bluuebunny/crossref_metadata_2025 huggingface dataset:
Risk factors for ectopic pregnancy: A case‐control study
Authors: KARAER Abdullah , AVSAR Filiz A. , BATIOGLU Sertac
Published: November 2006

Aim:  To identify the risk factors for ectopic pregnancy.

Methods:  We conducted a prospective case‐control study for the role of several risk factors in the occurrence of ectopic pregnancy in Turkey. A total of 225 cases and 375 controls were compared for sociodemographic characteristics, cigarette smoking, obstetric, gynaecological, surgical histories, the presence or absence of assisted conception and contraceptive usage.

Results:  The main risk factors for ectopic pregnancy were prior ectopic pregnancy (adjusted odds ratio (AOR): 13.1) and a history of infectious reproductive system (AOR for pelvic inflammatory disease: 6.8). Other risk factors found to be associated with an increased risk for ectopic pregnancy were multisexual partner (AOR: 3.5), history of infertility (AOR: 2.5), induced conception cycle (AOR: 3.4), current intrauterine device usage (AOR: 3.2), prior Caesarean section (AOR: 2.1) and cigarette smoking at the time of conception (AOR = 1.7). On the contrary, barrier methods were protective from ectopic pregnancy (AOR: 0.4).

Conclusions:  The increased awareness and knowledge of risk factors have enabled an early and accurate diagnosis of ectopic pregnancy. This study has found prior pelvic infection to be a major aetiological factor for ectopic pregnancy. Furthermore, other factors found to be associated with ectopic pregnancy, such as prior ectopic pregnancy, infertility history and induced conception cycle, may be the result of a previous pelvic infection that may cause tubal sequelae. These factors are potential targets for intervention and modification.
Uterine rupture: Preventable obstetric tragedies?
Authors: MISHRA Sangeeta K. , MORRIS Norman , UPRETY Dhruba Kumar
Published: November 2006

Background:  Although ruptured uterus is nowadays a rare obstetric emergency in Western countries, it is still alarmingly common in developing countries, where it remains a major cause of maternal mortality and morbidity.

Aims:  To review the recent experience of uterine rupture at a tertiary obstetric unit in eastern Nepal and to recommend improvements in the current management of labour, especially obstructed labour, in a poorly resourced country.

Methods:  All cases of uterine rupture managed from March 2002 to March 2006 were identified retrospectively, and details were retrieved from medical records.

Results:  Fifty‐two women suffered from uterine rupture during the four‐year period, approximately one woman per month. Most were unbooked multigravidae, with no antenatal care. They nearly all began labour at home in the absence of a skilled birth attendant. After prolonged labour, usually prolonged second stage, various interventions had often been attempted at home or in other health facilities before admission. Most were shocked and required urgent laparotomy and blood transfusion. Many required intensive care and ventilatory support. Forty‐six per cent required hysterectomy and 5.8% subsequently suffered from a urogenital fistula. The maternal mortality rate in this series was 13.5%, and the stillbirth rate was 94.2%.

Conclusions:  Unsafe obstetric practices were identified, especially the injudicious use of oxytocic drugs and fundal pressure in prolonged second stage. Several achievable improvements in obstetric care are recommended, particularly aimed at reducing the delay in women reaching emergency obstetric care when labour is prolonged.
Ratiometric Fluorescence‐Imaging Assays of Plant Membrane Traffic Using Polyproteins
Authors: Samalova Marketa , Fricker Mark , Moore Ian
Published: November 2006

Fluorescent protein markers are widely used to report plant membrane traffic; however, effective protocols to quantify fluorescence or marker expression are lacking. Here the 20 residue self‐cleaving 2A peptide from Foot and Mouth Disease Virus was used to construct polyproteins that expressed a trafficked marker in fixed stoichiometry with a reference protein in a different cellular compartment. Various pairs of compartments were simultaneously targeted. Together with a bespoke image analysis tool, these constructs allowed biosynthetic membrane traffic to be assayed with markedly improved sensitivity, dynamic range and statistical significance using protocols compatible with the common plant transfection and transgenic systems. As marker and effector expression could be monitored in populations or individual cells, saturation phenomena could be avoided and stochastic or epigenetic influences could be controlled. Surprisingly, mutational analysis of the ratiometric assay constructs revealed that the 2A peptide was dispensable for efficient cleavage of polyproteins carrying a single internal signal peptide, whereas the signal peptide was essential. In contrast, a construct bearing two signal peptide/anchors required 2A for efficient separation and stability, but 2A caused the amino‐terminal moiety of such fusions to be mis‐sorted to the vacuole. A model to account for the behaviour of 2A in these and other studies in plants is proposed.
Golgin‐160 Promotes Cell Surface Expression of the Beta‐1 Adrenergic Receptor
Authors: Hicks Stuart W. , Horn Troy A. , McCaffery J. Michael , Zuckerman David M. , Machamer Carolyn E.
Published: November 2006

Golgin‐160 is a ubiquitously expressed peripheral Golgi membrane protein that is important for transduction of certain pro‐apoptotic signals at the Golgi complex. However, the role of golgin‐160 in normal Golgi structure and function is unknown. Here, we show that depletion of golgin‐160 using RNA interference (RNAi) does not affect Golgi morphology or constitutive membrane traffic in HeLa cells. However, depletion of golgin‐160 leads to significantly decreased cell surface levels of exogenously expressed β1‐adrenergic receptor (β1AR), which can be rescued by expression of RNAi‐resistant forms of golgin‐160. Furthermore, overexpression of golgin‐160 leads to higher surface levels of β1AR. Golgin‐160 is localized mostly in the cis and medial regions of the Golgi stack by immunoelectron microscopy, suggesting that it does not directly promote incorporation of β1AR into transport vesicles at the trans Golgi network. Golgin‐160 interacts with β1AR in vitro, and we mapped the interaction to a region between residues 140 and 257 in the head of golgin‐160 and the third intracellular loop of β1AR. Our results support the idea that golgin‐160 may promote efficient surface delivery of a subset of cargo molecules.
Stress Overload: A New Diagnosis
Authors: Lunney Margaret
Published: November 2006

PURPOSE.  To describe the phenomenon of stress overload as a nursing diagnosis.

