Metagenomics analysis of the morphological aspects and bacterial composition of broiler feces.

In this descriptive study, we used metagenomics to analyze the relationship between the morphological aspects of chicken feces and its respective bacterial compositions. The microbiota composition was determined by sequencing the V4 region of the 16S rRNA genes collected from fresh broiler feces at 19 d old. In total, 48 samples were collected and divided into 8 groups of 6 samples each. The morphological changes studied were feed passage (FP) and reddish mucus (RM). Each was classified into 3 levels of intensity: 1 (slight), 2 (moderate), or 3 (intense). Thus, the 8 groups studied were feed passage (FP-1; FP-2; FP-3), reddish mucus (RM-1; RM-2; RM-3), normal ileal feces (NIF), and cecal discharge (CD). The alpha diversity (Shannon's index) revealed that the CD group showed greater diversity, and was significantly different from FP-2, FP-3, and RM-1. The beta diversity showed that the CD group samples were more homogeneous than the ileal feces groups. The relative abundance analysis revealed that Firmicutes and Proteobacteria were the most abundant phyla in the ileal feces groups. In CD, Firmicutes and Bacteroidetes were the most abundant. The relative abundance at the genus level revealed 136 different bacterial genera. In the ileal feces groups, the two most abundant genera were Lactobacillus and Escherichia/Shigella, except in the FP-1 and RM-2 groups, which had the opposite order. Unlike the others, the CD group had a higher abundance of Bacteroides and Faecalibacterium. When comparing the NIF group with the others, significant changes were found in the fecal microbiota, with nine genera for the FP groups, 19 for the RM groups, and 61 when compared to CD. The results of the present study suggest that evaluation of fecal morphology is a fundamental task that makes it possible to act quickly and assertively, as the morphological aspects of the feces may be related to the composition and structure of fecal microbiota.

3,3-Dimethyl-cis-9a,13a-diphenyl-2,3,9a,11,12,13a-hexa-hydro-1H-benzo[h][1,4]dioxino[2',3':5,6][1,4]dioxino[2,3-f]chromene.

In the title di-hydro-dioxin, C31H28O5, the dioxane ring has a chair conformation, whereas each of the pyran and dioxine rings has an envelope conformation with methyl-ene and quaternary C atoms, respectively, being the flap atoms. The phenyl rings are cis and form a dihedral angle of 82.11 (10)°. The molecular structure is stabilized by C-H⋯O contacts. In the crystal packing, supra-molecular layers parallel to (101) are sustained by C-H⋯π inter-actions.

Autoantibodies against inosine-5'-monophosphate dehydrogenase 2--characteristics and prevalence in patients with HCV-infection.

BACKGROUND: Indirect immunofluorescence (IIFT) on in house HEp-2 cell preparations revealed a novel antibody giving a granular cytoplasmic pattern not described before, which on two commercial cell preparations revealed a "rings and rods" pattern. This pattern was also observed in four HCV-RNA carriers and prompted the identification of the reactive antigen and the evaluation of the antibody prevalence in HCV-RNA carriers and control groups. METHODS: The antigen's molecular weight was determined by radioimmunoprecipitation of 35S-methionine labeled cell proteins. Expression library screening and sequencing was performed by standard techniques using an oligo(dT)-primed human HeLa cell cDNA expression library. Antibodies against the novel antigen Inositol-5'-monophosphatdehydrogenase 2 (IMPDH2) were analyzed by IIFT, western blot, line blot, and radioimmunoprecipitation assay (RIPA). IIFT was performed on commercial HEp-2 cells and cells cultivated in house for 24 - 60 hours, with or without the IMPDH2 inhibitors mycophenolic acid (MPA) or ribavirin, and subjected to various fixation conditions. Western and line blots were performed with IMPDH2 synthesized in E. coli, RIPA with 35S-methionine-IMPDH2 from in vitro transcription/translation products. Sera screened were positive for HCV-RNA (108), HBV-DNA (100), anti-mitochondrial (31), anti-actin (42), and anti-nuclear antibodies (51) and negative for HCV-RNA (100) and blood donors (100). RESULTS: IMPDH2 is capable of considerable intracellular rearrangements (upon action of inhibitors like MPA and ribavirin), which explains the contrasting immunofluorescence patterns in cells from different sources. By RIPA, proven to be the sole assay suitable for screening of anti-IMPDH2 in human sera, autoantibodies were found in 35.2% of HCV-RNA carriers and in low concentrations in 31% of anti-actin positive patients suspicious of autoimmune hepatitis. Antibodies reacted preferentially with conformational epitopes. Compared to the low concentration of anti-IMPDH2 found in other disease groups, high antibody concentrations were observed in HCV-RNA carriers. CONCLUSIONS: The common occurrence of anti-IMPDH2 in HCV-RNA carriers may be related to ribavirin therapy, causing intracellular aggregation of IMPDH2 thereby altering its immunogenicity. In this study the "rods and rings" immunofluorescence pattern observed could be ascribed to anti-IMPDH2. Anti-IMPDH2 may cause difficulties in interpretation of immunofluorescence patterns in routine autoantibody testing.

