The bacterial strains characterization problem

The accurate characterization of collections of bacterial strains is a major scientific challenge, since bacteria are indeed responsible of significant plant diseases and thus subjected to official control procedures (e.g., in Europe, Directive 2000/29/EC). The development of diagnostic tests is therefore an important issue in order to routinely identify strains of these species

rRNA gene restriction patterns as a characterization tool for Lactobacillus sake strains producing ropy slime

http://www.elsevier.nl/locate/01681605The rRNA gene restriction patterns (ribotypes) of 69 ropy slime producing Lactobacillus sake strains isolated mainly from vacuum-packaged meat products of ten meat plants were determined. Ribotypes of these spoilage bacteria were compared to the corresponding pat-terns of non-ropy L. sake strains, and also to other species of the genus Lactobacillus, Carnobacterium and Weissella associated with meat products. Ribotyping divided the ropy slime-producing L. sake strains into four characteristic groups corresponding to the phenotypic carbohydrate grouping. The major group was contaminating nine plants located in different parts of Finland and no association between certain ribotypes and individual plants was detected. Dif-ferences between ribotypes of slime producing and non-ropy strains of L. sake group sharing the same carbohydrate pattern were not found. Otherwise ribotyping distinguished the ropy slime producing strains from the non-ropy L. sake reference strains. All L. sake strains were distinguished from other species of the genus Lactobacillus, Carnobacterium and Weissella by characteristic band-ing patterns obtained especially with Hind III digestion. These results suggest that ribotyping is a suitable method for detection and surveillance of the contamination of ropy slime producing L. sake strains but the patterns alone cannot be used as markers of slime production capability. Comparison of ribotypes between differ-ent species of the genus Lactobacillus suggest that ribotyping may also be a suitable method for species identification within the genus Lactobacillus

Characterization of newly isolated lytic bacteriophages active against Acinetobacter baumannii

Based on genotyping and host range, two newly isolated lytic bacteriophages, myovirus vB_AbaM_Acibel004 and podovirus vB_AbaP_Acibel007, active against Acinetobacter baumannii clinical strains, were selected from a new phage library for further characterization. The complete genomes of the two phages were analyzed. Both phages are characterized by broad host range and essential features of potential therapeutic phages, such as short latent period (27 and 21 min, respectively), high burst size (125 and 145, respectively), stability of activity in liquid culture and low frequency of occurrence of phage-resistant mutant bacterial cells. Genomic analysis showed that while Acibel004 represents a novel bacteriophage with resemblance to some unclassified Pseudomonas aeruginosa phages, Acibel007 belongs to the well-characterized genus of the Phikmvlikevirus. The newly isolated phages can serve as potential candidates for phage cocktails to control A. baumannii infections

Fluorescence characterization of clinically-important bacteria

Healthcare-associated infections (HCAI/HAI) represent a substantial threat to patient health during hospitalization and incur billions of dollars additional cost for subsequent treatment. One promising method for the detection of bacterial contamination in a clinical setting before an HAI outbreak occurs is to exploit native fluorescence of cellular molecules for a hand-held, rapid-sweep surveillance instrument. Previous studies have shown fluorescence-based detection to be sensitive and effective for food-borne and environmental microorganisms, and even to be able to distinguish between cell types, but this powerful technique has not yet been deployed on the macroscale for the primary surveillance of contamination in healthcare facilities to prevent HAI. Here we report experimental data for the specification and design of such a fluorescence-based detection instrument. We have characterized the complete fluorescence response of eleven clinically-relevant bacteria by generating excitation-emission matrices (EEMs) over broad wavelength ranges. Furthermore, a number of surfaces and items of equipment commonly present on a ward, and potentially responsible for pathogen transfer, have been analyzed for potential issues of background fluorescence masking the signal from contaminant bacteria. These include bedside handrails, nurse call button, blood pressure cuff and ward computer keyboard, as well as disinfectant cleaning products and microfiber cloth. All examined bacterial strains exhibited a distinctive double-peak fluorescence feature associated with tryptophan with no other cellular fluorophore detected. Thus, this fluorescence survey found that an emission peak of 340nm, from an excitation source at 280nm, was the cellular fluorescence signal to target for detection of bacterial contamination. The majority of materials analysed offer a spectral window through which bacterial contamination could indeed be detected. A few instances were found of potential problems of background fluorescence masking that of bacteria, but in the case of the microfiber cleaning cloth, imaging techniques could morphologically distinguish between stray strands and bacterial contamination

Detection of genetic diversity among Indian strains of _Xanthomonas campestris_ pv. _mangiferaeindicae_ using PCR-RAPD