METHODS.  A qualitative study using case study method was conducted with nine adults experiencing stress overload to fully describe the experience and identify possible defining characteristics. Current literature sources on stress and its related factors were examined to support stress overload as a nursing diagnosis for inclusion in the NANDA International classification.

FINDINGS.  Stress overload, defined as excessive amounts and types of demands that require action, is a human response that is experienced as a problem and contributes to the development of other problems. The proposed defining characteristics are perceives situational stress as excessive, expresses a feeling of tension or pressure, expresses difficulty in functioning as usual, expresses problems with decision‐making, demonstrates increased feelings of anger and impatience, and reports negative effects from stress such as physical symptoms or psychological distress.

PRACTICE IMPLICATIONS.  Nursing interventions such as active listening and decision‐making support are needed to help people reduce stress levels. Studies are needed to further validate the defining characteristics and related factors of this new diagnosis.
Pathophysiology and management of dengue hemorrhagic fever
Authors: CHUANSUMRIT AMPAIWAN , TANGNARARATCHAKIT KANCHANA
Published: November 2006

Dengue infection is caused by any of four dengue virus serotypes. The clinical manifestations range from asymptomatic infection to undifferentiated fever, dengue fever and dengue hemorrhagic fever (DHF). DHF is characterized by sustained high fever for 2–7 days; bleeding diathesis such as positive tourniquet test, petechiae, epistaxis and hematemesis; thrombocytopenia with platelet counts ≤ 100 × 109/L and plasma leakage due to increased vascular permeability evidenced by hemoconcentration, pleural effusion and ascites. Bleeding diathesis is caused by vasculopathy, thrombocytopenia, platelet dysfunction and coagulopathy. The three stages of clinical presentations are classified as febrile, toxic and convalescent. The toxic stage, which lasts 24–48 hours, is the most critical period, with rapid plasma leakage leading to circulatory disturbance. The severity of DHF varies from mild (World Health Organization grades I and II), with minimal and transient change in vital signs, to severe (World Health Organization grades III and IV), with threatened shock (e.g. blood pressure 100/90 mmHg) or profound shock. There is no specific treatment for DHF. Intensive supportive care is the most important aspect of management. Early recognition of the disease and careful monitoring for circulatory disturbance are essential. Optimal fluid therapy to maintain the functions of the vital organs during the critical period and effective control of bleeding episodes will lead to favorable outcomes. Administration of recombinant activated factor VII is suggested whenever massive bleeding does not respond to blood component therapy.
Is there a link between intermittent hypoxia‐induced respiratory plasticity and obstructive sleep apnoea?
Authors: Mahamed Safraaz , Mitchell Gordon S.
Published: November 2006

Although neuroplasticity is an important property of the respiratory motor control system, its existence has been appreciated only in recent years and, as a result, its functional significance is not completely understood. The most frequently studied models of respiratory plasticity is respiratory long‐term facilitation (LTF) following acute intermittent hypoxia and enhanced LTF following chronic intermittent hypoxia. Since intermittent hypoxia is a prominent feature of sleep‐disordered breathing, LTF and/or enhanced LTF may compensate for factors that predispose to sleep‐disordered breathing, particularly during obstructive sleep apnoea (OSA). Long‐term facilitation has been studied most frequently in rats, and exhibits interesting properties consistent with a role in stabilizing breathing during sleep. Specifically, LTF: (1) is prominent in upper airway respiratory motor activity, suggesting that it stabilizes upper airways and maintains airway patency; (2) is most prominent during sleep in unanaesthetized rats; and (3) exhibits sexual dimorphism (greatest in young male and middle‐aged female rats; smallest in middle‐aged male and young female rats). Although these features are consistent with the hypothesis that upper airway LTF minimizes the prevalence of OSA in humans, there is little direct evidence for such an effect. Here we review advances in our understanding of LTF and its underlying mechanisms and present evidence concerning a potential role for LTF in maintaining upper airway patency, stabilizing breathing and preventing OSA in humans. Regardless of the relationship between LTF and OSA, a detailed understanding of cellular and synaptic mechanisms that underlie LTF may guide the development of new drugs to regulate upper airway tone, thereby offsetting the tendency for upper airway collapse characteristic of heavy snoring and OSA.
Modulation of ischaemic contracture in mouse hearts: a ‘supraphysiological’ response to adenosine
Authors: Reichelt Melissa E. , Willems Laura , Peart Jason N. , Ashton Kevin J. , Matherne G. Paul , Blackburn Michael R. , Headrick John P.
Published: November 2006

