Robert S. Wickert

Primary monophasic synovial sarcoma of the pleura: five cases confirmed by the presence of SYT-SSX fusion transcript.

This study reports five cases of primary pleural monophasic synovial sarcomas and assesses the role of the SYT-SSX fusion transcript in the differential diagnosis. Patients had a mean age of 47 years with no gender predilection. Chest pain and pleural-based masses with effusions characterized the clinical presentations. Each patient underwent a complete surgical resection of the mass. The mean follow-up was 9 months, available in four patients. They were all alive, with no evidence of disease. Histologically, neoplasms were composed of densely packed fusiform cells focally alternating with less cellular areas. No epithelial differentiation was identified at the hematoxylin and eosin level. Keratin and epithelial membrane antigen reactivity was focal and present in four and two tumors, respectively. There was no immunoreactivity for CD34. RT-PCR studies for the presence of a SYT-SSX1 or SYT-SSX2 fusion transcript were positive in every tumor. In comparison, 10 localized fibrous tumors were immunohistochemically negative for keratin and epithelial membrane antigen and positive for CD34. A SYT-SSX fusion transcript was not identified in any of five localized fibrous tumors tested. Identification of the synovial sarcoma-specific chimeric transcript (SYT-SSX1 or SYT-SSX2), in conjunction with immunoperoxidase studies, can be extremely helpful in identifying cases of pleural monophasic synovial sarcoma.

Clonality analysis of B‐lymphoid proliferations using the polymerase chain reaction

Polymerase chain reaction (PCR) based assays are becoming more reliable, simpler, and faster alternatives to traditional Southern blot hybridization (SBH) analysis for the detection of clonal immunoglobulin heavy chain (IgH) gene rearrangements. However, a variety of technical approaches have been reported with markedly different results.

Lymphomatous Polyposis: A Neoplasm of Either Follicular Mantle or Germinal Center Cell Origin

Lymphomatous polyposis (LP) is generally thought to be an expression of non-Hodgkins lymphoma (NHL) of follicular mantle cell (MC) origin. We report nine patients with LP from more than 3,500 cases of NHL studied by the Nebraska Lymphoma Study Group. Our patients differed from those reported previously in that LP represented a follicular center cell (FCC) NHL in two of the nine cases, with the remainder consisting of MC NHL. Three patients developed LP during a relapse of previously diagnosed and treated extraintestinal MC NHL (parotid gland, tonsil, and inguinal lymph node, respectively), whereas the other six patients presented with primary LP. In seven of the nine LP cases, a large mass predominated among a myriad of small polyps. The FCC cases were confined to the small intestine, whereas the MC cases were either pan-intestinal or colonic on their localization. Two MC cases studied by Southern blotting exhibited rearrangement of the bcl-1 locus. Bcl-2 rearrangement was not detected in any of the nine cases when studied by either a polymerase chain reaction-based assay (seven cases) or by Southern blotting (two cases). To date, four patients (three MC, one FCC) have experienced recurrent NHL in gastrointestinal sites. With follow-up ranging from 13 to 147 months, the entire group had a median survival of 41 months (primary MC LP:13, 13, 41, and 77 months; primary FCC LP:45 and 147 months; secondary MC LP:17, 41 and 76 months), and only one patient has died. We conclude that LP is a rare manifestation of NHL of either follicular MC or germinal center cell origin.

The t(14;18) and bcl-2 Expression Are Present in a Subset of Primary Cutaneous Follicular Lymphoma Association With Lower Grade

According to the European Organization for Research and Treatment of Cancer classification, primary cutaneous follicle center cell lymphoma is not associated with the t(14;18)(q32;q21) and only rarely expresses bcl-2 protein. To further investigate this issue, we evaluated a series of 20 patients (14 men, 6 women) with primary cutaneous follicular lymphoma (PCFL). The presenting skin lesion was located in the head and neck region in 16 of 20 patients. Most cases were grade 2 (6/20) or grade 3 (13/20), and all had a follicular architecture. Immunohistochemical analysis demonstrated bcl-2 expression in 8 cases (40%), and expression was inversely related to the grade. Of 7 grade 1 or 2 cases, 5 (71%) were positive, whereas only 3 (23%) of 13 grade 3 cases were positive for bcl-2. Clonal immunoglobulin heavy chain gene rearrangements were detected in 9 (45%) of 20 cases. In 4 (20%) of 20 cases, we identified the major breakpoint of the t(14;18) by polymerase chain reaction, 3 of which were grade 1 or 2. We conclude that bcl-2 protein expression and the t(14;18) are present in a subset of PCFL, particularly in lower grade cases.

Spontaneous Complete Regression of High Grade Non-Hodgkin's Lymphoma Morphologic, Immunohistochemical, and Gene Amplification Analyses

Background. Spontaneous regression of non‐Hodgkins lymphoma, occasionally reported in low grade groups, is a rare phenomenon in high grade groups. Clonal proliferation has not been confirmed in the majority of reported cases. In this woman, age 58 years, who had been diagnosed as having high grade immunoblastic lymphoma after excision of a single cervical lymph node, the remaining bilateral cervical, inguinal, and axillary adenopathy regressed completely without any cytotoxic treatments 22 days after biopsy. At the time of this writing, the patient has been free of disease for 24 months.

High-resolution analysis of immunoglobulin heavy-chain gene rearrangements using denaturing gradient gel electrophoresis.

