The role of imaging studies in urinary tract infection.

Technology has permitted significant advances in the imaging and early diagnosis and treatment of many serious urinary tract infections. The armamentarium is vast, with CT scan, ultrasound, nuclear scan, and magnetic resonance imaging on the horizon, with more studies certainly to follow. Nothing, however, can replace the value of proper assessment of clinical signs and symptoms and the logical pursuit of a diagnosis with knowledge of the capabilities and deficiencies of each study.

Differential antibody responses of individuals infected with AIDS-associated retroviruses surveyed using the viral core antigen p25gag expressed in bacteria.

Infection with the retrovirus that is the etiological agent of acquired immune deficiency syndrome (AIDS) is characterized by the development of antiviral antibodies. To generate reagents for studying immune responses to individual viral proteins, we have produced viral antigens in microorganisms by recombinant DNA techniques. Large amounts of the major core protein (p25gag) of an isolate of the AIDS retrovirus (AIDS-associated retrovirus; ARV-2) have been directly expressed in Escherichia coli. Recombinant p25gag (R-p25gag) has been purified and used in an enzyme-linked immunosorbent assay (ELISA) for antibodies to p25gag. Serum samples obtained from 100 individuals with AIDS, AIDS-related complex (ARC), or potential exposure to the virus through sexual contact with AIDS or ARC patients (contacts) were tested first in an ELISA with disrupted whole virus to determine which of the subjects had mounted an antibody response to the virus (virus seropositive) and then in the p25gag ELISA to determine if they had antibodies to this particular viral antigen. We observed a decrease in the proportion of virus seropositive individuals with antibodies to p25gag among patients groups in which the disease was more advanced; contacts were often positive (71%), ARC patients less frequently positive (48%), and AIDS patients only rarely positive (16%). Our results suggest that monitoring p25gag seropositivity of infected individuals may be useful for predicting either the prognosis or the stage of the disease.

Human Cu/Zn superoxide dismutase cDNA: isolation of clones synthesising high levels of active or inactive enzyme from an expression library.

The molecular cloning and nucleotide sequence of the cDNA for human Cu/Zn superoxide dismutase (SOD) is reported. The tacI promoter has been used to direct the synthesis in E. coli of this SOD which is soluble, stable, and of normal specific activity. The N-terminal methionine is removed from this protein. A construction with a ribosome binding site identical to that of the lacz gene 5' of the initiator methionine codon, resulted in low levels of SOD. An in vitro mutagenesis procedure was used to randomize the four nucleotides preceding the initiator methionine codon and the silent third positions of the codons specifying the second and third amino acids. Analysis of a sample of 500 clones showed that ca. 25 clones synthesised 5% or more of soluble cell protein as SOD. The nucleotide sequences of high level expressors showed a predominance of A and T residues in the variable positions 5' of the initiator methionine codon. An SOD mutant (ala4----gln) was discovered during the sequencing and shown to lack dismutation activity. Secondary structure predictions for the 5' regions of the mRNAs from high and low level expressors support the hypothesis that initiation of translation is much reduced if part of the region complementary to 16s rRNA is base paired in a stem structure.

Identification of a unique mouse mammary tumor virus in the BALB/cNIV mouse strain.

We examined the genetic structure, in terms of restriction endonuclease recognition sites, of the milk-transmitted, low-oncogenic mouse mammary tumor virus (MuMTV) of the BALB/cNIV mouse strain. An analysis with EcoRI documented the presence of acquired cNIV proviruses in the mammary tumor DNAs of BALB/cNIV animals. A comparison of tumor DNAs digested with PstI showed that both the cNIV MuMTV and C3Hf MuMTV proviruses lacked the 4.3- and 1.1-kilobase pair fragments characteristic of C3H MuMTV patterns. An examination of mammary tumor and normal, nonmammary tissue DNAs with BamHI supported the idea that the cNIV MuMTV is identical to the C3Hf MuMTV and demonstrated that these two low-oncogenic proviruses are identical to the high-oncogenic C3H MuMTV provirus with respect to a pair of BamHI sites which define a 1.3-kilobase pair fragment. For each of the three MuMTV strains, we also mapped DNAs generated in isolated virions by reverse transcription of their genomic RNAs. Our results showed that cNIV and C3Hf MuMTV are distinct entities by virtue of an additional PstI site within the cNIV long terminal repeat sequence. Another unique feature of cNIV MuMTV revealed by the analysis of virion-generated DNAs was the existence of a family of genomes within the cNIV population. We concluded that cNIV is distinct from its presumptive C3Hf MuMTV predecessor.

