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RESEARCH
Extremely high prevalence of multi-resistance among
uropathogens from hospitalised children in Beira,
Mozambique
B T van der Meeren,1 MSc; K D Chhaganlal,2 MD; A Pfeiffer,3 MD; E Gomez,2 MD; J J Ferro,3 MD; M Hilbink,4 PhD; C Macome,2 MD;
F J van der Vondervoort,5 BSc; K Steidel,3 MD; P C Wever,5 MD, PhD
1 Universidade Católica de Moçambique, Beira, Mozambique, and Department of Medical Microbiology and Infection Control, Jeroen Bosch Hospital, ’s-Hertogenbosch, The Netherlands 2 Hospital Central da Beira, Beira, Mozambique3 Universidade Católica de Moçambique, Beira, Mozambique4 Jeroen Bosch Academy, Jeroen Bosch Hospital, ’s-Hertogenbosch, The Netherlands5 Department of Medical Microbiology and Infection Control, Jeroen Bosch Hospital, ’s-Hertogenbosch, The Netherlands Corresponding author: B van der Meer
Objectives. A prospective surveil ance study was conducted to investigate the epidemiology and patterns of antibiotic resistance among
uropathogens from hospitalised children in Beira, Mozambique. Additional y, information regarding determinants of a urinary tract Methods. Bacterial species identification, antimicrobial susceptibility testing and extended-spectrum beta-lactamase testing were
performed for relevant bacterial isolates.
Results. Analysis of 170 urine samples from 148 children yielded 34 bacterial isolates, predominantly Escherichia coli and Klebsiella spp.,
causative of a urinary tract infection in 29 children; 30/34 isolates (88.2%) from 26/29 children (89.7%) were considered highly resistant micro-organisms (HRMOs). No significant determinants of urinary tract infection with HRMOs were detected when analysing gender, antibiotic use during hospital admission and HIV status.
Conclusion. This study shows, for the first time in Mozambique, an extremely high prevalence of HRMOs among uropathogens from
hospitalised children with a urinary tract infection.
S Afr Med J 2013;103(6):382-386. DOI:10.7196/SAMJ.5941 Bacteria are champions of evolution, and microbes Urinary tract infection (UTI) is one of the most common bacterial have adapted to a point where they pose serious clinical infections encountered by clinicians in developing countries and challenges for humans.[1] Emergence of antimicrobial the cause of significant morbidity and mortality.[8] Several studies resistance among bacteria is a worldwide problem. from the African continent have investigated the profile of common New mechanisms of bacterial resistance that have uropathogens and the pattern of their susceptibility to commonly used been detected include penicillin-binding protein defects, extended- antimicrobial agents in order to guide choice of empiric therapy. These spectrum beta-lactamases (ESBLs) and metallo-beta-lactamases. [2,3] studies reported the emergence of antibiotic-resistant Gram-negative Guidelines for first-line antibiotics in the treatment of different bacilli with special emphasis on ESBL-producing isolates. [9-11] infectious diseases have an empiric basis, but recommended To our knowledge, no studies have been published describing antibiotics are not always as successful as they are expected to antimicrobial resistance among uropathogens in Mozambique. A be. Collection of microbial data on a case-by-case basis is often small pilot study of the antimicrobial susceptibility pattern of necessary in choosing effective individual antimicrobial therapy, but common uropathogens conducted at the Universidade Católica de scarcity of microbiology laboratory capacity in developing countries Moçambique (UCM) showed results warranting further investigation. limits this approach. In these countries, the best option would be to A prospective surveil ance study was therefore conducted with the base antimicrobial therapy on local data, if available, regarding the aim of investigating the epidemiology and patterns of antibiotic susceptibility of common pathogens to antibiotics.[4] resistance of uropathogens from hospitalised children at the Hospital Mozambique, one of the poorest countries of the world, has a shortage of microbiology laboratory facilities, and choice of antimicrobial therapy often is guided solely by availability of antibiotics. Few studies Materials and methods
have addressed the emergence of antimicrobial-resistant bacteria in Study population
Mozambique. Increasing resistance to chloramphenicol among blood The study was conducted from 8 October to 10 December 2009 in culture isolates of non-typhi Salmonel a species, Escherichia coli, the two paediatric wards of HCB, a 728-bed referral hospital in Beira, Staphylococcus aureus and Haemophilus influenzae has been reported.[5] the second-largest city in Mozambique. A maximum of 150 and 60 Likewise, high resistance to tetracycline was reported among Neisseria children are treated in the 100-bed paediatric ward and the 24-bed gonorrhoeae isolates, while diarrhoeagenic E. coli isolates showed high malnutrition ward, respectively; one bed is therefore often occupied resistance to trimethoprim-sulphamethoxazole and ampicillin.[6,7] by more than one child. One family member is allowed to stay in the 382 June 2013, Vol. 103, No. 6 SAMJ
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hospital to take care of the child. For each child included in the study, status (determinants) on the one hand and UTI with HRMOs information regarding age, gender, antibiotic use during hospital (outcome variable) on the other hand. Two analyses were performed admission and HIV status was recorded on a data information sheet. for these relationships: a crude analysis and an adjusted analysis. A The study was approved by the local medical ethical committee.
