bluuebunny/crossref_metadata_2025 huggingface dataset:
Estimating Ground Water Flux into Large Lakes: Application in the Hamilton Harbor, Western Lake Ontario
Authors: Harvey F. Edwin , Rudolph David L. , Frape Shaun K.
Published: August 2005

Over the last two decades, there has been a growing recognition of the importance of evaluating ground water's contribution to lakes. As a result, a number of techniques have been developed for measuring hydraulic properties across lake bottoms, primarily in the littoral zone. However, for larger, deeper lakes such as the Great Lakes these techniques are impractical in the profundal zone. As a result, many water balance calculations in these settings omit the ground water component altogether owing to the difficulties encountered in making the necessary hydraulic observations across the deeper lake bottom sediments.

In this study, a methodology is developed for determining the flux into large, deep lakes using a combination of existing and recently developed techniques. The methodology is applied to the Hamilton Harbor, a natural bay at the western end of Lake Ontario, to estimate the ground water contribution to the harbor's water budget. Hydraulic gradients were monitored in 37 piezometers within the harbor during 1993 and 1994. Calculated hydraulic gradients, along with sediment hydraulic conductivities measured or estimated using a number of techniques, are used to estimate ground water flux to the harbor through Thiessen polygon weighting.

Measured hydraulic gradients ranged from −0.333 to 0.430, the majority being upward indicating ground water discharge conditions. Gradients were varied across the harbor and increased in magnitude closer to shore. The total ground water contribution to the harbor was estimated to be 2.1 × 107 m3/yr. Compared with other hydrological components, ground water was slightly larger than the yearly precipitation input, and approximately 8 % of the total surface inflows to the harbor, and 2 % of the total surface outflow through the Burlington ship canal, which connects the harbor to Lake Ontario.

The computed ground water flux suggests that despite the fact that ground water flux to large lakes may typically be a smaller input than surface water inputs, it is still a significant component of the overall water budget and should not be automatically omitted from water balance calculations.
Ground Penetrating Radar Imaging of an Aquifer During a Pumping Test
Authors: Endres Anthony L. , Clement William P. , Rudolph David L.
Published: August 2005

We have imaged the temporal and spatial response of an unconfined aquifer during a pumping test using ground penetrating radar (GPR) profiling. In particular, we have observed the transient behavior of the reflection generated by the water content variation occurring in the transition zone between the overlying residually saturated material and the water saturated capillary fringe below. The progressive arrival time delay of this reflection was measured and used to infer its drawdown that occurred during the pumping test. We also observed several other phenomena on the GPR profiles that are related to drainage: (1) the development of a series of diffractions indicating localized irregularities in water saturation and (2) the velocity pull‐up of stratigraphic reflections due to increased electromagnetic (EM) wave velocity in the overlying section.

Comparing the GPR profiling data and piezometer measurements, we have observed that the drawdown of the transition zone reflection is smaller and delayed relative to the measured hydraulic head drawdown. From the combined GPR and piezometer data, we have inferred that a 0.20 m increase in the combined thickness of the transition zone and capillary fringe occurred before the drawdown of the GPR transition zone reflection commenced. Once achieved, this 0.20 m increased thickness remained for the duration of the pumping test. Using the distance‐drawdown relationship obtained from GPR profiling, we have estimated the drained water volume due to the downward movement of the transition zone. The results of our analysis account for only a fraction of the pumping well production, approximately 45% on the first day and about 25% on the second day. Of the various reasons examined to explain this result, we have concluded that it is likely that the behavior of the GPR reflection originating from the transition zone is representative of the shallower, less saturated portion of the transition zone and undetected drainage is occurring in the deeper, more saturated portion of the transition zone.
Change in Field Turbidity and Trace Element Concentrations During Well Purging
Authors: Gibs Jacob , Szabo Zoltan , Ivahnenko Tamara , Wilde Franceska D.
Published: August 2005

Various physical and chemical properties were monitored sequentially in the field during well purging as indicators of stabilization of the composition of the water in the well. Turbidity was monitored on site during purging of oxic water from three wells with screened intervals open to an unconfined aquifer system in the Coastal Plain of southern New Jersey to determine if stabilization of turbidity is a reliable indicator of the optimum purge time required to collect unbiased trace element samples. Concurrent split (one filtered, one unfiltered) samples collected during purging of the wells were analyzed for concentrations of trace elements so that the relationships between trace element concentrations and turbidity could be compared.

Turbidity correlated with the whole water recoverable (WWR) concentration of trace element species, such as iron (Fe), aluminum (Al), and manganese (Mn) in the oxic ground water. Turbidity appeared to be independent of other field‐measured characteristics of water such as conductivity, pH, temperature, and dissolved oxygen. The WWR concentration of lead and copper, considered to be hydrophobic, correlated significantly with the sum of the WWR concentration of Fe, Al, and Mn. High values of field‐measured turbidity were a key indicator of an overestimate of ambient hydrophobic trace element WWR concentrations. Stabilization of turbidity was a better indicator of stable, unfiltered trace element concentrations than were the other commonly measured field characteristics. At one well, turbidity was a better indicator of stable, filtered trace element concentrations than the other commonly measured field characteristics. As analytical methods for trace elements improve resulting in smaller MRLs (method reporting levels) and better precision, turbidity of ground water at values of less than 10 NTU (nepheiometric turbidity units) will become important in interpreting the significance of both unfiltered and filtered sample results.
The Use of Principal Component Analysis for Interpreting Ground Water Hydrographs
Authors: Winter T.C. , Mallory S.E. , Allen T.R. , Rosenberry D.O.
Published: August 2005

