Drug resistance and proticinogenic types of Proteus mirabilis isolated from urinary tract infections

Proteus mirabilis strains (88 isolates) from hospitalised patients with urinary tract infection were tested for antibiotic susceptibility, ESBL production and their ability to produce proticin or on their susceptibility to proticin. Antibiotic susceptibility test was performed by standard disc diffusion method according to NCCLS. Proticin typing was made by the standard strain set from the B. W. Senior collection. Most (59%) strains belonged to ESBL producers and were more resistant to antibiotics than ESBL-negative strains. Predominant proticin patterns among the ESBL (+) strains were: P1,2(6)/SO (23%), P1,2/SO (13.5%). Among the ESBL-negative strains more frequent were P6/SO (16.6%), P1,2/SO (13.8%) and P3,6/SO (13.8%) proticin types.     

Typing ofSalmonella weltevreden strains by means of lysis patterns of symbiotic phages

A typing scheme forSalmonella weltevreden using the lysogenicity and lysis patterns of their carried phages is presented. Six strains ofS. weltevreden were selected for use as indicator strains for recognizing the lysis patterns of the carried phages. Two hundred and forty-five strains were examined and 207 were grouped in concurrence with the 15 lysis pattterns obtained out of 64 theoretically possible. Lysis pattern I (all 6 indicator strains lysed by the carried phage) included 24.5% of the strains. Thirty-eight strains (15.5%) were grouped as untypable because their lysates did not lyse any of the indicator strains. No correlation could be established between the lysis patterns of carried phages and the host and geographic distribution ofS. weltevreden.     

BOX-PCR is an Adequate Tool for Typing <Emphasis Type="Italic">Aeromonas</Emphasis> spp.

PCR-based methods of fingerprinting take advantage of the presence of repetitive sequences that are interspersed throughout the genome of diverse bacterial species. They include the repetitive extragenic palindromic (REP) sequence, the enterobacterial repetitive intergenic consensus sequence (ERIC) and the 154-bp BOX element. The combination of the three methods is used for fine discrimination of strains and is designated as rep-polymerase chain reaction (PCR). REP-PCR and ERIC-PCR have been shown to be useful for typing Aeromonas strains. To our knowledge, rep-PCR fingerprinting method using the BOXA1R primer has never been tested on aeromonads. In this study, the BOX-PCR fingerprinting technique was evaluated for the discrimination of strains of some Aeromonas species. All strains were typeable and the majority showed unique banding patterns. Four strains from culture collections were used to investigate the reproducibility of the method. According to our results, BOX-PCR fingerprinting is applicable for typing of Aeromonas strains and can be considered as a useful complementary tool for epidemiological studies of members of this genus.     

Pulsed-Field Gel Electrophoresis Is More Discriminating Than Multilocus Enzyme Electrophoresis and Random Amplified Polymorphic DNA Analysis for Typing Pyogenic Streptococci

The SmaI restriction endonuclease digestion patterns of chromosomal DNAs from 99 pyogenic streptococci belonging to Lancefield group A (41 Streptococcus pyogenes), group C (seven S. dysgalactiae, 11 \QS. equisimilis\W, three S. equi, eight S. zooepidemicus) and group G (25 human group G Streptococcus, four S. canis) were analyzed by pulsed-field gel electrophoresis (PFGE), and the results were compared with those previously obtained by multilocus enzyme electrophoresis (MLEE) and random amplified polymorphic DNA analysis (RAPD). PFGE revealed 93 distinct types among the 99 strains, and no patterns were common to strains of different species. The discriminatory power of PFGE was greater than that of MLEE and RAPD for groups A and G streptococci. The polymorphism among group C streptococci was similar with the three techniques. PFGE is, therefore, the most efficacious method for epidemiological typing of pyogenic streptococci. Received: 31 July 1996 / Accepted: 17 August 1996     