p-value of <0.05 (two-tailed) was considered to indicate statistical significance. Statistical analyses were performed using PASW 18.0 for Microbiological data
Windows (SPSS Inc., Chicago, IL, USA).
Laboratory investigations were all performed locally in UCM facilities in Beira. Nurses were instructed to provide midstream Results
clean-catch urine specimens, and to use urine collection bags Malnutrition ward
in the case of children who were not toilet trained. Suprapubic From the malnutrition ward, 88 urine samples were received from 76 bladder aspiration was considered too invasive. All urine samples children (47 males and 29 females; 61.8% and 38.2%). The median age from the paediatric and malnutrition wards were collected in was 2 years (range 3 months - 13 years). HIV status was documented sterile containers and analysed within the hour using a commercial in 62 of 76 children, of whom 29 were HIV-positive (46.8%; 18 males urine dipstick test (Combur 8 Test, Boehringer Mannheim GmbH, and 11 females). Table 1 shows the antibiotics used during hospital Mannheim, Germany). In the paediatric ward, urine analysis was stay of 75 of the 76 children admitted to the malnutrition ward.
requested if UTI was suspected on clinical grounds, whereas it was Dipstick analysis and Gram staining, the latter performed on 11 routinely performed in the malnutrition ward, typical y within a few samples, identified 56 urine samples for subsequent culture from days after admission. Urine samples for which leucocyte esterase 46 children (26 males and 20 females). Culture yielded 19 bacterial and/or nitrite tests were positive were inoculated on blood agar isolates causative of UTI from 16 urine samples from 16 children plates (Becton, Dickinson & Co., Sparks, MD, USA) and eosin (21.1% of all studied children; 10 males and 6 females). Cultured methylene blue agar plates (Becton, Dickinson & Co.), followed micro-organisms included E. coli (n=11) and Klebsiella spp. (n=8). by incubation for 18 - 24 hours at 35°C. The same process was Table 2 shows the AST results for these two pathogens.
undertaken when leucocyte esterase and nitrite tests were negative on dipstick analysis of turbid urine samples but Gram staining, Paediatric ward
only performed in these cases, showed leucocytes and/or bacteria. From the paediatric ward, 82 urine samples were received from Bacterial growth was evaluated according to American Society 72 children (41 males and 30 females; 57.7% and 42.3%; gender for Microbiology guidelines.[12] Relevant bacterial isolates were unknown for 1 child). The median age was 5 years (range 6 months - identified using commercial identification strips (API 20E and API 12 years). HIV status was documented in 70 of 72 children, of whom 20NE systems, bioMérieux, Marcy l’Etoile, France). Antimicrobial 14 were HIV-positive (20.0%; 8 males and 6 females). Table 1 shows susceptibility testing (AST) was performed by disc diffusion on the use of antibiotics during hospital stay of 72 children admitted to Mueller-Hinton II agar plates (Becton, Dickinson & Co.) with an the paediatric ward.
inoculum of 0.5 McFarland, using Neo-Sensitabs tablets (Rosco Dipstick analysis and Gram staining, the latter performed on 6 Diagnostica, Taastrup, Denmark). Inhibition zones were interpreted samples, identified 46 urine samples for subsequent culture from according to Clinical and Laboratory Standard Institute guidelines. 44 children (19 males and 24 females; gender unknown for 1 child). For Gram-negative isolates, the following Neo-Sensitabs tablets Culture yielded 15 bacterial isolates causative of a UTI from 13 urine were examined: amoxicillin 30 µg, piperacillin 100 µg, amoxicillin/ samples from 13 children (18.1% of studied individuals; 8 males and clavulanate 30/15 µg, piperacillin/tazobactam 100/10 µg, cefazolin 4 females; gender unknown for 1 child). Cultured micro-organisms 60 µg, cefuroxime 60 µg, cefotaxime 30 µg, cefotaxime/clavulanate included E. coli (n=3), Klebsiella spp. (n=9), Proteus mirabilis (n=1), 30/10 µg, ceftazidime 30 µg, ceftazidime/clavulanate 30/10 µg, Citrobacter freundii (n=1) and Enterococcus spp. (n=1). Table 2 shows cefepime 30 µg, cefepime/clavulanate 30/10 µg, imipenem 15 µg, the AST results for the two predominant pathogens, E. coli and amikacin 40 µg, kanamycin 100 µg, gentamicin 40 µg, nalidixic Klebsiella spp.
