bluuebunny/crossref_metadata_2025 huggingface dataset:
MAP Kinase Phosphatase As a Locus of Flexibility in a Mitogen-Activated Protein Kinase Signaling Network
Authors: Bhalla Upinder S. , Ram Prahlad T. , Iyengar Ravi
Published: October 2002

Intracellular signaling networks receive and process information to control cellular machines. The mitogen-activated protein kinase (MAPK) 1,2/protein kinase C (PKC) system is one such network that regulates many cellular machines, including the cell cycle machinery and autocrine/paracrine factor synthesizing machinery. We used a combination of computational analysis and experiments in mouse NIH-3T3 fibroblasts to understand the design principles of this controller network. We find that the growth factor–stimulated signaling network containing MAPK 1, 2/PKC can operate with one (monostable) or two (bistable) stable states. At low concentrations of MAPK phosphatase, the system exhibits bistable behavior, such that brief stimulus results in sustained MAPK activation. The MAPK-induced increase in the amounts of MAPK phosphatase eliminates the prolonged response capability and moves the network to a monostable state, in which it behaves as a proportional response system responding acutely to stimulus. Thus, the MAPK 1, 2/PKC controller network is flexibly designed, and MAPK phosphatase may be critical for this flexible response.
Helicobacter pylori
SabA Adhesin in Persistent Infection and Chronic Inflammation
Authors: Mahdavi Jafar , Sondén Berit , Hurtig Marina , Olfat Farzad O. , Forsberg Lina , Roche Niamh , Ångström Jonas , Larsson Thomas , Teneberg Susann , Karlsson Karl-Anders , Altraja Siiri , Wadström Torkel , Kersulyte Dangeruta , Berg Douglas E. , Dubois Andre , Petersson Christoffer , Magnusson Karl-Eric , Norberg Thomas , Lindh Frank , Lundskog Bertil B. , Arnqvist Anna , Hammarström Lennart , Borén Thomas
Published: October 2002

Helicobacter pylori
adherence in the human gastric mucosa involves specific bacterial adhesins and cognate host receptors. Here, we identify sialyl-dimeric-Lewis x glycosphingolipid as a receptor for
H
.
pylori
and show that
H
.
pylori
infection induced formation of sialyl-Lewis x antigens in gastric epithelium in humans and in a Rhesus monkey. The corresponding sialic acid–binding adhesin (SabA) was isolated with the “retagging” method, and the underlying
sabA
gene (JHP662/HP0725) was identified. The ability of many
H
.
pylori
strains to adhere to sialylated glycoconjugates expressed during chronic inflammation might thus contribute to virulence and the extraordinary chronicity of
H
.
pylori
infection.
Single-Cell Gene Expression Profiling
Authors: Levsky Jeffrey M. , Shenoy Shailesh M. , Pezo Rossanna C. , Singer Robert H.
Published: October 2002

A key goal of biology is to relate the expression of specific genes to a particular cellular phenotype. However, current assays for gene expression destroy the structural context. By combining advances in computational fluorescence microscopy with multiplex probe design, we devised technology in which the expression of many genes can be visualized simultaneously inside single cells with high spatial and temporal resolution. Analysis of 11 genes in serum-stimulated cultured cells revealed unique patterns of gene expression within individual cells. Using the nucleus as the substrate for parallel gene analysis, we provide a platform for the fusion of genomics and cell biology: “cellular genomics.”
Role of Genotype in the Cycle of Violence in Maltreated Children
Authors: Caspi Avshalom , McClay Joseph , Moffitt Terrie E. , Mill Jonathan , Martin Judy , Craig Ian W. , Taylor Alan , Poulton Richie
Published: October 2002

We studied a large sample of male children from birth to adulthood to determine why some children who are maltreated grow up to develop antisocial behavior, whereas others do not. A functional polymorphism in the gene encoding the neurotransmitter-metabolizing enzyme monoamine oxidase A (
MAOA
) was found to moderate the effect of maltreatment. Maltreated children with a genotype conferring high levels of
MAOA
expression were less likely to develop antisocial problems. These findings may partly explain why not all victims of maltreatment grow up to victimize others, and they provide epidemiological evidence that genotypes can moderate children's sensitivity to environmental insults.
Selection and Amplification of Hosts From Dynamic Combinatorial Libraries of Macrocyclic Disulfides
Authors: Otto Sijbren , Furlan Ricardo L. E. , Sanders Jeremy K. M.
Published: October 2002

We have discovered two receptors for two different guests from a single dynamic combinatorial library. Each of the two guests amplifies the formation of a tightly binding host at the expense of unfit library members. Small differences in host-guest binding translate into useful differences in amplification. The selected hosts could be readily synthesized using biased dynamic libraries that contain only the right ratio of those building blocks that were selected by the guests. These results establish dynamic combinatorial chemistry as a practical method not only for the discovery but also for the synthesis of new receptors.
Foraminiferal Calcification Response to Glacial-Interglacial Changes in Atmospheric CO
2
Authors: Barker Stephen , Elderfield Henry
Published: October 2002

A record of foraminiferal shell weight across glacial-interglacial Termination I shows a response related to seawater carbonate ion concentration and allows reconstruction of a record of carbon dioxide in surface seawater that matches the atmospheric record. The results support suggestions that higher atmospheric carbon dioxide directly affects marine calcification, an effect that may be of global importance to past and future changes in atmospheric CO
2
. The process provides negative feedback to the influence of marine calcification on atmospheric carbon dioxide and is of practical importance to the application of paleoceanographic proxies.
βAR Signaling Required for Diet-Induced Thermogenesis and Obesity Resistance
Authors: Bachman Eric S. , Dhillon Harveen , Zhang Chen-Yu , Cinti Saverio , Bianco Antonio C. , Kobilka Brian K. , Lowell Bradford B.
Published: October 2002

