Analyzing Multiclonality of Staphylococcus aureus in Clinical Diagnostics Using spa -Based Denaturing Gradient Gel Electrophoresis

Abstract

We present a novel denaturing gradient gel electrophoresis (DGGE) method which characterizes multiclonal communities of Staphylococcus aureus. The spa PCR-based DGGE method simultaneously separates strains that differ in only one base, thereby revealing multiclonal colonization and infections.Staphylococcus aureus causes a wide range of infections and is responsible for a considerable portion of hospital-acquired infections.In 30% to 70% of healthy individuals, S. aureus is a transient or persisting part of the residential flora (5, 10).Cespedes et al. investigated the frequency of simultaneous nasal carriage of multiple S. aureus strains by picking three bacterial colonies from plates derived from each colonized individual. Fewer than 7% of them were predicted to carry Ͼ1 strain (1). The simultaneous presence of an invasive and a carrier strain of methicillin-resistant S. aureus (MRSA) in one individual was reported by Soderquist and Berglund (12). The issue of multiclonal colonization is important, but conventional laboratory methods for detection of S. aureus are based on culture of a single colony. This might result in the identification of an antibiotic-susceptible commensal strain rather than a second, more resistant strain, which may impair the antibiotic treatment and bias epidemiological conclusions.We have developed a species-specific denaturing gradient gel electrophoresis (DGGE) method for S. aureus, utilizing spa, to characterize multiclonal colonization and infection. The novel assay was used to investigate a MRSA outbreak, revealing colonization with two different strains in some of the individuals.spa typing has been documented to be a useful tool in investigations of MRSA epidemiology (7) and for studies of S. aureus transmission (6). Thus, we based our DGGE method on spa and primer pairs, described by Kahl et al. (4), which were modified for DGGE analyses by the attachment of a GC clamp (11) at either the forward or the reverse primer. Annealing temperature and MgCl 2 concentrations for two primer combinations were optimized for PCR specificity and efficiency (data not shown).Eight S. aureus isolates of known spa types were acquired from the

Staphylococcus aureus colonization related to severity of hand eczema

“…This differs from the study by Haslund et al demonstrating that S. aureus of the same spa type always colonize the anterior nares and hand of a patient [6]. Simultaneous colonization with S. aureus of different spa types and simultaneous colonization with susceptible and resistant strains has been documented previously [36,37]. In case of an infection with multiple types, antibiotic susceptibility testing of solely one colony could lead to the detection of a susceptible rather than a resistant strain and thereby antibiotic treatment failure.…”

Prevalence and molecular epidemiology of Staphylococcus aureus in Swedish nursing homes – as revealed in the SHADES study

“…Furthermore, in epidemiological and transmission studies undetected carriage of multiple strains could lead to misinterpretation of data. However, analyses of dual colonization with different strains based on selecting several colonies from agar plates would be laborious and simplified methods are needed [ 32 ].…”

Large and finite sample properties of a maximum-likelihood estimator for multiplicity of infection

“…On the contrary, eukaryotic parasites such as Plasmodium , Trypanosoma , Toxoplasma , and Schistosoma [ 26 , 27 ] and bacteria such as Mycobacterium [ 28 ] evolve at a rate at which it is possible to determine a stable number of genetically distinct lineages during the course of an infection given a set of genetic markers. Even when restricted to such pathogens, due to difficulties naturally arising from confounding factors in ecological and epidemiological investigations [ 2 , 3 , 5 , 28 , 29 ], the concept of MOI is enervated. Estimating MOI and frequency spectra was dominated by ad-hoc methods, which are intuitive but typically introduce a bias, which cannot be quantified due to the lack of a statistical framework.…”