acid 130 µg, ciprofloxacin 10 µg, nitrofurantoin 260 µg, fosfomycin 70 µg, trimethoprim/sulfamethoxazole 5.2/240 µg, chloramphenicol Table 1. Use of antibiotics during hospital stay of children
60 µg, doxycycline 80 µg and tetracycline 80 µg. The following admitted to the malnutrition and paediatric wards
Neo-Sensitabs tablets were examined for Gram-positive isolates: amoxicillin 30 µg, amoxicillin/clavulanate 30/15 µg, cefoxitin 60 µg, Malnutrition ward Paediatric ward
ciprofloxacin10 µg, nitrofurantoin 260 µg, chloramphenicol 60 µg, (N=75)*
(N=72)
doxycycline 80 µg, clindamycin 25 µg, erythromycin 78 µg and n (%)
n (%)
vancomycin 5 µg. According to current Dutch guidelines, strains producing ESBL were defined as strains showing an increase of 5 mm or more in inhibition zones with cefotaxime/clavulanate and/ or ceftazidime/clavulanate and/or cefepime/clavulanate, compared with zones with their clavulanate-free counterparts. An ESBL-positive isolate was considered resistant to all penicillins and cephalosporins tested.[13] The guideline Measures to Prevent Transmission of Highly Resistant Microorganisms (HRMOs), published in 2005 by the Dutch Working Party on Infection Prevention, was used to identify Statistical analysis
Logistic regression was performed to investigate the relationships between gender, antibiotic use during hospital admission and HIV *Antibiotic use was not documented in 1 of 76 children admitted to the malnutrition ward.
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Table 2. Resistance to antibiotics* among Escherichia coli and Klebsiella spp. isolates from urine samples of children admitted to the
malnutrition and paediatric wards
Malnutrition ward
Paediatric ward
E. coli (N=11)
Klebsiella spp. (N=8)
E. coli (N=3)
Klebsiella spp. (N=9)
n (%)
n (%)
n (%)
n (%)
Combined aminoglycoside/quinolone resistance 9 (81.8) ESBL = extended-spectrum beta-lactamase; HRMO = highly resistant micro-organism.
*Resistance to a single antibiotic, ESBL production, combined resistance to aminoglycosides and quinolones, or fulfilment of the definition of HRMO.
Highly resistant micro-organisms
Determinants of urinary tract infection with highly
Overal , analysis of 170 urine samples from 148 children yielded resistant micro-organisms
34 bacterial isolates, predominantly E. coli and Klebsiella spp., In the crude analysis, gender, which was known for 147 of 148 causative of a UTI in 29 children. According to Dutch guidelines, children, was not associated with UTI with HRMOs (odds ratio (OR) 30/34 isolates (88.2%) from 26/29 children (89.7%) were considered 0.66; 95% confidence interval (CI) 0.26 - 1.63; p=0.37). Information HRMOs on the basis of ESBL production (n=26), combined on antibiotic use during hospital admission was obtained for 147 resistance to aminoglycosides and quinolones in E. coli and of 148 children. In children receiving either 0 - 2 (n=119) or 3 - 4 Klebsiella spp. (n=17), and combined resistance to aminoglycosides, (n=28) different antibiotics during admission, UTI with HRMOs quinolones and trimethoprim/sulfamethoxazole in P. mirabilis and was found in 19 (16.0%) and 7 (25.0%) children, respectively. Children receiving 3 - 4 different antibiotics during admission did 384 June 2013, Vol. 103, No. 6 SAMJ
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not have a significantly higher risk of UTI with HRMOs (OR 1.75; the malnutrition ward. This is in accordance with World Health 95% CI 0.65 - 4.70; p=0.26) compared with children receiving 0 - 2 Organization recommendations[20] in which oral treatment with antibiotics. HIV status was documented for 132 of 148 children. In trimethoprim/sulfamethoxazole is suggested for malnourished HIV-negative children (n=89) and HIV-positive children (n=43), children with no apparent signs of infection and no complications, UTI with HRMOs was found in 14 (15.7%) and 7 (16.3%) children, while systemic treatment with amoxicillin and gentamicin is respectively. HIV-positive children did not have a higher risk of UTI suggested for malnourished children with complications such as with HRMOs (OR 1.04; 95% CI 0.39 - 2.81; p=0.94) compared with septic shock, hypoglycaemia, hypothermia, or skin, respiratory or urinary tract infections, or for those who appear lethargic or sickly. In view of the worldwide development of antimicrobial resistance Discussion
