bluuebunny/crossref_metadata_2025 huggingface dataset:
A Whole-Genome Shotgun Optical Map of
Yersinia pestis
Strain KIM
Authors: Zhou Shiguo , Deng Wen , Anantharaman Thomas S. , Lim Alex , Dimalanta Eileen T. , Wang Jun , Wu Tian , Chunhong Tao , Creighton Robert , Kile Andrew , Kvikstad Erika , Bechner Michael , Yen Galex , Garic-Stankovic Ana , Severin Jessica , Forrest Dan , Runnheim Rod , Churas Chris , Lamers Casey , Perna Nicole T. , Burland Valerie , Blattner Frederick R. , Mishra Bhubaneswar , Schwartz David C.
Published: November 2002

Yersinia pestis
is the causative agent of the bubonic, septicemic, and pneumonic plagues (also known as black death) and has been responsible for recurrent devastating pandemics throughout history. To further understand this virulent bacterium and to accelerate an ongoing sequencing project, two whole-genome restriction maps (
Xho
I and
Pvu
II) of
Y. pestis
strain KIM were constructed using shotgun optical mapping. This approach constructs ordered restriction maps from randomly sheared individual DNA molecules directly extracted from cells. The two maps served different purposes; the
Xho
I map facilitated sequence assembly by providing a scaffold for high-resolution alignment, while the
Pvu
II map verified genome sequence assembly. Our results show that such maps facilitated the closure of sequence gaps and, most importantly, provided a purely independent means for sequence validation. Given the recent advancements to the optical mapping system, increased resolution and throughput are enabling such maps to guide sequence assembly at a very early stage of a microbial sequencing project.
AbiA, a Lactococcal Abortive Infection Mechanism Functioning in
Streptococcus thermophilus
Authors: Tangney Mark , Fitzgerald Gerald F.
Published: November 2002

The lactococcal abortive infection mechanisms AbiA and AbiG were introduced into
Streptococcus thermophilus
4035, and a range of phages capable of infecting this host were examined for sensitivity to these mechanisms. AbiA proved effective against six phages when examined at a growth temperature of 30°C but had no effect on any of the phages when tested at 37 or 42°C. AbiG failed to affect any of the
S. thermophilus
phages at 30, 37, or 42°C.
The Environmental Pathogen
Mycobacterium ulcerans
Grows in Amphibian Cells at Low Temperatures
Authors: Drancourt Michel , Jarlier Vincent , Raoult Didier
Published: November 2002

Mycobacterium ulcerans
, the etiological agent of Buruli ulcers, is an environmental pathogen. We cultivated it in an amphibian (XTC-2) cell line that grows at 28°C. By counting of Ziehl-Neelsen-stained mycobacteria and by quantitative PCR analysis, we found that
M. ulcerans
multiplies rapidly in association with XTC-2 cells. Transmission electron microscopy demonstrated the presence of intracellular
M. ulcerans
microorganisms. These data suggest an intracellular environmental niche, and we propose use of XTC-2 cells for isolation of
M. ulcerans
from environmental sources.
Phylogenetic Analysis of Acute Bee Paralysis Virus Strains
Authors: Bakonyi Tamás , Grabensteiner Elvira , Kolodziejek Jolanta , Rusvai Miklós , Topolska Gražyna , Ritter Wolfgang , Nowotny Norbert
Published: November 2002

Reverse transcription-PCR assays have been established for a quick, sensitive, and specific diagnosis of acute bee paralysis virus (ABPV), a common virus of the honeybee (
Apis mellifera
), directly from clinical samples. A 3,071-nucleotide fragment of the ABPV genome, which includes the entire capsid polyprotein gene, was amplified from Austrian, German, Polish, and Hungarian ABPV samples and sequenced, and the sequences were compared. The alignment of a smaller fragment with ABPV sequences from the United States and the United Kingdom revealed nucleotide identity rates between 89 and 96%, respectively. Phylogenetic trees which display the molecular relationship between the viruses of different geographic origin were constructed.
Reliability of Nested PCR for Detection of
Chlamydia pneumoniae
DNA in Atheromas: Results from a Multicenter Study Applying Standardized Protocols
Authors: Apfalter Petra , Assadian Ojan , Blasi Francesco , Boman Jens , Gaydos Charlotte A. , Kundi Michael , Makristathis Athanasios , Nehr Marion , Rotter Manfred L. , Hirschl Alexander M.
Published: November 2002

The present multicenter study was designed to find explanations for the discrepancies in the reported rates of detection of
Chlamydia pneumoniae
DNA in endarterectomy specimens. Coded identical sets of (i) a
C. pneumoniae
DNA dilution series (panel 1;
n
= 10), (ii) spiked control tissue specimens (panel 2;
n
= 10 specimens, including 5 negative controls), and (iii) endarterectomy specimens (panel 3; 15 atheromas, 5 negative controls) were analyzed at four laboratories by three standardized DNA extraction methods in each laboratory and a nested touchdown PCR protocol targeting the
ompA
gene of
C. pneumoniae
. Panel 1 samples were correctly identified as positive to levels of 0.3 inclusion-forming units (IFU)/PCR mixture (100%) and 0.03 IFU/PCR mixture (50%). All negative controls were correctly reported as negative. Panel 2 samples were identified as
C. pneumoniae
positive to levels of 0.01 IFU/PCR mixture (100%) and 0.005 IFU/PCR mixture (91%), independent of the DNA extraction method used, and only one false-positive result was reported. For panel 3 samples, 5 of 240 (2%) analyses (in which DNA extractions and PCR were performed at the same laboratory) were positive; the positive specimens were from three endarterectomy specimens and two negative controls. After exchange of DNA extracts between laboratories, 13 of 15 atheroma samples were
C. pneumoniae
DNA positive in at least 1 of a series of 48 analyses per atheroma sample; however, the overall positivity rate did not exceed 5% (33 of 720 analyses) and therefore was lower than that for the negative controls (8%; 19 of 240 analyses). Not a single positive result could be achieved when all panel 3 extracts (
n
= 240 analyses) were reamplified by a 16S rRNA PCR followed by hybridization with a
C. pneumoniae
-specific probe. Statistical analyses demonstrated that positive results did not occur in an independent and random fashion and could most likely be explained by amplicon carryover at the nested PCR level as well as amplicon introduction during DNA extraction, but not by the patterns of distribution of very low target levels or a certain DNA extraction protocol. The results of studies by nested PCR for detection of the prevalence of
C. pneumoniae
will always be questionable.
Microevolution of the Direct Repeat Region of
Mycobacterium tuberculosis
: Implications for Interpretation of Spoligotyping Data
Authors: Warren R. M. , Streicher E. M. , Sampson S. L. , van der Spuy G. D. , Richardson M. , Nguyen D. , Behr M. A. , Victor T. C. , van Helden P. D.
Published: November 2002