Identification and typing of Proteus penneri and Proteus vulgaris biogroups 2 and 3, from clinical sources, by computerized analysis of electrophoretic protein patterns

Seventy-six strains of the Proteus vulgaris complex ( Pr. penneri and Pr. vulgaris biogroups 2 and 3) were characterized by one-dimensional SDS-PAGE of cellular proteins. The protein patterns were highly reproducible. The strains came from various countries and were mainly of human origin: urine (28), respiratory tract (13), wounds (8), faeces (7), blood (3), miscellaneous sources (6) and unknown sources (11). The patterns of these strains, together with those of the type strains of seven Morganella, Proteus and Providencia species were subjected to two numerical analyses. In the first, in which the principal protein bands (in the 35.0–42.0 kDa range) were excluded, the strains of the Pr. vulgaris complex formed four clusters at the 83% similarity level. These corresponded to Pr. penneri, Pr. vulgaris biogroup 2, and two clusters (3a and 3b) represented biogroup 3. Each of these clusters was distinct from the Morganella, Proteus and Providencia reference strains. In the second analysis, which included all the protein bands, the 41 Pr. penneri strains showed little heterogeneity but 17 subphenons could be recognized among the 35 strains of Pr. vulgaris biogroups 2 and 3. These results support the division of biogroup 3 strains into at least two separate taxa. Other results indicate that biogroup 3 is heterogeneous and may contain further genomic groups. The method also provides a basis for typing clinical strains of Pr. vulgaris biogroups 2 and 3.     

Host specificity of Salmonella weltevreden typing phages

The host range of the six S. weltevreden typing phages was studied on 1469 strains belonging to 37 different Salmonella serotypes. In addition to S. weltevreden, only S. nchanga, S. give, S. lexington and S. anatum, all belonging to O group E₁, showed varying degrees of susceptibility to the action of some of the typing phages.Typing phage VI lysed only one strain other than S. weltevreden. All serotypes tested other than S. weltevreden were resistant to phages III and IV even at 1000 times the routine test dilution. Thus, typing phages III and IV were specific for S. weltevreden. The sensitivity patterns of S. weltevreden typing phages were not found to bear much correlation with either somatic of flagellar antigens of Salmonellae.     

Typing of Proteus mirabilis from clinical sources by computerized analysis of electrophoretic protein patterns

B. HOLMES, M. COSTAS AND A.C. WOOD. 1991. Seventy-five strains of Proteus mirabilis (mainly of human origin) were characterized by one-dimensional SDS-PAGE of cellular proteins. The strains came from various countries; 24 were from urine, 17 from faeces (including one from a dog and one from a monkey), 12 from the respiratory tract (including nine from sputum), four from a cerebral abscess, 12 from miscellaneous sources and six from unknown sources. The protein patterns, which contained 45 to 50 discrete bands, were highly reproducible. The patterns of the 75 P. mirabilis cultures plus those of the type strains of six Proteus and Providencia species were used as the basis for two numerical analyses. In the first, which included all the protein bands, the P. mirabilis cultures formed nine clusters at the 85% S level. In the second analysis, in which the principal protein bands (in the 34.0–44.6 kDa range) were excluded, 74 of the 75 cultures of P. mirabilis formed a single cluster at the 77% S level distinct from the six Proteus and Providencia reference strains. The P. mirabilis strain which failed to cluster with the others had a background band pattern typical of the species in the lower molecular weight region but appeared to be less typical in the heavier bands. It is concluded that high resolution PAGE combined with computerized analysis of protein patterns provides the basis for typing clinical strains of P. mirabilis. Reference strains of each of the nine PAGE types identified are available from NCTC for inclusion in future studies.     

