Nosocomial infections in post-operative wounds due to Staphylococcus aureus and Pseudomonas aeruginosa in Benue State Nigeria

Three hundred (300) post-operative wound swab specimens were aseptically collected from four hospitals and investigated. The four hospitals were Federal Medical Centre, Makurdi (FMCM), General Hospital, Gboko (GHG), General Hospital, Otukpo (GHO) and General Hospital North Bank, Makurdi (GHNBM). The swabs were cultured and organisms identified according to standard procedures. A prevalence rate of bacterial isolates (56.7%) was obtained from the post-operative wound sites investigated. Pseudomonas aeruginosa was the most encountered pathogen with 20.3% prevalence rate followed by Staphylococcus aureus (13.0%), while 8.3% accounted for co-infection of both organisms. Other organisms encountered included Klebsiella spp. (4.0%), Escherichia coli (3.3%), atypical coliform (2.7%), and Proteus spp. (2.3%). Enterococcus faecalis and Streptococcus pyogenes had the least prevalent rate of 1.3% each. Statistically, Chi square analysis showed that there was no significant difference in the number of isolates from FMCM, GHQ, GHO and GHNBM and in the occurrence of both organisms in relation to sex (p˃0.05). The incidence of P. aeruginosa was highest (38.4%) at Federal Medical Centre, Makurdi, compared with other collection points investigated while that of Staphylococcus aureus was highest (37.5%) at FMCM compared with all other collection points’ investigated. Antibiogram studies revealed that P. aeruginosa was most susceptible to levoxin to the magnitude of 98.4%. While P. aeruginosa was resistant to ampicillin, tetracycline and streptomycin, S. aureus was only resistant to tetracycline. The findings have revealed that nosocomial wound infections remain a menace in medical management of wounds.


INTRODUCTION
Nosocomial infection is an infection acquired in a hospital by a patient who was admitted for a reason other than that infection (Wenzel, 2011).These infections are acquired in the hospital but appear after discharge or still when on admission.The organisms that cause most hospital acquired infections are common in the general population and are relatively harmless.The most common are bacteria (Staphylococcus aureus, coagulase -negative Staphylococci, Enterococci, and Enterobacteria) including commensal bacteria, which are part of the normal flora, and pathogenic bacteria, which come from an exogenous source.Viruses including Hepatitis B and C, Respiratory Syncytial Virus, rotaviruses, and enteroviruses may also be transmitted nosocomially.During times of prolonged antibiotic treatment and severe immunosuppression, fungi such as Candida albicans, Aspergillus spp., Cryptococcus neoformans and Cryptosporidium including other opportunistic organisms can cause infections (Shittu et al., 2012).The organisms can be transferred from one patient to another (crossinfection).They can be part of a patient's own flora (endogenous infection), or they can be transferred from an inanimate object or from a substance recently contaminated by another human source (environmental transfer).Factors that increase a patient's susceptibility to nosocomial infections include age (e.g. the elderly), decreased immunity, underlying disease, therapeutic and diagnostic interventions (Mangram et al., 2011).
A number of studies in Nigeria have shown that nosocomial infections in post-operative wounds are endemic in parts of the country (Shittu et al., 2012;Kolmos et al., 2013).Akinjogula et al. (2010) reported that S. aureus was the leading etiologic agent of postoperative wound infection in Calabar and Uyo cities of Nigeria.
In a similar study in Benin City Nigeria, it showed that Proteus species were the leading etiologic agents in postoperative wound infections and P. aeruginosa was the prevalent agent in parts of South Eastern Nigeria (Shittu et al, 2012).Haghi et al. (2010) reported that S. aureus was the leading etiologic agent of post-operative wound infections in India, Thailand and Japan.They also found out that P. aeruginosa was more prevalent among microorganisms isolated from post-operative wounds in some parts of Jordan.
Data collected from this work will be used to establish the sanitary condition of hospitals where surgical operations are carried out.It will also establish the prevalent microorganisms involved in nosocomial infections.Antibiotic susceptibility test carried out will determine the drug of choice in the treatment of postoperative wound infections.In addition, the knowledge of these infections will help physicians to give adequate treatment when such infections occur and also advise on its prevention.
This study has become necessary to ascertain bacteria implicated in wound infections which delay the normal healing process.

Study population and area
Patients with post-operative wounds infections were targeted for this study.Three hundred (300) post-operative wounds swabs were collected from this population which comprised ninety-nine (99) from FMCM, seventy from GHG, sixty eight (68) from GHO, and sixty three (63) from GHNBM.Approval was obtained from ethical clearance committees and the Chief Medical Director of each hospital for all the samples used for the study.Confidentiality was maintained in accordance with standard medical practice.

Sample collection and processing
Sterile swab sticks were used to collect pus from the surgical sites of subjects under aseptic conditions.The samples were properly labeled and immediately conveyed to the laboratory for processing.Standard microbiological procedures for handling and transporting of specimens as enunciated by Cheesbrough (2002) were followed.