Neuromyelitis optica and NMO-IgG in European pediatric patients.

OBJECTIVE: Neuromyelitis optica (NMO) is currently considered a severe relapsing CNS demyelinating disorder that is associated with aquaporin-4 immunoglobulin G (NMO-IgG) while in earlier reports of NMO in childhood it has been described as a benign and monophasic disorder. This study was performed to analyze the prevalence and the clinical course of NMO in a European pediatric cohort of patients with demyelinating CNS disorders. METHODS: A cohort study was performed evaluating 118 pediatric patients presenting at the Center for Multiple Sclerosis in Childhood and Adolescents, Göttingen, Germany, with demyelinating CNS disorders between 2000 and 2009. In all patients, NMO-IgG status was determined. RESULTS: The majority of patients (94%) were diagnosed with remitting recurrent multiple sclerosis. Six patients fulfilled the clinical criteria for NMO but only 1 was seropositive for NMO-IgG. This patient had a severe relapsing course in contrast to the seronegative patients who showed a mild and in the majority of cases monophasic course. CONCLUSIONS: The diagnostic criteria clearly distinguished the patients with NMO from patients with other demyelinating CNS disorders. In the European pediatric population, NMO is very rare and in the majority of patients not associated with NMO-IgG. These seronegative cases have a benign and predominantly monophasic course and therefore do not need the immunosuppressant therapy that is recommended for NMO in the recent literature.

[A new genus of parasitic ciliata from tropical fishes]. / Novyi rod paraziticheskikh resnichnykh iz tropicheskikh ryb.

Ichthyophthirius schlotfeldti Yunchis, 1997 (Ophryoglenidae) was described by the junior author from aquarium fishes from South-East Asia. In the present paper this species is separated into a new genus Neoichthyophthirius gen. n. The reproduction of this species takes place within the epithelium of fishes and not in water as in Ichthyophthirius multifilius. Neoichthyophthirius schlotfeldti has a ring-shaped macronucleus with ends overlapping. Maximum temperature, when the reproduction is possible, is about +34 degrees. The parasite is pathogenous and causes a mortality of aquarium fishes.

Ureteric and pulmonary endometriosis.

Pulmonary lesions and ureter involvement are rare complications of endometriosis. We describe the first case with an involvement of both sites in this condition. The radiographs showed "pulmonary metastases" together with clinical findings of a hydronephrosis, mimicing a malignant metastatic gynecologic tumor of unknown primary. A laparoscopy showed severe endometriotic lesions; therefore the findings were most likely related to the condition of endometriosis. The patient was treated with gosereline acetate for six months (Zoladex). Control radiographs showed complete regression of the pulmonary lesions and the patient is symptom free at 36 months after diagnosis.

Limited ovarian stimulation (LOS), prevents the recurrence of severe forms of ovarian hyperstimulation syndrome in polycystic ovarian disease.

OBJECTIVES: We aimed to test the possibility that LOS could be used to avoid the risk of occurrence of severe OHSS in PCOD patients with past history of severe OHSS in their previous IVF treatment. STUDY DESIGN: A prospective study, with patients stood as their own controls. Twenty patients with history of severe OHSS in their previous IVF treatment were included in the study. The patients received LOS cycles during the six months period commencing January 1998, a full dose of human chorionic gonadotropin (10,000 IU) was administered when the leading follicle reached a mean diameter of 12 mm, and oocytes retrieval 36 h later was followed by intracytoplasmic sperm injection (ICSI) and embryo transfer. Signs and symptoms of severe OHSS were recorded. RESULTS: All patients produced mature oocytes, achieved fertilisation and eight clinical pregnancies were diagnosed. None of the patients re-experienced the symptoms of severe OHSS, and none required hospitalisation. CONCLUSION: We confirm that, in the studied group of patients, LOS helped in the prevention of the severe forms of OHSS in PCOD patients with past history of severe OHSS.

[Freshwater bryozoans (Bryozoa: Phylactolaemata) as vectors of salmonid disease]. / Presnovodnye mshanki (Bryozoa: Phylactolaemata)--perenoschiki bolezni lososevykh.

