were MSSA, and 71.72% were MRSA. For vancomycin, 58.62% were vancomycin sensitive, 20.68% were VISA, and 20.68% were VRSA; all of them were MRSA.

These results were similar to those of Thati et al. [24], who found that 79.6% of S. aureus isolates were identified as MRSA and 1.96% were VRSA and that all VRSA isolates were MRSA. However, the proportions found in this study were higher than those of Zaghlol [14] and Dubey et al. [25], who found that 32 and 42.94% of S. aureus isolates were MRSA, respectively.

In all, 81.73% of S. aureus isolates were MRSA and 23.86% were VRSA, as found in the study by Taha [26]. These results agree with ours.

In contrast, Cosgrove et al. [27] found that 65.7% of studied patients had MSSA bacteremia, and 34.3% had MRSA bacteremia. This difference from our study may be due to overcrowding in the wards, which may have led to a higher rate of S. aureus through nasal carriage.

The VRSA isolates in our work showed fair susceptibility to rifampicin (50%) and diminished susceptibility to erythromycin (13.3%), azithromycin (20%), ciprofloxacin (20%), gentamicin (30%), levofloxacin (33.3%), tetracycline (33.3%), septrin (36.6%), tigecycline (40%), amikacin (43.3%), and clindamycin (43.3%). All isolates were resistant to ampicillin, ampicillin-sulbactam, amoxicillin, and amoxiclav, and all were sensitive to linezolid.

Similarly, CDC [28] documented two VRSA isolates that were susceptible to linezolid, rifampicin, tetracycline, and tigecycline and were resistant to trimethoprim/sulfamethoxazole.

In addition, Taj et al. [29] documented that 22.2% of the 450 S. aureus isolates were VRSA; this isolate was found to be resistant to several other antimicrobials, such as gentamicin, tobramycin, amikacin, ciprofloxacin, erythromycin, and tetracycline.

Further, Wunderink et al. [30] documented that linezolid and other newer drugs are effective against both CA-MRSA and HA-MRSA. Linezolid is now felt to be the best drug for treating patients with MRSA, which agrees with our results.

In the present study, 10% of VRSA isolates had no plasmid, 26.7% had one plasmid, 50% had two plasmids, and 13.3% had three plasmids. Their molecular weight ranged from 40 (12%) to 120 (18%) kb. The most common (36%) was the 100 kb plasmid, which was present in 18 strains, and the least common (2%) was the 60 kb plasmid, which was present in one isolate.

This agrees with the study by Adebayo et al. [31], who stated that S. aureus isolates, and particularly those from hospitals, often carry one or more free or integrated plasmids. Out of seven strains, 57.14% contained two plasmids and 42.86% contained one plasmid.

One VRSA isolate was found by Saha et al. [32] and contained a large plasmid (53.4 kb) and four small plasmids of 6, 5.5, 5.1, and 1.5 kb. The large plasmid harbored the vancomycin-resistance gene cluster vanHAX - vanH (969 bp), vanA (1032 bp), and vanX (609 bp) - which was confirmed by PCR amplification.

Plasmids play an essential role in bacterial metabolism, pathogenesis, and evolution by harboring genes coding for diverse factors and facilitating their dissemination. Of particular importance is their role in the dissemination of resistance genes contributing to the current epidemic of resistant infections [33].

In contrast, Daini et al. [34] found that 47.05% of multiple resistant S. aureus isolates harbored plasmids with molecular weight ranging from 0.28 to 25.12 kb and plasmids were not detected in 18 (52.94%) of the resistant strains, indicating that their resistance was probably chromosomal.

Moreover, Udo and Jacob [35] found that the resistance of S. aureus to some antibiotics such as methicillin, benzyl penicillin, gentamicin, kanamycin, neomycin, streptomycin, tetracycline, trimethoprim, and ciprofloxacin was chromosomally mediated.

As regards the vanA gene, as detected by RFLP PCR, 51.9% of the isolates were positive and 48.1% were negative; this difference was of no significance. Also, 25 VRSA isolates had different gene restriction patterns. Three isolates collected simultaneously from the same department had the same pattern. Another two isolates from another department collected at the same time also had the same pattern, indicating a common infection source.

Our results agree with that of Zhu et al. [15], who found that 57.14% of studied VRSA isolates were positive for the vanA gene. Their HindIII plasmid restriction patterns showed that three isolates had the same pattern, indicating that these plasmids were related, and one isolate was different from the other three isolates, concluding that this plasmid was independent.

In contrast, Tiwari and Sen [36] found that none of the studied VRSA isolates had demonstrated the presence of the VanA/VanB gene by PCR.

There were two VRSA isolates documented by CDC [28] that were PCR positive for the vanA gene and showed different plasmid restriction patterns indicating that they had distinct plasmids.


  Conclusion Top


From these results we conclude that S. aureus was the most prevalent organism isolated from the patient group, being found in 26.1% of patients.

The VRSA isolates were significantly resistant to azithromycin, erythromycin, ciprofloxacin, tetracycline, and gentamicin, and all were sensitive to linezolid.

The plasmid profile for VRSA isolates showed that 10% had no plasmid, 26.7% had one plasmid, 50% had two plasmids, and 13.3% had three plasmids. The most common molecular weight of plasmids was 100 kb (36%) and the least was 60 kb (2%). The vanA gene was positive in 51.9% of isolated plasmids, as detected by RFLP PCR. Some of those VRSA isolates showed a similar resistance pattern, suggesting a common source of infection.

Recommendations

  1. The presence of VRSA isolates indicate that resistance to vancomycin will appear because of overuse. We recommend that this drug should not be used (prescribed) except if it is highly indicated and only after susceptibility tests.
  2. Most of the VRSA isolates in the present study were highly resistant to most of the tested antibiotics. This may be because control of antibiotic use is not strictly followed by clinicians, leading to selection of resistant strains, and therefore we recommend strict adherence to sensitivity test results.
  3. In this study, the routine disc diffusion method failed to detect strains categorized as intermediately sensitive to vancomycin. Therefore, we recommend the use of the E-test and Vitek-2 compact in screening of patients.
  4. For VRSA, we recommend the culture of the anterior nares of contacts on a regular basis.
  5. Molecular epidemiological tools are helpful for understanding transmission patterns and sources of infection; plasmid profile and RFLP PCR are molecular techniques that provide good discriminatory index in typing. Therefore, we recommend wider application of molecular typing in our hospitals, which should shed light on the epidemiology of hospital-acquired infections and thus allow for more effective control and prevention strategies.



  Acknowledgements Top


Conflicts of interest

There are no conflicts of interest.

 
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    Figures

  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5]
 
 
    Tables

  [Table 1], [Table 2], [Table 3], [Table 4]



 

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