Principal component analysis was used to define patterns in water table hydrographs at four small, lake‐watershed research sites in the United States. The analysis provided insights into (1) characteristics of ground water recharge in different parts of the watersheds; (2) the effect of seepage from lakes on water table fluctuations; and (3) the effect of differences in geologic properties on water table fluctuations. At two sites where all of the water table wells were completed in permeable deposits, glacial out‐wash in Minnesota and dune sand in Nebraska, the patterns of water table fluctuation primarily reflected timing and magnitude of recharge. The water table had more frequent and wider ranges in fluctuations where it was shallow compared with where it was deep. At two sites where the water table wells were completed in sand or till, a glaciated mountain valley in New Hampshire and stagnation moraine in North Dakota, the patterns of water table fluctuations were strongly related to the type of geologic unit in which the wells are completed. Furthermore, at the New Hampshire site, the patterns of water table fluctuations were clearly different for wells completed in sand downgradient of a lake compared with those completed in sandy terraces on a mountain‐side. The study indicates that principal component analysis would be particularly useful for summarizing large data sets for the purpose of selecting index wells for long‐term monitoring, which would greatly reduce the cost of monitoring programs.
Deterministic and Stochastic Modeling of Aquifer Stratigraphy, South Carolina
Authors: Miller Russell B. , Castle James W. , Temples Tom J.
Published: August 2005

Deterministic and stochastic methods of three‐dimensional hydrogeologic modeling are applied to characterization of contaminated Eocene aquifers at the Savannah River Site, South Carolina. The results address several important issues, including the use of multiple types of data in creating high‐resolution aquifer models and the application of sequence‐stratigraphic constraints. Specific procedures used include defining grid architecture stratigraphically, upscaling, modeling lithologic properties, and creating multiple equiprobable realizations of aquifer stratigraphy. An important question answered by the study is how to incorporate gamma‐ray borehole‐geophysical data in areas of anomalous log response, which occurs commonly in aquifers and confining units of the Atlantic Coastal Plain and other areas. To overcome this problem, gamma‐ray models were conditioned to grain‐size and lithofacies realizations. The investigation contributes to identifying potential pathways for downward migration of contaminants, which have been detected in confined aquifers at the modeling site.

The approach followed in this investigation produces quantitative, stratigraphically constrained, geocellular models that incorporate multiple types of data from borehole‐geophysical logs and continuous cores. The use of core‐based stochastic realizations in conditioning deterministic models provides the advantage of incorporating lithologic information based on direct observations of cores rather than using only indirect measurements from geophysical logs. The high resolution of the models is demonstrated by the representation of thin, discontinuous clay beds that act as local barriers to flow. The models are effective in depicting the contrasts in geometry and heterogeneity between sheet‐like nearshore‐transgressive sands and laterally discontinuous sands of complex shoreline environments.
Dispersivity Estimates from a Tracer Experiment in a Sandy Aquifer
Authors: Mallants Dirk , Espino Armando , Van Hoorick Marijke , Feyen Jan , Vandenberghe Noel , Loy Walter
Published: August 2005

The success or failure of transport models in predicting the migration of a contaminant plume in ground water depends to a large extent on the quality of flow and transport parameters used. In this study, we investigate the spatial variability in the tracer velocity and dispersivity in a shallow sandy aquifer in northern Belgium. Based on hydraulic conductivity measurements on cores sampled along a vertical profile, the aquifer was found to be mildly heterogeneous, i.e., with the variance of the log‐transformed conductivity K, ω2 InK, equal to 0.22. By means of a natural gradient tracer experiment, transport of a chloride tracer was investigated in a three‐dimensional network of multilevel point samplers (MLS). Least squares fitting of a two‐dimensional transport model to the individual breakthrough curves resulted in an average longitudinal dispersivity that was 10 times larger than the transverse dispersivity. The results further showed the existence of a dispersion‐scale effect whereby the depth‐averaged longitudinal dispersivity increases with increasing travel distance. The average longitudinal dispersivity corresponding to a travel distance of 10 m was equal to 0.2 m. We finally show that theoretical expressions for the macroscopic dispersivity tensor, which require input on hydraulic conductivity heterogeneity, could be used here to approximate the observed dispersive behavior. These conceptually simple models are useful to estimate macroscopic dispersivities when no tracer data are available.
Comparison of Parameter Estimation Design Criteria Using a Solute Transport Model with Matrix Diffusion
Authors: Nordqvist Rune
Published: August 2005

Three criteria for the experimental design of parameter estimation are compared by using the criteria to develop sampling patterns for determining parameter values in one‐dimensional transport models. The models include advection, dispersion, and matrix diffusion, the latter being an important process for nuclear waste isolation. The effectiveness of various designs is assessed by studying expected errors of parameter estimates and correlation between parameters. The correlations and expected errors are computed using parameter sensitivities, and the results demonstrate that the choice of design criterion can make a significant difference in the expected estimation error of parameter values in sparse designs. The best overall choice is D‐optimality, which is a measure of the volume of the joint confidence ellipsoid of the parameter estimates. E‐optimality, which minimizes the longest axis of the confidence ellipsoid, gives somewhat similar results but is not as robust for all of the estimated parameters. It is also shown that approaches that consider parameter sensitivities, but not correlation between parameters, may result in inferior designs.
Estimating In‐Situ Monod Biodegradation Parameters Using a Novel Explicit Solution of a One‐Dimensional Contaminant Transport Equation
Authors: Goudar Chetan T. , Strevett Keith A.
Published: August 2005

