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African Journal of Microbiology Research Vol. 4(13) pp. 1373-1382, 4 July, 2010 Available online http://www.academicjournals.org/ajmr ISSN 1996-0808 2010 Academic Journals Impact of multi drugs resistant bacteria on the
pathogenesis of chronic suppurative otitis media
Ihsan E. Alsaimary1*, Ahmed M. Alabbasi2 and Jassim M. Najim1
1Department of Microbiology, Col ege of Medicine, University of Basrah – Republic of Iraq. 2Department of Surgery, Col ege of Medicine, University of Basrah – Republic of Iraq. One hundred twenty patients with chronic suppurative otitis media (CSOM) in Basrah, 65 (54.2%) males
and 55 (45.8%) females, with male: females ratio (1.2:1) and 60 individual without otological problems as
control group were included in this study, which done during the period between March 2009 and
January 2010. This includes the collection of aural swab samples, culturing of samples, identification of
causative agent’s species and antibiotic sensitivity. Gram’s negative bacteria were the commonest
microorganism comprises (60%). Pseudomonas aeruginosa was common causative agent (19.04%),
followed by Staphylococcus aureus (16.7%) and Klebsiella spp. (14.3%). Mixed infection was found in
high percent (74%), in which P. aeruginosa and other microorganisms were more common. The
antibiotic sensitivity pattern showed that P. aeruginosa was sensitive to Ciprofloxacin, amoxicillin
+clavulanic acid and gentamicin, while other is appeared resistant, S. aureus was sensitive to
ciprofloxacin, amoxicillin+clavulanic acid, erthomycin, cephalexine and it is resistant to penicillin and
ampicillin, klebsiella species were sensitive to ciprofloxacin, amoxicillin +clavulanic acid, gentamicin,
while resistant to tetracycline.

Key words:
Pseudomonas aeruginosa, Staphylococcus aureus, Klebsiel a spp., chronic suppurative otitis
media, aural swab, antibiotic sensitivity, gram’s negative bacteria, otological.
INTRODUCTION
Otitis media is inflammation of the middle ear. This is commonly caused by the build up of fluid behind the ear CSOM, for the purposes of this document, defined as a drum, as a result of a blockage to the Eustachian tube. chronic inflammation of the middle ear and mastoid Otitis media is more common in children, as their cavity, which presents with recurrent ear discharges or Eustachian tube is shorter and more horizontal than otorrhoea through a tympanic perforation (Howard, adults and is made up of more flaccid cartilage, which 2007). The disease usual y begins in childhood as a can impair its opening (Bluestone and KLien, 2001). Otitis spontaneous tympanic perforation due to an acute media can cause a mild to moderate hearing loss, due to infection of the middle ear, known as acute otitis media the fluid interfering with the transmission of sound which presents with a rapid onset of signs and through to the inner ear. It can often affect the tympanic symptoms, such as pain, fever, irritability; a red bulging membrane causing it to retract or become inflamed. The ear drum and middle ear effusion (Jahn, 1991). fluid can cause the tympanic membrane to bulge and In CSOM the bacteria may be aerobic (e.g. become inflamed and occasional y the tympanic membrane wil perforate. There are three common types Staphylococcus aureus, Streptococcus pyogenes, of otitis media, acute purulent otitis media, otitis media Proteus mirabilis, Klebsiel a species) or anaerobic (e.g. with effusion and chronic suppurative otitis media Bacteroides, Peptostreptococcus, Proprionibacterium) (Saunders et al., 2009; Brook, 1996). The present studies aimed to identify the bacterial pathogens associated with CSOM, study the antibiotic susceptibility pattern of *Corresponding author. E-mail: ilsanalsaim@yahoo.com. antibiotic against bacterial pathogen, and determined the Table 1. Bacterial types isolated from healthy person (control
Sampling
Two groups were included in this study: Group (1) 120 aural swabs were taken from infected ear of CSOM patients. Group (2) 60 aural Microorganisms
No. of isolates
swab were taken from a control group. Swabs were taken under sterile condition and transfer immediately to the laboratory by brain heart broth for aerobic bacteria, thioglycol ate broth for anaerobic bacteria, and cultured on suitable media at 37˚C for 24 - 48 h. Primary isolation on (Blood agar, chocolate agar, nutrient agar), then on selective media identification and biochemical characteri- zation were carried out according to standard routine techniques (Fingole and Baron, 2002). Note: Al media are sterilized by autoclave (121˚C under 15 Ibs pressure for 15 min). Antibiotics disc Table 2. Bacterial type isolated from patients with CSOM.
