Antimicrobial activity among Pseudomonas and related strains of mineral water origin

Siderophore production in 382 Pseudomonas and related strains of mineral water origin were screened and the antimicrobial activities of 158 of these tested against nine target organisms of health significance. Presence of siderophores could be detected in 54·4% and the majority of strains tested (91·2%) inhibited at least one of the nine target strains. Staphylococcus, Escherichia coli and Aeromonas hydrophila were particularly sensitive. Addition of iron eliminated the inhibitory activity in 96·7% of cases ; the antagonistic effect should be largely determined by siderophore-mediated competition for iron. Most of the inhibitory strains produced siderophores, whereas the non-inhibitory strains did not. Few strains also produced bacteriocins showing activity against Pseudomonas aeruginosa and Aer. hydrophila. Strains isolated from mineral water have a broad antibacterial potential.     

山东地区钾矿物分解细菌的分离及生物学特性

不同土壤类型钾矿物分解细菌资源调查和高效稳定释钾、促生细菌的筛选鉴定有助于丰富微生物资源库,发掘和利用钾矿物分解细菌以及探究矿物生物风化机理等。作者采用以钾长石为唯一钾源的选择性细菌培养基, 从山东地区不同土壤和不同植物根际土壤中分离纯化了23株生长势良好的钾矿物分解细菌, 通过测定细菌代谢产物IAA和铁载体,研究其产生促生物质的能力, 通过摇瓶试验筛选高效释钾菌株, 采用16S rDNA限制性酶切多态性分析(amplified rDNA restriction analysis, ARDRA)方法研究了钾矿物分解细菌的遗传多样性, 根据16S rDNA同源性对高效释钾菌株进行了鉴定。结果表明, 供试菌株均产吲哚乙酸或其衍生物, 43.5%的分离菌株产极高量铁载体。ARDRA结果表明供试菌株在60%相似性水平上可分为11个基因型, 同一类型土壤上不同作物根际或不同类型土壤上同一作物根际的钾矿物分解细菌存在明显的遗传差异。摇瓶试验结果表明供试菌株中具有较显著释钾能力的菌株占17%, 39%的供试菌株无释钾能力。筛选到2株高效释钾菌株AFM2、AC2, 分别使溶液中钾含量增加了29.8%和25.4%。16S rDNA同源性分析表明菌株AC2、AHZ1与Bacillus mucilaginosus聚为一群, 该群与包含菌株AZH4的Paenibacillus sp.中的种聚为一大发育分支, 该分支在细菌分类地位上隶属于Firmicutes; 菌株AFM2与Rhizobium sp. 和Agrobacterium tumefaciens聚为另一大发育分支, 该分支在细菌分类地位上隶属于Alphaproteobacteria。     

Restriction endonucleases from various bacterial strains exhibit an ice-nucleating activity

Six strains containing site-specific endonucleases II were selected from a collection of 45 ice-nucleating bacterial strains isolated from rhizosphere of plants growing in various geographical regions. Endonucleases Pfl211I, Psp8I, and Psp23I were isolated and purified from two Pseudomonas sp. strains and a Pseudomonas fluorescens strain. Restriction endonucleases Pfl21I and Psp23I were shown to recognize and cleave the DNA nucleotide sequence 5'-CTGCA decrease G-3'. Endonuclease Psp8I recognized and cleaved the DNA nucleotide sequence 5'-G decrease GATCC-3'. These endonucleases were found to be true isoschizomers of PstI and BamHI, respectively.     

Restriction endonucleases from various bacterial strains displaying an ice-nucleating activity

Six strains containing site-specific endonucleases II were selected from a collection of 45 ice-nucleating bacterial strains isolated from rhizosphere of plants growing in various geographical regions. EndonucleasesPft211I,Psp8I, andPsp23I were isolated and purified from twoPseudomonas sp. strains and aPseudomonas fluorescens strain. Restriction endonucleasesPfl2lI andPsp23I were shown to recognize and cleave the DNA nucleotide sequence 5′-CTGCA↓G-3′. Endonuclease Psp81 recognized and cleaved the DNA nucleotide sequence 5′-G↓GATCC-3′. These endonucleases were found to be true isoschizomers of PstI andBamHI, respectively.     

Survival of Campylobacter jejuni strains of different origin in drinking water

AIMS: The aim of the study was to measure the survival of 19 Campylobacter jejuni strains of different origins, including two reference strains, four poultry-derived isolates, nine human isolates and four water isolates, in sterilized drinking water. METHODS AND RESULTS: Pure cultures of 19 C. jejuni strains were inoculated in sterile drinking water and incubated at 4 degrees C for 64 days. Survival was determined by culturability on both selective (Karmali agar) and non-selective [Columbia blood agar (CBA)] media. Culturability was shown to be strain and origin-dependent. Campylobacter jejuni showed prolonged survival on a non-selective than on a selective medium. CONCLUSIONS: The origin of the strain is a determining factor for the survival of C. jejuni in drinking water at 4 degrees C. Poultry isolates showed a prolonged survival, which could be an indication that these strains could play an important role in the transmission of campylobacteriosis through water. In addition, culture conditions are an important factor for evaluating the survival of C. jejuni in drinking water at 4 degrees C. The non-selective agar (CBA) allowed growth of C. jejuni over a longer period of time than the selective agar (Karmali). Furthermore, an enrichment broth (Bolton) allowed the recovery of all 19 C. jejuni strains during the 64 days of incubation at 4 degrees C. SIGNIFICANCE AND IMPACT OF THE STUDY: This study highlighted differences in culturability depending on culture conditions and on strain origin.     