and its relation to heavy use and misuse of antibiotics, the clinical This study shows, for the first time in Mozambique, an extremely practice of prescribing antibiotics to malnourished children without high prevalence of HRMOs in Gram-negative uropathogens from an apparent infection seems debatable, but is explained by the fact hospitalised children with a UTI. The recommended antibiotic that malnourished children with serious infections may not display for treatment of UTI in children hospitalised at HCB is currently fever and inflammation.[20] amoxicillin, which as this study shows cannot be expected to There is little in the literature on the impact of HIV on the be clinical y successful and should be reconsidered. Likewise, in presentation of UTI in children. We were unable to show an neighbouring South Africa, it has been recommended that because association between HIV status and the presence of UTI with of high resistance levels among Gram-negative uropathogens, HRMOs in hospitalised children, in keeping with previous results amoxicillin and trimethoprim/sulfamethoxazole should not be used showing no significant effect of HIV on the presentation of UTI in for empiric treatment of UTI at Dr George Mukhari Hospital in children.[21] We did not investigate the relationship between age and UTI with HRMOs because of the great variation in age in children A prevalence of 15 - 20% resistance to an antibiotic has been admitted to the malnutrition and paediatric wards.
considered an acceptable threshold for the antibiotic to be included In this study, we observed little use of third-generation in empiric coverage.[15] Suitable empiric options for treatment of cephalosporins such as ceftriaxone in either ward. Nevertheless, we UTI in hospitalised children at HCB therefore include amikacin, observed high-level resistance to third-generation cephalosporins imipenem, nitrofurantoin and fosfomycin. However, these antibiotics due to ESBL production. Currently there is great concern about are either not available in Mozambique, inappropriate because the possible relationship between heavy antibiotic use in chicken of broad-spectrum antimicrobial activity, restricted for use in farming and the presence of ESBL-producing Gram-negative bacilli uncomplicated UTI, or require blood level monitoring because of in humans.[22] Studies of antimicrobial resistance of Gram-negative side-effects. Both empiric and directed antimicrobial treatment of bacilli in poultry in Mozambique could give insight into the origin these UTIs is therefore a major clinical challenge in Mozambique. of the observed ESBL-producing strains in humans. Nosocomial It is, however, important to note that the study subjects were cross-contamination has also been reported as a major mode of hospitalised children, most of whom were receiving treatment with acquisition of multi-resistant bacteria, especial y among hospitalised multiple antibiotics. Presumably our results are not representative patients being treated with antibiotics.[23] Polymerase chain reaction of HRMOs in outpatients or the general population of the country, techniques would be required to characterise resistance genes and and it would be of value to investigate the prevalence in both these determine clonal relatedness of the isolated HRMOs in our study. groups. Such studies are, however, hampered by our lack of adequate However, whether or not clonal relatedness is apparent, the role clinical microbiology facilities. The absence of such facilities also of infection control practices in this situation cannot be over- means that we are not informed about the frequency of recurrent emphasised.
UTI in the study population. Likewise, renal imaging by ultrasound In conclusion, our results show an extremely high prevalence of to look for urinary tract anomalies is seldom performed. However, in HRMOs among uropathogens from hospitalised children with a UTI a study of malnourished black children in South Africa no anatomical abnormalities were demonstrated in a population with a high Acknowledgements. We thank the nurses of the malnutrition and
In this study, no resistance was found to amikacin despite the high paediatric wards and the laboratory technicians of the Hospital Central da level of resistance to the other aminoglycosides tested, kanamycin and gentamicin. This divergence is consistent with previously reported results from the African continent.[17] Amikacin is a derivative of kanamycin A with the amino group at position 1 acylated Funding sources. None.
by 4-amino-2-hydroxybutyrate. The high activity of amikacin in comparison with other aminoglycosides is attributed to the presence References
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