Pond sediment magnetite grains show a distinctive microbial community.

Formation of magnetite in anaerobic sediments is thought to be enhanced by the activities of iron-reducing bacteria. Geobacter has been implicated as playing a major role, as in culture its cells are often associated with extracellular magnetite grains. We studied the bacterial community associated with magnetite grains in sediment of a freshwater pond in South Korea. Magnetite was isolated from the sediment using a magnet. The magnetite-depleted fraction of sediment was also taken for comparison. DNA was extracted from each set of samples, followed by PCR for 16S bacterial ribosomal RNA (rRNA) gene and HiSeq sequencing. The bacterial communities of the magnetite-enriched and magnetite-depleted fractions were significantly different. The enrichment of three abundant operational taxonomic units (OTUs) suggests that they may either be dependent upon the magnetite grain environment or may be playing a role in magnetite formation. The most abundant OTU in magnetite-enriched fractions was Geobacter, bolstering the case that this genus is important in magnetite formation in natural systems. Other major OTUs strongly associated with the magnetite-enriched fraction, rather than the magnetite-depleted fraction, include a Sulfuricella and a novel member of the Betaproteobacteria. The existence of distinct bacterial communities associated with particular mineral grain types may also be an example of niche separation and coexistence in sediments and soils, which cannot usually be detected due to difficulties in separating and concentrating minerals.

A multiplex polymerase chain reaction for the simultaneous detection of the virus and satellite components associated with cotton leaf curl begomovirus disease complex.

Cotton leaf curl disease (CLCuD) is caused by a complex of several whiteflies (Bemisia tabaci Genn.)-transmitted begomovirus species, Cotton leaf curl Multan virus (CLCuMuV), Cotton leaf curl Kokhran virus (CLCuKoV) and Cotton leaf curl Alabad virus (CLCuAlV) by individual of mixed infection, associated with Cotton leaf curl Multan betasatellite (CLCuMB) and several alphasatellites. The disease causes major economic losses in cotton in the Indian subcontinent. For monitoring of epidemiology and development of management strategies of CLCuD, a quick, sensitive and effective method capable of detecting all the begomovirus, betasatellite and alphasatellite components associated with CLCuD is required. With this objective, a multiplex polymerase chain reaction (mPCR) assay was developed for the simultaneous detection of these three viral components associated with CLCuD of cotton. Primers for each component were designed based on the retrieved reference sequences from the GenBank. Each pair of primers, designed for each of the respective component, was evaluated for its sensitivity and specificity in both the component-specific simplex polymerase chain reaction (sPCR) and mPCR assay. This report identified three viral component-specific pairs of primers which, in all combinations, amplified simultaneously the CP gene (780 nts) of the begomovirus, the ßC1gene (375 nts) of the betasatellite and the Rep gene (452 nts) of the alphasatellite associated with CLCuD in the mPCR assays. The amplified products specific to each component produced by these assays were identified based on their amplicon sizes, and the identities of the viral components amplified were confirmed by cloning and sequencing the amplicons obtained in the mPCR. The mPCR assay was validated using naturally CLCuD-affected cotton plants of the fields. This assay will be useful for rapid detection of CLCuD-associated begomovirus, betasatellite and alphasatellite DNA in field samples, extensive resistance screening in resistance breeding programme, and also monitoring epidemiology for detection of virus and its components when symptoms are mild or absent in the plant.

An intracytoplasmic IL-10 receptor variant permits rapid reduction in STAT3 activation.

Within the interleukin-10 receptor 1 (IL10R1) gene, two common variants are associated with certain diseases: single-nucleotide polymorphism 3 (SNP3), a serine-138 to glycine mutation is in linkage disequilibrium with SNP4, a glycine-330 to arginine mutation, both of which are considered loss-of-function alleles. However, the molecular consequence of G330R is unknown. We investigated possible roles of G330R on the dynamics of IL10R1 surface expression and signal transducer and activator of transduction (STAT) phosphorylation. HeLa cells expressing the respective IL10R1 haplotype were stimulated with IL-10. Significant reduction of IL10R1 surface expression was observed after ligand binding. Receptor expression remained low on continuous incubation with IL-10. In contrast, when treated with an IL-10 pulse, IL10R1 surface expression returned to its resting state within 3-9 h irrespective of the haplotype. STAT3 was rapidly phosphorylated both in cells with wild-type (WT) or variant IL10R1, and maintained phosphorylated when cells were cultured with IL-10. On IL-10 pulse, however, STAT3 phosphorylation declined rapidly in cells expressing IL10R1-G330R but not IL10R1-WT or S138G. Similar dynamics were observed with STAT1 phosphorylation at Tyr701. No differences in janus kinase 1 (JAK1) activation were observed in cells with WT or variant IL10R1. Our results indicate that IL10R1-G330R does not alter surface expression but duration of STAT phosphorylation, indicating that the position of G330 is important in stabilizing the STAT signal.