The direct repeat (DR) region has been determined to be an important chromosomal domain for studying the evolution of
Mycobacterium tuberculosis
. Despite this, very little is known about microevolutionary events associated with clonal expansion and how such events influence the interpretation of both restriction fragment length polymorphism (RFLP) and spoligotype data. This study examined the structure of the DR region in three independently evolving lineages of
M. tuberculosis
with a combination of DR-RFLP, spoligotyping, and partial DNA sequencing. The results show that the duplication of direct variable repeat (DVR) sequences and single-nucleotide polymorphisms is rare; conversely, the deletion of DVR sequences and IS
6110
-mediated mutation is observed frequently. Deletion of either single or contiguous DVR sequences was observed. The deletion of adjacent DVR sequences occurred in a dependent manner rather than as an accumulation of independent events. Insertion of IS
6110
into either the direct repeat or spacer sequences influenced the spoligotype pattern, resulting in apparent deletion of DVR sequences. Homologous recombination between adjacent IS
6110
elements led to extensive deletion in the DR region, again demonstrating a dependent evolutionary mechanism. Different isolates from the same strain family and isolates from different strain families were observed to converge to the same spoligotype pattern. In conclusion, the binary data of the spoligotype are unable to provide sufficient information to accurately establish genotypic relationships between certain clinical isolates of
M. tuberculosis
. This has important implications for molecular epidemiologic strain tracking and for the application of spoligotype data to phylogenetic analysis of
M. tuberculosis
isolates.
M Types of Group A Streptococcal Isolates Submitted to the National Centre for Streptococcus (Canada) from 1993 to 1999
Authors: Tyrrell Gregory J. , Lovgren Marguerite , Forwick Betty , Hoe Nancy P. , Musser James M. , Talbot James A.
Published: November 2002

The National Centre for Streptococcus (NCS) (Canada) determined the group A streptococcal (GAS) M types of 4,760 Canadian isolates submitted between 1993 and 1999 by classic serotyping. The 10 most frequently identified M types were M1 (26.4%), M12 (9.8%), M28 (8.9%), M3 (6.8%), M4 (6.2%), M11 (4.8%), M89 (3.1%), M6 (3.0%), M2 (2.6%), and M77 (1.9%). Nontypeable isolates accounted for 15.4% of the collection. The province of Ontario submitted 51.1% of the isolates, followed by Quebec (21.2%) and Alberta (13.9%). Together, these three provinces constituted 71.3% of the Canadian population in 1996. The numbers of M types M1, M12, M28, and M3 occurred most frequently in subjects whose ages were <1 to 15 years and 25 to 45 years, as well as in the elderly (60 to 90 years). Further analysis found that the four most frequently identified M types from blood, brain, and cerebrospinal fluid were M1 (28.2%), M28 (9.2%), M12 (9.1%), and M3 (8.2%), with 13.4% of isolates being nontypeable. The four isolates from throats most frequently identified were M1 (19.5%), M12 (15.3%), M3 (8.6%), and M28 (5%) with 19.4% of isolates being nontypeable. Thesicgene of a subset of M1 strains (9.5% of the M1 collection) was sequenced. Of 36 sic types identified, the four most common were sic1.01 (22.8%), sic1.02 (14.9%), sic1.135 (10.5%), and sic1.178 (9.6%). Together these four sic types further characterized nearly 60% of the M1 strains sequenced. In summary, from the years 1993 to 1999, the NCS detected 54 M types, of which 10 different M types constituted 73.5% of the collection. M1 was the most common GAS M type circulating in the Canadian population, responsible for more than a quarter of the isolates typed. The most common throat isolates differed in M-type and proportion from those of invasive isolates. Sequencing thesicgene further characterized the most common M-type serotype 1 in a fashion that may be useful for epidemiologic investigations.
Genetic Diversity of Intimin Genes of Attaching and EffacingEscherichia coliStrains
Authors: Zhang W. L. , Köhler B. , Oswald E. , Beutin L. , Karch H. , Morabito S. , Caprioli A. , Suerbaum S. , Schmidt H.
Published: November 2002

