Plant-herbivorous beetle networks: molecular characterization of trophic ecology within a threatened steppic environment.

DNA barcoding facilitates many evolutionary and ecological studies, including the examination of the dietary diversity of herbivores. In this study, we present a survey of ecological associations between herbivorous beetles and host plants from seriously threatened European steppic grasslands. We determined host plants for the majority (65%) of steppic leaf beetles (55 species) and weevils (59) known from central Europe using two barcodes (trnL and rbcL) and two sequencing strategies (Sanger for mono/oligophagous species and Illumina for polyphagous taxa). To better understand the ecological associations between steppic beetles and their host plants, we tested the hypothesis that leaf beetles and weevils differ in food selection as a result of their phylogenetic relations (within genera and between families) and interactions with host plants. We found 224 links between the beetles and the plants. Beetles belonging to seven genera feed on the same or related plants. Their preferences were probably inherited from common ancestors and/or resulted from the host plant's chemistry. Beetles from four genera feed on different plants, possibly reducing intrageneric competition and possibly due to an adaptation to different plant chemical defences. We found significant correlations between the numbers of leaf beetle and weevil species feeding on particular plants for polyphagous taxa, but not for nonpolyphagous beetles. Finally, we found that the previous identifications of host plants based on direct observations are generally concordant with host plant barcoding from insect gut. Our results expand basic knowledge about the trophic relations of steppic beetles and plants and are immediately useful for conservation purposes.

Frequency of clonal intraepithelial T lymphocyte proliferations in enteropathy-type intestinal T cell lymphoma, coeliac disease, and refractory sprue.

BACKGROUND: Clonal T cell receptor (TCR) gene rearrangements and loss of T cell antigens such as CD8 and TCR-beta in intraepithelial lymphocytes (IELs) may indicate the development of an enteropathy-type intestinal T cell lymphoma (EITCL) in patients with refractory sprue. AIMS: To define the diagnostic value of these markers in duodenal biopsies from patients with villous atrophy as a result of various underlying disorders. PATIENTS AND METHODS: Duodenal biopsies from eight patients with coeliac disease and five patients with villous atrophy caused by defined disorders were compared with three patients with refractory sprue evolving into overt EITCL, two patients with ulcerative jejunitis, and with eight patients with overt EITCL, for expression of CD3, CD4, CD8, and TCR-beta in IELs using immunohistochemistry and for clonal TCR-gamma gene rearrangements using polymerase chain reaction. In addition, biopsies from six consecutive patients with refractory sprue of uncertain cause were examined. RESULTS: Clonal TCR-gamma gene rearrangements were found in all resected tumours of patients with EITCL, in 3/8 duodenal biopsies of patients with EITCL, in 2/2 patients with ulcerative jejunitis, in 2/3 patients with refractory sprue evolving into overt EITCL, and in 1/6 patients with refractory sprue. No rearrangements were found in biopsies from patients with refractory sprue caused by defined disorders or those with coeliac disease. Clonality in duodenal biopsies was associated with an abnormal phenotype of IELs in all cases and in all but one case in patients with evidence of underlying coeliac disease. Specificity for detection of an EITCL using immunohistology was 77% for CD8 and for TCR-beta staining, and 100% for detection of a clonal TCR-gamma gene rearrangement. Sensitivity was 62% for staining with CD8 and clonality investigation, while sensitivity reached 100% for TCR-beta staining in all investigated patients with EITCL. CONCLUSIONS: Clonal proliferations of phenotypically abnormal IELs in refractory sprue represent an early manifestation of EITCL, for which the term "sprue-like intestinal T cell lymphoma" is proposed. This constellation is also found in duodenal biopsies from patients with an overt EITCL and is not related to other sprue syndromes, resulting in a high specificity for detection of an EITCL or refractory sprue evolving into EITCL. Overt EITCL may develop directly from coeliac disease without a precursor lesion (refractory sprue with clonal IELs) being demonstrable in duodenal biopsies or via a "sprue-like intestinal T cell lymphoma". This latter entity is a complication of coeliac disease.

Gastric lymphomas: aspects of follow-up and after-care.