Cobalt-oxo core of a water-oxidizing catalyst film.

In photosynthesis, water is oxidized at a protein-bound Mn(4)Ca complex. Artificial water-oxidation catalysts that are similarly efficient and based on inexpensive and abundant materials are of great interest. Recently, assembly of a catalyst as an amorphous layer on inert cathodes by electrodeposition starting from an aqueous solution of cobalt ions and potassium phosphate has been reported. X-ray absorption spectroscopy on the cobalt catalyst film (CoCF) suggests that its central structural unit is a cluster of interconnected complete or incomplete Co(III)-oxo cubanes. Potassium ligation to Co-bridging oxygens could result in Co(3)K(mu-O)(4) cubanes, in analogy to the Mn(3)Ca(mu-O)(4) cubane motif proposed for the photosynthetic Mn complex. The similarities in function and oxidative self-assembly of CoCF and the catalytic Mn complex in photosynthesis are striking. Our study establishes a close analogy also with respect to the metal-oxo core of the catalyst.

Clinical profile and prevalence of rotavirus infection in children presented with acute diarrhea at tertiary care referral hospital at northern part of India.

A prospective analysis of 90 clinically diagnosed cases with acute diarrhea over a period of one year was carried out to determine the prevalence of rotavirus infection in children between 2 months to 2 years of age. Enzyme Linked Immunosorbent Assay (ELISA) and Polyacrylamide Gel Electrophoresis (PAGE) were used for detection of rotavirus from stool sample. Fourteen (15.6%) of them were found to be positive for group A rotavirus, 9 (23%) cases were between 6 months to 1 year of age. Rotavirus excretion was highest (50%) when all three symptoms (diarrhea, vomiting and fever) occurred in the same child. A planned study for surveillance of rotavirus serotypes is required from this area.

The DEAD box protein p68: a novel coactivator of Stat3 in mediating oncogenesis.

DEAD box RNA helicase p68 acts as a transcriptional coactivator of several oncogenic transcription factors apart from being a vital player of RNA metabolism. Signal transducer and activator of transcription 3 (Stat3) is a major oncogenic contributor of diverse cancers, including that of colon. Deciphering the mechanistic insights of coactivation of Stat3 transcriptional activity may aid in improved therapeutic strategies. Here we report for the first time a novel mechanism of alliance between p68 and Stat3 in stimulating transcriptional activity of Stat3. Interestingly, we observed that the expression of p68 and Stat3 bears strong positive correlation and significant colocalization in normal and colon carcinoma patient samples. We demonstrated that p68 directly interacts with Stat3 in HEK293 cells as well as multiple colon cancer cell lines. Additionally, p68 positively modulated both mRNA and protein expression levels of Stat3 target genes; promoter activity of Stat3 target gene Mcl-1 in multiple colon cancer cell lines. Also, p68 occupied the promoters of multiple Stat3 target genes in enhancing Stat3-dependent transcription. Moreover, the strong positive correlation between the abundance of p68 and Stat3 target genes in the same set of colon carcinoma samples further supported our observations. Enhanced expression levels of Stat3 target genes observed in primary tumors and metastatic lung nodules, generated in mice colorectal allograft model using syngeneic cells stably expressing p68, further reinforced our in vitro findings. Hence, this study unravels novel modes of p68-mediated oncogenesis through coactivation of Stat3 and enhancing Stat3 signaling.

The DEAD box protein p68: a crucial regulator of AKT/FOXO3a signaling axis in oncogenesis.