Molecular intraspecific characterization of Photobacterium damselae ssp. damselae strains affecting cultured marine fish

Aims: The aim of this study was to analyse the intraspecific variability of Photobacterium damselae ssp. damselae strains isolated from different cultured marine fish species using molecular typing methods. Methods and Results: Twenty P. damselae ssp. damselae strains isolated from marine fish species were used in this study. Phenotypic characterization of the strains was carried out using standard microbiological methods. Genetic characterization was conducted using three PCR‐based methods [random amplified polymorphic DNA (RAPD), enterobacterial repetitive intergenic consensus‐PCR (ERIC‐PCR) and repetitive extragenic palindromic‐PCR (REP‐PCR)]. Dice coefficient and the unweighted pair group method with average linkage were used for numerical analyses of banding patterns. At phenotypic level, the strains analysed showed seven different profiles, which could not be related to the host fish species, geographic area or outbreak of disease. Isolates were grouped into nine and eight clusters using the RAPD technique with primers 5 and 4, respectively. In both cases, the main cluster grouped 45% of strains. The techniques ERIC‐PCR and REP‐PCR were more discriminatory, both resulting in 14 different clusters, which grouped 15–20% of the isolates. Conclusions: In this study, the techniques tested are confirmed as good tools for molecular typing, because they allow discrimination between P. damselae ssp. damselae strains isolated within the same outbreak. In addition, ERIC‐PCR and REP‐PCR methods were more adequate for rapid typing of P. damselae ssp. damselae than RAPD, allowing the discrimination at strain level. Significance and Impact of the Study: The results, in agreement with previous studies, confirmed the high intraspecific variability among isolated P. damselae ssp. damselae strains at both phenotypic and genetic levels. This suggests the existence of different clonal lineages that coexist in the same geographic area, within a short period of time (2–3 years). The discrimination at strain level can be useful to study the traceability of infections.     

A new bacteriophage typing scheme forProteus mirabilis andProteus vulgaris strains

The lytic properties of 21 bacteriophages constituting a new typing set forProteus were examined in 507Proteus mirabilis and 29P. vulgaris strains isolated from patients and healthy subjects. Comparison of their morphological, serological, genetic and lytic properties showed that, in theMyoviridae andPodoviridae families, some phages were so closely related that the presence of all of them in the set was redundant. Analysis of the lytic properties revealed that some of the bacteriophages were not active enough to facilitate the differentiation ofProteus strains The size of the final typing set was reduced from 21 to 12 phages but it was suggested that, in order to improve the differentiation capacity of the set, new phages should be included. Second part:Folia Microbiol. 41, 137–140 (1996).     

Population growth dynamics of the rotifer Brachionus plicatilis cultured in non-limiting food condition

Population growth parameters in batch culture of Brachionus plicatilis under a continuous supply of freeze-dried microalgae powder have been determined. Two B. plicatilis strains (L- and S-types) and four microalgae species (Nannochloropsis oculata, Nannochloropsis gaditana, Nannochloris oculata and Tetraselmis suecica) have been tested, establishing the dynamics of growth at different daily food rations. Cultures showed a short lag phase, an exponential growth phase, a long post-exponential growth phase and long decline with episodic increases. In both rotifer strains, the best growth was obtained with Nannochloropsis oculata and the poorest with Nannochloris oculata.     

Characterization of a bacteriocin from Pediococcus acidilactici PC and comparison of bacteriocin-producing strains using molecular typing procedures

Summary Sixteen pediococcal strains, including eleven Pediococcus acidilactici and five P. pentosaceus strains were screened for inhibitory potential using a deferred overlay spot method against a limited collection of foodborne pathogens. Of those screened, P. acidilactici PC, an organism isolated from fermented sausage, was effective and subsequently screened for inhibitory potential against 46 foodborne pathogens and 28 other lactic acid bacteria. Strain PC produced an antimicrobial agent capable of inhibiting members of the genera Listeria, Clostridium, Leuconostoc and Pediococcus. Gram-negative microorganisms from seven genera, Lactococcus, Streptococcus and Lactobacillus strains were unaffected by the inhibitory substance. The inhibitory agent was sensitive to proteolytic enzymes and exhibited a bactericidal mode of action, confirming the identity as a bacteriocin. In addition, the partially purified bacteriocin was thermally stable up to 100°C for 60 min and maintained inhibitory potential over a wide range of pH values. Plasmid curing studies suggested linkage of bacteriocin production to a 5.5-MDa plasmid. Plasmid profiles were identical for P. acidilactici PC, PAC1.0 and PO2. Genetic analysis of total genomic DNA via DNA fingerprinting and ribosomal RNA (rRNA) typing provided further evidence that these strains were identical. DNA fingerprinting and rRNA typing also showed utility in discrimination between and within other species of pediococci.Published as paper no. 19574 of the contribution series of the Minnesota Agricultural Experiment Station Correspondence to: S. Harlander     

