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Insects resist infection by natural selection that favors the survival and reproduction of the fittest phenotypes. Although the genetic mechanisms mediating the evolution of insect resistance have been investigated, little is known about the contribution of epigenetic mechanisms. Gene expression in response to a pathogen selection pressure is regulated by different mechanisms affecting chromatin plasticity. Whether transgenerational inheritance of genome-wide epigenetic marks contributes to the heritable manifestation of insect resistance is presently debated. Here, we review the latest works on the contributions of chromatin remodeling to insect immunity and adaptation to pathogens. We highlight DNA methylation, histone acetylation, and microRNAs in mediating the transgenerational inherited transcriptional reprogramming of defense-related gene expression and the evolution of insect resistance.

The worldwide spread of toxin-producing and multi-drug resistant bacteria in water, food and the environment is considered a major threat to human health. Drinking water quality is controlled by inspection of fecal indicators presence whereby viable contaminants will be efficiently reduced by chlorination which is a common process for disinfection. However, the all-out efficiency is arguable, because bacterial regrowth has been documented after disinfection. In this study, we investigated the stability of Shiga toxin producing Escherichia coli (STEC) and β-lactamase expressing E. coli and Pseudomonas aeruginosa isolates, both equipped with multiple or single β-lactamase resistance genes. The aim of the study was to analyze the efficiency of chlorine (Cl2) disinfection against shigatoxigenic or β-lactamase producing bacteria. Cl2 reacts with the bacterial cells after first contact. Counts of antibiotic resistant E. coli were lower after short than upon extended Cl2 treatment. P. aeruginosa counts decreased moderately upon 15–60 min treatment with 1.2 mg Cl2/l, while cells adapted to tap water were not cultivable anymore. We assume that the bacterial physiology changed to a temporary non-cultivatable state at first Cl2 contact followed by resuscitation of some cells at later stages. STEC viability went down continuously at low Cl2 concentrations and these toxigenic E. coli isolates exhibited slightly increased stability to Cl2 treatment compared with non-toxigenic E. coli. Controlling the efficiency of disinfection, realistic counts of cultivatable cells are achieved after extended Cl2 action.

Fruits from Apium graveolens (Celery) are used traditionally in Persian and European medicine for the treatment of uncomplicated urinary tract infections. No data are available on A. graveolens extract on the interplay between uropathogenic E. coli and the eukaryotic host cells and on quorum sensing of the bacteria. The present study aimed to characterize an antiadhesive and anti quorum sensing effect of a characterized A. graveolens extract by specific in vitro assays and to correlate these effects with in vivo data obtained by an animal infection model. Hydroalcoholic extract CSE (EtOH-water, 1:1) from A. graveolens fruits was characterized by UHPLC/+ESI-QTOF-MS and investigated on antiproliferative activity against UPEC (strain NU14) and human T24 bladder cells. Antiadhesive properties of CSE were investigated within two different in vitro adhesion assays. For in vivo studies BALB/c mice were used in an UPEC infection model. The effect of CSE on bacterial load in bladder tissue was monitored within a 4- and 7 days pretreatment (200, 500 mg/kg) of the animals. CSE was dominated by the presence of luteolin-glycosides and related flavons besides furocoumarins. CSE had no cytotoxic effects against UPEC and bladder cells. CSE exerts a dose dependent antiadhesive activity against UPEC strains NU14 and UTI89. CSE inhibited in a concentration-dependent manner bacterial quorum sensing. 4- and 7-day pretreatment of animals with CSE transurethrally infected with UPEC NU14, significantly reduced the bacterial load in bladder tissue. CSE is assessed as an antiadhesive extract for which the traditional use in phytotherapy for UTI is justified.