While inhibition of ischaemic contracture was one of the first documented cardioprotective actions of exogenously applied adenosine, it is not known whether this is a normal function of endogenous adenosine generated during ischaemic stress. Additionally, the relevance of delayed contracture to postischaemic outcome is unclear. We tested the ability of endogenousversusexogenous adenosine to modify contracture (and postischaemic outcomes) in C57/Bl6 mouse hearts. During ischaemia, untreated hearts developed peak contracture (PC) of 85 ± 5 mmHg at 8.9 ± 0.8 min, with time to reach 20 mmHg (time to onset of contracture; TOC) of 4.4 ± 0.3 min. Adenosine (50 μm) delayed TOC to 6.7 ± 0.6 min, as did pretreatment with 10 μm2‐chloroadenosine (7.2 ± 0.5 min) or 50 nmof A1adenosine receptor (AR) agonist N6‐cyclohexyladenosine (CHA) (6.7 ± 0.3 min), but not A2AAR or A3AR agonists (20 nm2‐[4‐(2‐carboxyethyl) phenethylamino]‐5′ N‐methylcarboxamidoadenosine (CGS21680) or 150 nm2‐chloro‐N6‐(3‐iodobenzyl)‐adenosine‐5′‐N‐methyluronamide (Cl‐IB‐MECA), respectively). Adenosinergic contracture inhibition was eliminated by A1AR gene knockout (KO), mimicked by A1AR overexpression, and was associated with preservation of myocardial [ATP]. This adenosine‐mediated inhibition of contracture was, however, only evident after prolonged (10 or 15 min) and not brief (3 min) pretreatment. Ischaemic contracture was also insensitive to endogenously generated adenosine, since A1AR KO, and non‐selective and A1AR‐selective antagonists (50 μm8‐sulphophenyltheophylline and 150 nm8‐cyclopentyl‐1, 3‐dipropylxanthine (DPCPX), respectively), all failed to alter intrinsic contracture development. Finally, delayed contracture with A1AR agonism/overexpression or ischaemic 2,3‐butanedione monoxime (BDM; 5 μmto target Ca2+cross‐bridge formation) was linked to enhanced postischaemic outcomes. In summary, adenosinergic inhibition of contracture is solely A1AR mediated; the response is ‘supraphysiological’, evident only with significant periods of pre‐ischaemic AR agonism (or increased A1AR density); and ischaemic contracture appears insensitive to locally generated adenosine, potentially owing to the rapidity of contracture developmentversusthe finite time necessary for expression of AR‐mediated cardioprotection.
Effects of Age, Gender, and Race/Ethnicity on the Pharmacokinetics of Posaconazole in Healthy Volunteers
Authors: Sansone-Parsons Angela , Krishna Gopal , Simon Jason , Soni Peter , Kantesaria B. , Herron Jerry , Stoltz Randall
Published: November 2006

Posaconazole is a triazole antifungal for prophylaxis of invasive fungal infection and treatment of oropharyngeal candidiasis. We evaluated the effects of gender, age, and race/ethnicity (black or white) on the steady-state pharmacokinetics of posaconazole in two studies on healthy adult subjects (≥18 years of age). Additionally, we explored the effect of P-glycoprotein expression and MDR1 genotype on posaconazole pharmacokinetics in black and white subjects. Age, gender, and race/ethnicity had no clinically relevant effects on posaconazole pharmacokinetics. No association was observed between any MDR1 single-nucleotide polymorphism and the area under the concentration-time curve for posaconazole. Posaconazole was safe and well tolerated regardless of age, gender, or race/ethnicity. In conclusion, age, gender, and race/ethnicity have no clinically relevant effects on the steady-state pharmacokinetics of posaconazole in healthy adults; therefore, dosage adjustments based on these covariates are unnecessary.
Association of Fluconazole Area under the Concentration-Time Curve/MIC and Dose/MIC Ratios with Mortality in Nonneutropenic Patients with Candidemia
Authors: Pai Manjunath P. , Turpin Robin S. , Garey Kevin W.
Published: November 2006

The present study tested in vitro susceptibility of
Candida
bloodstream isolates to fluconazole to determine if the ratio of the fluconazole area under the concentration-time curve (AUC) or weight-normalized daily dose (dose
wn
) to MIC correlated with mortality. Fluconazole susceptibility and outcome data were determined for 77 patients with a positive
Candida
blood culture between 2002 and 2005. The most commonly isolated
Candida
species were
C. albicans
(64%),
C. glabrata
(14%),
C. parapsilosis
(8%),
C. tropicalis
(6%), and
C. lusitaniae
(4%). Only two isolates were classified as fluconazole resistant by the CLSI M27-A2 method. Fluconazole MICs were highest against
C. glabrata
relative to other
Candida
species. Overall the crude mortality assessed at hospital discharge was 19.4% (
n
= 15). Mortality rates by species were as follows:
C. albicans
, 16.3%;
C. glabrata
, 36.4%;
C. parapsilosis
, 0%;
C. tropicalis
, 0%;
C. lusitaniae
, 33.3%. A mortality rate of 50% was noted among patients infected with nonsusceptible isolates (MIC ≥ 16 μg/ml) compared to 18% for patients infected with susceptible (MIC ≤ 8 μg/ml) isolates (
P
= 0.17). The fluconazole dose
wn
/MIC (24-h) values were significantly higher for the 62 survivors (13.3 ± 10.5 [mean ± standard deviation]) compared to the 15 nonsurvivors (7.0 ± 8.0) (
P
= 0.03). The fluconazole AUC/MIC (24 h) values also trended higher for survivors (775 ± 739) compared to nonsurvivors (589 ± 715) (
P
= 0.09). These data support the dose-dependent properties of fluconazole. Underdosing fluconazole against less-susceptible
Candida
isolates has the potential to increase the risk of mortality associated with candidemia.
Identification and Structural Characterization of I84C and I84A Mutations That Are Associated with High-Level Resistance to Human Immunodeficiency Virus Protease Inhibitors and Impair Viral Replication
Authors: Mo Hongmei , Parkin Neil , Stewart Kent D. , Lu Liangjun , Dekhtyar Tatyana , Kempf Dale J. , Molla Akhteruzzaman
Published: November 2006