In this study, we determined the sequences of four intimin variant genes detected in attaching and effacingEscherichia coliisolates of human origin. Three of them were novel and were designatedeae-η (eta),eae-ι (iota), andeae-κ (kappa). The fourth was identical to the recently describedeae-ζ (zeta), isolated from a bovineE. coliO84:NM isolate. We compared these sequences with those of published intimin-α, intimin-β, intimin-γ1, intimin-γ2, intimin-ε, and intimin-θ alleles. Sequence analysis of these 10 intimin alleles confirmed extensive genetic diversity within the intimin gene family inE. coli.The genetic diversity was more prominent in the 3′ region (starting at bp 2112), which encodes the binding domain of intimin. Phylogenetic analyses revealed four groups of closely related intimin genes: α and ζ; β and κ; γ1 and γ2/θ; and ε and η. Calculation of homoplasy ratios of sequences of the 5′ region ofeae(positions 1 to 2111) revealed evidence for intragenic recombination. Split decomposition analysis also indicates that recombination events have played a role in the evolutionary history ofeae.In conclusion, we recommend aneaenomenclature system based on the Greek alphabet and provide an updated PCR scheme for amplification and typing ofE. coli eae.
Sequence Diversity of
Neisseria meningitidis
16S rRNA Genes and Use of 16S rRNA Gene Sequencing as a Molecular Subtyping Tool
Authors: Sacchi Claudio T. , Whitney Anne M. , Reeves Michael W. , Mayer Leonard W. , Popovic Tanja
Published: November 2002

We investigated the diversity of the primary sequences of 16S rRNA genes among
Neisseria meningitidis
strains (Men) and evaluated the use of this approach as a molecular subtyping tool. We aligned and compared a 1,417-bp fragment of the 16S rRNA gene from 264 Men strains of serogroups A, B, C, and Y (MenA, MenB, MenC, and MenY, respectively) isolated throughout the world over a 30-year period. Thirty-one positions of difference were found among 49 16S types: differences between types ranged from 1 to 14 positions (0.07 to 0.95%). 16S types and serogroups were highly associated; only 3 out 49 16S types were shared by two or more serogroups. We have identified 16S types that are exclusively associated with strains of certain hypervirulent clones: 16S type 5 with MenA subgroup III, 16S type 4 with the MenB electrophoretic type 5 (ET-5) complex, and 16S types 12 and 13 with MenC of the ET-37 complex. For MenC strains, 16S sequencing provided the highest sensitivity and specificity and the best overall association with the outbreak-related versus sporadic isolates when compared with pulsed-field gel electrophoresis, multilocus enzyme electrophoresis, and multilocus sequence typing. We demonstrated for the first time an unexpected diversity among 16S rRNA genes of Men strains, identified 16S types associated with well-defined hypervirulent clones, and showed the potential of this approach to rapidly identify virulent strains associated with outbreaks and/or an increased incidence of sporadic disease.
Direct and Rapid Identification and Genogrouping of Meningococci andporAAmplification by LightCycler PCR
Authors: Mölling Paula , Jacobsson Susanne , Bäckman Anders , Olcén Per
Published: November 2002

Invasive meningococcal infections are usually diagnosed by culture. This approach is far from optimal due to, e.g., treatment with precollection antibiotics. Molecular-genetics methods are therefore recognized as important tools for optimal laboratory confirmation of meningococcal infections as well as characterization of meningococci (Mc). The aims of the present study were to develop real-time PCRs for identification and genogrouping (A, B, C, Y, and W-135) of Mc andporAamplification that further can be used for subsequent genosubtyping. In a first run Mc were identified. In a second run they were genogrouped andporAgenes were amplified. All the Mc isolates (n= 71) but one and cerebrospinal fluid samples (n= 11) tested gave the expected positive results. The specificity, inter- and intra-assay variations, and effects of different amounts of DNA on the melting temperatures were also explored. The LightCycler PCR system was found to effectively detect and characterize Mc in a few hours. This testing, including the DNA sequencing of theporAgene to reveal the genosubtype, does not take more than a working day, and the results can be compared to those from culturing with phenotypic analysis, which takes at least a few days.
Normalized Quantification of Human Cytomegalovirus DNA by Competitive Real-Time PCR on the LightCycler Instrument
Authors: Stöcher Markus , Berg Jörg
Published: November 2002

The development of a novel normalized quantitative competitive real-time PCR on the LightCycler instrument (NQC-LC-PCR) and its application to the quantification of cytomegalovirus (CMV) DNA in clinical samples are described. A heterologous competitor DNA was spiked into test samples and served as an internal amplification control. The internal control (IC) DNA in the test samples was coamplified with the CMV DNA and was tested against a calibrator sample that contained equal amounts of IC DNA and CMV reference standard DNA. An algorithm was developed to normalize possible varying amplification efficiencies between the standard and the samples. After normalization, CMV DNA copy numbers were determined in absolute terms. In a routine clinical setting, normalized quantification by NQC-LC-PCR using a single IC concentration led to results ranging from 500 to 50,000 CMV DNA copies/ml. The results obtained with conventional real-time quantification on the LightCycler instrument were almost identical to those obtained with the NQC-LC-PCR-based quantification. This was the case only for samples in which the PCR was not inhibited. With partially inhibited samples, NQC-LC-PCR was still able to correctly quantify CMV DNA copy numbers even when the PCR was inhibited by about 70%. By analyzing 80 undefined clinical samples, we found that NQC-LC-PCR was suitable for the routine assessment of CMV DNA in clinical plasma samples. Since the ICs were already added to the samples during the DNA purification, almost the entire assay was controlled for sample adequacy. Thus, false negative results were precluded. The NQC-LC-PCR approach developed should be adaptable for additional microbiological applications.
Hepatitis E Virus (HEV) Capsid Antigens Derived from Viruses of Human and Swine Origin Are Equally Efficient for Detecting Anti-HEV by Enzyme Immunoassay
Authors: Engle R. E. , Yu C. , Emerson S. U. , Meng X.-J. , Purcell R. H.
Published: November 2002