Molecular fingerprinting of bacterial populations in groundwater and bottled mineral water

Monitoring the hygienic quality of drinking waters by determining the concentration of fecal indicators with traditional plate count techniques suffers from important drawbacks. In this work, the potential of PCR-DGGE (polymerase chain reaction - denaturing gradient gel electrophoresis) analysis of 16S rDNA genes to fingerprint the bacterial populations of mineral water and groundwater was investigated. A rapid and simple pretreatment to concentrate and release bacterial DNA prior to PCR was explored. This pretreatment was successful for commercially bottled mineral water. For groundwater, an additional resuscitation step was required to obtain a PCR signal. It was clear that the groundwater under scrutiny contained a more diverse bacterial community than the mineral water. A comparison was made between four kinds of mineral waters and one sample of groundwater using the developed procedures. For each kind of water, bacterial populations cultured on R2A plates were also subjected to PCR-DGGE. Comparison of the fingerprints of the plated samples and the original samples suggested the presence of viable but nonculturable bacteria in the waters. The obtained cluster dendrogram indicated that each kind of water was characterized by a specific molecular fingerprint. The sensitivity of the whole of the procedure was between 10(4) and 10(5) cfu ml(-1) as determined using a pure culture of Escherichia coli. The described PCR-DGGE method can constitute the basis of a new and interesting strategy to monitor in a relatively rapid way (less than 24 h) the bacterial quality of waters such as mineral water, groundwater and certain types of reclaimed water.     

Antibiotic susceptibility of bacterial strains isolated from patients with various infections

AIMS: Isolates from various samples obtained during 1998 and 1999 were identified and their susceptibility to third-generation cephalosporins, monobactams and/or cephamycins studied along with any production of ESBLs. METHODS AND RESULTS: Of these samples, bacteria most frequently isolated by the conventional techniques and Vitek GNI card were Escherichia coli (37%), Klebsiella pneumoniae (27%) and Enterobacter cloacae (16%). Using disk diffusion and double-disk synergy tests, we found that 71% strains produced ESBLs and 18% strains produced ESBLs and cephamycinases. Banding patterns of PCR amplification with the designed primers showed that 57% strains were capable of harbouring bla(SHV) genes. The bla(TEM), bla(CMY) and bla(AmpC) genes were harboured by 55%, 31% and 12% strains, respectively. Forty-five percent of strains contained more than two types of beta-lactamase genes. In particular, one strain contained bla(TEM), bla(SHV), bla(CMY) and bla(AmpC) genes. CONCLUSIONS: The percentage of ESBL-producing strains was high. The most prevalent beta-lactamase gene was bla(SHV) gene. The bla(CMY) genes have been prevalent in cephamycin-resistant strains. The multidrug-resistant strains resistant to third-generation cephalosporins and cephamycins were detected in high percentage. SIGNIFICANCE AND IMPACT OF THE STUDY: Resistance mechanisms to beta-lactams, comprising mostly extended-spectrum beta-lactamase (ESBL) production, lead to the resistance against even recently developed beta-lactams in enterobacteria, which is now a serious threat to antibiotic therapy. The high prevalence of bla(CMY) genes and multidrug-resistant genes may also cause therapeutic failure and lack of eradication of these strains by third-generation cephalosporins or cephamycins.     

Characterization of bacterial pectinolytic strains involved in the water retting process

Pectinolytic microorganisms involved in the water retting process were characterized. Cultivable mesophilic anaerobic and aerobic bacteria were isolated from unretted and water-retted material. A total of 104 anaerobic and 23 aerobic pectinolytic strains were identified. Polygalacturonase activity was measured in the supernatant of cell cultures; 24 anaerobic and nine aerobic isolates showed an enzymatic activity higher than the reference strains Clostridium felsineum and Bacillus subtilis respectively. We performed the first genotypic characterization of the retting microflora by a 16S amplified ribosomal DNA restriction analysis (ARDRA). Anaerobic isolates were divided into five different groups, and the aerobic isolates were clustered into three groups. 84.6% of the anaerobic and 82.6% of the aerobic isolates consisted of two main haplotypes. Partial 16S rRNA gene sequences were determined for 12 strains, representative of each haplotype. All anaerobic strains were assigned to the Clostridium genus, whereas the aerobic isolates were assigned to either the Bacillus or the Paenibacillus genus. Anaerobic isolates with high polygalacturonase (PG) activity belong to two clearly distinct phylogenetic clusters related to C. acetobutylicum-C. felsineum and C. saccharobutylicum species. Aerobic isolates with high PG activity belong to two clearly distinct phylogenetic clusters related to B. subtilisT and B. pumilusT.     