Increased abundance of proto-oncogene AKT and reduced expression of tumor suppressor Forkhead box O3 (FOXO3a), the downstream target of AKT, is frequent in carcinogenesis. Mechanistic insights of AKT gene regulation are limited. DEAD box RNA helicase p68 is overexpressed in various cancers and acts as a transcriptional co-activator of several transcription factors, including ß-catenin. Here, we report a novel mechanism of p68-mediated transcriptional activation of AKT, and its ensuing effect on FOXO3a, in colon carcinogenesis. Interestingly, we found that the expression of p68 and AKT exhibits strong positive correlation in normal and colon carcinoma patient samples. In addition, p68 increased both AKT messenger RNA (mRNA) and protein, enhanced AKT promoter activity in multiple colon cancer cell lines. Conversely, p68 knockdown led to reduced AKT mRNA and protein, diminished AKT promoter activity. Here, we demonstrated that p68 occupies AKT promoter with ß-catenin as well as nuclear factor-κB (NF-κB)and cooperates with these in potentiating AKT transcription. Furthermore, p68 and FOXO3a expression followed inverse correlation in the same set of colon carcinoma samples. We observed that p68 significantly reduced FOXO3a protein level in an AKT-dependent manner. Studies in primary tumors and metastatic lung nodules generated in mice colorectal allograft model, using syngeneic cells stably expressing p68, corroborated our in vitro findings. Hence, a new mechanism of oncogenesis is attributed to p68 by upregulation of AKT and consequent nuclear exclusion and degradation of tumor suppressor FOXO3a.

Increased expression of HIF2α during iron deficiency-associated megakaryocytic differentiation.

BACKGROUND: Iron deficiency is associated with reactive thrombocytosis; however, the mechanisms driving this phenomenon remain unclear. We previously demonstrated that this occurs alongside enhanced megakaryopoiesis in iron-deficient rats, without alterations in the megakaryopoietic growth factors thrombopoietin, interleukin-6, or interleukin-11. OBJECTIVES: The aim of this study was to evaluate megakaryocyte differentiation under iron deficiency in an in vitro model and to investigate potential genes involved in this process. METHODS: Human erythroleukemia and megakaryoblastic leukemia cell lines, as well as cord-blood derived hematopoietic stem cells were cultured under iron deficiency. Cell morphology, ploidy, expression of CD41, CD61, and CD42b, and proplatelet formation were assessed in iron-deficient cultures. Polymerase chain reaction arrays were used to identify candidate genes that were verified using real-time polymerase chain reaction. Hypoxia-inducible factor 1, α subunit (HIF2α) protein expression was assessed in bone marrow sections from iron-deficient rats and vascular endothelial growth factor (VEGF)-A in culture supernatants. RESULTS AND CONCLUSIONS: Iron deficiency enhanced megakaryoid features in cell lines, increasing ploidy and initiating formation of proplatelet-like structures. In cord blood cell cultures, iron deficiency increased the percentage of cells expressing megakaryopoietic markers and enhanced proplatelet formation. HIF2α and VEGF were identified as potential pathways involved in this process. HIF2α protein expression was increased in megakaryocytes from iron-deficient rats, and VEGF-A concentration was higher in iron-deficient culture supernatants. Addition of VEGF-A to cell cultures increased percentage expression of megakaryocyte CD41. In conclusion, the data demonstrate that iron deficiency augments megakaryocytic differentiation and proplatelet formation and a potential role of HIF2α in megakaryopoiesis.

Antinuclear antibodies in pleural fluid.

Systemic lupus erythematosus (SLE) frequently involves the pleura with resultant pleural effusion. Previous studies have reported that detection of antinuclear antibodies (ANA) in pleural fluid using animal tissue as substrate was a sensitive and specific method for distinguishing SLE pleural effusions from other etiologies. The HEp-2 ANA, which uses a human cell line as substrate, is now the preferred ANA test; however, to our knowledge, no studies on pleural fluid using this assay have been reported. To determine its sensitivity and specificity, when measured in pleural fluid, HEp-2 ANA levels were determined in pleural effusion samples associated with a variety of different etiologies, including SLE, malignancy, congestive heart failure, pneumonia, tuberculosis, and a miscellaneous group of diseases. Pleural fluid ANA results were positive in 14 of 82 samples. Six of the eight (75 percent) pleural fluid samples collected from patients with SLE were ANA positive, and all but one had high titers (> 1:160) with a homogenous staining pattern. The remaining two patients with SLE with negative pleural fluid ANA had recurrent pulmonary emboli and congestive heart failure, rather than lupus pleuritis. Eight of 74 patients (10.8 percent) without clinical evidence of SLE had a positive pleural fluid ANA, with the majority having a speckled pattern. High titers were noted in three. These results indicate that a negative or low titer ANA and a speckled staining pattern in pleural fluid from a patient suspected of lupus pleuritis suggest an alternative diagnosis. High pleural fluid titers (up to 1:640) were seen occasionally in patients with inflammatory pleural effusions in the absence of SLE.

