In 1991, the AIDS Task Force of the American Academy of Neurology published nomenclature and research case definitions to guide the diagnosis of neurologic manifestations of HIV-1 infection. Now, 16 years later, the National Institute of Mental Health and the National Institute of Neurological Diseases and Stroke have charged a working group to critically review the adequacy and utility of these definitional criteria and to identify aspects that require updating. This report represents a majority view, and unanimity was not reached on all points. It reviews our collective experience with HIV-associated neurocognitive disorders (HAND), particularly since the advent of highly active antiretroviral treatment, and their definitional criteria; discusses the impact of comorbidities; and suggests inclusion of the term asymptomatic neurocognitive impairment to categorize individuals with subclinical impairment. An algorithm is proposed to assist in standardized diagnostic classification of HAND.
Replication-defective acute leukemia viruses E26 and myeloblastosis virus (AMV) cause distinct leukemias although they belong to the same subgroup of oncogenic avian tumor viruses based on shared transformation-specific ( onc ) RNA sequences. E26 causes predominantly erythroblastosis in chicken and in quail, whereas AMV induces a myeloid leukemia. However, upon cultivation in vitro for >1 month, a majority of surviving hemopoietic cells of E26-infected animals bear myeloid markers similar to those of AMV-transformed cells. We have analyzed the genetic structure and gene products of E26 virus for a comparison with those of AMV. An E26/helper virus complex was found to contain two RNA species: a 5.7-kilobase (kb) RNA that hybridizes with cloned AMV-specific proviral DNA and hence is probably the E26 genome; and an 8.5-kb RNA that is unrelated to AMV and represents helper virus RNA. Thus, E26 RNA is smaller than 7.5-kb AMV RNA. Hybridization of size-selected poly(A)-terminating E26 RNA fragments with AMV-specific DNA indicated that the shared specific sequences are located in the 5′ half of the E26 genome as opposed to a 3′ location in AMV RNA. In nonproducer cells transformed in vitro by E26, a gag -related nonstructural 135,000-dalton protein (p135) was found. No gag (Pr76) or gag-pol (Pr180) precursors of essential virion proteins, which are present in AMV nonproducer cells, were observed. p135 was also found in cultured E26 virus producing cells of several leukemic chickens, and its intracellular concentration relative to that of the essential virion proteins encoded by the helper virus correlates with the ratio of E26 to helper RNA in virions released by these cells. p135 is phosphorylated but not glycosylated; antigenically it is not related to the pol or env gene products. It appears to be coded for by a partial gag gene and by E26-specific RNA sequences, presumably including those shared with AMV. Hence, AMV and E26 appear to use different strategies for the expression of related onc sequences: AMV is thought to encode a transforming protein via a subgenomic mRNA, whereas E26 codes for a gag -related polyprotein via genomic RNA. It is speculated that differences in the oncogenic properties of E26 and AMV are due to differences in their genetic structures and gene products.
The src genes of different Rous sarcoma virus (RSV) strains have been reported to be highly conserved by some investigators using RNA-cDNA hybridization, whereas others using oligonucleotide, peptide, and serological analyses have judged src genes to be variable in 30 to 50% of the respective markers. Moreover, distinctive src oligonucleotides and peptides of so-called recovered RSVs (rRSV's) whose src genes were reported to be experimentally transduced from the cell are thought to represent specific markers of host-derived src sequences. By contrast, we have pointed out previously that these markers may represent point mutations of parental equivalents. Here we have compared the src -specific sequences of eight RSV strains and of two rRSV's to each other and to a molecular clone of the src -related chicken locus. Our comparisons are based on RNase T 1 -resistant oligonucleotides of RNA hybridized to src -specific cDNA, which was prepared by hybridizing RSV cDNA with RNA of isogenic src deletion mutants, or to a cloned cellular src -related DNA. All of the approximately 20 src -oligonucleotides of a given RSV strain were recovered by src -specific cDNA's of all other RSV strains or by cellular src -related DNA. The number of oligonucleotides varied slightly with the length of the src deletion used to prepare src -specific cDNA, thus providing a measure for src deletion mutants. Our data indicate that the src genes of all RSV strains tested, including the two reportedly transduced from the cell, are about 98% conserved and completely allelic with only scattered single nucleotide differences in certain variable regions which are subject to point mutations. Hence, based on the src oligonucleotide markers analyzed by us and others, we