Evolution of Chlamydia trachomatis diversity occurs by widespread interstrain recombination involving hotspots

Chlamydia trachomatis is an obligate intracellular bacterium of major public health significance, infecting over one-tenth of the world's population and causing blindness and infertility in millions. Mounting evidence supports recombination as a key source of genetic diversity among free-living bacteria. Previous research shows that intracellular bacteria such as Chlamydiaceae may also undergo recombination but whether this plays a significant evolutionary role has not been determined. Here, we examine multiple loci dispersed throughout the chromosome to determine the extent and significance of recombination among 19 laboratory reference strains and 10 present-day ocular and urogenital clinical isolates using phylogenetic reconstructions, compatibility matrices, and statistically based recombination programs. Recombination is widespread; all clinical isolates are recombinant at multiple loci with no two belonging to the same clonal lineage. Several reference strains show nonconcordant phylogenies across loci; one strain is unambiguously identified as recombinantly derived from other reference strain lineages. Frequent recombination contrasts with a low level of point substitution; novel substitutions relative to reference strains occur less than one per kilobase. Hotspots for recombination are identified downstream from ompA, which encodes the major outer membrane protein. This widespread recombination, unexpected for an intracellular bacterium, explains why strain-typing using one or two genes, such as ompA, does not correlate with clinical phenotypes. Our results do not point to specific events that are responsible for different pathogenicities but, instead, suggest a new approach to dissect the genetic basis for clinical strain pathology with implications for evolution, host cell adaptation, and emergence of new chlamydial diseases.     

T-cell lymphomas in children

T-cell lymphomas in children are rare and compromise a rather limited spectrum of entities. Relatively frequently observed are precursor T-cell lymphomas mainly presenting as mediastinal tumors with or without leukemia and ALK-positive anaplastic large cell lymphomas (ALCL) with nodal or extra nodal manifestations. In contrast to adults, where peripheral T-cell lymphoma (PTCL) is the most frequent T-cell neoplasia, PTCL in children is exceptional and remains a challenging diagnosis even for experienced haematopathologists. Other rare T-cell lymphomas occasionally seen in children are panniculitis like T-cell lymphomas, NK/T-cell lymphomas and hepatosplenic gamma delta-lymphomas. Of note angioimmunoblastic T-cell lymphoma is never seen in children. Mimickers of T-cell lymphomas like viral or immunological disorders in children are more frequent than manifest T-cell lymphomas and knowledge of these reactive conditions and their differential diagnosis is essential. The recognition of typical histopathological and clinical features along with knowledgeable use and interpretation of immunohistochemical and molecular markers are mandatory for a reliable diagnosis of childhood T-cell lymphomas.     

Recombination pattern of the TCR γ locus in human peripheral T-cell lymphomas

The recombination events of the γ and β T-cell receptor (TCR) loci were analysed in a series of 39 peripheral T-cell lymphomas (PTCLs) in association with the expression of TCR chains. In TCR αβ PTCLs, 22/23 cases showed a γ-gene rearrangement while only 18/23 showed a concomitant β-gene rearrangement. The germline configuration of the β locus was found in angioimmunoblastic lymphadenopathy and lymphoepithelioid lymphomas. Three γδ PTCLs rearranged both γ and β genes. TCR silent PTCLs showed three different patterns of γ- and β-gene rearrangements. Three cases were in germline configuration for both loci; five cases had a rearranged γ and a germline β locus; and five cases had the two loci rearranged. Regarding the variable genes in the γ-rearranged alleles, members of the VγI subgroup were the most frequently presented (39/50), followed by VγII, VγIII, and VγIV (9/50, 1/50, and 1/50, respectively). Joining segment usage was as follows: J1 or J2 (32/50), JP1 or JP2 (17/50), and JP (1/50). Taken together, these data demonstrate that the γ locus is more frequently rearranged whatever the TCR expression. The γ-locus analysis provides a better diagnostic yield than the β locus in the study of PTCL clonality.     

Expression of the alpha/beta and gamma/delta T-cell receptors in 57 cases of peripheral T-cell lymphomas. Identification of a subset of gamma/delta T-cell lymphomas.