Two novel human immunodeficiency virus protease mutations, I84C and I84A, were identified in patient isolates. The mutants with I84C displayed high-level resistance (median, at least 56-fold) to nelfinavir and saquinavir, but the majority remained susceptible to lopinavir. In contrast, isolates with the I84A mutation exhibited ≥33-fold median increased levels of resistance to nelfinavir, indinavir, amprenavir, ritonavir, lopinavir, saquinavir, and atazanavir. Isolates with the I84A or I84C mutation tended to be more resistant than the isolates with the I84V mutation. Modeling of the structure of the mutant proteases indicated that the I84V, I84C, and I84A mutations all create unoccupied volume in the active site, with I84A introducing the greatest change in the accessible surface area from that of the wild-type structure.
Curing
Mycobacterium ulcerans
Infection in Mice with a Combination of Rifampin-Streptomycin or Rifampin-Amikacin
Authors: Lefrançois Sébastien , Robert Jérôme , Chauffour Aurélie , Ji Baohong , Jarlier Vincent
Published: November 2006

The curing activities of various durations of treatment with a combination of rifampin (RIF) and either streptomycin (STR) or amikacin (AMK) in murine
Mycobacterium ulcerans
infection were compared in two experiments. In the first experiment, treatment was begun 1 week after infection, when the inflammatory footpad lesion had not yet occurred (preventive model), and in the second experiment, treatment was begun 6 weeks after infection, when inflammatory footpad lesions were established (curative model). In the first experiment, 4 weeks of treatment with daily RIF-STR or RIF-AMK was able to postpone the occurrence of footpad lesion by 12 weeks (RIF-STR) or 17 weeks (RIF-AMK), thus demonstrating their promising bactericidal activities, but neither treatment was able to prevent the late occurrence of footpad lesions. In the second experiment, the overall cure rates, as assessed by the lack of rebound of inflammatory lesions or remultiplication of
M. ulcerans
, were only 62% after 2 weeks of treatment with RIF and an aminoglycoside and 85% after 4 weeks; but the cure rate reached 100% after 8 or 12 weeks of treatment. The cure rates were slightly higher with the AMK-containing combination than with the STR-containing combination, but the difference was at the limit of significance (
P
= 0.07). These results show that in the murine model of Buruli ulcer, 8 weeks is the optimal duration of treatment with a combination of RIF and an aminoglycoside.
In Vitro Antibacterial Activity of the Pyrrolopyrazolyl-Substituted Oxazolidinone RWJ-416457
Authors: Foleno Barbara D. , Abbanat Darren , Goldschmidt Raul M. , Flamm Robert K. , Paget Steve D. , Webb Glenda C. , Wira Ellyn , Macielag Mark J. , Bush Karen
Published: November 2006

RWJ-416457, an investigational pyrrolopyrazolyl-substituted oxazolidinone, inhibited the growth of linezolid-susceptible staphylococci, enterococci, and streptococci at concentrations of ≤4 μg/ml, generally exhibiting two- to fourfold-greater potency than that of linezolid. Time-kill studies demonstrated bacteriostatic effects for both RWJ-416457 and linezolid.
Antileishmanial Effect of 3-Aminooxy-1-Aminopropane Is Due to Polyamine Depletion
Authors: Singh Sushma , Mukherjee Angana , Khomutov Alex R. , Persson Lo , Heby Olle , Chatterjee Mitali , Madhubala Rentala
Published: November 2006

The polyamines putrescine, spermidine, and spermine are organic cations that are required for cell growth and differentiation. Ornithine decarboxylase (ODC), the first and rate-limiting enzyme in the polyamine biosynthetic pathway, catalyzes the conversion of ornithine to putrescine. As the polyamine biosynthetic pathway is essential for the growth and survival of
Leishmania donovani
, the causative agent of visceral leishmaniasis, inhibition of the pathway is an important leishmaniacidal strategy. In the present study, we examined for the first time the effects of 3-aminooxy-1-aminopropane (APA), an ODC inhibitor, on the growth of
L. donovani
. APA inhibited the growth of both promastigotes in vitro and amastigotes in the macrophage model, with the 50% inhibitory concentrations being 42 and 5 μM, respectively. However, concentrations of APA up to 200 μM did not affect the viability of macrophages. The effects of APA were completely abolished by the addition of putrescine or spermidine. APA induced a significant decrease in ODC activity and putrescine, spermidine, and trypanothione levels in
L. donovani
promastigotes. Parasites were transfected with an episomal ODC construct, and these ODC overexpressers exhibited significant resistance to APA and were concomitantly resistant to sodium antimony gluconate (Pentostam), indicating a role for ODC overexpression in antimonial drug resistance. Clinical isolates with sodium antimony gluconate resistance were also found to overexpress ODC and to have significant increases in putrescine and spermidine levels. However, no increase in trypanothione levels was observed. The ODC overexpression in these clinical isolates alleviated the antiproliferative effects of APA. Collectively, our results demonstrate that APA is a potent inhibitor of
L. donovani
growth and that its leishmaniacidal effect is due to inhibition of ODC.
Prevalence and Susceptibility Testing of New Species of
Pseudallescheria
and
Scedosporium
in a Collection of Clinical Mold Isolates
Authors: Alastruey-Izquierdo Ana , Cuenca-Estrella Manuel , Monzón Araceli , Rodriguez-Tudela Juan L.
Published: November 2006