In recent years, new insights into gastric lymphomas and their etiology and pathogenesis have been gained. The predominant role of infection with Helicobacter pylori in the pathogenesis as a pre-malignant condition of a special lymphoma entity [gastric lymphoma of the mucosa-associated lymphoid tissue (MALT) type], has defined new diagnostic procedures and concepts of treatment. Therefore, prognostic factors (e.g. stage of lymphoma, histopathologic grading, resectability etc.) are extremely important for the intensity and efficacy of follow-up and after-care. Surveillance programs in gastric lymphoma include sequelae of surgical resection, chemotherapy and radiotherapy, and the efficacy of follow-up procedures have to be measured by the prevention of tumor relapse in comparison to the intensity of diagnostic procedures. Since lymphoma relapse may occur both as local or disseminated recurrence in 13%-35% of cases, follow-up procedures have to regard both aspects during surveillance. While these follow-up programs are standardized in epithelial tumors (e.g. colon carcinoma), they are not yet established or comparable for each type of gastric lymphoma. Low-grade MALT lymphomas have to be considered as a new lymphoma entity. In addition, new diagnostic procedures (e.g. molecular parameters such as polymerase chain reaction (PCR) for clonality, endosonography, "gastric mapping") have been found to be important parameters for diagnosis and staging of gastric lymphoma and may therefore be relevant for the course of the disease. The definition of "lymphoma cure" and the impact of these procedures as prognostic factors will have to be discussed and may influence the follow-up of gastric lymphoma.

Efficacy, safety and toleration of fluconazole suppositories in the treatment of oral candidosis.

An open multicentre, non-comparative study was conducted in three countries to investigate the efficacy, safety and tolerance of fluconazole suppositories in the treatment of oropharyngeal candidosis. Patients received fluconazole 100 mg day-1 in the form of suppositories or capsules. Minimum duration of total treatment was 7 days, maximum total treatment duration was 14 days, and median duration of total treatment was 9.5 (7-14) days. After having received suppository-based treatment for at least 5 days, patients could be switched to oral treatment. Eighty-two male and 19 female patients with a mean age of 43 years were enrolled in the study. The median duration of suppository treatment was 8.9 (5-14) days. Patients were evaluated clinically and mycologically at regular intervals during and at the end of treatment. Seventy-nine of 101 patients enrolled in the study were considered efficacy-evaluable. Clinical cure was achieved in 75 of 79 (95%) patients and improvement was seen in four of 79 (5%) at the end of therapy. At follow-up after 1 month, clinical cure was observed in 48 of 63 (76%) patients. The results of this study demonstrates that the fluconazole suppository formulation is effective, safe and well tolerated.

Increased immunoglobulin G production by short term cultured duodenal biopsy samples from HIV infected patients.

BACKGROUND: Secretory immunity is a major defence mechanism against infections at mucosal surfaces which are common in HIV infected patients. AIMS: To analyse intestinal immunoglobulin production in HIV infection in comparison with that in saliva and serum. PATIENTS AND METHODS: Immunoglobulin G (IgG), A (IgA), and M (IgM) concentrations were determined in supernatants of short term cultured duodenal biopsy samples, serum, and saliva from HIV infected patients (n = 28) and controls (n = 14) by radial immunodiffusion. RESULTS: IgG was increased in the supernatants of short term cultured biopsy samples and saliva from HIV infected patients compared with controls (p < 0.01), but IgA and IgM levels were normal. In contrast, both IgG and IgA concentrations in serum were higher in HIV infected patients than in controls (p < 0.002). No correlation was found between IgA produced by duodenal biopsy specimens and serum IgA. CONCLUSION: Abnormalities in mucosal immunoglobulin production in HIV infection were surprisingly small, indicating that specific secretory immunity rather than quantitative immunoglobulin production may be impaired. However, increased production of IgG could contribute to mucosal inflammation by complement activation. Our findings of normal mucosal IgA production and the lack of correlation between serum and mucosal IgA argues against an intestinal origin for the increased serum IgA levels in HIV infected patients.

HIV-1 p24 but not proviral load is increased in the intestinal mucosa compared with the peripheral blood in HIV-infected patients.

