Strains In Clinical Samples Research Articles

Discrimination of Common E. coli Strains in Urine by Liquid Chromatography-Ion Mobility-Tandem Mass Spectrometry and Machine Learning.

Accurate identification of bacterial strains in clinical samples is essential to provide an appropriate antibiotherapy to the patient and reduce the prescription of broad-spectrum antimicrobials, leading to antibiotic resistance. In this study, we utilized the combination of a multidimensional analytical technique, liquid chromatography-ion mobility-tandem mass spectrometry (LC-IM-MS/MS), and machine learning to accurately identify and distinguish 11 Escherichia coli (E. coli) strains in artificially contaminated urine samples. Machine learning was utilized on the LC-IM-MS/MS data of the inoculated urine samples to reveal lipid, metabolite, and peptide isomeric biomarkers for the identification of the bacteria strains. Tandem MS and LC separation proved effective in discriminating diagnostic isomers in the negative ion mode, while IM separation was more effective in resolving conformational biomarkers in the positive ion mode. Using hierarchical clustering, the strains are clustered accurately according to their group highlighting the uniqueness of the discriminating biomarkers to the class of each E. coli strain. These results show the great potential of using LC-IM-MS/MS and machine learning for targeted omics applications to diagnose infectious diseases in various environmental and clinical samples accurately.

Molecular typing and antifungal susceptibility profile of Candida krusei bloodstream isolates from Türkiye.

Candida krusei also known as Pichia kudriavzevii is a potentially multidrug-resistant yeast because it is intrinsically resistant to fluconazole and develops acquired resistance to echinocandins and polyenes. Here, we aim to provide a better understanding of the epidemiology and transmission modes of C. krusei infections by comparing invasive bloodstream (n=35) and non-invasive vaginal (n=20) C. krusei isolates. The genetic relatedness of the isolates was assessed using a newly described short tandem repeat (STR) analysis and their sensitivity to eight antifungal compounds was evaluated by antifungal susceptibility testing using the CLSI microbroth dilution method. All C. krusei isolates revealed unique STR genotypes, indicating the absence of clonal transmission in the study group. Furthermore, no drug-resistant or non-wild-type isolates were identified. Our findings demonstrated high resolution of STR genotyping for the detection and simultaneous genetic analysis of multiple C. krusei strains in clinical samples and excellent in vitro activity of common antifungal agents against invasive strains.

Methods of Identifying Gordonia Strains in Clinical Samples.

Gordonia spp. are members of the family Gordoniacea in the suborder Corynebacteriales; their habitat, in most cases, is soil. Many representatives of this genus are human or veterinary pathogens. The main cause of the lack of a standardized approach to dealing with infections caused by Gordonia is their erroneous identification and little information regarding their susceptibility to antimicrobial drugs. This review presents the most common methods for identifying Gordonia strains, including modern approaches for identifying a species. The main prospects and future directions of this field of knowledge are briefly presented.

Isolation and Genetic Characterization of Vancomycin-resistant and mecC+ Methicillin-resistant Staphylococcus aureus Strains in Clinical Samples of Bojnurd, Northeastern Iran

Background: The emergence of antibiotic-resistant Staphylococcus aureus strains is one of the major concerns about the various staphylococcal infections. Vancomycin is one the most important effective antibiotics on staphylococcal lethal infections. To date, vancomycin-resistant strains are increasingly isolated in different parts of the world, and it is alerting. Objectives: The current study was designed to evaluate the prevalence, and antibiotic susceptibility pattern of methicillin-resistant S. aureus (MRSA) and vancomycin-resistant S. aureus (VRSA) isolates in the main tertiary hospital of Bojnurd, Iran. Methods: Staphylococcus aureus isolates were collected from different clinical samples in Imam Reza Hospital of Bojnurd. After identification of isolates through using conventional methods, they were evaluated by agar screening, disk diffusion, and minimum inhibitory concentration (MIC) methods to determine resistance to vancomycin and methicillin. We also performed polymerase chain reaction (PCR) for the detection of mecA, mecC, vanA, and vanB genes. After confirmation of vancomycin resistance, genetic analysis was performed using SCCmec, agr, and spa typing, and multilocus sequence typing (MLST) methods on VRSA isolates. Results: We found four vancomycin-resistant isolates (1.29%). Also, 75% of isolates were resistant to cefoxitin. Using the PCR method, mecA was found in 73%, mecC in 0.64%, and vanA in 1.29% of isolates. Interestingly, we found two mecC positive isolates in MRSA isolates. The alpha-hemolysin (81.81%) and enterotoxin C (27%) had the highest and lowest toxins percentage, respectively. Among mecA positive isolates, SCCmec IV (37%), SCCmec III (31.27%), SCCmec I (14%), SCCmec II (11%), and SCCmec V (5.7%) were the most prevalent SCCmec types, respectively. It should be noted that the two mecC positive isolates belonged to SCCmec XI. Agr I (76.29%) was the highest agr type. We recognized t037 as the dominant spa type, and ST239, ST6, ST97, and ST8 were found in VRSA isolates. Conclusions: In our study, the frequency of mecA genes in MRSA isolates was very high. It seems that the resistant isolates belonged to endemic clones of Iran.

