Mechanisms of bacterial resistance to antibiotics

The three fundamental mechanisms of antimicrobial resistance are (1) enzymatic degradation of antibacterial drugs, (2) alteration of bacterial proteins that are antimicrobial targets, and (3) changes in membrane permeability to antibiotics. Antibiotic resistance can be either plasmid mediated or maintained on the bacterial chromosome. The most important mechanism of resistance to the penicillins and cephalosporins is antibiotic hydrolysis mediated by the bacterial enzyme beta-lactamase. The expression of chromosomal beta-lactamase can either be induced or stably depressed by exposure to beta-lactam drugs. Methods to overcome resistance to beta-lactam antibiotics include the development of new antibiotics that are stable to beta-lactamase attack and the coadministration of beta-lactamase inhibitors with beta-lactam drugs. Resistance to methicillin, which is stable to gram-positive beta-lactamase, occurs through the alteration of an antibiotic target protein, penicillin-binding protein 2. Production of antibiotic-modifying enzymes and synthesis of antibiotic-insensitive bacterial targets are the primary resistance mechanisms for the other classes of antibiotics, including trimethoprim, the sulfonamides, the aminoglycosides, chloramphenicol, and the quinolone drugs. Reduced antibiotic penetration is also a resistance mechanism for several classes of antibiotics, including the beta-lactam drugs, the aminoglycosides, chloramphenicol, and the quinolones.     

Prevalence of the genes encoding extended-spectrum beta-lactamases, in Escherichia coli resistant to beta-lactam and non-beta-lactam antibiotics

The prevalences of extended-spectrum beta-lactamases (ESBL) and their encoding bla genes, TEM, SHV and CTX_M, were investigated in isolates of Escherichia coli that were resistant to beta-lactam and/or non-beta-lactam antibiotics. Of the 250 E. coli isolates investigated, all of which came from patients in a major hospital in southern Lebanon, 61 (13.3%) were found to have ESBL, their production of beta-lactamase being confirmed by the ceftazidime and ceftazidime/clavulanic-acid disc methods. All 61 ESBL isolates were resistant to beta-lactams and sensitive to imipenem, piperacillin/tazobactam and cefoxitime. Only 40% were resistant to fluoroquinolones, 33% were resistant to aminoglycosides, and 18% were considered to have multi-drug resistance.The results of the PCR-based amplification of the bla gene in DNA samples from the 61 ESBL isolates indicated that 11 (18%) of the isolates carried both the TEM and SHV genes, 37 (61%) carried the TEM gene but not the SHV, and 13 (21%) had the SHV gene but not the TEM. None of the isolates carried the CTX_M gene. Of the 37 TEM-positive/SHV-negative isolates, 43% were resistant to fluoroquinolones and 37% to aminoglycosides. Increased resistance to non-beta-lactam antibiotics was observed in the isolates harbouring both the TEM and SHV genes, of which 54% were resistant to all of the tested antibiotics except imipenem, 36% were only resistant to fluoroquinolones, and 9.1% only resistant to aminoglycosides.The possibility that the concomitant presence of TEM- and SHV-type beta-lactamases is associated with resistance to non-beta-lactam antibiotics requires further research. The prevalences of ESBL and their encoding genes in Gram-negative bacteria collected from various regions in Lebanon will now be investigated.     