Background Extracts from the leaves of Orthosiphon stamineus are used in phytotherapy for treatment of uncomplicated urinary tract infections. Purposes Evaluation of an aqueous extract against infection with uropathogenic Escherichia coli in vivo; investigation of underlying microbiological mechanisms. Study design In vivo studies in mice and in vitro investigations on cytotoxicity, antiadhesive potential, influence on bacterial gene expression and quorum sensing. Methods Extract OWE was prepared by hot water extraction. For in vivo studies BALB/c mice were used in an UPEC infection model. The effect of OWE on bacterial load in bladder/kidney tissue was monitored in pre- and posttreatment. Cytotoxicity of OWE against different UPEC strains, T24 bladder/A498 kidney cells, gene expression analysis, monitoring of phenotypic motility and quorum sensing was investigated by standard methods of microbiology. Results OWE was quantified (UHPLC) according to the content of rosmarinic acid, cichoric acid, caffeic acid. Three- and 5-day treatment of animals with OWE (750 mg/kg) after transurethral infection with UPEC CFT073 reduced the bacterial load in bladder and kidney, similar to norfloxacin. Four- and 7-day pretreatment of mice prior to the infection with UPEC NU14 reduced bacterial bladder colonization. In vitro investigations indicated that OWE (≤2 mg/ml) has no cytotoxic or proliferation-inhibiting activity against different UPEC strains as well as against T24 bladder and A498 kidney cells. OWE exerts a dose dependent antiadhesive activity against UPEC strains NU14 and UTI89. OWE reduced gene expression of fimH, but evoked increase of the expression of motility/fitness gene fliC. Increase of bacterial motility on gene level was confirmed by a changed bacterial phenotype by an increased bacterial motility in soft agar assay. OWE inhibited in a concentration-dependent manner bacterial quorum sensing. Conclusion OWE is assessed as a strong antiadhesive plant extract for which the traditional use in phytotherapy for UTI might be justified.

Tuberculosis (TB), caused by Mycobacterium tuberculosis (M. tuberculosis), is the leading infectious disease which claims one human life every 15-20s globally. The persistence of this deadly disease in human population can be attributed to the ability of the bacterium to stay in latent form. M. tuberculosis possesses a plethora of mechanisms not only to survive latently under harsh conditions inside the host but also modulate the host immune cells in its favour. Various M. tuberculosis gene families have also been described to play a role in this process. Recently, human bone marrow derived mesenchymal stem cells (MSCs) have been reported as a niche for dormant M. tuberculosis. MSCs possess abilities to alter the host immune response. The bacterium finds this self-renewal and immune privileged nature of MSCs very favourable not only to modulate the host immune system, with some help from its own genes, but also to avoid the external drug pressure. We suggest that the MSCs not only provide a resting place for M. tuberculosis but could also, by virtue of their intrinsic ability to disseminate in the body, explain the genesis of extra-pulmonary TB. A similar exploitation of stem cells by other bacterial pathogens is a distinct possibility. It may be likely that other intracellular bacterial pathogens adopt this strategy to 'piggy-back' on to ovarian stem cells to ensure vertical transmission and successful propagation to the next generation.

Uropathogenic Escherichia coli (UPEC) is the most common causal agent of urinary tract infections (UTIs) in humans. Currently, clinical detection methods take hours (dipsticks) to days (culturing methods), limiting rapid intervention. As an alternative, the use of molecular methods could improve speed and accuracy, but their applicability is complicated by high genomic variability within UPEC strains. Here, we describe a novel PCR-based method for the identification of E. coli in urine. Based on in silico screening of UPEC genomes, we selected three UPEC-specific genes predicted to be involved in pathogenesis (c3509, c3686 (yrbH) and chuA), and one E. coli-specific marker gene (uidA). We validated the method on 128 clinical (UTI) strains. Despite differential occurrences of these genes in uropathogenic E. coli, the method, when using multi-gene combinations, specifically detected the target organism across all samples. The lower detection limit, assessed with model UPEC strains, was approximately 104 CFU/ml. Additionally, the use of this method in a novel ultrafast PCR thermal cycler (Nextgen PCR) allowed a detection time from urine sampling to identification of only 52 min. This is the first study that uses such defined sets of marker genes for the detection of E. coli in UTIs. In addition, we are the first to demonstrate the potential of the Nextgen thermal cycler. Our E. coli identification method has the potential to be a rapid, reliable and inexpensive alternative for traditional methods.