Challenging Methicillin Resistance Detection in Bone and Joint Infections: Focus on the MRSA/SA SSTI® Strategy.

The genus Staphylococcus is the main causative agent of bone and joint infections (BJI) in which outcomes are impacted by both effective surgical and appropriate antimicrobial management. In this context, methicillin resistance (MR) detection is a microbiological challenge to optimize the anti-staphylococcal drug coverage and to secure the surgical procedure. During the last decade, molecular tools have been developed to rapidly detect bacterial-resistant strains in clinical samples. The GeneXpert MRSA/SA SSTI® assay (Cepheid, Sunnyvale, CA, USA) is a real-time PCR method aimed at detecting methicillin-resistant Staphylococcus aureus (MRSA) in skin and soft tissues infections. In the literature, this test has been reported to be diverted from its original purpose to be evaluated in surgical samples. Within the current review, we update the GeneXpert MRSA/SA SSTI® assay performance in staphylococcal species determination (i.e., S. aureus vs. coagulase-negative species) together with MR genotype detection, when performed in osteoarticular infections.

MgpB Types among Mycoplasma genitalium Strains from Men Who Have Sex with Men in Berlin, Germany, 2016-2018.

Mycoplasma genitalium is a cell wall-less bacterium causing urethritis and other sexually transmitted diseases. Despite a strongly conserved genome, strains in clinical samples can be typed by different methods. To obtain data from the risk population of men having sex with men, we analyzed the typing region in the gene coding for the MgpB adhesin of M. genitalium first in 163 and 45 follow-up samples among patients of two specialized practices in Berlin, Germany. Strains belong to 43 different mgpB types emphasizing the diversity of the genome region. With respect to 133 types previously described, 27 new types were found. However, the majority of strains (64.4%) were assigned to types 4, 6, 113, and 108, respectively. A correlation between mgpB type and the occurrence of mutations associated with macrolide and quinolone resistance was not demonstrated. Investigation of follow-up samples from 35 patients confirmed the same mgpB and, additionally, MG_309 types in 25 cases. In 10 cases, differences between types in subsequent samples indicated an infection with a genetically different strain in the period between samplings. MgpB/MG_309 typing is a useful method to compare M. genitalium strains in samples of individual patients as well as those circulating in different populations

Multi-Locus Next-Generation Sequence Typing of DNA Extracted From Pooled Colonies Detects Multiple Unrelated Candida albicans Strains in a Significant Proportion of Patient Samples.

The yeast Candida albicans is an important opportunistic human pathogen. For C. albicans strain typing or drug susceptibility testing, a single colony recovered from a patient sample is normally used. This is insufficient when multiple strains are present at the site sampled. How often this is the case is unclear. Previous studies, confined to oral, vaginal and vulvar samples, have yielded conflicting results and have assessed too small a number of colonies per sample to reliably detect the presence of multiple strains. We developed a next-generation sequencing (NGS) modification of the highly discriminatory C. albicans MLST (multilocus sequence typing) method, 100+1 NGS-MLST, for detection and typing of multiple strains in clinical samples. In 100+1 NGS-MLST, DNA is extracted from a pool of colonies from a patient sample and also from one of the colonies. MLST amplicons from both DNA preparations are analyzed by high-throughput sequencing. Using base call frequencies, our bespoke DALMATIONS software determines the MLST type of the single colony. If base call frequency differences between pool and single colony indicate the presence of an additional strain, the differences are used to computationally infer the second MLST type without the need for MLST of additional individual colonies. In mixes of previously typed pairs of strains, 100+1 NGS-MLST reliably detected a second strain. Inferred MLST types of second strains were always more similar to their real MLST types than to those of any of 59 other isolates (22 of 31 inferred types were identical to the real type). Using 100+1 NGS-MLST we found that 7/60 human samples, including three superficial candidiasis samples, contained two unrelated strains. In addition, at least one sample contained two highly similar variants of the same strain. The probability of samples containing unrelated strains appears to differ considerably between body sites. Our findings indicate the need for wider surveys to determine if, for some types of samples, routine testing for the presence of multiple strains is warranted. 100+1 NGS-MLST is effective for this purpose.

