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Immigrants from regions with a high incidence of tuberculosis (TB) are a risk group for TB in low-incidence countries such as Switzerland. In a previous analysis of a nationwide collection of 520 Mycobacterium tuberculosis isolates from 2000 to 2008, we identified 35 clusters comprising 90 patients based on standard genotyping (24-locus mycobacterial interspersed repetitive-unit–variable-number tandem-repeat [MIRU-VNTR] typing and spoligotyping). Here, we used whole-genome sequencing (WGS) to revisit these transmission clusters. Genome-based transmission clusters were defined as isolate pairs separated by ≤12 single nucleotide polymorphisms (SNPs). WGS confirmed 17/35 (49%) MIRU-VNTR typing clusters; the other 18 clusters contained pairs separated by >12 SNPs. Most transmission clusters (3/4) of Swiss-born patients were confirmed by WGS, as opposed to 25% (4/16) of the clusters involving only foreign-born patients. The overall clustering proportion was 17% (90 patients; 95% confidence interval [CI], 14 to 21%) by standard genotyping but only 8% (43 patients; 95% CI, 6 to 11%) by WGS. The clustering proportion was 17% (67/401; 95% CI, 13 to 21%) by standard genotyping and 7% (26/401; 95% CI, 4 to 9%) by WGS among foreign-born patients and 19% (23/119; 95% CI, 13 to 28%) and 14% (17/119; 95% CI, 9 to 22%), respectively, among Swiss-born patients. Using weighted logistic regression, we found weak evidence of an association between birth origin and transmission (adjusted odds ratio of 2.2 and 95% CI of 0.9 to 5.5 comparing Swiss-born patients to others). In conclusion, standard genotyping overestimated recent TB transmission in Switzerland compared to WGS, particularly among immigrants from regions with a high TB incidence, where genetically closely related strains often predominate. We recommend the use of WGS to identify transmission clusters in settings with a low incidence of TB.

During a 9-month period, 217 patients were newly diagnosed as methicillin-resistant Staphylococcus aureus (MRSA) carriers by using a commercial rapid PCR-based test (GeneXpert). However, no MRSA was recovered by culturing the second swab in 61 of these patients. Further analyses showed that 28 (12.9%) of the patients harbored S. aureus isolates with a staphylococcal cassette chromosome element lacking the mecA gene and were thus incorrectly determined to be MRSA carriers.

In order to improve the diagnosis of Listeria meningitis or meningoencephalitis, especially in patients who have received antibiotics before their cerebrospinal fluid (CSF) has been examined, two assays for the detection of Listeria monocytogenes based on the polymerase chain reaction (PCR) were evaluated. After a standard PCR, the amplified DNA was detected either by a second round of PCR with internal primers followed by gel electrophoresis and ethidium bromide staining (nested PCR) or by dot blot hybridization to an internal digoxigenin-labeled probe (PCR-dot blot). For PCR, two sets of primers within the invasion-associated protein gene (iap gene) were chosen. They allowed for the highly specific detection of all L. monocytogenes reference strains tested (serotypes 1/2a, 1/2b, 1/2c, 3a, 3b, 3c, 4a, 4b, 4c, 4d, and 7). These primers did not detect amplification products from various other gram-positive or gram-negative bacterial DNAs or human DNA. The sensitivities of both assays were assessed on sterile CSF samples that were artificially seeded with serial dilutions of L. monocytogenes serotype 4b cells. By both methods the limit of detection was less than 10 cells in the initial reaction. Since the nested PCR is more prone to contamination because of manipulation of the amplified products, a standard PCR assay followed by dot blot hybridization was applied to 52 CSF samples in a retrospective study. Of 28 CSF samples which were sterile or positive for bacteria other than Listeria species, 24 were PCR negative. In contrast, from 17 patients with culture-proven Listeria meningitis, 14 of 17 initial CSF samples were PCR positive, as were 3 of 7 culture-negative followup CSF samples taken after patients received antibiotics. These results support the usefulness of this approach in the diagnosis of Listeria meningitis, in particular, when antibiotic administration precedes culture of CSF.

There have been reports of increasing numbers of cases of malaria among migrants and travelers. Although microscopic examination of blood smears remains the “gold standard” in diagnosis, this method suffers from insufficient sensitivity and requires considerable expertise. To improve diagnosis, a multiplex real-time PCR was developed. One set of generic primers targeting a highly conserved region of the 18S rRNA gene of the genus Plasmodium was designed; the primer set was polymorphic enough internally to design four species-specific probes for P. falciparum , P. vivax , P. malarie , and P. ovale . Real-time PCR with species-specific probes detected one plasmid copy of P. falciparum , P. vivax , P. malariae , and P. ovale specifically. The same sensitivity was achieved for all species with real-time PCR with the 18S screening probe. Ninety-seven blood samples were investigated. For 66 of them (60 patients), microscopy and real-time PCR results were compared and had a crude agreement of 86% for the detection of plasmodia. Discordant results were reevaluated with clinical, molecular, and sequencing data to resolve them. All nine discordances between 18S screening PCR and microscopy were resolved in favor of the molecular method, as were eight of nine discordances at the species level for the species-specific PCR among the 31 samples positive by both methods. The other 31 blood samples were tested to monitor the antimalaria treatment in seven patients. The number of parasites measured by real-time PCR fell rapidly for six out of seven patients in parallel to parasitemia determined microscopically. This suggests a role of quantitative PCR for the monitoring of patients receiving antimalaria therapy.