Negative pressure wound therapy, staged excision and definitive closure with split-thickness skin graft for axillary hidradenitis suppurativa: a retrospective study.

OBJECTIVE: Bilateral axillary hidradenitis is a chronic, suppurative, and scarring disease that is most effectively treated by complete excision of all hair-bearing tissues. We assessed our staged procedure for excision and placement of a split-thickness skin graft for bilateral axillary hidradenitis in terms of costs, outcomes, and timing of excision. METHOD: An IRB approved retrospective case analysis was performed on patients that underwent bilateral axillary hidradenitis skin excision with eventual placement of split-thickness skin grafting using the current LSUHSC/University Health hidradenitis surgical treatment protocol. Using ICD-9 codes (705.83) and CPT codes (11041, 11042, 11451, 11600, 11601, 11602, 11603, 11604) we reviewed cases performed at our institution from 1 January 2008 to 24 Febuary 2014 and we selected patients based on bilateral axillary involvement (alone) and >1 year history of active disease. Patients were excluded if resection of tissue encompassed regions outside of the immediately adjacent axillary. RESULTS: A total of seven patients matching criteria for bilateral axillary hidradenitis were selected for analysis. Clinical course, cost and surgical techniques were assessed. Of the seven patients, six required admission throughout their treatment due to lack of funding making use of negative pressure wound therapy at home not possible. These patients stayed an average of 10 days with a mean hospital charge of $35,178 and a mean hospital provider charge of $10,019. No recurrence was demonstrated. All patients attained full range of motion, post grafting. No patient required a further operation due to graft failure. CONCLUSION: Split-thickness skin grafting without use of bilayer dermal regenerative templates yielded definitive results with acceptable cosmesis and functionality, without the added cost of treatments such as a bilayer dermal regenerative template.

Diagnostic monitoring of a changing environment: an alternative UK perspective.

Adaptive management of the marine environment requires an understanding of the complex interactions within it. Establishing levels of natural variability within and between marine ecosystems is a necessary prerequisite to this process and requires a monitoring programme which takes account of the issues of time, space and scale. In this paper, we argue that an ecosystem approach to managing the marine environment should take direct account of climate change indicators at a regional level if it is to cope with the unprecedented change expected as a result of human impacts on the earth climate system. We discuss the purpose of environmental monitoring and the importance of maintaining long-term time series. Recommendations are made on the use of these data in conjunction with modern extrapolation and integration tools (e.g. ecosystem models, remote sensing) to provide a diagnostic approach to the management of marine ecosystems, based on adaptive indicators and dynamic baselines.

Zinc is incorporated into cuticular "tools" after ecdysis: the time course of the zinc distribution in "tools" and whole bodies of an ant and a scorpion.

An understanding of the developmental course of specialized accumulations in the cuticular "tools" of arthropods will give clues to the chemical form, function and biology of these accumulations as well as to their evolutionary history. Specimens from individuals representing a range of developmental stages were examined using MeV - Ion microscopy. We found that zinc, manganese, calcium and chlorine began to accumulate in the mandibular teeth of the ant Tapinoma sessile after pre-ecdysial tanning, and the zinc mostly after eclosion; peak measured zinc concentrations reached 16% of dry mass. Accumulations in the pedipalp teeth, tarsal claws, cheliceral teeth and sting (aculeus) of the scorpion Vaejovis spinigeris also began after pre-ecdysial tanning and more than 48 h after ecdysis of the second instars. Zinc may be deposited in the fully formed cuticle through a network of nanometer scale canals that we observed only in the metal bearing cuticle of both the ants and scorpions. In addition to the elemental analyses of cuticular "tools", quantitative distribution maps for whole ants were obtained. The zinc content of the mandibular teeth was a small fraction of, and independent of, the total body content of zinc. We did not find specialized storage sites that were depleted when zinc was incorporated into the mandibular teeth. The similarities in the time course of zinc, manganese and calcium deposition in the cuticular "tools" of the ant (a hexapod arthropod) and those of the scorpion (a chelicerate arthropod) contribute to the evidence suggesting that heavy metal-halogen fortification evolved before these groups diverged.

Polymerization of 2'-fluoro- and 2'-O-methyl-dNTPs by human DNA polymerase alpha, polymerase gamma, and primase.