Caustive agents
No. of isolates
Statistical analysis
In order to determine the statistical significance among different variables, SPSS program (statistical program for social sciences) ver.11, was used for this purpose. The fol owing statistical testes were performed: Chi-square (x2) test and the difference between two proportions by T-test were used to assess the significance of difference between groups. P-value less than 0.05 was considered as statistical y significant (S), p-value < 0.01 as highly significant and (HS), p-value < 0.001 as extremely significant (ES). p-value more than 0.05 was considered as statistical y not significant (NS). mode of bacterial isolation and multi drugs resistant Table 1 show results of isolated bacterial from (60) healthy persons. The fol owing bacteria were isolated, Staphylococcus epidermidis 20 isolates (40%), fol owed by Corynebacterium species 15 isolate (30%). Other MATERIALS AND METHODS
types distributed according to species in Tables 3 - 8. Ten samples gave negative result for bacteria culture Patients
A total of 120 patients with CSOM were included in this study, the diagnosis of CSOM was carried out according to clinical examination by otoscopic and tuning fork examination, and audiological investigation Pathogenic bacteria isolated from patients with
(pure tone audiometry and tympanometry under supervision of specialists of ENT. Microbiological investigation includes (culture, identification of causative agents and antibiotic sensitivity. The study was carried out in Basrah General Hospital, out patients The occurrence of various bacterial isolate among CSOM E.N.T. clinic, during the period from March 2009 - January 2010. patients shown in Table 2 presents that P. aeruginosa was more frequently isolates 40 (19.41%), while S. aureus fol owed by Pseudomonas 35 (16.99%), Klebsiel a 30 (14.56%) Branhamel a catarrhalis 20 Control group
(9.70%), Proteus 20(9.70%), Heamophilus influenzae 20(9.70%), Streptococcal spp. 15(7.28%), E. coli A total of 60 individuals without otological problems, 30 males and 30 females in various age group, they were regarded as a control 10(4.85%), Corynebacterium 8 (3.88), and Bacil us 8 Table 3. Relationship between caustive agents and hearing loss.
No. of isolates with Hearing Loss (%)
Causative agent
No. of isolates
Not applicable
CHL: Conductive hearing loss, SNHL: Senserineural hearing loss and MXHL: Mixed hearing loss. Table 4. Standard antibiotic susceptibility test according to diameters of inhibition zone supplied by bioanalysis company.
Antimicrobial agent
Conc. mcg
Zone diameter (mm).
Sensitive
Resistant

Bacterial pathogens and hearing impairment
Amoxicil in + clavulanic acid (Augmentin) and Gentamicin are statistical y significantly higher than other types of The occurance of various caustive agents isolates among antibiotics. P < 0.01 in percentages of sensitivity between CSOM patients in three types of hearing loss (CHL, (50 - 75%) (p < 0.01), while 88% of P. aeruginosa SNHL, MXHL) are shown in Table 3. P. aeruginosa was isolates was resist by trimethoprim, 85% to Streptomycin, more frequently isolated in senserineural and profound and 80% to Vancomycin, while other pattern of resistance hearing loss (25 - 26.2%), while in conductive and mixed were between 25 - 78% of various antibiotics p < 0.01. hearing loss (16.7 - 20.4%) S. aureus isolates, appeared more frequently among CSOM patients with conductive and mixed hearing loss (20.4 - 25%) than in senserineura Antibiotic sensitivity of S. aureus
and profound hearing loss (12.5 - 15%) Klebsiel a species and other organisms isolated in various Table 6 shows that in each drugs group, the frequency percentages from these three types of hearing loss. sensitivity of Ciprofloaxacin, Augmenten, Cephalexin and Penicil in (57 - 80%) were statistical y significantly higher sensitive than other antibiotic. P < 0.01, while 83% of S. Antibiotic sensitivity of P. aeruginosa
aureus isolates was resist to trimethoprim, 83% to Streptomycin, and 83% to Vancomycin, while other Table 5 show that the frequency of Ciprofloxacin, pattern of resistance were between 20 - 77% of various Table 5. Antibiotic susceptibility pattern of Pseudomonas aeruginosa.