Excessive caloric intake is thought to be sensed by the brain, which then activates thermogenesis as a means of preventing obesity. The sympathetic nervous system, through β-adrenergic receptor (βAR) action on target tissues, is likely the efferent arm of this homeostatic mechanism. To test this hypothesis, we created mice that lack the three known βARs (β-less mice). β-less mice on a Chow diet had a reduced metabolic rate and were slightly obese. On a high-fat diet, β-less mice, in contrast to wild-type mice, developed massive obesity that was due entirely to a failure of diet-induced thermogenesis. These findings establish that βARs are necessary for diet-induced thermogenesis and that this efferent pathway plays a critical role in the body's defense against diet-induced obesity.
Laminar Growth of Ultrathin Metal Films on Metal Oxides: Co on Hydroxylated α-Al
2
O
3
(0001)
Authors: Chambers S. A. , Droubay T. , Jennison D. R. , Mattsson T. R.
Published: October 2002

Metals deposited in vacuum on metal oxides such as alumina normally grow as three-dimensional clusters because of weak adatom-substrate interactions. This tendency hinders our ability to form interfaces of ultrathin, laminar metal films on oxides for use in microelectronics and other technologies where nanostructural control is desired. We present experimental and theoretical results showing that room temperature Co deposition on fully hydroxylated clean sapphire (α-Al
2
O
3
) produces a surface chemical reaction that leads to laminar growth, despite a large mismatch in lattice constants. This process should be applicable to a wide range of metals and metal oxides.
Sour Grapes of Wrath
Authors: Grant Bruce S.
Published: October 2002

Of Moths and Men
Intrigue, Tragedy and the Peppered Moth. Judith Hooper. Fourth Estate, London, 2002. 397 pp. £15.99. ISBN 1-84115-392-3.

Of Moths and Men
The Untold Story of Science and the Peppered Moth. Norton, New York, 2002. 397 pp. $26.95, C$38.99. ISBN 0-393-05121-8.

Hooper presents a popular account of
Biston betularia
and the controversies surrounding Kettlewell's experiments on the moth's response to natural selection and pollution in the United Kingdom.
A Conserved p38 MAP Kinase Pathway in
Caenorhabditis elegans
Innate Immunity
Authors: Kim Dennis H. , Feinbaum Rhonda , Alloing Geneviève , Emerson Fred E. , Garsin Danielle A. , Inoue Hideki , Tanaka-Hino Miho , Hisamoto Naoki , Matsumoto Kunihiro , Tan Man-Wah , Ausubel Frederick M.
Published: October 2002

A genetic screen for
Caenorhabditis elegans
mutants with enhanced susceptibility to killing by
Pseudomonas aeruginosa
led to the identification of two genes required for pathogen resistance:
sek-1
, which encodes a mitogen-activated protein (MAP) kinase kinase, and
nsy-1
, which encodes a MAP kinase kinase kinase. RNA interference assays and biochemical analysis established that a p38 ortholog,
pmk-1
, functions as the downstream MAP kinase required for pathogen defense. These data suggest that this MAP kinase signaling cassette represents an ancient feature of innate immune responses in evolutionarily diverse species.
A General Synthetic Procedure for Heteropolyniobates
Authors: Nyman May , Bonhomme François , Alam Todd M. , Rodriguez Mark A. , Cherry Brian R. , Krumhansl James L. , Nenoff Tina M. , Sattler Amy M.
Published: October 2002

The heteropolyanions of W, Mo, and V, which have found numerous applications, are formed simply by acidification of solutions of their oxoanions. Under similar conditions, these oxoanion precursors are not available for Nb, and Nb-oxo chemistry is dominated by formation of the Lindquist ion [Nb
6
O
19
]
8–
only. However, heteropolyniobate formation is favored in hydrothermal reactions of aqueous, alkaline precursor mixtures. Here we give two examples of heteropolyniobates formed by this general reaction type: K
12
[Ti
2
O
2
][SiNb
12
O
40
]·16H
2
O [1], which contains chains of silicododecaniobate Keggin ions, and Na
14
[H
2
Si
4
Nb
16
O
56
]·45.5H
2
O [
2
], a new heteropolyanion structure type.
Moving and Mixing
Authors: Vigilant Linda
Published: October 2002

Mapping Human History
Discovering the Past Through Our Genes. Steve Olson. Houghton Mifflin, Boston, MA, 2002. 302 pp. $25. ISBN 0-618-09157-2.

Drawing on findings from molecular studies, Olson offers a popular account of the genealogy of extant humans and the role of historical accidents in producing the current diversity of peoples.
Interfaces Between the Detection, Signaling, and Repair of DNA Damage
Authors: Rouse John , Jackson Stephen P.
Published: October 2002

Left unrepaired, the myriad types of damage that can occur in genomic DNA pose a serious threat to the faithful transmission of the correct complement of genetic material. Defects in DNA damage signaling and repair result in genomic instability, a hallmark of cancer, and often cause lethality, underlining the importance of these processes in the cell and whole organism. The past decade has seen huge advances in our understanding of how the signal transduction pathways triggered by DNA damage radically alter cell behavior. In contrast, it is still unclear how primary DNA damage is detected and how this interfaces with signal transduction and DNA repair proteins.
Identification and Analysis of Amino Acid Mutations in Porin IB That Mediate Intermediate-Level Resistance to Penicillin and Tetracycline in
Neisseria gonorrhoeae
Authors: Olesky Melanie , Hobbs Marcia , Nicholas Robert A.
Published: October 2002