Fifty-seven cases of peripheral T-cell lymphoma were studied for cell expression of the T-cell receptor (TCR) chains, using monoclonal antibodies specific for the beta chain (beta F1) of the alpha/beta TCR, and for the delta chain (anti-TCR delta-1) of the gamma/delta TCR. Three different patterns were demonstrated: in 39 cases (69%), the phenotype (CD3+beta F1+TCR delta-1-) was that of most normal T cells. A second pattern was found on six cases (10%), which were of CD3+beta F1-TCR delta-1+ phenotype, and in which DNA analysis showed a clonal rearrangement of the delta locus in the five cases studied. It is suggested that these cases are the neoplastic counterpart of the small subpopulation of normal T cells that express gamma delta receptor. It is of considerable interest that these gamma delta lymphomas had unusual clinicopathologic presentations, as one case corresponded to a lethal midline granuloma and the five others to hepatosplenic lymphomas with a sinusal/sinusoidal infiltration in spleen, marrow, and liver. The fact that the distribution of the neoplastic gamma delta cells in the splenic red pulp resembles that of normal gamma delta cells reinforces the concept of a preferential homing of gamma delta T cells to this tissue. A third pattern (CD3 +/- beta F1-TCR delta-1-) was seen in 12 cases (21%), in which, by contrast to normal post-thymic T cells, no evidence of either alpha beta or gamma delta T cell receptor was found.     

淋巴结细胞毒性自然杀伤/T细胞淋巴瘤

目的 探讨淋巴结细胞毒性自然杀伤（NK/T）细胞淋巴瘤的临床病理学特征。方法 对5例淋巴结细胞毒性NK/T细胞淋巴瘤作临床病理观察及随访、用ISAB法做免疫表型分析（CD35RO、CD8、CD56、CD30、CD20、TIA-1）及EBER1/2原位杂交检测。结果 淋巴结细胞毒性NK/T细胞淋巴瘤的瘤 理组织学特点为：（1）淋巴结结构明显破坏并被瘤细胞所取代：（2）瘤细胞呈多形性;（3）我数肿瘤细胞表达淋巴细胞分化抗原。5例中CD45RO阳性的有4例,其中3例瘤细胞同时呈CD56阳性;1例为无标记细胞性;所有病例的TIA-1和EBER均为阳性。结论 淋巴结细胞毒性NK/T细胞淋巴瘤有特征性的形态改变和免疫表型。提示肿瘤进展及预后不良。     

T-cell and NK-cell lymphomas in the lung

While the lung is frequently involved by systemic lymphoma, primary pulmonary lymphoma accounts for less than 1% of all extranodal ymphomas. In particular, T-cell lymphoma is very rare in the lung, as a primary or secondary lesion. Patients with pulmonary T-cell lymphoma usually present with cough, dyspnea, pain, fever, recurrent infections, and hemoptysis. Typical radiologic features include pulmonary nodules, consolidation, solid pulmonary opacities, cystic changes, hilar adenopathy, and pleural effusions. Patients with these clinical and radiologic findings are frequently presumed to have pneumonia and initially treated with empirical antibiotics. Therefore, CT-guided needle biopsy, bronchoscopic examination, or even wedge biopsy should be considered when clinical symptoms show deterioration despite adequate antibiotic therapy. Precise pathologic diagnosis and molecular characterization are recommended in all cases, following the World Health Organization (WHO) classification. Principles of treatment typically vary with the different histologic types of T-cell lymphoma.     

Immunohistological quantitative analysis of S100 protein-positive cells in T-cell malignant lymphomas, especially in adult T-cell leukemia/lymphomas.