Drugs type
No. of isolates
Sensitive (%)
Resistant (%)
Table 6. Antibiotic susceptibility pattern of S. aureus.
Drugs type
No. of isolates
Sensitive (%)
Resistant (%)

Table 7. Antibiotic susceptibility pattern of Klebsiel a spp.
Drugs type
No of isolated
Sensitive
Resistant
Antibiotic sensitivity of Klebsiella spp.
Table 7 shows that in each drugs group, the frequency of sensitivity of Ciprofloxacin and Augmentin (67 - 70%) were statistical y significantly higher than other type of antibiotic drugs. (p < 0.01), while 73% of Klebsiel a spp isolates was resist to trimethoprim, 70% to Erythromycin, and 80% to Vancomycin, while other pattern of resistance were between 30 - 73% of various antibiotics p < 0.01. Antibiotic sensitivity of B. catarrhalis
Table 8 shows that in each drugs group, Ciprofloxacin, Augmentin, Cephalexin, Ampicil in, Gentamicin were statistical y significantly higher sensitivity (50 - 75%) Table 8. Antibiotic susceptibility pattern of Branhamel a spp.
Table 10. Antibiotic susceptibility pattern of Heamophilus spp.
Drugs type
Sensitive
Resistant
Drugs type
Sensitive
Resistant
isolates
isolates
Table 9. Antibiotic susceptibility pattern of Proteus spp.
30 - 60% of various antibiotics p < 0.01. Drugs type
Sensitive
Resistant
isolates
Antibiotic sensitivity of H. influenzae
Table 10 shows that in each drugs group, the frequency of Ciprofloxacin, Augmentin, Gentamicin, Vancomycin and Lincomycin (50 - 75%) were statistical y significantly higher sensitive drugs against H. influenzae than other (p < 0.01), while 80% of H. influenzae isolates was resist to Streptomycin, 70% Tetracycline, and 70% to Ampicil in, while other pattern of resistance were between 25 - 65% Antibiotic sensitivity of Streptococcus spp.
Table 11 shows that in each drugs group the frequency of Ciprofloxacin, Augmentin, Penicil in, Erythromycin and tetracycline were statistical y significant higher sensitive (67 - 80%) than other type of Antibiotics (p < 0.01), while against Branhamel a spp than other type of antibiotic (p < 60%, of Streptococcus spp isolates was resist to 0.01), while 75% of Branhamel a spp isolates was resist Trimethoprim, 53.33% to Streptomycin and 46% to to trimethoprim, 70% to Streptomycin, and 70% to Ampicil in, while other pattern of resistance were between Vancomycin, while other pattern of resistance were 20 - 40% of various antibiotics p < 0.01. between 25 - 60 of various antibiotics p<0.01. Antibiotic sensitivity of E. coli
Antibiotic sensitivity of Proteus spp.
Table 12 shows that in each drugs group, the frequency Table 9 shows that in each drugs group, the frequency of of Ciprofloxacin, Augmentin, Gentamicin, Lincomycin and Ciprofloxacin, Augmentin, Gentamicin and Trimethoprim Cephalexin were statistical y significantly higher sensitive were statistical y significantly higher effective against drugs (60 - 80%) against E. coli than other type of drugs Proteus spp than other type of Antibiotics, (60 - 70%) (p < 0.01), while 80%, of E. coli spp isolates was resist to sensitive (p < 0.01), while 75% of Proteus spp isolates Streptomycin, 60% to Trimethoprim and 60% to was resist to Ampicil in, 70% Erythromycin, and 75% to Ampicil in, Erythromycin and penicil in, while other pattern Penicil in, while other pattern of resistance were between of resistance were between 20 - 40% of various Table 11. Antibiotic susceptibility pattern of Streptococcus spp.
Drugs type
No. of isolates
Sensitive (%)
Resistant (%)

Table 12.
Antibiotic susceptibility pattern of E. coli.
Table 13. Antibiotic susceptibility pattern of Corynebacterium spp.