Studies were undertaken to assess the ability of human polymerase alpha (pol alpha) and polymerase gamma (pol gamma) to incorporate 2'-fluoro- and 2'-O-methyldeoxynucleotides into DNA. In vitro DNA synthesis systems were used to detect incorporation and determine K(m) and V(max) for 2'-FdATP, 2'-FdUTP, 2'-FdCTP, 2'-FdGTP, 2'-O-MedATP, 2'-O-MedCTP, 2'-O-MedGTP, 2'-O-MedUTP, dUTP, UTP, and FIAUTP, in addition to normal deoxynucleotides. Pol alpha incorporated all 2'-FdNTPs except 2'-FdATP, but not 2'-O-MedNTPs. Pol gamma incorporated all 2'-FdNTPs, but not 2'-O-MedNTPs. In general, 2'-fluorine substitution decreased V(max)/K(m) 2'-FdUTP. Because kinetics of insertion of pol alpha can be affected by the nature of the primer, we examined the ability of pol alpha to polymerize 2'-fluoro- and 2'-O-MedATP and dGTP when elongating a primer synthesized by DNA primase. Under these conditions, both 2'-FdATP and 2'-FdGTP were polymerized, but 2'-O-MedATP and 2'-O-MedGTP were not. Primase alone could not readily polymerize these analogs into RNA primers. Previous studies showed that 2'-deoxy-2'-fluorocytosine (2'-FdC) is incorporated by several non-human DNA polymerases. The current studies showed that human polymerases can polymerize numerous 2'-FdNTPs but cannot polymerize 2'-O-MedNTPs.

An evaluation of the toxicities of 2'-fluorouridine and 2'-fluorocytidine-HCl in F344 rats and woodchucks (Marmota monax).

The toxicities of 2'-fluorouridine (2'-FU) and 2'-fluorocytidine-HCl (2'-FC) were separately evaluated in 2 species, male Fischer 344 (F334) rats and woodchucks. Particular attention was focused on the ability of these nucleosides to induce toxicities similar to those induced by the antiviral drug fialuridine (FIAU). 2'-FU or 2'-FC was administered to F344 male rats by intravenous injection at doses of 5, 50, and 500 mg/kg/day for 90 consecutive days and to male and female woodchucks at doses of 0.75 and 7.5 mg/kg/day for 90 consecutive days. Clinical chemistry, hematology, and urinalysis (woodchuck only) profiles were assessed during and at the termination of the study. At necropsy, organs were weighed and tissues collected for routine histologic analysis. Cytochrome c oxidase activity, citrate synthase activity, and mitochondrial DNA content were measured, and micronucleus formation in the bone marrow (rats only) was evaluated. No adverse clinical effects were observed in either species. Rats treated with high doses of either 2'-FU or 2'-FC had body weights that were 90% of those of controls. 2'-FU and 2'-FC both induced a moderate decrease in the median lymphocyte count, and 2'-FC and 2'-FU induced a mild increase in mean corpuscular hemoglobin and mean corpuscular volume. Both compounds caused slight to moderate, reversible, histologic changes in the spleen and thymus. In the woodchuck, 2'-FC caused a slight increase in mean absolute lymphocytes, and 2'-FC and 2'-FU slightly increased hepatic periportal vacuolation and/or mononuclear cell infiltration. In summary, neither compound showed evidence of the toxicity induced by fialuridine in either species. Although compound effects were observed, none of these effects were considered to be adverse, and the no-observed adverse effect level was determined to be 500 mg/kg/day for both compounds in the male F344 rat and 7.5 mg/kg/day in the woodchuck.

Quantitative and qualitative differences in the metabolism of 14C-1,3-butadiene in rats and mice: relevance to cancer susceptibility.

1,3-Butadiene (butadiene) is a potent carcinogen in mice, but not in rats. Metabolic studies may provide an explanation of these species differences and their relevance to humans. Male Sprague-Dawley rats and B6C3F1 mice were exposed for 6 h to 200 ppm [2,3-14C]-butadiene (specific radioactivity [sa] 20 mCi/mmol) in a Cannon nose-only system. Radioactivity in urine, feces, exhaled volatiles and 14C-CO2 were measured during and up to 42 h after exposure. The total uptake of butadiene by rats and mice under these experimental conditions was 0.19 and 0.38 mmol (equivalent to 3.8 and 7.5 mCi) per kg body weight, respectively. In the rat, 40% of the recovered radioactivity was exhaled as 14C-CO2, 70% of which was trapped during the 6-h exposure period. In contrast, only 6% was exhaled as 14C-CO2 by mice, 3% during the 6-h exposure and 97% in the 42 h following cessation of exposure. The formation of 14C-CO2 from [2,3-14C]-labeled butadiene indicated a ready biodegradability of butadiene. Radioactivity excreted in urine accounted for 42% of the recovered radioactivity from rats and 71% from mice. Small amounts of radioactivity were recovered in feces, exhaled volatiles and carcasses. Although there was a large measure of commonality, the exposure to butadiene also led to the formation of different metabolites in rats and mice. These metabolites were not found after administration of [4-14C]-1,2-epoxy-3-butene to animals by i.p. injection. The results show that the species differences in the metabolism of butadiene are not simply confined to the quantitative formation of epoxides, but also reflect a species-dependent selection of metabolic pathways. No metabolites other than those formed via an epoxide intermediate were identified in the urine of rats or mice after exposure to 14C-butadiene. These findings may have relevance for the prediction of butadiene toxicity and provide a basis for a revision of the existing physiologically based pharmacokinetic models.