PenB
is the third resistance determinant in the stepwise acquisition of multiple resistance genes in chromosomally mediated resistant
Neisseria gonorrhoeae
(CMRNG). Alterations in

por
IB

, one of two alleles at the
por
locus that encodes the outer membrane protein porin IB (PIB), were recently reported to be responsible for the increased resistance to penicillin and tetracycline conferred by
penB
, but the specific mutations conferring antibiotic resistance were not identified experimentally. To determine which amino acids in PIB confer increased resistance, we transformed a recipient strain with chimeras of the

por
IB

genes from strains FA1090 and FA140 (
penB2
). These studies revealed that two amino acid changes, G120D and A121D, were both necessary and sufficient to confer increased resistance to penicillin and tetracycline. Site-saturation and site-directed mutagenesis of Gly-120 and Ala-121 revealed that both a single mutation, G120K, and the double mutations G120R A121H and G120P A121P also conferred antibiotic resistance to the recipient strain. The identical mutations in PIA increased penicillin and tetracycline resistance either moderately or not at all. Analysis of

por
IB

genes present in the GenBank database from 51 clinical isolates demonstrated that lysine and aspartate mutations at positions 120 and/or 121 also occur in nature. These studies demonstrate that charged amino acids at positions 120 and 121 in PIB are highly preferential for conferring resistance to penicillin and tetracycline in
N. gonorrhoeae
.
Immunomodulatory Effect of Zidovudine (ZDV) on Cytotoxic T Lymphocytes Previously Exposed to ZDV
Authors: Francke Sabine , Orosz Charles G. , Hsu Jason , Mathes Lawrence E.
Published: October 2002

In a previous study, zidovudine (ZDV) was shown to cause a concentration-dependent inhibition of antigen-specific cytotoxic T-lymphocyte (CTL) clonal expansion (S. Francke, C. G. Orosz, K. A. Hayes, and L. E. Mathes, Antimicrob. Agents Chemother. 44:1900-1905, 2000). However, this suppressive effect was lost if exposure to ZDV was delayed for 24 to 48 h during the antigen sensitization period, suggesting that antigen-primed CTL may be less susceptible than naive T lymphocytes to the suppressive effects of ZDV. The present study was undertaken to determine if naive T lymphocytes were more sensitive to the suppressive effects of ZDV than T lymphocytes previously exposed to antigen. The 50% inhibitory concentration (IC50) values of ZDV were determined on naive and antigen-primed T-cell responses in an alloantigen system. Lymphocyte cultures with continuous antigen exposure (double prime) were more resistant to ZDV suppression (IC50= 316 μM) than were naive lymphocytes (IC50= 87.5 μM). Interestingly, lymphocytes that were antigen primed but deprived of antigen during the final 7 days of culture (prime/hold) were exquisitely sensitive to ZDV suppression (IC50= 29.3 μM). The addition of 80 μM ZDV during the initial priming of the single-prime (prime/hold) and double-prime cultures did not select for a more drug-resistant cell population. The differences in ZDV sensitivities are likely a reflection of the physiological properties of the lymphocytes related to their activation state.
Efficacy of Quinupristin-Dalfopristin in Preventing Vascular Graft Infection Due to
Staphylococcus epidermidis
with Intermediate Resistance to Glycopeptides
Authors: Giacometti Andrea , Cirioni Oscar , Ghiselli Roberto , Orlando Fiorenza , Mocchegiani Federico , Riva Alessandra , Del Prete Maria Simona , Saba Vittorio , Scalise Giorgio
Published: October 2002

A rat model was used to investigate the efficacy of quinupristin-dalfopristin (Q-D) in the prevention of vascular prosthetic graft infection due to methicillin-resistant
Staphylococcus epidermidis
with intermediate resistance to glycopeptides. The in vitro activity of the compound was compared to that of vancomycin by MIC determination and time-kill study. Moreover, the efficacy of collagen-sealed Q-D-soaked Dacron was evaluated in a rat model of graft infection. Graft infections were established in the subcutaneous tissue of the backs of 120 adult male Wistar rats. The in vivo study included a control group, one contaminated group that did not receive any antibiotic prophylaxis, two contaminated groups that received grafts soaked with 10 and 100 μg of Q-D per ml, respectively, and two contaminated groups that received grafts soaked with 10 and 100 μg of vancomycin per ml, respectively. Rats that received Dacron grafts soaked with 100 μg of Q-D per ml showed no evidence of infection (<10 CFU/ml). In contrast, for rats that received Dacron grafts soaked with 10 μg of Q-D per ml and Dacron grafts soaked with 10 or 100 μg of vancomycin per ml, the quantitative graft cultures demonstrated 2.2 × 10
2
± 1.3 × 10
2
, 2.2 × 10
6
± 1.9 × 10
5
, and 5.6 × 10
2
± 0.3 × 10
2
CFU/ml, respectively. Taken together the results of the study demonstrate that the use of Dacron grafts soaked with Q-D can result in significant bacterial growth inhibition and show that this compound is potentially valuable for prevention of vascular prosthetic graft infection.
16S rRNA Mutation-Mediated Tetracycline Resistance in
Helicobacter pylori
Authors: Gerrits Monique M. , de Zoete Marcel R. , Arents Niek L. A. , Kuipers Ernst J. , Kusters Johannes G.
Published: October 2002

Most
Helicobacter pylori
strains are susceptible to tetracycline, an antibiotic commonly used for the eradication of
H. pylori
. However, an increase in incidence of tetracycline resistance in
H. pylori
has recently been reported. Here the mechanism of tetracycline resistance of the first Dutch tetracycline-resistant (Tet
r
)
H. pylori
isolate (strain 181) is investigated. Twelve genes were selected from the genome sequences of
H. pylori
strains 26695 and J99 as potential candidate genes, based on their homology with tetracycline resistance genes in other bacteria. With the exception of the two 16S rRNA genes, none of the other putative tetracycline resistance genes was able to transfer tetracycline resistance. Genetic transformation of the Tet
s
strain 26695 with smaller overlapping PCR fragments of the 16S rRNA genes of strain 181, revealed that a 361-bp fragment that spanned nucleotides 711 to 1071 was sufficient to transfer resistance. Sequence analysis of the 16S rRNA genes of the Tet
r
strain 181, the Tet
s
strain 26695, and four Tet
r
26695 transformants showed that a single triple-base-pair substitution, AGA
926-928
→TTC, was present within this 361-bp fragment. This triple-base-pair substitution, present in both copies of the 16S rRNA gene of all our Tet
r
H. pylori
transformants, resulted in an increased MIC of tetracycline that was identical to that for the Tet
r
strain 181.
Prospective Survey of β-Lactamases Produced by Ceftazidime- Resistant
Pseudomonas aeruginosa
Isolated in a French Hospital in 2000
Authors: De Champs Christophe , Poirel Laurent , Bonnet Richard , Sirot Danielle , Chanal Catherine , Sirot Jacques , Nordmann Patrice
Published: October 2002