OBJECTIVE: To investigate differences in viral and proviral load between the peripheral blood and the intestinal mucosal immune system in HIV-infected patients. DESIGN: HIV-1 p24 and HIV DNA content were compared in blood samples and intestinal biopsies from HIV-infected patients. METHODS: Intestinal biopsies and peripheral blood were simultaneously obtained from 27 HIV-infected patients undergoing diagnostic endoscopy. The p24 concentrations were measured in serum and homogenized intestinal biopsies by enzyme-linked immunosorbent assay after acid-dissociation of immune complexes. Proviral load was determined in blood and intestinal biopsies by a quantitative competitive polymerase chain reaction amplifying the HIV-1 nef gene from genomic DNA. RESULTS: No significant differences were found in proviral load comparing HIV copies per 1.5 x 10(5) cell equivalents in blood [2650 (600-44000)] and intestinal biopsies [4200 (1325-19 625)]. Paired analysis revealed a strong positive correlation between serum and mucosal proviral load. In contrast, HIV core protein p24 was detected in intestinal biopsies from 18 patients in much higher concentrations than in serum [858 (262-4111) pg/g versus 34 (9-242) pg/g; P < 0.005]. The p24 concentrations in serum and intestinal biopsies did not correlate and no significant correlation was observed in serum or intestinal biopsies between proviral load and p24 concentrations. No clear correlations were observed between clinical parameters and HIV DNA or HIV p24 levels in blood or biopsies. CONCLUSIONS: Our findings demonstrate a homogenous distribution of HIV proviral load in the peripheral blood and the intestinal mucosal immune system. The high viral antigen load in the intestine therefore indicates that mucosal HIV production is upregulated at the transcriptional and/or translational level. The intestinal mucosa is a major reservoir for HIV in HIV-infected patients.

[Current therapeutic strategy in primary intestinal non-Hodgkin's lymphoma]. / Aktuelle therapeutische Strategie des primären Intestinalen Non-Hodgkin-Lymphoms.

Of all surgical interventions of intestinal non-Hodgkin's lymphomas 58% (15 or 26 patients) are performed in an emergency situation. In 42% of cases, examination by ultrasonography, endosonography, intestinoscopy. Sellink's enema, thoracic, abdominal/pelvic CT and bone marrow puncture could determine the stage preoperatively. This could also be done by examining the regional and juxtaregional lymph nodes or performing a liver biopsy intraoperatively. Crucial for the therapy is in all cases the adequate staging even in emergency situations. Only special knowledge of the intestinal non-Hodgkin's lymphoma can lead to the necessary stage-adapted multimodal therapy--operation/irradiation/chemotherapy.

Mucosal abnormalities in microsporidiosis.

OBJECTIVE: To determine the prevalence of microsporidiosis in HIV-infected patients with and without diarrhoea and to characterize alterations in mucosal architecture and brush border enzyme activities in patients with microsporidiosis. PATIENTS: A total of 259 HIV-infected patients undergoing oesophago-gastroduodenoscopy because of diarrhoea (n = 123) or other symptoms (n = 136) were studied. METHODS: Patients were evaluated for the presence of microsporidia by electron microscopy of duodenal biopsies. Brush border enzyme activities were measured by histochemistry and mucosal architecture was determined by three-dimensional morphometry in biopsies from patients with microsporidiosis and compared with biopsies from a subgroup of HIV-infected patients with or without other enteropathogens. RESULTS: Enterocytozoon bieneusi was detected in 17 patients and Encephalitozoon intestinalis was detected in two patients. Microsporidiosis was significantly more frequent in patients with chronic diarrhoea (19.1%; P < 0.0001) or in patients with acute diarrhoea (7.2%; P = 0.04) than in patients without diarrhoea (1.5%). Microsporidiosis was associated with lactase deficiency (P = 0.03) and a reduced activity of alkaline phosphatase (P = 0.028) and alpha-glucosidase (P = 0.025) at the basal part of the villus compared with brush border enzymes in patients without enteropathogens. Patients with microsporidia had reduced villus height (P = 0.043) and a villus surface reduced by 40% (P = 0.004) compared with patients with enteropathogens other than microsporidia. CONCLUSIONS: Our study confirms the association between microsporidia and diarrhoea. The pathophysiologic mechanism by which microsporidia cause diarrhoea appears in part to be malabsorption, caused by a reduction of absorptive mucosal surface and impairment of enterocyte function.