Identification of novel metabolites of butadiene monoepoxide in rats and mice.

Differences in the metabolism of 1,3-butadiene (Bd) in rats and mice may account for the observed species difference in carcinogenicity. Previous studies of the metabolic fate of Bd have identified epoxide formation as a key metabolic transformation which gives 1, 2-epoxy-3-butene (BMO), although some evidence of aldehyde metabolites is reported. In this study, male Sprague-Dawley rats and male B6C3F1 mice received single doses of [4-14C]BMO at 1, 5, 20, and 50 mg/kg of body weight (0.014, 0.071, 0.286, and 0.714 mmol/kg of body weight). Analysis of urinary metabolites indicated that both species preferentially metabolize BMO by direct reaction with GSH when given by ip administration. The excretion of (R)-2-(N-acetyl-L-cystein-S-yl)-1-hydroxybut-3-ene (IIa), 1-(N-acetyl-L-cystein-S-yl)-2-(S)-hydroxybut-3-ene (IIb), 1-(N-acetyl-L-cystein-S-yl)-2-(R)-hydroxybut-3-ene (IIc), and (S)-2-(N-acetyl-L-cystein-S-yl)-1-hydroxybut-3-ene (IId) accounted for 48-64% of urinary radioactivity in rats and 46-54% in mice. The metabolites originating from the R-stereoisomer of BMO (IIc and IId) predominated over those arising from the S-stereoisomer (IIa and IIb) in both species. IIc was formed preferentially in mice and IId in rats. The corresponding mercaptoacetic acids, S-(1-hydroxybut-3-en-2-yl)mercaptoacetic acid (IIf) and S-(2-hydroxybut-3-en-1-yl)mercaptoacetic acid (IIg), were identified only in mouse urine (ca. 20% of the recovered radioactivity). 4-(N-Acetyl-L-cystein-S-yl)-1,2-dihydroxybutane (Ia), a metabolite derived from hydrolysis of BMO, accounted for 10-17% of the radioactivity in rat and 6-10% in mouse urine. 4-(N-Acetyl-L-cystein-S-yl)-2-hydroxybutanoic acid (Ib), 3-(N-acetyl-L-cystein-S-yl)propan-1-ol (Ic), and 3-(N-acetyl-L-cystein-S-yl)propanoic acid (Id), also derived from the hydrolysis of BMO, were only present in the rat. Metabolites of 1,2,3,4-diepoxybutane (DEB) were not detected after administration of BMO in rat or mouse urine. This study showed both quantitative and qualitative differences in the metabolism of BMO with varying doses and between species. The data aid in the safety evaluation of Bd and contribute to the interpretation of mathematical models developed for quantitative risk assessment and extrapolation of animals to humans.

Association study of the 5' flanking regions of endothelial-nitric oxide synthase and endothelin-1 genes in familial primary open-angle glaucoma.

