Iruka N. Okeke

Antimicrobial resistance in developing countries. Part I: recent trends and current status.

The global problem of antimicrobial resistance is particularly pressing in developing countries, where the infectious disease burden is high and cost constraints prevent the widespread application of newer, more expensive agents. Gastrointestinal, respiratory, sexually transmitted, and nosocomial infections are leading causes of disease and death in the developing world, and management of all these conditions has been critically compromised by the appearance and rapid spread of resistance. In this first part of the review, we have summarised the present state of resistance in these infections from the available data. Even though surveillance of resistance in many developing countries is suboptimal, the general picture is one of accelerating rates of resistance spurred by antimicrobial misuse and shortfalls in infection control and public health. Reservoirs for resistance may be present in healthy human and animal populations. Considerable economic and health burdens emanate from bacterial resistance, and research is needed to accurately quantify the problem and propose and evaluate practicable solutions. In part II, to be published next month, we will review potential containment strategies that could address this burgeoning problem.

Non-prescription antimicrobial use worldwide: a systematic review

In much of the world antimicrobial drugs are sold without prescription or oversight by health-care professionals. The scale and effect of this practice is unknown. We systematically reviewed published works about non-prescription antimicrobials from 1970-2009, identifying 117 relevant articles. 35 community surveys from five continents showed that non-prescription use occurred worldwide and accounted for 19-100% of antimicrobial use outside of northern Europe and North America. Safety issues associated with non-prescription use included adverse drug reactions and masking of underlying infectious processes. Non-prescription use was common for non-bacterial disease, and antituberculosis drugs were available in many areas. Antimicrobial-resistant bacteria are common in communities with frequent non-prescription use. In a few settings, control efforts that included regulation decreased antimicrobial use and resistance. Non-prescription antimicrobial and antituberculosis use is common outside of North America and northern Europe and must be accounted for in public health efforts to reduce antimicrobial resistance.

Antimicrobial resistance in developing countries. Part II: strategies for containment.

The growing threat from resistant organisms calls for concerted action to prevent the emergence of new resistant strains and the spread of existing ones. Developing countries have experienced unfavourable trends in resistance-as detailed in part I, published last month--and implementation of many of the containment strategies recommended by WHO is complicated by universal, as well as developing country-specific, factors. The control of selective pressure for resistance could potentially be addressed through educational and other interventions for orthodox and unorthodox prescribers, distributors, and consumers of antimicrobials. At national levels, the implementation of drug use strategies--eg, combination therapy or cycling--may prove useful to lengthen the lifespan of existing and future agents. Programmes such as the Integrated Management of Childhood Illnesses (IMCI) and directly observed short-course therapy (DOTS) for tuberculosis are prescriber-focused and patient-focused, respectively, and have both been shown to positively influence factors that contribute to the selective pressure that affects resistance. The institution of interventions to prevent the transmission of infectious diseases could also lead to beneficial effects on the prevalence of resistance, as has vaccination against Haemophilus influenzae type B and Streptococcus pneumoniae. There has been an upsurge in the number of organisations and programmes that directly address issues of resistance, and collaboration could be one way to stem the dire trend. Additional factors such as unregulated drug availability, inadequate antimicrobial drug quality assurance, inadequate surveillance, and cultures of antimicrobial abuse must be addressed to permit a holistic strategy for resistance control.

Enteroaggregative Escherichia coli

Enteroaggregative Escherichia coli (EAEC), an increasingly recognized cause of diarrhea in children in developing countries, has been particularly associated with persistent diarrhea (more than 14 days), a major cause of illness and death. Recent outbreaks implicate EAEC as a cause of foodborne illness in industrialized countries. The pathogenesis of EAEC infection is not well understood, but a model can be proposed in which EAEC adhere to the intestinal mucosa and elaborate enterotoxins and cytotoxins, which result in secretory diarrhea and mucosal damage. EAECs ability to stimulate the release of inflammatory mediators may also play a role in intestinal illness.