In 2000, at the Université d'Auvergne teaching hospital in Clermont-Ferrand, France, 44 (6.2%) strains of
Pseudomonas aeruginosa
were found to be resistant to ceftazidime. After genotyping, 34 strains were selected. Nine had an additional β-lactamase: OXA-21 (
n
= 6), PSE-1 (CARB-2) (
n
= 2), or PER-1 (
n
= 1). Ceftazidime resistance was related solely to the overproduction of the cephalosporinase in 30 strains. Sequencing of five
bla
AmpC
genes encoding cephalosporinases with different pIs showed 99% identity with the
ampC
gene of
P. aeruginosa
PAO1.
Clinical Isolates of
Staphylococcus aureus
with Ribosomal Mutations Conferring Resistance to Macrolides
Authors: Prunier Anne-Laure , Malbruny Brigitte , Tandé Didier , Picard Bertrand , Leclercq Roland
Published: October 2002

Six strains of
Staphylococcus aureus
isolated from cystic fibrosis patients after treatment with azithromycin were cross-resistant to azithromycin and erythromycin. None of the isolates contained
erm
or
msr
(A) genes, but they all carried either A2058G/U or A2059G mutations within the
rrl
genes, with a majority of the rRNA copies bearing the mutation. One strain displayed an additional mutation in the
rplV
gene, encoding the L22 ribosomal protein.
In Vitro and In Vivo Antibacterial Activities of DW286, a New Fluoronaphthyridone Antibiotic
Authors: Yun Hee-Jeong , Min Yu-Hong , Lim Jung-A , Kang Jin-Wook , Kim So-Young , Kim Mi-Jeong , Jeong Jae-Hee , Choi Yun-Jeong , Kwon Hyun-Jin , Jung Yong-Ho , Shim Mi-Ja , Choi Eung-Chil
Published: October 2002

The in vitro and in vivo activities of DW286, a novel fluoronaphthyridone with potent antibacterial activity, were compared with those of ciprofloxacin, gemifloxacin, sparfloxacin, and trovafloxacin. Against gram-positive bacteria, such as
Staphylococcus aureus
,
Staphylococcus epidermidis
,
Streptococcus pneumoniae
, and
Enterococcus faecalis
, the in vitro activity of DW286 was stronger than that of any other reference antibiotic. Against gram-negative bacteria, the activity of DW286 was similar to those of trovafloxacin and gemifloxacin but was weaker than that of ciprofloxacin. In a mouse systemic infection caused by three
S. aureus
strains, including methicillin-resistant
S. aureus
and quinolone-resistant
S. aureus
(QRSA), DW286 demonstrated the most potent activity, as found in vitro. Specially, DW286 is ≥8-fold more active against QRSA than the other fluoroquinolones. And the 50% protective doses for DW286 were correspondent with the in vitro activities.
Comparative Activities of the Oxazolidinone AZD2563 and Linezolid against Selected Recent North American Isolates of
Streptococcus pneumoniae
Authors: Baum Sue E. , Crawford Sharon A. , McElmeel M. L. , Whitney Cynthia G. , Jorgensen James H.
Published: October 2002

The activity of AZD2563 against 250 highly resistant pneumococci and 267 drug-susceptible isolates was determined. The AZD2563 MICs for 50 and 90% of the strains tested were 1 and 2 μg/ml and 0.5 and 1 μg/ml, respectively, for the two isolate groups. These MICs were within 1 log
2
dilution of those of linezolid.
Homologs of the Rml Enzymes from
Salmonella enterica
Are Responsible for dTDP-β-
l
-Rhamnose Biosynthesis in the Gram-Positive Thermophile
Aneurinibacillus thermoaerophilus
DSM 10155
Authors: Graninger Michael , Kneidinger Bernd , Bruno Katharina , Scheberl Andrea , Messner Paul
Published: October 2002

The glycan chains of the surface layer (S-layer) glycoprotein from the gram-positive, thermophilic bacterium
Aneurinibacillus
(formerly
Bacillus
)
thermoaerophilus
strain DSM 10155 are composed of
l
-rhamnose- and
d
-
glycero
-
d
-
manno
-heptose-containing disaccharide repeating units which are linked to the S-layer polypeptide via core structures that have variable lengths and novel O-glycosidic linkages. In this work we investigated the enzymes involved in the biosynthesis of thymidine diphospho-
l
-rhamnose (dTDP-
l
-rhamnose) and their specific properties. Comparable to lipopolysaccharide O-antigen biosynthesis in gram-negative bacteria, dTDP-
l
-rhamnose is synthesized in a four-step reaction sequence from dTTP and glucose 1-phosphate by the enzymes glucose-1-phosphate thymidylyltransferase (RmlA), dTDP-
d
-glucose 4,6-dehydratase (RmlB), dTDP-4-dehydrorhamnose 3,5-epimerase (RmlC), and dTDP-4-dehydrorhamnose reductase (RmlD). The rhamnose biosynthesis operon from
A. thermoaerophilus
DSM 10155 was sequenced, and the genes were overexpressed in
Escherichia coli
. Compared to purified enterobacterial Rml enzymes, the enzymes from the gram-positive strain show remarkably increased thermostability, a property which is particularly interesting for high-throughput screening and enzymatic synthesis. The closely related strain
A. thermoaerophilus
L420-91
T
produces
d
-rhamnose- and 3-acetamido-3,6-dideoxy-
d
-galactose-containing S-layer glycan chains. Comparison of the enzyme activity patterns in
A. thermoaerophilus
strains DSM 10155 and L420-91
T
for
l
-rhamnose and
d
-rhamnose biosynthesis indicated that the enzymes are differentially expressed during S-layer glycan biosynthesis and that
A. thermoaerophilus
L420-91
T
is not able to synthesize dTDP-
l
-rhamnose. These findings confirm that in each strain the enzymes act specifically on S-layer glycoprotein glycan formation.
Identification, Characterization, and Regulation of a Cluster of Genes Involved in Carbapenem Biosynthesis in
Photorhabdus luminescens
Authors: Derzelle Sylviane , Duchaud Eric , Kunst Frank , Danchin Antoine , Bertin Philippe
Published: October 2002