S100 protein-positive cells (S100+ cells) in 36 cases of T-cell lymphoma (T-ML) in the lymph node and 15 cases of T-ML in the skin were analyzed immunohistologically in order to study their quantitative features in adult T-cell leukemia/lymphoma (ATLL). The T-MLs were categorized according to the updated Kiel classification, and the T-cell pleomorphic type (Pleo) was subcategorized into 3 subtypes: Pleo-ATLL, Pleo-clear and Pleo-others. The population of S100+ cells and the first to fifth minimal distances of every S100+ cell were measured on micrographs of paraffin sections that had reacted to anti-S100 protein antibody according to the ABC method. Lymphoblastic and chronic lymphocytic leukemia types showed low populations of S100+ cells and long values of the first minimal distance. T-zone lymphoma without follicles and angioimmunoblastic lymphadenopathy with dysproteinemia-type T-ML had high populations and low values of the first minimal distance. Among the three subtypes of Pleo in the lymph node, Pleo-ATLL gave the highest population and the shortest value of the first minimal distance of S100+ cells, but this trend was not found in the skin. Clusters of more than five S100+ cells were more common in the Pleo-ATLL subtype than in the other two subtypes. The increase and clustering of S100+ cells in Pleo-ATLL suggests that the lymphoma cells act on S100+ cells as a helper.(ABSTRACT TRUNCATED AT 250 WORDS)     

Aberrant phenotypes in peripheral T cell lymphomas.

Seventy six peripheral T cell lymphomas were examined immunohistologically to test their reactivity with a panel of monoclonal antibodies against 11 T cell associated antigens (CD1-8, CD27, UCHL1, and the T cell antigen receptor). Sixty two (82%) lymphomas showed aberrant phenotypes, and four main categories were distinguished as follows: (i) lack of one or several pan-T cell antigens (49, 64% of the cases); (ii) loss of both the CD4 and CD8 antigens (11, 15% of the cases); (iii) coexpression of the CD4 and CD8 antigens (13, 17% of the cases); and (iv) expression of the CD1 antigen (eight, 11% of the cases). No correlation was seen between the occurrence of aberrant phenotypes and the histological subtype. It is concluded that the demonstration of an aberrant phenotype is a valuable supplement to histological assessment in the diagnosis of peripheral T cell lymphomas. It is recommended that the panel of monoclonal antibodies against T cell differentiation antigens should be fairly large, as apparently any antigen may be lost in the process of malignant transformation.     

Epstein-Barr virus in angioimmunoblastic T-cell lymphomas

Epstein-Barr virus (EBV) has been proposed as a possible infective agent involved in the pathogenesis of angioimmunoblastic lymphadenopathy (AIL), a progressive and often fatal lymphoproliferative disorder. We have studied 19 cases of AIL-like lymphomas for the presence of EBV using a sensitive in situ hybridization technique based on the detection of Epstein-Barr encoded RNAs with digoxigenin-labelled oligonucleotide probes. EBV was found in 11 cases; in seven of these EBV was detected in occasional cells. Immunocytochemical studies to investigate viral gene expression, revealed the presence of EBV-encoded latent membrane protein only in those cases which had appreciable numbers of positive cells by in situ hybridization. The intensity of staining varied from case to case and the overall proportion of cells staining for latent membrane protein in a given case was considerably less than that by in situ hybridization. In situ hybridization for cytomegalovirus and human herpes virus type-6 was negative in all cases. We discuss these findings in the light of the proposed role of EBV in the pathogenesis of AIL and conclude that the presence of EBV is a consequence of the disease rather than the cause.     

The blood microvasculature in T-cell lymphomas

Summary The microvasculature of lymph nodes of 55 cases of T-cell lymphoma was studied by light microscopy, immunohistochemistry and electron microscopy. A modified peroxidase-antiperoxidase (PAP) method was used for staining paraffin sections with lectin I of Ulex europaeus (UEA-I), which is a specific marker for vascular endothelial cells. The T-cell nature of each case was proven by immunohistochemistry, including immunoperoxidase staining of frozen sections with monoclonal T-cell antibodies. The cases were subclassified according to previously established criteria, but with the addition of a separate group showing a high content of clear cells. For the purpose of the present study, the small blood vessels were separated into two main variants, viz.: high endothelial venules (HEV) and all other types of vessels with flat endothelium (SVFE). The development of each of these variants and the extent of lymphocyte migration through the vascular wall were assessed semiquantitatively.The findings suggest that the blood microvasculature, as a whole, is similar in all types of T-cell lymphoma. There were distinct differences, however, in the development of the two main categories of small vessels between the various types. Chronic lymphocytic leukaemia of T-type (T-CLL) and Sézary's syndrome were poor in SVFE and rich in HEV, and there was considerable lymphocyte traffic through the latter. In contrast, T-immunoblastic and especially T-lymphoblastic lymphomas showed numerous SVFE, only a few or no HEV and minimal lymphocyte traffic. The appearance of the microvasculature varied markedly in the various subtypes of pleomorphic T-cell lymphoma. In the small cell subtype HEV predominated and SVFE represented only a small or moderate fraction of the microvasculature. As the size of the neoplastic lymphoid cells increased towards the medium-sized and large cell     