Isolation and antibiotic resistance pattern of vancomycin-resistant Enterococcus fascium strains in clinical samples in Tabriz hospitals.

Background: In recent decades, Enterococci have been identified as an important pathogen. Resistance of these bacteria to antibiotics causes major problems in the therapeutic process. Therefore, the aim of this study was to determine the antibiotic resistance pattern of Enterococcus fascium strain in clinical samples. Methods: In this research, 219 samples of Enterococci from clinical specimens including: Blood and urine were studied in Tabriz hospitals. All strains were identified by standard and bacteriological tests, and then antibiotic resistance patterns of bacteria isolated by Kirby-Bauer disc diffusion method were performed. Also, the minimum inhibitory concentration (MIC) was determined by E-Test method. Results: Of 219 strains of Enterococcus, 86 (39.26%) strains belong to the species of E. fascium. The highest resistance to antibiotics was tetracycline (83.7%) and erythromycin (65.1%), and lowest compared to linezolid (0%) and teicoplanin (18.6%). Based on the E-Test, 29 (33.7%) sample was resistant to vancomycin. Conclusion: Considering the increasing number of infectious diseases caused by resistant strains of Enterococcus genus and its dissemination in the hospital environment, it is very necessary to use infection control measures to eliminate the potential sources of infection and prevent it from developing.

Rotavirus genotype shifts among Swedish children and adults—Application of a real-time PCR genotyping

BackgroundIt is well known that human rotavirus group A is the most important cause of severe diarrhoea in infants and young children. Less is known about rotavirus infections in other age groups, and about how rotavirus genotypes change over time in different age groups. ObjectivesDevelop a real-time PCR to easily genotype rotavirus strains in order to monitor the pattern of circulating genotypes. Study designIn this study, rotavirus strains in clinical samples from children and adults in Western Sweden during 2010–2014 were retrospectively genotyped by using specific amplification of VP 4 and VP 7 genes with a new developed real-rime PCR. ResultsA genotype was identified in 97% of 775 rotavirus strains. G1P[8] was the most common genotype representing 34.9%, followed by G2P[4] (28.3%), G9P[8] (11.5%), G3P[8] (8.1%), and G4P[8] (7.9%) The genotype distribution changed over time, from predominance of G1P[8] in 2010–2012 to predominance of G2P[4] in 2013–2014. There were also age-related differences, with G1P[8] being the most common genotype in children under 2 years (47.6%), and G2P[4] the most common in those over 70 years of age (46.1%.). The shift to G2P[4] in 2013–2014 was associated with a change in the age distribution, with a greater number of rotavirus positive cases in elderly than in children. ConclusionsBy using a new real-time PCR method for genotyping we found that genotype distribution was age related and changed over time with a decreasing proportion of G1P[8].

Evaluation of matrix-assisted laser desorption ionization-time of flight mass spectrometry for identification of human oral Capnocytophaga species

Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) was evaluated for rapid identification of cfxA PCR positive and negative Capnocytophaga strains. Colonies were grown on blood agar, incubated anaerobically at 37 °C for 48 h, and were then evaluated by MALDI-TOF MS and 16S rRNA gene sequencing. Both methods identified all colonies to the genus level. The MALDI-TOF MS method gave the same result, at the species level, as 16S rRNA gene sequencing for 41/53 Capnocytophaga sp. strains (77.4%), but the limit of this technique was the absence of some species (C. leadbetteri, C. AHN) in the Biotyper-Bruker® database used in this study. Distinction between the cefotaxime resistant and susceptible strains was unsuccessful using the MALDI-TOF MS method. This technique had low discriminatory power to rapidly detect beta-lactamase-producing Capnocytophaga strains in clinical samples. However, the results from a score-oriented dendrogram confirmed MALDI-TOF MS is a rapid, inexpensive, and reliable method for Capnocytophaga species identification. Enrichment of the reference database used (Biotyper®) will improve future results.