The prevalence of new species of
Pseudallescheria
and
Scedosporium
in a collection of 46 clinical isolates was analyzed. Strain identification was done by morphological and molecular methods. Four
Scedosporium aurantiacum
isolates were detected among the panel of clinical strains. The susceptibility profile of
S. aurantiacum
was similar to that of
Scedosporium apiospermum
.
Pyrazinoic Acid and Its
n
-Propyl Ester Inhibit Fatty Acid Synthase Type I in Replicating Tubercle Bacilli
Authors: Zimhony Oren , Vilchèze Catherine , Arai Masayoshi , Welch John T. , Jacobs William R.
Published: November 2006

The activity of different analogs of pyrazinamide on
Mycobacterium tuberculosis
fatty acid synthase type I (FASI) in replicating bacilli was studied. Palmitic acid biosynthesis was diminished by 96% in bacilli treated with
n-
propyl pyrazinoate, 94% in bacilli treated with 5-chloro-pyrazinamide, and 97% in bacilli treated with pyrazinoic acid, the pharmacologically active agent of pyrazinamide. We conclude that the minimal structure of pyrazine ring with an acyl group is sufficient for FASI inhibition and antimycobacterial activity.
Shewanella oneidensis
MR-1 Fluxome under Various Oxygen Conditions
Authors: Tang Yinjie J. , Hwang Judy S. , Wemmer David E. , Keasling Jay D.
Published: November 2006

The central metabolic fluxes of
Shewanella oneidensis
MR-1 were examined under carbon-limited (aerobic) and oxygen-limited (microaerobic) chemostat conditions, using
13
C-labeled lactate as the sole carbon source. The carbon labeling patterns of key amino acids in biomass were probed using both gas chromatography-mass spectrometry (GC-MS) and
13
C nuclear magnetic resonance (NMR). Based on the genome annotation, a metabolic pathway model was constructed to quantify the central metabolic flux distributions. The model showed that the tricarboxylic acid (TCA) cycle is the major carbon metabolism route under both conditions. The Entner-Doudoroff and pentose phosphate pathways were utilized primarily for biomass synthesis (with a flux below 5% of the lactate uptake rate). The anaplerotic reactions (pyruvate to malate and oxaloacetate to phosphoenolpyruvate) and the glyoxylate shunt were active. Under carbon-limited conditions, a substantial amount (9% of the lactate uptake rate) of carbon entered the highly reversible serine metabolic pathway. Under microaerobic conditions, fluxes through the TCA cycle decreased and acetate production increased compared to what was found for carbon-limited conditions, and the flux from glyoxylate to glycine (serine-glyoxylate aminotransferase) became measurable. Although the flux distributions under aerobic, microaerobic, and shake flask culture conditions were different, the relative flux ratios for some central metabolic reactions did not differ significantly (in particular, between the shake flask and aerobic-chemostat groups). Hence, the central metabolism of
S. oneidensis
appears to be robust to environmental changes. Our study also demonstrates the merit of coupling GC-MS with
13
C NMR for metabolic flux analysis to reduce the use of
13
C-labeled substrates and to obtain more-accurate flux values.
Community Structure of Ammonia-Oxidizing Bacteria under Long-Term Application of Mineral Fertilizer and Organic Manure in a Sandy Loam Soil
Authors: Chu Haiyan , Fujii Takeshi , Morimoto Sho , Lin Xiangui , Yagi Kazuyuki , Hu Junli , Zhang Jiabao
Published: November 2006

The effects of mineral fertilizer (NPK) and organic manure on the community structure of soil ammonia-oxidizing bacteria (AOB) was investigated in a long-term (16-year) fertilizer experiment. The experiment included seven treatments: organic manure, half organic manure N plus half fertilizer N, fertilizer NPK, fertilizer NP, fertilizer NK, fertilizer PK, and the control (without fertilization). N fertilization greatly increased soil nitrification potential, and mineral N fertilizer had a greater impact than organic manure, while N deficiency treatment (PK) had no significant effect. AOB community structure was analyzed by PCR-denaturing gradient gel electrophoresis (PCR-DGGE) of the
amoA
gene, which encodes the α subunit of ammonia monooxygenase. DGGE profiles showed that the AOB community was more diverse in N-fertilized treatments than in the PK-fertilized treatment or the control, while one dominant band observed in the control could not be detected in any of the fertilized treatments. Phylogenetic analysis showed that the DGGE bands derived from N-fertilized treatments belonged to
Nitrosospira
cluster 3, indicating that N fertilization resulted in the dominance of
Nitrosospira
cluster 3 in soil. These results demonstrate that long-term application of N fertilizers could result in increased soil nitrification potential and the AOB community shifts in soil. Our results also showed the different effects of mineral fertilizer N versus organic manure N; the effects of P and K on the soil AOB community; and the importance of balanced fertilization with N, P, and K in promoting nitrification functions in arable soils.
Vulnerability of Pathogenic Biofilms to
Micavibrio aeruginosavorus
Authors: Kadouri Daniel , Venzon Nel C. , O'Toole George A.
Published: November 2006