1. Endothelium-derived substances are important regulators of the microcirculation. Endothelium-derived nitric oxide (NO), which is catalysed by nitric oxide synthase (NOS), is a potent modulator of vascular tone in the human ophthalmic artery, which is normally in a state of constant vasodilation due to the actions of NO. Endothelin-1 (ET-1) produces vasoconstriction of the anterior optic nerve vasculature and may be associated with glaucomatous optic neuropathy. The aetiology of primary open-angle glaucoma (POAG) remains largely unknown. Thus, alterations in the regulatory sequences of the genes coding for endothelium-derived NOS (eNOS) and ET-1 may have important effects in the development of POAG and were looked for in the present study. 2. In 56 patients with familial POAG and in 100 control subjects with no family history of hypertension or POAG, we examined the 5' flanking sequences of the eNOS and ET-1 genes, which contain many positive and negative regulatory regions affecting gene transcription, using polymerase chain reaction-based single strand conformation polymorphism analysis, to search for alterations. No variant in the promoter region of the ET-1 gene was observed in familial POAG or controls. Using three primer sets spanning 706 b.p. of the eNOS gene, we observed alterations in 11 of 56 (20%) familial POAG members and in seven of 100 (7%) controls. Sequence analysis demonstrated a C/T substitution at the 5' sequence position nucleotide (nt) -690 from the transcription start site, which lies between the cAMP regulatory element (nt -726 to -732) and an activator protein-1 binding domain (nt -655 to -661). 3. In summary, genotypic and allelic frequency analysis found no association between alterations in the promoter region of the ET-1 gene and familial POAG. A variant in the promoter region of the eNOS gene was seen in a significant percentage of familial POAG patients. Future expression studies will determine whether this polymorphism results in altered eNOS gene expression.

PCR-SSCP analysis of the glucocorticoid-responsive element of the atrial natriuretic peptide gene in familial primary open-angle glaucoma.

1. Familial primary open-angle glaucoma (POAG) is a heterogeneous disease of unknown aetiology and the elucidation of the underlying genetic mechanisms contributing to phenotypic expression will be essential if earlier diagnosis of at-risk individuals and more specific medical treatment can be achieved. In a significant percentage of patients with POAG, intraocular pressure increases in response to topical ocular glucocorticoids. 2. Atrial natriuretic peptide (ANP) assists in the regulation of intraocular pressure levels and binding of the glucocorticoid receptor dimer to the glucocorticoid-responsive element in intron 2 of the ANP gene has been shown to increase ANP mRNA levels in vitro. We amplified and examined this sequence in the ANP gene by PCR-SSCP analysis in 100 patients with familial POAG and in 60 normal control subjects. No base alterations in the amplified product were found. 3. Thus, the present study found no evidence for an alteration in the sequence of the glucocorticoid-responsive element of the ANP gene that could alter ANP gene transcription in patients with familial POAG. The mechanism responsible for the increase in intraocular pressure levels in response to glucocorticoids is most likely independent of the glucocorticoid-responsive element in the ANP gene.

Public health approaches to community-based needs. Boston's infant mortality crisis as a case study.

In light of a 10-year infant mortality crisis in Boston, a comprehensive public health approach was undertaken in which an extensive community-based needs assessment was used to develop a citywide maternal and child health improvement agenda. On the basis of the needs assessment, recommendations were made calling for community-based perinatal initiatives and midwifery services as critical elements in care for underserved communities and enhancement of perinatal services. A case description of one perinatal initiative illustrates the challenges of public health practice and describes a practice setting in which midwives provided leadership and guidance by using an interdisciplinary team approach in the implementation of a community empowerment project.

Insertion/deletion polymorphism of the angiotensin-converting enzyme gene and loss of the insertion allele in aldosterone-producing adenoma.

The genetic mechanisms responsible for the formation of adrenocortical adenomas which autonomously produce aldosterone are largely unknown. The adrenal renin-angiotensin system has been implicated in the pathophysiology of these tumours. Angiotensin-converting enzyme (ACE) catalyses the generation of angiotensin II, and the insertion/deletion (I/D) polymorphism of the ACE gene regulates up to 50% of plasma and cellular ACE variability in humans. We therefore examined the genotypic and allelic frequency distributions of the ACE gene I/D polymorphism in 55 patients with aldosterone-producing adenoma, APA, (angiotensin-unresponsive APA n = 28, angiotensin-responsive APA n = 27), and 80 control subjects with no family history of hypertension. We also compared the ACE gene I/D polymorphism allelic pattern in matched tumour and peripheral blood DNA in the 55 patients with APA. The frequency of the D allele was 0.518 and 0.512 and the I allele was 0.482 and 0.488 in the APA and control subjects respectively. Genotypic and allelic frequency analysis found no significant differences between the groups. Examination of the matched tumour and peripheral blood DNA samples revealed the loss of the insertion allele in four of the 25 patients who were heterozygous for the ACE I/D genotype. The I/D polymorphism of the ACE gene does not appear to contribute to the biochemical and phenotypic characteristic of APA, however, the deletion of the insertion allele of the ACE gene I/D polymorphism in 16% of aldosterone-producing adenomas may represent the loss of a tumour suppressor gene/s or other genes on chromosome 17q which may contribute to tumorigenesis in APA.

Biological monitoring of butadiene exposure by measurement of haemoglobin adducts.

