First Report of Kudzu (Pueraria montana) Infections by Tobacco ringspot virus in Mississippi.

Virus-like symptoms were observed in several kudzu patches in Mississippi during a survey of viruses infecting soybean carried out in late summer/fall of 2013 as a part of a project funded by the Mississippi Soybean Promotion Board. Symptomatology consisted of chlorotic mottle and ringspots, vein-associated feathering, necrosis, and leaf deformation, which were often observed in combination on the same plant. In order to identify the virus(es) involved in the disease, young leaves from a symptomatic kudzu sample collected in Kemper County were crushed in 10 volumes of 0.1 M phosphate buffer (pH 7.2) and mechanically inoculated onto celite-dusted leaves of two soybean varieties (Asgrow AG4605 and AG4730), each represented by 10 plants. Sap from an asymptomatic kudzu sample from Oktibbeha County was used as a control. Both varieties reacted by systemic mottle, stunting, and apical leaf necrosis approximately 2 weeks after inoculation, while no symptoms could be observed in controls. Partially purified preparations from both symptomatic soybean cultivars exhibited the presence of putative intact and empty spherical virus particles ~30 nm in diameter. ELISA tests with antisera to several soybean viruses were performed on the original kudzu sample and inoculated AG4605 and AG4730 soybean plants. These tests revealed the presence of Tobacco ringspot virus (TRSV) in all symptomatic samples. In order to better understand the incidence of this virus in kudzu in Mississippi, a total of 127 samples from 28 counties were collected during October 2013 and tested using ELISA. A total of 11 samples collected in 8 different counties were positive for TRSV. To further confirm these results, one step RT-PCR test was performed on total nucleic extracts from all ELISA-positive and four negative kudzu samples using TRSV-specific primers (3). A specific PCR product of 766 bp was present in all ELISA-positive samples and positive controls, whereas no visible bands were present in negative samples. PCR products generated from samples, collected in Kemper, Tippah, and Jefferson Davis counties, were cloned and custom sequenced. Pair-wise comparisons indicated conserved nucleotide (95 to 98%) and amino acid (98 to 99%) contents among sequenced products. Kudzu isolates from Mississippi shared 91 to 96% and 98 to 99% conserved nucleotides and amino acids, with TRSV sequences currently available in the NCBI/GenBank database. This is the first report of TRSV infection of kudzu in Mississippi. The possible implications to the soybean industry are yet to be determined since kudzu occupies approximately 202,000 ha in Mississippi and TRSV has historically been reported associated with bud blight in soybean (1). Nonetheless, results of our study, along with the recent report from Louisiana (2), strongly suggest that kudzu, due to its widespread distribution in the region, may represent a major reservoir of TRSV in the southeastern United States. References: (1) G. L. Hartman et al. Compendium of Soybean Diseases. American Phytopathological Society, St. Paul, MN, 1999. (2) Khankhum et al. Plant Dis. 97:561, 2013. (3) S. Sabanadzovic et al. Plant Dis. 94:126, 2010.

Comparative Susceptibility of Kudzu Accessions from the Southeastern United States to Infection by Phakopsora pachyrhizi.