The host specificity of the gram-negative exoparasitic predatory bacterium
Micavibrio aeruginosavorus
was examined.
M. aeruginosavorus
preyed on
Pseudomonas aeruginosa
, as previously reported, as well as
Burkholderia cepacia
,
Klebsiella pneumoniae
, and numerous clinical isolates of these species. In a static assay, a reduction in biofilm biomass was observed as early as 3 hours after exposure to
M. aeruginosavorus
, and an ∼100-fold reduction in biofilm cell viability was detected following a 24-h exposure to the predator. We observed that an initial titer of
Micavibrio
as low as 10 PFU/well or a time of exposure to the predator as short as 30 min was sufficient to reduce a
P. aeruginosa
biofilm. The ability of
Micavibrio
to reduce an existing biofilm was confirmed by scanning electron microscopy. In static and flow cell experiments,
M. aeruginosavorus
was able to modify the overall
P. aeruginosa
biofilm structure and markedly decreased the viability of
P. aeruginosa
. The altered biofilm structure was likely caused by an increase in cell-cell interactions brought about by the presence of the predator or active predation. We also conducted a screen to identify genes important for
P. aeruginosa-Micavibrio
interaction, but no candidates were isolated among the ∼10,000 mutants tested.
SO, a Protein Involved in Hyphal Fusion in
Neurospora crassa
, Localizes to Septal Plugs
Authors: Fleiβner André , Glass N. Louise
Published: November 2006

The colony of a filamentous ascomycete fungus typically grows as a multinucleate syncytium. While this syncytial organization has developmental advantages, it bears the risk of extensive damage caused by local injury of hyphae. Loss of cytoplasm in injured hyphae is restricted by the fast and efficient sealing of the central pores of hyphal crosswalls, or septa, by a peroxisome-derived organelle called the Woronin body. The formation of septal plugs is also associated with development and leads to separation of certain parts of the colony. Septal plugs associated with developmental processes or aging hyphae typically occur by the accumulation of sealing material. Here we report that in
Neurospora crassa
, a protein necessary for hyphal fusion and proper colony development called SO (SOFT) localizes to septal plugs. In response to injury, SO accumulates at the septal plug in a Woronin body-independent manner. However, the presence of the Woronin body affects the speed of accumulation of SO at the septal pore. We determined that SO contributes to, but is not essential for, septal plugging. SO accumulation was also observed at septal plugs formed during hyphal aging and during programmed cell death mediated by genetic differences at heterokaryon incompatibility (
het
) loci.
Characterization of the Outer Membrane Proteome of
Leptospira interrogans
Expressed during Acute Lethal Infection
Authors: Nally Jarlath E. , Whitelegge Julian P. , Bassilian Sara , Blanco David R. , Lovett Michael A.
Published: November 2006

Pathogenic
Leptospira
species adapt to a wide range of environmental conditions during disease transmission and infection. While the proteome of in vitro cultivated
Leptospira
has been characterized in several studies to date, relatively little is known of the proteome as expressed by
Leptospira
during disease processes. Isolates of
Leptospira
obtained from patients suffering the severe pulmonary form of leptospirosis cause acute lethal infection in guinea pigs and chronic asymptomatic infection in rats. Recent studies have demonstrated that protein and lipopolysaccharide constituents of
Leptospira
recovered from acutely infected guinea pig tissue differ from that of
Leptospira
in chronically infected rat tissue and in vitro cultivated
Leptospira
(J. E. Nally, E. Chow, M. C. Fishbein, D. R. Blanco, and M. A. Lovett, Infect. Immun. 73:3251-3260, 2005). In the current study, the proteome of
Leptospira
expressed during disease processes was characterized relative to that of in vitro cultivated
Leptospira
(IVCL) after enrichment for hydrophobic membrane proteins with Triton X-114. Protein samples were separated by two-dimensional gel electrophoresis, and antigens expressed during infection were identified by immunoblotting with monospecific antiserum and convalescent rat serum in addition to mass spectrometry. Results suggest a significant increase in the expression of the outer membrane protein Loa22 during acute infection of guinea pigs relative to other outer membrane proteins, whose expression is generally diminished relative to expression in IVCL. Significant amounts of LipL32 are also expressed by
Leptospira
during acute infection of guinea pigs.
Affinity-Purified Human Immunoglobulin G That Binds a Lacto-N-Neotetraose-Dependent Lipooligosaccharide Structure Is Bactericidal for Serogroup BNeisseria meningitidis
Authors: Estabrook Michele M. , Jarvis Gary A. , McLeod Griffiss J.
Published: November 2006

Despite technological advances, no vaccine to prevent serogroup B meningococcal disease is available. The failure to develop a vaccine has shifted the focus to an alternative outer membrane structure, lipooligosaccharide (LOS), because disseminated disease induces bactericidal immunoglobulin G (IgG) that binds LOS. The purpose of this study was to identify the LOS structure(s) that induces human bactericidal IgG by purification and characterization of these antibodies. Human LOS IgG antibodies were affinity purified by passage of intravenous immunoglobulin through purified, type-specific LOS having a known structure coupled to epoxy-activated Sepharose 6B. Pathogenic group B strains representing the major LOS serotypes were used to examine the binding and bactericidal activities of four LOS-specific IgG preparations. All four LOS-specific IgG preparations bound to strains expressing homologous, as well as heterologous, LOS serotypes as determined by flow cytometry and an enzyme-linked immunosorbent assay. With human complement, IgG that was purified with L7 LOS was bactericidal for strains expressing L3,7 and L2,4 LOS, serotypes expressed by the majority of disease-associated group B and C meningococci. In conclusion, we purified human LOS-specific IgG that binds meningococci across LOS glycose-specific serotypes. An antigen that is dependent on the glycose lacto-N-neotetraose induces IgG in humans that is bactericidal for L2, L3, L4, and L7 strains. A vaccine containing this antigen would have the potential to protect against the vast majority of group B meningococcal strains.
Hypothetical Protein Cpn0308 Is Localized in theChlamydia pneumoniaeInclusion Membrane
Authors: Luo Jianhua , Jia Tianjun , Flores Rhonda , Chen Ding , Zhong Guangming
Published: November 2006