Drugs type
Sensitive
Resistant
Drugs type
Sensitive
Resistant (%)
isolates
isolates
Trimethoprim were statistical y significantly higher sensitive drugs (75%) against Bacil us spp than other Antibiotic sensitivity of Corynebacterium spp.
type of drugs (p < 0.01), while 50%, of Bacil us spp isolates was resist to Lincomycin, 50% to Vancomycin Table 13 shows that in each drugs group, the frequency and 50% to Cephalexin, Penicil in and Streptomycin, of Ciprofloxacin, Cephalexin, Erythromycin, Ampicil in while other pattern of resistance were 25 - 37.5% of and Penicil in were statistical y significantly higher sensitive drugs (75%) against Corynebacterium spp (p < 0.01), while 63%, of Corynebacterium spp isolates was resist to Lincomycin, 63% to Vancomycin and 50% to Types of infection according to number of causative
Gentamicin and Cephalexin, while other pattern of resistance were between 25 - 38% of various antibiotics p Table 15 shows that the frequency of double causative agents (55 isolates, 45.83%) was statistical y significantly higher than single causative agent (38 isolates, 31.66%), Antibiotic sensitivity of Bacillus spp.
three causative agents (18 isolates, 15%) and more than three (9 isolates, 7.5%). There was no difference Table 14 shows that in each drugs group, the frequency between male and female in the frequency of various of Ciprofloxacin, Erythromycin, Ampicil in, and Table 14. Antibiotic susceptibility pattern of Bacil us spp.
Drugs type
No. of isolates
Sensitive (%)
Resistant (%)
Table 15. Modes of isolation of the bacterial pathogens among patients with CSOM.
Modes of isolated
No. of patients (%)

Bacterial agents and antibiotics
fluid build up and ear drum perforation inhibit the transmission or conduction of sound through the ear Table 16 shows that in each isolates group the frequency of susceptibility to antibiotic. P. aeruginosa was Our result goes with the study which was done by Guo statistical y significantly higher resistance than other (1994); Engel (1998), that show most patients with bacterial isolates (10.19%) fol owed by S. aureus CSOM infected by more than one pathogenic bacteria (8.73%), Klebsiel a (7.76%), B. catarrhalis, Proteus spp, leading to hearing loss, about 40 patients, (33.4%) of H. influenza (6.97%), Streptococcal spp. (4.85%), patients with CSOM suffered from bilateral hearing loss, Corynebacterium (0.9%) and Bacil us spp. (0.9%) p < 0.01. while (80 patients, 66.6) of patients with CSOM have unilateral hearing. Guo et al. (1994) studied found the effect of endotoxic damage to the strial vascularis and DISCUSSION
concluded that lipopolysaccharide induced by strial ototoxicity produced ion imbalance, causing changes in Chronic suppurative otitis media was develops from a endolymph composition and energy failure in the middle chronic bacterial infection. However, the bacteria that and inner ears organ explaining the pathogenesis of caused the initial episode of acute otitis media with perforation are usual y not those isolated from the chronic Engel et al. (1998) studied the passage of streptolysin- discharge when there is a chronic infection in the middle O and albumin through the round window membrane and ear and mastoid infection usual y polymicrobial and proposed that the passage of macromolecule, such as secondary in nature, derived from the external auditory protease, from a purulent middle ear effusion may be canal or commensal flora of nasopharynx (Bluestone and facilitated by pore forming toxins, resulting in middle and KLien, 2001). The infection causes a build up of fluid in inner ear organs damage and hearing loss. Karma et al. the middle ear. The pressure exerted by this fluid can (1978) have used gram stain not only to confirm the build up to the point where the ear drum perforated. The presence of cultured bacteria but to detected and identify
Table 16. Relationship between causative agents and antibiotics (resistance patterns).
