A Molecular Study on the Prevalence and Virulence Potential of Aeromonas spp. Recovered from Patients Suffering from Diarrhea in Israel

Background: Species of the genus Aeromonas are native inhabitants of aquatic environments and have recently been considered emerging human pathogens. Although the gastrointestinal tract is by far the most common anatomic site from which aeromonads are recovered, their role as etiologic agents of bacterial diarrhea is still disputed. Aeromonas-associated diarrhea is a phenomenon occurring worldwide; however, the exact prevalence of Aeromonas infections on a global scale is unknown. Methodology/Principal Findings: The prevalence and virulence potential of Aeromonas in patients suffering from diarrhea in Israel was studied using molecular methods. 1,033 diarrheal stools were sampled between April and September 2010 and Aeromonas species were identified in 17 (,2%) patients by sequencing the rpoD gene. Aeromonas species identity and abundance was: A. caviae (65%), A. veronii (29%) and Aeromonas taiwanensis (6%). This is the first clinical record of A. taiwanensis as a diarrheal causative since its recent discovery from a wound infection in a patient in Taiwan. Most of the patients (77%) from which Aeromonas species were isolated were negative for any other pathogens. The patients ranged from 1 to 92 years in age. Aeromonas isolates were found to possess different virulence-associated genes: ahpB (88%), pla/ lip/lipH3/apl-1 (71%), act/hlyA/aerA (35%), alt (18%), ast (6%), fla (65%), lafA (41%), TTSS ascV (12%), TTSS ascF-ascG (12%), TTSS-dependent ADP-ribosylating toxins aexU (41%) and aexT (6%) in various combinations. Most of the identified strain

Discrimination within Phenotypically Closely Related Definitive Types of Salmonella enterica Serovar Typhimurium by the Multiple Amplification of Phage Locus Typing Technique

Multilocus sequence typing (MLST) is a relatively new high-resolution typing system employed for epidemiological studies of bacteria, including Salmonella. Discrimination based on MLST of housekeeping genes may be problematical, due to the high identity of gene sequences of closely related Salmonella species. The presence of genomic sequences derived from stable temperate phages in Salmonella offers an alternative for MLST of Salmonella. We have used MLST of prophage loci in Salmonella enterica serovar Typhimurium to discriminate closely related isolates of serovar Typhimurium. We have compared these results to MLST of five housekeeping genes, as well as pulsed-field gel electrophoresis (PFGE). The presence or absence of prophage loci in the 73 serovar Typhimurium isolates tested, as well as allelic variation as detected by sequencing, provided greater discrimination between isolates than either MLST of housekeeping genes or PFGE. Amplification of prophage loci alone separated serovar Typhimurium isolates into 27 groups comprising multiple isolates or individual strains. Sequencing of isolates found within the clusters separated isolates even further. By contrast, PFGE could only divide the 73 isolates into five distinct groups. MLST using housekeeping genes did not provide any significant separation of isolates in comparison to amplification or MLST of prophage loci. The results demonstrate that the amplification and sequencing of prophage loci provides a high-resolution, objective method for the discrimination of closely related isolates of serovar Typhimurium. It is proposed that multiple amplification of phage locus typing may provide sufficient discrimination for epidemiological purposes without recourse to MLST

Use of AFLP and PFGE to discriminate between Salmonella enterica serovar Typhimurium DT126 isolates from separate food-related outbreaks in Australia.

In 2001 Salmonella enterica serovar Typhimurium definitive phage-type (DT) 126 was isolated at higher frequency in Australia compared to other S. Typhimurium phage types and in comparison to previous years. Associated with this increase was the implication of this phage type in a number of food-related outbreaks. We compared fluorescent amplified fragment length polymorphism (FAFLP) to pulsed-field gel electrophoresis (PFGE), the current 'gold standard' for molecular typing of Salmonella for the discrimination between outbreak-associated isolates and epidemiologically unrelated DT126 strains. FAFLP showed a greater ability to discriminate between isolates than PFGE, with 16 groups of clusters or individual isolates with < 90% similarity to each other compared to three groups as determined by PFGE. Both methods were able to discriminate between isolates from two separate outbreaks in South Australia and isolates associated with an outbreak at a restaurant in New South Wales. The resolving power of both methods was not sufficient to separate all epidemiologically unrelated DT126 isolates from the outbreak isolates. We conclude that amplified fragment length polymorphism is a useful tool to assist in the discrimination of S. Typhimurium DT126 isolates