中国小型猪细菌分离鉴定及药敏试验

目的对小型猪携带的细菌种群分布情况及耐药性进行调查分析。方法对小型猪采样进行细菌分离培养、生化鉴定和细菌16SrRNA基因PCR扩增、测序分析,并进行药敏试验。结果从25只小型猪中共分离到45株细菌。结果显示,小型猪携带的细菌种群主要有弯曲菌属(空肠弯曲菌)、螺杆菌属(幽门螺杆菌)、克雷伯菌属(肺炎克雷伯氏菌);肠杆菌属(阴沟肠杆菌);埃希菌属(大肠埃希菌、弗格森埃希菌);假单胞菌属(铜绿假单胞菌);窄食单胞菌属(嗜麦芽窄食单胞菌);葡萄球菌属(金黄色葡萄球菌、溶血葡萄球菌、模仿葡萄球菌);链球菌属(肺炎链球菌、猪链球菌、前庭链球菌、缓症链球菌、麻疹孪生球菌、绿色气球菌);肠球菌属(粪肠球菌);芽孢杆菌属(枯草芽孢杆菌、地衣芽孢杆菌、蜂房芽孢杆菌、巨大芽孢杆菌)。这些菌株对氨曲南、头孢噻吩等药物敏感,而对痢特灵等药物不敏感。结论小型猪携带的细菌种群具有多样性,研究结果为我国小型猪细菌学检测以及质量标准的制定提供了科学依据。     

Emergence and resurgence of meticillin-resistant Staphylococcus aureus as a public-health threat

Staphylococcus aureus is a gram-positive bacterium that colonises the skin and is present in the anterior nares in about 25-30% of healthy people. Dependent on its intrinsic virulence or the ability of the host to contain its opportunistic behaviour, S aureus can cause a range of diseases in man. The bacterium readily acquires resistance against all classes of antibiotics by one of two distinct mechanisms: mutation of an existing bacterial gene or horizontal transfer of a resistance gene from another bacterium. Several mobile genetic elements carrying exogenous antibiotic resistance genes might mediate resistance acquisition. Of all the resistance traits S aureus has acquired since the introduction of antimicrobial chemotherapy in the 1930s, meticillin resistance is clinically the most important, since a single genetic element confers resistance to the most commonly prescribed class of antimicrobials--the beta-lactam antibiotics, which include penicillins, cephalosporins, and carbapenems.     

Mechanisms of resistance of bacteria causing ventilator-associated pneumonia

The common causes of ventilator-associated pneumonia (Staphylococcus aureus, Pseudomonas aeruginosa, Klebsiella pneumoniae, Enterobacter cloacae, Acinetobacter spp) are frequently resistant to multiple antibiotic classes. S aureus develops resistance to all beta-lactam antibiotics by producing a modified penicillin-binding protein. Linezolid resistance arises by way of mutations in the 23S ribosomal subunit. Antibiotic resistance in gram-negative bacilli usually arises by way of beta-lactamase production, upregulation of efflux pumps, or loss of outer membrane proteins. The ability of bacteria to develop and disseminate new mechanisms of antibiotic resistance may outstrip the availability of new antibiotic classes.     

The prevention of antibiotic resistance during treatment

Prevention of emergence of antibiotic resistance during treatment is an important goal when prescribing antimicrobials. Antibiotic resistant bacteria can emerge in three main ways--by acquisition of new genes via transposons or horizontal gene transfer, by selection of resistant variants and by selection of naturally resistant strains. In order to minimize emergence of antibiotic resistance during therapy it is important to try and avoid antibiotics which encourage the transfer of resistance genes, to avoid selection of resistant variants from susceptible pathogens and to avoid ablation of antibiotic susceptible normal flora. However, implementing these objectives is not always easy. This paper discusses possible ways of limiting the emergence of resistant bacteria during treatment. It does not consider how to prevent the spread of these strains from person to person. The prevalence of antibiotic-resistant bacteria depends upon the selection of antibiotic-resistant strains and spread of these strains from person to person. Prevention therefore consists of two parts--the prevention of acquisition of resistance/selection of antibiotic-resistant variants and interrupting the mechanisms by which person-to-person spread can occur. This paper considers only the first of these two influences on prevalence of resistance.     