Combined Use of Ribotyping,PFGE Typing and IS431 Typing in the Discrimination of Nosocomial Strains of Methicillin-Resistant Staphylococcus aureus

We have previously reported the phenotypic characterization of methicillin-resistant Staphylococcus aureus (MRSA) clinical strains isolated in Malaya University Hospital in the period 1987 to 1989 using antibiogram, coagulase typing, plasmid profiles, and phage typing. Here, we report the analysis of the same strains with three genotyping methods; ribotyping, pulsed-field gel electrophoresis (PFGE) typing, and IS431 typing (a restriction enzyme fragment length polymorphism analysis using an IS431 probe). Ribotyping could discriminate 46 clinical MRSA strains into 5 ribotypes, PFGE typing into 22 types, and IS431 typing into 15 types. Since the differences of the three genotyping patterns from strain to strain were quite independent from one another, the combined use of the three genotyping methods could discriminate 46 strains into 39 genotypes. Thus, the powerful discriminatory ability of the combination was demonstrated.     

Improvement of strain discrimination by combination of RAPD with PFGE for the analysis of the swine isolates of Salmonella enterica serovar Choleraesuis

Salmonella enterica serovar Choleraesuis (S. Choleraesuis) can cause salmonellosis in pigs and humans. Currently, the most common method used for the subtyping of this Salmonella serovar is pulsed field gel electrophoresis (PFGE) using XbaI as a DNA digestion enzyme. In this study, we compared and combined PFGE with the randomly amplified polymorphic DNA method, for the typing of 95 S. Choloraesuis strains isolated from diseased pigs. Using PFGE with XbaI, with AvrII, and with SpeI digested DNA, 29, 74, and 40 patterns, respectively, were obtained. Also, 53, 15, and 35 strains, respectively, belong to the major patterns X1, A1, and S1. When these three digestion patterns were combined, 83 PFGE pattern combinations were obtained. On the other hand, using RAPD with selected primer alone generated 76 patterns, and 11 strains which fell within a single X1A1S1 PFGE combination pattern were discriminated into 10 patterns. Thus, for S. Choloraesuis, PFGE with AvrII allowed higher discrimination than PFGE with XbaI, and some of the PFGE groupings obtained by combining the XbaI, AvrII and SpeI digestion patterns were further subdivided by the RAPD method.     

Typing of Proteus mirabilis from clinical sources by computerized analysis of electrophoretic protein patterns.

Seventy-five strains of Proteus mirabilis (mainly of human origin) were characterized by one-dimensional SDS-PAGE of cellular proteins. The strains came from various countries; 24 were from urine, 17 from faeces (including one from a dog and one from a monkey), 12 from the respiratory tract (including nine from sputum), four from a cerebral abscess, 12 from miscellaneous sources and six from unknown sources. The protein patterns, which contained 45 to 50 discrete bands, were highly reproducible. The patterns of the 75 P. mirabilis cultures plus those of the type strains of six Proteus and Providencia species were used as the basis for two numerical analyses. In the first, which included all the protein bands, the P. mirabilis cultures formed nine clusters at the 85% S level. In the second analysis, in which the principal protein bands (in the 34.0-44.6 kDa range) were excluded, 74 of the 75 cultures of P. mirabilis formed a single cluster at the 77% S level distinct from the six Proteus and Providencia reference strains. The P. mirabilis strain which failed to cluster with the others had a background band pattern typical of the species in the lower molecular weight region but appeared to be less typical in the heavier bands. It is concluded that high resolution PAGE combined with computerized analysis of protein patterns provides the basis for typing clinical strains of P. mirabilis. Reference strains of each of the nine PAGE types identified are available from NCTC for inclusion in future studies.     