CS5 Pilus Biosynthesis Genes from Enterotoxigenic Escherichia coli O115:H40

We have sequenced the entire region of DNA required for the biosynthesis of CS5 pili from enterotoxigenic Escherichia coli O115:H40 downstream of the major subunit gene, designated csfA (for coli surface factor five A). Five more open reading frames (ORFs) (csfB, csfC, csfE, csfF, and csfD) which are transcribed in the same direction as the major subunit and are flanked by a number of insertion sequence regions have been identified. T7 polymerase-mediated overexpression of the cloned csf ORFs confirmed protein sizes based on the DNA sequences that encode them. The expression of only the csf region in E. coli K-12 resulted in the hemagglutination of human erythrocytes and the cell surface expression of CS5 pili, suggesting that the cluster contains all necessary information for CS5 pilus biogenesis and function

MLVA and phage typing as complementary tools in the epidemiological investigation of Salmonella enterica serovar typhimurium clusters

In South Australia serotyping and phage typing are employed for routine Salmonella surveillance. Molecular techniques such as Multiple-locus variable number tandem repeat analysis (MLVA) are increasingly utilized to aid outbreak investigations. During 2007 three Salmonella enterica serovar Typhimurium outbreaks involving phage types DT9, DT29, and DT44 were investigated. Human, food and environmental isolates were also typed by MLVA. In the DT9 outbreak cluster MLVA demonstrated distinct groupings that corresponded to epidemiological differences in time, place, and descriptive information on potential transmission mechanisms. In contrast, the human and food isolates of both the DT29 and DT44 clusters had identical MLVA profiles for all but one case. These data correlated with the epidemiology suggesting that these isolates were closely related and probably a single agent. These findings illustrate that phage typing and MLVA can provide different but complementary information for epidemiological investigations of Salmonella outbreaks.Ian L. Ross, Dianne E. Davos, Lillian Mwanri, Jane Raupach and Michael W. Heuzenroede

A Delphi study to identify content for a new questionnaire based on the 10 Principles of Dignity in Care

Published online: 16 August 2020AIM AND OBJECTIVE:To generate content for a new questionnaire, based on the 10 Principles of Dignity in Care. BACKGROUND:Older people in hospital are vulnerable and at risk of harm, including indignity. The 10 Principles of Dignity in Care, which undergird the United Kingdom's Dignity in Care Campaign, have been used to promote dignified care for older people in hospital. A 2006 recommendation of the campaign was to survey people on their experiences of dignity in care. To undertake such a survey, a questionnaire based on the 10 Principles of Dignity is required. DESIGN:Qualitative methods based on a modified Delphi technique, assessed against the CREDES checklist. METHODS:A Delphi panel of experts was convened that included: consumers, carers, clinicians, academics, policy experts and representatives from the National Dignity Council in the UK, Aboriginal people and people from culturally and linguistically diverse backgrounds. RESULTS:Fifty-seven experts consented to participate, over the three rounds of Delphi panel deliberations (response rate: R1 n = 49, R2 n = 47 and R3 n = 44). The Delphi panellists were asked to rank, rewrite, relocate or remove items and suggest additional items, under each of the 10 Principles of Dignity in Care. The initial list of 93 items, generated from a review of the literature, existing questionnaires and drafted by the authors, was reduced to 87 items in Round 2 and 69 items in Round 3. CONCLUSIONS:A panel of experts were able to determine, based on their own judgement, and through consensus, the 69-items and response categories to be included in the patient and carer versions of the Dignity in Care questionnaire, to progress to a pilot study.Louise Heuzenroeder, Faizal Ibrahim, Jyoti Khadka, Richard Woodman, Alison Kitso