Soybean rust, caused by Phakopsora pachyrhizi, was first discovered in the continental United States in the fall of 2004. The potential for economic loss in the United States hinges largely on whether or not the pathogen can survive winters in the absence of soybean. Kudzu (Pueraria lobata) is known to be a host for P. pachyrhizi in Asia and South America and is widely distributed in the southern United States. This study examined reactions of kudzu collected from several areas of the southeastern United States to three isolates of P. pachyrhizi, one each from Alabama, Louisiana, and Brazil. Susceptible tan (TAN) lesions, resistant reddish-brown (RB) lesions, and immune (IM) response, previously described on soybean, were produced on kudzu based on the evaluation of 125 plants. However, in contrast to soybean, the RB response on kudzu was common, with approximately 50% frequency. IM responses to at least one isolate were observed on five individual plants, and two plants were immune to all three pathogen isolates used in the test. TAN lesions averaged 3.2 uredinia per lesion with an average diameter per uredinium of 121 µm. In contrast, RB lesions had an average of 0.3 uredinia per lesion with an average uredinial diameter of 77 µm. In 25 of 39 (64%) instances in which multiple plants were tested from a site, each reacted the same to the individual pathogen isolates. This suggested a tendency for plants at specific sites to be genetically identical with respect to rust reaction. Only 19 of 125 (15%) individual plants produced a different reaction to one isolate than to the other two isolates. When four kudzu plants previously shown to produce only TAN lesions to P. pachyrhizi isolates Alabama 04-1, Brazil 01-1, and Louisiana 04-1 were inoculated with eight additional isolates from several areas of the world, all 11 isolates produced only TAN lesions. Likewise, when five other plants previously shown to produce only RB lesions when inoculated with the three isolates were inoculated with the 11 isolates, all produced only RB lesions. These results suggest that susceptibility or resistance to P. pachyrhizi in individual kudzu plants often is broad, extending over a wide range of P. pachyrhizi isolates.

Comparative Susceptibilities of Legume Species to Infection by Phakopsora pachyrhizi.

Knowledge of the host range of Phakopsora pachyrhizi is important to agriculture in the United States because of the distinct possibility that economic losses could occur to crops other than soybean. Furthermore, it is possible that alternative hosts could provide a means of overwintering of the pathogen, providing inoculum to initiate epidemics in future years. To clarify the potential importance of soybean rust on nonsoybean legumes and their role in overwintering of the disease, multiple accessions of clover, cowpea, pea, kudzu, lima bean, snap bean, and single accessions of coffee senna, Florida beggarweed, hemp sesbania, hyacinth bean, partridge pea, and showy crotalaria were inoculated under greenhouse conditions with urediniospores of P. pachyrhizi; infected soybean plants served as a control. The four criteria used to assess susceptibility were lesion density, proportion of lesions with sporulating uredinia, average number of uredinia per lesion, and average uredinia diameter, each determined 2 weeks following inoculation. Based on lesion densities, percentage of lesions with sporulation, and average numbers of uredinia per lesion, soybean, kudzu, and pea were the most susceptible species, followed by snap bean. However, because infected pea plants defoliated rapidly, urediniospore production presumably was limited, lessening the potential for epidemics on pea. Cultivars of snap bean produced numerous brown to reddish-brown lesions, many of which sporulated, but numbers of uredinia per lesion were lower than on soybean, kudzu, or pea. The presence of both tan (susceptible) and reddish-brown (resistant) lesions on kudzu demonstrated physiological differentiation on that host. Some kudzu plants appeared to be potentially excellent hosts for overwintering of the disease. The average number of uredinia per lesion appeared to be a valid measurement with which to compare host susceptibilities, and may have epidemiological significance. High susceptibility of a host was characterized by numerous uredinia with a wide range of sizes within individual lesions. In contrast, low susceptibility to rust was characterized by no or a few small uredinia.

Recurrence of dysplasia after loop electrosurgical excision procedures with long-term follow-up.