Relations of PKX [vector of proliferative kidney disease (PKD)] of reared and wild Salmonoidea and Tetraspora bryozoides, the only genus and species of Myxozoa found in fresh water Bryozoa, is discussed using recent publications. Both organisms were found in several European countries as well as in North America. It seems that PKX is a Bryozoa parasite which do not undergo full cycle of sporulation in fish.

Chemosensitivity of B cell chronic lymphocytic leukemia and correlated expression of proteins regulating apoptosis, cell cycle and DNA repair.

B cell chronic lymphocytic leukemia (B-CLL) cannot be cured with conventional chemotherapy. This clinical enigma appears to be at least partially due to the fact that B-CLL cells are resistant to programmed cell death (apoptosis) and that they are arrested in G0/G1 phase of the cell cycle. The reasons for the dysregulation of these two key cellular events in B-CLL are unclear. The present study aimed at determining correlations between the expression levels of proteins regulating apoptosis, cell cycle and DNA repair in B-CLL cells and normal B cells. In addition, the differential sensitivity of B-CLL cells to drug-induced apoptosis was quantified. We show that in B-CLL cells levels of the death-suppressor Bcl-2 correlated positively with those of the pro-apoptotic protein Bax and of the cyclin-dependent kinase (cdk) inhibitor p27Kip1. In B-CLL cells levels of the anti-apoptotic Bcl-xL showed a positive correlation with levels of the 80 kDa regulatory component (Ku80) of the DNA-dependent protein kinase that is involved in DNA double-stranded break repair. These correlations were not detected in normal B cells. The sensitivity of leukemic cells to FLUD but not to ADM, CPM or to DEX was reduced in pre-treated patients. These data support the hypothesis that in B-CLL cells death-modulators and molecules modulating cell cycle and DNA repair are regulated in a coordinated manner. Leukemia (2000) 14, 40-46.

Heterotopic pregnancy in a spontaneous cycle: do not forget about it!

Heterotopic pregnancies are estimated to be less frequent than 1:30000 if no assisted reproduction technologies (ART) are performed. After ART this entity is more frequent and in the range of 1:100. In the case reported here an ectopic pregnancy was detected in the right fallopian tube at 7+ 1 weeks of gestation. It was misdiagnosed as an ectopic singleton, and treated by laparoscopic salpingectomy, because of a previous ectopic in the same tube. Rising hCG after laparoscopy during the subsequent days followed by ultrasound evaluation revealed a viable intrauterine pregnancy. The pregnancy continued uneventfully and a healthy child was delivered at term. The problems, which lead to the misdiagnosis are discussed. The problem of rare cases in medicine, and the problems of a 'modern' medicine are discussed.

Fertilization potential of spermatozoa with abnormal morphology.

One of the best discriminators for the fertilization potential of human spermatozoa is sperm morphology. The problem in the assessment of the sperm morphological characteristics is their pleiomorphism. Examination of spermatozoa with the light microscope can provide only limited information on their internal structure. More detailed examination of sperm structure using electron microscopy can reveal major, often unsuspected ultrastructural abnormalities. Results and cut-off values for sperm analysis depend on the criteria for normal morphology. World Health Organization recommendations provide a classification suitable for clinical practice. Clinically reliable cut-off limits for normal sperm morphology according to strict Tygerberg criteria were suggested to be 4% in in-vitro fertilization procedures. Patients with severe sperm head abnormalities have a lower chance of establishing successful pregnancies, even though fertilization may be achieved. The outcome of intracytoplasmic sperm injection is not related to any of the standard semen parameters or to sperm morphology. Sperm decondensation defects and DNA anomalies may be underlying factors for the unrecognized derangements of the fertilizing capacity of spermatozoa, regardless of sperm morphology. Centrosome dysfunction may also represent a class of sperm defects that cannot be overcome simply by the insertion of a spermatozoon into the ooplasm. In this article an overview on the composition and ultrastructure of spermatozoa is presented, while emphasizing sperm ultrastructural and sperm DNA anomalies and their effects on fertilization.

Intracytoplasmic injection of round and elongated spermatids from azoospermic patients: results and review.