The measurement of haemoglobin (Hb) adducts is an important tool for monitoring exposures to industrial chemicals such as ethylene, ethylene oxide and propylene oxide. The aim of the present study was to use this method to monitor occupational exposure to butadiene. The methodology was evaluated with Hb samples obtained by reacting butadiene monoepoxide (BMO), the primary reactive metabolite of 1,3-butadiene, with blood and by dosing BMO to rats and mice. The Hb adducts: N-(2-hydroxy-3-buten-1-yl)valine and N-(1-hydroxy-3-buten-2-yl)valine were determined as the pentaflurophenylthiohydantoin derivatives using the modified Edman degradation procedure and gas chromatography with negative ion chemical ionisation mass spectrometry. The adducts were quantified using d4-N-(2-hydroxyethyl)valine as an internal reference chemical. Relatively high background signals were observed which made accurate quantitation difficult. The lower limit of detection was estimated as 10-20 pmol/g globin. The experiments have demonstrated that appropriate reference samples are required for accurate quantitation and the method requires further refinement to improve the sensitivity of the analysis.

The atrial natriuretic peptide gene in patients with familial primary open-angle glaucoma.

Family history is a major risk factor in the development of primary open-angle glaucoma. The atrial natriuretic peptide system has been implicated in the underlying pathophysiology of the disease. This study looked for any alterations in the ANP gene and 5' proximal promoter regions of the ANP gene, in 53 patients from familial primary open-angle glaucoma families. The ANP gene was amplified by a long-PCR technique from peripheral blood DNA. Gross insertions or deletions in the gene were sought and allelic frequencies at two restriction fragment length polymorphism (RFLP) sites within the gene (Sca I, Hpa II) were compared with allelic frequencies obtained from 60 normal controls with no known family history of glaucoma or ocular hypertension. PCR-based single strand conformation polymorphism analysis was then used to search for possible point mutations in the 5' proximal promoter region of the ANP gene, which is known to contain regulatory elements which modify gene transcription. No gross alterations in the ANP gene or differences in allelic frequencies at the RFLP sites within the gene were observed. PCR-SSCP analysis of the 5' proximal promoter region of the gene revealed mutations in 10 patients in the -595 to -384bp region (19% of patients). Mutations in the 5' proximal promoter region of the ANP gene may contribute to altered ANP transcription in at least a proportion of patients with familial glaucoma.

The effects of cross-link density and chemical exchange on magnetization transfer in polyacrylamide gels.

The effects of polymer structure and water-macromolecule interactions on proton relaxation in an aqueous model polymer have been investigated using quantitative measurements of magnetization transfer. Polyacrylamide gels composed of 95% water, 5% comonomers acrylamide and N,N'-methylene-bis-acrylamide were studied. The structure and rigidity were varied by changing the cross-linking density of the polymer. The polymer showed a biphasic change in transverse relaxation with increasing cross-linking density which was accompanied by a sudden increase in magnetization transfer above 40% cross linking. This change may be attributed to the formation of rigid domains in the polymer which exhibit solid-like behavior with a short T2 (11 microseconds) and a Gaussian lineshape. Water-macromolecule interactions were controlled by varying the pH of the gel. At high pH (> 8), there was an increase in magnetization transfer and transverse relaxivity consistent with a chemical-exchange-mediated interaction between water protons and the polymer. By analyzing the system as two proton reservoirs coupled by magnetization exchange, the proton populations, intrinsic relaxation rates, and exchange rates were estimated, for different degrees of cross linking and pH. This model affords useful insights into the relevance of both supramolecular structure and chemical exchange on relaxation in tissues.

Long-PCR of the ANP gene and PCR-SSCP analysis of the proximal promoter region of the ANP gene in patients with aldosterone producing adenoma.

Previous studies have shown a significant association between allelic frequencies at the ANP gene locus and aldosterone responsiveness to angiotensin in aldosterone-producing adenoma (APA). We searched for any gross insertions or deletions in the ANP gene in APA and any associations between allelic frequencies at the Hpa II and Sca I RFLP sites within the ANP gene and angiotensin-responsive and unresponsive APA and normal subjects. We also searched for possible point mutations in the promoter region of the ANP gene (-595 to transcription start site) in peripheral blood and tumor DNA from 59 patients with APA and in peripheral blood DNA from 39 normal subjects by polymerase chain reaction and single strand conformation polymorphism (PCR-SSCP) analysis. No large alterations in the ANP gene were observed, and no difference in allelic frequencies at the RFLP sites were seen between the two tumor subtypes, angiotensin-responsive and angiotensin-unresponsive APA, or between the APA group and normal subjects. SSCP analysis, however, did reveal mutations in the promoter region of the ANP gene (-375 to -595) in both peripheral blood and tumor DNA from 8 of 59 (14%) patients with APA, compared with only one of 39 normal controls (2.6%). This study suggests that alterations in the proximal promoter region of the ANP gene in APA may be important in the regulation of ANP transcription and may be involved in the underlying pathophysiology of aldosterone-producing adenoma in at least some patients.