Comparative evaluation of modified trichrome and Uvitex 2B stains for detection of low numbers of microsporidial spores in stool specimens.

At present, the laboratory diagnosis of intestinal infections caused by microsporidia depends on the detection of the typical spores either with a modified trichrome stain (MTS) or by staining with fluorochromes. The purpose of the present study was (i) to compare staining with MTS (MTS method) and the staining with the fluorochrome Uvitex 2B (U2B method) with respect to their sensitivities and specificities, particularly in the presence of low numbers of spores, and (ii) to evaluate their reliabilities under routine laboratory conditions. First, 30 negative human stool specimens as well as 30 specimens enriched with a low concentration of microsporidial spores were examined. The U2B and MTS methods detected 27 and 30, of the positive samples, respectively (95% confidence intervals for sensitivity, 0.73 to 0.98 for the U2B method and 0.88 to 1.00 for the MTS method) without yielding false-positive results (95% confidence intervals for specificity, 0.88 to 1.00 for the MTS and U2B methods). In addition, analysis of serial dilutions of 17 stool specimens from AIDS patients containing microsporidia revealed comparable detection thresholds (P = 0.52) for both methods. Finally, 40 slides prepared from one stool specimen containing very few microsporidia and 40 negative slides were included in the routine diagnostic program during 1 month in order to monitor laboratory handling and run-to-run variations. Again, both methods exhibited comparable sensitivities (95% confidence intervals, 0.83 to 0.99 for the MTS method and 0.91 to 1.00 for the U2B method) and specificities (95% confidence intervals, 0.91 to 1.00 for the MTS and U2B methods). In conclusion, MTS and U2B methods are equally useful in the diagnosis of microsporidiosis. However, since detection thresholds for both methods differed considerably in all diluted stool specimens, performance of a combination of both methods may be more sensitive than the performance of only one procedure in the event of very low numbers of microsporidial spores.

Fulminant primary manifestation of Crohn's colitis "Hot Crohn's disease".

Following the very short course of a disease with watery diarrhea, fever, nausea, meteorism and a severe feeling of general illness, a 22-year-old patient was diagnosed as having a toxic megacolon, and a subtotal colectomy was carried out. The postoperative progression was uncomplicated and the patient recovered quickly. The examination of the operation specimen revealed a serious ulcerous colitis with relative omission of the rectum and the distal sigmoid colon. After critical evaluation of the histological findings, it was judged to be a fulminant Crohn's colitis and, for the purposes of differential diagnosis, differentiated from ulcerative colitis and colitis indeterminate. The formal pathogenesis of the inflammatory-ulcerous processes is discussed, in particular with regard to the activation of the macrophages and the very short anamnesis in a clinically established primary manifestation of the disease.

Pathological and epidemiological aspects of skin lesions in histoplasmosis. Observations in an AIDS patient and badgers outside endemic areas of histoplasmosis.

As a consequence of HIV infection, histoplasmosis is increasingly occurring as an opportunistic infection with a systemic course outside histoplasmosis-endemic areas, e.g. in Europe. Accordingly, questions concerning the epidemiology of this mycosis arise. Two incidents involving histoplasmosis in man and badgers with prevailing involvement of the skin encouraged us to review the pathogenesis and epidemiology of this mycosis in Germany, where so far Histoplasma capsulatum has not been endemic. With a view to prevention, attention is drawn to the avoidance of microfoci of H. capsulatum in the newly introduced concept of biowaste and its composting plants in countries with modern waste management.

Malignant gastrointestinal lymphomas in patients with AIDS.