Single amino acid mutation in an ATP-binding cassette transporter gene causes resistance to Bt toxin Cry1Ab in the silkworm, Bombyx mori

Bt toxins derived from the arthropod bacterial pathogen Bacillus thuringiensis are widely used for insect control as insecticides or in transgenic crops. Bt resistance has been found in field populations of several lepidopteran pests and in laboratory strains selected with Bt toxin. Widespread planting of crops expressing Bt toxins has raised concerns about the potential increase of resistance mutations in targeted insects. By using Bombyx mori as a model, we identified a candidate gene for a recessive form of resistance to Cry1Ab toxin on chromosome 15 by positional cloning. BGIBMGA007792-93, which encodes an ATP-binding cassette transporter similar to human multidrug resistance protein 4 and orthologous to genes associated with recessive resistance to Cry1Ac in Heliothis virescens and two other lepidopteran species, was expressed in the midgut. Sequences of 10 susceptible and seven resistant silkworm strains revealed a common tyrosine insertion in an outer loop of the predicted transmembrane structure of resistant alleles. We confirmed the role of this ATP-binding cassette transporter gene in Bt resistance by converting a resistant silkworm strain into a susceptible one by using germline transformation. This study represents a direct demonstration of Bt resistance gene function in insects with the use of transgenesis.     

The epidemiology of antibiotic resistance in hospitals: paradoxes and prescriptions

A simple mathematical model of bacterial transmission within a hospital was used to study the effects of measures to control nosocomial transmission of bacteria and reduce antimicrobial resistance in nosocomial pathogens. The model predicts that: (i) Use of an antibiotic for which resistance is not yet present in a hospital will be positively associated at the individual level (odds ratio) with carriage of bacteria resistant to other antibiotics, but negatively associated at the population level (prevalence). Thus inferences from individual risk factors can yield misleading conclusions about the effect of antibiotic use on resistance to another antibiotic. (ii) Nonspecific interventions that reduce transmission of all bacteria within a hospital will disproportionately reduce the prevalence of colonization with resistant bacteria. (iii) Changes in the prevalence of resistance after a successful intervention will occur on a time scale of weeks to months, considerably faster than in community-acquired infections. Moreover, resistance can decline rapidly in a hospital even if it does not carry a fitness cost. The predictions of the model are compared with those of other models and published data. The implications for resistance control and study design are discussed, along with the limitations and assumptions of the model.     

How much do Pseudomonas biofilms contribute to symptoms of pulmonary exacerbation in cystic fibrosis?

Recent studies suggest that chronic Pseudomonas aeruginosa lung infections in cystic fibrosis (CF) involve anaerobic biofilms, and that these biofilms are the reason chronic infections are rarely eradicated by antibiotic therapy, regardless of the in vitro susceptibility of infecting bacteria. These observations led to the development of an in vitro method for testing antibiotic susceptibility of CF clinical isolates in biofilms (Moskowitz et al., J Clin Microbiol 2004;42:1915-1922) and unearthed an apparent paradox. Antibiotics that remain cornerstones of clinical management of CF pulmonary exacerbations (e.g., beta-lactam antibiotics) appear to have little to no activity at clinically achievable concentrations when tested in vitro against clinical P. aeruginosa isolates growing in biofilms. The proven clinical efficacy of beta-lactam antibiotics in treating exacerbations, and the selection for beta-lactam resistance in vivo, suggest that planktonic bacteria play a significant role in pulmonary exacerbations of CF. A model of infection architecture is proposed in which biofilm and planktonic compartments each play a role in infection persistence and pulmonary exacerbation, respectively. Infection architecture may partially account for the observed lack of correlation between in vitro antibiotic susceptibility testing and clinical response to antibiotic therapy.     