A Simple and Practical Method for Typing and Strain Differentiatin of Herpes Simplex Virus Using Infected Cell DNAs

A simple and practical method for typing and strain differentiation of herpes simplex virus (HSV) isolates, based upon analysis of the restriction endonuclease cleavage patterns of viral DNAs, was established by using unlabeled infected cell DNAs. The preparation of infected cell DNA is technically easier than that of purified viral DNA or of radiolabeled viral DNA. The method provides a powerful and practical tool for epidemiological and clinical studies of HSV infection, which can be performed in most diagnostic laboratories. In order to select suitable restriction endonucleases for the study of HSV isolates, the cleavage patterns of viral DNAs (strains MacIntyre, HF, UW-268, and SAV) with 12 enzymes were analyzed. Several enzymes, Bam HI, Kpn I, Pst I, Sal I, Sst I, and Xho I, were found to be useful for both typing and strain differentiation. With this method, HSV isolates from different individuals and from the same individual were analyzed by digestion of their infected cell DNAs with Bam HI. Six isolates from epidemiologically unrelated individuals were readily typed and differentiated from each other. Three isolates from the same individual showed very similar patterns. However, there was a small degree of difference between the first two isolates and the third isolate.     

2016-2017年宁夏地区牛源金黄色葡萄球菌的全基因组分析

[目的] 了解宁夏地区奶牛乳腺炎金黄色葡萄球菌（Staphylococcus aureus,SA）代表菌株的基因组序列基本特征,进一步探究其耐药基因型、毒力及进化关系,为兽医临床防治提供理论依据。[方法] 采用纸片法对97株金黄色葡萄球菌临床分离株进行抗菌药物敏感性试验,同时进行葡萄球菌蛋白A（Staphylococcus aureus protein A,spa）分型、多位点序列分型（multilocus sequence typing,MLST）,根据分型结果选取16株代表菌株进行全基因组测序,并对获得的测序序列进行处理分析。[结果] 药敏试验结果显示97株分离株对18种抗菌药物存在不同程度的耐药,其中9株耐甲氧西林金黄色葡萄球菌（methicillin-resistantStaphylococcus aureus,MRSA）对青霉素、氨苄西林、苯唑西林、头孢噻呋、磺胺异噁唑、红霉素、庆大霉素和克林霉素等8种抗菌药物完全耐药,甲氧西林敏感金黄色葡萄球菌（methicillin-sensitiveStaphylococcus aureus,MSSA）菌株对青霉素、氨苄西林、磺胺异噁唑耐药率较高。耐药基因数据库（antibiotic resistance genes database,ARDB）注释分析显示16株代表菌株共携带21种耐药基因,其中norA、tet38、bacA、mepA的携带率较高,与药敏试验结果具有一定的相关性。毒力基因数据库（virulence factors of pathogenic bacteria,VFDB）注释分析显示所有菌株携带多种与粘附、宿主免疫逃逸、分泌、胞外酶编码、铁摄取等疾病相关的毒力基因,MRSA菌株均携带较多毒力因子,MSSA菌株携带毒力因子数目不等。基因岛预测结果显示16株代表菌株存在不同数量的基因岛且MRSA菌株携带基因岛数目及毒力基因岛较多,但耐药基因岛数目与MSSA差异不明显。SNP分析结果显示部分分离株同源性较高,同源性较高的两株MRSA的全基因组基本序列特征差异较小,携带的耐药、毒力基因情况相似。[结论] 宁夏地区牛源SA分离株耐药性情况严重且具有较高的毒力水平,本研究为家畜相关MRSA（livestock-associated MRSA,LA-MRSA）与MSSA基因组序列信息的比较分析及宁夏地区SA感染的临床防控提供参考依据。     