The recombinant truncated ORF2 (capsid) antigen derived from the Meng strain of swine hepatitis E virus (HEV) differs from that of the Sar-55 strain of human HEV by approximately 5% at the amino acid level. Serial serum samples from two chimpanzees and six rhesus monkeys experimentally infected with HEV were tested with one enzyme immunoassay (EIA) based on the Sar-55 antigen and with a second EIA based on the Meng antigen. We obtained 98% agreement (κ = 0.952) by direct comparison. The virtually identical results obtained with these antigens in detecting seroconversion following infection with HEV suggests that they were reacting with antibodies that detect the same or very similar epitopes of HEV. We then tested human and swine serum samples for anti-HEV in EIAs that utilized one or the other of the two ORF2 antigens and showed that these results were also virtually identical. The specimens tested included swine sera from the United States, Canada, China, Korea, and Thailand and sera from veterinarians, U.S. and non-U.S. volunteer blood donors, and U.S. and non-U.S. animal handlers. We tested 792 swine sera and obtained 93% agreement (κ = 0.839). We similarly tested 882 human sera and obtained 99% agreement (κ = 0.938). Moreover, we found virtually no difference in the levels of prevalence of anti-HEV as measured by the two tests, again suggesting that the antigens derived from human and swine HEV contain the same immunodominant epitopes.
Strain Typing of
Borrelia burgdorferi
,
Borrelia afzelii
, and
Borrelia garinii
by Using Multiple-Locus Variable-Number Tandem Repeat Analysis
Authors: Farlow Jason , Postic Danielle , Smith Kimothy L. , Jay Zack , Baranton Guy , Keim Paul
Published: November 2002

Human Lyme borreliosis (LB) is the most prevalent arthropod-borne infection in temperate climate zones around the world and is caused by
Borrelia
spirochetes. We have identified 10 variable-number tandem repeat (VNTR) loci present within the genome of
Borrelia burgdorferi
and subsequently developed a multiple-locus VNTR analysis (MLVA) typing system for this disease agent. We report here the successful application of MLVA for strain discrimination among a group of 41 globally diverse
Borrelia
isolates including
B. burgdorferi
,
B. afzelii
, and
B. garinii
. PCR assays displayed diversity at these loci, with total allele numbers ranging from two to nine and Nei's diversity (
D
) values ranging from 0.10 to 0.87. The average
D
value was 0.53 across all VNTR loci. A clear correlation exists between the repeat copy number and the
D
value (
r
= 0.62) or the number of alleles (
r
= 0.93) observed across diverse strains. Cluster analysis by the unweighted pair-group method with arithmetic means resolved the 30 observed unique
Borrelia
genotypes into five distinct groups.
B. burgdorferi
,
B. afzelii
, and
B. garinii
clustered into distinct affiliations, consistent with current 16S rRNA phylogeny studies. Genetic similarity and diversity suggest that
B. afzelii
and
B. garinii
are close relatives and were perhaps recently derived from
B. burgdorferi
. MLVA provides both phylogenetic relationships and additional resolution to discriminate among strains of
Borrelia
species. This new level of strain identification and discrimination will allow more detailed epidemiological and phylogenetic analysis in future studies.
Improvement of Differentiation and Interpretability of Spoligotyping for
Mycobacterium tuberculosis
Complex Isolates by Introduction of New Spacer Oligonucleotides
Authors: van der Zanden A. G. M. , Kremer K. , Schouls L. M. , Caimi K. , Cataldi A. , Hulleman A. , Nagelkerke N. J. D. , van Soolingen D.
Published: November 2002

The direct repeat (DR) region in
Mycobacterium tuberculosis
complex strains is composed of multiple well-conserved 36-bp DRs interspersed with nonrepetitive DNA spacer sequences of similar size. Clinical isolates show extensive polymorphism in this DR region, and this has led to the development of a 43-spacer reversed line blot methodology: spoligotyping. Although this method has contributed significantly to the molecular epidemiology of tuberculosis in the last decade, the discriminatory power and the readability of this method were not found to be optimal. In order to improve the discriminatory power, the usefulness of 43 redesigned oligonucleotides and the usefulness of 51 new spacer oligonucleotides were evaluated. For 314
M. tuberculosis
complex strains isolated in the central part of The Netherlands over a 5-year period, 264 different IS
6110
RFLP types could be distinguished, and 160 different spoligotype patterns were identified by traditional spoligotyping. After the introduction of 51 new spacer oligonucleotides, 14 additional spoligotypes were recognized. This enabled us to split 11 clusters of isolates identified by the traditional spoligotyping. Furthermore, on the basis of the new spacer oligonucleotides a dichotomy was found among the Beijing genotype isolates. Among 76
Mycobacterium bovis
strains, 20 patterns were found by traditional spoligotyping and 30 patterns were found by novel probe spoligotyping, respectively. Nine
M. bovis
subsp.
caprae
isolates yielded six patterns by traditional spoligotyping and eight patterns by novel probe spoligotyping. A part of the redesigned oligonucleotides slightly improved the reading of spoligotype patterns. The reproducibility of spoligotyping, based on internal control probes, invariably yielded a high score; only 4 (1%) of the 314 patient isolates gave discrepant results. Analysis of a set of 31 duplicate
M. tuberculosis
complex strains demonstrated a 10% error rate for the identification of blinded duplicate samples. In a redundancy analysis, 40 essential spacer oligonucleotides of the 94-spacer sequences were selected, yielding the same number of spoligotype patterns. We propose to leave the traditional commercialized first-generation membrane for spoligotyping unchanged for current applications and to introduce a second-generation spoligotyping membrane whenever extended discrimination is required, e.g., for low-copy-number IS
6110
strains or for phylogenetic studies of Beijing genotype strains.
Simultaneous Identification of
Mycobacterium
Genus and
Mycobacterium tuberculosis
Complex in Clinical Samples by 5′-Exonuclease Fluorogenic PCR
Authors: García-Quintanilla Albert , González-Martín Julián , Tudó Griselda , Espasa Mateu , Jiménez de Anta María T.
Published: November 2002