The hypothetical protein encoded byChlamydia pneumoniaeopen reading frame cpn0308 was detected in inclusion membranes ofC. pneumoniae-infected cells using antibodies raised with Cpn0308 fusion proteins. The anti-Cpn0308 antibodies did not cross-react with IncA, a knownC. pneumoniaeinclusion membrane protein, although the anti-Cpn0308 antibody staining overlapped with the anti-IncA antibody labeling. The labeling of the inclusion membrane by the anti-Cpn0308 antibody was specifically blocked by the Cpn0308 but not IncA fusion proteins. The Cpn0308 antigen was detectable 24 h after infection and remained in the inclusion membrane throughout the infection course.
Differential Liver Protein Expression during Schistosomiasis
Authors: Harvie Marina , Jordan Thomas William , La Flamme Anne Camille
Published: November 2006

The arrival of eggs in the liver duringSchistosoma mansoniinfection initiates a protective granulomatous response; however, as the infection progresses, this response results in chronic liver fibrosis. To better understand the impact of schistosomiasis on liver function, we used a proteomic approach to identify proteins whose expression was significantly altered in schistosome-infected mice 8 weeks postinfection. Identification of differentially expressed proteins by mass fingerprinting revealed that schistosome infection markedly reduced the abundance of proteins associated with several normal liver functions (i.e., citric acid cycle, fatty acid cycle, and urea cycle), while proteins associated with stress responses, acute phase reactants, and structural components were all significantly more abundant. The expression patterns of several immunity-related proteins (peroxiredoxin 1, arginase 1, and galectin 1) suggested that different protein forms are associated with schistosome infection. These findings indicate that acute schistosomiasis has a significant impact on specific liver functions and, moreover, that the alterations in specific protein isoforms and upregulation of unique proteins may be valuable as new markers of disease.
Ribosome Depurination Is Not Sufficient for Ricin-Mediated Cell Death in
Saccharomyces cerevisiae
Authors: Li Xiao-Ping , Baricevic Marianne , Saidasan Hemalatha , Tumer Nilgun E.
Published: November 2006

The plant toxin ricin is one of the most potent and lethal substances
known. Ricin inhibits protein synthesis by removing a specific adenine
from the highly conserved α-sarcin/ricin loop in the large
rRNA. Very little is known about how ricin interacts with ribosomes and
the molecular mechanism by which it kills cells. To gain insight to the
mechanism of ricin-induced cell death, we set up yeast
(
Saccharomyces cerevisiae
) as a simple and genetically
tractable system to isolate mutants defective in cytotoxicity.
Ribosomes were depurinated in yeast cells expressing the precursor form
of the A chain of ricin (pre-RTA), and these cells displayed apoptotic
markers such as nuclear fragmentation, chromatin condensation, and
accumulation of reactive oxygen species. We conducted a large-scale
mutagenesis of pre-RTA and isolated a panel of nontoxic RTA mutants
based on their inability to kill yeast cells. Several nontoxic RTA
mutants depurinated ribosomes and inhibited translation to the same
extent as wild-type RTA in vivo. The mutant proteins isolated from
yeast depurinated ribosomes in vitro, indicating that they were
catalytically active. However, cells expressing these mutants did not
display hallmarks of apoptosis. These results provide the first
evidence that the ability to depurinate ribosomes and inhibit
translation does not always correlate with ricin-mediated cell death,
indicating that ribosome depurination and translation inhibition do not
account entirely for the cytotoxicity of
ricin.
Surface Proteins and Exotoxins Are Required for the Pathogenesis ofStaphylococcus aureusPneumonia
Authors: Wardenburg Juliane Bubeck , Patel Ravi J. , Schneewind Olaf
Published: November 2006

A model ofStaphylococcus aureus-induced pneumonia in adult, immunocompetent C57BL/6J mice is described. This model closely mimics the clinical and pathological features of pneumonia in human patients. Using this system, we defined a role forS. aureusstrain Newman surface proteins and secreted exotoxins in pneumonia-related mortality.
Two Distinct Cytotoxic Activities of Subtilase Cytotoxin Produced by Shiga-Toxigenic
Escherichia coli
Authors: Morinaga Naoko , Yahiro Kinnosuke , Matsuura Gen , Watanabe Masaharu , Nomura Fumio , Moss Joel , Noda Masatoshi
Published: November 2006

Subtilase cytotoxin (SubAB) is a recently identified AB5 subunit toxin produced by Shiga-toxigenic
Escherichia coli.
The A subunit is thought to be a subtilase-like, serine protease, whereas the B subunit binds to the toxin receptor on the cell surface. We cloned the genes from a clinical isolate; the toxin was produced as His-tagged proteins. SubAB induced vacuolation at concentrations greater than 1 μg/ml after 8 h, in addition to the reported cytotoxicity induced at a ng/ml level after 48 h. Vacuolation was induced with the B, but not the A, subunit and was dependent on V-type ATPase. The cytotoxicity of SubAB at low concentrations was associated with the inhibition of protein synthesis; the 50% inhibitory dose was ∼1 ng/ml. The A subunit, containing serine 272, which is thought to be a part of the catalytic triad of a subtilase-like serine protease, plus the B subunit was necessary for this activity, both in vivo and in vitro. SubAB did not cleave azocasein, bovine serum albumin, ovalbumin, or synthetic peptides. These data suggest that SubAB is a unique AB toxin: first, the B subunit alone can induce vacuolation; second, the A subunit containing serine 272 plus the B subunit inhibited protein synthesis, both in vivo and in vitro; and third, the A subunit proteolytic activity may have a strict range of substrate specificity.
Cell Wall Targeting of Laccase of
Cryptococcus neoformans
during Infection of Mice
Authors: Waterman Scott R. , Hacham Moshe , Panepinto John , Hu Guowu , Shin Soowan , Williamson Peter R.
Published: November 2006