Microinjection is established as the method of choice in the treatment of severe male factor infertility as well as in azoospermic patients. Recent studies have shown that fertilization and cleavage can be achieved by injection of ejaculated as well as testicular elongated spermatids into oocytes. Here we report on the two first pregnancies worldwide resulting from elongated spermatid injection from frozen-thawed testicular tissue. Four patients with complete Sertoli cell-only syndrome (SCOS) and two with spermatogenetic maturation arrest were included in our microinjection programme. Tissues from open testicular biopsies were cryopreserved until the time of follicle puncture. A total of 67 oocytes were harvested. In the two patients with maturation arrest, cryopreserved elongated spermatids were successfully injected, while in two of the other four SCOS patients only cryopreserved round spermatids were available to be injected into the oocytes. Out of 18 injected oocytes, 10 were fertilized in the first group, while nine out of 49 injected oocytes showed fertilization and cleavage in the second group. Two clinical pregnancies were achieved with elongated spermatids from frozen-thawed testicular tissue, while no pregnancy was established in the case of round spermatids. This study confirms that fertilization, cleavage and pregnancy can be successfully achieved in cases with spermatogenetic maturation arrest by injecting cryopreserved elongated spermatids into oocytes. The literature on pregnancies following spermatid injection, as well as the problems using this technique and possible risks, are discussed.

Transcervical falloscopic dilatation of proximal tubal occlusion. Is there an indication?

Proximal tubal occlusion (PTO), until recently a domain of microsurgery, can also be treated by a transcervical balloon dilatation and/or tubal recanalization. The aim of our study was to evaluate the possibility of transcervical tubal dilatation during transcervical Falloposcopy. Transcervical Falloposcopy and tubal dilatation was performed under laparoscopic control. During a period of 48 months a total of 157 Falloposcopies was performed. Out of a total of 157 patients, 42 patients had PTO confirmed by dye-pertubation during laparoscopy. All patients were referred because of primary or secondary tubal infertility. A total of 18 patients had bilateral PTO by dye-pertubation and of these six patients had successful bilateral and seven patients successful unilateral recanalization. The remaining five patients were unable to recanalize. A total of 24 patients had an unilateral PTO by dye-pertubation, 13 of these patients had a contralateral diseased tube, seven of which could be recanalized. Four patients had contralateral normal tubes, with successful recanalization in one patient. Seven patients had an occluded or missing contralateral tube, five of which could be recanalized. A total of 60 tubes with PTO were diagnosed, of which 32 (53.3%) tubes could be recanalized. 20 of these had normal tubes. Only patients with healthy Fallopian tubes carried pregnancies to term (five pregnancies, 12% of all patients). All patients conceived within a period of 3-6 months. We observed no ectopic pregnancy.

A simple, inexpensive and effective artificial cycle with exogenous transdermal oestradiol and vaginal progesterone for the transfer of cryopreserved pronucleated human oocytes in women with normal cycles.

Supernumerary pronucleated stage oocytes (PN) are usually cryopreserved. PN are transferred in spontaneous, stimulated or artificial cycles. In this study, an artificial cycle with a transdermal therapeutic system was used for oestradiol release (Estraderm TTS 100) in combination with a targeted drug delivery system for vaginal progesterone release (Crinone 8%). Patients started transdermal 17beta-oestradiol treatment on cycle day 1. Only one clinical monitoring was necessary on day 14 for confirmation of satisfactory endometrial development and exclusion of ovulation by transvaginal ultrasound and endocrine determinations (oestradiol, progesterone and luteinizing hormone). Embryo transfer was performed on the third day of progesterone treatment (day 17). The first 25 cycles were recently completed in a prospective study; no cycles were cancelled due to ovulation or unsatisfactory endometrial development. In comparison with the previous protocol of embryo transfer in stimulated cycles in our clinic which required extensive ultrasound and endocrine monitoring, the pregnancy rate in these oestrogen- and progesterone-supplemented cycles was nearly twice as high (34.8%). Two pregnancies were even achieved with zygotes after micro-injection of frozen-thawed late spermatids extracted from testicular tissue (cryo-TESE). In these cycles, the Estraderm TTS 100/Crinone 8% protocol seems to be superior to stimulation protocols and even to other protocols reported so far for artificial cycles with exogenous oestradiol and progesterone treatment.

Prediction of severe ovarian hyperstimulation syndrome by free serum vascular endothelial growth factor concentration on the day of human chorionic gonadotrophin administration.

In this prospective study the concentration of circulating vascular endothelial growth factor (VEGF) was followed in 10 patients with severe ovarian hyperstimulation syndrome (OHSS) after ovarian stimulation and in 15 patients without OHSS. VEGF was assayed by means of two different commercially available kits as either free or total VEGF in serum. The concentration of free VEGF was significantly higher on the days of human chorionic gonadotrophin (HCG) administration (309.4 +/- 165.0 versus 190.3 +/- 127.8 pg/ml, P < 0.05) and embryo transfer (315.0 +/- 125.2 versus 209.3 +/- 137. 2 pg/ml, P < 0.05) in the OHSS compared to the control group. No such difference existed with respect to total circulating VEGF. In addition, there was no significant rise in the free or in the total serum VEGF concentration in the OHSS patients or the controls from the day of HCG administration up to the days of oocyte retrieval or embryo transfer. A cut-off concentration of 200 pg/ml free serum VEGF concentration on the day of HCG treatment resulted in a sensitivity of 90% and a specificity of 80% for the prediction of OHSS development. This is the first report on the parallel measurement of free and total VEGF in serum following ovarian stimulation. The value of the proposed cut-off concentration should be confirmed in a study of a larger group of women.