We present a novel, explicit closed form solution using the Lambert W function to a steady‐state one‐dimensional contaminant transport equation incorporating the Monod expression for biodegradation. This new solution overcomes some of the limitations associated with using the existing implicit solution and provides a simple method of describing biodegradation at contaminated sites using the Monod equation. The field‐scale applicability of this approach was tested using data on phenol and toluene biodegradation from previously published studies. A review of published Monod kinetic parameters for hydrocarbon biodegradation indicated that in most cases, the half saturation coefficient was lower than 5 mg/L, suggesting that the commonly used first‐order approximation is valid only at low substrate concentrations. While the Monod equation does not have this limitation and can usually provide an accurate description of biodegradation at contaminated sites, its nonlinear nature presents computational and parameter estimation difficulties compared with the linear and relatively simple first‐ and zero‐order approximations. The explicit closed form solution to the Monod equation presented in this study greatly simplifies its use and appears to be an attractive approach to describe contaminant biodegradation in the field.
Hemodynamic Signals Correlate Tightly with Synchronized Gamma Oscillations
Authors: Niessing Jörn , Ebisch Boris , Schmidt Kerstin E. , Niessing Michael , Singer Wolf , Galuske Ralf A. W.
Published: August 2005

Functional imaging methods monitor neural activity by measuring hemodynamic signals. These are more closely related to local field potentials (LFPs) than to action potentials. We simultaneously recorded electrical and hemodynamic responses in the cat visual cortex. Increasing stimulus strength enhanced spiking activity, high-frequency LFP oscillations, and hemodynamic responses. With constant stimulus intensity, the hemodynamic response fluctuated; these fluctuations were only loosely related to action potential frequency but tightly correlated to the power of LFP oscillations in the gamma range. These oscillations increase with the synchrony of synaptic events, which suggests a close correlation between hemodynamic responses and neuronal synchronization.
Protein Structures Forming the Shell of Primitive Bacterial Organelles
Authors: Kerfeld Cheryl A. , Sawaya Michael R. , Tanaka Shiho , Nguyen Chau V. , Phillips Martin , Beeby Morgan , Yeates Todd O.
Published: August 2005

Bacterial microcompartments are primitive organelles composed entirely of protein subunits. Genomic sequence databases reveal the widespread occurrence of microcompartments across diverse microbes. The prototypical bacterial microcompartment is the carboxysome, a protein shell for sequestering carbon fixation reactions. We report three-dimensional crystal structures of multiple carboxysome shell proteins, revealing a hexameric unit as the basic microcompartment building block and showing how these hexamers assemble to form flat facets of the polyhedral shell. The structures suggest how molecular transport across the shell may be controlled and how structural variations might govern the assembly and architecture of these subcellular compartments.
A Swarm of Bee Books
Authors: Hosler Jay
Published: August 2005

Robbing the Bees
. A Biography of Honey: The Sweet Liquid Gold That Seduced the World.
By Holley Bishop
. Free Press, New York, 2005. 334 pp. $24, C$35. ISBN 0-7432-5021-4.

Letters from the Hive
. An Intimate History of Bees, Honey, and Humankind.
By Stephen Buchmann with Banning Repplier
. Bantam, New York, 2005. 288 pp. $24. ISBN 0-553- 80375-1.

The Hive
. The Story of the Honeybee and Us.
By Bee Wilson
. John Murray, London, 2004. 320 pp. £14.99. ISBN 0-7195-6409-3.

An enthusiastic writer who has recently come to beekeeping; a researcher who has long worked on bees, flowers, and their ecosystems; and a historian of ideas (and food writer) offer their diverse perspectives on bees, honey, and humans.
Experimental Infection of White-Tailed Deer with
Anaplasma phagocytophilum
, Etiologic Agent of Human Granulocytic Anaplasmosis
Authors: Tate Cynthia M. , Mead Daniel G. , Luttrell M. Page , Howerth Elizabeth W. , Dugan Vivien G. , Munderloh Ulrike G. , Davidson William R.
Published: August 2005

Serologic and molecular evidence of
Anaplasma phagocytophilum
has been demonstrated in white-tailed deer (WTD;
Odocoileus virginianus
), and deer are an important host for the tick vector
Ixodes scapularis
. In this study, we describe experimental infection of WTD with
A. phagocytophilum
. We inoculated four WTD with a human isolate of
A. phagocytophilum
propagated in tick cells. Two additional deer served as negative controls. All inoculated deer developed antibodies (titers, ≥64) to
A. phagocytophilum
, as determined by an indirect fluorescent antibody test, between 14 and 24 days postinfection [p.i.]), and two deer maintained reciprocal titers of ≥64 through the end of the 66-day study. Although morulae were not observed in granulocytes and
A. phagocytophilum
was not reisolated via tick cell culture of blood, 16S reverse transcriptase nested PCR (RT-nPCR) results indicated that
A. phagocytophilum
circulated in peripheral blood of three deer through at least 17 days p.i. and was present in two deer at 38 days p.i. Femoral bone marrow from one deer was RT-nPCR positive for
A. phagocytophilum
at 66 days p.i. There was no indication of clinical disease. These data confirm that WTD are susceptible to infection with a human isolate of
A. phagocytophilum
and verify that WTD produce detectable antibodies upon exposure to the organism. Because adults are the predominant life stage of
I. scapularis
found on deer and because adult
I. scapularis
ticks do not transmit
A. phagocytophilum
transovarially, it is unlikely that WTD are a significant source of
A. phagocytophilum
for immature ticks even though deer have a high probability of natural infection. However, the susceptibility and immunologic response of WTD to
A. phagocytophilum
render them suitable candidates as natural sentinels for this zoonotic tick-borne organism.
Comparison of Five Repetitive-Sequence-Based PCR Typing Methods for Molecular Discrimination of
Salmonella
enterica
Isolates
Authors: Rasschaert G. , Houf K. , Imberechts H. , Grijspeerdt K. , De Zutter L. , Heyndrickx M.
Published: August 2005

