Clinical Impact of Extended-Spectrum-Lactamase-Producing Bacteria

We have been forced to fight against the newly acquired antibiotic resistance of various bacteria. By the end of the 1970s, most Escherichia coli (E. coli) and Klebsiella pneumoniae (K. pneumoniae) strains contained plasmid-mediated, ampicillin-hydrolyzing -lactamases, such as TEM-1, TEM-2, and SHV-1, and could be eliminated by the use of third-generation cephalosporins. 1 TEM-1 and TEM-2 were detected mainly in E. coli, and SHV-1 was mainly detected in K. pneumoniae. 2 The emergence of K. pneumoniae strains with a gene encoding lactamase that hydrolyzes the extended-spectrum cephalosporins was first reported by a study from Germany in 1983. 3 The gene encoding the new -lactamase harbored a singlenucleotide mutation, as compared to the parental blaSHV-1 gene. In 1986, K. pneumoniae strains resistant to the third-generation cephalosporins were detected in France. 4 The resistance was attributed to a new -lactamase gene, which was closely related to TEM-1 and TEM-2. These newly detected -lactamases capable of hydrolyzing extended-spectrum -lactam antibiotics were named extended-spectrum -lactamases (ESBLs). 5 In 1989, the CTX-M type was reported as a new ESBL family member not belonging to either the TEM or SHV types. 6 Notably, the origin of CTX-M ESBLs is totally different from that of TEM or SHV ESBL. 7 Until the end of the 1990s, most of the ESBLs detected were either the TEM or SHV types and were usually associated with nosocomial outbreaks caused by K. pneumoniae. 1 In the new millennium, the worldwide spread of CTX-M-producing E. coli has been dramatic, and they are now considered to be the primary ESBL producers that are almost always associated with community-acquired infections. 8 ESBL-producing E.coli and Klebsiella spp. are now listed as one of the six drug-resistant pathogens for which few potentially effective drugs are available. 9 This chapter will outline the genetic aspects of TEM, SHV, and CTX-M ESBLs, including molecular epidemiology and mobile elements. In addition, we will also consider the impact of their genetic evolution on clinical aspects, including mode of infection and antibiotic resistance.


Global epidemiology: dissemination of ESBLs
ESBLs were first detected in the first half of the 1980s in Europe, and they later disseminated worldwide. 1Until the 1990s, the main producer of ESBLs was K. pneumoniae and nosocomial outbreaks caused by the organism were often reported. 1The number of ESBLproducing E. coli isolates has been dramatically increasing during the 21st century. 11A recent global surveillance database collected from Europe, North and South America, and Asia, showed that the detection frequencies for ESBL-producing K. pneumoniae and E. coli isolates were 7.5-44% and 2.2-13.5%,respectively. 12The prevalence of ESBL-producing isolates increased to a greater degree, particularly in Asia than in other regions, and one study conducted in 2007 showed that the frequencies of ESBL-producing K. pneumoniae and E. coli isolates exceeded 30% in both bacterial populations. 13A recent surveillance using samples collected from nine Asian countries showed ESBL producers accounted for 42.2% of K. pneumoniae isolates detected from patients with hospital-acquired pneumoniae. 14Our data collected from one institution in Japan showed that the detection rate of the E. coli isolates increased first, followed by increased detection rates of the K. pneumoniae and P. mirabilis isolates. 15(Figure1) These data suggest that K. pneumoniae, as well as E. coli, has been an important ESBL producer even in the last few years.
In the analysis of ESBL genotypes, TEM and SHV were predominantly observed until the 1990s, and it was most reported that SHV-producing K. pneumoniae strains showed clonal dissemination in hospitals. 1Recent studies show that TEM and SHV types have been frequently detected up to the present day.Interestingly, in some cases, SHV has been found in isolates expressing other ESBL types, such as TEM and CTX-M. 16Our study showed that multiple types of ESBLs, including TEM, SHV, and CTX-M, were most frequently detected in K. pneumoniae and E. coli. 15(Table1) These findings suggest that the genetic mechanism underlying dissemination of ESBL genes has become more divergent and complicated.After the first half of the 2000s, it was often reported that the number of CTX-M ESBLs detected was on the rise, that the main carrier was E. coli, and that most of the CTX-M-producing E. coli strains were acquired in the community, not in hospitals. 11The detection rate of CTX-M ESBLs has been dramatically rising, especially in the last 5 years. 17The mechanism behind the spread of blaCTX-M genes differs from that observed in the case of blaTEM and blaSHV genes.blaTEM and blaSHV ESBL genes are associated with the dissemination of particular clones, known as an "epidemic" pattern; however, the mechanism by which blaCTX-M ESBL genes disseminate reflects the simultaneous spread of multiple specific clones, known as an "allodemic" pattern. 18It has been indicated that various CTX-M-type ESBLs have spread Fig. 1.Frequencies of ESBL-producing organisms at Hara-Sanshin Hospital in Fukuoka (Japan) worldwide, and that specific CTX-M subgroups have been characterized in different geographic areas. 15,18,19In contrast, CTX-M-15 ESBLs, which belong to the CTX-M-1 group, have been found worldwide. 18Unlike in other countries, in the USA, ESBLs were rarely detected, until the first half of the 2000s; however, CTX-M ESBLs, specifically CTX-M-15, have frequently been encountered over the last 5 years. 20,21ble 1.Genotypes of ESBL-producing Eschrichia coli isolated from hospitalized patients www.intechopen.com

