The diversilab system is an automated platform that uses reppcr technology to provide standardized, reproducible dna fingerprinting and analysis of bacterial and fungal samples for complete isolate characterization. Comparison of a commercially available repetitiveelement. Yeast identification by sequencing, biochemical kits. Diversilab rep pcr used two different primer mixes for typing aspergillus and other moulds which resulted in all a. The costs of routinely performing pcr ribotyping with analysis of the pcr products using a diversilab chip compared to automated rep pcr diversilab is outlined in table 3. As already observed with mlst, the concordance of dl with pfge was low for all three ccs wallace coefficient rep pcr. The diversilab system, which includes microfluidicsbased detection, reagent kits, and software for data processing and analysis, is an automated method using repetitive sequencebased pcr reppcr for microbial strain typing. Repetitivesequencebased pcr using the diversilab system. Automated discrimination of mrsa and mssa using rep pcr.
The bacterial kit is designed to be used on all gram positive and gram negative bacteria, however it cannot. Conventional morphology versus pcr sequencing, rep pcr, and malditofms for identi. Interlaboratory reproducibility of diversilab rep pcr typing and clustering of acinetobacter baumannii isolates article pdf available in journal of medical microbiology 61pt 1. Comparison of an automated repetitive sequencebased pcr. It can perform same day strain typing for real time investigations. The diversilab system generated rep pcr dna fingerprints for each of the 26 fusarium isolates, and the similarity comparisons are shown as a dendrogram fig. Around 40% of the isolates expressed serotypes o6 20.
The diversilab salmonella kit was used for reppcr amplification of intergenic repetitive elements in the genomic dna. Diversilab, which is one of those methods, is a commercial repetitivesequencebased pcr reppcr typing system that amplifies strainspecific noncoding repetitive sequences. Pearsons correlation coefficient was used to create a pairwise percentage similarity. The automated the automated detection and standard algorithms for data analysis adds consistency to this molecular. Use of the diversilab semiautomated repetitivesequence. The system is particularly useful for bacterial genotyping and is used for source tracking of hospital and community infection, contamination and epidemiological surveillance. Interlaboratory reproducibility of diversilab reppcr.
Repetitivesequencepcrbased dna fingerprinting using the. Comparison of a commercially available repetitiveelement pcr system diversilab with pcr ribotyping for typing of clostridium difficile strains. Repetitivesequencebased pcr reppcr using the diversilab system was investigated for identification of aspergillus. Diversilab software, which has unlimited, passwordprotected access from any computer with an internet connection, generates userfriendly reports that can be customized for the testing laboratory. These kits contain the reagents and buffers needed to set up pcr reactions using rep pcr primers, starting from extracted genomic dna from microbial isolates. Alternative versus gold standard method for the typing of. Pcr the diversilab system and reported that cattle and wildlife were the main sources of e. It has more advantages than reppcr, such as higher reproducibility, standardization, and archiving fingerprinting patterns and reporting 3,1012. Based on patented reppcr technology, microbial isolates can quickly and. Recently, reppcr has been commercially adapted to an automated format known as the diversilab system to provide a reliable pcrbased typing system for clinical laboratories. Dna was extracted and genomic dna was amplified using the diversilab mycobacterium typing kit. Mitis salivarius with bacitracin msb, golds is a selective media commonly used for the isolation of streptococcus mutans sm, an organism associated with dental caries.
Dl bacteria kit 24 test dl reagents and consumables dl. On the basis of an 80 % similarity threshold, rep pcr consistently clustered like species and this set of isolates, along with five atcc reference strains, was used to create a dna fingerprint library with the diversilab software. For analysis, diversilab software utilized the pearson correlation coef. Use of the diversilab semiautomated repetitivesequencebased polymerase chain reaction for epidemiologic analysis on acinetobacter baumannii isolates in different italian hospitals. Diversilab was compared to pcr ribotyping of the 16s23s rna intergenic spacer of clostridium difficile cd as the gold standard method for cd typing. The diversilab system exploits the discriminatory power of reppcr. This media is currently being used in a largescale epidemiological study to genotype sm using repetitive extragenic palindromic pcr rep pcr. The resulting reppcrbased dna fingerprints were uploaded to and analysed using webbased diversilab software through pearsons correlation coefficient. Molecular fingerprinting of candida and aspergillus.