Early diagnosis of tuberculosis and screening of other mycobacteria is required for the appropriate management of patients. We have therefore developed a 5′-exonuclease fluorogenic PCR assay in a single-tube balanced heminested format that simultaneously detects
Mycobacterium tuberculosis
complex (MTC) and members of the
Mycobacterium
genus (MYC) using the 16S ribosomal DNA target directly on clinical samples. One hundred twenty-seven clinical samples (65 smear negative and 62 smear positive) with a positive culture result from 127 patients were tested, including 40 negative control specimens. The finding of both a positive MTC and probe value and a positive MYC probe value confirmed the presence of MTC or mycobacteria with a 100% positive predictive value. However, a negative value for MTC or MYC did not discount the presence of mycobacteria in the specimen. Interestingly, the addition of the MYC probe allowed the diagnosis of an additional 7% of patients with tuberculosis and rapid screening of nontuberculous mycobacteria (NTM). Thus, over 75% of the patients were diagnosed with mycobacterial disease by PCR. The sensitivity was much higher on smear-positive samples (90.3%) than smear-negative samples (49.2%) and was slightly higher for MTC than NTM samples. With regard to the origin of the sample, MTC pulmonary samples gave better results than others. In conclusion, we believe this test may be useful for the rapid detection of mycobacteria in clinical samples and may be a valuable tool when used together with conventional methods and the clinical data available.
Comparison of Two Commercial Assays with Expert Microscopy for Confirmation of Symptomatically Diagnosed Malaria
Authors: Iqbal Jamshaid , Khalid Nabila , Hira Parsotam R.
Published: November 2002

Conventional light microscopy has been the established method for malaria diagnosis. However, recently several nonmicroscopic rapid diagnostic tests have been developed for situations in which reliable microscopy may not be available. This study was conducted to evaluate the diagnostic performance of a recently introduced ICT Malaria Pf/Pv test. This assay detects
Plasmodium falciparum
histidine-rich protein 2 antigen (PfHRP-2) for
P. falciparum
diagnosis and pan-malarial antigen for
P. vivax
diagnosis. In this study we compared the performance of ICT Malaria Pf/Pv with microscopy of Giemsa-stained blood films and with an OptiMAL test that detects
Plasmodium
lactate dehydrogenase (pLDH) antigen. A total of 750 clinically suspected malaria patients were examined at local health centers in Kuwait. Both the antigen tests had a high degree of specificity (>98%) for detection of malaria infection. However, they were less sensitive than microscopy. Compared with microscopy the ICT Malaria PF/pf test failed to detect malaria infection in 93 (34%) of 271 malaria patients (11% of patients with
P. falciparum
and 37% of patients with
P. vivax
) and the OptiMAL test failed to detect malaria infection in 41 (15%) of 271 malaria patients (7% of patients with
P. falciparum
and 13% of patients with
P. vivax
). The sensitivities of the ICT Malaria Pf/Pv and OptiMAL tests for detection of
P. falciparum
infection were 81 and 87%, and those for detecting
P. vivax
were 58 to 79%, respectively. The sensitivity of the ICT Malaria Pf/Pv and OptiMAL tests decreased significantly to 23 and 44%, respectively, at parasite densities of <500/μl. Both of the tests also produced a number of false-positive results. Overall, the performance of the OptiMAL test was better than that of the ICT Malaria Pf/Pv test. However, our results raise particular concern over the sensitivity of the ICT Malaria Pf/Pv test for detection of
P. vivax
infection. Further developments appear necessary to improve the performance of the ICT Malaria Pf/Pv test.
Identification ofListeriaSpecies by Microarray-Based Assay
Authors: Volokhov Dmitriy , Rasooly Avraham , Chumakov Konstantin , Chizhikov Vladimir
Published: November 2002

We have developed a rapid microarray-based assay for the reliable detection and discrimination of six species of theListeriagenus:L. monocytogenes,L. ivanovii,L. innocua,L. welshimeri,L. seeligeri, andL. grayi. The approach used in this study involves one-tube multiplex PCR amplification of six target bacterial virulence factor genes (iap,hly,inlB,plcA,plcB, andclpE), synthesis of fluorescently labeled single-stranded DNA, and hybridization to the multiple individual oligonucleotide probes specific for eachListeriaspecies and immobilized on a glass surface. Results of the microarray analysis of 53 reference and clinical isolates ofListeriaspp. demonstrated that this method allowed unambiguous identification of all sixListeriaspecies based on sequence differences in theiapgene. Another virulence factor gene,hly, was used for detection and genotyping allL. monocytogenes, allL. ivanovii, and 8 of 11L. seeligeriisolates. Other members of the genusListeriaand threeL. seeligeriisolates did not contain thehlygene. There was complete agreement between the results of genotyping based on thehlyandiapgene sequences. AllL. monocytogenesisolates were found to be positive for theinlB,plcA,plcB, andclpEvirulence genes specific only to this species. Our data onListeriaspecies analysis demonstrated that this microarray technique is a simple, rapid, and robust genotyping method that is also a potentially valuable tool for identification and characterization of bacterial pathogens in general.
Apodemus
Species Mice Are Reservoir Hosts of
Borrelia garinii
OspA Serotype 4 in Switzerland
Authors: Huegli D. , Hu C. M. , Humair P.-F. , Wilske B. , Gern L.
Published: November 2002