Urinary tract infections are one of the most common bacterial infections and a major public health problem. The predominant causative agents are uropathogenic Escherichia coli. These strains differ from commensal E. coli by the presence of additional horizontally acquired chromosomal material, so-called pathogenicity islands, which encode traits that promote efficient bacterial colonization of the urinary tract. Uropathogenic model strain E. coli 536 possesses six archetypal pathogenicity islands. Bacteriophage-like integrases encoded by each pathogenicity island contribute to island instability. To learn more about the stability of these six islands and factors controlling their stability we constructed two chromosomal reporter systems for the measurement of island loss, as well as for the measurement of the promoter activity of the six island-associated integrase genes at the population level. We used these reporter gene modules to analyze the role of SOS response in island instability. Tests with subinhibitory concentrations of different antibiotics, including many drugs commonly used for the treatment of urinary tract infection, indicated that only SOS response-inducing antibiotics led to an increased loss of islands which was always associated with an increase in the bacterial subpopulations showing high integrase promoter activity. This suggests that island excision correlates with the expression of the cognate integrase. Our reporter modules are valuable tools to investigate the impact of various growth conditions on genome plasticity. Furthermore, a better understanding of the conditions, which affect bacterial integrase expression may open ways to specifically manipulate the genome content of bacterial pathogens by increasing pathogenicity island deletion rates in infecting or colonizing bacteria, thus leading to the attenuation of bacterial pathogens.

Autotransporter (AT) proteins are widespread surface-exposed or secreted factors in Escherichia coli. Several ATs have been correlated with pathogenesis or specific phylogenetic lineages. Therefore, an application as biomarkers for individual extraintestinal pathogenic E.coli (ExPEC) or intestinal pathogenic E.coli (IPEC) has been proposed. To put this assumption on a solid foundation, we analyzed 111 publicly available E. coli genome sequences and screened them bioinformatically for the presence of 18 ATs. We determined the highest AT prevalence per strain in phylogroup B2 isolates and showed that AT distribution correlates rather with phylogenetic lineages than with pathotypes. Although a strict dependence between AT prevalence and pathotype was not observed, EspP, EhaA, and EhaG cluster with IPEC of phylogroup B1 and E, respectively, whereas UpaH is predominantly present in ExPEC of phylogroup B2. Furthermore, PicU, SepA, UpaB, UpaI, and UpaJ were associated with phylogroup B2. We detected UpaI and its positional ortholog EhaC in 93% of the E.coli strains tested. This AT variant is thus the most prevalent in E.coli irrespective of pathotype or phylogenetic background. Compared with the ATs UpaB, UpaC, and UpaJ of uropathogenic E.coli strain 536, UpaI had redundant functions, contributing to autoaggregation, biofilm formation, and binding to extracellular matrix proteins. The functional redundancy and wide distribution of ATs among pathogenic and non-pathogenic E.coli indicates that ATs cannot generally be regarded as specific biomarkers and virulence factors. Our results demonstrate that phylogeny has a bigger impact on the distribution of AT variants in E.coli than initially thought, especially in ExPEC.

Cystic fibrosis (CF) patients suffer from chronic recurrent bacterial airway infections, which eventually lead to reduced life expectancy. Escherichia coli has not been considered as a CF pathogen. A total of 176 patients were observed over 5.6 years on average from 2002 to 2009 in two CF centers in Muenster, Germany. Sputum and throat swab cultures were screened for E. coli. E. coli isolates were analyzed for clinical microbiologic characteristics as well as strain identity, clonal distribution and phenotypic variability. In 45 patients (25.6%) E. coli was cultured at least once, mostly at medium to high bacterial load and primarily from patients less than 5 and older than 8 years. In 19 patients (10.8%) the same E. coli strain was isolated at least 3 times within a period of more than 6 months, with a mean persistence of 29 months. Multi-locus sequence typing revealed a distinctively strong association of CF E. coli with the B2 major clonal group. During persistence, long-term colonizing strains exhibited phenotypic variability known for typical CF pathogens such as surface capsule overproduction and changes in colony size or hemolytic activity. E. coli was occasionally or persistently isolated in a quarter of CF patients, mostly in very young or older patients. The relatively high bacterial load of E. coli colonization, the distinct association with the highly virulent extra-intestinal B2 clonal group and phenotypic variability in the long-term colonizing strains suggests a previously unrecognized clinical significance of E. coli as a CF pathogen.