Five repetitive-element PCR (rep-PCR) techniques [primer sets ERIC1R-ERIC2 and REP1R-REP2I and primers ERIC2, BOXA1R,
and (GTG)
5
] were evaluated for the discrimination of
Salmonella enterica
isolates at the serotype level. On the basis of number, even distribution over the whole fingerprint, and clarity of bands in the fingerprints, the enterobacterial repetitive intergenic consensus (ERIC) primer set and the (GTG)
5
primer
were chosen for use in the following experiments. For these two primer
sets, reproducibility was tested on different lysates of five selected
serotypes of
Salmonella
in the same PCR by using three
different PCR runs. Reproducibility was poor between different PCR runs
but high within the same PCR run. Furthermore, 80 different serotypes
and five isolates which were not typeable by serotyping were
fingerprinted. All strains were typeable by the ERIC primer set and the
(GTG)
5
primer and generated unique fingerprints, except for
some strains with incomplete antigenic codes. Finally, 55 genetically
different strains belonging to 10 serotypes were fingerprinted to
examine the genetic diversity of the rep-PCR within serotypes. This
experiment showed that one serotype did not always correlate to only
one ERIC or (GTG)
5
fingerprint but that the fingerprint
heterogeneity within a serotype was limited. In epidemiological
studies, ERIC- and/or (GTG)
5
-PCR can be used to limit the
number of strains that have to be serotyped. The reproducibility of
isolates in one PCR run, the discriminatory power, and the genetic
diversity (stability) of the fingerprint were similar for the Eric
primer set and the (GTG)
5
primer, so both are equally able
to discriminate
Salmonella
serotypes.
Genotype Distribution and Molecular Epidemiology of Hepatitis C Virus in Blood Donors from Southeast France
Authors: Cantaloube Jean-François , Gallian Pierre , Attoui Houssam , Biagini Philippe , De Micco Philippe , de Lamballerie Xavier
Published: August 2005

The genotype distribution of hepatitis C virus (HCV) in blood donors from southeast France was tracked for a period of 13 years (1991 to 2003). Virus genomes from 321 samples were analyzed by amplification and sequencing of the NS5b and E1 regions. The most frequent genotypes were 1b (30.2%), 1a (27.7%), and 3a (22.4%). Although it was less common, genotype 2 was characterized by the presence of strains belonging to 11 different subtypes, including 5 that had never been characterized. Genotypes 1a, 1b, 3a, and 4a presented typical “epidemic” profiles, with a large number of isolates per subtype and short mean genetic distances between isolates. Type 2 isolates displayed a typical “endemic” profile, with a large number of subtypes and very few isolates in each subtype. The epidemiology of HCV infection in southeast France changed radically during the study period in relation to modifications in the etiology of infection. We observed the emergence of new epidemic subtypes (subtypes 1a and 3) linked to intravenous drug use and a decrease in the types linked to blood transfusion and nosocomial infection (epidemic subtype 1b and endemic type 2). Comparison of strains from blood donors with strains from a cohort of inpatients in the same region during 2001 and 2002 demonstrated for the first time that the monitoring of blood donors is a generally valid indicator of HCV epidemiology in terms of genotype distribution.
Etiology of Diarrhea in Young Children in Denmark: a Case-Control Study
Authors: Olesen Bente , Neimann Jacob , Böttiger Blenda , Ethelberg Steen , Schiellerup Peter , Jensen Charlotte , Helms Morten , Scheutz Flemming , Olsen Katharina E. P. , Krogfelt Karen , Petersen Eskild , Mølbak Kåre , Gerner-Smidt Peter
Published: August 2005