Genetic characteristics
Genes harboring ESBLs are associated with several specific genetic structures.A variety of mobile genetic elements, such as transposons, insertion sequences, and integrons, play important roles in the dissemination of ESBL genes.blaTEM-type ESBL genes are acquired by the mutation of plasmid-mediated, parent blaTEM-1 and blaTEM-2 genes, and the main producer of TEM-type ESBLs is E. coli; these genes occur within the earliest bacterial transposons identified. 2,22blaSHV-type ESBL genes are the derivatives of chromosomal, parent blaSHV-1 genes, which occur mainly in K. pneumoniae, 23 and are likely acquired by the role of insertion sequences from chromosome to plasmid. 22Notably, blaTEM-type and blaSHV-type ESBL genes located in the integron structures have never been identified. 22The spread of blaCTX-M-type ESBL genes is associated with more complicated mobile elements, compared to that of blaTEM and blaSHV ESBL genes.blaCTX-M ESBL genes are not derivatives of K. pneumoniae or E. coli that contain original genes, as compared to blaTEM or blaSHV ESBL genes.blaCTX-M genes originate from the chromosomal -lactamase genes of Kluyvera species, which are environmental bacteria found worldwide, and are captured mainly by insertion sequence elements translocated from chromosome to plasmid. 18Original lactamase genes of Kluyvera species are identified in most CTX-M subgroups. 18This differential origin might be involved in the characteristic spread of blaCTX-M ESBL genes, that is, an "allodemic" pattern of spread.All blaCTX-M genes are associated with insertion sequences.Well-studied, CTX-M-associated insertion sequence elements include ISEcp1 and ISCR1, which are involved in the mobilization of blaCTX-M genes by a transposition mechanism. 24,25,26In addition, integron structures bearing insertion sequences and blaCTX-M genes can be linked to transposon elements, such as from the Tn21 family, which has been intensively studied.Transposons of the Tn21 family are disseminated worldwide in both environmental and clinical bacteria. 18,24These highly efficient mobile genetic elements may have influenced the rapid and easy dissemination of blaCTX-M ESBL genes.
An antibiotic resistance plasmid itself is responsible for the efficiency of gene transfer, as well as the mobile genetic elements described above.It has been shown that ESBL genebearing plasmids can be transferred to different bacterial species by conjugation. 27,28revious studies have shown that blaTEM and blaSHV ESBL genes are associated with plasmids belonging to a few specific incompatibility (Inc) groups. 18In contrast, blaCTX-M ESBL genes are carried by plasmids belonging to a variety of Inc groups including narrowand broad-host-range types. 18,29blaCTX-M-15 genes are located mainly on plasmids belonging to the IncF group. 29Interestingly, a recent study has described the diversity of ESBL genebearing plasmids, including SHV types. 30It was reported that a mosaic plasmid has been identified from a clonal CTX-M-producing E. coli isolate, suggesting genetic interactions among different plasmids. 31In ESBL gene-bearing plasmids, the genetic diversity has been constantly increasing through the mechanism of gene transfer and gene shuffling.