The randomly amplified polymorphic DNA (RAPD) technique was used to investigate the genetic diversity in 6 strains of _Xanthomonas campestris_ pv. _mangiferaeindicae_ (_Xcmi_), the causal pathogen of mango bacterial canker disease (MBCD). The RAPD analysis was also intended to identify molecular markers, specific to the species to develop PCR-based markers for detection of _Xcmi_ in mango field and planting materials. Twenty RAPD primers (CP 1-CP 20) were evaluated to establish molecular characters and genetic variability in the genome of _Xcmi_. Among these, only 4 were found efficient for development of reproducible banding pattern. It has been observed that the largest and smallest amplified RAPD products were of 2.036 and 0.201 kbp. A total of 136 bands were scored against 6 strains of _Xcmi_. There was 7.66 per cent polymorphism in individual isolates which indicates significant polymorphism among the evaluated strains, with mean difference of 0.33 (_Xcmi_ 2 vs. _Xcmi_ 8) and 0.29 (_Xcmi_ 10 vs. _Xcmi_ 12). However, the single linkage euclidean distances were statistically significant (P>0.05), i.e., 0.58. The markers CP 5, 10, 16 and 19 were amplified in all the strains with polymorphic alleles, which indicates that these markers could be used for rapid detection of genetic variability in _Xcmi_ strains

Mutation in the pssZ Gene Negatively Impacts Exopolysaccharide Synthesis, Surface Properties, and Symbiosis of Rhizobium leguminosarum bv. trifolii with Clover

Rhizobium leguminosarum bv. trifolii is a soil bacterium capable of establishing a nitrogen-fixing symbiosis with clover plants (Trifolium spp.). This bacterium secretes large amounts of acidic exopolysaccharide (EPS), which plays an essential role in the symbiotic interaction with the host plant. This polymer is biosynthesized by a multi-enzymatic complex located in the bacterial inner membrane, whose components are encoded by a large chromosomal gene cluster, called Pss-I. In this study, we characterize R. leguminosarum bv. trifolii strain Rt297 that harbors a Tn5 transposon insertion located in the pssZ gene from the Pss-I region. This gene codes for a protein that shares high identity with bacterial serine/threonine protein phosphatases. We demonstrated that the pssZ mutation causes pleiotropic effects in rhizobial cells. Strain Rt297 exhibited several physiological and symbiotic defects, such as lack of EPS production, reduced growth kinetics and motility, altered cell-surface properties, and failure to infect the host plant. These data indicate that the protein encoded by the pssZ gene is indispensable for EPS synthesis, but also required for proper functioning of R. leguminosarum bv. trifolii cells.Polish National Science Centre grant no. DEC-2012/07/B/NZ1/0009

Characterization of the role that bacterial surface polysaccharide poly N-acetyl glucosamine plays in nonvaccine serotypes of streptococcus pneumoniae colonization and pathogenicity

Poly N-acetyl glucosamine is a cell surface polysaccharide that has been characterized in Staphylococcus epidermidis and Staphylococcus aureus as involved in biofilm formation and implicated in virulence. Its role in bacterial colonization and pathogenicity is now being characterized in Streptococcus pneumoniae. The aim of this thesis was to produce PNAG-deficient S. pneumoniae mutants and to confirm differential levels of biofilm formation in PNAG-deficient mutants in comparison to their wild type strains. Using PCR, gel electrophoresis, and indirect immunofluorescence, successful transformations of PNAG-deficient mutants were confirmed. Biofilm assays provided preliminary data for further investigation of the role that PNAG plays in colonization in S. pneumoniae. A novel finding in PNAG genetic structure in S. pneumoniae was also discovered, providing a new avenue of research on PNAG

Combined endophytic inoculants enhance nickel phytoextraction from serpentine soil in the hyperaccumulator Noccaea caerulescens

This study assesses the effects of specific bacterial endophytes on the phytoextraction capacity of the Ni-hyperaccumulator Noccaea caerulescens, spontaneously growing in a serpentine soil environment. Five metal-tolerant endophytes had already been selected for their high Ni tolerance (6 mM) and plant growth promoting ability. Here we demonstrate that individual bacterial inoculation is ineffective in enhancing Ni translocation and growth of N. caerulescens in serpentine soil, except for specific strains Ncr-1 and Ncr-8, belonging to the Arthrobacter and Microbacterium genera, which showed the highest indole acetic acid production and 1-aminocyclopropane-1-carboxylic acid-deaminase activity. Ncr-1 and Ncr-8 co-inoculation was even more efficient in promoting plant growth, soil Ni removal, and translocation of Ni, together with that of Fe, Co, and Cu. Bacteria of both strains densely colonized the root surfaces and intercellular spaces of leaf epidermal tissue. These two bacterial strains also turned out to stimulate root length, shoot biomass, and Ni uptake in Arabidopsis thaliana grown in MS agar medium supplemented with Ni. It is concluded that adaptation of N. caerulescens in highly Ni-contaminated serpentine soil can be enhanced by an integrated community of bacterial endophytes rather than by single strains; of the former, Arthrobacter and Microbacterium may be useful candidates for future phytoremediation trials in multiple metal-contaminated sites, with possible extension to non-hyperaccumulator plants