Identification in Tissues

Cultures from infected tissue are often negative (Ribes et al., 2000) and the different zygomycetes shared similar morphological characteristics by histopathology. Furthermore, in some instances, differentiation of a zygomycetes with another hyalohyphomycete could be difficult by histopathology. Then, alternative methods for diagnosis of zygomycosis and for species identification directly from tissues are needed. For this purpose, molecular methods have been recently evaluated both on unfixed fresh/frozen material (Kobayashi et al., 2004; Iwen et al., 2005; Machouart et al., 2006; Lau et al., 2006; Schwarz et al., 2006) and on formalin-fixed paraffin-embedded biopsies (Lau et al., 2006; Bialek et al., 2005; Nagao et al., 2005; Rickerts et al., 2006b; Hayden et al., 2002; Kobayashi et al., 2004).

In an experimental model of zygomycosis in mice, it has been shown that molecular identification of the most common zygomycetes species could be done from frozen tissue samples by PCR amplification with panfungal primers followed by direct sequencing of the ITS region (Schwarz et al., 2006). Very recently, a similar approach has been successfully used in 7 tissue samples from three patients with proven zygomycosis (Lau et al., 2006). Molecular identification has also been used in selected case reports to identify zygomycetes such as Cunninghamella berthol-letiae (Kobayashi et al., 2004), Rhizomucor pusillus (Iwen et al., 2005), Rhizopus microsporus (Machouart et al., 2006), and Absidia corymbifera (Machouart et al., 2006) from various unfixed tissue samples.

Molecular detection of fungi from formalin-fixed paraffin-embedded tissues could be more difficult than from fresh specimens due to the degradation of DNA during processing of the samples. Nevertheless, paraffin-embedded tissues are often the only available samples, because zygomycosis has not been suspected initially. Several molecular techniques have been evaluated recently for identification of zygomycetes in such samples. In situ hybridization with panfungal and zygomycetes-specific oligonucleotide probes directed against 18S rDNA has been recently assessed in 13 samples from culture-proven cases of zygomycosis (Hayden et al., 2002). Although, good sensitivity and specificity was obtained, low staining intensity of the zygomyc-etes hyphae and sometimes high background staining limited the interpretation in some cases. A panfungal PCR assay targeting the ITS region has been recently evaluated on a large number of formalin-fixed paraffin-embedded tissue samples from patients with histologically proven fungal infection (Lau et al., 2006). Among the nine samples diagnosed as zygomycosis by histopathology, PCR amplification was negative in four samples, indicating that this molecular technique was less sensitive than the standard histological diagnosis. Nevertheless, in case of positive PCR, a molecular identification of the species was achieved while it is not possible by histology. In two other studies (Bialek et al., 2005; Rickerts et al., 2006b), an 18S-targeted semi-nested PCR specific for zygomycetes has been evaluated and showed promising results. Nevertheless, sensitivity was not optimal, as shown in one of the studies, with negative PCR for 9 out of 23 samples tested (Bialek et al., 2005). In some other clinical cases, molecular techniques have been successfully used for species identification (Kobayashi et al., 2004; Nagao et al., 2005). Overall, these results are very encouraging and warrant further studies to improve sensitivity.

Cure Your Yeast Infection For Good

Cure Your Yeast Infection For Good

The term vaginitis is one that is applied to any inflammation or infection of the vagina, and there are many different conditions that are categorized together under this ‘broad’ heading, including bacterial vaginosis, trichomoniasis and non-infectious vaginitis.

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