Molecular epidemiology of Corynebacterium diphtheriae from northwestern Russia and surrounding countries studied by using ribotyping and pulsed-field gel electrophoresis

Abstract

A selection of 100 Corynebacterium diphtheriae isolates from asymptomatic carriers and clinical cases from five regions in northwestern Russia were examined. Six additional isolates from patients in Finland and Estonia with epidemiological links to Russia were also examined. All isolates were characterized by biotyping, toxigenicity testing, ribotyping, and pulsed-field gel electrophoresis (PFGE). Hybridization of genomic DNA digested with BstEII revealed five ribotype patterns among the biotype gravis isolates (G1 through G5) and two patterns among the biotype mitis isolates (M1 and M2). PFGE using SfiI was not able to distinguish between ribotypes G1, G2, and G4. The predominant ribotype pattern, G1, found in cases of disease in all the areas studied, appears to be disseminating, in view of the isolates received from imported cases in Finland and Estonia. Among the 106 isolates examined, 68 produced pattern G1 and 24 produced pattern M1. Most of the M1 isolates were from the Leningrad Oblast region. Distinct ribotypes such as G2, G3, G4, G5, and M2 could represent endemic disease.

Diphtheria in the Republic of Georgia: Use of Molecular Typing Techniques for Characterization of Corynebacterium diphtheriae Strains

“…Six restriction enzymes (NotI, XbaI, AvrII, EcoRV, SpeI, and SfiI) and a variety of electrophoresis parameters were examined in order to determine the optimal conditions for PFGE typing of the C. diphtheriae strains. In agreement with a previous report (2), SfiI cleavage yielded the optimal number of bands (Ͼ10 bands [25]), and we used that enzyme in all subsequent PFGE typing experiments. The plugs were incubated (30 min, room temperature) with restriction enzyme buffer, the DNA in the plugs was digested by incubation (50°C, 5 h) of the plugs with SfiI (New England Biolabs, Beverly, Mass.…”

“…Analysis of PFGE and ribotyping patterns. C. diphtheriae isolates were separated into patterns on the basis of two band differences (PFGE) and one band difference (ribotyping) (2,25), and a dendrogram was constructed based on the PFGE patterns. The patterns were compared by means of the Dice coefficient with a Sun Microsystem (Bio Image, Ann Arbor, Mich.), and clustering of strains was based on the unweighted pair-group method with averages (a tolerance of 3% in the band position was applied).…”

“…Molecular typing techniques have been shown to be superior (i.e., to provide more specific and discriminating analysis) to serotyping and phage typing for studying the epidemiology of diphtheria outbreaks and for documenting the long-term persistence of C. diphtheriae strains in the population (1,21), but data pertaining to the superiority of any one molecular typing method over another are insufficient. Ribotyping was reported to be an excellent technique for the typing of C. diphtheriae strains (2). However, it is rather laborious and, in several laboratories, is performed with radiolabeled riboprobes.…”

Corynebacterium diphtheriae spoligotyping based on combined use of two CRISPR loci

“…However, a large diphtheria epidemic occurred in 1990s in Russia and other countries of the former Soviet Union with dramatic consequences (140 000 cases, 4000 deaths in 1991-1996 [1]). In 1990s, the available typing methods allowed a clonal group of closely related strains responsible for the epidemic to be identified [2][3][4]. These strains were indistinguishable by PFGE, randomly amplified polymorphic DNA (RAPD), and amplified fragment length polymorphism (AFLP), and were very similar in ribotyping (two principal profiles, 'Rossija' and 'Sankt-Peterburg', differing in one band [3,5]).…”

“…In 1990s, the available typing methods allowed a clonal group of closely related strains responsible for the epidemic to be identified [2][3][4]. These strains were indistinguishable by PFGE, randomly amplified polymorphic DNA (RAPD), and amplified fragment length polymorphism (AFLP), and were very similar in ribotyping (two principal profiles, 'Rossija' and 'Sankt-Peterburg', differing in one band [3,5]). However, multilocus enzyme electrophoresis (MLE), PFGE, and ribotyping are time-consuming and rather cumbersome methods, while RAPD lacks interlaboratory reproducibility and hence exchangeability of results.…”