Urinary tract infections (UTIs) are among the most frequently occurring infections and are mostly caused by extraintestinal pathogenic Escherichia coli. DNA microarrays are potent molecular diagnostic tools for rapid diagnosis of bacterial infections with high relevance for UTIs. In this study, we present the integration and application of two DNA chip modules for the simultaneous detection of single nucleotide polymorphisms in gyrA (quinolone resistance) and fimH (increased adhesion to urinary tract epithelium). The performance of the combined diagnostic chip was assessed by genotyping 140 E. coli strains. Resistance-causing mutations could only be identified in UTI isolates. A complete genotyping assay could be performed in

The species Escherichia coli comprises not only non-pathogenic or commensal variants that belong to the normal intestinal flora of most mammals, but also various pathogenic strains causing diverse intestinal and extraintestinal infections in man and animals. Virulence factors and mechanisms involved in pathogenesis have been successfully analyzed for many years resulting in a wealth of knowledge about many E. coli pathotypes. However, our knowledge on the genome content, diversity and variability between pathogenic and also non-pathogenic subtypes is only slowly accumulating. Pathotypes have been largely defined by the presence or absence of particular DNA segments that in most cases appear to have been acquired via horizontal gene transfer events. As these regions are frequently subjected to excisions, rearrangements, and transfers they contribute to the previously unexpected and underestimated rapid evolution of E. coli variants resulting in the development of novel strains and even pathotypes. In these studies various novel aspects of genome diversity and plasticity in extraintestinal and intestinal pathogenic E. coli pathotypes have been addressed and the results have been directly applied for the improvement of diagnostic methods.

A series of multivalent mannose containing compounds were prepared varying in size from small divalent, to 16-valent glycodenrimers and 21-valent glycopolymers. The molecules were approached via a common mannose building block. As scaffolds dendrimers and dendrons based on the 3,5-di-(2-aminoethoxy)-benzoic acid branching unit were used along with commercially available PAMAM dendrimers. To include larger structures, linear glycopolymers with varying amounts of mannose were prepared via radical polymerization. The compounds were tested for their biological activity using a newly developed ELISA based inhibition assay, for their ability to inhibit the binding of recombinant type I fimbriated E. coli to a monolayer of T24 cell line derived from human urinary bladder epithelium. All compounds showed enhanced affinity as compared to mannose with IC50’s down to the low micromolar range.

The growing knowledge of genetic diversity and whole genome organization in bacteria shows that pathogenicity islands (PAIs) represent a subtype of a more general genetic element, termed genomic island (GEI), which is widespread among pathogenic and non-pathogenic microbes. These findings mirror the importance of horizontal gene transfer, genome reduction and recombination events as fundamental mechanisms involved in evolution of bacterial variants. GEIs are part of the flexible gene pool and carry selfish genes, but also determinants which may be beneficial under certain conditions thus increasing bacterial fitness and consequently their survival or transmission. In this review, we focus on the role of mobile genetic elements that may also contain toxin-encoding genes for genome variability and evolution of bacteria.

Background & Aims: The microflora plays a crucial role in inflammatory bowel diseases (IBDs). Specific pathogen-free (SPF), but not germ-free, interleukin (IL)-2-deficient (IL-2−/−) mice develop colitis. The colitogenicity of commensal bacteria was determined. Methods: Gnotobiotic IL-2−/− and IL-2+/+ mice were colonized with Escherichia coli mpk, Bacteroides vulgatus mpk, or both bacterial strains, or with E. coli strain Nissle 1917. DNA arrays were used to characterize E. coli mpk. Colitis was analyzed by histology and real-time reverse-transcription polymerase chain reaction (RT-PCR) for interferon (IFN)-γ, tumor necrosis factor (TNF)-α, IL-10, and CD14 messenger RNA (mRNA) expression. Bacterial numbers in feces and bacterial localization in the colon was determined by culture and fluorescence in situ hybridization (FISH). Results: IL-2−/− but not IL-2+/+ mice monocolonized with E. coli mpk developed colitis, whereas mono-association with B. vulgatus mpk, or E. coli Nissle, or co-colonization with E. coli mpk and B. vulgatus mpk, did not induce colitis. DNA array experiments and cellular studies revealed that E. coli mpk is a nonpathogenic strain. FISH and culture methods revealed that the anticolitogenic effect of B. vulgatus mpk on E. coli mpk cannot be explained by a significant reduction in numbers of E. coli in the colon. E. coli mpk-induced colitis was associated with increased IFN-γ, TNF-α, CD14, and IL-10 mRNA expression in the colon. Conclusions: In IL-2−/− mice, B. vulgatus mpk protects against E. coli mpk-triggered colitis by an unknown mechanism. E. coli Nissle does not induce colitis. Various bacterial species common to the microflora differ in their ability to trigger IBD.