Laccase is a major virulence factor of the pathogenic fungus
Cryptococcus neoformans
, which afflicts both immunocompetent and immunocompromised individuals. In the present study, laccase was expressed in
C. neoformans lac1
Δ cells as a fusion protein with an N-terminal green fluorescent protein (GFP) using
C. neoformans
codon usage. The fusion protein was robustly localized to the cell wall at physiological pH, but it was mislocalized at low pH. Structural analysis of the laccase identified a C-terminal region unique to
C. neoformans
, and expression studies showed that the region was required for efficient transport to the cell wall both in vitro and during infection of mouse lungs. During infection of mice, adherence to alveolar macrophages was also associated with a partial mislocalization of GFP-laccase within cytosolic vesicles. In addition, recovery of cryptococcal cells from lungs of two strains of mice (CBA/J and Swiss Albino) later in infection was also associated with cytosolic mislocalization, but cells from the brain showed almost exclusive localization to cell walls, suggesting that there was more efficient cell wall targeting during infection of the brain. These data suggest that host cell antifungal defenses may reduce effective cell wall targeting of laccase during infection of the lung but not during infection of the brain, which may contribute to a more predominant role for the enzyme during infection of the brain.
Heterologous Priming-Boosting with DNA and Modified Vaccinia Virus Ankara Expressing Tryparedoxin Peroxidase Promotes Long-Term Memory againstLeishmania majorin Susceptible BALB/c Mice
Authors: Stober Carmel B. , Lange Uta G. , Roberts Mark T. M. , Alcami Antonio , Blackwell Jenefer M.
Published: November 2006

Leishmaniasis affects 12 million people, but there are no vaccines in routine clinical use. Th1 polarizing vaccines that elicit long-term protection are required to prevent disease in susceptible populations. We recently showed that heterologous priming-boosting with tryparedoxin peroxidase (TRYP) DNA followed by TRYP-modified vaccinia virus Ankara (TRYP MVA) protected susceptible BALB/c mice fromLeishmania major. Here we compared treatment with TRYP DNA with treatment with TRYP DNA/TRYP MVA. We found that equivalent levels of protection during the postvaccination effector phase correlated with equivalent levels of serum immunoglobulin G2a and gamma interferon (IFN-γ) in draining lymph nodes. In contrast, challenge infection during the memory phase revealed that there was enhanced clinical efficacy with TRYP DNA/TRYP MVA. This correlated with higher levels of effector phase splenic IFN-γ, sustained prechallenge levels of memory phase IFN-γ, and a more polarized post-L. majorchallenge Th1 response compared to the Th2/Tregresponse. Thus, TRYP DNA/TRYP MVA, but not TRYP DNA alone, provides long-term protection against murine leishmaniasis.
Trefoil Family Factor 2 Is Expressed in Murine Gastric and Immune Cells and Controls both Gastrointestinal Inflammation and Systemic Immune Responses
Authors: Kurt-Jones Evelyn A. , Cao LuCheng , Sandor Frantisek , Rogers Arlin B. , Whary Mark T. , Nambiar Prashant R. , Cerny Anna , Bowen Glennice , Yan Jing , Takaishi Shigeo , Chi Alfred L. , Reed George , Houghton JeanMarie , Fox James G. , Wang Timothy C.
Published: November 2006

Trefoil family factor 2 (TFF2), also known as spasmolytic peptide, is a low-molecular-weight protein that is upregulated in gastric tissues infected with
Helicobacter
or having other inflammatory conditions, but a precise function is yet to be elucidated. The role of TFF2 in the development of gastritis, colitis, and inflammatory cytokine responses was examined both in vivo and in vitro using wild-type and TFF2 knockout mice. TFF2 knockout and wild-type mice were infected with
Helicobacter felis (H. felis
) to induce gastritis. Colitis was induced in TFF2 knockout and wild-type mice by administering dextran sodium sulfate (DSS) in drinking water. Histopathology, clinical disease (colitis), and antibody levels (
H. felis
) were examined. TFF2 expression in tissues was determined by reverse transcriptase PCR, and the inflammatory and proliferative responses of TFF2-expressing macrophages and spleen cells were examined by cytokine enzyme-linked immunosorbent assay, thymidine incorporation, and gene array studies. TFF2 knockout mice have increased susceptibility to
H. felis
-induced gastritis, with enhanced gastric inflammation. They were also more susceptible to DSS-induced colitis, with prolonged colonic hemorrhage and persistent weight loss. Remarkably, TFF2 expression was not limited to the gastrointestinal tract, as suggested in previous studies, but was also present in macrophages and lymphocytes. The inflammatory and proliferative responses of these immune cell types were dysregulated in TFF2 knockout mice. TFF2
−/−
cells were hyperresponsive to interleukin 1 beta stimulation but showed normal responses to lipopolysaccharide, suggesting a specific role for TFF2 in interleukin 1 receptor but not Toll-like receptor 4 signaling via their Toll-interleukin 1 resistance domains. TFF2
−/−
lymphocytes also produced higher levels of interleukin 2 than wild-type cells. Thus, TFF2 was expressed in the gastrointestinal cells and in immune cells and was a negative regulator of gastrointestinal inflammation and immune cell cytokine responses. Our studies suggest that TFF2 not only controls gastrointestinal repair but also regulates mononuclear cell inflammatory responses.