The increasing incidence of aspergillosis, a life-threatening infection in immunocompromised patients, emphasizes the need to improve the currently limited diagnostic tools. We developed a two-step PCR assay that specifically amplifies a region of the 18S rRNA gene that is highly conserved in Aspergillus species. A number of primers with the least homology to equivalent human orCandida gene sequences were screened for the pairs that gave the highest sensitivity and specificity. No cross-reaction with the wide range of fungal and bacterial pathogens so far tested was observed. This assay allows direct and rapid detection of down to 10 fg of Aspergillus DNA corresponding to 1 to 5 CFU per ml of blood. A total of 315 blood and bronchoalveolar lavage samples from 140 subjects, including 93 patients at risk for invasive fungal disease, were screened. The result was a 100% correlation between positive histology, culture, or high-resolution computed tomography findings and PCR results. The test specificity was 89%. Our data point to the considerable potential clinical value of this simple, specific, rapid, and inexpensive PCR assay for improving the means of early diagnosis of systemic aspergillosis in high-risk patients.

Betaretroviruses exist in endogenous and exogenous forms in hosts that are widely distributed and evolutionarily distantly related. Here we report the discovery and characterization of several previously unknown betaretrovirus groups in the genomes of Mus musculus and Rattus norvegicus. Each group contains both mouse and rat elements, and several of the groups are more closely related to previously known betaretroviruses from nonmurine hosts. Some of the groups also include members from hosts which were not previously known to harbor betaretroviruses, such as the gray mouse lemur (Microcebus murinus) and Seba's short-tailed bat (Carollia perspicillata). Some of the mouse and rat elements possess intact open reading frames for gag, pro, pol, and/or env genes and display characteristics of having retrotransposed recently. We propose a model whereby betaretroviruses have been evolving within the genomes of murid rodents for at least the last 20 million years and, subsequent to (or concomitant with) the global spread of their murid hosts, have occasionally been transmitted to other species.Endogenous retroviruses are present in the genomes of all vertebrates (5). They are presumed to arise from germ line infection by exogenous retroviruses, although factors controlling endogenization are poorly understood. Endogenous proviruses accumulate mutations while in the germ line but can occasionally escape the germ line and infect other hosts, sometimes following recombination with other endogenous or exogenous retroviruses (5,20).The Betaretrovirus genus includes the viruses formerly known as type B and type D retroviruses (33). Betaretroviruses have been discovered in mammalian hosts of wide geographical and evolutionary diversity (Table 1). Mouse mammary tumor virus (MMTV), the prototype type B retrovirus, exists in closely related endogenous and exogenous forms, with variable distribution in both laboratory strains and wild species of mice (6,7,11,14). Jaagsiekte sheep retrovirus (JSRV), enzootic nasal tumor virus (ENTV), and endogenous sheep retrovirus are closely related endogenous and exogenous retroviruses of sheep and goats (9,12,35 (18), and a metatherian (marsupial) mammal, the Australian common brushtail possum (Trichosurus vulpecula endogenous retrovirus type D [TvERV-D]) (1). PCR approaches, using degenerate primers based on conserved regions of the retroviral pro and/or pol genes, have also been used to detect betaretrovirus-related elements in the genomes of pigs (10, 22), the bower bird, and the stripe-faced dunnart (13), although these elements have not been completely characterized. Many of the endogenous betaretroviruses appear to have entered the genomes of their hosts relatively recently (within the last ϳ10 million years) (Table 1). However, no satisfactory explanation as to how betaretroviruses could have become so widely distributed has been presented.The Muridae family of rodents comprises over 1,300 species and contains approximately one-quarter of all the known mammalian species (21). The family ...

Bronchoalveolar lavage (BAL) samples from 67 patients who were at high risk for invasive aspergillosis were examined using a recently developed 2-step polymerase chain reaction (PCR) that detects

Evolution over millions of years has adapted several thousand copies of retrovirus-like elements and over 10 times as many solitary long terminal repeats (LTRs) to their present location in the human genome. Transcription of these human endogenous retroviruses (HERVs) has been detected in various cells and tissues, and in some cases their transcriptional control elements have been recruited by cellular genes. We used a retroviral pol-specific expression array to obtain a HERV transcription profile in a variety of human cells such as epidermal keratinocytes, liver cells, kidney cells, pancreatic cells, lymphocytes, and lung fibroblasts. This rapid screening test revealed a distinct HERV pol-expression pattern in each cell type tested so far. About 40 different U3/R regulatory sequences from the HERV-H and HERV-W families were then amplified from actively transcribed 3'HERV LTRs of various cell lines and tissues. Their promoter activities were compared with LTR sequences of other known HERV families in 12 human cell lines using a transient luciferase reporter system. Expression of the isolated HERV LTRs varied significantly in these cell lines, in some cases showing strict cell type specificity. These results suggest that endogenous retroviral LTRs may be a valuable source of transcriptional regulatory elements for the construction of targeted retroviral expression vectors.