Diversity of DNA methylation pattern and total DNA restriction pattern in symbiotic Nostoc

Attempts were made to use total DNA restriction patterns and the response of purified DNA to treatment with restriction endonucleases to characterize several symbiotic Nostoc strains which had been isolated from different host plants cultivated in Italy. Among 27 restriction endonucleases tested, several did not cut any DNA and no significant variation in the susceptibility of the genomes to DNA restriction was seen among the strains. Therefore the Nostoc strains could not be separated into groups based on their different susceptibilities to the action of restriction endonucleases. However, in studies of total DNA restriction patterns, the restriction endonucleases BfrI and HpaI gave unique band patterns for each cyanobacterial isolate. Different profiles were even found in strains isolated from host plants belonging to the same species. The results do not support any definition of symbiotic Nostoc genomic groups or species and show that a tight specificity between the host plant and the cyanobacterium might not exist in the symbiotic associations involving Nostoc.     

Genomic DNA Fingerprinting of Oenococcus oeni Strains by Pulsed-Field Gel Electrophoresis and Randomly Amplified Polymorphic DNA-PCR

Genetic diversity of 60 Oenococcus oeni strains from different wines was evaluated by numerical analysis of (i) pulsed-field gel electrophoresis (PFGE) patterns with endonuclease ApaI and (ii) randomly amplified polymorphic DNA (RAPD)-PCR fingerprints with four oligonucleotide primers. Sixty-two percent of the strains could be distinguished by PFGE, whereas most strains were identified by distinct RAPD-PCR profiles and associated according to the geographical origin. Because of its rapidity and reliability, RAPD-PCR appeared to be a suitable method for typing and monitoring O. oeni strains in winemaking. Received: 3 November 1999 / Accepted: 8 December 1999     

Molecular Typing of Staphylococcus epidermidis and Other CNS with Repetitive Element Sequence-Based PCR

The chromosomal distribution of the repetitive DNA sequence found in Mycoplasma pneumoniae (REP-MP2) provides an ideal target for detecting DNA fragment patterns specific to individual Staphylococcus epidermidis and S. haemolyticus strains. A REP-MP2 sequence-based PCR (rep-PCR) was developed and applied to CNS isolates. We identified a 450 bp genomic DNA fragment which was common and specific to S. epidermidis isolates and not found in other CNS. In addition, S. epidermidis isolates showed several bands that could be grouped into 14 different fragment patterns. Similarly, S. haemolyticus isolates were classified into 10 groups. Significant correlations between the typing patterns of S. epidermidis and resistance to oxacillin (P<0.05), gentamicin (P<0.01), erythromycin (P<0.02), and sulfamethoxazole-trimethoprim (P<0.001) were found. The rep-PCR method is a rapid and reproducible discriminatory means for molecular typing of S. epidermidis and other CNS.     

1型CRISPR位点间区序列分析在嗜热链球菌菌株分型中的应用

为了建立适用于嗜热链球菌菌株资源多样性调查的菌株分型方法,尝试将1型CRISPR位点间区序列分析用于嗜热链球菌的菌株分型,并与常用ERIC-PCR指纹图谱方法进行了比较。结果表明,1型CRISPR位点间区序列分析可以把30株从三个不同样品中分离的嗜热链球菌分成三种差异明显的类型:不同类型菌株之间没有相同的间区序列;而同一类型菌株之间,间区序列的组成和排列则基本一致,并且上述分型的结果与用ERIC-PCR指纹图谱技术获得的结果完全一致。因此,1型CRISPR位点间区序列分析是嗜热链球菌分型鉴定的可靠方法,并适用于大量菌株的分型鉴定和多样性调查。