Amdinocillin treatment of catheter-associated bacteriuria in rabbits

The effect of the beta-lactam antibiotic, amdinocillin, on the bacterial biofilm adherent to the Foley catheter surface, the bacterial microcolonies attached to the urinary bladder mucosa, and on planktonic bacteria in the urine was studied in a rabbit model of the closed urinary catheter drainage system. Progressively increasing the dose of antibiotic in this experimental catheter-associated urinary tract infection model first eliminated the bacterial population adherent to the bladder mucosa and then the planktonic population in the urine. The bacterial biofilm on the Foley catheter could be eradicated only by the highest dose of antibiotic (400 mg/kg). Scanning electron microscopy showed a gradual deterioration of bacterial biofilm and reduction in bacterial numbers with increasing antibiotic dosages. These data suggest that antibiotics used in short-term catheterization may reduce the serious sequelae associated with catheter-related infections by clearing the potentially dangerous bladder mucosal bacterial populations and urine planktonic bacteria.     

Medicinal plant products targeting quorum sensing for combating bacterial infections

Traditional treatment of infectious diseases is based on compounds that aim to kill or inhibit bacterial growth. The bacterial resistance against antibiotics is a serious issue for public health. Today, new therapeutic targets other than the bacterial wall were deciphered. Quorum sensing or bacterial pheromones are molecules called auto-inducer secreted by bacteria to regulate some functions such as antibiotic resistance and biofilms formation. This therapeutic target is well-studied worldwide, nevertheless the scientific data are not updated and only recent researches started to look into its potential as a target to fight against infectious diseases. A major concern with this approach is the frequently observed development of resistance to antimicrobial compounds. Therefore, this paper aims to provide a current overview of the quorum sensing system in bacteria by revealing their implication in biofilms formation and the development of antibiotic resistance, and an update on their importance as a potential target for natural substances.     

Antimicrobial susceptibility patterns and distribution of blaOXA genes among Acinetobacter spp. Isolated from patients at Tehran hospitals

Multiple drug-resistant strains of Acinetobacter have created therapeutic problems worldwide. This study was conducted to determine the antimicrobial susceptibility patterns and prevalence of bla(OXA-type) carbapenemases among isolates of Acinetobacter spp. obtained from Iranian patients. Here, 128 Acinetobacter isolates were identified at the species level, and their susceptibilities to different antibiotics were determined using disk agar diffusion testing. Isolates were then subjected to multiplex-PCR targeting bla(OXA) genes. More than 50% of the isolates showed multidrug resistance to different antibiotics. The rates of susceptibility to imipenem, meropenem, piperacillin-tazobactam, and amikacin were 50.7, 50, 42.1, and 38.2%, respectively. The MICs of carbapenems for the resistant isolates ranged from 64 to > or = 256 microg/ml. All strains of Acinetobacter baumannii possessed a bla(OXA-51-like) gene. The co-existence of bla(OXA-51-like)/bla(OXA-23-like) and bla(OXA-51-like)/bla(OXA-24-like) was detected in 25% (n=32) and 17.9% (n=23) of the isolates, respectively. Over 70% of carbapenem-resistant strains contained at least two genes encoding OXA-type carbapenemase. Resistance to carbapenems in the population of Acinetobacter strains in Iran is high, with the majority of isolates showing multidrug resistance. A wide diversity of OXA genes exists among the strains of A. baumannii in Iran. Detection of bla(OXA-51-like) can be used as a simple and reliable method to differentiate A. baumannii strains from other species.     

Genetic mapping of resistance to Bacillus thuringiensis toxins in diamondback moth using biphasic linkage analysis

Transgenic plants producing environmentally benign Bacillus thuringiensis (Bt) toxins are deployed increasingly for insect control, but their efficacy will be short-lived if pests adapt quickly. The diamondback moth (Plutella xylostella), a worldwide pest of vegetables, is the first insect to evolve resistance to Bt toxins in open-field populations. A recessive autosomal gene confers resistance to at least four Bt toxins and enables survival without adverse effects on transgenic plants. Allelic variants of this gene confer resistance in strains from Hawaii, Pennsylvania, and the Philippines. Here we exploited the biphasic nature of Lepidopteran genetic linkage to map this gene in diamondback moth with 207 amplified fragment length polymorphisms as DNA markers. We also cloned and sequenced an amplified fragment length polymorphism marker for the chromosome containing the Bt resistance gene. The results provide a powerful tool for facilitating progress in understanding, monitoring, and managing resistance to Bt.     