Uropathogenic Escherichia coli (UPEC) are the most important group of microorganisms responsible for urinary tract infection. UPEC differ from non-pathogenic E. coli and from other E. coli pathotypes by the production of specific virulence factors, which enable the bacteria to adhere to uroepithelial cells and to establish urinary tract infections. Besides adherence factors, toxins, 'modulins', capsules, iron uptake systems and other bacterial products contribute to the virulence of the strains. The respective genes are frequently located on large pieces of DNA called 'pathogenicity islands' (PAIs). PAIs form (unstable) regions of the genome of UPECs, which are often associated with tRNA genes. Using various molecular techniques, the functions of PAI encoded gene products have been studied. The usage of DNA arrays give answers to questions on the distribution of PAIs among various enterobacteria and on the expression of the different genes under in vitro and in vivo conditions. In addition, assumptions can be made on the evolution of these important pathogens.

The exploitation of bacterial genome sequences has so far provided a wealth of new general information about the genetic diversity of bacteria, such as that of many pathogens. Comparative genomics uncovered many genome variations in closely related bacteria and revealed basic principles involved in bacterial diversification, improving our knowledge of the evolution of bacterial pathogens. A correlation between metabolic versatility and genome size has become evident. The degenerated life styles of obligate intracellular pathogens correlate with significantly reduced genome sizes, a phenomenon that has been termed "evolution by reduction". These mechanisms can permanently alter bacterial genotypes and result in adaptation to their environment by genome optimization. In this review, we summarize the recent results of genome-wide approaches to studying the genetic diversity of pathogenic bacteria that indicate that the acquisition of DNA and the loss of genetic information are two important mechanisms that contribute to strain-specific differences in genome content.

The uropathogenic Escherichia coli strain 536 (O6:K15:H31) carries four distinct DNA regions in its chromosome, termed pathogenicity islands (PAIs I536 to IV536). Each of these PAIs encodes at least one virulence factor. All four PAIs are associated with tRNA genes. PAI I536 and PAI II536 can be spontaneously deleted from the chromosome by homologous recombination between flanking direct repeats. The deletion of PAI II536 results in the truncation of the associated gene leuX encoding the tRNALeu. This tRNA influences the expression of various virulence traits. In order to get a deeper insight into the role of PAI I536/II536 and of the tRNA5LeU for the protein expression, the protein expression patterns of Escherichia coli 536 and different derivatives were studied. Differences in the protein expression patterns of the wild-type strain Escherichia coli 536, its mutants 536-21 (PAI I536-, PAI II536-, leuX-), 536delta102 (PAI I536+, PAI II536+, leuX-) as well as of the strain 536R3 (PAI I536-, PAI II536-, leuX+) were analyzed by two-dimensional polyacrylamide gel electrophoresis and MALDI-TOF mass spectrometry. We identified about 39 different intracellular proteins whose expression is markedly altered in the different strain backgrounds. These differences can be linked either to the presence or absence of the PAI I536 and PAI II536 or to that of the tRNA gene leuX. The identities of 34 proteins have been determined by MALDI-TOF-MS. The identification of five proteins was not possible. The results suggest that proteome analysis is an efficient approach to study differences in global gene expression. The comparison of protein expression patterns of the uropathogenic E. coli strain 536 and different derivatives revealed that in this strain the expression of various proteins including those encoded by many housekeeping genes is affected by the presence of PAI I536 and Pai II536 or by that of the tRNA5Leu.