Application of Fluorescence Based Molecular Assays for Improved Detection and Typing of Brucella Strains in Clinical Samples

Abstract Bacteria from the genus Brucella are causative agents of brucellosis - a zoonotic disease which affects many wild and domestic animal species and humans. Taking into account the significant socio-economic and public health impact of brucellosis, its control is of great importance for endemic areas. The chosen control strategy could be successful only if adapted to the current epidemiological situation. This implies that a choice of appropriate diagnostic procedures for detection and typing of Brucella spp. strains are of essential importance. Significant advancement of molecular techniques and their advantages compared to classical methods, give strong arguments in promotion of these techniques as a powerful tool for comprehensive diagnostics of brucellosis. Considering this, the major tasks of the study were to select and implement molecular tests for detection and genotyping Brucella spp. and evaluate their performances using DNA from cultivated brucellae (islolates) and limited number of tissue samples from seropositive animals. The obtained results confirmed that implemented real time PCR for Brucella spp. detection, as well as MLVA-16 used for genotyping, have excellent analytical sensitivity (4.2 fg of Brucella DNA were successfully detected and genotyped). Furthermore, compared to bacteriological cultivation of Brucella spp., real time PCR and MLVA-16 protocols showed superior diagnostic sensitivity and detected Brucella DNA in tissues from which Brucella could not be cultivated. Based on the summarized study results, we propose a diagnostic algorithm for detection and genotyping of Brucella spp. bacteria. Routine use of proposed diagnostic algorithm will improve the effectiveness of infection confirmation and help for accurate evaluation of epidemiological situation.

Near-infrared spectroscopy and variable selection techniques to discriminate Pseudomonas aeruginosa strains in clinical samples

Pseudomonas aeruginosa is a leading cause of nosocomial infections, ranking second among the negative Gram pathogens reported to the National Nosocomial Infection Surveillance System. P. aeruginosa may develop resistant during prolonged therapy with all antimicrobial agents. Therefore, isolates that are initially susceptible may become resistant within 3–4days after initiation of therapy. Testing of repeat isolates may be warranted. There is a need for sensitive and specific tests. We set out to determine whether near-infrared spectroscopy (NIR) combined with variable selection techniques employing successive projection algorithm – linear discriminant analysis (SPA-LDA) or genetic algorithm – (GA-LDA) could discriminate P. aeruginosa strains according to resistant vs. sensitive. The variables selected were then used for discriminating the strains. The influence of various spectral pre-treatments (Savitzky–Golay smoothing, multiplicative scatter correction (MSC), and Savitzky–Golay derivatives) was calculated. In addition, accuracy test results including sensitivity and specificity were determined. Sensitivity in the resistant category was 95% using a SPA-LDA model with 70 wavelengths. Sensitivity and specificity in both categories was 93% using a GA-LDA model with 32 wavelengths. We show that NIR spectroscopy of P. aeruginosa combined with variable selection techniques is a powerful tool for resistant vs. sensitive strains based on the unique spectral “fingerprints” of their biochemical microbial identification, emerging as an alternative for rapid and cost-effective identification of strains.

Detection of algD, oprL and exoA Genes by New Specific Primers as an Efficient, Rapid and Accurate Procedure for Direct Diagnosis of Pseudomonas aeruginosa Strains in Clinical Samples.

Background:Pseudomonas aeruginosa is an opportunistic pathogen that infects patients with cystic fibrosis, burning wounds, ophthalmic traumas and immunodeficiency.Objectives:The aim of the present study was to compare the efficiency of newly designed primer sets with some previously published primers for PCR detection of exoA, oprL and algD genes from P. aeruginosa.Materials and Methods:A total of 150 clinical specimens were inoculated into the routine and selective culture media for P. aeruginosa isolation. Specific primers were designed by bioinformatics analysis for detection of the virulence genes determinants, exoA, oprL and algD. The sequences of these three genes were obtained from NCBI and multiple alignments were performed to find the conserved sequences of each gene to design the primers. Multiple alignment and primer design steps were carried out by the AlleleID software, version 7.0.Results:Microbiological culture methods resulted 70 P. aeruginosa strains isolated from 70 of the 150 clinical specimens. The results of the PCR assay using the newly designed exoA, oprL and algD primer sets were positive in 68, 70 and 69 clinical samples which represent 97.2%, 100% and 98% sensitivity for each primer set, respectively. The PCR results using previously published exoA, oprL and algD primer sets were positive only in 57, 49 and 28 specimens that correspond to 81.5%, 70% and 40% sensitivity, respectively.Conclusions:The results of the present study showed that in comparison with previously published primer sets, P. aeruginosa infection can be diagnosed with higher sensitivity and specificity by the conventional PCR assay using the newly designed primers. It was also shown that the results of the PCR assay on clinical samples of severe infections became positive much earlier than the results of conventional culture method.