Corticosteroid rescue treatment of progressive fibroproliferation in late ARDS. Patterns of response and predictors of outcome.

Pulmonary fibroproliferation (PFP) is directly or indirectly the leading cause of death in patients with late ARDS. We previously reported our experience using intravenous corticosteroids (IVC) in 8 patients with late ARDS and now have expanded our observation to a total of 25 patients with severe fibroproliferation (mean lung injury score [LIS] 3) and progressive respiratory failure (RF). Thirteen patients had open-lung biopsy before treatment. Patients were started on IVC treatment (IVCT) an average of 15 +/- 7.5 days into mechanical ventilation (MV). Significant physiologic improvement (SPI) to IVCT was defined as a reduction in LIS of greater than 1 point or an increase in PaO2:FIO2 ratio of greater than 100. We observed three patterns of response: rapid responders (RR) had an SPI by day 7 (n = 15); delayed responders (DR) had an SPI by day 14 (n = 6); nonresponders (NR) were without SPI by day 14 (n = 4). Overall the following significant mean changes were seen within 7 days of IVCT: LIS from 3 to 2 (p = 0.001), PaO2:FIO2 from 162 to 234 (p = 0.0004), PEEP from 11 to 6.8 cm H2O (p = 0.001), chest radiograph score from 3.8 to 3.0 (p = 0.009), and VE from 16 to 13.6 L/min (p = 0.01). Development of pneumonia was related to the pattern of response. Surveillance bronchoscopy was effective in identifying pneumonia in eight afebrile patients. Nineteen of 25 (76 percent) patients survived the ICU admission. Comparisons were made between survivors (S) and nonsurvivors (NS) and among the three groups of responders. At the time ARDS developed, no physiologic or demographic variable could discriminate between S and NS. At the time of IVCT, only liver failure was more frequent in nonsurvivors (p = 0.035). Histologic findings at open-lung biopsy and pattern of physiologic response clearly predicted outcome. The presence of preserved alveolar architecture (p = 0.045), myxoid type fibrosis (p = 0.045), coexistent intraluminal bronchiolar fibrosis (p = 0.0045), and lack of arteriolar subintimal fibroproliferation (p = 0.045) separated S from NS. ICU survival rate was 86 percent in responders and 25 percent in nonresponders (p = 0.03). Only one death resulted from refractory respiratory failure.

Severe intravascular hemolysis associated with brown recluse spider envenomation. A report of two cases and review of the literature.

Envenomation by the brown recluse spider (loxoscelism) is classically associated with a necrotic ulcer. Systemic manifestations occur in a minority of cases, but are generally mild and self-limited. The hematologic complications of brown recluse spider bite range from mild hemolysis to fulminant intravascular hemolysis with or without evidence of disseminated intravascular coagulation. Intravascular hemolysis is a rare but occasionally lethal complication of brown recluse spider envenomation. This article presents two cases of severe hemolysis associated with loxoscelism occurring in two young women in Memphis, Tennessee. The second documented death in an adult from severe hemolysis due to a brown recluse spider bite is reported. A review of the literature emphasizing the pathogenic mechanisms of spider bite hemolysis is also included.

Association of Chlamydia trachomatis immunoglobulin gamma titers with dystrophic peritoneal calcification, psammoma bodies, adhesions, and hydrosalpinges.