Among
Borrelia burgdorferi
sensu lato isolates, seven outer surface protein A (OspA) serotypes have been described: serotypes 1 and 2 correspond to
B. burgdorferi
sensu stricto and
Borrelia afzelii
, respectively, and serotypes 3 to 7 correspond to
Borrelia garinii
. In Europe, serotype 4 has never been isolated from
Ixodes ricinus
ticks until recently, although this serotype has been frequently isolated from cerebrospinal fluid from patients. In Europe,
B. afzelii
and
B. burgdorferi
sensu stricto were found associated with rodents and
B. garinii
was found associated with birds. In this study, the reservoir role of
Apodemus
mice for
B. garinii
OspA serotype 4 was demonstrated by xenodiagnosis.
Apodemus
mice are the first identified reservoir hosts for
B. garinii
OspA serotype 4.
New Serological Assay for Detection of Putative
Helicobacter pylori
Virulence Factors
Authors: Park Chang-Young , Cho Yong-Kyun , Kodama Tadashi , El-Zimaity Hala M. T. , Osato Michael S. , Graham David Y. , Yamaoka Yoshio
Published: November 2002

We evaluated a new immunoblot assay (Helicoblot 2.1) for
Helicobacter pylori
infection and CagA and VacA status by using serum samples from 222 patients. The test accurately detected
H. pylori
infection and VacA status, but improvements in the interpretation criteria are required before it can be recommended for determination of CagA status.
Mycobacterium avium
subsp.
paratuberculosis
Strains from Cattle and Sheep Can Be Distinguished by a PCR Test Based on a Novel DNA Sequence Difference
Authors: Collins Desmond M. , De Zoete May , Cavaignac Sonia M.
Published: November 2002

A DNA sequence differing between sheep and cattle types of
Mycobacterium avium
subsp.
paratuberculosis
was identified and used to develop a PCR test. The test unequivocally distinguished all sheep types from cattle types and was negative for a wide range of other strains from the
Mycobacterium avium
-
Mycobacterium intracellulare
complex. The test will be useful for epidemiological purposes, particularly in hosts such as deer that can be easily infected with either type.
Loss of a 29-Kilodalton Outer Membrane Protein in
Acinetobacter baumannii
Is Associated with Imipenem Resistance
Authors: Limansky Adriana S. , Mussi María Alejandra , Viale Alejandro M.
Published: November 2002

We analyzed the possible causes of imipenem (IPM) resistance in multidrug-resistant isolates of
Acinetobacter baumannii
. Comparison of the outer membrane protein (OMP) profiles of two genomically related strains (Ab288 [IPM sensitive] and Ab242 [IPM resistant]) indicated the conspicuous loss of a 29-kDa polypeptide in the Ab242 strain. No carbapenemase activity was detected in any of these strains. The treatment of Ab288 with sodium salicylate resulted in IPM resistance and the loss of the 29-kDa OMP. In addition, IPM-resistant clones of Ab288 which were selected by repetitive culturing in increasing concentrations of this antibiotic also showed the absence of this 29-kDa OMP.
Roseomonas gilardii
Infection: Case Report and Review
Authors: Shokar Navkiran K. , Shokar Gurjeet S. , Islam Jamal , Cass Alvah R.
Published: November 2002

Roseomonas gilardii
is a bacterium that has been indicated as a rare cause of human infections. The case of a patient presenting with cellulitis and bacteremia secondary to
R. gilardii
is described together with the clinical characteristics of infection with this organism obtained from a review of cases previously reported.
Rotavirus Gastroenteritis and Central Nervous System (CNS) Infection: Characterization of the VP7 and VP4 Genes of Rotavirus Strains Isolated from Paired Fecal and Cerebrospinal Fluid Samples from a Child with CNS Disease
Authors: Iturriza-Gómara M. , Auchterlonie I. A. , Zaw W. , Molyneaux P. , Desselberger U. , Gray J.
Published: November 2002

Rotavirus RNA was detected in the cerebrospinal fluid (CSF) of a child with central nervous system disease symptoms associated with rotavirus gastroenteritis. The rotavirus isolates from the fecal and CSF samples were genotyped as G1P[8]. Sequence analysis of the VP7 and VP4 proteins derived from the fecal and CSF samples were remarkably similar to each other and to G1P[8] rotavirus strains commonly circulating in the community and associated with gastroenteritis.
Pulmonary Localization of
Enterocytozoon bieneusi
in an AIDS Patient: Case Report and Review
Authors: Botterel F. , Minozzi C. , Vittecoq D. , Bourée P.
Published: November 2002

Enterocytozoon bieneusi
is an agent of intestinal microsporidiosis leading to malabsorption syndrome and diarrhea in AIDS patients. Respiratory tract microsporidiosis due to
Encephalitozoon
spp. has been reported. To date, however, only two cases of pulmonary involvement of
E. bieneusi
have been documented for patients with intestinal microsporidiosis. We report here another pulmonary localization of
E. bieneusi
in a human immunodeficiency virus-infected patient. Clinical features of these three cases are reviewed.
E. bieneusi
can colonize the respiratory tract but could be considered a simple carriage associated with an intestinal infection.
The avoidability of head and neck injuries in ice hockey: an historical review
Authors: Biasca N , Wirth S , Tegner Y
Published: November 2002