Growing Problem of Multidrug-Resistant Enteric Pathogens in Africa

A disproportionate number of low-income persons are affected.

espC Pathogenicity Island of Enteropathogenic Escherichia coli Encodes an Enterotoxin

ABSTRACT At least five proteins are secreted extracellularly by enteropathogenic Escherichia coli (EPEC), a leading cause of infant diarrhea in developing countries. However only one, EspC, is known to be secreted independently of the type III secretion apparatus encoded by genes located within the 35.6-kb locus of enterocyte effacement pathogenicity island. EspC is a member of the autotransporter family of proteins, and the secreted portion of the molecule is 110 kDa. Here we determine that the espC gene is located within a second EPEC pathogenicity island at 60 min on the chromosome of E. coli. We also show that EspC is an enterotoxin, indicated by rises in short-circuit current and potential difference in rat jejunal tissue mounted in Ussing chambers. In addition, preincubation with antiserum against the homologous Pet enterotoxin of enteroaggregative E. coli eliminated EspC enterotoxin activity. Like the EAF plasmid, the espCpathogenicity island was found only in a subset of EPEC, suggesting that EspC may play a role as an accessory virulence factor in some but not all EPEC strains.

Etiology of Acute Diarrhea in Adults in Southwestern Nigeria

ABSTRACT Stool specimens from 113 adult outpatients with diarrhea in southwestern Nigeria and 63 controls were examined for bacterial and parasitic enteric pathogens. Enterohemorrhagic Escherichia coli (EHEC) (P < 0.02), enteroaggregative E. coli (EAEC) (P < 0.02), and Entamoeba histolytica (P < 0.0002) were significantly associated with diarrhea. Salmonella, Shigella, nontoxigenic Vibrio cholerae, other categories of diarrheagenic E. coli, as well as a variety of helminths were recovered more frequently from the stools of patients than from the stools of controls but did not show a significant association with disease. Multiple pathogens were recovered from 36.3% of specimens, and bloody diarrhea was commonly associated with E. histolytica and diarrheagenic E. coli infections. The majority of EHEC isolates were non-O157 strains that carried the stx2 gene. Of the 23 EHEC-infected patients, 12 (52.2%) presented during the 10th week of the study. EHEC strains isolated within this cluster were more likely to hybridize with the enterohemolysin gene probe, to be nonmotile and sorbitol positive, and to fail to agglutinate O157 antisera. Pulsed-field gel electrophoresis demonstrated that the only strains with XbaI profiles that occurred more than once were isolated during the 10th and 11th weeks of the study, suggesting an outbreak. The study has demonstrated that E. histolytica, EHEC, and EAEC are important diarrheal pathogens within the study area and that sporadic and epidemic EHEC infections occur in developing as well as developed countries. Routine surveillance for diarrheagenic E. coli, even only at the tertiary-care level, would be useful in identifying outbreaks and assist in identifying environmental reservoirs and transmission routes.

The Escherichia coli Common Pilus and the Bundle-Forming Pilus Act in Concert during the Formation of Localized Adherence by Enteropathogenic E. coli

Although the bundle-forming pilus (BFP) of enteropathogenic Escherichia coli (EPEC) mediates microcolony formation on epithelial cells, the adherence of BFP-deficient mutants is significantly abrogated, but the mutants are still adherent due to the presence of intimin and possibly other adhesins. In this study we investigated the contribution of the recently described E. coli common pilus (ECP) to the overall adherence properties of EPEC. We found that ECP and BFP structures can be simultaneously observed in the course (between zero time and 7 h during infection) of formation of localized adherence on cultured epithelial cells. These two pilus types colocalized at different levels of the microcolony topology, tethering the adhering bacteria. No evidence of BFP disappearance was found after prolonged infection. When expressed from a plasmid present in nonadherent E. coli HB101, ECP rendered this organism highly adherent at levels comparable to those of HB101 expressing the BFP. Purified ECP bound in a dose-dependent manner to epithelial cells, and the binding was blocked with anti-ECP antibodies, confirming that the pili possess adhesin properties. An ECP mutant showed only a modest reduction in adherence to cultured cells due to background expression levels of BFP and intimin. However, isogenic mutants not expressing EspA or BFP were significantly less adherent when the ecpA gene was also deleted. Furthermore, a DeltaespA DeltaecpA double mutant (unable to translocate Tir and to establish intimate adhesion) was at least 10-fold less adherent than the DeltaespA and DeltaecpA single mutants, even in the presence of BFP. A Delta bfp DeltaespA DeltaecpA triple mutant showed the least adherence compared to the wild type and all the isogenic mutant strains tested, suggesting that ECP plays a synergistic role in adherence. Our data indicate that ECP is an accessory factor that, in association with BFP and other adhesins, contributes to the multifactorial complex interaction of EPEC with host epithelial cells.