The luminescent entomopathogenic bacterium
Photorhabdus luminescens
produces several yet-uncharacterized broad-spectrum antibiotics. We report the identification and characterization of a cluster of eight genes (named
cpmA
to
cpmH
) responsible for the production of a carbapenem-like antibiotic in strain TT01 of
P. luminescens
. The
cpm
cluster differs in several crucial aspects from other
car
operons. The level of
cpm
mRNA peaks during exponential phase and is regulated by a Rap/Hor homolog identified in the
P. luminescens
genome. Marker-exchange mutagenesis of this gene in the entomopathogen decreased antibiotic production. The
luxS
-like signaling mechanism of quorum sensing also plays a role in the regulation of the
cpm
operon. Indeed,
luxS
, which is involved in the production of a newly identified autoinducer, is responsible for repression of
cpm
gene expression at the end of the exponential growth phase. The importance of this carbapenem production in the ecology of
P. luminescens
is discussed.
Comparison of In Vitro Cell Culture and a Mouse Assay for Measuring Infectivity of
Cryptosporidium parvum
Authors: Rochelle Paul A. , Marshall Marilyn M. , Mead Jan R. , Johnson Anne M. , Korich Dick G. , Rosen Jeffrey S. , De Leon Ricardo
Published: October 2002

In vitro cell cultures were compared to neonatal mice for measuring the infectivity of five genotype 2 isolates of
Cryptosporidium parvum
. Oocyst doses were enumerated by flow cytometry and delivered to animals and cell monolayers by using standardized procedures. Each dose of oocysts was inoculated into up to nine replicates of 9 to 12 mice or 6 to 10 cell culture wells. Infections were detected by hematoxylin and eosin staining in CD-1 mice, by reverse transcriptase PCR in HCT-8 and Caco-2 cells, and by immunofluorescence microscopy in Madin-Darby canine kidney (MDCK) cells. Infectivity was expressed as a logistic transformation of the proportion of animals or cell culture wells that developed infection at each dose. In most instances, the slopes of the dose-response curves were not significantly different when we compared the infectivity models for each isolate. The 50% infective doses for the different isolates varied depending on the method of calculation but were in the range from 16 to 347 oocysts for CD-1 mice and in the ranges from 27 to 106, 31 to 629, and 13 to 18 oocysts for HCT-8, Caco-2, and MDCK cells, respectively. The average standard deviations for the percentages of infectivity for all replicates of all isolates were 13.9, 11.5, 13.2, and 10.7% for CD-1 mice, HCT-8 cells, Caco-2 cells, and MDCK cells, respectively, demonstrating that the levels of variability were similar in all assays. There was a good correlation between the average infectivity for HCT-8 cells and the results for CD-1 mice across all isolates for untreated oocysts (
r
= 0.85,
n
= 25) and for oocysts exposed to ozone and UV light (
r
= 0.89,
n
= 29). This study demonstrated that in vitro cell culture was equivalent to the “gold standard,” mouse infectivity, for measuring the infectivity of
C. parvum
and should therefore be considered a practical and accurate alternative for assessing oocyst infectivity and inactivation. However, the high levels of variability displayed by all assays indicated that infectivity and disinfection experiments should be limited to discerning relatively large differences.
Association of Microbial Community Composition and Activity with Lead, Chromium, and Hydrocarbon Contamination
Authors: Shi W. , Becker J. , Bischoff M. , Turco R. F. , Konopka A. E.
Published: October 2002

Microbial community composition and activity were characterized in soil contaminated with lead (Pb), chromium (Cr), and hydrocarbons. Contaminant levels were very heterogeneous and ranged from 50 to 16,700 mg of total petroleum hydrocarbons (TPH) kg of soil
−1
, 3 to 3,300 mg of total Cr kg of soil
−1
, and 1 to 17,100 mg of Pb kg of soil
−1
. Microbial community compositions were estimated from the patterns of phospholipid fatty acids (PLFA); these were considerably different among the 14 soil samples. Statistical analyses suggested that the variation in PLFA was more correlated with soil hydrocarbons than with the levels of Cr and Pb. The metal sensitivity of the microbial community was determined by extracting bacteria from soil and measuring [
3
H]leucine incorporation as a function of metal concentration. Six soil samples collected in the spring of 1999 had IC
50
values (the heavy metal concentrations giving 50% reduction of microbial activity) of approximately 2.5 mM for CrO
4
2−
and 0.01 mM for Pb
2+
. Much higher levels of Pb were required to inhibit [
14
C]glucose mineralization directly in soils. In microcosm experiments with these samples, microbial biomass and the ratio of microbial biomass to soil organic C were not correlated with the concentrations of hydrocarbons and heavy metals. However, microbial C respiration in samples with a higher level of hydrocarbons differed from the other soils no matter whether complex organic C (alfalfa) was added or not. The ratios of microbial C respiration to microbial biomass differed significantly among the soil samples (
P
< 0.05) and were relatively high in soils contaminated with hydrocarbons or heavy metals. Our results suggest that the soil microbial community was predominantly affected by hydrocarbons.
Biodesulfurization of Naphthothiophene and Benzothiophene through Selective Cleavage of Carbon-Sulfur Bonds by
Rhodococcus
sp. Strain WU-K2R
Authors: Kirimura Kohtaro , Furuya Toshiki , Sato Rika , Ishii Yoshitaka , Kino Kuniki , Usami Shoji
Published: October 2002