OBJECTIVE: To correlate Chlamydia trachomatis immunoglobulin gamma (IgG) titers with psammoma bodies, dystrophic peritoneal calcification, degree of calcification, adhesions, and hydrosalpinges. DESIGN: This is a prospective single-blinded histologic analysis of tissue and retrospective analysis of historical laboratory and clinical variables. SETTING: Tertiary hospital and private practice patient charts. PATIENTS: Sixty consecutive patients with C. trachomatis IgG titers reported on the coding sheets of a previous study for endometriosis. MAIN OUTCOME MEASURES: The histologic slides were reviewed in a blinded fashion for calcification. Previously used data sheets were reviewed for C. trachomatis IgG titers. Historical data, adhesion scores, hystrosalpingogram findings, and laparoscopic findings were obtained from charts. RESULTS: Dystrophic calcification, psammoma bodies, moderate-to-severe dystrophic calcification and hydrosalpinges were associated with positive C. trachomatis IgG titers. CONCLUSION: This study suggests relationship of C. trachomatis with dystrophic calcification, psammoma bodies, adhesions, and hydrosalpinges. This relationship suggests that C. trachomatis IgG titers can be used as a marker to help determine those infertility patients who might best benefit from hysterosalpingogram or laparoscopy and in clinical studies of endometriosis, infertility, pain, or ovarian cancer. However, there is no current data to suggest a need for therapy on the basis of a positive C. trachomatis IgG titer or of dystrophic peritoneal calcification.

Placental metastasis from maternal carcinoma. A report of three cases.

Metastasis of a maternal neoplasm to the products of conception is extremely rare. Of the 54 reported cases in the world literature, only 14 (25%) showed fetal metastasis. More than 50% of the reported cases were not examined grossly or had no visually apparent tumor deposits. Malignant melanoma is the most common malignant maternal neoplasm to metastasize to the products of conception. We report three unusual maternal malignant neoplasms (one pancreatic and two breast cancer) with evidence of placental metastasis and discuss the risk factors for fetal involvement in these cases.

Frequency of sex chromatin in the buccal smear of newborn females and their mothers during first six days after delivery.

The frequencies of sex chromatin in the buccal smear of the newborn females and their mothers were low on the first post-partum day and it increased gradually during the second and third day. By the fourth and fifth day it stabilized and the incidence of sex chromatin both in the mothers and the children became similar, although the frequency on the first day was significantly lower in the newborn. The incidences of pyknotic cells in the buccal smears of the newborns and their mothers were highest on day one and these declined rapidly in the following days. The significance of these findings have been discussed.

csi2p modulates microtubule dynamics and organizes the bipolar spindle for chromosome segregation.

Proper chromosome segregation is of paramount importance for proper genetic inheritance. Defects in chromosome segregation can lead to aneuploidy, which is a hallmark of cancer cells. Eukaryotic chromosome segregation is accomplished by the bipolar spindle. Additional mechanisms, such as the spindle assembly checkpoint and centromere positioning, further help to ensure complete segregation fidelity. Here we present the fission yeast csi2+. csi2p localizes to the spindle poles, where it regulates mitotic microtubule dynamics, bipolar spindle formation, and subsequent chromosome segregation. csi2 deletion (csi2Δ) results in abnormally long mitotic microtubules, high rate of transient monopolar spindles, and subsequent high rate of chromosome segregation defects. Because csi2Δ has multiple phenotypes, it enables estimates of the relative contribution of the different mechanisms to the overall chromosome segregation process. Centromere positioning, microtubule dynamics, and bipolar spindle formation can all contribute to chromosome segregation. However, the major determinant of chromosome segregation defects in fission yeast may be microtubule dynamic defects.

Tumoral calcium pyrophosphate dihydrate crystal deposition disease: a rare diagnosis by fine-needle aspiration.

Calcium pyrophosphate dihydrate crystal deposition disease (CPPD) is a well-recognized inflammatory joint disorder characterized by presence of calcium pyrophosphate dihydrate crystals in intraarticular and periarticular tissue. We report here a case of a 48-year-old male who presented with painless right hand swelling. Clinical suspicion was that of malignant soft tissue tumor. Fine-needle aspiration (FNA) yielded chalky white gritty material. Microscopic examination showed large areas of basophilic calcified material, histiocytes, giant cells and characteristic rhomboid shaped crystals. At places, chondroid material was also identified, hence, diagnosis of CPPD was made. This was confirmed on histopathological examination. Tophaceous/ tumoral pseudogout is a rare form of CPPD and it is important to recognize that this form can be diagnosed in FNA cytology (FNAC) and misdiagnosis of benign or malignant cartilaginous lesions can be avoided.