The detection of clonality in B-cell lymphomas has been facilitated by polymerase chain reaction (PCR) analysis of the immunoglobulin heavy-chain gene (IgH) complementarity determining region 3 (CDR3) and size fractionation by polyacrylamide gel electrophoresis (PAGE). However, the detection of minor clonal populations and biallelic rearrangements and the isolation of monoclonal products from gels are sometimes problematic. This study evaluated whether denaturing gradient gel electrophoresis (DGGE), a technique that separates DNA based on nucleotide sequence rather than length, could alleviate these problems. A total of 32 selected cases was studied with a diagnosis of monoclonal (n = 10). polyclonal (n = 9). and indeterminate (n = 13) IgH gene rearrangements, which were determined by analysis of seminested IgH CDR3 PCR products in 8% PAGE. These cases were evaluated using DGGE of seminested IgH CDR3 PCR products that included a 40-bp GC clamp on the Jh primer. DGGE allowed the discrimination of monoclonal populations in 9 of 13 cases where 8% PAGE results were indeterminate. In addition, DGGE demonstrated biallelic IgH rearrangements in three cases where 8% PAGE revealed only one predominant product. DGGE facilitated the purification and isolation of clonal IgH CDR3 products for sequencing without prior cloning. As an adaptation of current IgH PCR protocols. DGGE can enhance the construction of tumor-specific CDR3 primers/probes for investigations of minimal residual disease.

EWS/FLI-1 fusion signal inserted into chromosome 11 in one patient with morphologic features of Ewing sarcoma, but lacking t(11;22)

A reciprocal t(11;22)(q24;q12) is found in 85% of Ewing sarcomas (ES) cases. We report a case of a child with ES, in whom trisomy 8 was observed as the sole chromosomal abnormality. Fluorescence in situ hybridization---using a set of probes that localize to 22q12 (EWS) and 11q24 (FLI-1) and usually show the translocation as fusion (red-green) signal on der(22)---showed a fusion signal on der(11) suggesting an insertion as the mechanism that led to the EWS-FLI-1 gene rearrangement. Reverse transcriptase-polymerase chain reaction studies revealed the presence of two EWS/FLI1 fusion gene products.

Molecular identification of Mycobacterium chimaera as a cause of infection in a patient with chronic obstructive pulmonary disease

This report describes a case of Mycobacterium chimaera infection in a patient with a history of chronic obstructive pulmonary disease where the organism was identified by using molecular methods. M. chimaera was identified from fresh lung tissue and from an instrument-negative mycobacterial growth indicator tube broth culture. The utility of using sequence analysis of the internal transcribed spacer region for the rapid identification of a slow-growing nontuberculous Mycobacterium spp. where conventional culture methods were not successful was shown.

An Outbreak of Avian Mycobacteriosis Caused by Mycobacterium intracellulare in Little Blue Penguins (Eudyptula minor)

Abstract Mycobacterium intracellulare (MIT) was diagnosed postmortem by culture and supporting histopathology in seven birds from a flock of little blue penguins (Eudyptula minor) at the Henry Doorly Zoo (HDZ). These birds represented 20% of the deaths in the population over a 4 yr period. Clinical signs in affected birds included severe respiratory distress characterized by open-mouth breathing with chronic debilitation. On exam, plaques were noted in the larynx, trachea, and soft tissue of the caudal oropharynx. Index cases were identified on necropsy in two birds on loan to another institution in 2003. Following a case confirmed antemortem at the HDZ, a three-drug protocol of rifampin (15 mg/kg p.o. s.i.d.), ethambutol (15 mg/kg p.o. s.i.d.), and clarithromycin (10 mg/kg p.o. s.i.d.) was started on this bird in 2004 and extended to the entire flock in 2005. Gastric wash, fecal samples, and throat plaques were obtained antemortem on five birds within the flock, selected because of the presence of oral plaques, and tested by culture followed by a polymerase chain reaction assay. MIT was detected in gastric washes from four birds and in throat plaques from all five. Three more birds died during treatment. After the seventh bird died, antimicrobial susceptibility testing performed in July 2007 indicated that the MIT was now resistant to most antibiotics tested, including rifampin and ethambutol. The treatment regimen was changed to minocycline (10 mg/kg p.o. b.i.d.) and clarithromycin (10 mg/kg p.o. s.i.d.). Oral plaques were not seen on monthly rechecks of the flock through November 2008. The proposed mechanism of transmission is exposure to wild birds but the source has not been determined. These cases of avian mycobacteriosis caused by MIT are the first known cases reported in little blue penguins.

HTLV-I sequence in lymphoproliferative disorders.

Several recent studies reported the detection of partially deleted HTLV-I provirus in biopsies of lesions from patients with mycosis fungoides (MF) and T-cell anaplastic large-cell lymphoma. We studied lesions from 59 patients (21 B-cell lymphomas: 16 diffuse and five follicular; 11 cutaneous T-cell lymphomas, including seven MF; one T-immunoblastic lymphoma; 10 diffuse anaplastic large-cell lymphomas: two B, four T, and four of indeterminate phenotype; three Hodgkin s lymphomas; eight atypical lymphoid proliferations; four other lymphoid lesions, and one squamous-cell carcinoma) using primers to the gag, pol and pX regions of HTLV-I in the polymerase chain reaction (PCR) to detect relevant sequences. A total of 10 patients showed one or more PCR-amplifiable products, including five of 11 patients with cutaneous T-cell lymphomas (45%) as compared with one of 21 patients with B-cell lymphomas (4.3%). We did not find a high incidence of positivity in anaplastic large-cell lymphomas, as reported previously. Detectable HTLV-I sequences were not limited to any subtype of lymphoma, and a pX sequence was detected in a squamous-cell carcinoma. Sequence analysis of one amplified product from each of the three regions studied showed a 94.2, 100, and 98.9% homology to the corresponding prototypical gag. pol, and pX HTLV-I sequences, respectively, indicating that the amplified sequences were derived from HTLV-I or a very closely related virus. HTLV-I sequences were detected in a significant proportion of patients with cutaneous T-cell lymphoma, but their role in the pathogenesis of the neoplasm is still unclear.