Naphtho[2,1-
b
]thiophene (NTH) is an asymmetric structural isomer of dibenzothiophene (DBT), and in addition to DBT derivatives, NTH derivatives can also be detected in diesel oil following hydrodesulfurization treatment.
Rhodococcus
sp. strain WU-K2R was newly isolated from soil for its ability to grow in a medium with NTH as the sole source of sulfur, and growing cells of WU-K2R degraded 0.27 mM NTH within 7 days. WU-K2R could also grow in the medium with NTH sulfone, benzothiophene (BTH), 3-methyl-BTH, or 5-methyl-BTH as the sole source of sulfur but could not utilize DBT, DBT sulfone, or 4,6-dimethyl-DBT. On the other hand, WU-K2R did not utilize NTH or BTH as the sole source of carbon. By gas chromatography-mass spectrometry analysis, desulfurized NTH metabolites were identified as NTH sulfone, 2′-hydroxynaphthylethene, and naphtho[2,1-
b
]furan. Moreover, since desulfurized BTH metabolites were identified as BTH sulfone, benzo[
c
][1,2]oxathiin
S
-oxide, benzo[
c
][1,2]oxathiin
S
,
S
-dioxide,
o
-hydroxystyrene, 2-(2′-hydroxyphenyl)ethan-1-al, and benzofuran, it was concluded that WU-K2R desulfurized NTH and BTH through the sulfur-specific degradation pathways with the selective cleavage of carbon-sulfur bonds. Therefore,
Rhodococcus
sp. strain WU-K2R, which could preferentially desulfurize asymmetric heterocyclic sulfur compounds such as NTH and BTH through the sulfur-specific degradation pathways, is a unique desulfurizing biocatalyst showing properties different from those of DBT-desulfurizing bacteria.
Differentially Regulated, Vegetative-Mycelium-Specific Hydrophobins of the Edible Basidiomycete
Pleurotus ostreatus
Authors: Peñas María M. , Rust Brian , Larraya Luis M. , Ramírez Lucía , Pisabarro Antonio G.
Published: October 2002

Three different hydrophobins (Vmh1, Vmh2, and Vmh3) were isolated from monokaryotic and dikaryotic vegetative cultures of the edible fungus
Pleurotus ostreatus
. Their corresponding genes have a number of introns different from those of other
P. ostreatus
hydrophobins previously described. Two genes (
vmh1
and
vmh2
) were expressed only at the vegetative stage, whereas
vmh3
expression was also found in the fruit bodies. Furthermore, the expression of the three hydrophobins varied significantly with culture time and nutritional conditions. The three genes were mapped in the genomic linkage map of
P. ostreatus
, and evidence is presented for the allelic nature of
vmh2
and
POH3
and for the different locations of the genes coding for the glycosylated hydrophobins Vmh3 and POH2. The glycosylated nature of Vmh3 and its expression during vegetative growth and in fruit bodies suggest that it should play a role in development similar to that proposed for SC3 in
Schizophyllum commune
.
Isolation of
Shigella dysenteriae
Type 1 and
S. flexneri
Strains from Surface Waters in Bangladesh: Comparative Molecular Analysis of Environmental
Shigella
Isolates versus Clinical Strains
Authors: Faruque Shah M. , Khan Rasel , Kamruzzaman M. , Yamasaki Shinji , Ahmad Q. Shafi , Azim Tasnim , Nair G. Balakrish , Takeda Yoshifumi , Sack David A.
Published: October 2002

Bacillary dysentery caused by
Shigella
species is a public health problem in developing countries including Bangladesh. Although, shigellae-contaminated food and drinks are often the source of the epidemic's spread, the possible presence of the pathogen and transmission of it through environmental waters have not been adequately examined. We analyzed surface waters collected in Dhaka, Bangladesh, for the presence of shigellae by a combination of PCR assays followed by concentration and culturing of PCR-positive samples. Analysis of 128 water samples by PCR assays for
Shigella
-specific virulence genes including
ipaBCD
,
ipaH
, and
stx1
identified 14 (10.9%) samples which were positive for one or more of these virulence genes. Concentration of the PCR-positive samples by filtration followed by culturing identified live
Shigella
species in 11 of the 14 PCR-positive samples. Analysis of rRNA gene restriction patterns (ribotype) showed that the environmental isolates shared ribotypes with a collection of clinical isolates, but in contrast to the clinical isolates, 10 of the 11 environmental isolates were either negative or carried deletions in the plasmid-encoded invasion-associated genes
ipaB
,
ipaC
, and
ipaD
. However, all environmental
Shigella
isolates were positive for the chromosomal multicopy invasion-associated gene
ipaH
and all
Shigella dysenteriae
type 1 isolates were positive for the
stx1
gene in addition to
ipaH
. This study demonstrated the presence of
Shigella
in the aquatic environment and dispersion of different virulence genes among these isolates which appear to constitute an environmental reservoir of
Shigella
-specific virulence genes. Since critical virulence genes in
Shigella
are carried by plasmids or mobile genetic elements, the environmental gene pool may contribute to an optimum combination of genes, causing the emergence of virulent
Shigella
strains which is facilitated in particular by close contact of the population with surface waters in Bangladesh.
Ability of Nonpathogenic
Fusarium oxysporum
Strain Fo47 To Induce Resistance against
Pythium ultimum
Infection in Cucumber
Authors: Benhamou Nicole , Garand Chantal , Goulet Alain
Published: October 2002