OBJECTIVE: The aim of this study was to determine the rates of recurrent dysplasia with longer follow-up durations and to determine whether margin status and other variables were associated with recurrence. STUDY DESIGN: A retrospective chart review was performed for all women who underwent a loop electrosurgical excision procedure at Wilford Hall Medical Center, Lackland Air Force Base, Texas, between January 1993 and December 1994. Extracted information included age, parity, indication for the loop electrosurgical excision procedure, histologic classification of the loop electrosurgical excision procedure specimen, margin status, and whether a "deep" (endocervical) pass had been performed. Follow-up data included findings of repeated cytologic examination, colposcopy, and biopsy if performed. RESULTS: The mean duration of follow-up for all women was 24 months. Margins were positive in 28%, with 73% of these being endocervical. The overall recurrent dysplasia rate was 31%, with a mean time to recurrence of 11.9 months. Participants with any positive margins had a higher recurrence rate than did those with negative margins (47% vs 26%; P = .009). High-grade lesions at the margin were more commonly associated with recurrence than were low-grade lesions relative to those with clear margins (high-grade lesion vs negative margins, 55% vs 26%; P = .003; low-grade lesion vs negative margins, 36% vs 26%; P = .34). Recurrence was not associated either with the performance of an endocervical pass or with the histologic diagnosis of the loop electrosurgical excision procedure specimen. CONCLUSION: With comprehensive long-term follow-up, positive margins on loop electrosurgical excision procedure specimens were shown to be a risk factor for recurrence of cervical dysplasia, particularly when high-grade lesions were seen at the margin. Recurrence was also considerable among women with negative margins. Women should be counseled regarding this risk, and the importance of follow-up should be emphasized.

K-ras mutations in Müllerian inclusion cysts associated with serous borderline tumors of the ovary.

OBJECTIVE: Müllerian inclusion cysts (MIC) are small benign appearing glands that are occasionally noted in lymph nodes and peritoneal biopsies. They occur most frequently in women with serous ovarian tumors, with borderline tumors (SBOT) having a higher incidence than invasive cancers. The aim of this study was to examine whether MIC and SBOT have identical K-ras mutations, which would suggest that they are related. Methods. Six patients in whom adequate tissue was available from SBOT, MIC, and normal tissue were identified from a consecutive series of patients with SBOT who underwent lymph node sampling from 1992 to 1997 at Duke University Medical Center. DNA extraction was performed using laser capture microdissection. Exon 1 of the K-ras gene was amplified using PCR and subjected to single-strand conformation analysis to screen for mutations. Shifted bands were sequenced to confirm the presence of mutations. RESULTS: Mutations in codon 12 of K-ras were found in three of six (50%) SBOT. In two of these three cases, the identical mutation was found in the SBOT and the MIC (gly to val in both cases), but not in the corresponding normal DNA. In one case, a mutation was seen in the ovarian tumor (gly to asp), but not in the corresponding MIC. CONCLUSIONS: Mutations in codon 12 of the K-ras gene are a hallmark of serous borderline tumors. The presence of identical K-ras mutations in some SBOT and their associated MIC suggests that they are related processes. Both may arise due to a field effect, or alternatively some MIC may represent metastases from the primary ovarian tumor.

Differential localization of placental extracellular superoxide dismutase as pregnancy progresses.

OBJECTIVE: The aim of this study was to determine placental localization and activity of extracellular superoxide dismutase, a nitric oxide modulator, during early gestation and to correlate these characteristics with fetal vascular development. STUDY DESIGN: First-trimester (n = 10) and second-trimester (n = 10) villi were obtained at elective pregnancy termination. Extracellular superoxide dismutase was localized by means of an immunoperoxidase method. Activity was measured by determining the inhibition of cytochrome c reduction at pH 10 and messenger ribonucleic acid expression by in situ hybridization. RESULTS: Extracellular superoxide dismutase was intracellular within villous trophoblasts until 17 weeks' gestation, when it relocated to the villous extracellular matrix. Activities were similar between first- and second-trimester villi. In situ hybridization confirmed extracellular superoxide dismutase messenger ribonucleic acid within trophoblasts throughout gestation. CONCLUSION: Extracellular superoxide dismutase is produced by trophoblasts early in pregnancy, but it remains intracellular until 17 weeks' gestation, which may be related to fetal vascular development.

Some mullerian inclusion cysts in lymph nodes may sometimes be metastases from serous borderline tumors of the ovary.