Infectious gastroenteritis is one of the most common diseases in young children. To clarify the infectious etiology of diarrhea in Danish children less than 5 years of age, we conducted a 2-year prospective case-control study. Stools from 424 children with diarrhea and 870 asymptomatic age-matched controls were examined, and their parents were interviewed concerning symptoms. Rotavirus, adenovirus, and astrovirus were detected by enzyme-linked immunosorbent assay, and norovirus and sapovirus were detected by PCR.
Salmonella
, thermotolerant
Campylobacter
,
Yersinia
,
Shigella
, and
Vibrio
spp. were detected by standard methods. Shiga toxin-producing (STEC), attaching-and-effacing (A/EEC), enteropathogenic (EPEC), enterotoxigenic, enteroinvasive, and enteroaggregative
Escherichia coli
were detected by using colony hybridization with virulence gene probes and serotyping. Parasites were detected by microscopy. Overall, a potential pathogen was found in 54% of cases. More cases than controls were infected with rotavirus,
Salmonella
, norovirus, adenovirus,
Campylobacter
, sapovirus, STEC, classical EPEC,
Yersinia
, and
Cryptosporidium
strains, whereas A/EEC, although common, was not associated with illness. The single most important cause of diarrhea was rotavirus, which points toward the need for a childhood vaccine for this pathogen, but norovirus, adenovirus, and sapovirus were also major etiologies.
Salmonella
sp. was the most common bacterial pathogen, followed by
Campylobacter
, STEC,
Yersinia
, and classical EPEC strains. A/EEC not belonging to the classical EPEC serotypes was not associated with diarrhea, underscoring the importance of serotyping for the definition of EPEC.
Identification of Immunodominant Epitopes on the Membrane Protein of the Severe Acute Respiratory Syndrome-Associated Coronavirus
Authors: He Yuxian , Zhou Yusen , Siddiqui Pamela , Niu Jinkui , Jiang Shibo
Published: August 2005

Similar to other coronaviruses, the membrane (M) protein of severe acute respiratory syndrome-associated coronavirus (SARS-CoV) is a major transmembrane glycoprotein with multiple biological functions. To date, limited information is available about its antigenic properties. In this study, we identified two major immunodominant epitopes on the M protein located in the extreme N-terminal region (residues 1 to 31) and the interior C-terminal region (residues 132 to 161), respectively, by Pepscan analyses against convalescent-phase sera from SARS patients and antisera from virus-immunized mice and rabbits. Synthetic peptides M1-31 derived from the N-terminal epitope and M132-161 derived from the C-terminal epitope were highly reactive with all of the convalescent-phase sera from 40 SARS patients but not with 30 control serum samples from healthy blood donors, suggesting their potential application for serologic diagnosis of SARS. We showed that both peptides (M1-31 and M132-161) were able to induce high titers of antibody responses in the immunized rabbits, highlighting their antigenicity and immunogenicity. These findings provide important information for developing SARS diagnostics and vaccines.
Molecular Subtyping of
Treponema pallidum
from North and South Carolina
Authors: Pope Victoria , Fox Kimberley , Liu Hsi , Marfin Anthony A. , Leone Peter , Seña Arlene C. , Chapin Johanna , Fears Martha B. , Markowitz Lauri
Published: August 2005

Patients from five clinics in North and South Carolina who had lesions suggestive of primary or secondary syphilis were evaluated using molecular techniques that allow the differentiation of
Treponema pallidum
strains on the basis of two variable genes,
tpr
and
arp
. Lesion samples were screened for the presence of
T. pallidum
DNA using PCR for
polA
, which represents a segment of the polymerase I gene that is unique to the spirochete. Twenty-seven of 154 lesion samples were found to contain
T. pallidum
, 23 of which had typeable DNA. Seven molecular subtypes were found (10f, 12f, 13f, 14f, 14g, 15f, and 16f); one to four subtypes were identified at each clinic. Subtype 14f was found in 52% of the typeable specimens and was distributed in four of the five clinics. Subtype 16f was found in 22% of specimens and was concentrated at one clinic. Further data are needed to define the role of this technique in examining the epidemiology of syphilis.
International and Multicenter Comparison of EUCAST and CLSI M27-A2 Broth Microdilution Methods for Testing Susceptibilities of
Candida
spp. to Fluconazole, Itraconazole, Posaconazole, and Voriconazole
Authors: Espinel-Ingroff A. , Barchiesi F. , Cuenca-Estrella M. , Pfaller M. A. , Rinaldi M. , Rodriguez-Tudela J. L. , Verweij P. E.
Published: August 2005

The aim of this study was to compare MICs of fluconazole, itraconazole, posaconazole, and voriconazole obtained by the European Committee on Antibiotic Susceptibility Testing (EUCAST) and CLSI (formerly NCCLS) methods in each of six centers for 15
Candida albicans
(5 fluconazole-resistant and 4 susceptible-dose-dependent [S-DD] isolates), 10
C. dubliniensis
, 7
C. glabrata
(2 fluconazole-resistant isolates), 5
C. guilliermondii
(2 fluconazole-resistant isolates), 10
C. krusei
, 9
C. lusitaniae
, 10
C. parapsilosis
, and 5
C. tropicalis
(1 fluconazole-resistant isolate) isolates. CLSI MICs were obtained visually at 24 and 48 h and spectrophotometric EUCAST MICs at 24 h. The agreement (within a 3-dilution range) between the methods was species, drug, and incubation time dependent and due to lower EUCAST than CLSI MICs: overall, 94 to 95% with fluconazole and voriconazole and 90 to 91% with posaconazole and itraconazole when EUCAST MICs were compared against 24-h CLSI results. The agreement was lower (85 to 94%) against 48-h CLSI endpoints. The overall interlaboratory reproducibility by each method was ≥92%. When the comparison was based on CLSI breakpoint categorization, the agreement was 68 to 76% for three of the four species that included fluconazole-resistant and S-DD isolates; 9% very major discrepancies (≤8 μg/ml versus ≥64 μg/ml) were observed among fluconazole-resistant isolates and 50% with voriconazole (≤1 μg/ml versus ≥4 μg/ml). Similar results were observed with itraconazole for seven of the eight species evaluated (28 to 77% categorical agreement). Posaconazole EUCAST MICs were also substantially lower than CLSI MIC modes (0.008 to 1 μg/ml versus 1 to ≥8 μg/ml) for some of these isolates. Therefore, the CLSI breakpoints should not be used to interpret EUCAST MIC data.
Bifidobacterial Lipoglycan as a New Cause for False-Positive Platelia
Aspergillus
Enzyme-Linked Immunosorbent Assay Reactivity
Authors: Mennink-Kersten Monique A. S. H. , Ruegebrink Dorien , Klont Rocus R. , Warris Adilia , Gavini Françoise , Op den Camp Huub J. M. , Verweij Paul E.
Published: August 2005