Cultivation, isolation and identification
All the swabs collected for bacteriological investigations were treated according to the methods of Isenberg et al. (2011).MacConkey, blood and chocolate agars (Oxoid, England) were prepared following the manufacturer's instructions and allowed to solidify.The samples were inoculated onto the agar plates and incubated at 37C for 24 h.Incubation period was extended to 48 h if there was no bacterial growth within 24 h.Investigations such as characteristics, Gram stain and biochemical reactions of the organisms were carried out in line with standard operating procedures.Identification and biochemical testing of isolates were carried out following standard procedures (Cheesbrough, 2012).

Antimicrobial susceptibility test
Kirby-Bauer disc diffusion susceptibility technique as documented by Isenberg et al. (2011) was adopted for the susceptibility assay.Only P. aeruginosa and S. aureus isolates obtained were used for this assay.In this technique, a well dried agar plate was seeded with appropriate inoculum.Filter paper discs impregnated with various antibiotics were placed at specific locations on the seeded agar plate.The plates were incubated at 37°C for 18 hrafter which susceptibility to antimicrobial agents was measured in millimeter as zones of inhibition, around the antibiotic discs.

RESULTS
With respect to all the hospitals where samples were processed, 170 (56.7%) bacteria were isolated, with P. aeruginosa having the highest 61 (20.3%) followed by S. aureus 39(13.0%),E. faecalis and S. pyogenes the least *Corresponding author.E-mail: unata71@yahoo.com.
Author(s) agree that this article remain permanently open access under the terms of the Creative Commons Attribution License 4.0International License  1).Table 2 reveals that appendicetomy has 12and 8% of P. aeruginosa and S. aureus respectively according to type of surgery while Prostatectomy the least with prevalence of 0.3% for both pseudomonas aeruginosa and S. aureus.Coinfection was highest in appendicetomy (3.7%) and lowest in amputation (0.3%), mastectomy (0.3%) and prostatectomy (0.3%) (Table 3).
Among the health facilities investigated, FMC has the highest incidence of P. aeruginosa (11.0%) with GHO the least (4.7%) (Table 4).Similarly, S. aureus has 8.0% prevalence in FMCM while both GHO and GHNBM were lowest (4.0%).The rate of co infection in the various hospitals was more at FMCM (3.0%) with General hospitals Otukpo coming last (1.3%)(Table 5).
Table 6 indicated the number of males and females infected with P. aeruginosa as 15.7 and 13.0% respectively.In contrast, the rates of S. aureus infection in males were 9.7% and females 11.6%.Table 7 shows the distribution of P. aeruginosa and S. aureus infections according to age group with both organisms showing higher occurrence of infections in the young people compare to the elderly.Tables 8 and 9 also show the occurrence of co-infections of both organisms in age and sex of patients respectively with female having a higher prevalence of 4.6% and male 3.7%.