The influence exerted by nonpathogenic
Fusarium oxysporum
strain Fo47 in triggering cucumber protection against infection by
Pythium ultimum
was investigated ultrastructurally. Macroscopic and microscopic observations of the pathogen colony in dual cultures revealed that reduction of
Pythium
growth was associated with marked disorders, including generalized disorganization of the host cytoplasm, retraction of the plasmalemma, and complete loss of the protoplasm. Cytochemical labeling of cellulose with an exoglucanase-gold complex showed that the cellulose component of the host cell walls was structurally preserved at a time when the host cytoplasm had undergone complete disorganization. A similar antagonistic process was observed at the root cell surface. Most striking and interesting was the finding that mycoparasitism, as evidenced by the frequent occurrence of Fo47 hyphae within nearly empty cells of the pathogen, occurred not only at the root surface but also within the invaded root tissues. The specific labeling pattern obtained with the exoglucanase-gold complex confirmed that Fo47 successfully penetrated cells of the pathogen, both in the rhizosphere and inside the root tissues.
Pythium
cells that could evade the first defensive line in the rhizosphere could penetrate the root epidermis, but their growth was restricted to the outermost tissues. Positive correlations between Fo47 treatment and induced resistance to infection by
P. ultimum
in cucumber were confirmed by (i) the reduction of pathogen viability; (ii) the elaboration of newly formed barriers, a phenomenon which was not seen in Fo47-free plants, where the pathogen proliferated in all root tissues within a few days; and (iii) the occlusion of intercellular spaces with a dense material likely enriched in phenolics. Taken together, our observations provide the first convincing evidence that Fo47 exerts a direct inhibitory effect on
P. ultimum
through a combination of antibiosis and mycoparasitism, in addition to being a strong inducer of plant defense reactions.
Internal Spatiotemporal Population Dynamics of Infection with ThreeWolbachiaStrains in the Adzuki Bean Beetle,Callosobruchus chinensis(Coleoptera: Bruchidae)
Authors: Ijichi Nobuyuki , Kondo Natsuko , Matsumoto Rena , Shimada Masakazu , Ishikawa Hajime , Fukatsu Takema
Published: October 2002

The adzuki bean beetle,Callosobruchus chinensis, is infected with three distinct lineages of endosymbiotic bacteria belonging to the genusWolbachia, which were designated wBruCon, wBruOri, and wBruAus. In an attempt to understand the mechanisms underlying the infection with these three organisms, the spatiotemporal infection dynamics of the threeWolbachiastrains was investigated in detail by using a quantitative PCR technique. During the development ofC. chinensis, the wBruCon, wBruOri, and wBruAus infection levels consistently increased but the growth patterns were different. The levels of infection plateaued at the pupal stage at approximately 3 × 108, 2 × 108, and 5 × 107wspcopy equivalents per insect for wBruCon, wBruOri, and wBruAus, respectively. At the whole-insect level, the population densities of the threeWolbachiatypes did not show remarkable differences between adult males and females. At the tissue level, however, the total densities and relative levels of the threeWolbachiatypes varied significantly when different tissues and organs were compared and when the same tissues derived from males and females were compared. The histological data obtained by in situ hybridization and electron microscopy were concordant with the results of quantitative PCR analyses. Based on the histological data and the peculiarWolbachiacomposition commonly found in nurse tissues and oocytes, we suggest that theWolbachiastrains are vertically transmitted to oocytes not directly, but by way of nurse tissue. On the basis of our results, we discuss interactions among the three coinfectingWolbachiatypes, reproductive strategies ofWolbachia, and factors involved in the different cytoplasmic incompatibility phenotypes.
Extent of Variation of the
Bacillus thuringiensis
Toxin Reservoir: the Case of the Geranium Bronze,
Cacyreus marshalli
Butler (Lepidoptera: Lycaenidae)
Authors: Herrero Salvador , Borja Marisé , Ferré Juan
Published: October 2002

Despite the fact that around 200
cry
genes from
Bacillus thuringiensis
have already been cloned, only a few Cry proteins are toxic towards a given pest. A crucial step in the mode of action of Cry proteins is binding to specific sites in the midgut of susceptible insects. Binding studies in insects that have developed cross-resistance discourage the combined use of Cry proteins sharing the same binding site. If resistance management strategies are to be implemented, the arsenal of Cry proteins suitable to control a given pest may be not so vast as it might seem at first. The present study evaluates the potential of
B. thuringiensis
for the control of a new pest, the geranium bronze (
Cacyreus marshalli
Butler), a butterfly that is threatening the popularity of geraniums in Spain. Eleven of the most common Cry proteins from the three lepidopteran-active Cry families (Cry1, Cry2, and Cry9) were tested against the geranium bronze for their toxicity and binding site relationships. Using
125
I-labeled Cry1A proteins we found that, of the seven most active Cry proteins, six competed for binding to the same site. For the long-term control of the geranium bronze with
B. thuringiensis
-based insecticides it would be advisable to combine any of the Cry proteins sharing the binding site (preferably Cry1Ab, since it is the most toxic) with those not competing for the same site. Cry1Ba would be the best choice of these proteins, since it is significantly more toxic than the others not binding to the common site.
Antimicrobial Resistance among Enterococci from Pigs in Three European Countries
Authors: Aarestrup Frank Møller , Hasman Henrik , Jensen Lars Bogø , Moreno Miguel , Herrero Inmaculada A. , Domínguez Lucas , Finn Maria , Franklin Anders
Published: October 2002