Detection of acquired provirus sequences in mammary tumors from low-expressor, low-risk mice.

Murine mammary tumor virus (MuMTV) provirus sequences in the DNA from early-occurring (average age 10 mo) and late occurring (age greater than 20 mo) tumors in BALB/cfC3H mice were analyzed by Eco RI restriction endonuclease mapping procedures. All early tumors were MuMTV antigen-positive mammary adenocarcinomas that contained the 0.92- and 4.0-kilo base (kb) exogenous C3H MuMTV-specific Pst I restriction endonuclease fragments. All but 1 of the late mammary adenocarcinomas had MuMTV antigens detected by peroxidase antiperoxidase staining, and all contained the 0.92- and 4.0-kb exogenous virus Pst I fragments. Three late nonmammary tumors lacked both MuMTV antigens and acquired provirus sequences. Greater numbers of MuMTV sequences were detected in both early and late-arising mammary tumors by Eco RI restriction endonuclease mapping than were detected in tissues from uninfected BALB/c mice. However, neither the number nor the location of MuMTV proviruses correlated with tumor latent period.

Identification and partial characterization of an endogenous form of mouse mammary tumor virus that is transcribed into the virion-associated RNA genome.

Restriction mapping demonstrated the presence of several distinct proviral forms of mouse mammary tumor virus in the genome of GR mice. One of these proviruses (GR-MTV-2) was highly amplified in GR 3A cells, a cell line derived from a GR mammary tumor. By the criterion of restriction mapping, the amplified GR-MTV-2 provirus found in GR 3A cells was identical to the provirus found in M1.19 cells, a rat cell line infected with virions obtained from GR 3A culture fluid. This result clearly implies that the GR-MTV-2 provirus in GR 3A cells was transcribed into the virion-associated viral RNA genome. Cleavage of either GR 3A or M1.19 cell DNAs with the restriction enzyme Bg1 II gave rise to a 2.6 x 10(6) dalton GR-MTV-2 proviral fragment (ca. 45% of the viral genome). This fragment was isolated and mapped with thirteen restriction enzymes.

Selective amplification of mouse mammary tumor virus in mammary tumors of GR mice.

DNAs extracted from the mammary tumors of GR mice were analyzed for mouse mammary tumor virus proviral sequences by the restriction enzyme-Southern blot procedure. The tumor DNAs contain more proviral copies of mouse mammary tumor virus than DNA from a nonmalignant tissue. The degree of proviral amplification is small (ca. one to five additional copies) and appears to be variable from tumor to tumor. The restriction patterns of the amplified proviral sequences suggest a clonal origin for the tumor mass. In addition, the restriction patterns observed after digestion with the enzymes BglII and SacI indicate that only one of the proviruses endogenous to GR mice is amplified. The amplified provirus found in GR mammary tumors is identical to the provirus that is missing in GR-Mtv-2- mice, a congenic line exhibiting a low mammary tumor incidence.

Intracellular synthesis of mouse mammary tumor virus polypeptides: indication of a precursor glycoprotein.

Mouse mammary tumor virus polypeptides were detected in the cytoplasm of mouse mammary tumor cell cultures using immunological precipitation techniques. The anti-mouse mammary tumor virus serum precipitated the major virion glycoproteins gp49 and gp37.5/33.5 and a viral-related nonvirion glycoprotein of 76,000 daltons. Subcellular fractionation studies revelaed that the cell-associated virion glycoproteins were present in the membrane fraction. Pulsechase experiments indicated that a viral-related nonvirion glycoprotein of 76,000 daltons may be a precursor to one or more of the virion glycoproteins.

Identification of a precursor protein to the major glycoproteins of mouse mammary tumor virus.

Mouse mammary tumor virus-producing cultures of mouse mammary tumor cells synthesize a viral-related polypeptide of molecular weight of 73,000 (gp 73) which is rapidly labeled during a short pulse but disappears during the chase concomitantly with the appearance of label in the virion glycoproteins gp 49 and gp 37.5/33.5. The addition of the protein synthesis-inhibitor cycloheximide to the chase medium has little effect on this conversion. Treatment of the proposed precursor with alpha-chymotrypsin leads to the formation of a polypeptide of molecular weight 49,000, similar to the major virion glycoprotein. A comparison of tryptic digest maps of the glycoproteins involved supports the hypothesis that both the viral glycoproteins gp 49 and gp 37.5/33.5 are derived from gp 73.