Optimization of one-step real-time reverse transcription-polymerase chain reaction assays for norovirus detection and molecular epidemiology of noroviruses in Thailand

Noroviruses (NoVs) are an important human pathogen associated with acute viral gastroenteritis worldwide. NoVs display a significant amount of genetic heterogeneity, making it difficult to develop comprehensive detection assays. In this study, primer sets and probes were designed for a TaqMan®-based real-time reverse transcription-polymerase chain reaction (RT-PCR) for norovirus detection purposes. The assay was optimized and utilized as a multiplex real-time RT-PCR assay for genogroup I (GI) detection, and a singleplex real-time RT-PCR assay for genogroup II (GII) detection. The assays showed high specificity for NoV detection and no cross-reactivity was observed between GI and GII. The detection limit of the assay was as low as 10 and 50 RNA copies per reaction for GI and GII, respectively. The optimized protocol was employed to assess the presence of NoV strains in clinical samples collected throughout Thailand during December 2005 to November 2006. The percentage of NoV infections among children with acute gastroenteritis (case) was 23.8% (119/500) and for children without acute gastroenteritis (control) it was 6.8% (30/441). The frequency of NoV infections varied geographically, with the highest frequency observed in the central region and the lowest frequency in the northern region (P>0.0001). Of the 149 positive case and control specimens, GII was found to be the predominant genogroup (98.6%). Partial capsid sequences were successfully obtained from 67 NoV-positive specimens and a phylogenetic analysis was performed to genotype the viral strains. GII.4 was the most common genotype detected.

Cycling probe technology to quantify and discriminate between wild-type varicella-zoster virus and Oka vaccine strains

Rapid differentiation between wild-type varicella zoster virus (VZV) and Oka-vaccine (vOka) strains is important for monitoring side reactions of varicella vaccination. To develop a high-throughput molecular diagnostic method for the differentiation of wild-type VZV and vOka strains based on cycling probe technology. The primers were designed to amplify common sequences spanning a single nucleotide polymorphism (SNP) in gene 62 of VZV. DNA–RNA chimeric probes (cycling probes) were designed to detect the SNP at nucleotide 105705. The cycling probe real-time PCR assays for VZV wild-type and vOka strains specifically amplified plasmids containing target sequences that ranged between 10 and 1×106 copies per reaction. The inter- and intra-assay coefficients of variation were less than 5%. After initial validation studies, the clinical reliability of this method was evaluated using 38 swab samples that were collected from patients suspected of being zoster. Compared to the loop mediated isothermal amplification method, which is defined as the gold standard, cycling probe real-time PCR was highly sensitive and specific. The cycling probe real-time PCR technology is a reliable tool for differentiating between wild-type VZV and vOka strains in clinical samples.

Detection of Salmonella strains in clinical samples from Saudi Arabia by invA and hilA polymerase chain reaction (PCR)-based assays

In this study, 23 Salmonella isolates were detected by polymerase chain reaction (PCR) from human fecal samples obtained during 2010 from local hospitals and clinical laboratories in Kingdom of Saudi Arabia (KSA). The bacteria were cultured, serotyped and biochemically characterized by the analytical profiling index (API 20E). The invA and hilA gene primers were selected specifically for the detection of Salmonella to amplify a 284 and 845 bp DNA fragments, respectively. Only 3 isolates out of 23 were not identified or typed neither by API 20E nor serological tests. One of these isolates (S5) had 98% sequence similarity with the invA gene sequences of Salmonella typhimurium strain LT2, 14028S and SL1344. The results showed that invA or hilA PCRs specificity was 66.6% compared to API 20E and serology for Salmonella enterica identification in clinical specimens. The PCR assays compared to both biochemical and serological tests were able to specifically detect all of 23 Salmonella isolates (100% sensitivity). Key words: Salmonella, analytical profiling index (API 20E), serological tests, polymerase chain reaction (PCR), invA or hilA gene, DNA sequences, clinical specimens.

Direct Serogrouping of Streptococcus pneumoniae Strains in Clinical Samples by Use of a Latex Agglutination Test

Pneumotest-Latex (Statens Seruminstitut) was evaluated for direct serogrouping of Streptococcus pneumoniae strains in clinical samples from patients with invasive disease. The technique was accurate to its level of discrimination for 62 of 67 clinical samples (92.5%). Pneumotest-Latex would be a useful alternative for direct serogrouping of pneumococci in clinical samples.