Glandular inclusions that appear morphologically benign are occasionally found in lymph nodes as well as in peritoneal and omental biopsies. In patients with gynecologic malignancies, the nature and significance of these mullerian inclusion cysts (MIC) present a diagnostic challenge with regard to whether they are benign and incidental or are related to the coincident tumor for which surgery is being performed. Sixty-two cases of MIC were prospectively identified during a 6-year period. The frequencies were calculated and stratified by lymph node chain distribution, primary tumor site, and primary tumor type. MIC appeared as small cysts lined by a serous (mullerian)-type, cytologically bland, cuboidal to columnar epithelium with a simple architecture. Among 62 women, MIC was found in lymph nodes (27 cases), pelvic peritoneum (19 cases), omentum (16 cases), bowel serosa (9 cases), uterine serosa (8 cases), and parametrial connective tissues (4 cases). Among a set of 417 consecutive cases in which lymphadenectomy was performed, 46 (11%) women had MIC. The MIC involved multiple sites (26 cases in the peritoneum/omentum and 27 in lymph nodes). The primary tumor was in the ovary in 32 of the 46 women with MIC (70%) and of these, 17 were borderline serous (53%). Sixty-two of 6,154 lymph nodes examined contained MIC (1.0%). 3.2% of nodes contained MIC in which the primary tumor arose in the ovary, but only 0.1% with either endometrial or cervical tumors (chi2, p <0.00001). The lymph nodes most often involved by MIC were from para-aortic sites (40%), which reflect the primary drainage route from the ovary. Not uncommonly, neighboring areas in the same lymph node group with MIC disclosed separate foci of obvious metastatic borderline tumor (4 of 10; 40%). In summary, the increased frequency of MIC in lymph nodes sampled for primary ovarian malignancies suggests that MIC in some cases, rather than being benign, incidental inclusions, are more likely bland-appearing forms of metastatic tumor. The preponderance of inclusions occurs with serous ovarian tumors of borderline malignancy, and the inclusions are overrepresented in the lymph nodes that primarily receive drainage from the ovary.

Signet ring adenocarcinoma metastatic to the bronchus and mimicking goblet cell hyperplasia. A case report.

BACKGROUND: Goblet cells in the lower respiratory tract are metaplastic bronchial epithelial cells usually associated with asthma or chronic bronchitis. Goblet cells acquire their name by a tendency to distend with mucus, with subsequent distortion in cell shape. Due to similarity of shape, metaplastic goblet cells and signet ring cells can be easily confused in cytologic samples. CASE: A 55-year-old male with a history of gastrointestinal adenocarcinoma underwent brushing, washing and biopsy of a bronchial lesion. The bronchial wash and brush samples showed a very cellular specimen, with large aggregates of distended columnar cells. These were arranged in long strips, thick bundles and occasional three-dimensional aggregates. Some aggregates contained numerous rounded cells with markedly distended cytoplasm. The rounded cells were slightly larger than the distended columnar cells. These cells had a relatively large but innocuous-appearing nucleus displaced to the periphery of the cell. The corresponding bronchial biopsy revealed signet ring adenocarcinoma, presumably metastatic from the gastrointestinal primary. CONCLUSION: Signet ring adenocarcinoma, either primary or metastatic, can be difficult to diagnose in cytologic and histologic specimens. There are numerous mimics of signet ring cells, both benign and neoplastic. In respiratory cytologic specimens, one of the benign imposters is goblet cell metaplasia.

Goblet-cell mucinous epithelium lining the endometrium and endocervix: evidence of metastasis from an appendiceal primary tumor through the use of cytokeratin-7 and -20 immunostains.

Differential staining with cytokeratin (CK)-7 and CK-20, two members of a complex family of proteins in human epithelial cells, proved critical in showing that the extremely well-differentiated goblet-cell (intestinal) mucinous epithelium lining the surface of the endometrium and endocervix in two patients and the fallopian tube in one was identical to that of the coincident appendiceal neoplasms. One of these patients also had a large ovarian tumor that grossly and microscopically resembled a mucinous cystadenoma of borderline malignancy and would have been considered primary except for the CK stains (CK-20 positive and CK-7 negative), which suggested metastasis from the appendix, presumably by a transtubal route.