Nosocomial infection
Up to the end of the 1990s, clinical infections caused by ESBL-producing bacteria were associated with nosocomial outbreaks, where the chief ESBL producer was K. pneumoniae, but not E. coli.In addition, the ESBL genotypes detected in the nosocomial setting were almost always TEM and SHV, but not CTX-M types. 1 SHV-producing K. pneumoniae strains were intensively examined in the analysis of clonal dissemination in hospitals.Clonally related SHV-4-producing K. pneumoniae isolates were shown to have spread to multiple hospitals within the specific region. 32This phenomenon indicates that, at that time, the mode of spread for SHV-producing K. pneumoniae was the dissemination of particular clones, that is to say, an "epidemic" pattern.The number of nosocomial outbreaks caused by TEM-or SHV-producing K. pneumoniae strains have been decreased during this century; however, as observed in many studies, these organisms are frequently identified in many hospitals worldwide.Interestingly, the derivatives of TEM and SHV types have been reported to be more divergent in K. pneumoniae strains isolated in European hospitals. 16,33oreover, the new variants of blaSHV genes were detected from an Algerian hospital. 34

Community-acquired infection
The mode of ESBL-related infection has dramatically changed since the 2000s.Communityacquired infections caused by ESBL-producing bacteria have been increasingly documented. 8CTX-M-producing E. coli strains are chiefly responsible for community-acquired infections, which are related to an increase in the number of ESBL carriers in the general population. 11,35One report describes a significant increase in the prevalence of ESBL carriers in a specific population from 2001 to 2006. 36The interfamilial dissemination of ESBLproducing bacteria has also been suggested. 37Notably, animals used as food and or pets are reported to carry CTX-M ESBLs, 38,39,40,41 and this finding may explain the dramatic spread of ESBLs in the community.The community-onset dissemination of ESBLs in both humans and animals may suggest that blaCTX-M ESBL genes detected in pathogenic bacteria are acquired from environmental bacteria.Branger et al showed that many CTX-M ESBLs were associated with the phylogenetic group D2 that lacked a virulence factor. 42,43 he specific features may be related to the colonization and spread among the general population.The spread of ESBLs in the community is linked to the emergence of ESBL-related infections in outpatients, in whom urinary tract infections are most often reported along with bacteremia.44,45, 46 One study has described the detection of CTX-M-producing K. pneumoniae in outpatients, 33 A nosocomial outbreak was caused by CTX-M-producing K. pneumoniae isolates from foods, suggesting the influx of ESBL-producing K. pneumoniae into a hospital. 47hese reports may account for the dissemination of CTX-M ESBL genes from E. coli to other bacteria in the community.

Antibiotic resistance
Antibiotic resistance is of utmost importance for the clinical impact of ESBL-producing bacteria.A meta-analysis showed increased mortality and delay in effective antibiotic use in ESBL-related bacteremia, 48 indicating the importance of constant surveillance for an antibiotic resistance pattern in organisms with ESBLs.ESBL-producing bacteria are resistant to almost all -lactam antibiotics, except carbapenems, as indicated by their definition.In addition, most ESBL-producing bacteria, particularly those with the TEM, SHV, and CTX-M genotypes, exhibit co-resistance to aminoglycosides, tetracyclines, and sulfonamides. 18rganisms with CTX-M genotypes, such as those with CTX-M-9, -14, and -15, are reported to be resistant to fluoroquinolones. 18This additional resistance is induced by the main mechanism that blaCTX-M genes are directly linked to quinolone resistance genes, qnr genes.ISCR1, a mobile element for blaCTX-M genes, is associated with qnr genes, 26 indicating an effective transfer of quinolone resistance genes together with blaCTX-M genes.This genetic finding is interesting for clinical reasons.Selective pressure by the use of fluoroquinolones may induce the emergence of CTX-M ESBL-producing bacteria.As a consequence, the therapeutic options for infections caused by ESBL-producing bacteria may be more limited.Tigecycline has been shown to be microbiologically active against ESBLproducing E. coli and K. pneumoniae, 49,50 whereas, fosfomycin has been reported to be effective against urinary tract infections caused by ESBL-producing E. coli. 51,52