Nowhere is the relationship between microorganisms and eukaryotes as diverse, intimate and clinically relevant as in the gastrointestinal tract. An estimated 500–1000 mostly anaerobic species reside in the intestine, approaching enormous densities of 10 11 –10 12 organisms per gram colon content. The commensal interactions between intestinal microorganisms and animal hosts have been difficult to study in the past because of the diversity of microorganisms involved and because of the lack of culturability that accompanies many microbial consortia. However, recent work has provided new insights into these interactions.

Background Adenomyomatosis of gallbladder (GB) is considered as benign condition, but several reports suggest association with malignancies, especially for segmental subtype. Because of this chance, patients suspected to have segmental or diffuse type adenomyomatosis on the imaging study may undergo cholecystectomy. But, in some cases, histopathology reveals only chronic inflammation. In this study, we aimed to analyze radiologic and pathologic features of chronic cholecystitis and adenomyomatosis for more accurate diagnosis. Methods Patients who performed cholecystectomy during recent four years in one medical center were reviewed retrospectively. Among those, patients who were suspected to have adenomyomatosis on their abdominal imaging but with no adenomyoma detected on the histopathologic examination were analyzed for their radiologic, pathologic and clinical features compared to patients who had consistent results between abdominal imaging and histopathologic examination. Results Among 3144 patients who underwent cholecystectomy, 358 patients were suspected to have GB adenomyomatosis on the abdominal imaging study. Among them (358 patients), only 92 patients had histopathologic findings compatible with adenomyomatosis. Remaining 266 patients (74.3%) did not have adenomyomatosis. Instead, many GB specimens showed chronic cholecystitis. In this group, diffuse subtype of adenomyomatosis was significantly frequent compared to 92 patients who had coherent adenomyoma finding for both imaging study and pathology. The frequency of accompanying GB stones was significantly higher in mismatching group. Radiologically, contour of GB wall thickening, intralesional cystic area and inner layer enhancement pattern was helpful in differentiating adenomyomatosis from chronic cholecystitis. Conclusion Chronic cholecystitis mimicking adenomyomatosis is difficult to diagnose solely based on the abdominal imaging, and this can lead to unnecessary cholecystectomy. Existence of gallstones and several radiologic features can help differentiate these challenging entities.

“Epidemiology is bread and butter for infectious diseases esearch”. This sentence was given by Stanley Falkow during a eeting on molecular pathogenesis research in the late nineties. lthough meanwhile there have been a number of breakthroughs, .g., genome sequencing approaches and others, the sentence is still alid. Therefore, it is necessary to summarize new aspects of moleclar epidemiology in infectious diseases research from different iewpoints. In this issue Wolfgang Witte and co-workers describe the recontruction of a phylogenetic tree for Staphylococcus aureus, one of he most important pathogens. Allessandro Muzzi and Claudio onati describe the pan-genome of Streptococcus pneumoniae. The an-genome hypothesis, first described by Rino Rappuoli and coorkers, is a very stimulating approach and it is interesting to iscuss this hypothesis in the light of different species. Alexander omasz and co-workers also describe staphylococci and pneumoocci from a clinical viewpoint. Furthermore, J.A. Wagenaar is in avour of the “one health” approach which is an important idea in rder to bring together veterinary medicine and human medicine. rom the viewpoint of veterinary medicine, Lothar Wieler and coorkers describe animals as a reservoir for pathogenic microbes, taphylococci, enterobacteria and others. Ulrich Dobrindt tries to nswer the question “What is an extraintestinal pathogen?”. Data rom genome analysis give new answers regarding this interesting uestion. A similar approach is described by Bjorn-Arne Lindstdt regarding enteropathogens appearing in Norway. Plasmids are mportant mobile elements of pathogenic microbes. Allessandra arattoli describes the distribution of resistance plasmids of gram egative bacteria. Last but not least Ulrich Vogel describes the Euroean efforts in order to harmonize the typing of meningococci, mportant pathogens of humans. Altogether, in the issue a number of leading scientists in the field f molecular epidemiology present their recent data. We hope that