Complications of hematopoietic stem cell transplantation: Bacterial infections

Bacterial infections remain a common complication of hematopoietic stem cell transplantation (HSCT), especially in the pre-engraftment phase. The risk of bacterial infections is mainly related to neutropenia, mucositis, and the presence of vascular lines. Most parts of the world have witnessed a shift in epidemiology toward Gram-negative bacteria; a large proportion of which are resistant to fluoroquinolones, extended-spectrum beta-lactams, carbapenems, and in some units even colistin. Meticulous infection control practices are essential for prevention of bacterial infections in HSCT. The role of routine prophylactic antibiotics is limited in settings with high rates of bacterial resistance. If used, prophylactic antibiotics should be limited to high-risk patients, and the agents are selected based on local resistance profiles. Neutropenic fever is a medical emergency in most HSCT recipients. Prompt clinical evaluation is paramount, along with the intravenous administration of appropriate empiric antimicrobials, typically an antipseudomonal beta-lactam agent. Glycopeptides should only be considered if the patient is hemodynamically unstable or Gram-positive infection is suspected. Additional Gram-negative agents, such as colistin or aminoglycosides, may be added if extensive Gram-negative resistance is expected. To mitigate increasing bacterial resistance, empiric antibiotic regimens should be rationalized or discontinued as soon as possible.     

鲍曼不动杆菌17种β内酰胺酶基因研究

目的了解鲍曼不动杆菌(Acinetobacter baumannii.Ab)β内酰胺酶耐药基因存在状况。方法根据美国CLSI2004年版要求进行抗生素敏感性判断,应用PCR技术对Ab菌进行17种β内酰胺酶基因测定。结果Ab对13种抗菌药物耐药严重。27株Ab菌中blaTEM阳性22株(81.5%)、blaOXA-23群阳性12株(44.4%),blaADC阳性23株(85.2%)。其中12株为blaTEM、blaOXA-23群、blaADC3种β内酰胺酶基因均阳性,10株为blaTEM、blaADC2种β内酰胺酶基因阳性。结论我院分离的Ab不仅具有多重耐药性,而且blaTEM、blaOXA-23群、blaADC等β内酰胺酶基因携带率高。     

Antimicrobial therapy of febrile complications after high-dose chemo-/radiotherapy and autologous hematopoietic stem cell transplantation--guidelines of the Infectious Diseases Working Party (AGIHO) of the German Society of Hematology and Oncology (DGHO)

Infectious complications occur in 60-100% of patients following high-dose chemotherapy (HDC) and autologous hematopoietic stem cell transplantation (HSCT), and are commonly caused by Gram-negative aerobic bacteria (such as Pseudomonas aeruginosa and enterobacteriacea e) and Gram-positive cocci (such as enterococci, streptococci and staphylococci), which should be covered by empiric first-line antibiotic therapy. Less frequently, infections are caused by fungi and anaerobic bacteria, and initial therapy does not necessarily have to cover coagulase-negative staphylococci, oxacillin-resistant S. aureus (MRSA), anaerobic bacteria and fungi. Patients who already receive antibiotics and develop pulmonary infiltrates should immediately be treated with systemic antifungals. Patients with fever and diarrhea or other signs and symptoms of gastrointestinal or perianal infection should be treated with antibiotics covering anaerobic bacteria and enterococci. Clinically stable patients with skin infections or central venous catheter-related infections can be treated with standard empiric antibiotic therapy including a beta-lactam active against Pseudomonas aeruginosa with or without an aminoglycoside, and should only receive glycopeptides if they do not respond to first-line therapy within 72 hours, become clinically unstable, have severe mucositis, or when resistance against the empiric antibiotics is demonstrated. Recombinant hematopoietic growth factors should not be added routinely but may be considered in life-threatening situations such as invasive pulmonary mycoses or sepsis.     