Substrate affinity from bacterial strains and distribution water biofilms

Biodegradable dissolved organic carbon (BDOC) is almost the only limiting factor of bacterial growth in drinking water. Two representative bacterial strains were isolated from a two-column reactor for the determination of BDOC (from respectively the first and the second column) and used to estimate their growth kinetics in two different types of water: finished water from a water treatment plant and water collected at the outlet of the first column of the BDOC reactor. K _s, the substrate saturation constant, was calculated by analogy with Michaelis-Menten enzyme kinetics, and for each strain a lower K _s value was observed for the type of water that it normally receives.
K _s for in situ biofilms, from both first and second column and the same types of water, were also calculated according to a technique which uses a curve-matching method with kinetic results from the non-steady-state experiments. In this case, the results were similar for the first and second column biofilms. However, a higher affinity was observed for finished water than for the water from the outlet of the first column in both biofilms.     

Short Communication: Leaching of Chromite overburden with various native bacterial strains

Four heterotrophic bacterial strains were isolated from soil and water samples from a chromite mine in Sukinda valley in Orissa. They were identified as Lactobacillus acidophilus, Staphylococcus lactis, Lactobacillus sp. and Propionibacterium acnes. Each strain was used for leaching of metals from chromite overburden of Sukinda. Comparatively, Lactobacillus sp. showed maximum capability for metal solubilization. It leached a maximum of 5.98% of nickel in 35 days of contact time.     

Study of the bacterial flora of a non-carbonated natural mineral water

Natural mineral water from a UK spring was monitored at various stages after it was pumped from the ground, through to bottling and during shelf life before consumption. Samples were collected in commercial PVC bottles, in PVC bottles previously sterilized and hand-filled and in glass bottles. The bacterial flora was counted on plate count agar (PCA) and on PCA diluted 10 times (PCA/10). The predominant bacteria were identified to genus level. Growth rates and nutrient types of isolates were determined by the nutrient-tolerance test (NT). The plate counts at the prebottling stage were low. During storage larger numbers of bacteria grew in glass than PVC bottles; the largest number grew in PVC bottles filled by hand. Most of the pigmented bacteria isolated were oligocarbotolerant.     

Evaluation of the specificity of Salmonella PCR primers using various intestinal bacterial species

AIMS: Nine sets of PCR primers targeting Salmonella were evaluated for their specificity with pure cultures of intestinal-associated bacteria prior to their application to Salmonella detection in faecal samples. METHODS AND RESULTS: Gene targets of PCR primers included: 16S rDNA, a Salmonella pathogenicity island I virulence gene, Salmonella enterotoxin gene (stn), invA gene, Fur-regulated gene, histidine transport operon, junction between SipB and SipC virulence genes, Salmonella-specific repetitive DNA fragment, and multiplex targeting invA gene and spvC gene of the virulence plasmid. Fifty-two Salmonella strains were used to determine sensitivity; five strains from related genera and 45 intestinal bacteria were used to evaluate specificity. All primers amplified DNA from Salmonella strains, although two primer sets failed to amplify Salmonella DNA from either Salmonella bongori (hilA) or subgroups VI or VII (16S rDNA). There was no detected amplification of DNA from related bacterial genera with any of nine PCR assays. Six of the PCR assays amplified DNA for some intestinal bacteria. CONCLUSIONS: Only three primer pairs were determined to be suitable for application of PCR amplification of Salmonella in faecal samples - 16S rDNA, stn and histidine transport operon. We are currently evaluating their sensitivity of detection of Salmonella in faecal samples. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrated the importance of internal lab validation of PCR primers prior to application to the type of samples of interest. Information from this evaluation can be applied in other labs to facilitate choosing Salmonella PCR primers.     

Study of the bacterial flora of a non-carbonated natural mineral water.

Natural mineral water from a UK spring was monitored at various stages after it was pumped from the ground, through to bottling and during shelf life before consumption. Samples were collected in commercial PVC bottles, in PVC bottles previously sterilized and hand-filled and in glass bottles. The bacterial flora was counted on plate count agar (PCA) and on PCA diluted 10 times (PCA/10). The predominant bacteria were identified to genus level. Growth rates and nutrient types of isolates were determined by the nutrient-tolerance test (NT). The plate counts at the pre-bottling stage were low. During storage larger numbers of bacteria grew in glass than PVC bottles; the largest number grew in PVC bottles filled by hand. Most of the pigmented bacteria isolated were oligocarbotolerant.     

Comparative study of the homology of DNA in thermophilic and mesophilic lactococcal strains of various origin

A comparative study of DNA homology among 12 strains of thermophilic streptococci currently used as ferments in the dairy industry in different regions of the CIS and 16 strains of mesophilic lactococci obtained from different sources was performed by the method of optical reassociation in solution. These strains were found to form three groups with DNA homology generally not exceeding 30-35% between each other. These data indicate that strains commonly classified as Streptococcus thermophilus may belong to different taxa. At the same time, the DNA base composition was similar in all these strains (the G + C content was 38-40 mol %). Among 16 strains of mesophilic lactococci DNA homology was generally higher than 50%, i.e., all these strains indeed belong to the same species, Lactococcus lactis.