We previously hypothesized that a lipoglycan of
Bifidobacterium bifidum
subsp.
pennsylvanicum
cross-reacts with the Platelia
Aspergillus
(PA) enzyme-linked immunosorbent assay (ELISA) based on the presence of galactofuranosyl epitopes in the cell wall (M. A. S. H. Mennink-Kersten, R. R. Klont, A. Warris, H. J. M. Op den Camp, and P. E. Verweij, Lancet 363:325-327, 2004). We tested this hypothesis by testing bacterial suspensions of different bifidobacterial species and other gram-positive and -negative bacteria with the PA ELISA, which is used to detect circulating galactomannan for the serodiagnosis of invasive aspergillosis. Furthermore, neonatal fecal samples were enumerated for bifidobacteria by fluorescence in situ hybridization (FISH) and tested for PA ELISA reactivity. All bifidobacteria, except
B. infantis
and
B. adolescentis
, showed reactivity 6- to 600-fold higher compared to the controls (i.e.,
Micrococcus luteus
and
Propionibacterium freudenreichii
, which contain a cell wall lipomannan).
Eggerthella lenta
showed a 25-fold-higher reactivity. ELISA reactivity was clearly shown to be associated with bacterial lipoglycans containing a β-1,5-galactofuranosyl chain. All neonatal feces showed PA ELISA reactivity and associated numbers of bifidobacteria. Since high concentrations of bifidobacteria are present in the human gut, these bacteria or excreted lipoglycan may cause false serum PA ELISA reactivity in selected patient groups, especially neonates.
In Vitro Cultivation of a ZoonoticBabesiasp. Isolated from Eastern Cottontail Rabbits (Sylvilagus floridanus) on Nantucket Island, Massachusetts
Authors: Holman Patricia J. , Spencer Angela M. , Droleskey Robert E. , Goethert Heidi K. , Telford Samuel R.
Published: August 2005

ABabesiasp. found in eastern cottontail rabbits (Sylvilagus floridanus) on Nantucket Island, Massachusetts, is the same organism that caused human babesiosis in Missouri and Kentucky, on the basis of morphology and identical small-subunit rRNA (SSU rRNA) gene sequences. Continuous cultures of the rabbit parasite were established from infected blood samples collected from two cottontail rabbits livetrapped on Nantucket Island. HL-1 medium or minimal essential medium alpha medium supplemented with 20% human serum best supported in vitro propagation of the parasite in human or cottontail erythrocytes, respectively. Parasite growth was not sustained in domestic-rabbit erythrocytes or in medium supplemented with domestic-rabbit serum. The cultured parasites were morphologically indistinguishable from the Kentucky human isolate. Transmission electron microscopy revealed similar fine structures of the parasite regardless of the host erythrocyte utilized in the cultures. Two continuous lines of the zoonoticBabesiasp. were established and confirmed to share identical SSU rRNA gene sequences with each other and with the Missouri and Kentucky humanBabesiaisolates.
Molecular Evolutionary History of Tubercle Bacilli Assessed by Study of the Polymorphic Nucleotide within the Nitrate Reductase (
narGHJI
) Operon Promoter
Authors: Goh Khye Seng , Rastogi Nalin , Berchel Mylène , Huard Richard C. , Sola Christophe
Published: August 2005

A well-characterized collection of
Mycobacterium tuberculosis
complex (MTC) isolates, representing all known subspecies as well as some relevant genotypic families of
M. tuberculosis
, was analyzed for the newly discovered
narGHJI
−215 C-to-T promoter single-nucleotide polymorphism (SNP). This point mutation has been shown in earlier studies to be responsible for the differential nitrate reductase activity of
M. tuberculosis
versus
M. bovis
. As previously defined by the presence or the absence of the TbD1 genetic locus, the group included both the “modern” W-Beijing, Haarlem, and Central-Asian1 (CAS1) families as well as the “ancestral” East-African-Indian (EAI) clade. Interestingly, among “modern”
M. tuberculosis
isolates, those previously classified as Principal Genetic Group 1 (PGG1) organisms by
katG

463

-gyrA

95

polymorphism analysis did not present the two-banded
narGHJI
restriction fragment length polymorphism analysis of PCR products pattern common to the other PGG1 MTC members, including the “ancestral”
M. tuberculosis
isolates. Instead, they showed a one-banded pattern, aligning them with other evolutionarily recent
M. tuberculosis
isolates of the PGG2 and PGG3 groups, such as Haarlem, Latin-American and Mediterranean (LAM), and X families. The presence of a nitrate reductase producer phenotype in “
Mycobacterium canettii”
and some “ancestral”
M. tuberculosis
isolates, despite a two-band −215C genotype, argues in favor of an alternate mechanism to explain the differential nitrate reductase activity of certain PGG1 subspecies of the MTC. Overall, these findings may help to establish the precise evolutionary history of important genotype families such as W-Beijing and suggest that the −215T genotype may have contributed the virulence, spread, and evolutionary success of “modern”
M. tuberculosis
strains compared to the remaining MTC organisms.
Simultaneous Visual Detection of Multiple Viral Amplicons by Dipstick Assay
Authors: Dineva Magda Anastassova , Candotti Daniel , Fletcher-Brown Fiona , Allain Jean-Pierre , Lee Helen
Published: August 2005

