A novel nested multiplex polymerase chain reaction (PCR) assay for differential detection of Entamoeba histolytica, E. moshkovskii and E. dispar DNA in stool samples

<p>Abstract</p> <p>Background</p> <p><it>E. histolytica</it>, a pathogenic amoeba, is indistinguishable in its cyst and trophozoite stages from those of non-pathogenic <it>E. moshkovskii </it>and <it>E. dispar </it>by light microscopy. We have developed a nested multiplex PCR targeting a 16S-like rRNA gene for differential detection of all the three morphologically similar forms of <it>E. histolytica</it>, <it>E. moshkovskii </it>and <it>E. dispar </it>simultaneously in stool samples.</p> <p>Results</p> <p>The species specific product size for <it>E. histolytica</it>, <it>E. moshkovskii </it>and <it>E. dispar </it>was 439, 553 and 174 bp respectively, which was clearly different for all the three <it>Entamoeba </it>species. The nested multiplex PCR showed a sensitivity of 94% and specificity of 100% for the demonstration of <it>E. histolytica</it>, <it>E. moshkovskii </it>and <it>E. dispar </it>DNA in stool samples. The PCR was positive for <it>E. histolytica</it>, <it>E. moshkovskii </it>and <it>E. dispar </it>in a total of 190 out of 202 stool specimens (94% sensitive) that were positive for <it>E. histolytica/E. dispar</it>/<it>E. moshkovskii </it>by examination of stool by microscopy and/or culture. All the 35 negative control stool samples that were negative for <it>E. histolytica/E. dispar</it>/<it>E. moshkovskii </it>by microscopy and culture were also found negative by the nested multiplex PCR (100% specific). The result from the study shows that only 34.6% of the patient stool samples that were positive for <it>E. histolytica/E. dispar</it>/<it>E. moshkovskii </it>by examination of stool by microscopy and/or culture, were actually positive for pathogenic <it>E. histolytica </it>and the remaining majority of the stool samples were positive for non-pathogenic <it>E. dispar </it>or <it>E. moshkovskii </it>as demonstrated by the use of nested multiplex PCR.</p> <p>Conclusion</p> <p>The present study reports a new nested multiplex PCR strategy for species specific detection and differentiation of <it>E. histolytica</it>, <it>E. dispar </it>and <it>E. moshkovskii </it>DNA in stool specimens. The test is highly specific, sensitive and also rapid, providing the results within 12 hours of receiving stool specimens.</p

Detection of excretory Entamoeba histolytica DNA in the urine, and detection of E. histolytica DNA and lectin antigen in the liver abscess pus for the diagnosis of amoebic liver abscess

<p>Abstract</p> <p>Background</p> <p>Amoebic liver abscess (ALA) and pyogenic liver abscesses (PLA) appear identical by ultrasound and other imaging techniques. Collection of blood or liver abscess pus for diagnosis of liver abscesses is an invasive procedure, and the procedure requires technical expertise and disposable syringes. Collection of urine is a noninvasive procedure. Therefore, there has been much interest shown towards the use of urine as an alternative clinical specimen for the diagnosis of some parasitic infections. Here, we report for the first time the detection of <it>E. histolytica </it>DNA excreted in the urine for diagnosis of the cases of ALA.</p> <p>Results</p> <p><it>E. histolytica </it>DNA was detected in liver abscess pus specimen of 80.4% of ALA patients by a nested multiplex polymerase chain reaction (PCR) targeting 16S-like r RNA gene. The nested PCR detected <it>E. histolytica </it>DNA in all 37 (100%) liver abscess pus specimens collected prior to metronidazole treatment, but were detected in only 53 of 75 (70.6%) pus specimens collected after therapy with metronidazole. Similarly, the PCR detected <it>E. histolytica </it>DNA in 21 of 53 (39.6%) urine specimens of ALA patients. The test detected <it>E. histolytica </it>DNA in only 4 of 23 (17.4%) urine specimens collected prior to metronidazole treatment, but were detected in 17 of 30 (56.7%) urine specimens collected after treatment with metronidazole. The enzyme-linked immunosorbent assay (ELISA) for the detection of lectin <it>E. histolytica </it>antigen in the liver abscess pus showed a sensitivity of 50% and the indirect haemagglutination (IHA) test for detection of amoebic antibodies in the serum showed a sensitivity of 76.8% for the diagnosis of the ALA.</p> <p>Conclusion</p> <p>The present study for the first time shows that the kidney barrier in ALA patients is permeable to <it>E. histolytica </it>DNA molecule resulting in excretion of <it>E. histolytica </it>DNA in urine which can be detected by PCR. The study also shows that the PCR for detection of <it>E. histolytica </it>DNA in urine of patients with ALA can also be used as a prognostic marker to assess the course of the diseases following therapy by metronidazole. The detection of <it>E. histolytica </it>DNA in urine specimen of ALA patients provides a new approach for the diagnosis of ALA.</p