The number of minor traumatic brain injury (mTBI), cerebral concussions, is increasing and cannot be eliminated by any kind of equipment. Prevention strategies, such as the introduction of “checking from behind” rules have become effective in decreasing the number of severe spinal injuries. A new “head checking” rule should reduce mTBI in the same way in the following years. Mouthguards should be mandatory as an effective device for the prevention of dental and orofacial injuries, as well as reducing the incidence and severity of mTBI. A new internet database system, the International Sports Injury System (ISIS) should improve epidemiological analysis of head, face, and spinal injuries worldwide. ISIS should provide an internationally compatible system for continuous monitoring of risk factors, protective effects of equipment, and protective effects of equipment and effects of changes in rules through the years.
Patterns of illness and injury encountered in amateur ocean yacht racing: an analysis of the British Telecom Round the World Yacht Race 1996–1997
Authors: Price C J S , Spalding T J W , McKenzie C
Published: November 2002

Objectives:To quantify the incidence and type of medical problem arising during an amateur circumnavigation yacht race, the BT Global Challenge.

Methods:All cases from 14 participating yachts in a confidential medical log completed by an appointed medic were reported.

Results:A total of 685 cases were reported, of which 299 (43.6%) were injuries and 386 (56.4%) illnesses. The subtype of injury, illness, and three evacuations at sea are described.

Conclusion:Injury and other forms of medical problem are relatively common in an amateur long distance ocean yacht race. Most can be adequately managed at sea, provided that optimal communication, training, and equipment are provided and maintained.
Unilateral vulval hypertrophy in competitive female cyclists
Authors: Humphries D
Published: November 2002

Four cases of unilateral vulval hypertrophy in competitive female cyclists are reported. This condition is unusual in the general population and often associated with serious pathology, including infection and neoplasm. It is previously unreported in female cyclists and should be included in the list of conditions that may affect them.
FRACTALS AND COMBAT MODELING: USING MANA TO EXPLORE THE ROLE OF ENTROPY IN COMPLEXITY SCIENCE
Authors: LAUREN MICHAEL K. , STEPHEN ROGER T.
Published: November 2002

Combat data collected from World War II and a cellular automaton combat model called MANA are shown to display fractal properties. This strongly supports our earlier hypotheses as to the nature of combat attrition data. It also provides a method by which we can judge a combat model's ability to produce realistic synthetic combat data. The data appear to display properties extremely similar to those of the fractal cascade models used to describe turbulent dynamics. Interestingly, the fractal parameters appear to depend on how the model is set up, implying that they are determined by the boundary and initial conditions. Examination of the dynamical rules used in the MANA model simulation suggests that the model entities need to respond to changes in the level of order on the battlefield grid for fractal behavior to occur. Such data imply that the entropy of the battlefield is dependent on the scale at which it is examined. We speculate that such formations in a military case effectively act to isolate the highest level of command from disorder in the lowest. If disorder within a force grows to the point where that force can no longer maintain a fractal-like distribution, the force distribution may tend to become uniformly random, effectively destroying its viability as a combat unit.
FRACTAL ANALYSIS OF THE HOURLY TIME VARIABILITY IN SELF-POTENTIAL AND FLOW VARIATIONS CONCOMITANTLY MEASURED IN THE SOOS NATURE PARK (CZECH REPUBLIC)
Authors: BALASCO MARIANNA , HEINICKE JENS , KOCH ULRICH , MARTINELLI GIOVANNI , TELESCA LUCIANO
Published: November 2002

Scaling properties of self-potential and carbon dioxide flow rate data, concomitantly recorded in a CO 2 degassing area at Soos, Bohemia (Czech Republic), have been investigated by means of several fractal tools. The power spectra, behaving as power law function of the frequency, show the presence of colored noise-type dynamics. The Higuchi analysis reveals the self-potential and flow signals to have the property of fractal curves. The variograms, analyzing time-scales stretching from one hour to almost 50 hours, reveal the presence of approximately three time-scaling ranges, characterized by different features in terms of persistence of the processes underlying the signals.
A Computationally Efficient Evolutionary Algorithm for Real-Parameter Optimization
Authors: Deb Kalyanmoy , Anand Ashish , Joshi Dhiraj
Published: November 2002

Due to increasing interest in solving real-world optimization problems using evolutionary algorithms (EAs), researchers have recently developed a number of real-parameter genetic algorithms (GAs). In these studies, the main research effort is spent on developing an efficient recombination operator. Such recombination operators use probability distributions around the parent solutions to create an offspring. Some operators emphasize solutions at the center of mass of parents and some around the parents. In this paper, we propose a generic parent-centric recombination operator (PCX) and a steady-state, elite-preserving, scalable, and computationally fast population-alteration model (we call the G3 model). The performance of the G3 model with the PCX operator is investigated on three commonly used test problems and is compared with a number of evolutionary and classical optimization algorithms including other real-parameter GAs with the unimodal normal distribution crossover (UNDX) and the simplex crossover (SPX) operators, the correlated self-adaptive evolution strategy, the covariance matrix adaptation evolution strategy (CMA-ES), the differential evolution technique, and the quasi-Newton method. The proposed approach is found to consistently and reliably perform better than all other methods used in the study. A scale-up study with problem sizes up to 500 variables shows a polynomial computational complexity of the proposed approach. This extensive study clearly demonstrates the power of the proposed technique in tackling real-parameter optimization problems.
Fludarabine, mitoxantrone, dexamethasone (FND) compared with an alternating triple therapy (ATT) regimen in patients with stage IV indolent lymphoma
Authors: Tsimberidou Apostolia M. , McLaughlin Peter , Younes Anas , Rodriguez Maria A. , Hagemeister Fredrick B. , Sarris Andreas , Romaguera Jorge , Hess Mark , Smith Terry L. , Yang Ying , Ayala Ana , Preti Alejandro , Lee Ming-Sheng , Cabanillas Fernando
Published: November 2002

