Quorum-sensing control of bacteriocin production in olive fermentations

Bacteriocin production in many lactic acid bacteria is controlled by specific peptides called autoinducers (AIPs) via a sophisticated mechanism which senses cell density and is known as quorum sensing (QS).Peer Reviewe

Predominance of multidrug-resistant bacteria causing urinary tract infections among symptomatic patients in East Africa : a call for action

Background In low- and middle-income countries, antibiotics are often prescribed for patients with symptoms of urinary tract infections (UTIs) without microbiological confirmation. Inappropriate antibiotic use can contribute to antimicrobial resistance (AMR) and the selection of MDR bacteria. Data on antibiotic susceptibility of cultured bacteria are important in drafting empirical treatment guidelines and monitoring resistance trends, which can prevent the spread of AMR. In East Africa, antibiotic susceptibility data are sparse. To fill the gap, this study reports common microorganisms and their susceptibility patterns isolated from patients with UTI-like symptoms in Kenya, Tanzania and Uganda. Within each country, patients were recruited from three sites that were sociodemographically distinct and representative of different populations. Methods UTI was defined by the presence of >104 cfu/mL of one or two uropathogens in mid-stream urine samples. Identification of microorganisms was done using biochemical methods. Antimicrobial susceptibility testing was performed by the Kirby–Bauer disc diffusion assay. MDR bacteria were defined as isolates resistant to at least one agent in three or more classes of antimicrobial agents. Results Microbiologically confirmed UTI was observed in 2653 (35.0%) of the 7583 patients studied. The predominant bacteria were Escherichia coli (37.0%), Staphylococcus spp. (26.3%), Klebsiella spp. (5.8%) and Enterococcus spp. (5.5%). E. coli contributed 982 of the isolates, with an MDR proportion of 52.2%. Staphylococcus spp. contributed 697 of the isolates, with an MDR rate of 60.3%. The overall proportion of MDR bacteria (n = 1153) was 50.9%. Conclusions MDR bacteria are common causes of UTI in patients attending healthcare centres in East African countries, which emphasizes the need for investment in laboratory culture capacity and diagnostic algorithms to improve accuracy of diagnosis that will lead to appropriate antibiotic use to prevent and control AMR.Peer reviewe

Treatment seeking behaviours, antibiotic use and relationships to multi-drug resistance : a study of urinary tract infection patients in Kenya, Tanzania and Uganda

Antibacterial resistance (ABR) is a major public health threat. An important accelerating factor is treatment-seeking behaviour, including inappropriate antibiotic (AB) use. In many low- and middle-income countries (LMICs) this includes taking ABs with and without prescription sourced from various providers, including health facilities and community drug sellers. However, investigations of complex treatment-seeking, AB use and drug resistance in LMICs are scarce. The Holistic Approach to Unravel Antibacterial Resistance in East Africa (HATUA) Consortium collected questionnaire and microbiological data from adult outpatients with urinary tract infection (UTI)-like symptoms presenting at healthcare facilities in Kenya, Tanzania and Uganda. Using data from 6,388 patients, we analysed patterns of self-reported treatment seeking behaviours (‘patient pathways’) using process mining and single-channel sequence analysis. Among those with microbiologically confirmed UTI (n = 1,946), we used logistic regression to assess the relationship between treatment seeking behaviour, AB use, and the likelihood of having a multi-drug resistant (MDR) UTI. The most common treatment pathway for UTI-like symptoms in this sample involved attending health facilities, rather than other providers like drug sellers. Patients from sites in Tanzania and Uganda, where over 50% of patients had an MDR UTI, were more likely to report treatment failures, and have repeat visits to providers than those from Kenyan sites, where MDR UTI proportions were lower (33%). There was no strong or consistent relationship between individual AB use and likelihood of MDR UTI, after accounting for country context. The results highlight the hurdles East African patients face in accessing effective UTI care. These challenges are exacerbated by high rates of MDR UTI, suggesting a vicious cycle of failed treatment attempts and sustained selection for drug resistance. Whilst individual AB use may contribute to the risk of MDR UTI, our data show that factors related to context are stronger drivers of variations in ABR.Peer reviewe

Production of Plantaricin NC8 by Lactobacillus plantarum NC8 is induced in the Presence of Different Types of Gram-positive Bacteria