Spread of CTX-M-15-producing ST131 E. coli clones
The dissemination of CTX-M-15 producing E. coli strains has become a major concern of research in antibiotic resistance.The first isolation of CTX-M-15-type ESBLs was reported in India in 2001. 53CTX-M-15 is derived from CTX-M-3, belonging to the CTX-M-1 group, differing by one amino acid substitution.blaCTX-M-15 genes are transferred mainly by the IncF group plasmids, which are well adapted to E. coli and have acquired many antibiotic resistance genes. 54,55,56Recently, Mnif et al reported that the IncF plasmids carrying blaCTX-M-15 genes contained many addiction systems, which could contribute to their maintenance in E. coli host strains. 57The detection rate of the CTX-M-15 producing E. coli strains with multidrug resistance has been dramatically increasing worldwide since the 2000s. 56This CTX-M-15-producing E. coli strain is often thought to be associated with ST131 clones. 56ost of the CTX-M-15-producing E. coli strains isolated from three continents were O25:H4-ST131 clones that show highly similar PFGE profiles, suggesting a recent emergence of these clones. 58The emergence of the CTX-M-15-producing ST131 E. coli clones is highly related to the recent dissemination of ESBLs in the USA. 59,60 he worldwide spread of the multi-drugresistant ST131 E. coli clones can be explained by the acquisition of IncFII plasmids harboring blaCTX-M-15 genes and many other antibiotic resistance genes.Interestingly, these ST131 E. coli clones belong to the highly virulent, phylogenetic group B2. 56Over the past 5 years, CTX-M-15-producing ST131 E. coli clones have become an important causative agent for community-acquired ESBL infections, mainly urinary tract infections and bacteremia. 45

Clinical impact on immunodeficient patients
The sufficient therapy for ESBL-related infections is important, especially in immunodeficient patients.One study has shown that approximately 13% of E. coli-related bacteremia cases detected in patients with cancer and neutropenia were caused by ESBLs, that CTX-M types were predominant among the ESBLs, and that the bacteremia induced by ESBL-producing E. coli strains was linked to inadequate empirical antibiotic therapy. 61In our institution, the detection rate of ESBL-related bacteremia has been increasing in febrile neutropenic patients with hematological malignancies, and consequently, we have been forced to use carbapenems for the therapy. 62,63In immunodeficient patients, such as those undergoing chemotherapy, serious ESBL-related infections may result in a poor prognosis owing to the failure of the initial therapy.Recently, M. D. Anderson Cancer Center has reported an interesting finding that pyomyositis was caused by ESBL-producing E. coli strains in neutropenic patients with hematological malignancies, and that the E. coli strains were ST131 clones belonging to phylogenetic group B2. 64This notable finding implies that ESBL-producing ST131 E. coli clones cause fatal damage in the case of immunodeficient patients because of their high virulence.

Conclusions
The spread of ESBL-producing bacteria in the community has begun influencing outpatient therapy.Community-acquired bacteremia, due to ESBL-producing E. coli strains, is becoming a critical concern for outpatients, because inappropriate use of empirical antibiotics, such as cephalosporins and fluoroquinolones, has resulted in high mortality. 65,66ne study has shown that the resistance of CTX-M-15-producing ST131 E. coli strains isolated from the community to fosfomycin has increased. 67In the near future, we may be forced to use carbapenems as the first choice for the empirical therapy of patients with community-acquired infections due to ESBL-producing bacteria.The identification of carbapenemase-producing E coli and K. pneumoniae strains has been frequently documented as evidence for additional -lactamases-producing bacteria other than the ESBL-producing bacteria. 68The study of NDM-1-type carbapenemase-producing E coli and K. pneumoniae is currently a topic of much interest in multidrug-resistant bacteria research.Notably, some of the NDM-1-type-producing E coli and K. pneumoniae strains express blaCTX-M-15 ESBL genes in a single isolate. 69,70,71A worldwide surveillance recently showed that many NDM-1-producing bacteria detected carried additional ESBL genes. 72The acquisition of efficient mobile elements has accelerated the transfer of various antibiotic resistance genes.Potentially, a "super bug," resistant to almost all licensed antibiotics, may emerge in the future.Constant and careful worldwide surveillance for multidrug-resistant bacteria is urgently warranted.