Treatment for patients with stage IV indolent lymphoma ranges from watchful waiting to intensive chemotherapy and stem cell transplantation. In this trial we compared 2 induction regimens followed by 1 year of interferon maintenance therapy. Fludarabine, mitoxantrone (Novantrone), and dexamethasone (FND) were compared with an alternating triple therapy (ATT) regimen (CHOD-Bleo, ESHAP, and NOPP). Maintenance interferon/dexamethasone was given for 1 year in both treatment arms. Endpoints were comparisons of remission rates, survival, failure-free survival (FFS), molecular response rates, and toxicities. One hundred forty-two patients with previously untreated stage IV indolent lymphoma were evaluable (73 on FND; 69 on ATT). The overall response rates were 97% for FND and 97% for ATT (P = .9). The median follow-up is 5.9 years. The 5-year survival rates were 84% with FND and 82% with ATT (P = .9); the 5-year FFS rates were 41% with FND and 50% with ATT (P = .02). In a multivariate analysis, factors predicting for longer FFS were β2-microglobulin less than 3 mg/L (P = .01) and ATT treatment (P = .03). ATT was associated with a substantially higher rate of grade 3-4 toxicities than FND. In conclusion, both regimens were associated with high rates of response and survival. ATT was associated with substantially longer FFS, but it was more toxic than FND.
Induction of platelet thrombi by bacteria and antibodies
Authors: Sjöbring Ulf , Ringdahl Ulrika , Ruggeri Zaverio M.
Published: November 2002

We have characterized 2 distinct mechanisms through which infectious agents may promote platelet adhesion and thrombus formation in flowing blood, thus contributing to the progression of disease. In one case, the process initiates when the integrin αIIbβ3 mediates platelet arrest onto immobilized bacterial constituents that have bound plasma fibrinogen. If blood contains antibodies against the bacteria, immunoglobulin (Ig) G may cluster on the same surface and activate adherent platelets through the FcγRIIA receptor, leading to thrombus growth. As an alternative, bacteria that cannot bind fibrinogen may attach to substrates, such as immobilized plasma proteins or components of the extracellular matrix, which also support platelet adhesion. As a result of this colocalization, IgG bound to bacteria can activate neighboring platelets and induce thrombus growth regardless of their ability to initiate platelet-surface contact. Our results demonstrate that intrinsic constituents of infectious agents and host proteins play distinct but complementary roles in recruiting platelets into thrombi, possibly contributing to complications of acute and chronic infections.
Transgenic targeting with regulatory elements of the humanCD34 gene
Authors: Radomska Hanna S. , Gonzalez David A. , Okuno Yutaka , Iwasaki Hiromi , Nagy Andras , Akashi Koichi , Tenen Daniel G. , Huettner Claudia S.
Published: November 2002

The human CD34 gene is expressed on early progenitor and stem cells in the bone marrow. Here we report the isolation of the human CD34 locus from a human P1 artificial chromosome (PAC) library and the characterization and evaluation of this genomic fragment for expression of reporter genes in stable cell lines and transgenic mice. We show that a 160-kb fragment spanning 110 kb of the 5′ flanking region and 26 kb of the 3′ flanking region of theCD34 gene directs expression of the human CD34gene in the bone marrow of transgenic mice. The expression of human CD34 transgenic RNA in tissues was found to be similar to that of the endogenous murine CD34 gene. Colony-forming cell assays showed that bone marrow cells staining positive for human CD34 consist of early progenitor cells in which expression of CD34 decreased with cell maturation. In order to test the construct for its ability to express heterologous genes in vivo, we used homologous recombination in bacteria to insert the tetracycline-responsive transactivator protein tTA. Analysis of transgenic human CD34-tTA mice by cross breeding with a strain carrying Cre recombinase under control of a tetracycline-responsive element demonstrated induction of Cre expression in mice in a pattern consistent with the expression of the human CD34 transgene.
Chimeric IgA antibodies against HLA class II effectively trigger lymphoma cell killing
Authors: Dechant Michael , Vidarsson Gestur , Stockmeyer Bernhard , Repp Roland , Glennie Martin J. , Gramatzki Martin , van de Winkel Jan G. J. , Valerius Thomas
Published: November 2002

Antibodies against human leukocyte antigen (HLA) class II, such as 1D10 or Lym-1, are currently being evaluated for the treatment of B-cell lymphomas. Previous studies have demonstrated that, in addition to IgG Fc receptors, the human myeloid IgA receptor (FcαRI, CD89) also effectively triggered tumor cell killing. Therefore, we used the variable light and heavy chain sequences from another murine anti–HLA class II hybridoma, F3.3, to generate a panel of chimeric human/mouse antibodies, including human immunoglobulin A1 (IgA1), IgA2, IgG1, IgG2, IgG3, and IgG4. Antibody production was accomplished by stable transfection of baby hamster kidney cells, and binding activity and specificity were confirmed by enzyme-linked immunosorbent assay (ELISA) and Western blotting. All constructs demonstrated similar binding to HLA class II. Functional studies revealed that chimeric IgG1, IgA1, and IgA2 triggered similar levels of tumor cell lysis. Analyses of effector populations, however, demonstrated that killing by chimeric IgG1 constructs was triggered mainly by human mononuclear cells and complement, while IgA1 and IgA2 mediated effective lysis by polymorphonuclear neutrophils. Importantly, IgG1 and both IgA isotypes were equally effective at killing freshly isolated human chronic lymphocytic leukemia cells. Chimeric IgA antibodies against HLA class II may constitute attractive reagents for lymphoma therapy.