Aberrant recombination and the origin of Klinefelter syndrome

Trisomy is the most commonly identified chromosome abnormality in humans, occurring in at least 4% of all clinically recognized pregnancies; it is the leading known cause of pregnancy loss and of mental retardation. Over the past decade, molecular studies have demonstrated that most human trisomies originate from errors at maternal meiosis I. However, Klinefelter syndrome is a notable exception, as nearly one-half of all cases derive from paternal non-disjunction. In this review, the data on the origin of sex chromosome trisomies are summarized, focusing on the 47,XXY condition. Additionally, the results of recent genetic mapping studies are reviewed that have led to the identification of the first molecular correlate of autosomal and sex chromosome non-disjunction; i.e. altered levels and positioning of meiotic recombinational events.     

鼻NK/T细胞淋巴瘤瘤细胞分化状态的探讨

目的 探讨鼻自然杀伤(NK)／T细胞淋巴瘤肿瘤细胞的分化状态。方法 收集已确诊的88例鼻NK／T细胞淋巴瘤的临床资料。免疫组织化学染色选用的抗体有：T细胞分化抗原(CD3ε、CD5、CD1a)、NK细胞相关抗原(CD56、CD57),还有CD34和CD38抗原。结果(1)病变部位以鼻腔最常见,其次为咽部,主要体征为溃疡或黏膜糜烂。本组88例鼻NK／T细胞淋巴瘤中有62例(70．5％)之瘤细胞呈弥漫性增生和浸润,瘤细胞中等大小者有71例(80．7％);(2)88例NK／T细胞淋巴瘤中,T细胞标记CD3ε阳性者78例(88．6％),CD5阳性者56例(63．6％),NK细胞标记CD56阳性者25例(28．4％);CD57、CD1a、CD34和CD38均为阴性。结论 鼻NK／T细胞淋巴瘤瘤细胞的分化可能已超越前体细胞阶段,但尚未达到成熟T淋巴细胞或NK细胞阶段。     

Peripheral T-cell lymphomas of the intestine.

Twenty-seven cases of primary peripheral T-cell lymphomas of the intestine (PTLI) were investigated. Seven patients had histories of malabsorption. The most frequent symptoms at presentation were weight loss, abdominal pain, and acute abdomen. The jejunum was the most common site of lymphoma and multifocal disease was found in 72% of the cases. Twenty-two patients (92%) presented with localized disease confined to the intestine and abdominal lymph nodes, only two patients had generalized disease. According to the pattern of lymphoma infiltration and the morphology of the uninvolved small intestinal mucosa, 21 cases were separated histologically into three categories; 1) enteropathy-associated T-cell lymphoma (EATCL, n = 9) showing predominant intramucosal lymphoma spread and villous atrophy of uninvolved mucosa with high density of intraepithelial lymphocytes (IEL), 2) EATCL-like lymphoma without enteropathy (EATCL-LLWE, n = 5) but with an infiltration pattern similar to EATCL, and 3) T-cell lymphoma without features of EATCL (Non-EATCL, n = 7). Distinctive features of EATCL were the high incidence of malabsorption states, multifocal intestinal disease in all cases, and the high frequency of intestinal recurrences. On frozen sections four of eight PTLI showed the phenotype CD3+ CD4- CD8- HML-1+, which is also expressed on a small subset of normal IEL. The morphologic and immunomorphologic findings suggest that the majority of PTLI is derived from mucosal T lymphocytes. This derivation may be responsible for certain biologic features, such as the preferential spread to and relapse of PTLI at small intestinal sites.     