The forgotten child--a case of heterotopic, intra-abdominal and intrauterine pregnancy carried to term.

Heterotopic pregnancies are estimated to be less frequent than one in 30,000 if no assisted reproduction technologies are performed. Here we report a case which occurred in Tanzania. An abdominal pregnancy at term was first misdiagnosed as an ovarian tumour and diagnosed on the first post-partum day of the intrauterine fetus, which was delivered spontaneously. The abdominal pregnancy was then treated by laparotomy and removal of the placenta. The fetus was alive and healthy. The follow-up of the twins was normal.

Ovarian abscess and heterotopic triplet pregnancy: two complications after IVF in one patient.

A patient is reported, who suffered from ovarian abscess after ovarian puncture of a functional ovarian cyst. The cyst has developed after administration of a GnRH agonist depot preparation in the preceeding luteal phase. She was planned to be stimulated for IVF according to the long luteal protocol. The abscess was removed by laparoscopy. and stimulation started two months later after administration of two further GnRH against depot preparations. The patient got pregnant after embryo transfer of three embryos. and a heterotopic triplet pregnancy, with intrauterine twins and a tubal singleton was established. Bilateral salpingectomy was performed, because of bilateral haematosalpinx and previously described bilateral tubal occlusion. The further pregnancy was uncomplicated.

Ovarian hyperstimulation syndrome (OHSS) in a spontaneous pregnancy with fetal and placental triploidy: information about the general pathophysiology of OHSS.

An ovarian hyperstimulation syndrome (OHSS) in spontaneous pregnancies is a very rare event. Hence, clinicians might make wrong decisions, such as laparotomy, because of suspicion of an ovarian carcinoma, or severe complications such as renal insufficiency may develop because the diagnosis and treatment of OHSS are delayed. Here we report a case of a woman in the 15th week of gestation, presenting with a partial hydatidiform mole and a triploidy of fetus and placenta, with an ongoing and severe OHSS even after legal induced abortion. A low vascular endothelium growth factor (VEGF) concentration (50 ng/ml) was measured when human chorionic gonadotrophin (HCG) exceeded 1000 IU/l and was followed by VEGF concentrations >900 ng/ml, when the OHSS developed. The literature on spontaneous pregnancies associated with OHSS is reviewed, and possible reasons for the clinical course presented in this study and the reviewed reports are discussed. The findings in this case contribute to our theory that VEGF is a causative factor of OHSS, but has no impact on the course of this disease.

Treatment of uterine fibroids with a slow-release formulation of the gonadotrophin releasing hormone antagonist Cetrorelix.

A depot preparation of the third-generation gonadotrophin-releasing hormone (GnRH) antagonist Cetrorelix (SB-75) was used for preoperative treatment in twenty premenopausal patients with symptomatic uterine fibroids who were to undergo surgery. In a prospective, open, randomized setting 60 mg of Cetrorelix pamoate salt was administered i.m. on cycle day 2. Patients were randomized for a second dose of 30 or 60 mg of Cetrorelix depot, which was administered according to the degree of oestradiol suppression (<50 pg/ml) on treatment day 21 or 28. Surgery was done after 6 or 8 weeks of treatment, depending on second dosage administration. Weekly transvaginal sonography (TVS) and magnetic resonance imaging (MRI) before and after treatment was performed, for fibroid volume assessment. Sixteen patients showed satisfactory suppression of gonadotrophins and sex steroid secretion, avoiding any initial flare-up effect. In these patients a mean shrinkage rate of largest fibroid volume of 33.5% at the end of treatment could be observed according to TVS, while the mean shrinkage rate obtained after 14 days of treatment was 31.3%. In good responders (shrinkage >20%) largest fibroid volume at day 14 was approximately 56.7% of basic assessment. Although MRI showed minor mean shrinkage rates of only 25.4% of the initial volume, these differences in comparison to TVS assessment were not statistically significant. The avoidance of any initial flare-up in gonadotrophin secretion may explain this extremely fast reduction in fibroid size. The advantages of GnRH antagonist treatment in this indication consist in the short treatment time with a fast restoration of the ovarian function. The rate of poor responders may be reduced by using an improved slow release preparation.