cannot distinguish between a cellular and viral origin of rRSV's. However, the following are not compatible with a cellular origin of rRSV's. (i) The only putative oligonucleotide marker which is exclusively shared by the two rRSV's studied and which differs from a parental counterpart in a single base was not detectable in cellular src -related DNA. (ii) The number of different allelic src markers observed by us and others in rRSV's was too large to derive from one or two known cellular src -related loci. (iii) The known absence of linkage of the cellular src -related locus with other virion sequences was extended to all non- src oligonucleotides, including some mapping directly adjacent to src . This is difficult to reconcile with the claim that transformation-defective, partial src deletion mutants of RSV which contain both, one, or, as we show here, possibly no src termini nevertheless transduce at the same frequencies, even though homologous, single or double illegitimate recombinations would be involved. Given (i) our evidence that src genes are subject to point mutation under selective conditions similar to those prevailing when rRSV's were generated and (ii) the lack of absolute evidence for the clonal purity of the transformation-defective, partial src deletion mutants of RSV used to generate rRSV's, we submit that the src genes of rRSV's could have been generated by cross-reactivation of nonoverlapping src deletions or mutation of src variants possibly present in transformation-defective, partial src deletion mutants of RSV. To prove experimental transduction, unambiguous markers need to be identified, or it would be necessary to generate rRSV's with molecularly cloned transformation-defective, partial src deletion mutants of RSV. Although our evidence casts doubt on the idea that specific src sequences of rRSV's originated by transduction, the close relationship between viral src and cellular src -related sequences argues that src genes originated at one time in evolution from the cell by events that involved illegitimate recombination and deletion of non- src sequences that interrupt the cellular src locus.
ABSTRACT We report the generation of a multimodal cell census and atlas of the mammalian primary motor cortex (MOp or M1) as the initial product of the BRAIN Initiative Cell Census Network (BICCN). This was achieved by coordinated large-scale analyses of single-cell transcriptomes, chromatin accessibility, DNA methylomes, spatially resolved single-cell transcriptomes, morphological and electrophysiological properties, and cellular resolution input-output mapping, integrated through cross-modal computational analysis. Together, our results advance the collective knowledge and understanding of brain cell type organization: First, our study reveals a unified molecular genetic landscape of cortical cell types that congruently integrates their transcriptome, open chromatin and DNA methylation maps. Second, cross-species analysis achieves a unified taxonomy of transcriptomic types and their hierarchical organization that are conserved from mouse to marmoset and human. Third, cross-modal analysis provides compelling evidence for the epigenomic, transcriptomic, and gene regulatory basis of neuronal phenotypes such as their physiological and anatomical properties, demonstrating the biological validity and genomic underpinning of neuron types and subtypes. Fourth, in situ single-cell transcriptomics provides a spatially-resolved cell type atlas of the motor cortex. Fifth, integrated transcriptomic, epigenomic and anatomical analyses reveal the correspondence between neural circuits and transcriptomic cell types. We further present an extensive genetic toolset for targeting and fate mapping glutamatergic projection neuron types toward linking their developmental trajectory to their circuit function. Together, our results establish a unified and mechanistic framework of neuronal cell type organization that integrates multi-layered molecular genetic and spatial information with multi-faceted phenotypic properties.
The polymerase chain reaction (PCR) has many potential applications in the field of nucleic acid diagnostics. In particular, it has been successfully applied to the detection of pathogens present in low copy numbers such as the human immunodeficiency virus type 1. Here we describe a time-resolved fluorescence-based hybridization assay which, combined with the PCR, offers an extremely sensitive method for the detection of nucleic acids. In this assay format, the PCR is run by standard procedures and the subsequent hybridization reaction is carried out in solution by using two oligonucleotide probes, one biotinylated and one labeled with europium (Eu3+). The sandwich hybrids are then collected onto a streptavidin-coated microtitration well, and the bound Eu3+ is measured in a time-resolved fluorometer. This assay is rapid, user friendly, and quantitative and lends itself to automation. The application of this assay to the detection of human immunodeficiency virus type 1 is described.
