Commonality of adherent-invasive Escherichia coli isolated from patients with extraintestinal infections, healthy individuals and the environment.

Adherent-invasive Escherichia coli (AIEC) has been implicated as a microbiological factor in inflammatory bowel disease (IBD) pathogenesis. These strains are defined by their ability to adhere to and invade intestinal epithelial cells, and to survive and replicate in macrophages. We postulated that AIEC strains may commonly inhabit the gut of healthy individuals (HI), cause extraintestinal infections, and be found in sewage treatment plants (STP) and surface waters (SW). A total of 808 E. coli strains isolated from HI; patients with community-acquired urinary tract infection (CA-UTI), septicaemia and urosepsis; STP; and SW, showing a diffuse adhesion pattern to Caco-2 cells were included in this study. Typing of the strains using a combination of RAPD-PCR and PhPlate fingerprinting grouped them into 48 common clones (CCs). Representatives of each CC were tested for the ability to invade Caco-2 cells, survive and replicate in macrophages, and for the presence of six virulence genes commonly found among AIEC strains. Twenty CCs were deemed AIEC based on their ability to survive and replicate in macrophages, while encoding htrA, dsbA and clbA genes. These CCs primarily originated from HI and CA-UTI patients but were also detected in secondary locations including STP and SW. Strains lacking intramacrophagic survival and replication abilities were regarded as diffusely adhering E. coli (DAEC). Certain clones of AIEC are common in the gut of HI whilst promoting CA-UTI. The survival and persistence of AIEC in STP and SW may have serious public health ramifications for individuals predisposed to IBD.

Invasion and translocation of uropathogenic Escherichia coli isolated from urosepsis and patients with community-acquired urinary tract infection.

Uropathogenic Escherichia coli (UPEC) strains are found in high numbers in the gut of patients with urinary tract infections (UTIs). We hypothesised that in hospitalised patients, UPEC strains might translocate from the gut to the blood stream and that this could be due to the presence of virulence genes (VGs) that are not commonly found in UPEC strains that cause UTI only. To test this, E. coli strains representing 75 dominant clonal groups of UPEC isolated from the blood of hospitalised patients with UTI (urosepsis) (n = 22), hospital-acquired (HA) UTI without blood infection (n = 24) and strains isolated from patients with community-acquired (CA)-UTIs (n = 29) were tested for their adhesion to, invasion and translocation through Caco-2 cells, in addition to the presence of 34 VGs associated with UPEC. Although there were no differences in the rate and degree of translocation among the groups, urosepsis and HA-UTI strains showed significantly higher abilities to adhere (P = 0.0095 and P < 0.0001 respectively) and invade Caco-2 cells than CA-UTI isolates (P = 0.0044, P = 0.0048 respectively). Urosepsis strains also carried significantly more VGs than strains isolated from patients with only UTI and/or CA-UTI isolates. In contrast, the antigen 43 allele RS218 was found more commonly among CA-UTI strains than in the other two groups. These data indicate that UPEC strains, irrespective of their source, are capable of translocating through gut epithelium. However, urosepsis and HA-UTI strains have a much better ability to interact with gut epithelia and have a greater virulence potential than CA-UPEC, which allows them to cause blood infection.

Comparison between virulence characteristics of dominant and non-dominant Escherichia coli strains of the gut and their interaction with Caco-2 cells.

Escherichia coli strains are normal inhabitants of the gut and are normally found in the faeces of the host at different population sizes. We characterised faecal E. coli of 45 healthy male (n = 17) and female (n = 28) volunteers by testing 28 isolates from each individual. These isolates were typed and divided into dominant (if constituted >50% of the population tested) and non-dominant types in each individual. Representative strains of each dominant and non-dominant type were tested for their virulence gene profiles, their ability to form biofilm, adhere to, invade and translocate through a gut epithelial cell line (Caco-2 cells). Strains belonging to dominant types adhered significantly more to Caco-2 cells than non-dominant strains (5.7 ± 0.3 versus 4.3.± 0.13 CFU/cell mean ± SEM, P = 0.0003). They also invaded (135 ± 6 versus 63 ± 13 CFU) and translocated through Caco-2 cells (84 ± 5 versus 32 ± 9 CFU) significantly more than non-dominant strains (P < 0.0001 and P = 0.0002, respectively). Moreover, dominant strains showed the ability to form significantly more biofilm than non-dominant strains (1.1 ± 0.01 versus 0.5 ± 0.1 OD600, P < 0.0001). Majority (51%) of the strains belonged to phylogroup D followed by B2 (23%). Furthermore, out of 25 virulence genes tested, kpsMTII, papC and papG allele III were found to be significantly higher among dominant than non-dominant strains. Our results suggest that E. coli strains dominating the gut may have virulence properties that enable them to efficiently interact with the gut epithelium and translocate under predisposing conditions of the host.

Genes and associated peptides involved with aestivation in a land snail.

Some animals can undergo a remarkable transition from active normal life to a dormant state called aestivation; entry into this hypometabolic state ensures that life continues even during long periods of environmental hardship. In this study, we aimed to identify those central nervous system (CNS) peptides that may regulate metabolic suppression leading to aestivation in land snails. Mass spectral-based neuropeptidome analysis of the CNS comparing active and aestivating states, revealed 19 differentially produced peptides; 2 were upregulated in active animals and 17 were upregulated in aestivated animals. Of those, the buccalin neuropeptide was further investigated since there is existing evidence in molluscs that buccalin modulates physiology by muscle contraction. The Theba pisana CNS contains two buccalin transcripts that encode precursor proteins that are capable of releasing numerous buccalin peptides. Of these, Tpi-buccalin-2 is most highly expressed within our CNS transcriptome derived from multiple metabolic states. No significant difference was observed at the level of gene expression levels for Tpi-buccalin-2 between active and aestivated animals, suggesting that regulation may reside at the level of post-translational control of peptide abundance. Spatial gene and peptide expression analysis of aestivated snail CNS demonstrated that buccalin-2 has widespread distribution within regions that control several physiological roles. In conclusion, we provide the first detailed molecular analysis of the peptides and associated genes that are related to hypometabolism in a gastropod snail known to undergo extended periods of aestivation.

Differential peptide expression in the central nervous system of the land snail Theba pisana, between active and aestivated.

Hypometabolism is a physiological state of dormancy entered by many animals in times of environmental stress. There are gaps in our understanding of the molecular components used by animals to achieve this metabolic state. The availability of genomic and transcriptome data can be useful to study the process of hypometabolism at the molecular level. In this study, we use the land snail Theba pisana to identify peptides that may be involved in the hypometabolic state known as aestivation. We found a total of 22 neuropeptides in the central nervous system (CNS) that were differentially produced during activity and aestivation based on mass spectral-based neuropeptidome analysis. Of these, 4 were upregulated in active animals and 18 were upregulated in aestivation. A neuropeptide known to regulate muscle contractions in a variety of molluscs, the small cardioactive peptide A (sCAPA), and a peptide of yet unknown function (termed Aestivation Associated Peptide 12) were chosen for further investigation using temporal and spatial expression analysis of the precursor gene and peptide. Both peptides share expression within regions of the CNS cerebral ganglia and suboesophageal ganglia. Relative transcript abundance suggests that regulation of peptide synthesis and secretion is post-transcriptional. In summary, we provide new insights into the molecular basis of the regulation of aestivation in land snails through CNS peptide control.