A sensitive, simple, and instrument-independent method for the visual detection and identification of multiple nucleic acid amplicons by dipstick has been developed. This method is based on nucleic acid hybridization on the dipstick membrane and a signal amplification system to allow visual detection. With hepatitis B virus (HBV), hepatitis C virus (HCV), and human immunodeficiency virus type 1 (HIV-1) as model analytes, it is demonstrated that the visual dipstick test combined with multiplex reverse transcription (RT)-PCR for the amplification of viral nucleic acid provides a specific and sensitive detection method. The RT-PCR products were detected by the dipstick with an efficiency similar to that of a complex, expensive, and instrument-dependent method based on fluorogenic oligonucleotide probes. The detection limits of the dipstick combined with multiplex RT-PCR were 50, 125, and 500 IU/ml for HBV DNA, HCV RNA, and HIV-1 RNA, respectively. The dipstick assay detected with similar efficiencies amplicons derived from strains of HBV genotypes A through F, HCV genotypes 1 to 6, and HIV-1 subtypes A through H as well as CRF02 circulating recombinant forms of HIV-1. Analysis of 295 clinical samples and 19 pools of 10 plasma specimens from blood donors revealed that multiplex dipstick detection was reproducible, sensitive, and specific. The visual dipstick detection of multiple amplicons thus provides an attractive alternative to complex, instrument-dependent detection methods currently in use for nucleic acid testing. This new and sensitive method for nucleic acid detection should increase the availability of genomic screening in resource-limited settings and its applicability to near-patient testing.
Value of Serological Testing for Diagnosis of Legionellosis in Outbreak Patients
Authors: Rojas Almudena , Navarro M. Dolores , Fornés Francisca E. , Serra Estefanía , Simarro Encarnación , Rojas José , Ruiz Joaquín
Published: August 2005

Serum antibody detection tests and a urine antigen detection technique were compared in samples from 116 patients epidemiologically characterized as belonging to a legionellosis outbreak. Sera were tested by enzyme-linked immunosorbent assays (ELISAs) for immunoglobulin M (IgM) and IgG plus IgM and by immunofluorescent assays (IFAs) for IgG, IgM, IgA, and polyimmunoglobulin using commercial kits (Vircell); concentrated urines were tested with the Binax NOW
Legionella
test. ELISA for IgM, ELISA for IgG plus IgM, antigenuria detection, and IFA for IgM were able to diagnose 72.3%, 60.5%, 53.3%, and 51.4%, respectively, of patients. Antigenuria was present in 53.8% of first samples, ELISA detected IgM in 29.7%, ELISA detected IgG plus IgM in 7.9%, and IFA detected IgM in 3.9%. Ten antigenuria-negative first samples tested serologically positive, 9 of them to IgM by ELISA. Despite the single source of the samples included in the study, detection of IgM using a sensitive technique such as ELISA seems to be a suitable complement to antigenuria detection for the diagnosis of legionellosis.
Novel 5-Flucytosine-Resistant Clade of
Candida dubliniensis
from Saudi Arabia and Egypt Identified by Cd25 Fingerprinting
Authors: Al Mosaid Asmaa , Sullivan Derek J. , Polacheck Itzhack , Shaheen Faisal A. , Soliman Osama , Al Hedaithy Saleh , Al Thawad Sahar , Kabadaya Motaz , Coleman David C.
Published: August 2005

DNA fingerprinting of
Candida dubliniensis
isolates using the species-specific probe Cd25 previously showed that this species consists of two distinct groups, termed Cd25 group I and Cd25 group II. The present study investigated the population structure of 30
C. dubliniensis
oral isolates from Saudi Arabia and Egypt using Cd25 fingerprinting and rRNA gene internal transcribed spacer region-based genotyping. Cd25 fingerprinting analysis of these isolates revealed two distinct populations, the first of which consisted of 10 closely related genotype 1 isolates (average similarity coefficient [
S
AB
] value, 0.86). The second population of 20 isolates was much more heterogeneous (average
S
AB
value, 0.35) and consisted of two distinct subpopulations, one of which consisted of genotype 3 isolates (
n
= 13) and the other of genotype 4 isolates (
n
= 7). A mixed dendrogram generated from the fingerprint data from the 30 Saudi Arabian and Egyptian isolates, 5 Israeli isolates, and 51 previously characterized international isolates (32 of Cd25 group I and 19 of Cd25 group II) revealed the presence of three distinct main clades. The first corresponded to the previously described Cd25 group I and contained all the Saudi Arabian, Egyptian, and Israeli genotype 1 isolates mixed with international isolates. The second clade corresponded to the previously described Cd25 group II and contained three Israeli isolates, one genotype 2 isolate, one genotype 3 isolate, and a genotype 4 variant isolate, which were mixed with international isolates. The third clade has not been described before and consisted solely of the 20 Saudi Arabian and Egyptian genotype 3 and 4 isolates identified in this study and a previously described genotype 4 Israeli isolate. All 20 Cd25 group III isolates exhibited high-level resistance to 5-flucytosine (MIC ≥ 128 μg/ml), whereas all Cd25 group I and Cd25 group II isolates tested (10 Saudi Arabian and Egyptian, 16 Israeli, and 24 international) were susceptible to 5-flucytosine (MIC ≤ 0.125 μg/ml). The results of this study show for the first time the presence of a novel 5-flucytosine-resistant clade of
C. dubliniensis
(Cd25 group III) that is predominant among isolates from Saudi Arabia and Egypt and absent from a previously characterized international collection of 98 isolates from 15 countries.
Evaluation of the ReSSQ Assay in Relation to the COBAS AMPLICOR CMV MONITOR Test and an In-House Nested PCR Method for Detection of Cytomegalovirus DNA
Authors: Wirgart Benita Zweygberg , Andersson Pia , Grillner Lena
Published: August 2005