Cholera Outbreaks in Nigeria Are Associated with Multidrug Resistant Atypical El Tor and Non-O1/Non-O139 Vibrio cholerae

Background The current millennium has seen a steep rise in the number, size and case-fatalities of cholera outbreaks in many African countries. Over 40,000 cases of cholera were reported from Nigeria in 2010. Variants of Vibrio cholerae O1 El Tor biotype have emerged but very little is known about strains causing cholera outbreaks in West Africa, which is crucial for the implementation of interventions to control epidemic cholera. Methodology/Principal Findings V. cholerae isolates from outbreaks of acute watery diarrhea in Nigeria from December, 2009 to October, 2010 were identified by standard culture methods. Fifteen O1 and five non-O1/non-O139 strains were analyzed; PCR and sequencing targeted regions associated with virulence, resistance and biotype were performed. We also studied genetic interrelatedness among the strains by multilocus sequence analysis and pulsed-field gel electrophoresis. The antibiotic susceptibility was tested by the disk diffusion method and E-test. We found that multidrug resistant atypical El Tor strains, with reduced susceptibility to ciprofloxacin and chloramphenicol, characterized by the presence of the SXT element, and gyrA Ser83Ile/parC Ser85Leu alleles as well CTX phage and TCP cluster characterized by rstR ElTor, ctxB-7 and tcpA CIRS alleles, respectively, were largely responsible for cholera outbreaks in 2009 and 2010. We also identified and characterized a V. cholerae non-O1/non-O139 lineage from cholera-like diarrhea cases in Nigeria. Conclusions/Significance The recent Nigeria outbreaks have been determined by multidrug resistant atypical El Tor and non-O1/non-O139 V. cholerae strains, and it seems that the typical El Tor, from the beginning of seventh cholera pandemic, is no longer epidemic/endemic in this country. This scenario is similar to the East Africa, Asia and Caribbean countries. The detection of a highly virulent, antimicrobial resistant lineage in Nigeria is worrisome and points to a need for vaccine-based control of the disease. This study has also revealed the putative importance of non-O1/non-O139 V. cholerae in diarrheal disease in Nigeria.

Multi-locus sequence typing of enteroaggregative Escherichia coli isolates from Nigerian children uncovers multiple lineages

Background Enteroaggregative Escherichia coli (EAEC) are defined by their stacked-brick adherence pattern to human epithelial cells. There is no all-encompassing genetic marker for EAEC. The category is commonly implicated in diarrhea but research is hampered by perplexing heterogeneity. Methodology/Principal Findings To identify key EAEC lineages, we applied multilocus sequence typing to 126 E. coli isolates from a Nigerian case-control study that showed aggregative adherence in the HEp-2 adherence assay, and 24 other EAEC strains from diverse locations. EAEC largely belonged to the A, B1 and D phylogenetic groups and only 7 (4.6%) isolates were in the B2 cluster. As many as 96 sequence types (STs) were identified but 60 (40%) of the EAEC strains belong to or are double locus variants of STs 10, 31, and 394. The remainder did not belong to predominant complexes. The most common ST complex, with predicted ancestor ST10, included 32 (21.3%) of the isolates. Significant age-related distribution suggests that weaned children in Nigeria are at risk for diarrhea from of ST10-complex EAEC. Phylogenetic group D EAEC strains, predominantly from ST31- and ST394 complexes, represented 38 (25.3%) of all isolates, include genome-sequenced strain 042, and possessed conserved chromosomal loci. Conclusions/Significance We have developed a molecular phylogenetic framework, which demonstrates that although grouped by a shared phenotype, the category of ‘EAEC’ encompasses multiple pathogenic lineages. Principal among isolates from Nigeria were ST10-complex EAEC that were associated with diarrhea in children over one year and ECOR D strains that share horizontally acquired loci.