The automated diversilab system provides high performance genotypic characterization of bacteria, yeast and mold. Diversilab system dl biomerieux, marcy etoile, france was introduced a semiautomated reppcr system. The genomic profile, determined by diversilab system rep pcr based technique, was compared with the p. The reppcr technology and reppcr primers are covered by u. Antimicrobial resistance and genomic reppcr fingerprints. Species identification and strain differentiation of. Molecular fingerprinting of clinical candida isolates. Ninetyfive clinical isolates, identified by conventional methods, and five atcc strains were tested. Pfge was performed on 31 isolates representative of the most relevant dlrpcs clustered within the 3 major mlst ccs cc17, cc23 and cc1. Laurent, quebec, canada utilizing a 2100 bioanalyzer agilent technologies, santa clara, ca operating the diversilab v1. These kits contain the reagents and buffers needed to set up pcr reactions using reppcr primers, starting from extracted genomic dna from microbial isolates. After pcr amplification of genomic dna regions between noncoding intergenic repetitive elements, amplified fragments are separated by microfluidic electrophoresis using agilent technology.
Six and eight distinct fingerprinting types were found with primer mixes for. Similarly, vogel and colleagues 2007 used automated rep. Predicting salmonella enterica serotypes by repetitive. Comparative evaluation of an automated repetitivesequence. Pdf yeast identification by sequencing, biochemical kits.
The diversilab reppcr assay is a rapid, semiautomated method based on pcr amplification of specific regions between noncoding repetitive sequences in the bacterial genome. Using a commercial diversilab semiautomated repetitive. The dendrograms were generated automatically using the diversilab software and pairwise groupings with the aspergillus dna fingerprint patterns. The diversilab system biomerieux is an automated dna fingerprinting system based on the reppcr electrophoresis technique. Figure figure1 1 shows rep pcr profiles of the aspergillus isolates. Routine molecular typing using rep pcr would not only increase annual material costs by 32%, but it would also require an additional 0. Evaluation of automated repetitivesequencebased pcr. Use of the diversilab system for species and strain. Osf repetitive extragenic palindromic pcr reppcr of. The rep pcr results apparently suggested a higher level of genetic variability, and both the dendrogram and the similarity matrix generated by the diversilab software fig.
Molecular characterization of multidrugresistant klebsiella pneumoniae isolates xianghua hou1, xiuyu song2, xiaobo ma2, shiyang zhang3,4, jiaqin zhang2,3 1department of nephrology, the first affiliated hospital of xiamen university, xiamen, china. The amplified fragments of various sizes and fluorescence intensities were separated and detected using a microfluidics chip with an agilent 2100 bioanalyzer. Diversilab fingerprinting kits previously called bacterial bar codes, a type of repetitiveelement pcr rep pcr, is a rapid, semiautomated, pcr based, commercial typing system that can be easily performed by bench technologists with some molecular experience. Microbial dna typing by automated repetitivesequence. Conventional morphology versus pcr sequencing, rep. Use of the diversilab repetitive sequencebased pcr system for genotyping and identification of archaea article in journal of microbiological methods 732.
The performance of repetitivesequencebased pcr reppcr using the diversilab system for identification of dermatophytes commonly isolated in a clinical laboratory was assessed by comparing results to those of conventional tests colony morphology, microscopic examination of slide cultures, and, for suspected trichophyton species, use of additional media. Strain typing is suited to track the source of an exact contaminant. Th e analysis utilized the the rep pcr technology and rep pcr primers are covered by u. Using pcr technology, diversilab enables you to quickly differentiate microbial isolates at a subspecies and strain level in case of contamination. On the basis of an 80% similarity threshold, rep pcr consistently clustered like species and this set of isolates, along with five atcc reference strains, was used to create a dna fingerprint library with the diversilab software. The dna amplicons were detected, and analyses were performed with the diversilab system and accompanying software v. For analysis, diversilab software utilized the pearson correlation coefficient and the unweighted pairgroup methods of averages upgma to automatically compare the rep pcr profiles and create corresponding dendrograms healy et al. Use of the diversilab repetitive sequencebased pcr system. Detection and analysis of the rep pcr products were performed using the diversilab software version 3.
The diversilab system spectral genomics, houston, tx is a rapid, technically simple method that uses repetitivesequencebased pcr reppcr to determine relatedness of many organisms and to identify aspergillus and candida to the species level 2, 7. Based on patented rep pcr technology, microbial isolates can quickly and accurately be distinguished at the subspecies and strain level. Reppcr and rapdpcr fingerprinting of aspergillus terreus. Rep pcr based dendrogram and virtualgel image fingerprints for 71 isolates of different salmonella serovars following typing using the diversilab system with the salmonella kit.
Clinical evaluation of the diversilab microbial typing. We describe the adaptations made to automate reppcr and explore the performance and reproducibility of the system as a molecular genotyping tool for bacterial strain typing. The amplified fragments were separated and detected using a microfluidics chip with agilent 2100. The diversilab dna fingerprinting kits are designed to generate reppcr dna fingerprints from purecultured microbial samples. Yeast identification by sequencing, biochemical kits, malditof ms and rep pcr dna fingerprinting.
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