The p53-inducible gene 3 (PIG3) contributes to early cellular response to DNA damage.

The p53-inducible gene 3 (PIG3) is originally isolated as a p53 downstream target gene, but its function remains unknown. Here, we report a role of PIG3 in the activation of DNA damage checkpoints, after UV irradiation or radiomimetic drug neocarzinostatin (NCS). We show that depletion of endogenous PIG3 sensitizes cells to DNA damage agents, and impaired DNA repair. PIG3 depletion also allows for UV- and NCS-resistant DNA synthesis and permits cells to progress into mitosis, indicating that PIG3 knockdown can suppress intra-S phase and G2/M checkpoints. PIG3-depleted cells show reduced Chk1 and Chk2 phosphorylation after DNA damage, which may directly contribute to checkpoint bypass. PIG3 exhibited diffuse nuclear staining in the majority of untreated cells and forms discrete nuclear foci in response to DNA damage. PIG3 colocalizes with gamma-H2AX and 53BP1 to sites of DNA damage after DNA damage, and binds to a gamma-H2AX. Notably, PIG3 depletion decreases the efficient induction and maintenance of H2AX phosphorylation after DNA damage. Moreover, PIG3 contributes to the recruitment of 53BP1, Mre11, Rad50 and Nbs1 to the sites of DNA break lesions in response to DNA damage. Our combined results suggest that PIG3 is a critical component of the DNA damage response pathway and has a direct role in the transmission of the DNA damage signal from damaged DNA to the intra-S and G2/M checkpoint machinery in human cells.

The 3' --> 5' exonuclease of T4 DNA polymerase removes premutagenic alkyl mispairs and contributes to futile cycling at O6-methylguanine lesions.

We have studied the processing of O(6)-methylguanine (m6G)-containing oligonucleotides and N-methyl-N-nitrosourea (MNU)-treated DNA templates by the 3' --> 5' exonuclease of T4 DNA polymerase. In vitro biochemical analyses demonstrate that the exonuclease can remove bases opposite a defined m6G lesion. The efficiency of excision of a terminal m6G.T was similar to that of m6G.C, and both were excised as efficiently as a G.T substrate. Partitioning assays between the polymerase and exonuclease activities, performed in the presence of dNTPs, resulted in repeated incorporation and excision events opposite the m6G lesion. This idling produces dramatically less full-length product, relative to natural substrates, indicating that the 3' --> 5' exonuclease may contribute to DNA synthesis inhibition by alkylating agents. Genetic data obtained using an in vitro herpes simplex virus-thymidine kinase assay support the inefficiency of the exonuclease as a "proofreading" activity for m6G, since virtually all mutations produced by the native enzyme using MNU-treated templates were G --> A transitions. Comparison of MNU dose-response curves for exonuclease-proficient and -deficient forms of T4 polymerase reveals that the exonuclease efficiently removes 50-86% of total premutagenic alkyl mispairs. We propose that idling of exonuclease-proficient polymerases at m6G lesions during repair DNA synthesis provides the biochemical explanation for cellular cytotoxicity of methylating agents.

Anecdotal association of endosalpingiosis with chlamydia trachomatis IgG titers and Fitz-Hugh-Curtis adhesions.

STUDY OBJECTIVE: To correlate Chlamydia trachomatis IgG titers with endosalpingiosis. DESIGN: Prospective, single-blind, histologic analysis of tissue, and retrospective analysis of historical laboratory and clinical variables. SETTING: A tertiary care patient practice. PATIENTS: Sixty consecutive patients with C. trachomatis IgG titers, 28 positive and 32 negative. INTERVENTIONS: The second review of histology slides and charts of these women. MEASUREMENTS AND MAIN RESULTS: One patient was found to have endosalpingiosis. She had Fitz-Hugh-Curtis (FHC) adhesions and a positive C. trachomatis IgG titer, but no endometriosis. CONCLUSIONS: This review confirmed an anecdotal association of endosalpingiosis with positive C. trachomatis IgG titers and FHC adhesions, but failed to demonstrate a statistically significant relationship due to the size of the sample.