Comparative microcytometric analysis of European peripheral T-cell malignant lymphomas (EPTL) and adult T-cell leukemia/lymphomas (ATLL) in Japan

A simultaneous microcytometric analysis of nuclear DNA content and size was performed in 8 European peripheral T cell lymphomas (EPTL) and 8 adult T-cell leukemia/lymphomas (ATLL), comparing their patterns on the nuclear density scattergram (NDS) of DNA content versus nuclear size. The intermingling lymphocytes with or without irregular-shaped nuclei in EPTL and ATLL were interpreted as stimulated reactive. Medium-sized cell-dominated T-zone lymphomas and ATLL pleomorphic medium-sized cell type showed two distribution patterns in the diploid range. The first was oblique zonal cluster (OZC) with the second high concentration in the middle part of it, suggesting mixed proliferation with stimulated reactive lymphocytes, and the second was the high concentration in the lower region of the NDS with some cells corresponding to the growth fraction. In large cell-dominated T-zone lymphomas and ATLL pleomorphic large cell type, the third decreasing pattern in the cell density from the lower part of the OZC to its upper part was found upto the hypertetraploid range. The wide distribution on DNS and giant cells with aneuploid high DNA content were found only in the pleomorphic large cell type and the pleomorphic medium-sized and large cell type of ATLL. The T-immunoblastic type of EPTL showed the third pattern and the pleomorphic large cell type of EPTL, characteristic in the clear cytoplasm, showed the second pattern.     

Experimental evidence that RNA recombination occurs in the Japanese encephalitis virus

Due to the lack of a proofreading function and error-repairing ability of genomic RNA, accumulated mutations are known to be a force driving viral evolution in the genus Flavivirus, including the Japanese encephalitis (JE) virus. Based on sequencing data, RNA recombination was recently postulated to be another factor associated with genomic variations in these viruses. We herein provide experimental evidence to demonstrate the occurrence of RNA recombination in the JE virus using two local pure clones (T1P1-S1 and CJN-S1) respectively derived from the local strains, T1P1 and CJN. Based on results from a restriction fragment length polymorphism (RFLP) assay on the C/preM junction comprising a fragment of 868 nucleotides (nt 10-877), the recombinant progeny virus was primarily formed in BHK-21 cells that had been co-infected with the two clones used in this study. Nine of 20 recombinant forms of the JE virus had a crossover in the nt 123-323 region. Sequencing data derived from these recombinants revealed that no nucleotide deletion or insertion occurred in this region favoring crossovers, indicating that precisely, not aberrantly, homologous recombination was involved. With site-directed mutagenesis, three stem-loop secondary structures were destabilized and re-stabilized in sequence, leading to changes in the frequency of recombination. This suggests that the conformation, not the free energy, of the secondary structure is important in modulating RNA recombination of the virus. It was concluded that because RNA recombination generates genetic diversity in the JE virus, this must be considered particularly in studies of viral evolution, epidemiology, and possible vaccine safety.     

Primary natural killer/T-cell lymphomas of the oral cavity are aggressive neoplasms

Thirty-four cases of primary non-Hodgkin’s lymphoma of the oral cavity were investigated for their clinical findings, histopathological features, immunophenotypes and association with Epstein-Barr virus (EBV). Four cases (12%) were natural killer/T-cell lymphomas, 3 (9%) were T-cell lymphomas and 27 (79%) were B-cell lymphomas. Compared with T- and B-cell lymphomas, NK/T-cell lymphomas had a male predominance (M:F 4:0), and most presented as ulceration of the palate and/or maxillary gingiva. Histologically, the lesions showed diffuse infiltration of medium-sized or large lymphoid tumour cells. Angiocentricity and/or angioinvasion were found in all 4 cases. The immunophenotypes of the NK/T-cell lymphomas were CD3+, CD43+, CD45RO+, CD56+ and TIA-1+. EBV was detected in 2 NK/T-cell lymphomas by in situ hybridization (ISH) and polymerase chain reaction (PCR) methods, and was not detected in T- and B-cell lymphomas. The survival rate of patients with NK/T-cell lymphoma was zero, but the survival rates for patients with T-cell and B-cell lymphomas were 67% and 38%, respectively. It appears that NK/T-cell lymphomas of the oral cavity have a predilection for originating in the palate and maxillary gingiva and are aggressive neoplasms. EBV positivity might be associated with more aggressive behaviour. Received: 21 January 1999 / Accepted: 14 April 1999