The evaluation of methapyrilene for bacterial mutation with metabolic activation by Aroclor-induced, methapyrilene-induced and noninduced rat-liver S9.

The antihistamine methapyrilene (MP) has been shown to be a potent hepatocarcinogen in rats. However, it has demonstrated little genotoxic activity in a wide variety of short-term tests. In this study, Fischer 344 rats were fed a carcinogenic dose of 0.1% methapyrilene in the diet for 10 weeks prior to sacrifice. S9 was prepared from the livers of the control, MP-treated and Aroclor-induced Fischer 344 rats. Each type of S9 was analyzed for mixed function oxidase activity, cytochrome P-450, and protein content. MP was then evaluated for mutagenicity in 6 strains of S. typhimurium (TA1535, TA1537, TA98, TA100, TA2638 and TA104) and one strain of E. coli (WP2uvrA-) using the standard plate-incorporation assay. MP was not mutagenic in any of the 7 bacterial strains when tested at concentrations < or = 10 mg/ml in the presence of each type of S9. However, in the absence of metabolic activation, an approximate 2-fold increase in revertants was noted with strain TA1535. The data from this study show that MP was not converted to a mutagenic metabolite by any of the three S9 types examined. However, the "weak" positive response with strain 1535 in the absence of metabolic activation indicates that further research is needed to elucidate the mechanism of action of this rat carcinogen.

The activity of the coat protein promoter of chloris striate mosaic virus is enhanced by its own and C1-C2 gene products.

DNA fragments from the bidirectional promoter region of the geminivirus chloris striate mosaic virus (CSMV) were cloned into the pUC18-based vector, pG1 producing transcriptional fusions with the beta-glucuronidase (GUS) gene and nopaline synthase terminator sequence. The relative activity of each promoter construct was analyzed by a GUS expression assay of extracts from Zea mays (maize) Black Mexican Sweet protoplasts coelectroporated with the GUS reporter constructs and constructs in which individual CSMV open reading frames (ORFs) were placed under control of a cauliflower mosaic virus 35 S promoter. Weak promoter activity was observed for the promoter of the C1 and C2 ORFs (C1-C2 gene) and for the promoter of the V1 ORF. The activity of these promoters was unaffected by coelectroporation with the CSMV ORF constructs. Moderate activity was observed for the promoter of the V2 ORF (coat protein gene) which was enhanced by coelectroporation of the C1-C2 ORF construct. Sequences within the C1-C2 gene responsible for transactivation of the V2 ORF promoter were mapped close to the A site of a conserved NTP-binding sequence pattern within the C2 ORF. To a lesser extent activity for the promoter of the V2 ORF was enhanced by the V2 ORF construct providing evidence for positive autoregulation of the CSMV coat protein gene.

The nucleotide sequence of the infectious cloned DNA component of tobacco yellow dwarf virus reveals features of geminiviruses infecting monocotyledonous plants.

An infectious clone of the Australian geminivirus tobacco yellow dwarf virus (TobYDV) was constructed from virus-specific double-stranded DNA isolated from infected tobacco and used to demonstrate a single-component genome. The nucleotide sequence of TobYDV DNA comprises 2580 nucleotides. TobYDV DNA has three coding regions, two in the virion sense and one in the complementary sense, homologous to those identified for other geminiviruses, particularly those infecting monocotyledonous (monocot) plants. The complementary sense coding region is comprised of two overlapping reading frames, with an intron of 86 nucleotides. Efficient splicing of the mRNA for this coding region was observed in the infected dicotyledonous (dicot) hosts bean and tobacco despite the intron having an A + U content (57%) more typical of geminiviruses of monocot plants. TobYDV encapsidates a small oligonucleotide able to prime synthesis of the complementary DNA strand in vitro. The TobYDV genome organization, low A + U intron, and encapsidated oligonucleotide primer resemble those of the monocot-infecting geminiviruses. These results strongly suggest that TobYDV is a monocot geminivirus which has become adapted to dicot hosts.