DISCUSSION
The results of this study show that the prevalence of P. aeruginosa (20.3%) and S. aureus (13.0%) postoperative wound infections differed.This finding agrees      The microbial analysis revealed that P. aeruginosa and S. aureus were the leading etiologic agents of postoperative infection in this study.Similar results were obtained by et al. (1992) in Bombay town of India, Konno (2011) and Akinjogunla et al. (2010).The virulence of the microorganisms may be responsible for their high infection rates as suggested by Coffin et al. (2011).
The rate of P. aeruginosa and S. aureus were higher at Federal Medical Centre Makurdi than in other hospitals.In Federal Medical Centre, patients on admission stay long in the overcrowded wards, and are therefore exposed to cross infections The prevalent rate of P. aeruginosa was higher than S. aureus in all the hospitals and this finding agrees with  Joshi et al. (2011) andCheadle W (2010).In another study by Prinsloo et al. (2010) and Burke (2012), P. aeruginosa was reported to be responsible for most nosocomial infections.This could be as a result of its ability to grow in disinfectants, sinks, water and other materials in the hospitals.It is also possible that patients may have developed immunity to S. aureus infection but this assumption contradicts the report of Johnson et al. (2013) where patients were more infected with S. aureus than P. aerginosa infection.
Incidence of P. aeruginosa was higher in males (except in General hospital Gboko) than females who were more infected mostly with S. aureus (except in General hospital Otukpo).This result is consistent with the reports of Kolmos et al. (2013) and Dulworth and Pyenson (2012), but contrary to that of Church et al. (2010) in which females were more infected with P. aeruginosa.It is possible that there are differences in hygienic practice of both males and females including the hospitals environment.
Patients within the age groups 10-19 years, 50-59 years and ≥60 years are at the highest risk of infections.Maltezou et al. (2012) in southern Uganda had reported that the age groups 10-19 years and ≥50 years were the most infected.According to Joshi et al. (2011) in south east Nigeria, children less than 13 years old were infected with post-operative wound infections whcich is also common in other parts of the world.Our results also agree with findings of Dantas et al. (2013) that infections were more common among the young and debilitated elderly people.From this study also, the age group 30-39 years had the least rate of infection in most of the hospitals.This may be due to good hygienic practices and avoidance of cross-infections exhibited by these groups of patients.
The rate of co-infection of P. aeruginosa and S. aureus according to health facility, gender and age differs but was not statistically significant.This result implies that coinfection is not influenced by these factors.
Chi square analysis at 99% confidence limit did not show any significant difference in the number of organisms isolated from the four hospitals.
The susceptibility rate of P. aeruginosa and S. aureus isolates to the eight antibiotics tested in vitro were relatively low compared to the sensitivity pattern to different anti pseudomonal and staphylococcal drugs reported worldwide (Haghi et al., 2010).In this study, P. aeruginosa isolates was highly susceptible to levoxin (97.7%) followed by ciprocin (81.4%) and norbactin (70.9%) (Table 11), while S.aureus isolates was also highly susceptible to levoxin (98.4%), ciprocin (93.8%), and norbactin (81.3%) (Table 12).Other drugs showed very low percentage of susceptibility.The non-hygienic measures in hospitals, the ability of some bacteria to grow in hospital materials or indiscriminate use of antibiotics, fake drugs, and self-prescription among patients are favourable conditions which overtime encourages the development of antibiotic resistant bacteria.The isolates were completely resistant to three of the antibiotics (ampicillin, tetracycline and streptomycin) tested in vitro, which is much higher compared to a Belgian study (Prinsloo et al., 2010) but lower than the Turkish study where one third of the isolates were multidrug resistant.This could be due to misuse of these drugs without running sensitivity tests thereby resulting to development of resistant organisms.
The prevalence and sensitivity of P. aeruginosa and S. aureus often varies between communities, hospitals in the same community and among different patient populations in the same hospital (Kolmos et al., 2013).Faced with these variations, the physician in clinical practice has the responsibility of making clinical judgments, and should have access to recent data on the prevalence and antimicrobial resistance pattern of commonly encountered pathogens.It is therefore important to institute a system for the surveillance of antimicrobial resistance that will involve the clinical collection of The concentration of each antibiotic was 10µg.The number of isolates tested against each antibiotic was 86.The concentration of each antibiotic was 10 µg.The number of isolates tested against each antibiotic was 64.
microbiological data.Shittu et al. (2012) found that patients were the sources of bacteria in all cases of wound infection and that increase in post-operative infections was due to high penicillin resistant carrier rate in hospital personnel and patients as a result of widespread use of Penicillin.However, Kolmos et al. (2013) reported that cleaners and patients were the major source of wound contamination.
The high incidence of P. aeruginosa and S. aureus may be related to indiscriminate use of antibiotics without laboratory diagnosis and antibiotic sensitivity report.This single factor could eliminate the normal flora and provide a non-competitive environment for P. aeruginosa and Staphylococcus aureus to occur.The resistance of the organism to antimicrobial agents, nutritional versatility and the difficulties encountered in maintaining proper hygienic standards especially among personnel involved in wound dressing and general care of patients may have contributed to the high rate of P. aeruginosa and S. aureus infections.

Table 1 .
Prevalence of bacterial isolates in 300 post-operative wounds examined in Benue State.

Table 2 .
Prevalence of pseudomonas aeruginosa and staphylococcus aureus in wounds according to type of surgical operation.

Table 3 .
Co-Infections of Pseudomonas aeruginosa and Staphylococcus aureus in different types of surgical operation.

Table 4 .
Prevalence of Pseudomonas aeruginosa and Staphylococcus aureus in relation to health facilities.

Table 5 .
Prevalence OF Pseudomonas aeruginosa and Staphylococcus aureus CO-infection in relation to health facilities.

Table 6 .
Prevalence of pseudomonas aeruginosa and Staphylococcus aureus according to sex of patients.

Table 7 .
Prevalence of Pseudomonas aeruginosa and Staphylococcus aureus infections according to age group.

Table 8 .
Co-infection of Pseudomonas aeruginosa and Staphylococcus aureus among age group.

Table 9 .
Co-infection of Pseudomonas aeruginosa and Staphylococcus aureus in relation to sex of patients.
obtained increasing prevalence of P. aeruginosa and S. aureus in post-operative wound infections especially in recent years.It is thus clear that the prevalence of P. aeruginosa and S. aureus obtained in this study is in agreement with what is obtained in other hospitals in Nigeria.

Table 10 .
Distribution of Pseudomonas aeruginosa and Staphylococcus aureus in relation to age of patients in various hospitals.Federal Medical Centre Makurdi; GHG = General Hospital Gboko; GHO = General Hospital Otukpo; GHNBM= General Hospital North Bank Makurdi.P.aeru =Pseudomonas aeruginosa; S.aur = Staphylococcus aureus; No. = Number; exam.= examined the reports of

Table 11 .
Susceptibility pattern of Pseudomonas aeruginosa isolates to common antibiotics.

Table 12 .
Susceptibility pattern of staphylococcus aureus isolates to common antibiotics.