The ReSSQ CMV assay is a novel commercially available kit for quantification of cytomegalovirus (CMV), based on real-time PCR with a peptide nucleic acid probe coupled with a single dye. In combination with the LightCycler, the ReSSQ CMV assay was evaluated with respect to specificity, PCR inhibition, linearity, reproducibility, and sensitivity. All nontested CMV materials were negative, and the assay was not inhibited by the use of different anticoagulants or other factors that may influence blood samples. The dynamic range was between 10 and 5 × 10
8
copies/PCR, and intra- and interassay variabilities were below 0.10 and 0.12 log
10
standard deviations, respectively. Assay sensitivity was validated by analysis of 24 samples from a proficiency panel and by comparison to a nested in-house CMV PCR and the COBAS AMPLICOR CMV MONITOR test, using 159 clinical samples. Results from the proficiency panel were well in accordance with input values over the entire range of viral concentrations tested (50 to 31,250 copies/ml). The association between the ReSSQ CMV assay and the in-house PCR was in agreement in 90% of the clinical samples, and discordant results were found for all types of sample materials analyzed. The ReSSQ CMV and COBAS AMPLICOR assays showed no significant differences for samples containing >1,000 CMV copies/ml, but results differed to a greater extent at lower viral concentrations. The results demonstrate that the ReSSQ CMV assay is a CMV-specific, robust, and reproducible method and hence is well suited for routine use in clinical virology laboratories.
Use of a Novel Panel of Nine Short Tandem Repeats for Exact and High-Resolution Fingerprinting of
Aspergillus fumigatus
Isolates
Authors: de Valk Hanneke A. , Meis Jacques F. G. M. , Curfs Ilse M. , Muehlethaler Konrad , Mouton Johan W. , Klaassen Corné H. W.
Published: August 2005

Here we describe a new panel of short tandem repeats (STRs) for a novel exact typing assay that can be used to discriminate between
Aspergillus fumigatus
isolates. A total of nine STR markers were selected from available genomic
A. fumigatus
sequences and were divided into three multicolor multiplex PCRs. Each multiplex reaction amplified three di-, tri-, or tetranucleotide repeats, respectively. All nine STR markers were used to analyze 100 presumably unrelated
A. fumigatus
isolates. For each marker, between 11 and 37 alleles were found in this population. One isolate proved to be a mixture of at least two different isolates. With the remaining 99 isolates, 96 different fingerprinting profiles were obtained. The Simpson's diversity index for the individual markers ranged from 0.77 to 0.97. The diversity index for the multiplex combination of di-, tri-, and tetranucleotide repeats ranged from 0.9784 to 0.9968. The combination of all nine markers yielded a Simpson's diversity index of 0.9994, indicative of the high discriminatory power of these new loci. In theory, this panel of markers is able to discriminate between no less than 27 × 10
9
different genotypes. The multicolor multiplex approach allows large numbers of markers to be tested in a short period of time. The exact nature of the assay combines high reproducibility with the easy exchange of results and makes it a very suitable tool for large-scale epidemiological studies.
Simultaneous Detection of Four Human Pathogenic Microsporidian Species from Clinical Samples by Oligonucleotide Microarray
Authors: Wang Zheng , Orlandi Palmer A. , Stenger David A.
Published: August 2005

Microsporidian species have been rapidly emerging as human enteric pathogens in immunocompromised and immunocompetent individuals in recent years. Routine diagnostic techniques for microsporidia in clinical laboratories are laborious and insensitive and tend to underestimate their presence. In most instances, they are unable to differentiate species of spores due to their small sizes and similar morphologies. In this study, we report the development of another protozoan oligonucleotide microarray assay for the simultaneous detection and identification to the species level of four major microsporidian species:
Enterocytozoon bieneusi
,
Encephalitozoon cuniculi
,
Encephalitozoon hellem
, and
Encephalitozoon intestinalis
. The 18S small-subunit rRNA gene was chosen as the amplification target, labeled with fluorescence dye, and hybridized to a series of species-specific oligonucleotide probes immobilized on a microchip. The specificity and sensitivity of the microarray were clearly demonstrated by the unique hybridization profiles exhibited by each species of microsporidian tested and its ability to detect as few as 10 spores. In order to assess the applicability of this microarray in a clinical setting, we conducted microarray assays of 20 fecal samples from AIDS patients. Twelve of these samples were positive for the presence of microsporidia and could be confidently identified; 11 of them were positive for more than one species. Our results suggested that this microarray-based approach represents an attractive diagnostic tool for high-throughput detection and identification of microsporidian species in clinical and epidemiological investigations.