Enterococci from pigs in Denmark, Spain, and Sweden were examined for susceptibility to antimicrobial agents and copper and the presence of selected resistance genes. The greatest levels of resistance were found among isolates from Spain and Denmark compared to those from Sweden, which corresponds to the amounts of antimicrobial agents used in food animal production in those countries. Similar genes were found to encode resistance in the different countries, but the
tet
(L) and
tet
(S) genes were more frequently found among isolates from Spain. A recently identified transferable copper resistance gene was found in all copper-resistant isolates from the different countries.
Asaia
sp., an Unusual Spoilage Organism of Fruit-Flavored Bottled Water
Authors: Moore John E. , McCalmont Mark , Xu Jiru , Millar B. Cherie , Heaney Neville
Published: October 2002

A gram-negative bacillus was isolated from a batch of fruit-flavored bottled water, which had spoiled as a result of bacterial overgrowth (>10
6
CFU/ml). The spoilage organism was extremely difficult to identify phenotypically and was poorly identified as
Pasturella
sp. (78.7% identification profile) employing the API 20NE identification scheme, which gave the profile 5040000. Molecular identification through PCR amplification of a partial region of the 16S rRNA gene followed by direct automated sequencing of the PCR amplicon allowed identification of the organism. Due to the sequence identity (100%) between the spoilage organism and a reference strain in GenBank, the spoilage isolate was considered to be an
Asaia
sp., a recently described genus and member of the acetic acid bacteria. This is the first report of
Asaia
sp. causing spoilage of a foodstuff and highlights the benefits of molecular identification techniques based on 16S rRNA gene sequences in the identification of unusual spoilage organisms.
Fluorescent Amplified Fragment Length Polymorphism and Repetitive Extragenic Palindrome-PCR Fingerprinting Reveal Host-Specific Genetic Diversity of
Vibrio halioticoli
-Like Strains Isolated from the Gut of Japanese Abalone
Authors: Sawabe Tomoo , Thompson Fabiano L. , Heyrman Jeroen , Cnockaert Margo , Hayashi Karin , Tanaka Reiji , Yoshimizu Mamoru , Hoste Bart , Swings Jean , Ezura Yoshio
Published: October 2002

When analyzed by fluorescent amplified fragment length polymorphism and repetitive extragenic palindrome-PCR fingerprinting, a total of 47
Vibrio halioticoli
strains isolated from four Japanese abalone species and one turban shell species formed three clusters that roughly reflect the different species of host abalone from which they were isolated. The
V. halioticoli
isolates from turban shells were distributed evenly among the clusters. Representative isolates from two clusters were deemed separate species or subspecies by DNA-DNA hybridization.
Accelerated Death Kinetics ofAspergillus nigerSpores under High-Pressure Carbonation
Authors: Shimoda M. , Kago H. , Kojima N. , Miyake M. , Osajima Y. , Hayakawa I.
Published: October 2002

The death kinetics ofAspergillus nigerspores under high-pressure carbonation were investigated with respect to the concentration of dissolved CO2(dCO2) and treatment temperature. All of the inactivation followed first-order death kinetics. TheDvalue (decimal reduction time, or the time required for a 1-log-cycle reduction in the microbial population) in the saline carbonated at 10 MPa was 0.16 min at 52°C. The logDvalues were linearly related to the treatment temperature and the concentration of dCO2, but a significant interaction was observed between them.
Inactivation of
Cryptosporidium parvum
Oocysts in Fresh Apple Cider by UV Irradiation
Authors: Hanes D. E. , Worobo R. W. , Orlandi P. A. , Burr D. H. , Miliotis M. D. , Robl M. G. , Bier J. W. , Arrowood M. J. , Churey J. J. , Jackson G. J.
Published: October 2002

This study evaluated the efficacy of UV irradiation on the inactivation of
Cryptosporidium parvum
oocysts in fresh apple cider. Cider was inoculated with oocysts and exposed to 14.32 mJ of UV irradiation/cm
2
. Oocyst viability was assessed with the gamma interferon gene knockout (GKO) mouse and infant BALB/cByJ mouse models. All GKO mice challenged with UV-treated cider demonstrated no morbidity or mortality, and infant BALB/c mice challenged with treated cider were negative for the presence of
C. parvum
. In contrast, the GKO mice challenged with non-UV-treated inoculated cider died and the parasite was detected in the ileums of all challenged infant mice. This study shows that UV irradiation can be used to inactivate
C. parvum
in fresh apple cider.
Human Salivary Histatin 5 Causes Disordered Volume Regulation and Cell Cycle Arrest inCandida albicans
Authors: Baev Didi , Li Xuewei S. , Dong Jin , Keng Peter , Edgerton Mira
Published: October 2002

Human salivary histatin 5 (Hst 5) is a nonimmune salivary protein with antifungal activity against an important human pathogen,Candida albicans.The candidacidal activity of histatins appears to be a distinctive multistep mechanism involving depletion of theC. albicansintracellular ATP content as a result of nonlytic ATP efflux. Hst 5 caused a loss of cell viability concomitant with a decrease in cellular volume as determined both by a classical candidacidal assay with exogenous Hst 5 and by using a genetically engineeredC. albicansstrain expressing Hst 5. Preincubation ofC. albicanscells with pharmacological inhibitors of anion transport provided complete or substantial protection from Hst 5-induced killing and volume reduction of cells. Moreover, intracellular expression of Hst 5 resulted in a reduction in the population mean cell volume that was accompanied by an increase in the percentage of unbudded cells andC. albicanscells in the G1phase. Following expression of Hst 5, the smallest cells sorted by fluorescence-activated cell sorting from the total population did not replicate and were exclusively in the G1phase. Cells with intracellularly expressed Hst 5 had greatly reduced G1cyclin transcript levels, indicating that they arrested in the G1phase before the onset of Start. Our data demonstrate that a key determinant in the mechanism of Hst 5 toxicity inC. albicanscellsis the disruption of regulatory circuits for cell volume homeostasis that is closely coupled with loss of intracellular ATP. This novel process of fungicidal activity by a human salivary protein has highlighted potential interactions of Hst 5 with volume regulatory mechanisms and the process of yeast cell cycle control.