Porphyromonas gingivalis Strain Diversity

Porphyromonas gingivalis is implicated in the etiology of chronic periodontitis. Genotyping studies suggest that genetic variability exists among P. gingivalis strains; however, the extent of variability remains unclear and regions of variability remain largely unidentified. To assess P. gingivalis strain diversity, we previously used heteroduplex analysis of the ribosomal operon intergenic spacer region (ISR) to type strains in clinical samples and identified 22 heteroduplex types. Additionally, we used ISR sequence analysis to determine the relatedness of P. gingivalis strains to one another and demonstrated a link between ISR sequence phylogeny and the disease-associated phenotype of the strains. In the current study, heteroduplex analysis of the ISR was used to determine the worldwide genetic variability and distribution of P. gingivalis, and microarray-based comparative genomic hybridization (CGH) analysis was used to more comprehensively examine the variability of major heteroduplex type strains by using the entire genome. Heteroduplex analysis of clinical samples from geographically diverse populations identified 6 predominant geographically widespread heteroduplex types (prevalence, > or = 5%) and 14 rare heteroduplex types (prevalence, <2%) which are found in one or a few locations. CGH analysis of the genomes of seven clinically prevalent heteroduplex type strains identified 133 genes from strain W83 that were divergent in at least one of the other strains. The relatedness of the strains to one another determined on the basis of genome content (microarray) analysis was highly similar to their relatedness determined on the basis of ISR sequence analysis, and a striking correlation between the genome contents and disease-associated phenotypes of the strains was observed.

Diagnostic discrimination of live attenuated influenza vaccine strains and community-acquired pathogenic strains in clinical samples

Live vaccines can generate false-positive results on common influenza assays including reverse transcriptase-PCR (RT-PCR), culture and antigen tests. This threatens the integrity of epidemiological data and may misdirect treatment and control efforts. We report the development of RT-PCR tests that distinguish live FluMist™ vaccine (FMV) strains from circulating influenza strains in clinical samples. Primers were validated using influenza-positive samples from unvaccinated patients, packaged FMV, and one PCR-positive asymptomatic vaccine. Furthermore, the assay was used to experimentally test our lab's collection of influenza-positive samples from the 2004–05 and 2005–06 influenza seasons and several 2005 preseason isolates to determine the rate of vaccine-derived false-positive results under differing epidemiological conditions. Analytical and clinical validations show that the assay is both sensitive and specific. Experimental results demonstrate that 51 out of 51 influenza-positive samples collected during influenza season from ill, previously-vaccinated military personnel represent real infections with circulating strains. Finally, the assay shows that four preseason influenza-positive samples were false positives resulting from vaccine shedding. The vaccine-discriminatory RT-PCR methods described here provide the first test designed to distinguish FMV strains from circulating strains. The results show that the test is effective, and demonstrate the importance of such tests in the age of live vaccines.

Global identification of three major genotypes of varicella-zoster virus: longitudinal clustering and strategies for genotyping.

By analysis of a single, variable, and short DNA sequence of 447 bp located within open reading frame 22 (ORF22), we discriminated three major varicella-zoster virus (VZV) genotypes. VZV isolates from all six inhabited continents that showed nearly complete homology to ORF22 of the European reference strain Dumas were assigned to the European (E) genotype. All Japanese isolates, defined as the Japanese (J) genotype, were identical in the respective genomic region and proved the most divergent from the E strains, carrying four distinct variations. The remaining isolates carried a combination of E- and J-specific variations in the target sequence and thus were collectively termed the mosaic (M) genotype. Three hundred twenty-six isolates collected in 27 countries were genotyped. A distinctive longitudinal distribution of VZV genotypes supports this approach. Among 111 isolates collected from European patients, 96.4% were genotype E. Consistent with this observation, approximately 80% of the VZV strains from the United States were also genotype E. Similarly, genotype E viruses were dominant in the Asian part of Russia and in eastern Australia. M genotype viruses were strongly dominant in tropical regions of Africa, Indochina, and Central America, and they were common in western Australia. However, genotype M viruses were also identified as a minority in several countries worldwide. Two major intertypic variations of genotype M strains were identified, suggesting that the M genotype can be further differentiated into subgenotypes. These data highlight the direction for future VZV genotyping efforts. This approach provides the first simple genotyping method for VZV strains in clinical samples.