Evolution and spread of antibiotic resistance

Antibiotic resistance is a clinical and socioeconomical problem that is here to stay. Resistance can be natural or acquired. Some bacterial species, such as Pseudomonas aeruginosa, show a high intrinsic resistance to a number of antibiotics whereas others are normally highly antibiotic susceptible such as group A streptococci. Acquired resistance evolve via genetic alterations in the microbes own genome or by horizontal transfer of resistance genes located on various types of mobile DNA elements. Mutation frequencies to resistance can vary dramatically depending on the mechanism of resistance and whether or not the organism exhibits a mutator phenotype. Resistance usually has a biological cost for the microorganism, but compensatory mutations accumulate rapidly that abolish this fitness cost, explaining why many types of resistances may never disappear in a bacterial population. Resistance frequently occurs stepwise making it important to identify organisms with low level resistance that otherwise may constitute the genetic platform for development of higher resistance levels. Self-replicating plasmids, prophages, transposons, integrons and resistance islands all represent DNA elements that frequently carry resistance genes into sensitive organisms. These elements add DNA to the microbe and utilize site-specific recombinases/integrases for their integration into the genome. However, resistance may also be created by homologous recombination events creating mosaic genes where each piece of the gene may come from a different microbe. The selection with antibiotics have informed us much about the various genetic mechanisms that are responsible for microbial evolution.     

From the Cover: Increased survival of western corn rootworm on transgenic corn within three generations of on-plant greenhouse selection

To delay evolution of insect resistance to transgenic crops producing Bacillus thuringiensis (Bt) toxins, nearby “refuges” of host plants not producing Bt toxins are required in many regions. Such refuges are expected to be most effective in slowing resistance when the toxin concentration in Bt crops is high enough to kill all or nearly all insects heterozygous for resistance. However, Bt corn, Zea mays, introduced recently does not meet this “high-dose” criterion for control of western corn rootworm (WCR), Diabrotica virgifera virgifera. A greenhouse method of rearing WCR on transgenic corn expressing the Cry3Bb1 protein was used in which approximately 25% of previously unexposed larvae survived relative to isoline survival (compared to 1–4% in the field). After three generations of full larval rearing on Bt corn (Constant-exposure colony), WCR larval survival was equivalent on Bt corn and isoline corn in greenhouse trials, and the LC₅₀ was 22-fold greater for the Constant-exposure colony than for the Control colony in diet bioassays with Cry3Bb1 protein on artificial diet. After six generations of greenhouse selection, the ratio of larval recovery on Bt corn to isoline corn in the field was 11.7-fold greater for the Constant-exposure colony than the Control colony. Removal from selection for six generations did not decrease survival on Bt corn in the greenhouse. The results suggest that rapid response to selection is possible in the absence of mating with unexposed beetles, emphasizing the importance of effective refuges for resistance management.     

New antibiotic strategies in patients with cirrhosis and bacterial infection

Early diagnosis and adequate empirical antibiotic treatment of bacterial infections in advanced cirrhosis is essential to improve outcomes given the high risk of developing severe sepsis, multiple organ failure and death. β-lactams and quinolones are nowadays frequently ineffective in nosocomial and healthcare associated infections, due to the increasing prevalence of multidrug resistant (MDR) bacteria reported across different geographical areas. Recent antibiotic exposure also increases the risk of developing MDR bacterial infections. Initial antibiotic strategies should therefore be tailored according to the presence or absence of risk factors of MDR bacteria and to the severity of infection and should consider the local epidemiology. Empirical treatment in the population at high risk of MDR bacterial infections requires the use of broad-spectrum antibiotics (carbapenems or tigecycline) and of drugs active against specific resistant bacteria (glycopeptides, linezolid, daptomycin, amikacin, colistin). Early de-escalation policies are recommended to prevent the spread of MDR bacteria in cirrhosis.