Lactobacillus plantarum NC8 was shown to produce plantaricin NC8 (PLNC8), a recently purified and genetically characterized inducible class IIb bacteriocin, only when it was co-cultured with other gram-positive bacteria. Among 82 strains belonging to the genera Bacillus, Enterococcus, Lactobacillus, Lactococcus, Leuconostoc, Listeria, Pediococcus, Staphylococcus, and Streptococcus, 41 were shown to induce PLNC8 production in L. plantarum NC8. There was apparently no relationship between the sensitivity of the strains and their ability to induce the bacteriocin, indicating that the inducer and sensitive phenotypes may not be linked. In some instances, induction was promoted by both living and heatkilled cells of the inducing bacteria. However, no PLNC8-inducing activity was found in the respective cell-free, pure culture supernatants. Inducer strains also promoted the production of a PLNC8-autoinducing activity by L. plantarum NC8, which was found only in the cell-free co-culture supernatants showing inhibitory activity. This PLNC8-autoinducing activity was diffusible, heat resistant, and of a proteinaceous nature, and was different from the bacteriocin itself. Taken together, the results suggest that the presence of specific gram-positive bacteria acts as an environmental stimulus activating both PLNC8 production by L. plantarum NC8 and a PLNC8-autoinducing activity, which in turn triggers or maintains bacteriocin production in the absence of inducing cells.This work was supported by the Spanish Government through MCYT project AGL2000-1611-CO3-01. A.M. was the recipient of a grant from MCYT, Spain.Peer reviewe

Knockout of three-component regulatory systems reveals that the apparently constitutive plantaricin-production phenotype shown by Lactobacillus plantarum on solid medium is regulated via quorum sensing

It has been found that many bacteriocins from lactic acid bacteria (LAB) are only produced in broth cultures when specific growth conditions are achieved and a dedicated three-component regulatory system, involved in a quorum sensing (QS) mechanism, is switched on. Surprisingly, bacteriocin production in LAB occurs in an apparently constitutive manner on solid media. This study addresses the question of constitutive versus regulated bacteriocin production on solid media in two different QS-regulated plantaricin-producing strains: Lactobacillus plantarum NC8 and L. plantarum WCFS1. Construction of knockout mutants for their respective regulatory operons revealed that bacteriocin production is controlled through a QS mechanism in both strains, on solid as well as in liquid media. These results could be extensible to other bacteriocins from LAB which are only produced on agar plates and not in broth cultures. Our findings suggest that QS-regulated bacteriocin production in LAB has evolved for competing on solid supports rather than in liquid media. In practice, this could be of major importance in vegetable fermentations, where the solid substrate itself provides an enormous surface where bacteria can attach to and produce biofilms. Therefore, QS-regulated bacteriocinogenic LAB growing in biofilms are under the optimum conditions to produce bacteriocins. Selection of strains to be used as starter cultures for vegetable fermentations should take into account these facts.This study was supported by the Spanish Government through the MEC Projects AGL2003-00642 and AGL2006-00763.Peer reviewe

Small-scale total DNA extraction from bacteria and yeast for PCR applications

In this study, we describe a very simple and rapid method for extraction of both plasmid and chromosomal DNA from colonies or pellets of bacteria and yeast that is suitable for subsequent PCR. Our method involves only chloroform as the cell disrupting and extracting agent, also contributing to DNA preservation from inherent and contaminating nucleases.This study was supported by the Spanish government through MCYT projects AGL2000-1611-C03-01 and AGL2003-00642.Peer reviewe

Molecular analysis of the 21-kb bacteriocin-encoding plasmid pEF1 from Enterococcus faecium 6T1a

The complete 21,344-bp DNA sequence of the bacteriocin-encoding plasmid pEF1 from Enterococcus faecium 6T1a was determined. Thirty-four putative open reading frames which could code for proteins longer than 42 amino acids were found. Those included the structural genes encoding for the previously described bacteriocins enterocin I and J (also named as enterocins L50A and L50B). After comparison to sequences in public databases, analysis of the gene organization of pEF1 suggests a modular structure with three diVerent functional domains: the replication region, the bacteriocin region and the mobilization plus UV-resistance region. This genetic mosaic structure most probably evolved through recombination events promoted by transposable elements. The hypothesis that the bacteriocin cluster on pEF1 could act as a functional plasmid stabilization module in E. faecium 6T1a is discussed.This work was supported by MEC-CICYT Projects ALI97-0658-CO3-01 and AGL2003-00642. B.F. was the recipient of a contract from the Spanish Ministry of Education and Science. A.M.B. was the recipient of a grant from the Spanish Ministry of Science and Technology.Peer reviewe

Genome Sequence of Lactobacillus pentosus IG1, a Strain Isolated from Spanish-Style Green Olive Fermentations

Lactobacillus pentosus is the most prevalent lactic acid bacterium in Spanish-style green olive fermentations. Here we present the draft genome sequence of L. pentosus IG1, a bacteriocin-producing strain with biotechnological and probiotic properties isolated from this food fermentations.This research was funded by the Spanish Ministry of Science and Innovation (MICINN) through Project AGL2009-07861 and by the Junta de Andalucía Excellence Project AGR-04621. A.M.-B. and H.L.-P. were the recipients of grants awarded by the Spanish National Research Council (CSIC) through the JAE Doc and JAE predoctoral Programmes, respectivelyPeer reviewe