Charge and critical density of strange quark matter

The electric charge of strange quark matter is of vital importance to experiments. A recent investigation shows that strangelets are most likely highly negatively charged, rather than slightly positively charged as previously believed. Our present study indicates that negative charges can indeed lower the critical density, and thus be favorable to the experimental searches in heavy ion collisions. However, too much negative charges can make it impossible to maintain flavor equilibrium.Comment: 4 pages, LATeX with REVTeX style, one PS figure. To be published in Phys. Rev. C 59(6), 199

Vasorelaxation to Eugenol, Curcumin and Nanocurcumin mediated by differential augmentation of Na+, K+-ATPase activity in middle uterine artery of Capra hircus

59-67Sodium-potassium ATPase (Na, K-ATPase) is an integral protein universally distributed in the plasma membrane of all eukaryotic cells. It regulates trans-epithelial sodium ion transport, muscle contraction, nerve excitability, secretary functions and intracellular signaling. We investigated the mechanism of vasorelaxation effect of Eugenol, Curcumin and Nanocurcumin in the middle uterine artery (MUA) of goat to evaluate their possible therapeutic potential in hypertension during pregnancy or pre-eclampsia. The UA rings were isometrically contracted by K+-free MKHS to plateau and relaxed with KCl (10 &mu;M-10 mM) following standard protocol. The K+ -vasorelaxation response (VR) was taken either in presence of ouabain with or without Eugenol/Curcumin/Nanocurcumin (10 &mu;M). Rmax (maximum % vasorelaxation) of K+-induced VR-curve elicited in UA rings was reduced (from 6.90% and 26.55%) to 4-5% in presence of Ouabain in both NP and P goats. Rmax obtained from KVR curve elicited in UA rings were augmented in presence of Eugenol (NP: 16.22%, P: 42.67%) or Curcumin (NP:21.23%, P:35.49%) or Nanocurcumin (NP:6.91%, P:34.88%). K+ vasorelaxation was inhibited in presence of Ouabain and Eugenol/ Curcumin/Nanocurcumin (Rmax: 6.11%, 9.18%, 4.11% in non-P, respectively) and (Rmax: 5.40%, 3.39%, 6.00% in P, respectively). The results indicated that (i) there is increased function of &alpha;1- Na+, K+-ATPase in UA of P goats (ii) Eugenol, Curcumin and Nano-curcumin differentially augmented the function and expression of &alpha;1 isoenzymes of sodium pump in UA of non-P and P goats. Thus, the polyphenols potentially activate &alpha;1 isoenzymes of Na+-K+-ATPase and could possibly be useful as therapeutic in hypertension during pregnancy

Mass formulas and thermodynamic treatment in the mass-density-dependent model of strange quark matter

The previous treatments for strange quark matter in the quark mass-density-dependent model have unreasonable vacuum limits. We provide a method to obtain the quark mass parametrizations and give a self-consistent thermodynamic treatment which includes the MIT bag model as an extreme. In this treatment, strange quark matter in bulk still has the possibility of absolute stability. However, the lower density behavior of the sound velocity is opposite to previous findings.Comment: Formatted in REVTeX 3.1, 5 pages, 3 figures, to appear in PRC6

Thermodynamics, strange quark matter, and strange stars

Because of the mass density-dependence, an extra term should be added to the expression of pressure. However, it should not appear in that of energy according to both the general ensemble theory and basic thermodynamic principle. We give a detail derivation of the thermodynamics with density-dependent particle masses. With our recently determined quark mass scaling, we study strange quark matter in this new thermodynamic treatment, which still indicates a possible absolute stability as previously found. However, the density behavior of the sound velocity is opposite to the previous finding, but consistent with one of our recent publication. We have also studied the structure of strange stars using the obtained equation of state.Comment: 6 pages, 6 PS figures, REVTeX styl

Curvature energy effects on strange quark matter nucleation at finite density

We consider the effects of the curvature energy term on thermal strange quark matter nucleation in dense neutron matter. Lower bounds on the temperature at which this process can take place are given and compared to those without the curvature term.Comment: PlainTex, 6 pp., IAG-USP Rep.5

Tissue Invasion by Entamoeba histolytica: Evidence of Genetic Selection and/or DNA Reorganization Events in Organ Tropism

Entamoeba histolytica infection may have various clinical manifestations. Nine out of ten E. histolytica infections remain asymptomatic, while the remainder become invasive and cause disease. The most common form of invasive infection is amebic diarrhea and colitis, whereas the most common extra-intestinal disease is amebic liver abscess. The underlying reasons for the different outcomes are unclear, but a recent study has shown that the parasite genotype is a contributor. To investigate this link further we have examined the genotypes of E. histolytica in stool- and liver abscess-derived samples from the same patients. Analysis of all 18 paired samples (16 from Bangladesh, one from the United States of America, and one from Italy) revealed that the intestinal and liver abscess amebae are genetically distinct. The results suggest either that E. histolytica subpopulations in the same infection show varying organ tropism, or that a DNA reorganization event takes place prior to or during metastasis from intestine to liver