Susceptibility to drugs
Bacterial isolate Type
Isolates
(2) drugs
(3) drugs
More than (3)
them as wel , gram stain smear were obtain from 108 ear with suppurative otitis media, fol owed by methicil in swab; in 98 (91%) of them bacteria were found, seven of resistant S. aureus (3%) and P. aeruginosa (1%). In our the 108 ear swab (6%) were devoid of bacteria both in study, we found that Klebsiel a species isolated from culture and in the gram stain. Papastavros et al. (1986) patients suffering from chronic suppurative otitis media indicated that this practice considerable error, because was (30 isolates, 14.56%). our patients infected by non viable bacteria can be as equal y incriminated as the Enterobacteriaecea such as Klebsiel a species, most of main pathogens present, furthermore, if the patients is them are among children and infants group, because the under antimicrobial treatment. In our study, we found Eustachian tube in children are shorter and wider than that, the different type of bacterial flora in the external adult. Bluestone et al. (1974) showed that young children canal were founded, S. epidermidis is the most common have shorter, straighter and more compliant Eustachian (20 isolates, 40%), fol owed by Corynebacterium species tube than adult; this permits a reflex from nasopharynx to about (15 isolates, 30%), while other type have various the middle ear with the consequence of bacterial percentages of isolation. Our result is agreed with (Pelton contamination. Brook and Yocum (1989) found that et al., 1980; Brook et al., 1996), while, it is against the result Klebsiel a species (6.2%) isolate from patients with is of (Saunders et al., 2009). Pelton et al., (1980); Brook et CSOM, while Ostfeld and Rubinstein (1980) stated that al. (1981) showed that the predominant microflora were S. (20%) of Klebsiel a species presented in young infant epidermidis, diphtheroid, and S. aureus. with acute otitis media, but rarely appear in the middle In the present study, the number of P. aeruginosa ear effusion of older children with otitis media. isolates was (40 isolates, 19.41%). our result agreement In our work, we found that B. catarrhalis was (20 with studies done by (Aslam et al., 2004); (Verhoeff, isolates, 9.7%). Faden (1994) found that, Moraxel a 2006) that Pseudomonas most common agents in catarrhalis or B. catarrhalis were common organisms, patients with CSOM, and not approved with (Saunders et Diplococcus are considered as part of the normal flora of al., 2009) found S. epidermidis most common causative human upper respiratory tract, classified as causative agents. Aslam et al. (2004) showed that P. aeruginosa is agents to middle ear infection; it had constituted the most common isolates from infected mastoid cavity approximately 10% of al isolates. Hanan (2000) showed and chronic otitis media and the most common aerobic bacteria isolated from chronic suppurative otitis media. (cephalosporinases) may protect these bacteria and Verhoeff et al. (2006), stated that P. aeruginosa was the other type from antimicrobial agents to which the second most prevalent bacteriological agent in chronic otitis target pathogen ordinarily might be susceptible, which media, fol owed by S. aureus. Saunders et al. (2009) can be differentiated from the other Neisseriae spp by its stated that S. epidermidis species was the most lack of carbohydrate fermentation and by its DNase prevalence bacteriological agent in chronic otitis media. production. In our study, we found proteus species In this study we found that S. aureus (35 isolates, isolated (20 isolates 9.7%). Iseh and Adegbite (2004) 16.99%) fol owed P. aeruginosa in their incidence, our found that proteus species (12.8) isolated from 41 result agree with study done by (Aslam et al., 2006), patients with acute suppurative otitis media. Vaishnav while against the study done by (Saunders et al., 2009). and changani (1981), found that Proteus species with Saunders et al. (2009) found that S. epidermidis (6%) highest incidence (44%) of isolates from 100 cases with was the most common bacteria isolated from patients In our result, we found that H. influenzae was (20 Topical treatment is better than systemic therapy; this is isolates 9.7%), while S. pneumonia (15 isolates, 7.28%). probably because a higher local concentration of anti- Yamanaka et al. (2008) showed that H. influenzae and biotic is achieved. Macfadyen et al. (2006) the antibiotic Streptococcus pneumonia are the most prevalent should have activity against gram negative bacteria, organisms responsible for acute otitis media. However, especial y Pseudomonas, and gram positive bacteria, most studies from different parts of Africa suggest various especial y S. aureus. The amino glycosides and the bacterial pathogens as accusatives agents. Hence, S. fluoroquinolones, both of them meet these criteria but the aureus and S. pyogenes appear to be the most dominant former may be ototoxic, failures of the antibiotic are causative organisms among Africans Hussain et al. usual y due to failure to penetration of the debris rather (1991). Bluestone and Klein (2001) found that S. than bacterial resistance. Marais et al. (1998). Amino- pneumonia and H. influenzae are the most common glycosides are contraindicated; there is evidence that bacteria species causing middle ear infection in acute they may cause hearing loss (Bance et al., 2005). otitis media. Some European studies found H. influenzae to be the most common organism fol owed by S. pneumonia and B. catarrhalis (Gray and Canter, 1997). In REFERENCE
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