HIV-associated malignant lymphomas are a common complication in late HIV infection, and there is a high percentage of gastrointestinal tract involvement. Non-Hodgkin's lymphoma was found in 108 of 2,750 HIV-positive patients (3.9%) in our institution, whereas gastrointestinal manifestation was diagnosed in 48 of 108 patients (44.4%). 44 of these cases were found during endoscopy of the upper and lower gastrointestinal tract (or by laparotomy or laparoscopy in 4 cases). Endoscopy is a reliable procedure for the diagnosis of lymphoma. Unusual manifestations such as oral, esophageal or perianal lesions and multifocal disease were common findings. Life-threatening complications such as gastrointestinal bleeding, perforation, and obstruction occurred in 37.5%. High-grade B-cell lymphomas were found in all cases including mainly lymphoblastic, immunoblastic, centroblastic and Burkitt subtypes. 52% of the patients had disseminated lymphoma with Ann Arbor stage III or IV. Standard chemotherapy with cyclophosphamide, adriamycin, vincristine and prednisone was started in 25 patients and resulted in a mean survival time of 4.8 months. The prognosis of AIDS patients presenting with malignant gastrointestinal lymphoma depends mainly on the presence or absence of previous AIDS-defining diseases, not CD4 cells, lymphoma-associated gastrointestinal complications or the histopathologic lymphoma type at the time of diagnosis.

Abnormal predominance of IgG in HIV-specific antibodies produced by short-term cultured duodenal biopsy specimens from HIV-infected patients.

The aim of our study was to analyze HIV-specific humoral immunity in the intestinal mucosa at different stages of HIV infection in comparison with serum and saliva. Duodenal biopsy specimens from 30 AIDS patients and 9 HIV-infected patients without AIDS were cultured for 48 hours. Culture supernatants, as well as simultaneously obtained serum and saliva samples, were adjusted to the same immunoglobulin concentrations and tested for HIV-specific IgG and IgA by Western blot. The HIV antigen pattern differed clearly between IgA and IgG but was similar for each isotype independent of its origin (i.e., serum, saliva, or biopsy specimen supernatants). Short-term cultured duodenal biopsy specimens from HIV-infected patients at all stages produced predominantly IgG, which was broadly reactive with HIV antigens. Lower titers of HIV-specific IgA, which recognized few antigens, were found, mostly the glycoprotein gp160. At later stages of the disease compared with earlier stages, the reaction pattern of mucosal IgA from saliva and biopsy supernatants was even more restricted; secretory component was frequently absent. The abnormal predominance of HIV-specific IgG over IgA in mucosal secretions may result from abnormal antibody production in the mucosa rather than from serum leakage. Mucosal inflammation induced by HIV-IgG immune complexes and insufficient immune exclusion by secretory IgA may not only lead to increased mucosal HIV replication but may also contribute to gastrointestinal disease in HIV-infected patients.

Intestinal T-cell lymphoma: a reassessment of cytomorphological and phenotypic features in relation to patterns of small bowel remodelling.

Intestinal T-cell lymphoma (ITCL) is an uncommon entity among primary gastrointestinal lymphomas. In this study we evaluated tumours from 20 patients presenting with (n = 8) or without (n = 12) a history of coeliac disease (CD). Neoplastic lesions were composed of predominantly small (n = 4), small-to-medium (n = 2), medium/mixed-to-large (n = 7) or large and anaplastic (n = 7) cells. Different patterns of tumour growth and remodelling of the small bowel wall were observed. Pattern a (n = 4) was characterized by an intramucosal spread of small tumour cells with a small growth fraction. This pattern resembles mucosal inflammation in CD. In pattern b (n = 2), ulcerated solitary or multiple tumours composed of small to medium-sized cells were observed. The adjacent or distant mucosa showed a nearly normal architecture. In pattern c (n = 7), ulcerated lesions were composed of medium-sized to large cells. Mucosal flattening occurred in all segments infiltrated by lymphoma. In pattern d (n = 7), bowel remodelling was observed along the small intestine even at sites not affected by lymphoma. The main neoplastic lesions were composed of pleomorphic large or anaplastic cells frequently expressing the CD30 molecule. Intramucosal spread of a small epitheliotropic T-cell population was observed in the vicinity or even at distant segments of the small bowel. The demonstration of clonal rearrangements of T-cell receptor genes helped to trace widespread occurrence of this small intraepithelial neoplastic component. We suggest that different features of tumour cells such as the expression of activation antigens may contribute to the remodelling of small bowel mucosa. The addition of immunophenotyping data to macroscopic and microscopic features of specimens provided evidence that this uncommon lymphoma exhibits a spectrum in cytological composition and growth patterns. However, despite the considerable heterogeneity of the cases analysed, most of them shared a characteristic immunohistochemical profile (CD3+, CD8+/-, CD103+), further substantiating the view that ITCL is the neoplastic equivalent of an intraepithelial T-cell subset of the small intestine. This phenotype and the intraepithelial accumulation of lymphoma cells observed in the surviving mucosa are clues to the diagnosis of this clinicopathological lymphoma entity characterized by a broad range of morphological expressions.

Stool viruses, coinfections, and diarrhea in HIV-infected patients. Berlin Diarrhea/Wasting Syndrome Study Group.

To examine the prevalence of stool viruses and their role in the pathogenesis of diarrhea in HIV infection, we evaluated biopsies and repeated stool samples of 256 HIV-infected patients undergoing diagnostic endoscopy because of diarrhea (n = 136) or other symptoms (n = 120) for bacterial, protozoal, and viral enteropathogens. In 70% of the patients with diarrhea, at least one potential enteropathogen was detected. Stool virus was detected by electron microscopy in 17% (44 of 256), adenovirus in 6.6% (17 of 256), and coronavirus in 11.3% (29 of 256) of the patients. Adenovirus and coronavirus were detected more frequently in patients with diarrhea than in patients without diarrhea [adenovirus 10% (13 of 136) vs. 3.3% (4 of 120), p = 0.0129; coronavirus 15% (21 of 136) vs. 6.6% (8 of 120), p = 0.0142]. Sixty-one percent of patients harboring stool virus were coinfected by another enteropathogen. Pathogens other than stool virus were detected more frequently in patients harboring adenovirus (82%) than in patients without stool virus (48%, p < 0.025). Adenovirus and coronavirus are frequently detected in stools of HIV- infected patients and may contribute to diarrhea. Adenovirus infection may facilitate the occurrence of other intestinal pathogens. Due to frequent coinfections, detection of stool viruses reduces the rate of diarrhea of unknown origin only by approximately 5%.

Foscarnet 5 versus 7 days a week treatment for severe gastrointestinal CMV disease in HIV-infected patients.

In a randomized open trial foscarnet 90 mg/kg b.i.d. 5 days for 3 weeks was compared to 90 mg/kg b.i.d. daily in severe gastrointestinal cytomegalovirus disease in HIV-infected patients. Thirty-eight patients were randomized, 36 were evaluable (all male, age 24-54 years, median 40 years; CD4/microliter 0-150, median 10). Treatment efficacy was evaluated based on a score consisting of symptoms, endoscopic and histologic examination. In the 5-day treatment group 10/16 (62%) patients responded to treatment, in the 7-day treatment group 13/20 (65%), with symptoms resolving in most patients after 1 week. Side effects and adverse events were seen in 13 patients in the 5-day treatment group and in 15 patients in the 7-day treatment group. Laboratory abnormalities were common in both groups, in one patient reversible renal insufficiency developed. Efficacy and safety of treatment 5 days a week was comparable to the standard regimen.

High-level sensitivity of quantitative pp65 cytomegalovirus (CMV) antigenemia assay for diagnosis of CMV disease in AIDS patients and follow-up.

Cytomegalovirus (CMV) antigenemia was evaluated in 174 patients positive for human immunodeficiency virus. Antigenemia could be detected in 96.7% of patients with CMV disease, 76.9% of patients suffering from a relapse of the disease, and 11.4% of asymptomatic patients with CD4 levels of < 100 cells per microliter. No antigenemia was detected in patients with CD4 levels of 250 to 500 cells per microliter. Specificity and the positive predictive value for CMV disease were increased only if more than 5 positive cells per slide were considered. However, CMV disease may also occur in patients with low-grade antigenemia.